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Spontaneous Congenital Iris Cyst
S P O N T A N E O U S CONGENITAL IRIS CYST FREDERICK H.
ROY, M.D.,
A ND C A L V I N H A N N A , P H . D .
Little Rock, Arkansas This paper presents a case of spontaneous congenital iris cyst. I n addition, the micro scopic, ultramicroscopic, autoradiographic, and free amino acid findings of the cyst are also reported. C A S E REPORT
This Negro boy was born to a 20-year-old woman, gravida 3, para 3, by forceps delivery on March 19, 1969. His birth weight was 3.17 kg and he had an Apgar rating of 10 at five minutes. On examination, the infant was adjudged to be normal and he was discharged at three days of age. He was readmitted eight weeks of age with a diagnosis of diarrhea and dehydration. At that time, the left eye, which had remained closed for the previous three weeks, revealed an abnormality. An examination re vealed a normal right eye and a bilobated, slightly turbid, pigmented cystic structure which occupied about the lower half of the anterior chamber of the left eye. It touched the endothelium of the lower cornea and was present on the surface of the iris. Over a period of two weeks, the cyst appeared to fill 75% of the anterior chamber (Fig. 1). The hori zontal corneal diameter in both eyes was 11 mm, while the intraocular pressure (Schiotz) was two scale readings higher in the left eye than in the right. Under general anesthesia (halothane), a limbal-based conjunctival flap was laid inferior from four to six o'clock. A syringe with a 27-gauge nee dle was used to aspirate 100 ml of fluid from the cyst through a 4 mm incision of the limbus. Smooth, non-toothed forceps were used to bring the cyst out through the opening. The cyst appeared to be attached to the inferior mid-portion of the iris and a sector iridectomy was made inferiorly to re move the attachment of the cyst with the iris. The cyst was not attached to the pupillary margin and there was no lens defect. Following surgery, the in ferior cornea of the left eye was hazy and by three months after surgery the inferior cornea was slightly cloudy, the intraocular pressure was 7/7 (7.5) mm Hg and a small semitransparent mass From the Departments of Ophthalmology and Pharmacology, Veterans Administration Hospital and University of Arkansas Medical Center, Little Rock, Arkansas. This investigation was supported in part by USPHS Research Grants EY—00239 and EY—00270 and NB—10041 from the Na tional Eye Institute. Reprint requests to Frederick H. Roy, M.D., Veterans Administration Hospital, 300 East Roose velt, Little Rock, Arkansas.
Fig. 1 (Roy and Hanna). Left eye, with bilobated, slightly turbid and pigmented cystic structure oc cupying lower 75% of anterior chamber. Light at superior temporal limbus is artifact. was noted at six o'clock with a contact lens. One year later this mass was not further enlarged. M A T E R I A L S A N D METHODS
O n removal of the cyst, the tissue was im mersed in a solution of tissue culture medium ( T C - 1 9 9 ) containing 1 ixCi/ml of 3 H-thymidine and 5u.CI/ml of 3 H-l-leucine both of sp. act 5 Ci/mmole.* After one hour's incubation at 37°C, the tissue was fixed for one day in freshly prepared 4 % formaldehyde. T h e tis sue was cut into six pieces and stained with osmium tetroxide and then embedded in epoxy (Araldite) plastic for microscopic, ultramicroscopic, and autoradiographic stud ies. T h e thick sections (0.1 to 4\i) were usu ally stained with toluidine blue, the thin sec tions (about 0.06 [j.) were stained with uranyl acetate and lead citrate, and for autoradiography thick sections were coated with N T B 2 Kodak liquid emulsion. 1 T h e cyst fluid sample was precipitated with 1 5 % ( w / v ) aqueous sulfosalicylic acid, centrifuged and subjected * Nuclear-Chicago Corp., Des Plaines, Illinois.
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Fig. 3 (Roy and Hanna). Normal iris tissue adjacent to iris cyst which contains pigmented epithelial cells on the posterior surface lower right corner (PE), smooth muscle cells in center (M), and pigmented stromal cells, upper left (toluidin blue, X800).
Fig. 2 (Roy and Hanna). Iris cyst wall with two to three layers of nonpigmented epithelial cells outer most with one or two layers of pigmented stromal cells (toluidine blue, X900).
to ion exchange chromatography, using the Beckman Spinco Model 120C amino acid ana lyzer.2 RESULTS
The cyst wall consisted of two or three layers of nonpigmented epithelial cells on the anterior side overlying one or two layers of pigmented cells (Fig. 2). The cyst was at tached to the anterior surface of the iris be low which were located normal appearing iris cells (Fig. 3). On the posterior side, remnants of the embryonic ciliary processes were found at the root of the iris (Fig. 4).
