Optimization of the Laser Doppler Signal

original ARTICLES Optimization of the Laser Doppler Signal Acquisition Timing for Pulp Vitality Evaluation Mariana-Ioana Miron 1, Dorin Dodenciu 2, Mircea Calniceanu 1, Laura Maria Filip 1, Darinca Carmen Todea 1 REZUMAT Scop: S-a evaluat influenţa timpului de achiziţie asupra semnalelor laser Doppler la nivelul dinţilor maxilari anteriori. Complexitatea tehnicii de achiziţie presupune o poziţionare precisă şi constantă în timp, impunând minimalizarea mişcărilor involuntare ale pacientului. Scopul studiului a fost stabilirea intervalului optim al timpului de achiziţie şi analiza statistică a semnalului laser doppler, în care nivelul artefactelor de mişcare este minim. Material si Metodă: A fost utilizat un grup de 15 pacienţi, la care s-a determinat fluxul sanguin pulpar la nivelul frontalilor maxilari vitali. Semnalele au fost înregistrate cu ajutorul unui laser Doppler MoorLab, utilizând o probă optică, MP3b, poziţionată cu ajutorului unei amprente siliconice, perpendicular pe suprafaţa vestibulară a dintelui, în treimea cervicală. Pentru fiecare pacient s-a realizat o înregistrare de 3 minute, iar ulterior, fiecare traseu a fost divizat în intervale de 30 secunde. S-au obtinut astfel 6 intervale de timp de achiziţie a semnalului, începând de la punctul 0. A fost efectuată o analiză comparativă a mediilor si deviaţiilor standard, cu scopul analizării gradului de reproductibilitate şi încredere a celor 6 intervale luate in studiu. Rezultate: Conform analizei statistice, prin aplicarea testului Friedman si Wilcoxon signed-rank test, nu există diferenţe semnificative din punct de vedere statistic între primul minut de achiziţie si celelalte intervale considerate în studiu. Concluzii: Timpul de achiziţie a semnalului pentru testul de vitalitate pulpară poate şi trebuie să fie optimizat, iar studiul de faţă a arătat că timpul de achiziţie al semnalului laser Doppler poate fi redus la un minut, fără afectarea preciziei testării. Cuvinte cheie: timpul optim de achiziție, laser Doppler, fluxul sanguin pulpar ABSTRACT Objective: We evaluated the influence of acquisition time on the laser Doppler signals in vital upper anterior teeth. Due to the complexity of this technique, accurate signal acquisition requires a fixed and constant position, minimizing involuntary patient movement. The purpose of the study was to find optimal timing for signal acquisition and statistical data analysis, in which movement artifacts level are minimal. Materials and Method: In a group of 15 patients, pulp blood flow was determined in healthy maxillary anterior teeth. Signals were recorded with aid of a laser Doppler MoorLab and a straight optical probe, MP3b, held in place with a silicon impression, perpendicular on the vestibular surface of the tooth, in the cervical third. For each patient the signal was acquired during three minutes and subsequently processed: the signal was divided in six epochs of 30 seconds for each patient. Six acquisition intervals were thus obtained, starting from 0 point. The signal mean and standard deviations were analyzed and compared between these intervals in order to observe how reproducible and reliable a shorter acquisition can be. Results: Following statistical analysis, by applying the Friedman and Wilcoxon signed-rank test, there are no significant differences, from a statistical point of view, betweeen the first acquisition minute and the other intervals included in the study. Conclusions: The timing of the signal acquisition for a vitality test should and can be optimized, and this study concluded that acquisition time for laser Doppler signals could be reduced to a minute, without losing accuracy. Key words: optimal time of acquisition, laser Doppler flowmetry, pulpar blood flow. 1 Department of Oral Rehabilitation and Dental Emergencies, 2 Department of Medical Biophysics, Victor Babeş Medicine and Pharmacy University, Timisoara Correspondence to: Mariana-Ioana Miron Timisoara, 300627, Torontalului 15, sc. C, 2nd floor, ap. 7, Romania Phone: 0722644842 E-mail: mariana.miron@yahoo.co.uk Received for publication: Oct. 02, 2010. Revised: Nov. 07, 2009. 