 |
|
A Study of Iridectomy Histopathologic Features of Latanoprost- and Non–Latanoprost-Treated Patients
Daniel M. Albert, MD, MS;
Ronald E. Gangnon, PhD;
Michele L. Zimbric, BS;
Christine M. Damico, BS;
Marian R. Fisher, PhD;
Joel Gleiser, MD;
Hans E. Grossniklaus, MD;
W. Richard Green, MD
Arch Ophthalmol. 2004;122:1680-1685.
ABSTRACT
| |
Objectives To examine the histopathologic features of iridectomy specimens from patients undergoing glaucoma surgery and to compare histologic abnormalities in a group of patients with a history of latanoprost therapy with those in a group of patients who had no history of prostaglandin therapy (controls).
Methods Iridectomy specimens and patient history forms were submitted to the central Latanoprost Pathology Center. These were independently examined by 3 ophthalmic pathologists in a masked fashion. Specimens were evaluated for malignant, premalignant, and other changes including differences in levels of pigmentation, degrees of cellularity, inflammation, vascular abnormalities, and changes in the iris pigment epithelium.
Results Specimens were received from 449 patients with a history of latanoprost treatment and 142 patients who had no history of treatment with latanoprost or other prostaglandin analogues. No evidence of malignant or premalignant changes was found. In latanoprost-treated irides, the prevalence of iris freckles was higher (P = .001) than in control irides, as was the combined number of stromal fibroblasts and melanocytes (P<.001). In a subgroup of specimens received through June 2002, there was no significant difference in mean melanocyte counts (P=.35) obtained by immunohistochemical staining techniques between the latanoprost-treated and control groups.
Conclusions These findings support previous studies indicating that latanoprost-induced eye color changes are due to an increased amount of melanin within the iris stromal melanocytes. The increased numbers of freckles may be a focal manifestation of this effect.
INTRODUCTION
New glaucoma drugs including latanoprost, isopropyl unoprostone, travoprost, and bimatoprost are associated with increased pigmentation of the iris in some patients.1 Our knowledge of this phenomenon is based primarily on clinical observations, with relatively limited histopathologic investigation.2-3 This study was designed to evaluate, in a systematic manner, key histopathologic findings in iris specimens from patients undergoing glaucoma surgery and to compare the results in groups defined by a history of latanoprost therapy or no history of therapy with latanoprost or any prostaglandin analogues. Assessments of malignant or premalignant changes including the presence of melanoma, increased cell number, and mitotic figures were recorded. Other characteristics evaluated included the degree of pigmentation of the anterior border layer and stroma, inflammation, abnormalities of the stromal blood vessels, and pigment epithelial changes. This is the first large histopathologic study to address the effects of latanoprost on the human iris, including the safety aspects of latanoprost-induced iris pigmentation.
METHODS
Iris specimens, obtained at the time of iridectomy during glaucoma surgery, were received in 10% neutral-buffered formalin at the Latanoprost Pathology Center (LPC) and labeled by the LPC coordinator with a unique study accession number. The formalin-fixed tissue specimens were processed overnight on an automated tissue processor, embedded in paraffin, serially sectioned at 5 µm, and mounted onto treated slides. The first 3 slides were stained with hematoxylin-eosin, the next 3 slides were bleached with potassium permanganate prior to staining with hematoxylin-eosin, and the following 7 slides were left unstained. All slides were labeled with the unique study accession number assigned to the specimen. If any tissue remained after sectioning, the paraffin block was archived.
Sets of microscopic slides (1 hematoxylin-eosin slide, 1 bleached hematoxylin-eosin slide, and 1 unstained slide) were sent to 3 reviewing pathologists (D.M.A., H.E.G., and W.R.G.) who examined the set independently and with no knowledge of therapy history. They completed the accompanying pathology grading form, including an assessment of the quality of the specimens, their location, and their orientation. These forms were returned to the LPC coordinator, who completed a composite grading form using specific rules for combining data from the 3 grading forms.
An iris freckle (ephelis) was identified by the pathologists as a focal area of increased pigmentation in the anterior border layer melanocytes without an increased number of melanocytes.4 This is consistent with the general definition of an iris freckle in the field of eye pathology. Using all specimens received by June 2002 and the techniques previously described,5 1 of the unstained slides was stained with S100a and fluorescein isothiocyanate. This stained slide was examined by a single reviewer (in the laboratory of D.M.A.) who was masked to treatment history and identified melanocytes and other cells (fibroblasts and clump cells) by their staining characteristics, counted them, and entered the counts into a spreadsheet. Specimens were adjudicated using a protocol formulated by the 3 pathologists prior to the beginning of the study.