The epithelial cells covering the cyst took on many shapes. The innermost epithelial cells contained larger round nuclei and they closely resembled the basal epithelial cells of the cornea. No goblet or mucin cells were found. The amino acid leucine is incorporated al most exclusively into newly synthesized pro tein. To estimate the relative rates of overall protein synthesis by various cell types, the cyst was incubated for a short period of time in leucine with a radioactive tracer ( 3 H-leucine). The autoradiograph was used to esti mate the relative uptake of the radioactive
Fig. 4 (Roy and Hanna). Posterior surface of iris containing villi projecting into the posterior chamber These villi appear to be remnants of the embryonic ciliary process that may extend to the posterior surface of the iris. The surface of the villi are covered with nonpigmented epithelial cells. The outermost epi thelial cells are covered by microvilli. Inside the villi are brightly red-staining collagen fibers (the red stain is so light that it appears in this photograph as white). Below the collagen layer are pigmented stromal cells. Further toward the pupil, the nonpigmented epithelial cells and villi disappear and become pigmented epithelial cells on the posterior side of the iris (basic f uchsin and methylene blue, X800).
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Fig. 5 (Roy and Hanna). The number of reduced silver grains (black dots) overlying the incorporating H-leucine into protein was higher for epithelial cells than for the pigmented stromal cells (inferior portion of photograph). In this autoradiographic photograph the focus is on the silver grains, and as a result the possibly confusing melanin granules are out of focus (autoradiogram stained with toluidine blue, X900). 3
leucine taken up into protein. The cells of the cyst incorporated 3H-leucine into protein (Figs. 5 and 6 ) . There was a relatively high grain count from the autoradiogram over the individual epithelial cells of the cyst that was similar to that of normal epithelial cells of the cornea.3 The free surface of the epithelial cells contained microvilli (Figs. 7-10). The cyto plasm of the epithelial cells contained vari ous types of junctions, i.e., desmosomes, tight and gap junctions (Figs. 8-11), and intracellular filaments (Figs. 7-14). Similar junctions between stromal cells or epithelial and stromal cells were not noted (Figs. 11, 12, and 15) and the extracellular space be tween the epithelial and stromal cells con tained banded filaments (Figs. 12-13). The pigmented stromal cells of the cyst contained nuclei that were of varying shapes (Figs. 2, 12, and 15). These cells incorpo rated 3H-leucine into protein to give a low grain count in comparison with the epithelial cells of the cyst (Figs. 5, 6). These stromal cells incorporated 3H-leucine at a rate com parable to the stromal cells of the cornea.3 The pigmented stromal cells were sur
rounded by collagen fibrils (Figs. 12, 13, and 15). These pigmented stromal cells did not have the characteristics of the pigment ep ithelial layer found on the posterior surface of the iris (Fig. 3). A piece of the iris that was tightly attached to the cyst was removed with the cyst. This iris tissue contained two layers of pigmented epithelial cells on the posterior surface, several layers of nonpig mented epithelial cells over the anterior sur face and heavily pigmented and lightly pig mented (stromal) cells in the remainder of the tissue along with muscle cells (dilator iris) above the pigmented epithelium (Fig. 3). Only a few mitotic figures were observed in the sections of the cyst. Only an occa sional basal epithelial cell, and rarely a stro mal cell, of the cyst incorporated 3 H-thymidine into newly forming DNA. This indi cates that a slow rate of cell division was taking place in the cells of the cyst compared to basal epithelial cells of the normal cornea that are undergoing a moderate rate of cell turnover.3 The piece of iris tissue adjacent to the cyst contained several layers of nonpigmented
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Fig. 6 (Roy and Hanna). The number of reduced silver grains over the endothelial cells surrounding a blood vessel (B) that had incorporated 3H-leucine into protein was similar to the number of reduced silver grains over the surface epithelium (top) overlying the iris (autoradiogram stained with toluidine blue, X900).
epithelial cells on the anterior surface and two layers of heavily pigmented epithelial cells on the posterior surface. Between these layers were pigmented stromal cells. Heavily and lightly pigmented stromal cells were
present beneath the epithelial layer (Fig. 15). These stromal cells were not generally in contact with adjacent cells. Collagen fibrils were found between the stromal cells and no cell junctions or tonofibrils were found.
