Assessment of apical extrusion of irrigation in mandibular molars during application of the novel Gentle Wave system in a simulated environment

Assessment of apical extrusion of irrigation in mandibular molars during application of the novel Gentle Wave system in a simulated environment by Karine Charara A thesis submitted in conformity with the requirements for the degree of Master of Sciences Graduate Department of Endodontics University of Toronto Copyright by Karine Charara 2015

Assessment of apical extrusion of irrigation in mandibular molars during application of the novel Gentle Wave system in a simulated environment Karine Charara Degree of Master of Sciences Abstract Graduate Department of Endodontics University of Toronto 2015 This study assessed apical extrusion during treatment with GentleWaveTM (GW), conventional open- ended 30G needle (CN) or EndoVac (EV) in root canals enlarged to different dimensions with and without apical constriction. No extrusion occurred with GW and EV, while frequency of extrusion with CN was 33%. Mean extruded water mass using CN ranged in mesial canals from 0.000±0.000 g (OI) to 0.047±0.098 g (MI) and in distal canals from 0.123±0.191 g (MI) to 0.505±0.490 g (OI). With TI and OI instrumentation, extruded mass in distal canals was significantly higher than in mesial canals (p<0.002) and than in distal canals with MI (p<0.020). Within this study's limitations, root canal treatment with GentleWaveTM and irrigation with EndoVac was not associated with extrusion. ii

Table of content List of Tables... v List of Figures... vi Introduction... 1 1. Apical periodontitis... 1 1.1. Apical periodontitis: a disease of biofilm origin... 1 1.2. Treatment of apical periodontitis... 3 1.3. Mechanical instrumentation... 4 2. Irrigation... 5 2.1. Sodium hypochlorite... 6 a. Mechanism of action of sodium hypochlorite... 7 b. Concentration of sodium hypochlorite... 8 c. Sodium hypochlorite cytotoxicity and related accidents... 9 d. Sodium hypochlorite accident recognition and management... 11 2.2. Convention Syringe and Needle for Irrigation... 12 2.3. Vapour lock... 14 2.4. Additional Cleaning Techniques... 15 a. Manual and Machine- assisted Irrigation Techniques... 16 b. Continuous and Intermittent Flushing Techniques... 17 2.5. Apical Negative Pressure... 17 2.6. The EndoVac System... 18 a. Debris Removal... 20 b. Microbial control... 22 c. Smear Layer removal... 24 d. Calcium hydroxide removal... 25 e. Safety... 26 3. GentleWave System... 27 a. GentleWave characteristics... 28 b. Tissue dissolution... 29 c. Calcium hydroxide removal... 29 Rationale... 31 Aim... 31 Hypothesis... 31 JOE Article... 32 Statistics... 39 iii

1.1. Amount of Extrusion and Canal Size... 39 1.2. Correlation of Extrusion Masses and Canal Size... 39 1.3. CNI Extrusion masses for Mesial vs. Distal Canals... 40 Discussion... 42 1. Methodology... 42 1.1. Experimental set- up... 42 1.2. Back pressure... 44 1.3. Tooth selection... 44 1.4. Instrumentation levels... 45 1.5. Validation and control groups... 45 1.6. Solution choice... 46 2. Results... 46 2.1. Needle... 46 2.2. GentleWave... 47 2.3. EndoVac... 48 Conclusion... 50 References... 51 Figures... 61 Figure 1... 61 Figure 2... 62 Figure 3... 63 Figure 4... 64 Figure 5A and 5B... 65 Figure 6... 66 Figure 7... 67 Figure 8... 68 Figure 10... 70 Figure 11... 71 Tables... 72 Table 1... 72 Table 2... 73 Table 3... 74 Table 4... 75 iv

List of Tables Table 1: Summary of Wilcoxon t- test comparing CNI and MU for each canal size. Table 2: Summary of the Correlation Analysis using Pearson s r coefficient. Table 3: Summary of the chance of extrusion for each endodontic treatment device for all canals and all canal sizes. Table 4: Mean extruded irrigation mass (gram) in each of 16 specimens tested v

List of Figures Figure 1: Sodium Hypochlorite accident Figure 2: EndoVac system components Figure 3: Final irrigation protocol using EndoVac system Figure 4: Gentlewave system Figure 5: Experimental set- up Figure 6: Pilot study Figure 7: Amount of extrusion; mesial canals Figure 8: Amount of extrusion; distal canals Figure 9: Percent of mass extruded in mesial canals Figure 10: Percent of mass extruded in distal canals Figure 11: Average of extruded fluid during 30 s with conventional needle irrigation (NI) vi

Introduction 1. Apical periodontitis 1.1. Apical periodontitis: a disease of biofilm origin Periapical tissues consist of cementum, periodontal ligament and alveolar bone. The response of the periapical tissues to various injuries is manifested as an immune- inflammatory reaction resulting in apical periodontitis (AP) (1). AP has been proven to be of bacterial origin and strongly associated with bacterial biofilms (2 4). AP is a tangible manifestation of the incapacity of the immune response to control the irritation caused by root canal s microbes. Infected root canal counts of between 103 and 108 bacteria of different species often agglomerate into a biofilm. A biofilm is a well- organized micro- community of mono or multiple bacterial species embedded in a self- produced matrix of extracellular polymeric substance (EPS) and attached to a surface. Within the biofilm, microorganisms are more resistant to their eradication compared to their planktonic form (5). AP is the inflammation and destruction of the periapical tissues that is of pulpal origin. Radiographically it is manifest as periapical radiolucency. Although the microbial infection of the pulp is the primary cause of AP, the pathologic changes are not directly associated with the bacteria themselves but rather with their toxins, metabolic by- products and necrotic pulp remnants (1). The initiation of AP is mainly caused by bacterial lipopolysaccharides (LPS) and other microbial toxins released during bacterial cell growth and death entering into the periapex. Those irritants are capable of inducing an innate and 1

adaptive immune response. When entering the periapical tissues, pathogen- associated molecular patterns (PAMPs) found on pathogens such as LPS, a bacterial toxin, activate the innate immune system. PAMPs are recognized by different pattern recognition receptors (PRRs) or toll like receptors (TLRs) on host cells such as phagocytes, dendritic cells and B lymphocytes. Once activated, these local cells increase the permeability of the blood vessel, induce vasodilatation and activate cytokine production, including RANKL, IL- 1, IL- 6, TNF- α, and PGE2, and consequently promote osteoclastogenesis and bone loss (6). Periapical inflammation is a direct result of interactions between the bacteria in an untreated infected root canal system and the host s defense or immune system (7). It begins as an acute inflammatory response but it is a dynamic situation that can change spontaneously throughout the disease process. As there is no longer any blood supply to a necrotic pulp or into the root canal system in a pulpless tooth, the host s defense cells can not reach the source of the irritation (i.e., the bacteria in the canal) and therefore the body is unable to eliminate the infection. Hence, a chronic inflammatory response develops in the periapical region and the intra- canal bacteria survive with nutrients being obtained from tissue fluid and inflammatory exudate that seep in to the root canal system through the apical foramen (8). Saliva and food substances may also penetrate through the original pathway of entry of the bacteria (i.e. caries, cracks or broken- down restoration margins) to help supply nutritional elements for the organisms. Once an infection is established within the root canal system, the bacterial numbers will gradually increase through normal cell reproduction and proliferation mechanisms (8). To eliminate the infection, proper debridement of the root canal system has to be performed. 2

1.2. Treatment of apical periodontitis To eliminate bacteria and their toxic metabolites from the root canal system and to prevent their diffusion into the periapical tissues, which may cause bone destruction and AP, endodontic treatment is indicated. The primary goal of endodontic treatment is to prevent or to treat AP. The endodontic treatment of a tooth with irreversible pulp inflammation aims to prevent infection from developing in the periapical tissue, as the apical third is generally free of bacteria (9). In the presence of a necrotic infected pulp or a previously treated root canal with persistent or recurrent infection, AP has likely developed. Achieving adequate debridement of the necrotic pulp tissue and bacterial elimination are critical procedural goals when treating infected root canals. The reduction of the bacterial load to a number below the threshold necessary to sustain AP is correlated with a favourable outcome (9,10). Sjögren et al. (10) demonstrated that the bacteria remaining in the root canal will influence the endodontic outcome of teeth with AP. Periapical healing was observed five years after the completion of the treatment and healing occurred in 94% of teeth with no- growth cultures compared to 68% for teeth with positive cultures prior to root filling. Because culturing procedures have their limitations, a no- growth culture is not a confirmation of complete disinfection. It can be associated with improved prognosis relative to teeth with a positive culture (11). Therefore, bacterial reduction in the root canal system to the extent of achieving a no- growth culture should be a clinical goal to significantly improve the potential for healing of the periapical tissues. The biomechanical preparation of the root canal system is the primary manner used by clinicians in 3

attempting to eradicate bacteria, debris, vital and necrotic tissues from the root canal system (12). 1.3. Mechanical instrumentation Endodontic mechanical instrumentation includes different endodontic hand files, rotary files, reamers and broaches that clinicians utilize according to their preference to achieve the following goals: - - - - - - Removal of vital and necrotic tissue from the main root canal(s). Creation of sufficient space for irrigation and medication. Preservation of the integrity and location of the apical canal anatomy. Avoidance of iatrogenic damage to the canal system and root structure. Facilitation of canal filling. Avoidance of further irritation and/or infection of the periapical tissues. - Preservation of sound root dentine to allow long- term function of the tooth. (12) Byström et al. (13), demonstrated that mechanical instrumentation alone is not sufficient to predictably reduce the level of bacteria, while mechanical instrumentation combined with antibacterial irrigation is. The disinfection improves when mechanical instrumentation is coupled with antibacterial irrigation. In addition, mechanical debridement leaves at least 35% of the canals walls untouched (14). Hence, cleaning of the canal in terms of soft tissue removal and elimination of bacteria relies heavily on the adjunctive action of chemically active irrigating solutions and is crucial in terms of reaching those untreated areas. 4

2. Irrigation Mechanical instrumentation coupled with irrigation of the root canal with sodium hypochlorite (NaOCl) appreciably reduces the bacteria load in the root canal (13). A reduction has been reported in the proportion of positive bacterial cultures of between 11% and 53% and a bacterial count from an initial order of 10 3 to 10 7 to a level of 10 2 to 10 6 (15). The intricate nature of the root canal anatomy complicates the ability of endodontic instrumentation to substantially remove biofilms (16). Micro computed tomography (17,18) and tooth decalcification anatomic studies (19,20) reveal root canal systems that are complex with multiple isthmuses and an abundant number of lateral and accessory canals creating pathways to the periapical tissues. This complex root canal system is unreachable with mechanical instrumentation and requires irrigation to be properly disinfected. There are five well established aims of root canal irrigation (21): 1) To deliver irrigation throughout the entire length of the root canal so as to be in direct contact with debris and bacteria. 2) To provide lubrication for instrumentation. 3) To maintain a high concentration of the active component(s) by frequent refreshing. 4) To create wall shear stress in order to disrupt biofilm. 5) To prevent apical extrusion (irrigation should be restricted to within the root canal). Irrigation is defined as the action of washing an organ or wound with the continuous flow of water or medication. In endodontics, it represents chemical debridement of the root canal 5

system. Several solutions are available to achieve this aim. The desired characteristics of an irrigation solution are as follows: effective germicide and fungicide, non- irritating to periapical tissues, stable in solution, prolonged antimicrobial effect, active in the presence of blood, serum and protein derivatives of tissue, low surface tension, non- detrimental to repair of periapical tissues, non- staining of tooth structure, possible to inactivate in a culture medium, non- inducing a cell- mediated immune response, able to completely remove the smear layer, able to disinfect the underlying dentin and its tubules, non- antigenic, non- toxic, and non- carcinogenic to tissue cells surrounding the tooth, no adverse effects on the physical properties of exposed dentin, no adverse effects on the sealing ability of filling materials, conveniently applied and relatively inexpensive (1). 2.1. Sodium hypochlorite Currently, the irrigation solution of choice is sodium hypochlorite (NaOCl). It has excellent antibacterial properties and the capacity to dissolve organic tissue. However, it is very cytotoxic to the periapical tissues (22). NaOCl was developed in France around 1789 and labelled as Eau de Javel. At first, it was used as a bleaching agent for cotton but eventually its antiseptic properties were recognized. Dakin recommended NaOCl as an antiseptic agent in hospitals and as a rinse to treat the wounds of the soldiers during the first world war (23). The manufacture of NaOCl requires several steps. In the process, sodium hypochlorite 6