The patient history form accompanying the specimen included iris color (blue, hazel, or brown); type of glaucoma; ocular history; type of surgery; treatment history, including the start of medical treatment; medication history, including specific medications used; duration of latanoprost therapy (<3 months, 3 months to 3 years, or >3 years); other glaucoma medications used; and presence or absence of increased pigmentation after glaucoma medication. The latanoprost group was defined by a history of latanoprost therapy, and the control group was defined by no history of therapy with latanoprost or any other prostaglandin analogue.
After review by the LPC coordinator, the patient history form and the composite grading form, each identified only by the unique accession number, were sent to the Statistical Data Analysis Center (Department of Biostatistics and Medical Informatics, University of Wisconsin, Madison) for data entry (Oracle database; Oracle, Redwood Shores, Calif) and statistical analysis. The numbers of melanocytes and other cells from the subset of specimens were transferred separately from the laboratory for their inclusion in this article. This study received institutional review board approval from the University of Wisconsin Health Sciences Human Subjects Committee.
Comparisons of groups were based on the Wilcoxon rank sum test for continuous and ordinal variables and the Pearson  2 test for categorical variables. Analyses were performed using analysis of variance models for continuous and ordinal variables and logistic regression models for dichotomous variables, which included sex and iris color as covariates. Statistical significance was set at P<.05. No formal adjustment for multiple comparisons was used.
RESULTS
Between September 1998 and January 2003, 591 iris specimens were received at the LPC through the cooperation of 41 glaucoma surgeons and 22 centers, listed in Table 1. Specimens were received from 449 patients with a history of latanoprost treatment and 142 patients who had no history of treatment with latanoprost or other prostaglandin analogues. The duration of latanoprost treatment was recorded for 346 (77%) of the 449 patients in the latanoprost group; 57 used latanoprost for less than 3 months, 247 for 3 months to 3 years, and 42 for more than 3 years. Table 2 summarizes, by treatment group, relevant information from the patient history form and the year of completion of the pathology grading form. Race and ethnicity, iris color, type of glaucoma, and ocular history were statistically similar for the latanoprost and control groups; however, there were more women in the control group compared with the latanoprost group (P = .01). With regard to the year of completion of the pathology grading form, there was a statistically significant difference (P<.001) in the distribution of specimens by treatment group.
|
|
|
|
Table 1. Surgeons and Centers Contributing Iris Specimens
|
|
|
|
|
|
|
Table 2. Patient History Forms Summary*
|
|
|
Evaluation of the composite pathology grading forms indicated that the latanoprost and control groups were comparable in terms of quality, location, and orientation of the iridectomy specimens (data not shown). All specimens were obtained from the peripheral iris.
No melanomas were reported on the pathology grading form in either the latanoprost or control group (data not shown). No statistically significant differences in the presence of nevi were noted either in all irides or in blue, hazel, or brown irides (Figure 1). There was, however, a statistically significant difference in the presence of freckles in the latanoprost group compared with the control group in all irides (P = .001) and in the hazel (P = .01) and brown (P<.001) irides (Figure 2). This difference remained statistically significant in multivariable logistic regression models, which included sex and iris color (P = .009).
|
|
|
|
Figure 1. Proportion of specimens with nevi by treatment group for all irides and for subgroup based on iris color.
|
|
|
|
|
|
|
Figure 2. Proportion of specimens with freckles by treatment group for all irides and for subgroups based on iris color.
|
|
|
There was a statistically significant difference between treatment groups in the number of combined stromal fibroblasts and melanocytes in all irides (P<.001) and in the brown irides (P<.001). In analyses by sex, there was no statistically significant difference between groups in the number of stromal fibroblasts and melanocytes in the irides of all men (P = .24) or in the irides of brown-eyed men (P = .32); there were statistically significant differences between groups in the irides of all women (P<.001) and in the irides of brown-eyed women (P<.001). (Table 3).
|
|
|
|
Table 3. Grading Form for Stromal Fibroblasts and Melanocytes in the Irides*
|
|
|
No statistically significant differences in the degree of pigmentation in the melanocytes of the anterior border layer or stroma, in the mean thickness, or in many stromal features were noted between the latanoprost and control groups (Table 4). Free pigment granules in the stroma (P = .05) were not statistically significantly different; rubeosis, although rare, was statistically significantly different (P = .002) (Table 4). The proportion of specimens determined to have a normal iris did not differ between the 2 groups (data not shown).
|
|
|
|
Table 4. Pathology Grading Form Summary*
|
|
|
Prior to iridectomy, 21 patients were reported to have darkening of the iris. All of these patients had received latanoprost treatment. In 6 cases, there was photographic documentation of the darkening. Data from patients with clinically observed darkening of the iris were included with data from other latanoprost-treated patients without darkening. No histopathologic abnormalities were seen in these specimens. It is presumed that subclinical or unnoticed eye color changes occurred in additional patients.