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Fig. 7 (Roy and Hanna). The surface epithelial cells of the cyst contain striated or brush borders or microvilli (V). These microvilli are covered with plethora of delicate branching filaments (F) which forms an almost continuous coating. These filaments appear to be continuous with the unit membrane of the microvilli (X45,000).
Fig. 9 (Roy and Hanna). Epithelila cells of cyst sectioned through area shown in Figure 2. Note the microvilli on the right side X 15,000). The anterior chamber fluid contains a full spectrum of free amino acids similar to that found in the peripheral blood. The cyst it self, being an almost avascular sphere, may contain a fluid that has limited nutrients. The concentration of free amino acids in the cyst fluid was low. Free amino acids are presented in Table 1. With the exception of histidine and cystine,
the concentration of free amino acids in the aqueous humor of the adult human2 was in all cases higher than that of the cyst. COMMENT
T h e origin of spontaneous iris cysts is still controversial. 4 " 7 Since both the cyst from the iris stroma and from the neural epithelium may occur in the stroma of the iris, Duke-
Fig. 8 (Roy and Hanna). Junction between two epithelial cells, left center, demonstrating desmosome (macula adherens) (D) surrounded by prominent tonofibrils. Part of a microvillus (V) with branching filaments (F) can be seen in upper right corner. Dark black spots throughout the cell are ribosomes (X 100,000).
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Fig. 10 (Roy and Hanna). Sur face epithelium of the cyst contain unusually long microvilli and typical microvilli, cell junctions (tight junc tion, zonula occluden) (T), and many ribosomes (X4S.000).
Elder 4 states that "differentiation between them on clinical grounds is impossible, and on histological grounds is difficult, the exis tence and separation of the two types rest more upon a theoretical than on an observa tional basis." Even with electron microscopic studies, we feel that one cannot positively determine how the nonpigmented epithelial cells arrived over the stromal cells of the cyst. T h e epithelial cells covering the cyst
and part of the iris appear to be typical ep ithelial cells from the standpoint of ultrastructure with numerous polysomes, tonofibrils, desmosomes, and the high rate of in corporation of leucine into protein compared to adjacent stromal cells. T h e pigmented fibrocytes of the cyst contain intracellular collagen fibrils of various sizes. These cells could have a similar origin as did the stromal cells of the iris to which the cyst was at-
Fig. 12 (Roy and Hanna). Lack of contact between pigment stromal cells on lower right and nonpig mented epithelial cells on upper left. The upper right shows collagen (C) fibrils (X 15,000).
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Fig. 11 (Roy and Hanna). Epithelial cells adjacent to pigmented cells or iris cyst from Figure 9, illustrat ing desmosomes with apparently innerconnecting tonofibrils (T) (Xl5,000).
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Fig. 13 (Roy and Hanna). Extracellular space surrounding a pigment stromal cell of Figure 12 at upper left, illustrating pigmentcd vesicles (P) and collagen fibrils (C) (X30,000).
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Fig. 15 (Roy and Hanna). Pigmented stromal cells near junction of iris and cyst near the pupil showing lack of cell junctions. Instead these cells are surrounded by extracellular collagen fibrils (X 5,000).
tached. The resultant of the uptake and in corporation of 3H-leucine into protein by pigmented cells of the cyst and stromal cells of the noncystic part of the iris were also similar. Several possibilities should be considered in the histogenesis of the cyst: It may derive from (1) pigment epithelium, (2) conjunctival epithelium, (3) ectopic lacrimal gland ductal epithelium, (4) from nonpigmented neuroepithelium, or (5) from iris stroma. Since the epithelial layer of this cyst lacks pigment granules, mucous glands (goblet cells),6 and glandular tissue, the first three possibilities are unlikely. The most likely histogenesis of this spontaneous congenital
cyst is nonpigmented neuroepithelium cov ering stromal cells derived from the iris. Fine 7 reported a free-floating pigmented cyst in the anterior chamber of a 22-yearold man. In his patient, the cyst was composed of pigmented epithelial cells that were delineated on one side by a basement membrane and contained diastase sensitive granules (glycogen deposits). In the study reported here, neither pigmented epithelial cells, a basement membrane, nor glycogen deposits were detected in the cells of the cyst. The extent and length of the microvilli covering the epithelial cells to the cyst were extraordinary. These microvilli were visible
Fig. 14 (Roy and Hanna). Enlarged view of filaments and the rounded ribosomes from an epithelial cell (X 100,000).
