adenoma of the retinal pigment epithelium

ADENOMA OF THE RETINAL PIGMENT EPITHELIUM HlSTOCHEMICAL AND ELECTRON MICROSCOPIC OBSERVATIONS

R A M O N L. F O N T , M.D., LORENZ E. Z I M M E R M A X , M.Ü., AND B E N S. F I N E , M.D.

Washington, D.C.

While neoplastic lesions of the retinal pigment epithelium (RPE) are exceedingly rare, benign reactive proliferations of this tissue are frequently observed as a result of a variety of stimuli, including trauma, retinal detachment, high myopia, disciform macular degeneration, and many others.1

Few case reports of tumors of the RPE dealing mainly with light microscopic studies have been recorded in the literature.2-4 Morris and Henkind5 collected and summarized 14 histologically verified cases from the literature and reported on two additional neoplasms of the pigment epithelium of the iris.

From unpublished observations made on approximately 25 pigmented tumors arising from the retinal and ciliary epithelium on file in the Registry of Ophthalmic Pathology, one type was found to be characterized by a prominent vacuolization of many of the pigmented tumor cells. In this report we describe some histochemical and electron microscopic studies made on one such tumor.

CASE REPORT

A 63-year-old white man complained of decreased vision and flashing lights of two months' duration in his left eye. His visual acuity was RE: 20/40 and LE :20/200. The intraocular tension was normal in each eye. The left eye was enucleated with a diagnosis of intraocular tumor and secondary retinal detachment. The fellow eye was essentially normal.

From the Registry of Ophthalmic Pathology, Armed Forces Institute of Pathology, Washington, D.C. This study was supported in part by Public Health Service Training Grant 5 T01-EY-00032 and Research Grants EY-00397 and EY-00133 from the National Eye Institute.

The opinions or assertions contained herein are the private views of the authors and are not to be construed as social or as reflecting the views of the Department of the Army or the Department of Defense.

Reprint requests to Ramon L. Font, M.D., Armed Forces Institute of Pathology, Washington, D.C. 20305.

Fig. 1 (Font, Zimmerman, and Fine). A jet-black tumor is present between the equator and the ora serra ta (arrows). A secondary retinal detachment with white, coagulated subretinal exu-date is also seen (AFIP Neg. 69-4201-2).

The specimen consisted of a firm left eye measuring 24 X 23 X 22 mm with 1 mm of optic nerve attached. The cornea was clear and measured 11 X 10 mm. Upon transillumination there was a shadow measuring 9 X 10 mm in the superior temporal quadrant. The globe was opened vertically. The anterior chamber was deep and clear, and the angle was open. Superiorly, there was a jet-black tumor that measured 4 x 4 x 3 mm projecting inward from the choroid between the ora serrata and the equator (Fig. 1). The lens appeared normal. The vitreous was partially liquefied. The retina was detached and elevated over the tumor. A small focus of hemorrhage measuring 1 X 1 mm was present in the subretinal fluid adjacent to the tumor. There was peripheral micro-cystoid degeneration of the retina. The optic nerve was normal.

L I G H T MICROSCOPIC OBSERVATIONS

No significant changes were observed in the tissues of the anterior segment. A serous exudate was present in the subretinal space, and the photoreceptor layer of the retina was degenerated. Arising from the retinal pig-

544

VOL. 73, NO. 4 RETINAL PIGMENT EPITHELIUM ADENOMA 545

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Fig. 2 (Font, Zimmerman, and Fine). Under low magnification the pigmented tumor has a flat base toward the sciera (bottom of picture). Cells with vacuolization are more numerous toward the apical portion than toward the base. A focal hemorrhage is present in the vitreous at the posterior edge of the tumor (hematoxylin and eosin, χ28, AFIP Neg. 70-7096).

ment epithelium superiorly behind the ora serrata was a densely pigmented tumor projecting into the subretinal space. The base of the tumor rested upon Bruch's membrane, and there was no invasion of the adjacent choroid. The tumor cells were heavily pigmented and exhibited polygonal outlines with distinct cell borders. Large intracyto-plasmic vacuoles were present throughout the cells of the tumor, more numerous toward its apical portion than at its base (Fig. 2) . No mitotic figures were seen. A slightly basophilic granular material was present in many of the vacuoles. The pigment granules within the cells were large and ellipsoid, resembling those of the normal retinal pigment epithelium. In addition to routine hematoxylin and eosin stains, sections of the tumor were bleached with potassium permanganate to facilitate more detailed study of the cytologie features of the heavily pigmented tumor cells. Bleached preparations revealed

