Dry eye disease (DED) is one of the most common reasons. Decreased Corneal Sensitivity in Patients with Dry Eye

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1 Decreased Corneal Sensitivity in Patients with Tristan Bourcier, 1,2 M. Carmen Acosta, 2 Vincent Borderie, 1 Fernando Borrás, 3 Juana Gallar, 2 Thierry Bury, 1 Laurent Laroche, 1 and Carlos Belmonte 2 From the 1 Quinze-Vingts National Center of Ophthalmology, University of Paris 6, Paris, France; the 2 Instituto de Neurociencias de Alicante, Universidad Miguel Hernández Consejo Superior de Investigaciones Científicas, San Juan de Alicante, Spain; and the 3 Centro de Investigación Operativa, Universidad Miguel Hernández, Elche, Spain. Submitted for publication December 6, 2004; revised March 9, 2005; accepted March 17, Disclosure: T. Bourcier, None; M.C. Acosta, None; V. Borderie, None; F. Borrás, None; J. Gallar, None; T. Bury, None; L. Laroche, None; C. Belmonte, None The publication costs of this article were defrayed in part by page charge payment. This article must therefore be marked advertisement in accordance with 18 U.S.C solely to indicate this fact. Corresponding author: Tristan Bourcier, Hôpital Quinze-Vingts, Service du Professeur Laurent Laroche, 28 rue de Charenton, Paris, France; bourcier@quinze-vingts.fr. PURPOSE. To explore changes in corneal sensitivity that develop in patients with dry eye and the relationship between sensibility and severity of the dry eye disease. METHODS. Experiments were performed in 44 patients with dry eye and 42 healthy individuals. Corneal sensitivity was measured with the Belmonte noncontact gas esthesiometer. Mechanical (air jets at flow rates from 0 to 200 ml/min, reaching the corneal surface at 34 C), thermal (cold or warm air at subthreshold flow rates changing corneal basal temperature 1 C), and chemical stimuli (air containing 0% to 50% CO 2 at subthreshold flow rate and temperature at the cornea of 34 C) were applied to the center of the cornea to determine the sensitivity threshold for each stimulus modality. The clinical state of the ocular surface was also explored, measuring the fluorescein tear break-up time, the degree of corneal staining with fluorescein and Lissamine green, and tear production with the Schirmer test. RESULTS. Both in control subjects and patients with dry eye, the corneal thresholds for mechanical, chemical, and thermal stimulation increased with age. Moreover, the thresholds for the three modalities of stimuli were significantly higher in patients with dry eye than in control subjects. In both groups, individual mechanical, chemical, and thermal thresholds correlated significantly. Also, high thresholds in patients with dry eye correlated with the intensity of fluorescein and Lissamine green corneal staining but not with the results of the Schirmer test. CONCLUSIONS. Patients with dry eye exhibit corneal hypoesthesia after mechanical, thermal, and chemical stimulation that appears to be related to damage to the corneal sensory innervation. (Invest Ophthalmol Vis Sci. 2005;46: ) DOI: /iovs Dry eye disease (DED) is one of the most common reasons that patients seek clinical examination by the ophthalmologist. Ocular irritation in DED may cause considerable discomfort and reduce the patient s quality of life. Epidemiologic studies investigating the prevalence of DED in the general population indicate that up to 33% of adults aged 50 years or more experience dry eye symptoms. 1 4 DED has been recently defined as a disease attributable to disturbances in the normal function and protective mechanisms of the ocular surface that lead to unstable tear film during the open-eye state. 5 Tear film instability or tear deficiency causes several corneal and conjunctival epithelial disturbances (keratoconjunctivitis sicca, KCS) and also reduces corneal sensitivity to mechanical stimulation measured with the Cochet-Bonnet esthesiometer. 