(NaOCl) and sodium chloride (NaCl) are formed when chlorine is passed into cold and dilute sodium hydroxide solution. It is prepared industrially by electrolysis with minimal separation between the anode and the cathode. The solution must be kept below 40 C to prevent the undesired formation of sodium chlorate. NaOCl production chemical reaction Cl2 + 2 NaOH NaCl + NaOCl + H2O Hence, chlorine is simultaneously reduced and oxidized; this process is known as disproportionation. Commercial solutions always contain significant amounts of sodium chloride (common salt) as the main by- product, as seen in the equation above. Sodium hypochlorite is a clear, slightly yellowish solution with a characteristic odour. It has a relative density of 1.1 g/cm 3 (5,5% aqueous solution). As a bleaching agent for domestic use it usually contains ±5% sodium hypochlorite with a ph of around 11. a. Mechanism of action of sodium hypochlorite Physico- chemical characteristics of NaOCl are important for the explanation of its mechanism of action. The antimicrobial process consists of saponification and enzymatic inactivation. Amino acid neutralization and chloramination reactions occur in the presence of microorganisms and organic tissue lead to the saponification. NaOCl also promotes irreversible inactivation by hydroxyl ions and a chloramination reaction related to bacterial essential enzymatic sites. The antimicrobial mechanism of NaOCl is also enhanced with the high ph of sodium hypochlorite that interferes with the cytoplasmic membrane integrity with an irreversible enzymatic inhibition, biosynthetic alterations in cellular metabolism and phospholipid degradation and lipidic peroxidation. The organic dissolution action can 7

be observed in the saponification reaction when sodium hypochlorite degrades lipids and fatty acids resulting in the formation soap and glycerol (24). b. Concentration of sodium hypochlorite In endodontics, NaOCl is used in concentrations of between 0.5% and 6%. There is considerable variation in the literature regarding the influence of the concentration on the antibacterial effect of NaOCl. Several clinical and laboratory studies have failed to demonstrate any significant difference in antibacterial effect between different NaOCl concentrations after root canal sampling (25). Other studies report considerably longer times for the killing of the same species when a lower concentration is utilized (26,27). Byström and Sundqvist studied the irrigation of root canals that were necrotic and contained a mixture of anaerobic bacteria. These investigators showed that using 0.5% or 5% NaOCl resulted in considerable reduction of bacterial counts in the canal when compared with irrigation with saline (13,28,29). Siqueira and colleagues (25) reported similar results using root canals infected with Enterococcus faecalis. The studies failed to show a significant difference in the antibacterial efficacy between the low and high concentrations of NaOCl (26). These in vitro studies may not be applicable to the clinical reality where the root canal is partially filled with organic tissue. It has been demonstrated that the presence of organic tissue during the killing experiments has a negative effect on the antibacterial activity of NaOCl. Inflammatory exudate, tissue remnants and microbial biomass consume NaOCl and weaken its effect. NaOCl in higher concentrations have better tissue dissolving ability but 8

Haapasalo and colleagues (30) showed that the presence of dentin caused marked delay in the killing of E. faecalis by 1% NaOCl. To counteract the impact of an environment such as an infected tooth, continuous replenishment of NaOCl and adequate time exposure are recommended. An emphasis is placed on the importance of repeated exchanges of large volumes of irrigation to maintain adequate antibacterial effectiveness, which compensates for a lower concentration (25). Regarding the ability of NaOCl to dislodge biofilm; an in vitro biofilm study (31) demonstrated a significant difference in the effectiveness against biofilm bacteria with 3% and 6% NaOCl, the higher concentration being more effective. Nevertheless, the clinician must keep in mind that a higher concentration is more toxic to the periapical tissue. c. Sodium hypochlorite cytotoxicity and related accidents In light of the cytotoxicity of the NaOCl, its extrusion from the root canal will affect the periapical tissue and may cause the patient a series of complications of a variable clinical significance, including post- operative pain (32). Although devastating endodontic NaOCl accidents are rare (33), the cytotoxic effects of NaOCl on vital tissue are well established (34). The associated sequelae of NaOCl extrusion have been reported to include life- threatening airway obstructions (35), facial disfigurement requiring multiple corrective surgical procedures (36), permanent paraesthesia with loss of facial muscle control (37) and tooth loss (38). Although the exact aetiology of a NaOCl accident is still uncertain, based on the evidence from actual accidents and the location of the associated tissue trauma, it would appear that 9

an intravenous injection might be the main cause. The patient shown in figure 1 (73) demonstrates a widespread area of tissue trauma that is in contrast to the characteristics of NaOCl accident reported by Pashley (34,39). This extensive trauma, particularly involving the pattern of ecchymosis around the eye, could only have occurred if the NaOCl had been introduced intravenously to a vein close to the root apex through which extrusion of the NaOCl occurred and the NaOCl then travelled to the venous complex. This would require positive pressure apically exceeding the venous pressure, for which the mean value is higher than 5.88 mmhg, the central venous pressure (22). In other words, NaOCl extrusion into the venous system is more likely to occur when the apical pressure of irrigation solution is greater than venous pressure or when there is a decrease in the apical resistance (22). Female bone density has a reduced resistance to apical extrusion (40). Moreover, the thickness of the cortical bone is thinner in women than in men (40). This could explain why NaOCl accidents are seen more often in women and maxillary teeth, more specifically the maxillary pre- molars (40). One in vitro study where a positive- pressure needle irrigation technique was used to mimic clinical conditions and techniques, demonstrated that the apical pressure generated easily exceeded the value of central venous pressure(41). The results of this study suggested that a combination of factors was necessary for a severe NaOCl accident to occur. The hypothesis that involves intravenous infusion of extruded NaOCl into the facial vein via non- collapsible venous sinusoids within the cancellous bone has been suggested (12). This does not imply that NaOCl can or should be excluded as an endodontic solution; in fact, its use is essential to achieve adequate chemical debridement. Therefore it must be 10

delivered safely. With traditional root canal irrigation, clinicians must be careful when determining how far an irrigation needle is placed into the canal. Recommendations for avoiding NaOCl accidents include not binding the needle in the canal, adequate establishment of the working length, not placing the needle close to working length (WL) (i.e. 1 mm away from WL with a closed- ended needle and 3 mm away from WL with an open- ended needle), and using a gentle flow rate when delivering the solution with a positive pressure device (39). d. Sodium hypochlorite accident recognition and management When a surprising unfortunate accident happens, the clinician must be prepared to provide the best care to the patient. First, the clinician must be able to recognize that extrusion of sodium hypochlorite has occurred (23). Most of the time, a sharp pain despite the action of the local anaesthetic manifests it. The depolarization of the transient receptor potential vanilloid one (TRPV1) is suggested to cause the sudden painful sensation (42). TRPV1 is a non- selective cation channel that may be activated by a wide variety of exogenous and endogenous physical and chemical stimuli. The activation of TRPV1 leads to a painful, burning sensation. TRPV1 receptors are found mainly in the nociceptive neurons of the peripheral nervous system, but they have also been described in many other tissues, including the central nervous system (42). TRPV1 is involved in the transmission and modulation of pain (nociception). A profuse bleeding from the root canal follows the painful sensation felt by the patient (22). The clinician should reassure the patient, generously flush the root canal with saline and consider a local block with a long- acting local anaesthetic such as Bupivacaine HCl 0.5% with epinephrine 1:200,000. The patient should 11

be kept under observation for thirty minutes while removing the drainage (blood) with high volume to enhance further drainage. If the tooth drains after 30 min, the clinician should consider leaving it open for twenty- four hours. Amoxicillin 500 mg tid for 5 days is indicated in more severe cases. Other antibiotics can be prescribed if patient has an allergy to penicillin. In all cases, clinicians should prescribe analgesics and patients should be told to apply cold compresses for six hours (43). Some authors (43,44) mention that corticosteroids or anti- histamines would help minimizing the ensuing inflammatory process; therefore, they could be added to the list of prescribed drugs. The clinician should call the patient the night of the accident; the patient should be seen the day after and close follow- up should be maintained until complete resolution of the symptoms. If any kind of paresthesia persists for more than 10 weeks, the patient should be referred to a neurologist or an oral surgeon (43). 2.2. Convention Syringe and Needle for Irrigation NaOCl is conventionally delivered via a positive- pressure syringe fitted with a needle, a variety of which are available (26). A 5 ml syringe has been recommended as a reasonable compromise between less frequent refilling and ease of use. This size of syringe allows the clinician to keep good control of the plunger and to not constantly having to refill the syringe. A smaller syringe could be responsible of a higher flow rate if the clinician applies the same force than when using a larger size of syringe. The irrigation flow rate is proportional to the pressure difference between the atmospheric pressure and the pressure built within the syringe by the force applied on the plunger. A larger syringe can be tiring as 12

the amount of force that needs to be applied to create the same flow rate is higher. A syringe of a small size is easier to press than one of a larger size. The needle size also influences the pressure of the delivered irrigation. For the same pressure difference, the flow through a smaller needle will be much less than through a larger needle. In other words, a larger difference is required to achieve the same flow rate through a smaller needle. Most of the time in endodontic treatment, irrigation is delivered with either a 30- or 27- G endodontic slot tipped needle placed into the canal until just short of the binding point. The difficulty with this technique is that the depth of needle penetration is dependent on the size and morphology of each canal. With the current endodontic protocols, total elimination of the root canal bacteria is unpredictable. Nair et al. (45) looked at the microbial status of the apical root canal system of necrotic, mandibular first molars with apical periodontitis after single- visit endodontic therapy during which NaOCl was delivered using a conventional needle (CN) for irrigation. After removal of the apical root segment and evaluation using correlative light and transmission electron microscopy, they found that 14 of 16 teeth had residual intracanal bacteria (45). They noted that the microbes were located in inaccessible recesses, isthmuses and accessory canals. Even with the use of NaoCl, obtaining predictably root canal cleanliness and no- growth cultures remained unpredictable (46). Several studies (47 49) evaluated the root canal after initial root canal treatment and biofilms were located in areas inaccessible to root canal instrumentation. With bacteria frequently surviving the cleaning and shaping procedure and debris compromising cleanliness, adjuvant techniques 13