large polyhedral cells with eccentric round to oval nuclei with prominent nucleoli and abundant granular cytoplasm (Fig. 3) . Thin fibrovascular septa containing scattered melanophages were present throughout the tumor. Occasional cystlike spaces were filled with melanophages. The large intracyto-plasmic vacuoles were more readily seen after bleaching of the melanin pigment. A fold of retinal tissue, which partially covered the posterior half of the tumor, contained blood. The adjacent retina showed areas of hemor-rhagic necrosis and contained a blood-filled space that was separated from the vitreous by a thin glial membrane. Vitreous strands were attached to the surface of the tumor anteriorly. Scattered drusen were seen along Bruch's membrane. The choroid and sciera were normal.

Diagnosis—Adenoma ' of retinal pigment epithelium with secondary retinal detachment.

546 AMERICAN JOURNAL OF OPHTHALMOLOGY APRIL, 1972

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Fig. 3 (Font, Zimmerman, and Fine). A bleached preparation reveals large polyhedral cells with eccentric round to oval pyknotic nuclei and abundant granular cytoplasm. Thin fibrovascular septa are present. Large äntracytoplasmic vacuoles are seen in many of the tumor cells (χ!65, AFIP Neg. 70-7088).

Fig. 4 (Font, Zimmerman, and Fine). Thin sections (—■ 1.5 μ) reveal the presence of light and dark cells. The intracelluar vacuoles are mostly localized to the latter cells. Several melanophages are seen in a cys-toid space just above the center of the picture (arrow) (paraphenylenediamine, X24S, AFIP Neg. 70-7091).

VOL. 73, NO. 4 R E T I N A L P I G M E N T E P I T H E L I U M A D E N O M A 547

Fig. S (Font, Zimmerman, and Fine). Intracytoplasmic vacuole ( V ) containing granular material. A prominent basement membrane (BM) is present (uranyl acetate and lead citrate, xll,5Q0, A F I P Neg. 71-2004).

HlSTOCHEMICAL FINDINGS

Several histochemical procedures were used in an effort to characterize the contents of the intracytoplasmic vacuoles of the tumor cells, including periodic acid-Schiff reaction with and without diastase digestion; alcian blue at pH 2.5 and 0.4, alcian blue at pH 2.5 after digestion with hyaluronidase and sialidase, and aldehyde-fuchsin at pH 1.7 and 1.0. The histochemical reactions (Table 1) demonstrated the presence of a nonsulfated, diastase-resistant, partially hyaluronidase-sensitive mucopolysaccharide containing sialic acid.6

Several additional adenomas of the retinal pigment epithelium of the vacuolated type were also studied. In none of the tumors did

TABLE 1 HISTOCHEMICAL REACTIONS OF THE CONTENTS OF THE

I N T R A C Y T O P L A S M I C V A C U O L E S I N A N A D E N O M A O F

THE RETINAL PIGMENT EPITHELIUM

Staining Method

Alcian blue pH2.S p H 0 . 4 pH 2.5 after hyaluronidase

digestion pH 2.5 after sialidase di

gestion

Aldehyde-fuchsin pH 1.7 pH 1.0

Periodic acid-Schiff Before diastase After diastase

Reaction

Positive ( 3 + ) Negative

Weakly positive (1 +) ·

Negative

Negative Negative

Positive ( 3 + ) Positive ( 3 + )


Fig. 7 (Font, Zimmerman, and Fine). Higher magnification of a portion of dark cell shown in Figure 6. Most of the melanin granules are mature melanosomes. Rough-surfaced endoplasmic reticulum is present (uranyl acetate and lead citrate, χ30,000, AFIP Neg. 71-2004).

all of the vacuoles stain, and in some of the tumors, none of the vacuoles stained. Our interpretation of these findings is that some of the material in the vacuoles was lost during fixation and/or subsequent processing of the tissue.

ELECTRON MICROSCOPIC OBSERVATIONS

Formalin-fixed wet tissue of a portion of the tumor that was present in one of the calottes of the eye was processed for electron

microscopy as has been previously reported from our laboratory.7 The sections, embedded in Epon, were stained with uranyl acetate and lead citrate and examined with an RCA 3G electron microscope.

Thin sections (1.5 to 2μ) stained with par-aphenylenediamine revealed the presence of dark and light cells (Fig. 4) . Most of the dark tumor cells showed large intracyto-plasmic vacuoles displacing the pigment granules to one side of the cells, thus ac-

Fig. 6 (Font, Zimmerman, and Fine). The dark cell below contains large numbers of mature melanosomes. Elongated cytoplasmic bridges separate the intracytoplasmic vacuoles. The bridges rupture, leaving short cytoplasmic tags (see inset at upper right, X 15,000). The light cell, right, above, contains very few mature melanosomes (uranyl acetate and lead citrate, χ7,500, AFIP Neg. 71-2004).