44 TMJ 2010, Vol. 60, No. 1 INTRODUCTION The diagnosis of dental pulp status is frequently given insufficient attention by many dentists. Vitality testing is an essential aid for dental pulp health status monitoring, especially after traumatic injuries and for the correct diagnosis concerning pulp disease and apical periodontitis. 1 Current routine methods include thermal stimulation, electrical or direct dentine stimulation, assessment of the integrity of the Aδ nerve fibers in the dentine-pulp complex by briefly applying the stimulus to the outer surface of the tooth and indicate that the

nerve fibers are functioning, but does not give any indication of blood flow within the pulp. These testing methods have the potential to produce an unpleasant and occasionally painful sensation and inaccurate results (false positive or negative can be obtained in many instances). In addition, each test is subjective, depending on the patient s perceived response to a stimulus as well as the dentist s interpretation of that response. 2 Many studies have shown that blood circulation and not innervation is the most accurate determinant in assessing pulp vitality, as it provides an objective differentiation between necrotic and vital pulp tissue. 3 In order to achieve an early diagnosis of pulpal inflammation, sensitive, specific and precise exploring methods are necessary for the detection of initial, pre-clinic signs of inflammation, thereby allowing rapid treatment with maximal chances of success. Consequently, medical research of international value is attempting to develop new, less invasive, precise and objective methods for the early detection of vascular inflammation, in order to permit assessment of the degree of pulpal deterioration by exploration of blood flow dynamics. Laser Doppler flowmetry is an established technique for the real-time measurement of microvascular red blood cell perfusion in tissue. Pulpal blood flow can now be measured non-invasively in the clinic by means of laser Doppler flowmetry. 4,5 The method is noninvasive, and the probe needs not actually touch the surface of the tissue. Laser Doppler signals from the tissue are recorded in Blood Perfusion Units, which is a relative, arbitrary units scale defined using a carefully controlled motility standard, comprising a suspension of latex spheres, polystyrene microspheres in water undergoing Brownian motion. Physical laser Doppler parameters analyzed in the study were Flow and DC. Flow is related to the product of average speed and concentration of moving red blood cells in the tissue sample volume. It is the parameter most widely reported in Laser Doppler publications. DC gives an indication of the backscattered laser light intensity. It can be used to check the efficiency of light collection by the laser Doppler probes. The validation criteria of the pulp vitality test achieved through the laser Doppler technique are related to the level of the flux signal and the presence of the pulsatile character of the acquired signal, synchronised with the cardiac frequency. The DC level indicates a correct positioning of the optical probe, showing the reflected laser radiation level from the level of the concerned area. The variations of this parameter are in direct correlation with the movements of the optical probe, this generating artifacts in the Flux recording also. Due to these facts, the DC signal is the one that indicates the mechanic stability of the optical probe placed at the level of the acquisition area. We evaluated the influence of acquisition time on the variability of laser Doppler signals in vital upper central incisors. Due to the complexity of this technique, accurate signal acquisition requires a fixed and constant position, minimizing involuntary patient movement. For the acquisition of laser Doppler signals, the acquisition time most frequently used in the specialty literature is of 3 minutes. 