Using the subset of specimens received by June 2002, no statistically significant differences in melanocyte counts were noted. However, there was a statistically significant difference in other cell counts (fibroblasts and clump cells) between the latanoprost and control groups for all irides (P = .005; n = 371), brown irides (P = .02; n = 182), and men with blue irides (P = .03; n = 56) (Table 5).
|
|
|
|
Table 5. Melanocyte and Other Cell Counts for Irides*
|
|
|
COMMENT
Differences in the iris color of normal eyes are the result of variable amounts of melanin granules within a constant number of melanocytes in the superficial stroma of the iris.5-8 Iris melanocytes seem to reach their genetically determined amount of melanin in early childhood, and their melanin content usually remains constant in adulthood. Iris color, however, can be affected by a variety of ocular disorders.9
Latanoprost, a phenyl-substituted analogue of prostaglandin F2 , is effective for lowering intraocular pressure. In early trials in the United States, United Kingdom, and Scandinavia, it was associated with increased melanogenesis in the irides of patients treated in clinical trials.10 Typically this change was manifested by a concentric increase in iris pigmentation appearing after about 6 months of treatment.10 The incidence of increased pigmentation is apparently related to the color of the iris and has been reported to range from 5% to 70%.11-12 More recently, other prostaglandin analogues introduced for the treatment of glaucoma have been associated with increased pigmentation of the iris in some individuals.1 Most of the data regarding this phenomenon involve latanoprost, which was the first prostaglandin analogue available for the treatment of glaucoma.
In vitro studies in which human iris tissue or melanocyte cultures were incubated with latanoprost indicate that the iris darkening associated with latanoprost treatment is caused by the induction of tyrosinase expression.13-16 In an additional in vitro study, Dutkiewicz et al17 demonstrated that latanoprost induced tyrosinase activity but did not increase the mitotic index in a panel of human uveal and cutaneous melanoma cell lines. That study also suggested that the adverse effect of latanoprost on in vivo iris pigmentation did not result from increased cell division but from elevated tyrosinase activity.
The cynomolgus monkey has been described as an effective animal model for studying the increased pigmentation of the iris with latanoprost treatment.10, 18-19 Darkening of the iris was noted after about 2 months of treatment, with latanoprost inducing increased pigmentation in sympathectomized eyes. Nevi and freckles on the iris conjunctiva or eyelids were not affected. The authors concluded that latanoprost treatment increased the normal low melanin synthesis in the iridial melanocytes of the cynomolgus monkey. In latanoprost-treated eyes, the amount of eumelanin increased from 3- to 7-fold, whereas the variation of pheomelanin did not exceed 25%.20
Zhan et al21 used Dutch-belted rabbits following unilateral superior cervical ganglionectomy as an experimental model for studying prostaglandin-induced iris color darkening. These investigators demonstrated that sympathetic innervation is required for age-related physiologic darkening of iris color and that prostoglandins may compensate for sympathetic denervation to produce the darkening.
Individual case reports and small series of iris specimens studied histopathologically following darkening from latanoprost treatment have been reported.2-3,22-24 These light and electron microscopic studies indicate that latanoprost-induced eye color change is due to an increased amount of melanin within the iris stromal melanocytes rather than any increase in melanocyte number.
In our study, no evidence of malignant or premalignant changes was observed in either group based on the absence of melanomas, atypical nevi, cells with an atypical appearance, or increased numbers of mitotic figures. There was an increased prevalence of freckles in the latanoprost-treated group. We suggest that the increased number of freckles is a manifestation of focal increased tyrosinase expression and that the same pathogenesis exists for the more diffuse darkening. An extreme of this change may be the latanoprost-associated, diffuse, uniform dark velvet-brown appearance simulating a diffuse iris melanoma, as described by Tsai et al.2 We do not believe that the increase in iris freckles has malignant potential or can lead to any adverse clinical effect on the eye.
There was a statistically significant increase in the number of combined stromal melanocytes and fibroblasts, most strongly evident in brown-eyed women. It should be noted that with the hematoxylin-eosin staining used for this study, stromal melanocytes and fibroblasts cannot be reliably differentiated. Because of this, immunohistochemical staining was performed for the subset of specimens received by June 2002, and no statistically significant difference in melanocyte count was seen in all irides or any subgroup. The combined stromal and fibroblast increase noted in the hematoxylin-eosin–stained slides appears to be due to an excess of fibroblasts and/or clump cells but not melanocytes.