6 However, this device has limited accuracy and only stimulates mechanosensory nerve fibers. Hence, the level of impact on the various functional types of corneal sensory receptors during KCS has not been established in detail. Belmonte et al., 7 in 1998, developed a noncontact gas esthesiometer that stimulates corneal nerve endings with a gas jet of adjustable flow, temperature, and CO 2 concentration, thus allowing the application of stimuli of different modalities (mechanical, thermal, and chemical) and of variable intensity and duration to the eye surface. 7 The gas esthesiometer enables exploration of the sensations evoked by selective activation of the three main classes of corneal sensory receptors mechanoreceptors, polymodal receptors, and thermoreceptors in different clinical conditions. 8 In the present study, we used this instrument to explore the corneal sensitivity of patients with dry eye and its relationship with the severity of keratoconjunctivitis sicca (KCS). METHODS Patients Forty-four patients with dry eye (14 men, 30 women; mean age, years; range, 18 92) and 42 age- and sex-matched normal subjects (14 men, 28 women; mean age, years; range, 22 77) were included in the study. Fourteen of the 44 patients with dry eye had primary or secondary Sjögren syndrome. The experiments adhered to the tenets of the Declaration of Helsinki. Patients signed an informed consent to a protocol approved by the Quinze-Vingts National Center of Ophthalmology and were free to discontinue the sessions at any time. Clinical Examination and Ocular Surface Tests Patients with dry eye reported having irritation, photophobia, and decreased vision of variable intensity. Before detailed ocular examination, patients were asked to record their symptoms in a standardized questionnaire. Three symptoms burning or sandy sensation, itching, and stinging were quantitatively scored by the patients. Zero corresponded to the absence of a symptom and 1, 2, 3, and 4 to minimal, mild, moderate, and severe intensity of the symptom, respectively. Clinical examinations included fluorescein tear break-up time (BUT), fluorescein and Lissamine green corneal staining, and Schirmer test. Subjects were included in the group of patients with DED when tear BUT was 6 seconds. Subsequently, corneal fluorescein and Lissamine green stainings were performed and graded according to the Oxford scheme 9 : 0, no staining; 1, minimal; 2, mild; 3, moderate; 4, diffuse; and 5, severe staining. The Schirmer 1 test (5 minutes, no topical anesthesia) was also performed in all patients. Clinical exploration and diagnostic tests were always performed before esthesiometry. The patients with dry eye were not receiving any topical treatment at the time of examination, nor were they using artificial tear substitutes or cyclosporine. None of the patients included in this group was being treated with topical nonsteroidal anti-inflammatory drugs. Investigative Ophthalmology & Visual Science, July 2005, Vol. 46, No. 7 Copyright Association for Research in Vision and Ophthalmology 2341

2 2342 Bourcier et al. IOVS, July 2005, Vol. 46, No. 7 TABLE 1. Distribution of Subjects and Patients with Group 1 (n 10) Group 2 (n 18) Group 3 (n 16) Group 1 (n 11) Group 2 (n 14) Group 3 (n 17) Mean age (y) Male (n) Female (n) Sjögren syndrome (n) Patients were distributed in three subgroups according to age: group 1 (below 40 years), group 2 (between 40 and 55 years), and group 3 (older than 55 years). Subjects included in the control group were recruited among the hospital staff and patients who entered the clinic for routine eye examination (ametropias, cataracts). Individuals with pathologic ocular symptoms, a history of contact lens wear or previous ocular surgery, use of eye drops, or the presence of a systemic disease that could interfere with tear film production were excluded. In control patients, tear BUT was more than 7 seconds, corneal fluorescein staining and Lissamine green staining were absent, and the Schirmer 1 test results were more than 15 mm. Esthesiometry Measurement of corneal sensation was made with the Belmonte noncontact gas esthesiometer. 