to conventional root canal treatment cleaning protocols may predicate better root canal cleaning and disinfection. 2.3. Vapour lock In 1971, Senia was the first to describe vapor lock in root canal. Since roots are surrounded by the periodontium, unless the root canal foramen is open, the root canal behaves like a close- ended channel. This produces an apical vapour lock that resists displacement during instrumentation and final irrigation, thus preventing the flow of irrigation solution into the apical region and adequate debridement of the root- canal system (50,51). Apical vapour lock also results in gas entrapment in the apical one third (52). During irrigation, NaOCl reacts with organic tissue in the root- canal system, and the resulting hydrolysis liberates abundant quantities of ammonia and carbon dioxide (53). This gaseous mixture is trapped in the apical region and quickly forms a column of gas into which further fluid penetration is impossible. Extension of instruments into this vapour lock does not reduce or remove the gas bubble (54), just as it does not enable adequate flow of irrigation solution. The phenomenon of apical vapour lock has been confirmed in studies in which roots were embedded in a polyvinylsiloxane impression material to restrict fluid flow through the apical foramen, simulating a close- ended channel (55). The results in these studies was found to be an incomplete debridement of the apical part of the canal walls with the use of a positive- pressure syringe delivery technique (55). Micro- CT scanning and histological tests conducted by Tay et al. (55) have also confirmed the presence of apical vapour lock. In fact, 14

studies conducted without ensuring a close- ended channel cannot be regarded as conclusive on the efficacy of irrigation solutions and the irrigation system (56 58). The apical vapour lock may also explain why in a number of studies investigators were unable to demonstrate a clean apical third in sealed root canals (59 61). In a paper published in 1983, Chow determined that traditional positive- pressure irrigation had virtually no effect apical to the orifice of the irrigation needle in a closed root- canal system (62). Fluid exchange and debris displacement were minimal. Equally important to his primary findings, Chow set forth an infallible paradigm for endodontic irrigation: For the solution to be mechanically effective in removing all the particles, it has to: (a) reach the apex; (b) create a current (force); and (c) carry the particles away. (62) The apical vapour lock and consideration for the patient s safety have always prevented the thorough cleaning of the apical 3 mm. It is critically important to determine which irrigation system will effectively irrigate the apical third, as well as isthmuses and lateral canals (63), and do it in a safe manner that prevents the extrusion of irrigation solution. 2.4. Additional Cleaning Techniques Several adjuvant techniques to the conventional chemico- mechanical regiment have been suggested. For instance, increasing apical enlargement to ISO size 60 achieves 93% of no- growth cultures [10]; however, it also produces higher risk of canal transportation (12). Some postulate that inter- appointment medication such as calcium hydroxide has the ability to decrease the percentage of positive cultures [11]. The drawback of this approach 15

is that it requires a second treatment session (64). Moreover, two randomized control trials (65,66) found no significant difference between the percentage of no- growth cultures after instrumentation and calcium hydroxide application inside the root canal. In recent years several systems have been developed to activate and to deliver NaOCl into the root canal. Optimizing delivery of NaOCl throughout the root canal by continuous replenishment while avoiding periapical extrusion has been a subject of research and development in recent years (67 71). a. Manual and Machine- assisted Irrigation Techniques Root- canal irrigation systems can be divided into two categories: manual irrigation techniques and machine- assisted irrigation techniques (52). Manual irrigation techniques include the positive- pressure syringe fitted with a variety of needle designs and the manual dynamic agitation using a gutta- percha point. Machine- assisted irrigation techniques include sonics and ultrasonics, as well as newer systems such as the EndoVac (SybronEndo) designed to generate apical negative pressure, the GentleWave (Sonendo) based on multisonic pressure wave formation, the plastic rotary F File (Plastic Endo), the Vibringe sonically agitated syringe (Vibringe), the Rinsendo (Air Techniques) and the EndoActivator (Dentsply Tulsa Dental Specialties). Two important factors that should be considered during the process of irrigation are whether the irrigation systems can deliver irrigation to the apical terminus, and whether irrigation is capable of debriding areas that could not be reached with mechanical instrumentation, such as lateral/accessory canals, isthmuses and 16

deltas. b. Continuous and Intermittent Flushing Techniques Two flushing methods are currently employed to irrigate root canal systems: intermittent and continuous (52). With the intermittent flushing technique, irrigation is injected in the root canal space with a syringe and the solution can then be activated; the canal is filled several times after each activation cycle. Inversely, the continuous flush techniques provide an uninterrupted supply of fresh irrigation solution into the root canal. This technique can provide more effective results and reduce the time required for final irrigation when compared with intermittent irrigation devices. Taking into consideration that chloride (responsible for dissolving the organic tissues and NaOCl s antibacterial property) is unstable and quickly consumed, a continuous flow of irrigation would make intuitive sense (52). 2.5. Apical Negative Pressure Pressure is defined as a force per area. During root canal treatment, pressure is exerted onto the root canal wall when the irrigation is delivered into the root canal space. Negative pressure refers to a situation in which an enclosed volume has lower pressure than its surroundings. 17

2.6. The EndoVac System The EndoVac system was developed to safely and predictably deliver irrigation to the apical terminus thereby allowing a better penetration of the irrigation solution into the intricate anatomy and morphology of the root canal system such as isthmuses, inter- canal and intra- canal communications, curvatures and oval shaped canals (63). Apical negative pressure systems for irrigation have the ability to suction, thereby drawing and delivering the irrigation solution passively to the apex (52). The EndoVac system delivers the chosen irrigation solution passively to the apex (63,72) and positively addresses the problem of irrigation penetration past the apex into the periapical tissue which may result in treatment complications (73,74). The EndoVac apical negative- pressure irrigation system has three active component parts (figure 2): the Master Delivery Tip (MDT), the macro cannula and the micro cannula. The MDT accommodates a syringe of irrigation solution, which is expressed through a 20- gauge needle. There is also a plastic suction hood attached around the 20 gauge needle which is connected to clear plastic tubing which inserts into a multiport adaptor which in turn is inserted into the high volume suction (54). As such the MDT can simultaneously deliver and evacuate any excess irrigation solution that may flow over from the pulp chamber. The macro cannula is used to draw irrigation solution by way of suction from the chamber to the coronal and middle segments of the canal while irrigation solution is simultaneously delivered to the pulp chamber directed towards an axial wall and never towards a canal orifice. The macro cannula or micro cannula is connected via clear plastic tubing to the high- speed suction of the dental unit via the multiport adaptor. The plastic macro cannula 18

has an external diameter of 0.55 mm and an internal diameter of 0.35 mm. It is made of blue translucent plastic, has a 0.02 taper and is meant for single use only. It is attached snug to an autoclavable aluminium handpiece and is used in an up and down pecking motion while the irrigation solution is simultaneously delivered passively into the pulp chamber in the manner mentioned above. It is used to remove the gross debris and tissue left behind during instrumentation. The stainless- steel micro cannula has an external diameter of 0.32 mm and zero taper. It has four sets of three laser- cut, laterally positioned offset holes adjacent to its closed end, each 100 µ in diameter and spaced 100 µ apart (71). While the micro cannula acts to evacuate debris at full working length, these micro holes act as filters to prevent the clogging of the internal lumen of the micro cannula which is 0.20 mm in diameter. The micro cannula is attached to an autoclavable aluminum finger piece and is used for irrigation of the apical part of the canal when it is positioned at working length. The micro cannula has a closed end and should be taken to the full working length to aspirate irrigation solution and debris. Because of the dimensions of the micro cannula, canals need to be enlarged to ISO size 35 with 0.04 taper or larger. Alternatively, the manufacturer recommends an enlargement of the root canal to ISO size 40/0.02. During irrigation, the MDT delivers irrigation solution to the pulp chamber and siphons off the excess irrigation solution to prevent overflow. Both the macro cannula and micro cannula exert negative pressure that pulls fresh irrigation solution from the chamber, down the canal to the tip of the cannula, into the cannula, and out through the suction hose. Thus, a constant flow of fresh irrigation solution is delivered by negative pressure to working length, allowing the reaction of hydrolysis to continually occur. Figure 3 present the final 19

irrigation protocol using EndoVac system. a. Debris Removal Several studies were carried out to evaluate the EndoVac system s ability to remove debris within the root canal system after instrumentation with rotary files (70,75 79). Debridement is a principal objective of root canal treatment and remains a challenge especially in the apical portion of the canal and within the isthmuses and lateral and accessory canals. Debridement is the elimination of organic and inorganic substances as well as microorganisms from the root canal by mechanical and/or chemical means (80). When compared to traditional syringe and side- vented needle irrigation, the EndoVac system has demonstrated better control to reach the last millimetre of the root canal. Some in vitro and in vivo studies have demonstrated greater removal of debris from the apical walls and a statistically cleaner result using negative apical pressure irrigation in closed root- canal systems with sealed apices. In an in vivo study of 22 teeth by Siu and Baumgartner (81), less debris remained at 1 mm from the working length using negative apical pressure compared to the use of traditional needle irrigation. Shin et al. (78) found in an in vitro study of 69 teeth comparing traditional needle irrigation with negative apical pressure that these methods both resulted in clean root canals, but that apical negative pressure resulted in less debris remaining at 1.5 and 3.5 mm from working length (70,78,81). When comparing root- canal debridement using manual- dynamic agitation (using a well fitted gutta- percha cone in an up and down motion in the canal) or the EndoVac system for final agitation in a closed system and an open system, it was found that 20

the presence of a sealed apical foramen adversely affected debridement efficacy when manual- dynamic agitation was used, but did not adversely affect results when the EndoVac system was used. Apical negative- pressure irrigation is an effective method to overcome the fluid- dynamic challenges inherent in closed root- canal systems (55,82). The ability of the EndoVac system to significantly clean more debris from the mechanically inaccessible recess of the in vitro root canal model may be caused by bubble formation during irrigation delivery, creating higher wall shear stresses by a two- phase air liquid flow phenomenon that is well known in other industrial debridement systems (83). To enhance cleanliness of the root canal system, EndoVac system has the ability to safely deliver irrigation solution to working length (70) by pulling the irrigation solution into the canal and removing it by negative pressure (70). This vacuum action enhances the volume of solution and the circulation of the irrigation solution in the apical end of the root canal. Moreover, the negative pressure avoids air entrapment in the apical third (79) and promotes a regular replenishment of the irrigation solution apically (79). A recent study demonstrated that the volume of irrigation solution delivered apically was significantly higher than the volume delivered by conventional syringe needle irrigation within the same period (70), and resulted in significantly more debris removal at 1 mm from working length than did needle irrigation. One study is not in agreement with those positive outcomes discussed above (76). Jiang et al. (76) evaluated the EndoVac system s ability to remove dentin debris from artificially made grooves in standardized root canals. The model was made of a single tooth root in which an apical groove comparable to an ovoid apical canal was created and packed with 21

dentine debris. They compared several devices to activate the irrigation solution. Once the irrigation regimen was completed, they viewed the grooves through a stereomicroscope to evaluate the residual dentine debris. A score between 0 and 3 was given to each specimen, 0 = the groove is empty, 1 = less than half of the groove is filled with debris. 2 = More than half of the groove is filled with debris. 3 = the complete groove is filled with debris. The specimens irrigated with the EndoVac System had their groove completely filled with debris (score 3) 65% of the time while 35% had less than half filled with debris (76). It is important to note that Jiang et al. (76) failed to follow the manufacturer s instructions by failing to use the critical macro cannula, an error that could easily cause the micro cannula to clog and become ineffective. A weaker capacity of the EndoVac system to remove apical debris could be attributed to the minimal turbulence intensity produced within the canal by the micro cannula (84). This evidence of low wall shear stress values causes a minimum physical interaction between the irrigation solution and the root canal walls (84). This absence of interaction may explain the difficulty of the irrigation solution to reach the root canal s lateral canals and anastomoses(72). b. Microbial control The effective removal of organic and inorganic tissues would allow better access and elimination of endodontic pathogens, responsible for apical periodontitis, localized in the root canal system. Hockett et al. (85) tested the ability of apical negative pressure irrigation to remove a thick biofilm of E. faecalis in mesial roots of mandibular molars, finding that these specimens rendered no- growth cultures after 48 hours of incubation, while some of those irrigated using traditional positive- pressure irrigation were positive at 48 hours (85). 22