6, 7 Bearing in mind that a valid and correct acquisition requires a complex technique, which implies mainly, the precise positioning of the probe as well as a special disposition of the patient throughout the recording (i.e. relaxation, absence of any movement) in order to avoid movement artifacts, 3 minutes represents a long period of time. Given the situation, a legitimate question arises: what is the minimum acquisition interval of a laser Doppler signal, necessary for the establishment of a correct diagnostic regarding pulp vitality? The purpose of the study was to find optimal timing for signal acquisition and statistical data analysis, in which noise and disturbance levels are minimal. Materials and Method The study was carried out on 15 healthy, single rooted anterior teeth, corresponding to 15 patients (aged between 18 and 28). The study was performed at the Department of Oral Rehabilitation and Dental Emergencies, School of Dentistry, Victor Babes University of Medicine and Pharmacy, Timisoara. The in vivo study was carried out with the approval of the Local Ethics Committee and, after the presentation of the written protocol the patients signed their written participation agreement. All teeth were decay free and were either intact or had only a small restoration. Radiographic examination and electrical pulp stimulation confirmed that they were vital and healthy. Measurement of collected data took place using a MoorLab laser Doppler equipment and a straight optical probe, MP3b, 10 mm. To stabilize the probe, a silicon impression was used. This was fixed perpendicularly on the vestibular surface of the tooth, in the cervical third (Fig 1). The Moor Instruments MoorLab laser Doppler monitor uses laser radiation generated by a semi conductor laser diode operating at a wavelength Mariana-Ioana Miron et al 45

of 780+10nm and a maximum accessible power of 1.6mW. The programmed bandwidth of the recorded laser Doppler signal was 20 Hz-20 khz, while sampling frequency displayed a value of 40 Hz. Calibration took place according to the instructions of the manufacturer. movements. Figure 2. The acquisition technique of laser Doppler signals. Figure 1. The positioning of the laser probe. As materials, there were also used: silicon impression materials; rotary instruments for the preparation of the access canals; pulp test, a PC system for collecting and processing the data; insulation material. The data were processed using the statistical analysis software SPSS v11.0.1. The work technique On the patients first visit, after signing the participation agreement, an electrical pulp stimulation and radiographic examination of their teeth was made in order to confirm that they were vital and healthy. Then, for every patient involved in the study, a double silicon impression of the anterior maxillary teeth was made. With the help of a drill of 1,5 mm in diameter, the vestibular-oral impression was chiseled perpendicularly, in the cervical third of the tooth involved in the study. On the second visit, every patient sat comfortably on the dentist s chair; the ambient temperature was approximately 21 C, and the lamp from the dentist s unit was positioned at a distance of between 45 and 55 cm from the patient. Prior to being tested, the subjects were asked to rest in the dental chair for approximately 10 min. Then the impression was positioned and the laser Doppler probe was fixed by means of the tunnel. (Fig 