7 This instrument was used to apply to the center of the cornea 3-second air jets of adjustable flow rate, composition, and temperature, separated by 15-second intervals. Selective mechanical stimulation consisted of a series of pulses of air at variable flow (0 200 ml/min). Air was heated at the tip of the probe to 50 C so that it reached the ocular surface at 34 C, to prevent a change in corneal temperature caused by the airflow. Thermal stimulation was performed with pulses of air previously heated or cooled to produce increases (heat stimulation) or decreases (cold stimulation) in the corneal surface temperature of 1 C, at a basal temperature of 34 C. 10 To prevent mechanical stimulation by thermal stimuli, the stimuli were applied at a flow that was 10 ml/min below mechanical threshold. Chemical stimulation was performed with air pulses, at subthreshold gas flow and a neutral temperature, containing increasing concentrations of CO 2 (0% 50%). In all experiments, pulses of different intensity were randomly applied. The esthesiometer s probe was adapted to a slit lamp table and the tip of the probe placed perpendicular to the apex of the cornea, at a distance of 5 mm, as measured with a transparent ruler. The patient was allowed to blink freely in the interstimulus intervals. An audible click produced by the opening of the gas valve identified the onset of the gas pulse. Immediately after each stimulation pulse, the subject was asked to report the presence or absence of sensation. Mechanical, thermal, and chemical thresholds were determined by recording the levels. 7,11,12 In each exploration, the stimulation sequence was always the same and started with the right eye followed by the left eye: mechanical, heat, cold, and chemical. All experiments were performed during the morning hours (between 8 and 12 AM) by the same physician. The temperature and humidity of the room were kept constant (20 C/45%). Statistical Analysis Threshold sensitivities in the dry eye and control groups were expressed as the mean SD. Because corneal sensitivity varies with age (Acosta MC, et al. IOVS 2004;45:ARVO E-Abstract 3946), normal subjects and patients with dry eye were grouped into three different age subgroups: group 1, 40 years; group 2, years; and group 3 55 years. Statistical analysis was performed on computer (Statistica ver. 6.1 software; StatSoft France, Maison-Alfort, France; and SPSS ver ; SPSS Inc., Chicago, IL). Irritation symptoms, ocular surface test results, and esthesiometry results were analyzed with the t-test or the Mann-Whitney test. Correlation coefficients were determined with the nonparametric Spearman test. P 0.05 was considered statistically significant. RESULTS Eighty-six patients (172 eyes) were examined. Table 1 summarizes some of the general characteristics of the subjects included in the control and the dry eye groups. Mean age, size of age subgroups, and proportion of men and women were matched in both groups of patients. Clinical Symptoms Table 2 lists the incidence and severity of irritation symptoms and the mean results of dry eye tests in the dry eye and control groups. There were no significant differences between the values of the different parameters in the men and women of both groups or between left and right eyes. Therefore, results in each subject are expressed as the mean of both eyes. No symptoms of irritation were reported by the subjects in the control group, whereas the symptoms were present in all patients with dry eye, and in them, the severity of the burning and stinging symptoms correlated significantly with the degree of fluorescein and Lissamine green stainings (Table 3). However, no correlation was observed between Lissamine corneal staining and BUT (Table 3). Itching did not correlate with any of the ocular surface test results, nor did age correlate with the severity of dry eye symptoms. Patients with Sjögren syndrome TABLE 2. Ocular Irritation Symptoms and Mean Results of Ocular Tests Symptoms Itching Burning Stinging Fluorescein Lissamine green BUT Schirmer test Symptoms were scored by the patient from 0 (absence of symptoms) to 4 (severe symptoms). Fluorescein and Lissamine green corneal staining were graded after slit-lamp examination (from 0 to 5) according to the Oxford scheme. Tear break-up time is expressed in seconds, Schirmer results in millimeters. Data are the mean SD. Differences between dry eye and control groups were statistically significant for all the irritation symptoms and tests (P 0.001).