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TABLE 1
SUMMARY
COMPARISON OF AMINO ACID CONCENTRATIONS ( M G / L I T E R ) IN AQUEOUS FLUID OF 19 ADULTS JUST PRIOR TO CATARACT EXTRACTION AND IN THE FLUID OF THE CYST
A spontaneous cyst in the anterior chamber and attached to the iris was found in a n eight-week-old infant. Microscopic, ultramicroscopic, and autoradiographic features of the cyst are presented. T h e cyst wall is composed of several layers of pigmented stromal cells covered by multiple layers of nonpigmented epithelial cells. T h e surface epithelial cells were covered with unusually long microvilli and there was no evidence of goblet cells. A n occasional epithelial cell was observed to be undergoing D N A synthesis and a few mitotic figures were found; how ever, these cells readily incorporated radio active leucine into protein. The free amino acid content of the cyst fluid was low com pared to the free amino acids in the aqueous fluid. W e concluded that the cyst developed from the anterior surface of the iris and the cells covering it are derived from the nonpig mented neuroepithelium.
A.
T3.U.
0.708 0.330 0.250 0.180 0.129 0.127 0.123 0.110 0.101 0.098 0.093 0.061 0.056 0.048 0.040 0.021 0.012
0.059 0.032 0.017 0.040 0.063 0.025 0.031 0.025 0.017 0.050 0.021 0.019 0.021 0.033 0.015 0.014
Aqueous Aqueous 1. Asparagine and glutamine 2. Alanine 3. Valine 4. Serine 5. Threonine 6. Leucine 7. Lysine 8. Arginine 9. Phenylalanine 10. Isoleucine 11. Tyrosine 12. Taurine 13. Histidine 14. Glycine 15. Ornithine 16. Glutamic acid 17. Aspartic acid 18. 2-amino-isobutyric acid 19. Methionine 20. Tryptophane 21. Proline 22. Citrolline 23. J Cystine 5 Cystine and urea
0.011 0.027 0.002 0.002 0.002 0.001 2.714
— —
0.009
— — — —
0.785
LySt
Fluid 0.0001 0.0001 0.0043
— —
0.0004
— — —
0.0003 0.0001 0.0007 0.0660 0.0004
— —
0.0001
—
0.0001
— — —
0.0043 0.0080
under microscopic observation. Such villi have not previously been reported for iris cysts. T h e presence of very low free amino acids in the fluid of the cyst, compared to the normal anterior chamber fluid, suggests that the cyst contained no opening to the anterior chamber, or at the most, only a small open ing. The microvilli are similar in structure to the absorptive cells of the small intestine 8 and the concentration of nutrients inside the cyst might be expected to be low. It is of in terest that in the presence of a very low free amino acid concentration, the cells lining the inside of the cyst were viable. Possibly this is due to the fact that the pigmented cells lin ing the cyst are undergoing a low rate of protein synthesis as indicated by the rela tively small uptake and incorporation of 3 H leucine into the protein of these cells.
ACKNOWLEDGMENTS
Jerome Sherman and Catherine Dodd, Depart ment of Anatomy, aided in the electron microscopic observation, and Juan Sanchez, Department of Ophthalmology, determined the free amino acid composition of the cyst. We wish to acknowledge their contributions to this study. REFERENCES
1. Hanna, C, and O'Brien, J. E.: Lens epithe lium cell proliferation and migration in radiation cataracts. Radiat. Res. 19:1, 1963. 2. Sanchez, J., and Hanna, C.: Effects of acetazolamide on concentration of free amino acids in aqueous fluid. T. Arkansas Med. Soc. 66:144, 1969. 3. Hanna, C, Blocker, D., and Allen, J. H.: RNA and protein synthesis by cells of human eye bank corneas. Ann. Ophth. 1:53, 1969. 4. Duke-Elder, S.: System of Ophthalmology, vol. 3. St. Louis, Mosby, 1965, pt. 2, p. 603. 5. Vail, D. T., and Merz, E. H.: Embryonic intraepithelial cyst of the ciliary processes. Tr. Am. Ophth. Soc. 49:167, 1952. 6. Naumann, G., and Green, W. R.: Spontaneous non-pigmented iris cysts. Arch. Ophth. 78:496, 1967. 7. Fine, B. S.: Free-floating pigmented cyst in the anterior chamber. Am. J. Ophth. 67:493, 1969. 8. Brunser, O., and Luft, J. H.: Fine structure of the apex of absorption cells from rat small intes tine. J. Ultrastr. Res. 31:291, 1970.
