Conjunctival Impression Cytology in Patients With Glaucoma Using Long-term Topical Medication James D. Brandt, M.D., John R. Wittpenn, M.D., L. Jay Katz, M.D., William N. Steinmann, M.D., and George L. Spaeth, M.D.
Increasing evidence indicates that longterm use of topically administered medications can induce changes in the conjunctiva and ocular surface. We used the technique of conjunctival impression cytology to evaluate the conjunctival changes that develop with long-term use of topically administered antiglaucoma medications. Patients with glaucoma who were on a stable regimen of one, two, or three topically administered medications were recruited for study; glaucoma suspects who were not using topically administered medications served as controls. Eyes with clinical or historical evidence of external eye disease or conjunctival surgery were excluded. Impression cytology specimens, collected from the bulbar and palpebral conjunctiva, were coded and subsequently graded by a masked observer. We examined specimens from 72 eyes by using this technique. Aggregate scores for the bulbar conjunctiva were compiled, using a previously described grading system with a range of 0 (normal) to 3 (diffuse, severe metaplasia). The results show statistically significant degrees of conjunctival metaplasia associated with the number of glaucoma medications used. These results suggest that the long-term use of antiglaucoma medications induces changes in the conjunctival surface. These changes may be Accepted for publication May 16, 1991. From the Glaucoma Service, Wills Eye Hospital, Philadelphia, Pennsylvania (Drs. Brandt, Katz, Steinmann, and Spaeth); and the Department of Ophthalmology, School of Medicine, State University of New York, Stony Brook, New York (Dr. Wittpenn). This study was supported in part by the Richard G. Scobee Foundation. Dr. Brandt was a Heed Foundation Fellow (1988-1989). Reprint requests to James D. Brandt, M.D., Department of Ophthalmology, University of California at Davis, 1603 Alhambra Blvd., Sacramento, CA 958167051.
©AMERICAN JOURNAL OF OPHTHALMOLOGY
112:297,
related to the medications themselves, the preservatives in the commercial preparations, or the duration of topical treatment. The clinical relevance of these changes remains unknown. ALTHOUGH TOPICALLY ADMINISTERED medications remain the usual treatment in primary open-angle glaucoma, these drugs are not without adverse effects, both ocular and systemic. The systemic adverse effects of several classes of these drugs (for example, systemic ~-adren ergic blockade) have been studied extensively. Although some ocular adverse effects (for example, cystoid macular edema in aphakic patients receiving epinephrine) have been identified, remarkably little is known about the effects of long-term use of these drugs on the conjunctiva. Although clinicians view the conjunctiva as a passive, semipermeable barrier that allows drugs to enter the eye, it is nevertheless a living dynamic tissue that can respond to stress with inflammation, scarring, keratinization, and neovascularization. Virtually all topically administered medications are known to cause conjunctival reactions in some patients. The topically administered drugs used in the treatment of glaucoma are no exception. Particular offenders include echothiophate iodide.' dipivefrin hydrochloride." and pilocarpine.t-" although the most frequently used class of glaucoma medications, the ~-adrenergic antagonists, are also known to cause conjunctival keratinization.' Much anecdotal evidence suggests that the conjunctiva, the ocular surface, or both, of patients using topically administered antiglaucoma medications are altered. Many patients with glaucoma complain of dry eye symptoms when using their medications, yet their tear
SEPTEMBER,
1991
297
298
September, 1991
AMERICAN JOURNAL OF OPHTHALMOLOGY
production as measured by Schirmer testing is normal, suggesting that the lipid composition of the tear film has been altered. Conjunctival goblet cells are lost with long-term treatment with pilocarpine." and light and electron microscopy disclose conjunctival changes with long-term ~-antagonist treatment." There is in vitro evidence that many of the antiglaucoma drugs (and the preservatives used in their formulation) are toxic to conjunctival epithelium in culture. 7 The conjunctival surface is known to possess regional variations in both cellular distribution and morphologic characteristics. Because of these regional variations, incisional biopsy is limited as a method to examine the conjunctival surface; it is invasive and provides only a crosssectional view of a single point on the ocular surface. Conjunctival impression cytology is a method of obtaining cytologic specimens from the conjunctival surface.":" It is safe, painless, and simple, and has provided insights into the conjunctival causes of a number of ocular surface disorders, including xerophthalmia.l':" keratoconjunctivitis sicca," squamous metaplasia, 15 conjunctivitis," ocular pemphigoid, StevensJohnson syndrome, and others." Electron microscopy of the specimens is possible" and sequential study is not difficult. In this study, we evaluated the conjunctival changes associated with long-term use of antiglaucoma medications, using the technique of conjunctival impression cytology.