3 IOVS, July 2005, Vol. 46, No. 7 Corneal Sensitivity in Patients 2343 TABLE 3. Correlation between Ocular Irritation Symptoms and Mean Results of Ocular Tests Correlation Coefficient Itching Burning Stinging Fluo LG BUT Schirmer Itching * 0.324* Burning ** 0.572** 0.588** Stinging 0.333* 0.574** 0.376* 0.322* Fluo 0.324* 0.572** 0.376* 0.741** 0.431** 0.517** LG ** 0.322* 0.741** * BUT ** ** Schirmer ** 0.337* 0.400** Ocular irritation sensations (itching, burning, and stinging), fluorescein (Fluo) and Lissamine green (LG) corneal stainings, tear break-up time (BUT), and Schirmer test results are compared using the Spearman correlation test. *P 0.05, ** P reported a more severe ocular burning sensation than did patients with dry eye but not Sjögren syndrome (P 0.05). Fluorescein staining scores were significantly higher in patients with Sjögren compared with those without (P 0.05). Local treatments significantly decreased fluorescein staining and burning and stinging symptoms (P 0.05), but did not affect other test results. Tear BUT and Schirmer test scores decreased with age in the control (P 0.05), but not in the dry eye group. Esthesiometry No significant differences were observed between the results obtained in left and right eyes, and thus for each subject, the mean of the results in both eyes was used. The mean thresholds for mechanical, thermal, and chemical stimulation are shown in Table 4. Mechanical, thermal and chemical thresholds were significantly higher in patients with dry eye (P 0.001). Mechanical, chemical, and thermal thresholds correlated significantly within both groups (P 0.05) that is, patients with comparatively higher mechanical thresholds also exhibited higher thermal and chemical thresholds. In patients with dry eye, sensitivity thresholds increased in parallel with the scores of the Lissamine green and fluorescein corneal staining tests (P 0.05, Spearman test). However, the tear BUT and Schirmer test results did not correlate with thresholds obtained with esthesiometry. The intensity of the stinging and burning symptoms, but not of itching, also correlated significantly with sensitivity thresholds (P 0.05). No clear sex-related differences were noted in these results. Although the mechanical threshold was slightly higher in Sjögren dry eyes ( ml/min) compared with non-sjögren dry eyes ( ml/min), differences did not reach the level of significance. The same was true of the other stimulus modalities tested. Patients receiving topical treatments did not differ in sensitivity thresholds from the rest of the patients with DED in the same age group. TABLE 4. Sensation Thresholds for Mechanical, Thermal, and Chemical Stimulation of the Cornea Mechanical threshold (ml/min) * Chemical threshold (%CO 2 ) * Heat threshold ( C) * Cold threshold ( C) * Data are expressed as means SD. Mean results in both eyes of each subject were used. * P 0.001, t-test or Mann-Whitney test. Mechanical and chemical thresholds increased with age in both the control and dry eye groups; however, thermal (cold and heat) stimulus thresholds did not clearly correlate with age in the control group (Table 5, Fig. 1). In all age groups, mechanical and chemical thresholds were significantly higher in patients with dry eye than in the control subjects (Table 5). Thresholds for heat and cold stimuli were also higher in patients with dry eye, although significance levels in comparison with control age-matched subjects were attained in groups 1 and 2 but not in group 3 (Table 5). DISCUSSION In the present study, corneal sensitivity to different stimulus modalities was significantly reduced in patients with dry eye when compared with age-matched normal subjects. The findings also evidenced subtle differences in corneal sensitivity associated with age and the modality of stimulus applied (mechanical, thermal, and chemical) that were present in both groups of patients. Significant reductions of corneal sensitivity measured with the Cochet-Bonnet esthesiometer have been previously re- TABLE 5. Sensation Thresholds for Mechanical, Thermal, and Chemical Stimulation of the Cornea in the Three Different Age Groups Mechanical threshold (ml/min) Group * Group Group Chemical threshold (%CO 2 ) Group * Group Group Heat threshold ( C) Group Group * Group (P 0.098) Cold threshold ( C) Group Group * Group (P 0.06) Data are expressed as means SD. Data for each dry eye group were compared with those for the corresponding control group. * P 0.01, P P Probabilities were by t-test or Mann-Whitney test.