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Fig. 8 (Font, Zimmerman, and Fine). Another tumor cell showing mixture of immature (early and late) and mature me'annsomes. Premelanosomes were not observed (uranyl acetate and lead citrate, χ22,000, AFIP Neg. 71-2004).


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Fig. 9 (Font, Zimmerman, and Fine). A light cell has produced a basement membrane (arrow) as well as fine collagen filaments (CF) (uranyl acetate and lead citrate, χ20,000, AFIP Neg. 71-2004).

counting for their dark appearance. The light tumor cells did not contain vacuoles and showed a more even distribution of the melanin granules.

Electron microscopy revealed, in addition to intracellular vacuolization (Fig. 5), the presence of a granular material within the vacuoles. These vacuoles had apparently coalesced by thinning and rupture of cyto-plasmic bridges (Fig. 6) , seen occasionally on the edges of the vacuoles as retracted, round, short cytoplasmic tags (inset, Fig. 6) . The vacuoles, as they enlarged, appeared to compress the melanin granules peripherally, producing in part the dark appearance of the cells as seen in thicker sections. In addition, we observed that the pigment granules of the dark cells were predominantly mature melanosomes.8 The dark cells also contained prominent rough-surfaced endoplasmic re-ticulum (Fig. 7) , a finding believed to be in

dicative of metabolically more active cells capable of elaborating mature melanin granules but also capable of forming secretory vacuoles containing the mucopolysaccharide previously described. The light cells contained mostly immature (early and late) or incompletely developed melanosomes and very few mature melanosomes.8 Some tumor cells contained an almost equal mixture of immature and mature melanosomes (Fig. 8) . No pre-melanosomes, as may be seen in human em-byronic pigment epithelium,8 were observed in either the dark or the light cells.

The pigmented tumor cells produced basement membranes (Fig. 9) as well as fine collagen filaments. They were attached to each other by desmosomes (Fig. 10) and junc-tional complexes, the latter including both zonula occludens and zonula adherens (Fig. H ) ·

Tn areas where the apical portions of adja-


Fig. 10 (Font, Zimmerman, and Fine). The tumor cells are attached to each other by desmosomes (arrows). Fine collagen filaments are present between the tumor cells (CF) (uranyl acetate and lead

citrate, X 16,500, AFIP Neg. 71-2004).

cent cells converged, short microvilli were commonly present (inset, Fig. 11). An occasional cell showed a reversion of the normal polarity, with the zonula adherens located toward the apical portion of the cell (near the microvilli).

COMMENTS

Light and electron microscopic study of the tumor cells of an adenoma of the retinal pigment epithelium revealed both similarities and differences as compared with the normal retinal pigment epithelium. Both cell types have basement membranes, junctional complexes (i.e., zonulae occludens and adherens), and microvilli. Although no desmosomes or fine collagen filaments are seen in the plane of the normal retinal pigment epithelium, both were present in the adenoma of the retinal pigment epithelium. Both findings have also been observed in prolifer

ated retinal pigment epithelium.9'10 Of great interest was the presence in the tumor cells of secretory intracellular vacuoles containing a sialomucin. Mucopolysaccharides containing sialic acid (sialomucin) are present in normal lacrimal gland tissue,11·12 intestinal mucosa, and other epithelia. The significance of this finding in an adenoma of the retinal pigment epithelium is unknown. The exact localization of mucopolysaccharide containing sialic acid in the intraocular structures is uncertain. Farkas and co-workers13 reported the presence of a mucopolysaccharide containing sialic acid in drusen. Additional h'stochemical studies are indicated to confirm their results and to elucidate the functional significance, if any, of the presence of a sialomucin in the normal human eye. It is also possible that the pigment epithelial cells may normally contribute a small quantity of sialomucin to the interreceptor mucoid matrix.


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Fig. 11 (Font, Zimmerman, and Fine). Another tumor cell shows junctional complexes containing a zonula occludens (ZO) and a zonula adherens-like configuration (ZA). Numerous microvilli are present at places where the apical portions of several cells meet (inset, χ20,000) (uranvl acetate and lead citrate, χ21,000, AFIPNeg. 71-2004).

SUMMARY

The enucleated left eye of a 63-year-old white man contained a jet-black mass that was classified histopathologically as an adenoma of the retinal pigment epithelium.