One in vivo dog study found that negative apical pressure irrigation with 2.5% NaOCl resulted in similar bacterial reduction than the use of apical positive pressure irrigation combined with seven days of intracanal medication with the triple- antibiotic paste (86). The triple- antibiotic Trimix (metronidazole, ciprofloxacin, and minocycline) has been utilised for pulpal regeneration/revascularisation in teeth with incompletely formed apices (87). The antibiotic medication is applied in regeneration cases to safely kill bacteria. Since the triple- antibiotic versus the use of EndoVac with NaOCl were statistically equivalent for mineralised tissue formation and the repair process (86); the study (86) suggests that EndoVac may overcome the need for intracanal medication. Further research is required to evaluate this potential. Using apical negative pressure with NaOCl also decreases the risk of drug resistance, tooth discoloration, and allergic reactions often seen with the administration of antibiotics (88,89). A recent randomized controlled clinical trial (90) compared the antimicrobial effectiveness of EndoVac system and the traditional positive pressure syringe and needle for irrigation. From the sixteen mandibular molars treated with the conventional method, no- growth culture was found in 67% compared to 100% among the apical negative pressure irrigation group. A second clinical study (91) demonstrated a higher frequency of obtaining no- growth cultures with the EndoVac system compared to a syringe with regular needle. Unlike Cohenca et al. (90), Pawar et al. (91) did not report a significant difference between the two clinical groups. However, Pawar et al. (91) added an overriding codicil in their discussion: The original EndoVac protocol recommends using a concentration of 5.25% NaOCl. Almost all studies investigating the efficacy of EndoVac have used NaOCl at concentrations ranging from 2.5%- 6%. The use of 0.5% NaOCl [a 1,000 % dilution from the manufacturer s instructions] in this study could be 23

considered responsible for the lack of significant differences in antimicrobial efficacy between EndoVac irrigation and standard irrigation (35). c. Smear Layer removal The smear layer is created when the dentinal walls of the root canal system interact with endodontic instruments (92). The smear layer is comprised of inorganic and organic material such as dentin filings and pulp tissue remnants (93). This deposit can be penetrated by bacteria and may offer protection to biofilms adhering to the root canal walls (94). Furthermore, the smear layer interferes with the tight adaptation of currently used root canal sealers to dentin walls and may therefore promote micro- leakage (95). Torabinejad et al. (96) suggested that removal of the smear layer would decrease bacteria and improve adaptation of root filling materials to the canal walls. Another study (97) showed that the smear layer produced during root canal preparation promoted adhesion and colonization of P. nigrescens to the dentin matrix and might increase the likelihood of canal reinfection. Removing the smear layer reduces the potential for microleakage (77,98) and improves sealer penetration in dentinal tubules (99). When the manufacturer s recommendations are followed, the EndoVac system delivers a sufficient volume of irrigation to enable removal of the smear layer (77,100,101). Compared to passive ultrasonic irrigation, negative apical pressure irrigation and manual dynamic irrigation are more efficient in removing the smear layer in the apical one third (101). A possible explanation for this is that both techniques reach full working length of instrumented canals, eliminate the apical vapour lock at the apex and hence allow adequate 24

solution replacement (100,101). When evaluating irrigation of the apical one third, the phenomenon of apical vapour lock should be considered (55,102,103). d. Calcium hydroxide removal As stated previously, the debridement of the root canal system consists of elimination of organic, inorganic and microbial components, thus accomplished by mechanical instrumentation supported by various irrigation regimens and placement of intracanal medication. Calcium hydroxide (Ca(OH)2) is a commonly used intracanal medication (104) that has antimicrobial activity proven to contribute to bacterial endotoxin neutralization (105) and to periapical repair(106). However, to provide a maximum interface between the root canal walls and the filling material, Ca(OH)2 has to be removed (107) otherwise the bond strength (108) of the sealer and its penetration into the dentine tubules could be reduced (109). Conventional irrigation methods have demonstrated limited capacity to remove Ca(OH)2 from the apical third of the root canal (110). A scanning electron microscopic evaluation of longitudinally sectioned canines demonstrated that EndoVac system performs better than the traditional syringe irrigation in removing Ca(OH)2 from the apical one third of root canals (111). The results were similar to another study(112) where EndoVac system was compared to the traditional syringe irrigation and the ProUltra Piezoflow TM ultrasonic irrigation needle (Dentsply Tulsa, Tulsa, OK, USA). The EndoVac system left significantly less calcium hydroxide compared to the traditional syringe irrigation and were found to be as efficient as the PiezoFlow TM in removing Ca(OH)2 (112). Although the EndoVac system improves the removal of Ca(OH)2, the apical portion of the canal was not completely free of intracanal medication. Therefore, the use of the master 25

apical file in combination with the EndoVac system may result in better removal of Ca(OH)2 (112). e. Safety As stated previously, with traditional root canal irrigation, clinicians must be careful when determining how far an irrigation needle is placed into the canal. Recommendations for avoiding NaOCl accidents include not binding the needle in the canal, not placing the needle close to working length, and using a gentle flow rate when using positive pressure irrigation (39). In contrast, the EndoVac system pulls irrigation solution into the canal to working length and the solution and debris are removed by negative pressure. Negative apical pressure has been shown to enable irrigation solutions to safely reach the apical one third and help overcome apical vapour lock (70,78). Apart from being able to avoid air entrapment, the EndoVac system is also advantageous in its ability to deliver irrigation solution safely to working length without causing their undue extrusion into the periapex (70,71), thereby avoiding NaOCl accidents. It is important to note that it is possible to create positive pressure in the pulp canal if the MDT is misused, which would create the risk of a NaOCl accident. The manufacturer s instructions must be followed for correct use of the MTD by never directing towards the orifice of a canal. In order to compare the safety of six current intra- canal irrigation delivery devices, an in vitro test was conducted using the worst- case scenario of apical extrusion, with neutral atmospheric pressure and an open apex (71). The study concluded that the EndoVac system 26

did not extrude irrigation solution even after deep intra- canal delivery and suctioning of the solution from the chamber to full working length, whereas other devices did. The EndoActivator extruded only a very small volume of solution, the clinical significance of which is not known. Mitchell and Baumgartner (73) tested NaOCl extrusion from a root canal sealed with a permeable agarose gel. Significantly less extrusion occurred using the EndoVac system compared with positive- pressure needle irrigation. A well- controlled study by Gondim et al. (32) found that patients experienced less post- operative pain, measured objectively and subjectively, when apical negative- pressure irrigation was performed (EndoVac system) than with apical positive- pressure irrigation (32). Furthermore, PiezoFlow TM shows a greater potential to cause apical extrusion compared with EndoVac system s safety. When positioned within the last five millimetres of the root canal, the ultrasonic activated needle could cause apical extrusion of irrigation solution (67). 3. GentleWave System Recently, a novel system to clean the root canal was developed. The GentleWave system (Sonendo, Laguna Hills, CA) is composed of a console and a handpiece (Figure 4). The console includes several electrical circuits, three irrigation solution containers, one waste canister, a degassing system and a pressure generator. GentleWave continuously delivers energized treatment solution throughout the root canal system via a handpiece positioned on an accessed occlusal tooth surface (113,114). A built- in impingement plate disrupts the 27

irrigation stream redirecting the resulting mist to flow across the pulp chamber and over canal orifices. When the liquid interacts with the stationary liquid inside the pulp chamber it generates a shear stress strong enough to induce acoustic streaming and cavitation. The sound waves then reverberate against the dentinal wall and the energy is propagated apically. The GentleWave handpiece collects excess of fluid from the canals via its three- point vented suction and removes the outflowing fluid creating negative pressure within the root canal system. The fluid motion induced also generates fluid motion such as vortices and swirls. Three different irrigation solutions can be charged in the system. The manufacturer suggests the combination sodium hypochlorite and ethylenediaminetetraacetic acid (EDTA) with a rinse of distilled water between each of them to avoid potential damaging chemical reaction. Each irrigation solution is degassed before being delivered by the hand piece. The degassing process eliminated air in the liquid, therefore potentially decreases the occurrence of a phenomenon called vapour lock. a. GentleWave characteristics GentleWave s manufacturer suggests that their novel technology avoids the creation of vapour lock and promotes a constant exchange of solution of the entire length of the root canal. According to their previous in house experimentations, canals need not be enlarged beyond ISO size 15 for the GentleWave system to work effectively. 28

b. Tissue dissolution One study reported the superiority of GentleWave in dissolving vital tissue when compared to passive ultrasonic activation, EndoVac or a positive- pressure conventional needle coupled to a syringe for irrigation (113). This study has several limitations and the conclusion should be interpreted with caution. c. Calcium hydroxide removal According to Ma et al. (114), the GentleWave system has the capacity to remove calcium hydroxide along the entire canal length and inside root canal complexities. Mesial and distal canals of 30 mandibular molars were prepared with the WaveOne Primary (25/.08) and WaveOne Large (40/.08) instruments (Dentsply Tulsa Dental Specialties, Tulsa, OK), respectively. All canals were then filled with Ca(OH)2. The teeth were divided into the following 3 irrigation protocols: (1) needle irrigation, (2) passive ultrasonic activation, and (3) GentleWave system. The irrigation time in each group was 7.5 minutes. To further test the efficiency of the GentleWave system, shorter times of 90 seconds were used. Reconstructed micro- computed tomographic scans measured the volume of the canals and Ca(OH)2 after instrumentation, after filling of Ca(OH)2, and after its removal. The percentage of Ca(OH)2 remaining in the canals was calculated. None of the 10 teeth in the conventional irrigation and passive ultrasonic activation groups were completely cleaned of Ca(OH)2 and the GentleWave system removed significantly more Ca(OH)2 with 100% and 98.78% in the mesial and distal canals, respectively. Additional experiments in 10 teeth revealed that the GentleWave system removed 99.85% and 99.97% of Ca(OH)2 within 90 seconds in the mesial and distal canals, respectively. 29

30

Rationale Because the GentleWave system delivers energized irrigation solution, including NaOCl, the risk of periapical extrusion must be considered and assessed as a measure of safety in clinical usage. Aim This study assessed apical extrusion during treatment with GentleWaveTM (GW), conventional open- ended 30G needle (CN) or EndoVac (EV) in root canals enlarged to different dimensions with and without apical constriction. Hypothesis The null hypothesis was that incidence and mass of extrusion would not differ significantly between groups GW and CN for all instrumentation subgroups. 31