Prostaglandin F2 as well as prostaglandins E1 and E2 has been shown to act as an extracellular factor to regulate cell proliferation.25 Prostaglandin F2 stimulated DNA synthesis and cell proliferation in quiescent Swiss mouse 3T3 cell cultures25-26 and in 1 of 2 clones of 3T3 cells from the National Institutes of Health (Bethesda, Md).26 In addition, prostaglandin F2 can modulate the growth and differentiation of corneal epithelial cells cultured in vitro. Consequently, it is not surprising that an increase in the number of iris fibroblasts and/or clump cells occurs in some latanoprost-treated eyes.27
In summary, our results are consistent with previous findings that latanoprost-induced eye color change is due to an increased amount of melanin within the iris stromal melanocytes and does not involve an increase in melanocyte number. Further examination of several histopathologic characteristics, as noted previously, shows no adverse histopathologic effects in latanoprost-treated irides as compared with controls.
AUTHOR INFORMATION
Correspondence: Daniel M. Albert, MD, MS, Department of Ophthalmology and Visual Sciences, F4/344 Clinical Science Center, 600 Highland Ave, Madison WI 53792-3284 (dalbert{at}wisc.edu).
Submitted for Publication: July 17, 2003; final revision received January 26, 2004; accepted April 21, 2004.
Funding/Support: This study was supported by a grant from Pharmacia Corporation, a Pfizer, Inc, company (New York, NY).
Acknowledgment: We thank Kirsten L. Hope, BA, for her diligent effort as an editor.
Financial Disclosure: None.
Author Affiliations: Department of Ophthalmology and Visual Sciences (Drs Albert and Gleiser and Mss Zimbric and Damico) and Department of Biostatistics and Medical Informatics (Drs Gangnon and Fisher), University of Wisconsin Medical School, Madison; Department of Ophthalmology, Emory University, Atlanta, Ga (Dr Grossniklaus); and Department of Ophthalmology, The Johns Hopkins University, Baltimore, Md (Dr Green).
REFERENCES
| |
1. Stjernschantz JW, Albert DM, Hu D, Drago F, Wistrand PJ. Mechanism and clinical significance of prostaglandin-induced iris pigmentation. Surv Ophthalmol. 2002;47:S162-S175.
2. Tsai JC, Sivak-Callcott JA, Haik BG, Zhang J, McLean IW. Latanoprost-induced iris heterochromia and open-angle glaucoma: a clinicopathologic report. J Glaucoma. 2001;10:411-413.
FULL TEXT
|
WEB OF SCIENCE
| PUBMED
3. Grierson I, Pfeiffer N, Cracknell KP, Appleton P. Histology and fine structure of the iris and outflow system following latanoprost therapy. Surv Ophthalmol. 2002;47:S176-S184.
4. Yanoff M, Fine BS. Ocular Pathology: A Text and Atlas. 2nd ed. Philadelphia, Pa: Harper & Row Publishers; 1982:822.
5. Wilkerson CL, Syed NS, Fisher MR, Robinson NL, Wallow IH, Albert DM. Melanocytes and iris color: light microscopic findings. Arch Ophthalmol. 1996;114:437-442.
FREE FULL TEXT
6. Eagle RC Jr. Iris pigmentation and pigmented lesions: an ultrastructural study. Trans Am Ophthalmol Soc. 1988;86:581-687.
PUBMED
7. Imesch PD, Bindley CD, Khademian Z, et al. Melanocytes and iris color: electron microscopic findings. Arch Ophthalmol. 1996;114:443-447.
FREE FULL TEXT
8. Albert DM, Green WR, Zimbric ML, et al. Iris melanocyte numbers in Asian, African-American, and Caucasian irides. Trans Am Ophthalmol Soc. 2003;101:217-222.
PUBMED
9. Imesch PD, Wallow IH, Albert DM. The color of the human eye: a review of the morphologic correlates and of some conditions that affect iridial pigmentation. Surv Ophthalmol. 1997;41:S117-S123.
10. Wistrand PJ, Stjernschantz J, Olsson K. The incidence and time-course of latanoprost-induced iridial pigmentation as a function of eye color. Surv Ophthalmol. 1997;41(suppl 2):S129-S138.
11. Teus MA, Arranz-Marquez E, Lucea-Suescun P. Incidence of iris colour change in latanoprost treated eyes. Br J Ophthalmol. 2002;86:1085-1088.