2). The influence of acquisition time on the level and the variability of the laser Doppler signals registred at the level of the anterior maxillary teeth were assessed. One of the major objectives of the laser Doppler signal acquisition technique at the level of pulp chamber requires a precise and time-constant positioning of the optical probe, imposing minimum involuntary patient 46 TMJ 2010, Vol. 60, No. 1 After probe calibration, for each patient the signal was acquired during 3 minutes (7200 numerical values) and subsequently processed: the signal was divided into six epochs of 30 seconds for each patient. Thus 6 acquisition intervals were obtained starting from 0 point: 30 ; 1 ; 1 30 ; 2 ; 2 30 ; 3, for each acquired signal (Fig 3). Figure 3. The marking of the 6 intervals for each aquired signal researched in the study. Results and statistical analysis The acquisition of laser Doppler signals for 3 minutes allowed the recording of 7200 values for each acquired recording, all 15 of them being submitted to the descriptive statistical analysis (Table I). As a result of the analysis of the experimentally obtained values it can be observed that the DC parameter mean values are different in each case. This can be physically explained in that each tooth displays different individual optical characteristics and through the optical coupling appropriate for each case. Hence, the chosen parameter for the statistical analysis was the standard deviation that reflected the best the stability, in time, of the DC signal. In other words, the DC parameter was selected because it does not have a pulsatile character as the Flux parameter and it relates directly to the modification of the

optical probe position. Any modification of the DC shows the changing of the optical probe position in relation to the tooth, situation which brings about modifications of the Flux values. Therefore the ideal would be that the DC value should have a variability as low as possible. Table 1. Experimental results N of subject Time intervals N of values Mean Std. Deviation 1 30 1200 26.270.2531 1 2400 27.079.4587 1 30 3600 27.493.5563 2 4800 27.862.6259 2 30 6000 28.021.6545 3 7200 28.101.6618 2 30 1200 36.488.0842 1 2400 36.317.1388 1 30 3600 36.165.1673 2 4800 36.226.1587 2 30 6000 36.162.1703 3 7200 36.116.1717 3 30 1200 38.912.1143 1 2400 38.970.1236 1 30 3600 39.115.1508 2 4800 39.122.1477 2 30 6000 39.046.1386 3 7200 38.995.1357 4 30 1200 27.449.1088 1 2400 27.611.1767 1 30 3600 27.494.1568 2 4800 27.450.1530 2 30 6000 27.453.1437 3 7200 27.563.1407 5 30 1200 40.013.1388 1 2400 40.111.1267 1 30 3600 40.054.1215 2 4800 40.206.1402 2 30 6000 40.305.1538 3 7200 40.300.1557 6 30 1200 41.939.0993 1 2400 41.973.1017 1 30 3600 41.991.0966 2 4800 42.016.0992 2 30 6000 42.003.0988 3 7200 42.116.1091 7 30 1200 28.593.6416 1 2400 31.315 1.5889 1 30 3600 31.225 1.5271 2 4800 31.757 1.4724 2 30 6000 31.761 1.3817 3 7200 32.254 1.3579 8 30 1200 24.876.0926 1 2400 25.009.1214 1 30 3600 25.199.1648 2 4800 25.116.1537 2 30 6000 25.180.1533 N of subject Time intervals N of values Mean Std. Deviation 3 7200 25.194.1524 9 30 1200 33.129.1027 1 2400 33.058.1055 1 30 3600 33.010.1042 2 4800 33.055.0956 2 30 6000 32.991.0972 3 7200 33.110.0947 10 30 1200 26.301.1328 1 2400 26.255.1174 1 30 3600 26.400.1233 2 4800 26.334.1171 2 30 6000 26.227.1179 3 7200 26.114.1422 11 30 1200 33.182.0806 1 2400 33.212.0946 1 30 3600 33.139.1017 2 4800 33.150.1043 2 30 6000 33.243.1142 3 7200 33.253.1124 12 30 1200 35.728.2501 1 2400 35.209.3278 1 30 3600 35.046.3387 2 4800 34.921.3446 2 30 6000 34.942.3350 3 7200 35.104.3093 13 30 1200 42.605.4709 1 2400 43.192.4507 1 30 3600 43.389.4390 2 4800 43.217.3902 2 30 6000 43.661.4202 3 7200 43.475.4037 14 30 1200 50.535.2154 1 2400 50.960.2919 1 30 3600 51.034.2943 2 4800 51.116.2908 2 30 6000 51.196.2902 3 7200 