JOE Article 32

Basic Research Technology 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 ½Q9Š ½Q1Š Assessment of Apical Extrusion during Root Canal Irrigation with the Novel GentleWave System in a Simulated Apical Environment Karine Charara, DMD, Shimon Friedman, DMD, Adria Sherman, MSc, Anil Kishen, BDS, MDS, PhD, Gevik Malkhassian, DDS, MSc, Mehrzad Khakpour, PhD, and Bettina Basrani, DDS, PhD Abstract Introduction: This study assessed apical extrusion during treatment with GentleWave (GW; Sonendo, Laguna Hills, CA), a conventional open-ended 30-G needle (CN), or Endovac (EV; SybronEndo, Orange, CA) in root canals enlarged to different dimensions with and without apical constriction. Methods: Sixteen mandibular molars were mounted in an in vitro apparatus. Roots were immersed in a pressure-regulated chamber containing distilled water with pressure kept at 5.88 0.15 mm Hg to simulate periapical back pressure. Mesiobuccal (curved #30 ) and distal (straight) canals were instrumented to the working length (WL) as follows: minimal instrumentation (MI, #15/.04), traditional instrumentation (#35/.06), or overinstrumentation (OI, #35/.06, to the WL + 1 mm). Canals were tested 5 times each with distilled water using GW, CN (at WL-3 mm), or EV and the mass (g) of extruded water recorded. Extrusion frequency and mean extruded mass were compared for each canal, irrigation group, and canal instrumentation mode (Wilcoxon t test, P <.05). Results: No extrusion occurred with GW and EV, whereas the frequency of extrusion with CN was 33%. Mean extruded water mass using CN ranged in mesial canals from 0.000 0.000 g (OI) to 0.047 0.098 g (MI) and in distal canals from 0.123 0.191 g (MI) to 0.505 0.490 g (OI). With traditional instrumentation and OI instrumentation, extruded mass in distal canals was significantly higher than in mesial canals (P <.002) and distal canals with MI (P <.020). Conclusions: Within this study s limitations, root canal treatment with GW and irrigation with EV was not associated with extrusion. Extruded irrigation mass using the open-ended 30-G needle depended on the canal type and enlargement. These results have to be interpreted with caution, and further investigations are warranted to evaluate the possibility of extrusion using GW in different tooth types and clinical situations. (J Endod 2015;-:1 6) Key Words Apical extrusion, apical foramen, Endovac, GentleWave, hypochlorite accidents, needle irrigation Debridement of necrotic pulp tissue and bacterial elimination are critical procedural goals when treating infected root canals (1). Complex root canal anatomy (2) frequently makes it impossible to achieve these goals by instrumentation alone (3) and necessitates the adjunctive use of root canal irrigation with topical antiseptics. Currently, the irrigation solution of choice is sodium hypochlorite (NaOCl), which is commonly used in concentrations of 0.25% 6.0% (2). Although NaOCl dissolves necrotic tissue and is bactericidal, it requires frequent replenishment to be effective, and it can cause tissue damage if extruded periapically during irrigation (2, 4). Optimizing the delivery of NaOCl throughout the root canal by continuous replenishment while avoiding periapical extrusion has been a subject of research and development in recent years (5 9). NaOCl is conventionally delivered via a positive-pressure syringe fitted with a needle, a variety of which are available (3). An open-ended 30-G needle is commonly used, which has, on occasion, resulted in periapical extrusion of NaOCl (10) and consequent cytotoxic effects (11 13) clinically manifested by pain, localized, or widespread swelling and ecchymosis (12). Despite its drawbacks (9, 14, 15), positive-pressure irrigation remains the most commonly used method for root canal irrigation (11, 12, 15 17). The negative-pressure irrigation device Endovac (EV; SybronEndo, Orange, CA) uses a suction cannula inserted to the canal working length (WL) to deliver irrigation solutions throughout the canal length (18) while avoiding extrusion (8, 9). For proper use, the EV requires canals to have a minimal width of 350 mm at the WL. The novel GentleWave (GW) system (Sonendo, Laguna Hills, CA) continuously delivers energized treatment solution throughout the root canal system via a handpiece positioned on an accessed occlusal tooth surface (19, 20). The handpiece delivers a stream of treatment fluid into the pulp chamber, and the built-in suction removes the outflowing fluid creating negative pressure within the root canal system. The manufacturer suggests that canals need not be enlarged beyond ISO size 15 for the GW system to work effectively. Because the GW system delivers energized irrigation solution, From the University of Toronto, Faculty of Dentistry, Toronto, Ontario, Canada. Address requests for reprints to Dr Karine Charara, University of Toronto, Faculty of Dentistry, 124 Edward Street, #348C, Toronto, Ontario, Canada M5G 1G6. E-mail address: kcharara@me.com 0099-2399/$ - see front matter Copyright ª 2015 American Association of Endodontists. http://dx.doi.org/10.1016/j.joen.2015.04.009 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78 79 80 81 82 83 84 85 86 87 88 89 90 91 92 93 94 95 96 97 98 99 100 101 102 103 104 105 106 JOE Volume -, Number -, - 2015 Apical Extrusion during Root Canal Irrigation 1 FLA 5.2.0 DTD Š JOEN3131_proof Š 8 May 2015 Š 6:08 am Š ce BT 33

107 108 109 110 111 112 113 114 115 116 117 118 119 120 121 122 123 124 125 126 127 128 129 130 131 132 133 134 135 136 137 138 139 140 141 142 143 144 145 146 147 148 149 150 151 152 153 Basic Research Technology including NaOCl, the risk of periapical extrusion must be considered and assessed as a measure of safety in clinical usage. Therefore, this study evaluated the extrusion of solution during use of the GW system in root canals enlarged to different dimensions with and without preservation of the apical constriction. Materials and Methods Specimens The University of Toronto Research Ethics Board approved the study protocol (#29320). To estimate the sample size, the mass of extruded solution using an open-ended needle was determined in a pilot study (not reported) with a setup similar to that described later. The difference between the extruded irrigation mass and 0 (assumed for GW) suggested that 12 canals/group would support analysis with 80% power and alpha of 5%. To accommodate variation of data, a repeated measures design was used, and the sample was increased to 16 canals/group, which was consistent with that reported in previous studies on apical extrusion (21, 22). Extracted human mandibular molars with fully developed roots; a pulp chamber height of $2 mm; and no root caries, cracks, fractures, or internal and external resorption were stored in 4 C phosphatebuffered saline until use (23). Micro computed tomographic imaging was performed to standardize the samples and identify 16 molars with 2 independent and moderately curved (23 36 ) mesial canals and a single relatively straight (<20 ) distal canal (24). The pulp chamber was accessed conventionally, and the mesiobuccal and distal canals were located. The WL for each canal was established 1 mm short of the emergence of a size 10 K-type file (Dentsply Maillefer, Ballaigues, Switzerland) through the major foramen. A glide path was prepared with size 13 PathFile instruments (Dentsply Maillefer), and the canals were enlarged with Vortex instruments (Dentsply Tulsa Dental Specialties, Tulsa, OK) and irrigated with distilled water with a 30-G open needle in 3 successive steps: 1. Minimal instrumentation (MI): size 15/.04 Vortex instruments were used to the WL, and apical patency was confirmed 1 mm longer with size 10 K-type files. 2. Traditional instrumentation (TI): Vortex instruments were used sequentially to size 35/.06 at the WL, and apical patency was confirmed 1 mm longer with size 10 K-type files. 3. Overinstrumentation (OI): Vortex instruments were used to size 35/.06 to 1 mm beyond the WL. In addition to simulating inadvertent OI, this step also simulated canals with large apical foramina. Apical Extrusion Apparatus and Measurement After each canal instrumentation step (MI, TI, and OI), 1 tooth specimen at a time was mounted in an airtight chamber filled with 154 155½F1Š distilled water (Fig. 1A and B). To enable independent extrusion measurements for each canal, all other apical foramina were sealed with hot 156 157 glue. The tooth crown remained exposed to apply different devices, 158 whereas the roots were submerged in the water-filled chamber. 159 The airtight chamber was connected to a custom pressure vessel 160 by a 12 VDC precision solenoid valve (01540-01; Cole-Palmer, Vernon 161 Hills, IL). Pressure within the vessel was maintained at 5.88 0.15 mm 162 Hg to simulate periapical back pressure (25 27). A pressure generator 163 (SP600EC-HR-L; Schwarzer, Essen, Germany), a pressure valve (SS- 164 SS2-VH; Swagelok, Solon, OH), and a transducer (PXM409-350HCGI; 165 Omega, Stamford, CT) were used to generate and verify constant 166 pressure. 167 During irrigation, any pressure generated at the canal terminus 168 was transferred throughout the distilled water acting as an incompressible Newtonian fluid medium (density r = 998.2 kg/m 3 ; constant viscosity m = 1.0 U 10 3 kg/ms 1 ). When pressure exceeded 5.88 0.15 mm Hg, the solenoid valve opened allowing extrusion of solution through the apical foramen, causing water displacement from the chamber into the pressure vessel. A load cell (FSH02536; Futek, Irvine, CA) mounted within the pressure vessel weighed the displaced water (g). Displaced water mass data were recorded by a linked computer equipped with a Data-Acquisition DAQ Board (USB6008; National Instruments Corp, Austin, TX) and LabView software (National Instruments Corp). Pressure readings obtained from the experimental apparatus were calibrated with the hydrostatic pressures determined for 5 different water columns in a 10-mL pipette attached to a water-filled chamber. The coefficient of determination (R 2 ) between the theoretic apical pressure and the measured apical pressure was 0.9997. Groups To avoid NaOCl damage to the experimental apparatus, canals were irrigated with distilled water. A pilot study (not reported) confirmed that extruded masses were similar for water and NaOCl. Teeth (N = 16) were randomly assigned to different pre-established sequences of the following 3 groups and positive control: 1. GW: the delivery tip was positioned in the center of the access cavity 1 2 mm coronal to the pulpal floor, strictly avoiding placement at canal orifices and contact with cavity walls. Treatment fluid was dispensed for 30 seconds at 45 ml/min. 2. Conventional needle (CN): a 30-G conventional open-ended needle (Navitip; Ultradent, South Jordan, UT) connected to a peristaltic pump (New Era Pump, Farmingdale, NY) was inserted 3 mm short of the WL or slightly coronal to that point if it bound in the canal. Irrigation was dispensed for 30 seconds at 5 ml/min. The CN group was used to ascertain the ability to record extrusion with the needle kept within the root canal as previously reported (5, 9, 10, 21, 23). 3. EV: the microcannula was inserted to the WL. The master delivery tip was connected to the peristaltic pump and irrigation dispensed for 30 seconds at 5 ml/min while moving the cannula from the crown to apex at a low amplitude every 6 seconds (23). The EV group was considered as the negative control based on previous reports of no extrusion during its use (8, 9). It could not be used in canals with MI. 4. Positive control (n = 9): a 30-G open-ended needle attached to the peristaltic pump set at a flow rate of 5 ml/min was inserted 2 mm beyond the apical foramen, and the access cavity was sealed with vinyl polysiloxane impression material. Apical extrusion was determined for a period of 30 seconds. Validation Experiment Because the manufacturer of GW recommends continuous application of NaOCl for 5 minutes, an experiment was performed to ensure that irrigation extrusion over a 30-second period did not underestimate the risk of extrusion during irrigation for 5 minutes. Canals of 8 teeth with OI were irrigated with GW for 5 minutes. The data were compared with that of the GW group. Analysis To compensate for the systematic error of the experimental setup (0.2 g), all negative extrusion values were replaced with 0, and 0.2 g was subtracted from all positive values. Data were analyzed with Graph- Pad Prism (GraphPad Software Inc, La Jolla, CA,). Extrusion occurrence and mass (g) were recorded 5 times/canal for all groups and the incidence and mean mass calculated for each canal and group. Groups were compared with the Wilcoxon t test (P <.05). Because the ½Q2Š ½Q3Š 169 170 171 172 173 174 175 176 177 178 179 180 181 182 183 184 185 186 187 188 189 190 191 192 193 194 195 196 197 198 199 200 201 202 203 204 205 206 207 208 209 210 211 212 213 214 215 216 217 218 219 220 221 222 223 224 225 226 227 228 229 230 2 Charara et al. JOE Volume -, Number -, - 2015 FLA 5.2.0 DTD Š JOEN3131_proof Š 8 May 2015 Š 6:08 am Š ce BT 34