The tumor was composed of heavily pig-mented, large polyhedral cells containing in-tracellular vacuoles. Histochemical stains demonstrated the presence of a nonsulfated diastase-resistant mucopolysaccharide containing sialic acid (sialomucin) in these vacuoles.

Sections 1.5 to 2μ thick were examined by light microscopy and revealed dark and light cells. When examined by electron microscopy the dark cells that contained intracyto-plasmic vacuoles showed a predominance of mature melanosomes as compared with the light cells, which did not have vacuoles but did contain a predominance of immature melanosomes.

Electron microscopic studies showed the

presence of basement membranes and fine collagen filaments, desmosomes, and junctional complexes with formation of microvilli by the tumor cells. The intracellular vacuoles containing the mucopolysaccharides enlarged by coalescence of smaller vacuoles and subsequent rupture of cytoplasmic bridges. Some of the tumor cells revealed both immature and mature melanosomes, but no premelanosomes were observed.

ACKNOWLEDGMENT

This case was contributed to the Registry of Ophthalmic Pathology by Thomas P. Stratford, M.D., who kindly gave his permission for its publication.

REFERENCES

1. Kurz, G. H., and Zimmerman, L. E. : Vagaries of the retinal pigment epithelium. Int. Ophth. Clin. 2:441, 1962.

2. Greer, C. H. : Epithelial tumors of the retinal pigment epithelium. Trans. Ophth. Soc. U.K. 72: 265, 1952.

3. Reese, A. B.: Tumors of the Eye, 2nd ed.


New York, Harper and Row, Hoeher Medical Division, 1963, p. 45.

4. Garner, A. : Tumors of the retinal pigment epithelium. Brit. J. Ophth. 54:715, 1970.

5. Morris, D. A., and Henkind, P. : Neoplasms of the iris pigment epithelium. Am. J. Ophth. 66:31, 1968.

6. Johnson, W. C, and Helwig, E. B. : Histochemistry of the acid mucopolysaccharides of skin in normal and in certain pathologic conditions. Am. J. Clin. Path. 40:123, 1963.

7. Fine, B. S., Ts'o, M. O. M., Font, R. L., and Zimmerman, L. E. : Electron microscopy of pathologic ocular tissues. Int. Ophth. Clin. In press.

8. Mund, M. L., Rodrigues, M. M., and Fine, B. S. : Light and electron microscopic observations of the developing pigmented layers of the human eye. Am. J. Ophth. In Press.

9. Ts'o, M. O. M., Fine, B. S., and Zimmerman, L. E. : Photic maculopathy produced by the indirect ophthalmoscope. I. Clinical and histopathologic study. Am. J. Ophth. In press.

10. Wallow, I. H. L., and Ts'o, M. O. M. : Pathology of proliferated retinal pigment epithelium in degenerative retinopathy. Am. J. Ophth. In press.

11. Jensen, O. A., Falbe-Hansen, L, Jacobsen, T., and Michelsen, A. : Mucosubstances of the acini of the human lacrimal gland (orbital part). I. Histo-chemical identification. Acta Ophth. 47:605, 1969.

12. Falbe-Hansen, I., Jensen, O. A., and Degn, J. K. : Mucosubstances of the acini of the human lacrimal gland (orbital part). II. Biochemical identification. Acta Ophth. 47:620, 1969.

13. Farkas, T. G., Sylvester, V., Archer, D., and Altona, M. : The histochemistry of drusen. Am. J. Ophth. 71:1206, 1971.

O P H T H A L M I C MINIATURE

The careful measurements of the curvature of the cornea, first made by Senff of Dorpat, next, with a better adapted instrument, the writer's ophthalmometer, and afterwards carried out in numerous cases by Bonders, Knapp, and others, have proved that the cornea of most human eyes is not a perfectly symmetrical curve, but is variously bent in different directions. I have also devised a method of testing the "centering" of an eye during life, i.e. ascertaining whether the cornea and crystalline lens are symmetrically placed with regard to their common axis. By this means I discovered in the eyes I examined slight but distinct deviations from accurate centering. The result of these two defects of construction is the condition called astigmatism which is found in more or less all human eyes, and prevents our seeing vertical and horizontal lines at the same distance perfectly clearly at once. If the degree of astigmatism is excessive, it can be obviated by the use of glasses with cylindrical surfaces, a circumstance which has lately much attracted the attention of oculists.

Hermann von Helmholtz Popular Lectures on Scientific Subjects

Longmans, Green, 1893

adenoma of the retinal pigment epithelium

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