Subjects and Methods
The patient protocol and associated informed-consent documents were reviewed and approved by the Institutional Review Board of the Wills Eye Hospital. Consecutive subjects were recruited from the Glaucoma Service of the Wills Eye Hospital. Patients asked to participate included the following: (1) patients with glaucoma who were on a long-term (greater than three months) stable regimen of topically administered glaucoma medications that included one or more of the three basic classes of glaucoma medications: epinephrine or dipivefrin, ~-adrenergic antagonists, and miotics; and (2) patients being monitored as glaucoma suspects who were receiving no topical treatment. Patients were excluded from the study for the following reasons: (1) age under 18 years; (2) any history of surgery or other interventions
such as cryotherapy that might cause conjunctival scarring; (3) use of any topically administered medications not among the classes previously described, including artificial tear preparations; or (4) any history or slit-lamp examination evidence of ocular surface disorder (that is, ocular pemphigoid, keratoconjunctivitis sicca, or herpes) predating the diagnosis and treatment of glaucoma. Informed consent was obtained from the patients who participated. On entering the study, a detailed ocular history was obtained, emphasizing medication history, as well as symptoms and clinical signs of ocular surface disorders. Each patient received a careful slit-lamp examination by one of us to identify any ocular surface abnormalities or evidence of external eye disease. Data were recorded in a standardized format and coded for later retrieval. We performed impression cytologic examination of the topically anesthetized conjunctiva according to the technique described by Nelson." Small (6.2 mm) disks of cellulose acetate filter paper were placed on the bulbar conjunctiva after topically administered anesthesia was achieved. Four bulbar conjunctival specimens were obtained just adjacent to the corneoscleral limbus nasally, temporally, inferiorly, and superiorly. Specimens of the inferior palpebral conjunctiva at the center of the lower eyelid were obtained. The five specimens from each eye were then attached to microscope slides with double-sided tape so that their positions on the slide corresponded to the location from which the specimen was obtained. Specimens were then fixed with cytologic spray fixative and stored for later staining. The specimens were stained, examined, and graded by one of us (J.R.W.) according to the technique and grading scheme outlined by Nelson.'? The examiner was masked to the clinical or medical history of the patient from whom the specimen was taken. The impression cytology grades from the four bulbar specimens were averaged for an individual bulbar score; the single palpebral conjunctival specimen grade was analyzed alone. Statistical comparison between cases and controls was performed by use of Student's unpaired t-test. Results
We recruited 50 patients to participate in the study. Specimens in which two or more of the
Vol. 112, No.3
Conjunctival Cytology in Treated Glaucoma