ROY, M.D.,
A ND C A L V I N H A N N A , P H . D .
Little Rock, Arkansas This paper presents a case of spontaneous congenital iris cyst. I n addition, the micro scopic, ultramicroscopic, autoradiographic, and free amino acid findings of the cyst are also reported. C A S E REPORT
This Negro boy was born to a 20-year-old woman, gravida 3, para 3, by forceps delivery on March 19, 1969. His birth weight was 3.17 kg and he had an Apgar rating of 10 at five minutes. On examination, the infant was adjudged to be normal and he was discharged at three days of age. He was readmitted eight weeks of age with a diagnosis of diarrhea and dehydration. At that time, the left eye, which had remained closed for the previous three weeks, revealed an abnormality. An examination re vealed a normal right eye and a bilobated, slightly turbid, pigmented cystic structure which occupied about the lower half of the anterior chamber of the left eye. It touched the endothelium of the lower cornea and was present on the surface of the iris. Over a period of two weeks, the cyst appeared to fill 75% of the anterior chamber (Fig. 1). The hori zontal corneal diameter in both eyes was 11 mm, while the intraocular pressure (Schiotz) was two scale readings higher in the left eye than in the right. Under general anesthesia (halothane), a limbal-based conjunctival flap was laid inferior from four to six o'clock. A syringe with a 27-gauge nee dle was used to aspirate 100 ml of fluid from the cyst through a 4 mm incision of the limbus. Smooth, non-toothed forceps were used to bring the cyst out through the opening. The cyst appeared to be attached to the inferior mid-portion of the iris and a sector iridectomy was made inferiorly to re move the attachment of the cyst with the iris. The cyst was not attached to the pupillary margin and there was no lens defect. Following surgery, the in ferior cornea of the left eye was hazy and by three months after surgery the inferior cornea was slightly cloudy, the intraocular pressure was 7/7 (7.5) mm Hg and a small semitransparent mass From the Departments of Ophthalmology and Pharmacology, Veterans Administration Hospital and University of Arkansas Medical Center, Little Rock, Arkansas. This investigation was supported in part by USPHS Research Grants EY—00239 and EY—00270 and NB—10041 from the Na tional Eye Institute. Reprint requests to Frederick H. Roy, M.D., Veterans Administration Hospital, 300 East Roose velt, Little Rock, Arkansas.
Fig. 1 (Roy and Hanna). Left eye, with bilobated, slightly turbid and pigmented cystic structure oc cupying lower 75% of anterior chamber. Light at superior temporal limbus is artifact. was noted at six o'clock with a contact lens. One year later this mass was not further enlarged. M A T E R I A L S A N D METHODS
O n removal of the cyst, the tissue was im mersed in a solution of tissue culture medium ( T C - 1 9 9 ) containing 1 ixCi/ml of 3 H-thymidine and 5u.CI/ml of 3 H-l-leucine both of sp. act 5 Ci/mmole.* After one hour's incubation at 37°C, the tissue was fixed for one day in freshly prepared 4 % formaldehyde. T h e tis sue was cut into six pieces and stained with osmium tetroxide and then embedded in epoxy (Araldite) plastic for microscopic, ultramicroscopic, and autoradiographic stud ies. T h e thick sections (0.1 to 4\i) were usu ally stained with toluidine blue, the thin sec tions (about 0.06 [j.) were stained with uranyl acetate and lead citrate, and for autoradiography thick sections were coated with N T B 2 Kodak liquid emulsion. 1 T h e cyst fluid sample was precipitated with 1 5 % ( w / v ) aqueous sulfosalicylic acid, centrifuged and subjected * Nuclear-Chicago Corp., Des Plaines, Illinois.
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Fig. 3 (Roy and Hanna). Normal iris tissue adjacent to iris cyst which contains pigmented epithelial cells on the posterior surface lower right corner (PE), smooth muscle cells in center (M), and pigmented stromal cells, upper left (toluidin blue, X800).
Fig. 2 (Roy and Hanna). Iris cyst wall with two to three layers of nonpigmented epithelial cells outer most with one or two layers of pigmented stromal cells (toluidine blue, X900).
to ion exchange chromatography, using the Beckman Spinco Model 120C amino acid ana lyzer.2 RESULTS
The cyst wall consisted of two or three layers of nonpigmented epithelial cells on the anterior side overlying one or two layers of pigmented cells (Fig. 2). The cyst was at tached to the anterior surface of the iris be low which were located normal appearing iris cells (Fig. 3). On the posterior side, remnants of the embryonic ciliary processes were found at the root of the iris (Fig. 4).