4 2344 Bourcier et al. IOVS, July 2005, Vol. 46, No. 7 B D FIGURE 1. Relationship between sensitivity threshold and age, in normal subjects and patients with dry eye. Mechanical (A) and chemical (B) thresholds increased with age in both groups; however, the thermal (cold, C; heat, D) stimulus threshold did not clearly correlate with age in the control group. (F) Dry eyes; (E) control eyes. ported in patients with dry eye. 6 This instrument however explores only mechanical sensitivity and has limitations in its sensitivity and the reproducibility of its measurements. In the present study, corneal sensitivity was measured using the gas esthesiometer developed by Belmonte et al., 7 which applies controlled mechanical, chemical, and thermal stimuli to the corneal surface and allows separate determinations of mechanical, thermal, and chemical irritation sensations. When only air at the temperature of the corneal surface is applied, corneal mechanoreceptors are predominantly stimulated, accompanied by a moderate recruitment of polymodal nociceptors with the strongest stimuli. CO 2 mixtures induce a decrease of local ph at the corneal surface that is proportional to the CO 2 concentration. 10 This constitutes a selective stimulus for polymodal nociceptors of the cornea of intensity proportional to the local decline in ph. On the one hand, the warmth of the air jet applied to the cornea raises the normal temperature of 34 C of this tissue and selectively stimulates polymodal nociceptors, silencing simultaneously the cold receptors. On the other hand, moderate cooling stimulates the cold receptors solely, beginning to recruit polymodal nociceptors only when corneal temperatures below 29 C are attained. 7,13 In normal subjects, corneal sensitivity to mechanical and chemical stimulation decreased with age, as reflected by the significant increase in threshold to mechanical and chemical stimuli. This confirms previous observations with the Cochet- Bonnet esthesiometer 14,15 and the more recent findings of Acosta et al. (IOVS 2004;45:ARVO E-Abstract 3946), also with the gas esthesiometer. In addition, we detected a significant increase in threshold with all modalities of stimuli in patients with dry eye when compared with normal subjects of the same age, indicating that patients with DED have hypoesthesia extended to mechanical, chemical, and, to a lesser degree, thermal sensitivity that increases with age. The decreased sensitivity to these three modalities of stimuli showed crosscorrelation, suggesting that the damage to sensory nerve endings was unspecific and affected to a similar degree the different functional types of sensory receptors of the cornea. Within the group of patients with dry eye, corneal sensitivity appeared further reduced in parallel with the severity of DED. Changes in threshold to mechanical and chemical stimuli seem to reflect this reduction more reliably than thermal stimulation with cold or warm air. In patients with dry eye, the thickness and the composition of the tear film are disturbed. Therefore, the possibility that the altered tear film changes the final intensity of the stimulus reaching the corneal nerve endings must be considered. In the case of mechanical stimuli, the normal tear film is expected to act as a limited filter for mechanical forces. Decreases in its thickness and/or elastoviscosity would at best reduce this filtering effect, enhancing the transmission of force to the nerve endings so that the same stimulus would be more intensely felt in dry eyes. Despite this, in patients with DED, mechanical sensitivity was significantly lower. Chemical stimulation with CO 2 is conceivably mediated by the protons, resulting in the local formation of carbonic acid in the microenvironment of the nerve endings that are located at the intercellular space between epithelial cells. CO 2 diffuses very rapidly through cell membranes. Carbonic acid formation is proportional to CO 2 concentration and this in turn depends on the partial pressure and solubility coefficient of CO 2. These factors are not affected by the thickness of the tear film. Therefore, no differences in the magnitude of the decrease in ph caused by a given concentration of CO 2 are expected to occur between control and dry eyes. In the case of stimuli with cold air, evaporation can be a contributing factor to the cooling effect. It is difficult to establish the importance of evaporation in the final tempera-

5 IOVS, July 2005, Vol. 46, No. 7 Corneal Sensitivity in Patients 2345 ture change that is mainly caused by convection. The magnitude of evaporation depends on the temperature gradient between the gas jet and the cornea and the vapor pressure, and these factors should not vary much between normal corneas and those affected by and dry eye. However, it has been reported that in dry eyes, corneal evaporation can be faster than in normal eyes. 5,6 Thus, if anything, the temperature decrease caused by a given cold stimulus would be comparatively larger in dry eyes. There were no significant differences in sensitivity between patients with dry eye who had Sjögren syndrome and those without. This surprising finding suggests that the degree of functional impairment of transducing properties of the corneal nerve endings is similar in both conditions, although the higher