Basic Research Technology 231 232 233 234 235 236 237 238 239 240 241 242 243 244 245 246 247 248 249 250 251 252 253 254 255 256 257 258 259 260 261 262 263 264 265 266 267 268 269 270 271 272 273 274 275 276 277 278 279 280 281 282 283 284 285 286 print & web 4C=FPO ½Q8Š Figure 1. (A and B) The apical extrusion measurement apparatus. (A) The tooth is mounted on a customized cap and secured into an airtight chamber filled with incompressible fluid (B). If the periapical pressure, detected by pressure transducer (C), is greater than the simulated periapical back pressure, fluid passes through the extrusion line (D), which consists of a 12 VDC precision solenoid valve. Extruded fluid is then collected onto a glass dish (E) positioned on a load cell. The load cell is placed inside a pressure vessel (F). The pressure vessel is pressurized via a pressure generator (G) that simulates the periapical back pressure. The pressure within the vessel is measured by another pressure transducer (H) and is regulated by a pressure valve (I). extrusion mass data were nonparametric, a correlation of extrusion and canal size was explored with Spearman rho coefficients (P >.4 represents a strong correlation). The null hypothesis was that incidence and mass of extrusion would not differ significantly between the GW and CN groups for MI, TI, and OI. Results Extrusion occurred in all 9 positive controls (100%) and did not occur in any of the canals in the EV group (negative control), validating the experimental model. In the GW group, no extrusion occurred 287 288½T1Š throughout the experiments (0%). In the CN group (Table 1), extrusion 289 incidence varied among MI, TI, and OI groups and between mesial and 290 distal canals. Overall, extrusion incidence was higher in distal canals 291 (27/48, 47%) than in mesial canals (5/48, 10%). In distal canals, 292 the highest extrusion incidence was recorded for TI and OI (10/16, 63%), and in mesial canals, the highest extrusion incidence was recorded for MI (4/16, 15%). In the CN group, the mass of extruded solution across all specimens ranged from 0.000 1.373 g. In mesial canals, the mean water mass ranged from 0.000 0.000 g (OI) to 0.047 0.098 g (MI), and it did not differ significantly with the GW and EV groups (P >.125). In distal canals, the mean extruded water mass ranged from 0.123 0.191 g (MI) to 0.505 0.490 g (OI) and was significantly higher than in the GW and EV groups in MI, TI, and OI (P <.015). Comparing the 3 instrumentation modes, the mean extruded water mass in distal canals was significantly higher in TI and OI than in MI (P <.018). In TI and OI, the extruded water mass was significantly higher in distal canals than in mesial canals (P <.002). The application of GW for 5 minutes yielded 0 apical extrusion, validating the results obtained during the 30-second activation. All the measurements made were within the error limits of the experimental setup (data not reported). 293 294 295 296 297 298 299 300 301 302 303 304 305 306 307 308 309 310 311 312 313 314 315 316 317 318 319 320 321 322 323 324 325 326 327 328 329 330 331 332 333 334 335 336 337 338 339 340 341 342 343 344 345 346 347 348 349 350 351 352 353 354 JOE Volume -, Number -, - 2015 Apical Extrusion during Root Canal Irrigation 3 FLA 5.2.0 DTD Š JOEN3131_proof Š 8 May 2015 Š 6:08 am Š ce BT 35

355 356 357 358 359 360 361 362 363 364 365 366 367 368 369 370 371 372 373 374 375 376 377 378 379 380 381 382 383 384 385 386 387 388 389 390 391 392 393 394 395 396 397 398 399 400 401 402 403 404 405 406 407 408 409 410 411 412 413 414 415 416 Basic Research Technology TABLE 1. Mean Extruded Irrigation Mass (g) in Each of 16 Specimens Tested, Having Canals Instrumented Consecutively with 3 modes (MI, TI, and OI) and Irrigated with an Open-ended 30-G Needle Inserted 3 mm Short of the Working Length Tooth no. Discussion Serious NaOCl accidents during endodontic treatment have been rare and mostly occurred during syringe-delivered irrigation using open-ended needles (16, 28). Over the years, needle designs have been modified and negative pressure delivery systems introduced to enhance irrigation delivery throughout the canal volume while also preventing inadvertent extrusion (10, 22, 23, 29, 30). GW is a novel root canal cleaning system shown to deliver treatment fluid to the canal terminus, as shown by its ability to remove calcium hydroxide throughout the length of molar canals instrumented to size 25/.08 and 40/.08 (20). Considering that the GW system uses NaOCl for treatment, it was deemed necessary to assess the risk of periapical extrusion during its use in a variety of simulated clinical conditions. The experimental apparatus was developed to mimic the resistance to extrusion afforded by periapical tissue. Because the exact apical pressure that might result in a NaOCl accident is not known (31), thesafetylimitinthisstudywas set to not exceed the central venous pressure of 5.88 mm Hg (25), suggestedtopreventthe occurrence of intravenous accidents (25). In maxillary molars and incisors, reduced apical resistance may be affected by bone porosity, fenestrations, anatomic variationssuchasdrainageofa tooth by the facial vein (12), orlocationofrootapiceswithin the maxillary sinus without bone or Schneiderian membrane cover (32). In these situations, there may be an increased risk of NaOCl accidents (12). Becauseofthisconcern,themanufacturerofthe GW system considers its use contraindicated in maxillary teeth contacting the sinus. Keeping the roots in pressurized medium provided a quantitative assessment of extrusion, which could occur only when irrigation pressure at the apical foramen exceeded the set back pressure (10, 22). This model offered a lower risk of overestimating the amount of apical extrusion than previously reported when an airfilled vial was used (10, 22, 30). Thesameairtightchamberand pressure transducer were previously used to measure the apical pressure generated in canals of mandibularmolarsbydifferent needle designs (31). Mean extruded irrigation mass (g) per canal in each specimen Minimal instrumentation (MI) Traditional instrumentation (TI) Over-instrumentation (OI) Mesial canal Distal canal Mesial canal Distal canal Mesial canal Distal canal 1 0.139 0.000 0.000 0.000 0.000 0.957 2 0.000 0.573 0.000 0.754 0.000 0.000 3 0.000 0.225 0.000 0.281 0.000 0.682 4 0.000 0.000 0.000 0.000 0.000 0.359 5 0.000 0.537 0.000 0.966 0.000 1.132 6 0.045 0.000 0.000 0.000 0.000 0.000 7 0.255 0.124 0.000 0.149 0.000 1.164 8 0.000 0.360 0.000 0.728 0.000 0.139 9 0.000 0.000 0.000 0.000 0.000 0.000 10 0.000 0.000 0.000 0.000 0.000 0.000 11 0.305 0.000 0.338 0.590 0.000 1.041 12 0.000 0.022 0.000 0.009 0.000 0.726 13 0.000 0.000 0.000 0.253 0.000 0.505 14 0.000 0.130 0.000 0.105 0.000 0.000 15 0.000 0.000 0.000 0.000 0.000 0.000 16 0.000 0.000 0.000 0.468 0.000 1.373 Mean mass (SD) 0.047 (0.098) 0.123 (0.191) 0.021 (0.084) 0.269 (0.318) 0.000 (0.000) 0.505 (0.490) SD, standard deviation. Each canal was tested 5 times. Molars were used for the study primarily because the GW system had only a molar handpiece available at the time of the study. The selection of mandibular molars afforded 3 canals per tooth, including the moderately curved mesial canals and relatively straight distal canals. The same 16 teeth were used in random sequence for the 3 irrigation groups to avoid impacts of varying canal anatomy on extrusion, as previously reported (22). To simulate different clinical situations, 3 modes of instrumentation were tested, representing minimally invasive preparation (MI), conventional preparation (TI), and the absence of apical constriction (OI) as might occur in overinstrumented canals and externally resorbed and immature roots. For the negative control, it was deemed appropriate to use the negative-pressure EV system because it has been shown to not cause extrusion during use (5, 8, 9, 23). Irrigation with a needle extending beyond the apex was the positive control. Consistent extrusion in the positive control and none with EV validated the experimental model. In addition, irrigation with an open-ended needle positioned 3 mm short of the WL, shown to cause minimal extrusion (10), was also tested to ascertain that the experimental apparatus was adequately sensitive to detect such minimal amounts of extrusion. The absence of extrusion with the use of GW and EV systems suggested that both devices generated apical pressures during use that did not exceed 5.88 0.15 mm Hg. However, syringe irrigation with an open-ended needle did cause significant extrusion of irrigation solution in distal canals, partially rejecting the null hypothesis. The amount of extruded irrigation solution in distal canals was positively correlated with the extent of canal instrumentation, corroborating suggestions in previous studies (20 22). The GW system is composed of a console and a sterile disposable handpiece. It delivers a stream of treatment solution from the tip of the handpiece into the pulp chamber while excess fluid is simultaneously removed from the chamber by the built-in vented suction through the handpiece into a waste canister inside the console. According to the manufacturer, upon initiation of flow through the treatment tip of the handpiece, the stream of the treatment fluid interacts with the stationary fluid inside the pulp chamber creating a strong shear force, which causes hydrodynamic ½Q4Š 417 418 419 420 421 422 423 424 425 426 427 428 429 430 431 432 433 434 435 436 437 438 439 440 441 442 443 444 445 446 447 448 449 450 451 452 453 454 455 456 457 458 459 460 461 462 463 464 465 466 467 468 469 470 471 472 473 474 475 476 477 478 4 Charara et al. JOE Volume -, Number -, - 2015 FLA 5.2.0 DTD Š JOEN3131_proof Š 8 May 2015 Š 6:08 am Š ce BT 36