The epithelial cells covering the cyst took on many shapes. The innermost epithelial cells contained larger round nuclei and they closely resembled the basal epithelial cells of the cornea. No goblet or mucin cells were found. The amino acid leucine is incorporated al most exclusively into newly synthesized pro tein. To estimate the relative rates of overall protein synthesis by various cell types, the cyst was incubated for a short period of time in leucine with a radioactive tracer ( 3 H-leucine). The autoradiograph was used to esti mate the relative uptake of the radioactive
Fig. 4 (Roy and Hanna). Posterior surface of iris containing villi projecting into the posterior chamber These villi appear to be remnants of the embryonic ciliary process that may extend to the posterior surface of the iris. The surface of the villi are covered with nonpigmented epithelial cells. The outermost epi thelial cells are covered by microvilli. Inside the villi are brightly red-staining collagen fibers (the red stain is so light that it appears in this photograph as white). Below the collagen layer are pigmented stromal cells. Further toward the pupil, the nonpigmented epithelial cells and villi disappear and become pigmented epithelial cells on the posterior side of the iris (basic f uchsin and methylene blue, X800).
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Fig. 5 (Roy and Hanna). The number of reduced silver grains (black dots) overlying the incorporating H-leucine into protein was higher for epithelial cells than for the pigmented stromal cells (inferior portion of photograph). In this autoradiographic photograph the focus is on the silver grains, and as a result the possibly confusing melanin granules are out of focus (autoradiogram stained with toluidine blue, X900). 3
leucine taken up into protein. The cells of the cyst incorporated 3H-leucine into protein (Figs. 5 and 6 ) . There was a relatively high grain count from the autoradiogram over the individual epithelial cells of the cyst that was similar to that of normal epithelial cells of the cornea.3 The free surface of the epithelial cells contained microvilli (Figs. 7-10). The cyto plasm of the epithelial cells contained vari ous types of junctions, i.e., desmosomes, tight and gap junctions (Figs. 8-11), and intracellular filaments (Figs. 7-14). Similar junctions between stromal cells or epithelial and stromal cells were not noted (Figs. 11, 12, and 15) and the extracellular space be tween the epithelial and stromal cells con tained banded filaments (Figs. 12-13). The pigmented stromal cells of the cyst contained nuclei that were of varying shapes (Figs. 2, 12, and 15). These cells incorpo rated 3H-leucine into protein to give a low grain count in comparison with the epithelial cells of the cyst (Figs. 5, 6). These stromal cells incorporated 3H-leucine at a rate com parable to the stromal cells of the cornea.3 The pigmented stromal cells were sur
rounded by collagen fibrils (Figs. 12, 13, and 15). These pigmented stromal cells did not have the characteristics of the pigment ep ithelial layer found on the posterior surface of the iris (Fig. 3). A piece of the iris that was tightly attached to the cyst was removed with the cyst. This iris tissue contained two layers of pigmented epithelial cells on the posterior surface, several layers of nonpig mented epithelial cells over the anterior sur face and heavily pigmented and lightly pig mented (stromal) cells in the remainder of the tissue along with muscle cells (dilator iris) above the pigmented epithelium (Fig. 3). Only a few mitotic figures were observed in the sections of the cyst. Only an occa sional basal epithelial cell, and rarely a stro mal cell, of the cyst incorporated 3 H-thymidine into newly forming DNA. This indi cates that a slow rate of cell division was taking place in the cells of the cyst compared to basal epithelial cells of the normal cornea that are undergoing a moderate rate of cell turnover.3 The piece of iris tissue adjacent to the cyst contained several layers of nonpigmented
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Fig. 6 (Roy and Hanna). The number of reduced silver grains over the endothelial cells surrounding a blood vessel (B) that had incorporated 3H-leucine into protein was similar to the number of reduced silver grains over the surface epithelium (top) overlying the iris (autoradiogram stained with toluidine blue, X900).
epithelial cells on the anterior surface and two layers of heavily pigmented epithelial cells on the posterior surface. Between these layers were pigmented stromal cells. Heavily and lightly pigmented stromal cells were
present beneath the epithelial layer (Fig. 15). These stromal cells were not generally in contact with adjacent cells. Collagen fibrils were found between the stromal cells and no cell junctions or tonofibrils were found.