five cytologic specimens obtained per eye that were judged unreadable were discarded. After excluding 28 individual eyes that met the exclusion criteria listed, we analyzed the impression cytology data from a total of 72 eyes. Comparison of the four groups disclosed similar ages (Table). The control group, consisting of patients suspected of having glaucoma and who were receiving no topically administered medication, had the lowest cumulative impression cytology score, a finding that was consistent with a healthy conjunctiva. The impression cytology grade of three of the four treatment groups (~-blockers, ~-blockers and pilocarpine, and maximal treatment) all demonstrated a statistically significant increase in cytologic grade compared to the control group (Table). The group of patients receiving a combination of a ~-blocker and epinephrine/ dipivefrin had a cytologic grade only slightly higher than the control group, and this difference was not statistically significant (P = .175). Palpebral and bulbar conjunctival cytologic grades were determined in all five groups (Figure). The largest difference between treatment group and control is seen in those patients receiving maximal treatment, suggesting a relationship between the number of medications and the cytologic grade. Discussion
Our results confirm the clinical impression that the ocular surfaces of patients receiving long-term topical treatment for glaucoma are abnormal. Our data suggest a relationship between the number of medications and the severity of conjunctival changes observed. In obtaining medication histories, we found
299
that most patients could not recall the specific duration of individual treatments. This information was not available from the patients' charts, as most had received much of their care elsewhere before being referred to us. Because most patients receiving maximal treatment had been under medical treatment longer than those on single-drug treatment, our findings may be more closely related to the overall duration of topical treatment rather than to the individual components of that treatment. One strength of this study was that the grading of specimens was done in a masked fashion. However, interpreting the statistical (and clinical) relevance of the findings is hampered because the grading system we used is a subjective one that simply divides cytologic appearance into four categories. We do not fully understand the natural history of conjunctival metaplasia. If grades 2 and 3 represent metaplastic changes that are at the far end of the cytologic spectrum (for example, 95 and 99 on a scale of 100), then our findings are perhaps more significant than P values alone would indicate. Conversely, if grade 2 represents metaplasia at the lower portion of the cytologic spectrum, our results may be less significant. The cause for the apparent effect of long-term topical treatment for glaucoma on the conjunctiva is not known. The glaucoma drugs themselves or the preservative common to their commercial preparations, benzalkonium chloride, may be responsible. We suspect that benzalkonium chloride has a large role in the development of the conjunctival changes we have demonstrated. Of importance is that the only treatment group that did not demonstrate a statistically significant increase in cytologic grade was that in which a ~-adrenergic antagonist was combined with an adrenergic agonist (epinephrineydipivefrin). Perhaps some of the
TABLE
AGGREGATE IMPRESSION CYTOLOGY SCORES BY TREATMENT GROUP
TREATMENT GROUP
Controls p-blockers alone p-blockers and epinephrine/dipivefrin p-blockers and pilocarpine Maximal treatment "P = .05 compared to control. tp = .001 compared to control.
AGE
BULBAR
PALPEBRAL
NO. OF EYES
(YRs)
CONJUNCTIVA
CONJUNCTIVA
19 13
69.5 ± 12.8 67.2 ± 7.6
0.75 ± 0.11 1.23 ± 0.11"
0.05 ± 0.05 0.39 ± 0.18"
10
73.6 ±
9.8
1.00 ± 0.13
0.30 ± 0.15
22 8
67.8 ± 73.9 ±
8.3 6.4
1.35 ± 0.12t 1.44 ± 0.13t
0.46 ± 0.11" 0.63 ± 0.18t
300
September, 1991
AMERICAN JOURNAL OF OPHTHALMOLOGY
2
o
Palpebral
•
Bulbar
Figure (Brandt and associates). Graph comparing the conjunctival impression cytologic grade of both bulbar and palpebral conjunctivae in the patient groups studied.