51.064.2747 15 30 1200 50.264.6278 1 2400 50.749.5629 1 30 3600 50.447.4845 2 4800 50.603.4502 2 30 6000 50.742.4430 3 7200 50.610.4468 We were particularly interested in comparing the 6 acquisition epochs (30 ; 1 ; 1 30 ; 2 ; 2 30 ; 3 ), starting with the 0 point. The signal mean and the standard deviations were analyzed and compared between epochs in order to observe how reproducible and reliable a shorter acquisition can be. In order to compare the standard deviations at the 6 time intervals (table II), the nonparametric Friedman test was applied (the test was applied to series of paired values that represent standard deviations) and we obtained p=0.204, i.e. there were no significant Mariana-Ioana Miron et al 47

differences among the 6 time intervals from a statistical point of view. (Table III). Table 2. The standard deviations for the 6 acquisition intervals studied, for the 15 subjects. case1 case2 case3 case4 case5 case6 case7 case8 case9 case10 case11 case12 case13 case14 case15 30 1 1 30 2 2 30 3 0,253 0,459 0,556 0,626 0,655 0,662 0,084 0,139 0,167 0,159 0,170 0,172 0,114 0,124 0,151 0,148 0,139 0,136 0,109 0,177 0,157 0,153 0,144 0,141 0,139 0,127 0,122 0,140 0,154 0,156 0,099 0,102 0,097 0,099 0,099 0,109 0,642 1,589 1,527 1,472 1,382 1,358 0,093 0,121 0,165 0,154 0,153 0,152 0,103 0,106 0,104 0,096 0,097 0,095 0,133 0,117 0,123 0,117 0,118 0,142 0,081 0,095 0,102 0,104 0,114 0,112 0,250 0,328 0,339 0,345 0,335 0,309 0,471 0,451 0,439 0,390 0,420 0,404 0,215 0,292 0,294 0,291 0,290 0,275 0,628 0,563 0,485 0,450 0,443 0,447 Table 3. The Friedman Test, test Statistics N 15 Chi-Square 7,231 df 5 Asymp. Sig.,204 Later, the times considered in the study were compared two by two, by applying the Wilcoxon signed-rank test (based on the same reasons as for the Friedman test), the results of the probabilities and their interpretations being given in Table IV: Discussions Laser Doppler flowmetry has been successfully employed for the estimation of pulp vitality in adults and children 8-11 differential diagnosis of 48 TMJ 2010, Vol. 60, No. 1 apical radiolucencies 12,13 examining reactions to pharmacological agents or electrical and thermal stimulation 14,15 and monitoring of pulpal responses to orthodontic procedures 16, or traumatic injuries. 17-22 Because this test produces no noxious stimuli, apprehensive or distressed patients accept it more readily than current methods of pulp vitality assessment, although the assessments may be highly susceptible to environmental and technique related factors. 23 Table 4. Wilcoxon signed-rank test Compared times p value 30 with 1 0,016s with 1 30 with 2 with 2 30 0,024 s 0,031 s 0,049 s 0,056 ns 1 with 1 30 0,866 ns with 2 with 2 30 0,784 ns 0,633 ns 0,587 ns 1 30 with 2 0,915 ns with 2 30 0,757 ns 0,708 ns 2 with 2 30 0,839 ns 0,788 ns 2 30 0,948 ns Interpretation: s significant differences with the significant threshold a=0.05, ns insignificant differences with the significant threshold a=0.05 Due to the complexity of this technique, accurate signal acquisition requires a fixed and constant position, minimizing involuntary patient movement. In the specialty literature, an acquisition time of 3 minutes is used most frequently, especially due to the indications of the manufacturer. 6,7 This time interval causes a real discomfort to the patient as he has to keep still during the acquisition, otherwise the movement artifacts appear and these can invalidate the testing, even if the technique of the probe fixation at tooth level is perfect. This situation causes real problems to the investigator, i.e. the doctor who performs the test. The analysis of the DC values stability, within this study, indicated a very low variation in time, which indicates a very stable fixation of the optical probe in the case of a use of a double silicone positioning system.