FREE FULL TEXT
12. Camras CB, Wax MB, Ritch R, et al. Latanoprost treatment for glaucoma: effects of treating for 1 year and of switching from timolol. Am J Ophthalmol. 1998;126:390-399.
FULL TEXT
|
WEB OF SCIENCE
| PUBMED
13. Lindsey JD, Jones HL, Hewitt EG, Angert M, Weinreb RN. Induction of tyrosinase gene transcription in human iris organ cultures exposed to latanoprost. Arch Ophthalmol. 2001;119:853-860.
FREE FULL TEXT
14. Drago F, Marino A, La Manna C. Alpha-methyl-p-tyrosine inhibits latanoprost-induced melanogenesis in vitro. Exp Eye Res. 1999;68:85-90.
FULL TEXT
|
WEB OF SCIENCE
| PUBMED
15. Hu D-N, Stjernschantz J, McCormick SA. Effect of prostaglandins A2E, E1,F2 and latanoprost on cultured human iridial melanocytes. Exp Eye Res. 2000;70:113-120.
FULL TEXT
|
WEB OF SCIENCE
| PUBMED
16. Stjernschantz J, Ocklind A, Wentzel P, Lake S, Hu DN. Latanoprost-induced increase of tyrosinase transcription in iridial melanocytes. Acta Ophthalmol Scand. 2000;78:618-622.
FULL TEXT
|
WEB OF SCIENCE
| PUBMED
17. Dutkiewicz R, Albert DM, Levin LA. Effects of latanoprost on tyrosinase activity and mitotic index of cultured melanoma lines. Exp Eye Res. 2000;70:563-569.
FULL TEXT
|
WEB OF SCIENCE
| PUBMED
18. Lindquist NG, Larsson BS, Stjernschantz J. Increased pigmentation of iridial melanocytes in primates induced by a prostaglandin analogue. Exp Eye Res. 1999;69:431-436.
FULL TEXT
|
WEB OF SCIENCE
| PUBMED
19. Selen G, Stjernschantz J, Resul B. Prostoglandin-induced iridial pigmentation in primates. Surv Ophthalmol. 1997;41(suppl 2):S125-S128.
20. Prota G, Vincensi MR, Mapolitano A, Selen G, Stjernschantz J. Latanoprost stimulates eumelanogenesis in iridial melanocytes of cynomolgus monkeys. Pigment Cell Res. 2000;13:147-150.
FULL TEXT
|
WEB OF SCIENCE
| PUBMED
21. Zhan GL, Toris CB, Camras CB, Wang YL, Bito LZ. Prostaglandin-induced iris color darkening: an experimental model. Arch Ophthalmol. 1998;116:1065-1068.
FREE FULL TEXT
22. Grierson I, Lee WR, Albert DM. The fine structure of an iridectomy specimen from a patient with latanoprost-induced eye color change. Arch Ophthalmol. 1999;117:394-396.
FREE FULL TEXT
23. Grierson I, Cracknell KP, Pfeiffer N. The iris after prostanoid treatment. Curr Opin Ophthalmol. 2001;12:112-118.
FULL TEXT
| PUBMED
24. Pfeiffer N, Grierson I, Goldsmith H, Hochgesand D, Winkgen-Bohres A, Appleton P. Histological effects in the iris after 3 months of latanoprost therapy: the Mainz 1 study. Arch Ophthalmol. 2001;119:191-196.
FREE FULL TEXT
25. De Asua LJ, Clingan D, Rudland PS. Initiation of cell proliferation in culture mouse fibroblasts by prostaglandin F2alpha. Proc Natl Acad Sci U S A. 1975;72:2724-2728.
FREE FULL TEXT
26. Black FM, Wakelam MJ. Activation of inositol phospholipid breakdown by prostaglandin F2 alpha without any stimulation of proliferation in quiescent NIH-3T3 fibroblasts. Biochem J. 1990;266:661-667.
WEB OF SCIENCE
| PUBMED
27. Conconi MT, Spinazzi R, Tommasini M, Limoli A, Parnigotto PP. Prostaglandin F2 alpha can modulate the growth and the differentiation of bovine corneal epithelial cells cultured in vitro. Ann Anat. 2001;183:567-573.
WEB OF SCIENCE
| PUBMED
 CiteULike  Connotea  Del.icio.us  Digg  Reddit  Technorati  Twitter
What's this?
THIS ARTICLE HAS BEEN CITED BY OTHER ARTICLES
A Study of Histopathological Features of Latanoprost-Treated Irides With or Without Darkening Compared With Non-Latanoprost-Treated Irides
Albert et al.
Arch Ophthalmol 2008;126:626-631.
ABSTRACT
| FULL TEXT
|