o.lL-L--
Control
B-Blocker
B-Blocker/Epi
B-Blocker/Pllo
Maximum Rx
Treatment Group
conjunctival changes we have demonstrated are mediated at the adrenergic receptor. The impact of these conjunctival changes on the outcome of filtering surgery is clinically important, because most patients undergo surgery only after months or years of topical drug treatment. Successful glaucoma-filtering surgery depends on many technical and host factors, not the least of which is the existence of a healthy conjunctiva. Long-term conjunctival inflammation and scarring, whether results of underlying ocular disease, previous surgery, or drug-induced toxicity such as pseudopemphigoid, are known to predispose patients to poor outcome after filtration surgery." Recent animal studies" demonstrated an increase of myofibroblastic cell proliferation in fistulized rabbit conjunctiva treated with glaucoma medications or a preserved artificial tear compared with untreated controls. Sherwood and associates" reported on the morphologic effects of antiglaucoma drugs on the conjunctiva and Tenon's capsule in patients undergoing trabeculectomy. Results of biopsies in patients who had been on long-term topical treatment showed an increase in macrophages, fibroblasts, lymphocytes, and mast cells and decreased conjunctival goblet cells compared with patients who had received little if any preoperative drug treatment. In a related report, Lavin and assoctates" found that filtering surgery was more successful in patients who had received an average of only two weeks of preoperative medical treatment compared with patients who had received at least one year of medical treatment.
It is often difficult to identify those patients who are at increased risk of surgical failure after trabeculectomy. Our study results suggest that impression cytology may be useful in identifying conjunctival changes related to longterm drug use, and further studies will be needed to determine whether these changes are predictive of surgical outcome.
References 1. Fiore, P. M., Jacobs, I. H., and Goldberg, D. B.: Drug-induced pemphigoid. A spectrum of diseases. Arch. Ophthalmol. 105:1660, 1987. 2. Liesegang, T. J.: Bulbar conjunctival follicles associated with dipivefrin therapy. Ophthalmology
92:228,1985.
3. Johnson, D. H., Kenyon, K. R., Epstein, D. L., and Van Buskirk, E. M.: Corneal changes during pilocarpine gel therapy. Am. J. Ophthalmol. 101:13,
1986.
4. Derous, D., de Keizer, R. J., de WolffRouendaal, D., and Soudijn, W.: Conjunctival keratinization. An abnormal reaction to an ocular betablocker. Acta Ophthalmol. (Copenh.) 67:333, 1989. 5. Gerstenberger, A., and Marquardt, R.: Die becherzelldicte unter pilocarpineinflu[3 [The density of goblet cells modified by pilocarpine]. Fortschr. Ophthalmol. 83:46, 1986. 6. Varga, M., and Follmann, P.: Feinstrukturelle Untersuchungen der Bindehautoberflache nach Langzeitbehandlung mit Timolol [Ultrastructural studies of the conjunctival surface following longterm treatment with timolol]. Fortschr. Ophthalmol.
83:155, 1986.
7. Takahashi, N.: Cytotoxic effects of antiglaucoma
Vol. 112, No.3
Conjunctival Cytology in Treated Glaucoma
agents on cultured human conjunctival cells (author's transl.). Nippon Ganka Gakkai Zasshi 85:1046, 1981. 8. Egbert, P. R., Lauber,S., and Maurice, D. M.: A simple conjunctival biopsy. Am. J. Ophthalmol. 84:798, 1977. 9. Nelson, J. D., Havener, V. R., and Cameron, J. D.: Cellulose acetate impressions of the ocular surface. Dry eye states. Arch. Ophthalmol. 101:1869, 1983. 10. Nelson, J.: Impression cytology. Cornea 7:71, 1988. 11. Wittpenn, J., Tseng,S., and Sommer, A.: Detection of early xerophthalmia by impression cytology. Arch. Ophthalmol. 104:237, 1986. 