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Fig. 7 (Roy and Hanna). The surface epithelial cells of the cyst contain striated or brush borders or microvilli (V). These microvilli are covered with plethora of delicate branching filaments (F) which forms an almost continuous coating. These filaments appear to be continuous with the unit membrane of the microvilli (X45,000).
Fig. 9 (Roy and Hanna). Epithelila cells of cyst sectioned through area shown in Figure 2. Note the microvilli on the right side X 15,000). The anterior chamber fluid contains a full spectrum of free amino acids similar to that found in the peripheral blood. The cyst it self, being an almost avascular sphere, may contain a fluid that has limited nutrients. The concentration of free amino acids in the cyst fluid was low. Free amino acids are presented in Table 1. With the exception of histidine and cystine,
the concentration of free amino acids in the aqueous humor of the adult human2 was in all cases higher than that of the cyst. COMMENT
T h e origin of spontaneous iris cysts is still controversial. 4 " 7 Since both the cyst from the iris stroma and from the neural epithelium may occur in the stroma of the iris, Duke-
Fig. 8 (Roy and Hanna). Junction between two epithelial cells, left center, demonstrating desmosome (macula adherens) (D) surrounded by prominent tonofibrils. Part of a microvillus (V) with branching filaments (F) can be seen in upper right corner. Dark black spots throughout the cell are ribosomes (X 100,000).
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Fig. 10 (Roy and Hanna). Sur face epithelium of the cyst contain unusually long microvilli and typical microvilli, cell junctions (tight junc tion, zonula occluden) (T), and many ribosomes (X4S.000).
Elder 4 states that "differentiation between them on clinical grounds is impossible, and on histological grounds is difficult, the exis tence and separation of the two types rest more upon a theoretical than on an observa tional basis." Even with electron microscopic studies, we feel that one cannot positively determine how the nonpigmented epithelial cells arrived over the stromal cells of the cyst. T h e epithelial cells covering the cyst
and part of the iris appear to be typical ep ithelial cells from the standpoint of ultrastructure with numerous polysomes, tonofibrils, desmosomes, and the high rate of in corporation of leucine into protein compared to adjacent stromal cells. T h e pigmented fibrocytes of the cyst contain intracellular collagen fibrils of various sizes. These cells could have a similar origin as did the stromal cells of the iris to which the cyst was at-
Fig. 12 (Roy and Hanna). Lack of contact between pigment stromal cells on lower right and nonpig mented epithelial cells on upper left. The upper right shows collagen (C) fibrils (X 15,000).
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Fig. 11 (Roy and Hanna). Epithelial cells adjacent to pigmented cells or iris cyst from Figure 9, illustrat ing desmosomes with apparently innerconnecting tonofibrils (T) (Xl5,000).
106
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JULY, 1971
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Fig. 13 (Roy and Hanna). Extracellular space surrounding a pigment stromal cell of Figure 12 at upper left, illustrating pigmentcd vesicles (P) and collagen fibrils (C) (X30,000).
VOL. 72, NO. 1
SPONTANEOUS CONGENITAL IRIS CYST
107
Fig. 15 (Roy and Hanna). Pigmented stromal cells near junction of iris and cyst near the pupil showing lack of cell junctions. Instead these cells are surrounded by extracellular collagen fibrils (X 5,000).
tached. The resultant of the uptake and in corporation of 3H-leucine into protein by pigmented cells of the cyst and stromal cells of the noncystic part of the iris were also similar. Several possibilities should be considered in the histogenesis of the cyst: It may derive from (1) pigment epithelium, (2) conjunctival epithelium, (3) ectopic lacrimal gland ductal epithelium, (4) from nonpigmented neuroepithelium, or (5) from iris stroma. Since the epithelial layer of this cyst lacks pigment granules, mucous glands (goblet cells),6 and glandular tissue, the first three possibilities are unlikely. The most likely histogenesis of this spontaneous congenital
cyst is nonpigmented neuroepithelium cov ering stromal cells derived from the iris. Fine 7 reported a free-floating pigmented cyst in the anterior chamber of a 22-yearold man. In his patient, the cyst was composed of pigmented epithelial cells that were delineated on one side by a basement membrane and contained diastase sensitive granules (glycogen deposits). In the study reported here, neither pigmented epithelial cells, a basement membrane, nor glycogen deposits were detected in the cells of the cyst. The extent and length of the microvilli covering the epithelial cells to the cyst were extraordinary. These microvilli were visible
Fig. 14 (Roy and Hanna). Enlarged view of filaments and the rounded ribosomes from an epithelial cell (X 100,000).