level of dryness in patients with Sjögren increased the spontaneous activity of injured nerve fibers, thus evoking more discomfort. In a recent study, De Paiva and Pflugfelder 16 used a modified Belmonte gas esthesiometer to explore corneal sensitivity to mechanical and chemical (CO 2 ) stimulation in healthy and DED-affected subjects. The absolute threshold gas flow and CO 2 concentrations that they reported, both in normal subjects and in patients with dry eye were lower than in the present work. The differences in absolute thresholds are presumably attributable to variations in the physical characteristics of the stimulus delivered by the instrument used in each case, which present differences in size of the tip, tip distance to the cornea, and possibly the extension of the corneal area stimulated by the gas jet. All these factors introduce variations in the final intensity of the stimulus reaching the nerve endings and in the number of endings that are recruited by the stimulus. Variability among instruments is difficult to avoid until a homologated gas esthesiometer is available. Other factors such as diurnal oscillations of corneal sensitivity 17 or changes in the environmental conditions 18 may also contribute to the differences in threshold reported in these studies. The present work confirms in dry eyes the previously reported lack of correlation between subjective symptoms, tear deficiency (as measured by the Schirmer test), and ocular surface damage (evaluated with fluorescein and Lissamine green staining). 5,19,20 The degree of corneal staining with fluorescein and Lissamine green were the only parameters that correlated well with the esthesiometry thresholds reported in this study, further suggesting that the reduction in sensitivity found in DED is mainly attributable to a decrease in the number of functionally intact nerve endings, consecutive to the pathologic changes that occur in the superficial layers of the cornea. In parallel with the decrease in corneal sensitivity, patients with dry eye experienced discomfort and irritation symptoms that were equally associated with the importance of the disturbance of the ocular surface. The capability of sensory nerve endings to transduce a physical or chemical stimulus of a given intensity into a discharge of nerve impulses that propagates to the brain, giving rise to a conscious sensation, depends on the density and functional integrity of sensory nerve endings in the stimulated area. When corneal endings are injured, as seems to occur to a portion of them in dry eye conditions, they lose their transducing properties, and this results in a reduced number of intact endings able to signal natural stimuli and a number of damaged axons in various stages of regeneration. These form neuromas and show development of abnormal impulse activity. 8 It is likely that this altered excitability is the origin of the dysesthesia and the subjective symptoms reported by our patients with DED. 21 The results obtained in this study indicate that gas esthesiometry may serve as a more refined clinical procedure than those available hitherto to evaluate the integrity of the corneal innervation in DED and eventually also to assess the efficacy and effects on corneal sensibility of the various treatments applied in this disease. References 1. Moss SE, Klein R, Klein BE. Incidence of dry eye in an older population. Arch Ophthalmol. 2004;122: Schaumberg DA, Sullivan DA, Buring JE, Dana MR. Prevalence of dry eye syndrome among US women. Am J Ophthalmol. 2003; 136: Lin PY, Tsai SY, Cheng CY, Liu JH, Chou P, Hsu WM. Prevalence of dry eye among an elderly Chinese population in Taiwan: the Shihpai eye study. Ophthalmology. 2003;110: Chia EM, Mitchell P, Rochtchina E, Lee AJ, Maroun R, Wang JJ. Prevalence and associations of dry eye syndrome in an older population: the Blue Mountain Eye Study. Clin Exp Ophthalmol. 2003;31: Brewitt H, Sistani F. Dry eye disease: the scale of the problem. Surv Ophthalmol. 2001;45(suppl 2):S199 S Xu KP, Yagi Y, Tsubota K. Decrease in corneal sensitivity and change in tear function in dry eye. Cornea. 1996;15: Belmonte C, Acosta MC, Schmelz M, Gallar J. Measurement of corneal sensitivity to mechanical and chemical stimuli with a CO 2 esthesiometer. Invest Ophthalmol Vis Sci. 1999;40: Belmonte C, Acosta MC, Gallar J. Neural basis of sensation in intact and injured corneas. Exp Eye Res. 2004;78: Bron AJ, Evans VE, Smith JA. Grading of corneal and conjunctival staining in the context of other dry eye tests. Cornea. 2003;22: Chen X, Gallar J, Pozo MA, Baeza M, Belmonte C. 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Am J Ophthalmol. 2004;137: Du Toit R, Vega JA, Fonn D, Simpson T. Diurnal variation of corneal sensitivity and thickness. Cornea. 2003;22: Bourcier T, Viboud C, Cohen JC, et al. Effects of air pollution and climatic conditions on the frequency of ophthalmological emergency examinations. Br J Ophthalmol. 2003;87: Baudouin C. The pathology of dry eye. Surv Ophthalmol. 2001; 45(suppl 2):S211 S Macri A, Pflugfelder S. Correlation of the Schirmer 1 and fluorescein clearance tests with the severity of corneal epithelial and eyelid disease. Arch Ophthalmol. 2000;118: Belmonte C, Aracil A, Acosta MC, Luna C, Gallar J. Nerves and sensations from the eye surface. Ocul Surface. 2004;2:29 34.