12. Amedee-Manesme, 0., Luzeau, R., Wittpen, J., Hanck, A., and Sommer, A.: Impression cytology detects subclinical vitamin A deficiency. Am. J. Clin. Nutr. 47:875, 1988. 13. Keenum, D. G., Semba, R. D., Wirasasmita, 5., Natadisastra, G., Muhilal, West, K. P., and Sommer, A.: Assessment of vitamin A status by a disk applicator for conjunctival impression cytology. Arch. Ophthamol. 108: 1436, 1990. 14. Nelson, J., and Farris, R.: Sodium hyaluronate and polyvinyl alcohol artificial tear preparations. A comparison in patients with keratoconjunctivitis sicca. Arch. Ophthalmol. 106:484, 1988. 15. Tseng,S.: Staging of conjunctival squamous
301
metaplasia by impression cytology. Ophthalmology 92:728, 1985. 16. Hershenfeld, 5., Kazdan, J., Mancer, K., Feugas, P., Basu, P., and Avaria, M.: Impression cytology in conjunctivitis. Can. J. Ophthalmol. 16:76, 1981. 17. Nelson, J. D., and Wright, J. c.. Conjunctival goblet cell densities in ocular surface disease. Arch. Ophthalmol. 102:1049, 1984. 18. Maskin, 5., and Bode, D.: Electron microscopy of impression-acquired conjunctival epithelial cells. Ophthalmology 93:1518, 1986. 19. Rockwood, E. J., Parrish, R. K., II, Heuer, D. K., Skuta, G. L., Hodapp, E., Palmberg, P. F., Gressel, M. G., and Feuer, W.: Glaucoma filtering surgery with 5-fluorouracil. Ophthalmology 94:1071,1987. 20. Young, T. L., Higginbotham, E. J., Zou, X" and Farber, M. D.: Effects of topical glaucoma drugs on fistulized rabbit conjunctiva. Ophthalmology 97:1423, 1990. 21. Sherwood, M. B., Grierson, I., Millar, L., and Hitchings, R. A.: Long-term morphologic effects of antiglaucoma drugs on the conjunctiva and Tenon's capsule in glaucomatous patients. Ophthalmology 96:327, 1989. 22. Lavin, M. J., Wormald, R. P. L., Migdal, C. 5., and Hitchings, R. A.: The influence of prior therapy on the success of trabeculectomy. Arch. Ophthalmol. 108:1543,1990.
Increasing evidence indicates that longterm use of topically administered medications can induce changes in the conjunctiva and ocular surface. We used the technique of conjunctival impression cytology to evaluate the conjunctival changes that develop with long-term use of topically administered antiglaucoma medications. Patients with glaucoma who were on a stable regimen of one, two, or three topically administered medications were recruited for study; glaucoma suspects who were not using topically administered medications served as controls. Eyes with clinical or historical evidence of external eye disease or conjunctival surgery were excluded. Impression cytology specimens, collected from the bulbar and palpebral conjunctiva, were coded and subsequently graded by a masked observer. We examined specimens from 72 eyes by using this technique. Aggregate scores for the bulbar conjunctiva were compiled, using a previously described grading system with a range of 0 (normal) to 3 (diffuse, severe metaplasia). The results show statistically significant degrees of conjunctival metaplasia associated with the number of glaucoma medications used. These results suggest that the long-term use of antiglaucoma medications induces changes in the conjunctival surface. These changes may be Accepted for publication May 16, 1991. From the Glaucoma Service, Wills Eye Hospital, Philadelphia, Pennsylvania (Drs. Brandt, Katz, Steinmann, and Spaeth); and the Department of Ophthalmology, School of Medicine, State University of New York, Stony Brook, New York (Dr. Wittpenn). This study was supported in part by the Richard G. Scobee Foundation. Dr. Brandt was a Heed Foundation Fellow (1988-1989). Reprint requests to James D. Brandt, M.D., Department of Ophthalmology, University of California at Davis, 1603 Alhambra Blvd., Sacramento, CA 958167051.