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TABLE 1
SUMMARY
COMPARISON OF AMINO ACID CONCENTRATIONS ( M G / L I T E R ) IN AQUEOUS FLUID OF 19 ADULTS JUST PRIOR TO CATARACT EXTRACTION AND IN THE FLUID OF THE CYST
A spontaneous cyst in the anterior chamber and attached to the iris was found in a n eight-week-old infant. Microscopic, ultramicroscopic, and autoradiographic features of the cyst are presented. T h e cyst wall is composed of several layers of pigmented stromal cells covered by multiple layers of nonpigmented epithelial cells. T h e surface epithelial cells were covered with unusually long microvilli and there was no evidence of goblet cells. A n occasional epithelial cell was observed to be undergoing D N A synthesis and a few mitotic figures were found; how ever, these cells readily incorporated radio active leucine into protein. The free amino acid content of the cyst fluid was low com pared to the free amino acids in the aqueous fluid. W e concluded that the cyst developed from the anterior surface of the iris and the cells covering it are derived from the nonpig mented neuroepithelium.
A.
T3.U.
0.708 0.330 0.250 0.180 0.129 0.127 0.123 0.110 0.101 0.098 0.093 0.061 0.056 0.048 0.040 0.021 0.012
0.059 0.032 0.017 0.040 0.063 0.025 0.031 0.025 0.017 0.050 0.021 0.019 0.021 0.033 0.015 0.014
Aqueous Aqueous 1. Asparagine and glutamine 2. Alanine 3. Valine 4. Serine 5. Threonine 6. Leucine 7. Lysine 8. Arginine 9. Phenylalanine 10. Isoleucine 11. Tyrosine 12. Taurine 13. Histidine 14. Glycine 15. Ornithine 16. Glutamic acid 17. Aspartic acid 18. 2-amino-isobutyric acid 19. Methionine 20. Tryptophane 21. Proline 22. Citrolline 23. J Cystine 5 Cystine and urea
0.011 0.027 0.002 0.002 0.002 0.001 2.714
— —
0.009
— — — —
0.785
LySt
Fluid 0.0001 0.0001 0.0043
— —
0.0004
— — —
0.0003 0.0001 0.0007 0.0660 0.0004
— —
0.0001
—
0.0001
— — —
0.0043 0.0080
under microscopic observation. Such villi have not previously been reported for iris cysts. T h e presence of very low free amino acids in the fluid of the cyst, compared to the normal anterior chamber fluid, suggests that the cyst contained no opening to the anterior chamber, or at the most, only a small open ing. The microvilli are similar in structure to the absorptive cells of the small intestine 8 and the concentration of nutrients inside the cyst might be expected to be low. It is of in terest that in the presence of a very low free amino acid concentration, the cells lining the inside of the cyst were viable. Possibly this is due to the fact that the pigmented cells lin ing the cyst are undergoing a low rate of protein synthesis as indicated by the rela tively small uptake and incorporation of 3 H leucine into the protein of these cells.
ACKNOWLEDGMENTS
Jerome Sherman and Catherine Dodd, Depart ment of Anatomy, aided in the electron microscopic observation, and Juan Sanchez, Department of Ophthalmology, determined the free amino acid composition of the cyst. We wish to acknowledge their contributions to this study. REFERENCES
1. Hanna, C, and O'Brien, J. E.: Lens epithe lium cell proliferation and migration in radiation cataracts. Radiat. Res. 19:1, 1963. 2. Sanchez, J., and Hanna, C.: Effects of acetazolamide on concentration of free amino acids in aqueous fluid. T. Arkansas Med. Soc. 66:144, 1969. 3. Hanna, C, Blocker, D., and Allen, J. H.: RNA and protein synthesis by cells of human eye bank corneas. Ann. Ophth. 1:53, 1969. 4. Duke-Elder, S.: System of Ophthalmology, vol. 3. St. Louis, Mosby, 1965, pt. 2, p. 603. 5. Vail, D. T., and Merz, E. H.: Embryonic intraepithelial cyst of the ciliary processes. Tr. Am. Ophth. Soc. 49:167, 1952. 6. Naumann, G., and Green, W. R.: Spontaneous non-pigmented iris cysts. Arch. Ophth. 78:496, 1967. 7. Fine, B. S.: Free-floating pigmented cyst in the anterior chamber. Am. J. Ophth. 67:493, 1969. 8. Brunser, O., and Luft, J. H.: Fine structure of the apex of absorption cells from rat small intes tine. J. Ultrastr. Res. 31:291, 1970.