The statistical analysis of the experimental data shows that the increase with over 1 minute of time acquisition does not result in higher test accuracy but simply leads to the increase of the level of the disturbing signals due to patient s fatigue who, towards the end of acquisition, becomes eager for the test to finish. The reduction of acquisition interval under 1 minute affects the accuracy of the obtained values. In conclusion, the results of our study clearly showed that one minute acquisition of the laser Doppler signal in view of testing pulp vitality of frontal monoradicular teeth is sufficient in order to establish the vitality diagnosis. This decrease in the testing time from 3 minutes to 1 minute leads to a higher accuracy of the acquired signal through the reduction of the action time of the independant variables (i. e. signal disturbing factors: movement artifact, patient body position, external light sources, ambient temperature and humidity) on the laser Doppler signal acquisition as well as in higher patient comfort. References 1. Noblett WC, Wilcox LR, Scammon F, et al. Detection of pulpal circulation in vitro by pulse oximetry. J Endod 1996;22:1-5. 2. Ehrmann EH. Pulp testers and pulp testing with particular reference to the use of dry ice. Austral Dent J 1977;22:272-9. 3. Samraj RV, Indira R, Srinivasan MR, Kumar A. Recent advances in pulp vitality testing. Endodontology 2003;15:14 9. 4. Gazelius B, Olgart L, Edwall L. Non-invasive recording of blood flow in human denatal pulp. Endod Dent Traumatol 1986;2:219-21; 5. Gazelius B, Olgart L, Edwall L. Restored vitality in luxated teeth assessed by laser Doppler flowmetry. Endod Dent Traumatol 1988;4:265-8; 6. Roy E, Alliot-Licht B, Dajean-Trutaud S, et al. Evaluation of the ability of laser Doppler flowmetry for the assessment of pulp vitality in general dental practice. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2008;106(4):615-20. 7. Ikawa M, Komatsu H, Ikawa K, et al. Age-related changes in the human pulpal blood flow measured by laser doppler flowmetry. Dent Traumatol 2003;19:36-40. 8. Roeykens H, Van Maele G, De Moor R, et al. Reliability of Laser Doppler flowmetry in a 2-probe assessment of pulpal blood flow, Oral Surg Oral Med Oral Pathol Oral Radiol Endod 1999;87:742-8; 9. Sigurdsson A. Pulpal diagnosis. Endod Top 2003;5:12 5. 10. Camp JH. Diagnosis Dilemmas in Vital Pulp Therapy: Treatment for the Toothache is Changing, Especially in Young, Immature Teeth. Pediatr Dent 2008; 30(3):197-205. 11. Gopikrishna V, Pradeep G, Venkateshbabu N. Assessment of pulp vitality: a review. Int J Ped Dent 2009;19(1):3 15. 12. Kress B, Buhl Y, Hähnel S, et al. Age- and tooth-related pulp cavity signal intensity changes in healthy teeth: a comparative magnetic resonance imaging Analysis. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2007;103(1):134-7. 13. Walton RE, Chandler NP, Love RM, et al. Laser Doppler flowmetry - An aid in differential diagnosis of apical radiolucencies. Oral Surg Oral Med Oral Pathol Oral Radiol Endod. 1999; 87(5):613-6. 14. Goodis HE, Winthrop V, White JM. Pulpal Responses to Cooling Tooth Temperatures. J Endod, 2000;26(5):263-7. 15. Ikawa M, Ikawa K, Horiuchi H. The effects of thermal and mechanical stimulation on blood flow in healthy and inflamed gingiva in man. Arch Oral Biology 1998;43(2):127-132. 16. Bauss O, Röhling J, Meyer K, et al. Pulp Vitality in Teeth Suffering Trauma during Orthodontic Therapy. Angle Orthod 2009;79(1):16671. 17. Emshoff R, Moschen I, Strobl H. Use of laser Doppler flowmetry to predict vitality of luxated or avulsed permanent teeth Oral Surg Oral Med Oral Pathol Oral Radiol Endod. 2004;98(6):750-5. 18. Eshoff R, Emshoff I, Moschen I, et al. Laser Doppler low measurements of pulpal blood flow and severity of dental injury. Int Endod J 2004;37:463 7. 19. Olgart L, Gazelius B, Lindh-Stroemberg U. Laser Doppler flowmetry in assessing vitality in luxated teeth. Int. Endod J 1988;21:300-6; 20. Lee JY, Yanpiset K, Sigurdsson A, Vann WF. Laser Doppler flowmetry for monitoring traumatized teeth. Dent Traumatol 2001;17:231-5; 21. Strobl H, Haas M, Norer B, Gerhard S, Emshoff R. Evaluation of pulpal blood flow after tooth splinting of luxated permanent maxillary incisors. Dent Traumatol 2004;20:36-41. 22. Ikawa M, Fujiwara M, Horiuchi H, et al. The effect of short-term tooth intrusion on human pulpal blood flow measured by laser Doppler flowmetry. Arch Oral Bio 2001;46:781 7. 23. Jafarzadeh H. Laser Doppler flowmetry in endodontics: a review. Int Endod J 2009;42(6):476 90 Mariana-Ioana Miron et al 49