©AMERICAN JOURNAL OF OPHTHALMOLOGY
112:297,
related to the medications themselves, the preservatives in the commercial preparations, or the duration of topical treatment. The clinical relevance of these changes remains unknown. ALTHOUGH TOPICALLY ADMINISTERED medications remain the usual treatment in primary open-angle glaucoma, these drugs are not without adverse effects, both ocular and systemic. The systemic adverse effects of several classes of these drugs (for example, systemic ~-adren ergic blockade) have been studied extensively. Although some ocular adverse effects (for example, cystoid macular edema in aphakic patients receiving epinephrine) have been identified, remarkably little is known about the effects of long-term use of these drugs on the conjunctiva. Although clinicians view the conjunctiva as a passive, semipermeable barrier that allows drugs to enter the eye, it is nevertheless a living dynamic tissue that can respond to stress with inflammation, scarring, keratinization, and neovascularization. Virtually all topically administered medications are known to cause conjunctival reactions in some patients. The topically administered drugs used in the treatment of glaucoma are no exception. Particular offenders include echothiophate iodide.' dipivefrin hydrochloride." and pilocarpine.t-" although the most frequently used class of glaucoma medications, the ~-adrenergic antagonists, are also known to cause conjunctival keratinization.' Much anecdotal evidence suggests that the conjunctiva, the ocular surface, or both, of patients using topically administered antiglaucoma medications are altered. Many patients with glaucoma complain of dry eye symptoms when using their medications, yet their tear
SEPTEMBER,
1991
297
298
September, 1991
AMERICAN JOURNAL OF OPHTHALMOLOGY
production as measured by Schirmer testing is normal, suggesting that the lipid composition of the tear film has been altered. Conjunctival goblet cells are lost with long-term treatment with pilocarpine." and light and electron microscopy disclose conjunctival changes with long-term ~-antagonist treatment." There is in vitro evidence that many of the antiglaucoma drugs (and the preservatives used in their formulation) are toxic to conjunctival epithelium in culture. 7 The conjunctival surface is known to possess regional variations in both cellular distribution and morphologic characteristics. Because of these regional variations, incisional biopsy is limited as a method to examine the conjunctival surface; it is invasive and provides only a crosssectional view of a single point on the ocular surface. Conjunctival impression cytology is a method of obtaining cytologic specimens from the conjunctival surface.":" It is safe, painless, and simple, and has provided insights into the conjunctival causes of a number of ocular surface disorders, including xerophthalmia.l':" keratoconjunctivitis sicca," squamous metaplasia, 15 conjunctivitis," ocular pemphigoid, StevensJohnson syndrome, and others." Electron microscopy of the specimens is possible" and sequential study is not difficult. In this study, we evaluated the conjunctival changes associated with long-term use of antiglaucoma medications, using the technique of conjunctival impression cytology.
Subjects and Methods
The patient protocol and associated informed-consent documents were reviewed and approved by the Institutional Review Board of the Wills Eye Hospital. Consecutive subjects were recruited from the Glaucoma Service of the Wills Eye Hospital. Patients asked to participate included the following: (1) patients with glaucoma who were on a long-term (greater than three months) stable regimen of topically administered glaucoma medications that included one or more of the three basic classes of glaucoma medications: epinephrine or dipivefrin, ~-adrenergic antagonists, and miotics; and (2) patients being monitored as glaucoma suspects who were receiving no topical treatment. Patients were excluded from the study for the following reasons: (1) age under 18 years; (2) any history of surgery or other interventions
such as cryotherapy that might cause conjunctival scarring; (3) use of any topically administered medications not among the classes previously described, including artificial tear preparations; or (4) any history or slit-lamp examination evidence of ocular surface disorder (that is, ocular pemphigoid, keratoconjunctivitis sicca, or herpes) predating the diagnosis and treatment of glaucoma. Informed consent was obtained from the patients who participated. On entering the study, a detailed ocular history was obtained, emphasizing medication history, as well as symptoms and clinical signs of ocular surface disorders. Each patient received a careful slit-lamp examination by one of us to identify any ocular surface abnormalities or evidence of external eye disease. Data were recorded in a standardized format and coded for later retrieval. We performed impression cytologic examination of the topically anesthetized conjunctiva according to the technique described