479 480 481 482 483 484 485 486 487 488 489 490 491 492 493 494 495 496 497 498 499 500 501 502 503 504 505 506 507 508 509 510 511 512 513 514 515 516 517 518 519 520 521 522 523 524 525 526 527 528 529 530 531 532 533 534 535 536 537 538 539 540 ½Q5Š cavitation in the form of a cavitation cloud.thecontinuousformation and implosion of thousands of microbubbles inside the cavitation cloud generate an acoustic field with broadband frequency spectrum that travels through the fluid into the entire root canal system. Throughout the treatment, the fluid starts with 3% NaOCl and changes to 8% EDTA with a water rinse in between. The treatment tip of the handpiece is designed to deflect the stream of treatment fluid in such a way to generate a flow over the orifices of the root canals. This flow induces gentle vortical flow as well as a slight negative pressure within the root canal system. The energy and the vortical flow dissipate as they travel apically into the root canal system. One reason to explain the energy dissipation would be the lack of hard tissue in the periapical space that is needed for the reflection of the sound waves. Furthermore, the positive pressure created by the periapical tissue when encountered by the negative pressure inside the root canal system would prevent apical extrusion. Independent studies to show the mechanism of action and the flow dynamics of the GW system are warranted. EV simultaneously delivers irrigation solution and draws it out of the canal close to the apical level, preventing it from expressing to the apical tissue. Extrusion has been tested in a variety of in vitro conditions with consistently negative results (5, 8, 9, 21, 23), as also shown herein. Syringe irrigation with a 30-G open-ended needle occasionally results in irrigation extrusion characterized by a broad range of volumes and large standard deviation values (30). Similar inconsistency observed herein could be attributed to variations in irrigation flow dynamics caused by differences in root canal anatomy and needle position, impacting apical pressure and resulting extrusion (10, 33). Moreover, involuntary operator movement leading to changes in needle position and angulation might also have contributed to extrusion (10, 31). This study was designed to answer a very specific research question, and its results should not be extrapolated to clinical application or different conditions than those tested herein. Extrusion of irrigation with the GW system was assessed as 1 measure of usage safety specifically in intact molars. As the GW system is further adapted for use in other tooth types, further akin assessments are warranted. Furthermore, the results of this study may not apply to teeth presenting with reduced apical resistance, proximity of the mental foramen, undiagnosed vertical root fracture, perforating resorptive defects, and carious lesions communicating with the pulp chamber. These conditions require specific evaluation before the assessment of GW safety can be considered complete. Conclusion Within the limitations of this in vitro study, GW and EV did not cause treatment fluid extrusion in the apparatus with an apical pressure threshold of 5.88 0.15 mm Hg. Occasional extrusion during irrigation with a 30-G conventional open-ended needle was noted and associated with canal size. These results have to be interpreted with caution, and the safety of GW needs to be investigated further. Acknowledgments The authors thank Dr Calvin Torneck for his insights into this article. This study was supported by grants from the American Association of Endodontists Foundation, the Canadian Academy of Endodontics Endowment Fund, and Sonendo Inc. Basic Research Technology The authors report that a financial affiliation exists for this study because one author is an employee of Sonendo Inc (Mehrzad Khakpour). The other authors deny any conflicts of interest related to this study. References 1. Cano V, Nair PN, Henry S, Vera J. Microbial status of apical root canal system of human mandibular first molars with primary apical periodontitis after one-visit endodontic treatment. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2005; 99:231 52. 2. Basrani B, Haapasalo M. Update on endodontic irrigating solutions. Endod Topics 2012;27:74 102. 3. Haapasalo M, Shen Y, Qian W, Gao Y. Irrigation in endodontics. Dent Clin North Am 2010;54:291 312. 4. Pashley EL, Birdsong NL, Bowman K, Pashley DH. Cytotoxic effects of NaOCl on vital tissue. J Endod 1985;11:30 3. ½Q6Š 5. Malentacca A, Uccioli U, Zangari D, et al. Efficacy and safety of various active irrigation devices when used with either positive or negative pressure: an in vitro study. J Endod 2012;38:1622 6. 6. Boutsioukis C, Lambrianidis T, Kastrinakis E, Bekiaroglou P. Measurement of pressure and flow rates during irrigation of a root canal ex vivo with three endodontic needles. Int Endod J 2007;40:504 13. 7. Boutsioukis C, Gogos C, Verhaagen B, et al. The effect of root canal taper on the irrigant flow: evaluation using an unsteady Computational Fluid Dynamics model. Int Endod J 2010;43:909 16. 8. Nielsen BA, Craig Baumgartner J. Comparison of the EndoVac system to needle irrigation of root canals. J Endod 2007;33:611 5. 9. Desai P, Himel V. Comparative safety of various intracanal irrigation systems. J Endod 2009;35:545 9. 10. Psimma Z, Boutsioukis C, Kastrinakis E, Vasiliadis L. Effect of needle insertion depth and root canal curvature on irrigant extrusion ex vivo.jendod2013;39: 521 4. 11. Motta MV, Chaves-Mendonca MA, Stirton CG, Cardozo HF. Accidental injection with sodium hypochlorite: report of a case. Int Endod J 2009;42:175 82. 12. Zhu W, Gyamfi J, Niu L, et al. Anatomy of sodium hypochlorite accidents involving facial ecchymosis a review. J Dent 2013;41:935 48. ½Q7Š 13. Pashley EL, Birdsong NL, Bowman K, Pashley DH. Cytotoxic effects of NaOCl on vital tissue. J Endod 1985;11:525 8. 14. Ehrich DG, Brian JD, Walker WA. Case report sodium hypochlorite accident: inadvertent injection into the maxillary sinus. J Endod 1993;19:180 2. 15. Markose G, Cotter CJ, Hislop WS. Facial atrophy following accidental subcutaneous extrusion of sodium hypochlorite. Br Dent J 2009;206:263 4. 16. Kleier DJ, Averbach RE, Mehdipour O. The sodium hypochlorite accident: experience of diplomates of the American Board of Endodontics. J Endod 2008;34: 1346 50. 17. Dutner J, Mines P, Anderson A. Irrigation trends among American Association of Endodontists members: a web-based survey. J Endod 2012;38:37 40. 18. De Gregorio C, Paranjpe A, Garcia A, et al. Efficacy of irrigation systems on penetration of sodium hypochlorite to working length and to simulated uninstrumented areas in oval shaped root canals. Int Endod J 2012;45:475 81. 19. Haapasalo M, Wang Z, Shen Y, et al. Tissue dissolution by a novel multisonic ultracleaning system and sodium hypochlorite. J Endod 2014;40:1178 81. 20. Ma J, Shen Y, Yang Y, et al. In vitro study of calcium hydroxide removal from mandibular molar root canals. J Endod 2015;41:553 8. 21. Mitchell RP, Baumgartner JC, Sedgley CM. Apical extrusion of sodium hypochlorite using different root canal irrigation systems. J Endod 2011;37:1677 81. 22. Psimma Z, Boutsioukis C, Vasiliadis L, Kastrinakis E. A new method for real-time quantification of irrigant extrusion during root canal irrigation ex vivo. Int Endod J 2013;46:619 31. 23. Mitchell RP, Yang SE, Baumgartner JC. Comparison of apical extrusion of NaOCl using the EndoVac or needle irrigation of root canals. J Endod 2010;36:338 41. 24. Schneider SW. A comparison of canal preparations in straight and curved root canals. Oral Surg Oral Med Oral Pathol 1971;32:271 5. 25. Goode N, Khan S, Eid AA, et al. Wall shear stress effects of different endodontic irrigation techniques and systems. J Dent 2013;41:636 41. 26. Lyons RH, Kennedy JA, Burwell CS. The measurement of venous pressure by the direct method. Am Heart J 1938;16:675 93. 27. Baumann UA, Marquis C, Stoupis C, et al. Estimation of central venous pressure by ultrasound. Resuscitation 2005;64:193 9. 541 542 543 544 545 546 547 548 549 550 551 552 553 554 555 556 557 558 559 560 561 562 563 564 565 566 567 568 569 570 571 572 573 574 575 576 577 578 579 580 581 582 583 584 585 586 587 588 589 590 591 592 593 594 595 596 597 598 599 600 601 602 JOE Volume -, Number -, - 2015 Apical Extrusion during Root Canal Irrigation 5 FLA 5.2.0 DTD Š JOEN3131_proof Š 8 May 2015 Š 6:08 am Š ce BT 37

Basic Research Technology 603 604 605 606 607 608 609 610 611 612 613 614 615 616 617 618 619 620 621 622 623 624 625 626 627 628 629 630 631 632 633 634 635 636 637 638 639 640 641 642 643 644 645 646 647 648 649 650 651 652 653 654 655 656 657 658 659 660 661 662 663 664 28. Mehdipour O, Kleier DJ, Averbach RE. Anatomy of sodium hypochlorite accidents. Compend Contin Educ Dent 2007;28:544 6. 548, 550. 29. Spoorthy E, Velmurugan N, Ballal S, Nandini S. Comparison of irrigant penetration up to working length and into simulated lateral canals using various irrigating techniques. Int Endod J 2013;46:815 22. 30. Brown DC, Moore BK, Brown CE, Newton CW. An in vitro study of apical extrusion of sodium hypochiorite during endodontic canal preparation. J Endod 1995;21:587 91. 31. Park E, Shen Y, Khakpour M, Haapasalo M. Apical pressure and extent of irrigant flow beyond the needle tip during positive-pressure irrigation in an in vitro root canal model. J Endod 2013;39:511 5. 32. Hauman CH, Chandler NP, Tong DC. Endodontic implications of the maxillary sinus: a review. Int Endod J 2002;35:127 41. 33. Khan S, Niu L, Eid A, et al. Periapical pressures developed by nonbinding irrigation needles at various irrigation delivery rates. J Endod 2013;39:529 33. 665 666 667 668 669 670 671 672 673 674 675 676 677 678 679 680 681 682 683 684 685 686 687 688 689 690 691 692 693 694 695 696 697 698 699 700 701 702 703 704 705 706 707 708 709 710 711 712 713 714 715 716 717 718 719 720 721 722 723 724 725 726 6 Charara et al. JOE Volume -, Number -, - 2015 FLA 5.2.0 DTD Š JOEN3131_proof Š 8 May 2015 Š 6:08 am Š ce BT 38

Statistics The purpose of the following section is to describe the various statistical analyses of the data acquired for this study. 1.1. Amount of Extrusion and Canal Size The mean mass of extruded irrigation in relation to canal size is summarized in Figures 7 and 8 for mesial and distal canals, respectively. Percent mass extruded is represented in Figures 9 and 10. In order to test the null hypothesis regarding differences between CN and GW in the mean mass of extruded irrigation, a Wilcoxon t- test was used as the data was paired (a result from the repeated measures study design) but normality could not be assumed. Different Wilcoxon t- tests were performed to compare the mesial and distal canals at different canal sizes. Results are summarized in Table 1. EV was not compared to GW because these devices both demonstrated zero extrusion. In the mesial canals tested, there was no significant difference in mean mass of extrusion for CN and GW for any canal size. In the distal canals tested, there was a significant difference in the mean mass of extrusion between CN and GW for all canal sizes. 1.2. Correlation of Extrusion Masses and Canal Size In order to determine the correlation between mass of extruded irrigation and canal size, Spearman s rho coefficients were calculated to measure the degree of dependence between 39

two variables. A Spearman s test (non- parametric) was used instead of a Pearson s test (parametric) because normality tests of the data (D Agostino & Pearson omnibus normality test, Shapiro Wilk normality test, and KS normality test) concluded that the extrusion mass was non- parametric. The larger the absolute value of the rho coefficient represents a stronger relationship. The sign of the rho coefficient indicates the direction of the relationship. Table 2 summarizes the coefficients for various comparisons and whether or not these rho coefficients were significant. In distal canals, CN showed a positive correlation between mass of extrusion and canal size. Thus, it was extrapolated that as canal size increased, the amount of extrusion increased as well. GW and EV were not associated with any extrusion of irrigation regardless of canal size. 1.3. CNI Extrusion masses for Mesial vs. Distal Canals Because CN was the only device that resulted in extrusion of irrigation from the apex, a test was done to see if there was a significant difference between type of canal and extrusion mass (Figure 11 a, b, c). A Wilcoxon test was performed due to the nonparametric and paired nature of the data set. 40

Use of CN resulted in greater extrusion in distal canals than in mesial canals. There was a significant difference in the amount of extruded irrigation by CN in mesial canals compared to distal canals for traditional instrumentation (TI) and over instrumentation (OI) (Figure 9b, c). 41