by Nelson." Small (6.2 mm) disks of cellulose acetate filter paper were placed on the bulbar conjunctiva after topically administered anesthesia was achieved. Four bulbar conjunctival specimens were obtained just adjacent to the corneoscleral limbus nasally, temporally, inferiorly, and superiorly. Specimens of the inferior palpebral conjunctiva at the center of the lower eyelid were obtained. The five specimens from each eye were then attached to microscope slides with double-sided tape so that their positions on the slide corresponded to the location from which the specimen was obtained. Specimens were then fixed with cytologic spray fixative and stored for later staining. The specimens were stained, examined, and graded by one of us (J.R.W.) according to the technique and grading scheme outlined by Nelson.'? The examiner was masked to the clinical or medical history of the patient from whom the specimen was taken. The impression cytology grades from the four bulbar specimens were averaged for an individual bulbar score; the single palpebral conjunctival specimen grade was analyzed alone. Statistical comparison between cases and controls was performed by use of Student's unpaired t-test. Results
We recruited 50 patients to participate in the study. Specimens in which two or more of the
Vol. 112, No.3
Conjunctival Cytology in Treated Glaucoma
five cytologic specimens obtained per eye that were judged unreadable were discarded. After excluding 28 individual eyes that met the exclusion criteria listed, we analyzed the impression cytology data from a total of 72 eyes. Comparison of the four groups disclosed similar ages (Table). The control group, consisting of patients suspected of having glaucoma and who were receiving no topically administered medication, had the lowest cumulative impression cytology score, a finding that was consistent with a healthy conjunctiva. The impression cytology grade of three of the four treatment groups (~-blockers, ~-blockers and pilocarpine, and maximal treatment) all demonstrated a statistically significant increase in cytologic grade compared to the control group (Table). The group of patients receiving a combination of a ~-blocker and epinephrine/ dipivefrin had a cytologic grade only slightly higher than the control group, and this difference was not statistically significant (P = .175). Palpebral and bulbar conjunctival cytologic grades were determined in all five groups (Figure). The largest difference between treatment group and control is seen in those patients receiving maximal treatment, suggesting a relationship between the number of medications and the cytologic grade. Discussion
Our results confirm the clinical impression that the ocular surfaces of patients receiving long-term topical treatment for glaucoma are abnormal. Our data suggest a relationship between the number of medications and the severity of conjunctival changes observed. In obtaining medication histories, we found
299
that most patients could not recall the specific duration of individual treatments. This information was not available from the patients' charts, as most had received much of their care elsewhere before being referred to us. Because most patients receiving maximal treatment had been under medical treatment longer than those on single-drug treatment, our findings may be more closely related to the overall duration of topical treatment rather than to the individual components of that treatment. One strength of this study was that the grading of specimens was done in a masked fashion. However, interpreting the statistical (and clinical) relevance of the findings is hampered because the grading system we used is a subjective one that simply divides cytologic appearance into four categories. We do not fully understand the natural history of conjunctival metaplasia. If grades 2 and 3 represent metaplastic changes that are at the far end of the cytologic spectrum (for example, 95 and 99 on a scale of 100), then our findings are perhaps more significant than P values alone would indicate. Conversely, if grade 2 represents metaplasia at the lower portion of the cytologic spectrum, our results may be less significant. The cause for the apparent effect of long-term topical treatment for glaucoma on the conjunctiva is not known. The glaucoma drugs themselves or the preservative common to their commercial preparations, benzalkonium chloride, may be responsible. We suspect that benzalkonium chloride has a large role in the development of the conjunctival changes we have demonstrated. Of importance is that the only treatment group that did not demonstrate a statistically significant increase in cytologic grade was that in which a ~-adrenergic antagonist was combined with an adrenergic agonist (epinephrineydipivefrin). Perhaps some of the
TABLE
AGGREGATE IMPRESSION CYTOLOGY SCORES BY TREATMENT GROUP
TREATMENT GROUP
Controls p-blockers alone p-blockers and epinephrine/dipivefrin p-blockers and pilocarpine Maximal treatment "P = .05 compared to control. tp = .001 compared to control.
AGE
BULBAR
PALPEBRAL
NO. OF EYES
(YRs)
CONJUNCTIVA
CONJUNCTIVA
19 13
69.5 ± 12.8 67.2 ± 7.6
0.75 ± 0.11 1.23 ± 0.11"
0.05 ± 0.05 0.39 ± 0.18"
10
73.6 ±
9.8
1.00 ± 0.13
0.30 ± 0.15
22 8
67.8 ± 73.9 ±
8.3 6.4
1.35 ± 0.12t 1.44 ± 0.13t
0.46 ± 0.11" 0.63 ± 0.18t
300
September, 1991
AMERICAN JOURNAL OF OPHTHALMOLOGY
2
o
Palpebral
•
Bulbar
Figure (Brandt and associates). Graph comparing the conjunctival impression cytologic grade of both bulbar and palpebral conjunctivae in the patient groups studied.
o.lL-L--
Control
B-Blocker
B-Blocker/Epi
B-Blocker/Pllo
Maximum Rx
Treatment Group
conjunctival changes we have demonstrated are mediated at the adrenergic receptor. The impact of these conjunctival changes on the outcome of filtering surgery is clinically important, because most patients undergo surgery only after months or years of topical drug treatment. Successful glaucoma-filtering surgery depends on many technical and host factors, not the least of which is the existence of a healthy conjunctiva. Long-term conjunctival inflammation and scarring, whether results of underlying ocular disease, previous surgery, or drug-induced toxicity such as pseudopemphigoid, are known to predispose patients to poor outcome after filtration surgery." Recent animal studies" demonstrated an increase of myofibroblastic cell proliferation in fistulized rabbit conjunctiva treated with glaucoma medications or a preserved artificial tear compared with untreated controls. Sherwood and associates" reported on the morphologic effects of antiglaucoma drugs on the conjunctiva and Tenon's capsule in patients undergoing trabeculectomy. Results of biopsies in patients who had been on long-term topical treatment showed an increase in macrophages, fibroblasts, lymphocytes, and mast cells and decreased conjunctival goblet cells compared with patients who had received little if any preoperative drug treatment. In a related report, Lavin and assoctates" found that filtering surgery was more successful in patients who had received an average of only two weeks of preoperative medical treatment compared with patients who had received at least one year of medical treatment.
It is often difficult to identify those patients who are at increased risk of surgical failure after trabeculectomy. Our study results suggest that impression cytology may be useful in identifying conjunctival changes related to longterm drug use, and further studies will be needed to determine whether these changes are predictive of surgical outcome.
References 1. Fiore, P. M., Jacobs, I. H., and Goldberg, D. B.: Drug-induced pemphigoid. A spectrum of diseases. Arch. Ophthalmol. 105:1660, 1987. 2. Liesegang, T. J.: Bulbar conjunctival follicles associated with dipivefrin therapy. Ophthalmology
92:228,1985.
3. Johnson, D. H., Kenyon, K. R., Epstein, D. L., and Van Buskirk, E. M.: Corneal changes during pilocarpine gel therapy. Am. J. Ophthalmol. 101:13,
1986.
4. Derous, D., de Keizer, R. J., de WolffRouendaal, D., and Soudijn, W.: Conjunctival keratinization. An abnormal reaction to an ocular betablocker. Acta Ophthalmol. (Copenh.) 67:333, 1989. 5. Gerstenberger, A., and Marquardt, R.: Die becherzelldicte unter pilocarpineinflu[3 [The density of goblet cells modified by pilocarpine]. Fortschr. Ophthalmol. 83:46, 1986. 6. Varga, M., and Follmann, P.: Feinstrukturelle Untersuchungen der Bindehautoberflache nach Langzeitbehandlung mit Timolol [Ultrastructural studies of the conjunctival surface following longterm treatment with timolol]. Fortschr. Ophthalmol.
83:155, 1986.
7. Takahashi, N.: Cytotoxic effects of antiglaucoma
Vol. 112, No.3
Conjunctival Cytology in Treated Glaucoma
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