Discussion Serious NaOCl accidents during endodontic treatment have been rare and most occurred during syringe- delivered irrigation using open- ended needles (33). Over the years, needle designs have been modified and negative pressure delivery systems have been introduced to enhance irrigation delivery throughout the canal volume while also preventing inadvertent extrusion (63,72,73,81,115). GentleWave is a novel irrigating system shown in preliminary experiments using dye in transparent teeth to deliver irrigation to the level of the apical foramen (113). In a previous study aiming to access the ability of GentleWave to remove calcium hydroxide throughout the length of molar canals instrumented to size 25/.08 and 40/.08, it was demonstrated that the treatment fluid reaches to the canal terminus, as all the calcium hydroxide was removed (114). Considering that GentleWave utilizes NaOCl for irrigation, it was deemed necessary to assess the risk of periapical extrusion during its use in a variety of simulated clinical conditions. 1. Methodology 1.1. Experimental set- up Although different, the design of the experimental apparatus was inspired by Psimma et al. (116). Keeping the roots in pressurized medium provided a quantitative assessment of extrusion that could occur only when irrigation pressure at the apical foramen exceeded the predetermined back- pressure (116). Psimma et al. (116) had a set- up able to measure apical extrusion in a real- time manner. The teeth were placed into a chamber filled with 42

distilled water and extrusion was measured as the amount of electrolytes diffused in the water. Using a conductivity probe of high sensitivity has been proven to be accurate. With their system, an open valve allowed the quantification of apical extrusion in a compliant system, which aimed to mimic a clinical situation where a sinus tract is present. Keeping the valve closed decreased the compliance of the experimental set- up and significantly less extrusion was seen when compared the open valve measurements. Psimma et al. (116) did not control the periapical backpressure. Disregarding the backpressure produced by the periapical tissue by evaluating extrusion into vials filled with air possibly overestimates the extruded amount of irrigation (71,116). It also creates an obvious discrepancy with in vivo conditions because periapical tissues act as a barrier against irrigation extrusion. Another method suggested to quantify apical extrusion of irrigation is to embed teeth in an agarose gel which changes color when NaOCl contacts the gel (73,74). This method has a clear advantage over the previous one described; however, the quantification of extruded irrigation is difficult and the system is affected by chemical diffusion. In the present study, the experimental apparatus was developed to mimic the resistance to extrusion afforded by periapical tissues. The backpressure was constantly applied and controlled. Moreover, the quantification of extruded irrigation was not influenced by chemical diffusion. 43

1.2. Back pressure Because the exact apical pressure that might result in a NaOCl accident is not known (117), the safety limit in this study was set to not exceed the central venous pressure of 5.88 mmhg (83), suggested to prevent occurrence of intravenous accidents (83). The pressure detected in dogs periapical lesions also supports this threshold (118). In maxillary molars and incisors, reduced apical resistance may be effected by bone porosity, fenestrations, anatomic variations such as drainage of a tooth by the facial vein (22) or location of root apices within the maxillary sinus without covering bone or Schneiderian membrane (119). In these situations, there may be an increased risk of NaOCl accidents (22). Because of this concern, the manufacturer of the GentleWave system considers its use contraindicated in maxillary teeth contacting the sinus. Keeping the roots in pressurized medium provided a quantitative assessment of extrusion, which could occur only when irrigation pressure at the apical foramen exceeded the set back- pressure (116,120). This model offered a lower risk of overestimating the amount of apical extrusion than previously reported when an air- filled vial was used (116,120,121). The same air- tight chamber and pressure transducer were previously used to measure the apical pressure generated in canals of mandibular molars by different needle designs (117). 1.3. Tooth selection Molars were used for the study primarily because the GentleWave system had only a molar handpiece available at the time of the study. The selection of mandibular molars 44

afforded three canals per tooth, including the moderately curved mesial canals and relatively straight distal canals. The same 16 teeth were used in random sequence for the three irrigation groups to avoid impact of changes in root canal anatomy on extrusion, as previously reported (116). 1.4. Instrumentation levels Furthermore, to simulate different clinical situations, three modes of instrumentation were tested: MI represented a minimally- invasive preparation (MAF: #15/.02), TI represented conventional preparation (MAF: #35/.04) and OI represented absence of apical constriction as might occur in over- instrumented, externally resorbed and immature roots (MAF: #35/.04 WL +1mm). 1.5. Validation and control groups In addition, to test extrusion with the GentleWave system, it was deemed appropriate to use the negative- pressure EndoVac system as a negative control as it is reported to not cause extrusion during use (67,70,71,73,74). Irrigation with a needle extending beyond the apex was the positive control. Consistent extrusion in the positive control and none with EndoVac validated the experimental model. In addition, irrigation with an open- ended needle positioned 3 mm short of working length and shown to cause minimal extrusion (120), was also tested to ascertain that the experimental model was adequately sensitive to detect such minimal amounts of extrusion. The absence of extrusion with the use of GentleWave and EndoVac systems suggested 45

that both devices generated apical pressures during use that did not exceed 5.88 ± 0.15 mmhg. 1.6. Solution choice Distilled water was used for irrigation to avoid corrosion of the experimental apparatus by NaOCl. A pilot study confirmed that NaOCl and distilled water resulted in similar extrusion values (figure 6). 2. Results Syringe irrigation with an open- ended needle did cause significant extrusion of irrigation in distal canals, partially rejecting the null hypothesis. 2.1. Needle The amount of extruded irrigation in distal canals was positively correlated with the extent of canal instrumentation, in agreement with previous studies (20 22). Syringe irrigation with the 30 G open- ended needle has occasionally resulted in irrigation extrusion characterized by a broad range of volume and a large standard deviation (121). Such inconsistency was also seen in this study and could be attributed to variations in irrigation flow dynamics caused by differences in root canal anatomy and needle position, impacting on apical pressure and the risk of extrusion (120 122). Moreover, involuntary operator movement leading to changes in needle position and angulation, has also been reported as a 46

possible contributor to extrusion of irrigation (120). 2.2. GentleWave The GentleWave system is composed of a console and a sterile disposable handpiece. It delivers a stream of treatment solution from the tip of the handpiece into the pulp chamber while excess fluid is simultaneously removed from the chamber by the built- in vented suction through the handpiece into a waste canister inside the console. According to the manufacturer, upon initiation of flow through the treatment tip of the handpiece, the stream of the treatment fluid interacts with the stationary fluid inside the pulp chamber creating a strong shear force that causes hydrodynamic cavitation in the form of a cavitation cloud. The continuous formation and implosion of thousands of microbubbles inside the cavitation cloud generates an acoustic field with broadband frequency spectrum that travels through the fluid into the entire root canal system. Throughout the treatment, the fluid starts with 3% Sodium Hypochlorite and changes to 8% EDTA following a water rinse in between. The treatment tip of the handpiece is designed to deflect the stream of treatment fluid in such a way to generate a flow over the orifices of the root canals. This flow induces gentle vortical flow as well as a slight negative pressure within the root canal system. The energy and the vortical flow dissipate as they travel apically into the root canal system. One explanation for the energy dissipation would be the lack of hard tissue in the periapical space which is needed for the reflection of the sound waves. Further, the positive pressure 47

created by the periapical tissue when encountered by the negative pressure inside the root canal system would prevent apical extrusion. Independent studies to demonstrate the mechanism of action and the flow dynamics of the GentleWave system are warranted. 2.3. EndoVac EndoVac has been proven to be safe because it simultaneously delivers irrigation and draws it away from the canal and apical tissue. EndoVac has been tested under a multitude of in- vitro conditions and the results have consistently shown that no extrusion of irrigation occurred (67,70,71,73,74). Apart from being able to avoid air entrapment, the EndoVac system is also advantageous in its ability to deliver irrigation safely to working length without causing undue extrusion into the periapex (70,71), thereby avoiding NaOCl accidents. It is important to note that it is possible to create positive pressure in the pulp canal if the Master Delivery Tip is misused, which would create the risk of a NaOCl accident. The manufacturer s instructions must be followed for correct use of the Master Delivery Tip by never directing it towards the orifice of a canal. In order to compare the safety of six current intra- canal irrigation delivery devices, an in vitro test was conducted using the worst- case scenario of apical extrusion, with neutral atmospheric pressure and an open apex (71). The study (50) concluded that the EndoVac system did not extrude irrigation even after deep intra- canal delivery and suctioning of the irrigation from the chamber to full working length, whereas other tested devices did. The EndoActivator extruded only a very small volume of irrigation, the clinical significance of which cannot be speculated on. Mitchell and Baumgartner (73) tested irrigation (NaOCl) extrusion from a root canal sealed 48

with a permeable agarose gel. Significantly less extrusion occurred using the EndoVac system compared with positive- pressure needle irrigation (54). A well- controlled study by Gondim et al. (32) found that patients experienced less post- operative pain, measured objectively and subjectively, when apical negative- pressure irrigation was performed (EndoVac system) than with apical positive- pressure irrigation. Furthermore, PiezoFlow TM showed a greater potential to cause apical extrusion compared with EndoVac system s safety (112). When positioned within the last five millimetres of the root canal, the ultrasonic activated needle could cause apical extrusion of irrigation solution (67). This study was designed to answer a very specific research question and its results should not be extrapolated to clinical application or different conditions than those tested herein. Extrusion of irrigation with the GentleWave system was assessed as one measure of usage safety specifically in intact molars. As the GentleWave system is further adapted for use in other tooth types, further akin assessments are warranted. Furthermore, this study results might not apply to teeth presenting with reduced apical resistance, proximity of the mental foramen, undiagnosed vertical root fracture, perforating resorption defects and carious lesions communicating with the pulp chamber. These conditions require specific evaluation for safety of use of GentleWave. Furthermore, because clinical application of irrigation systems may be influenced by additional factors other than those that can be assessed in vitro, more comprehensive assessment of the safety of GentleWave in clinical use is warranted. 49

Conclusion Within the limitations of this in vitro study, GentleWave and EndoVac did not cause extrusion of irrigation in the apparatus with apical pressure threshold of 5.88 ± 0.15 mmhg. Occasional extrusion during irrigation with a 30G conventional open- ended needle was noted and associated with canal size. These results have to be interpreted with caution and the safety of GentleWave to be further investigated. 50

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Figures Figure 1 Clinical aspect of emphysema related to extravasation of sodium hypochlorite solution associated with endodontic treatment, with ecchymosis and severe swelling of the right side of the face. These symptoms appeared after a root canal treatment of the upper right canine (Reproduced with permission from Elsevier). 61

Figure 2 The components of the EndoVac system: the Master Delivery Tip (MDT) accommodates different sizes of syringes, the macrocannula is attached to the autoclavable aluminium handpiece and the microcannula is attached to an autoclavable aluminium fingerpiece. The macrocannula, the microcannula and the MDT are connected via clear plastic tubing. The tubes are connected to the high- volume suction of the dental chair via the multiport adaptor (Courtesy John Schoeffel). 62

Figure 3 Final irrigation protocol using EndoVac system 63

Figure 4 GentleWave system (Sonendo, Laguna Hills, CA) 64

Figure 5A and 5B Experimental apparatus A) The tooth is mounted on a customized cap and secured into an air- tight chamber filled with incompressible fluid (B). If the periapical pressure, detected by pressure transducer (C), is greater than the simulated periapical back pressure, fluid passes through the extrusion line (D), which consists of a 12 VDC precision solenoid valve. Extruded fluid is then collected onto a plexi glass dish (E) positioned on a load cell. The load cell is placed inside a pressure vessel (F). The pressure vessel is pressurized via a pressure generator (G) that simulates periapical back pressure. The pressure within the vessel is measured by another pressure transducer (H) and regulated by a pressure valve (I). 65

Figure 6 Pilot study comparing the apical extrusion of 0.5% NaOCl and water. 66