KERATINIZATION OF THE CONJUNCTIVA* BY A. Edward Maumenee, MD THE CORNEAL
EPITHELIUM IS A FIVE-TO-SIX-LAYERED STRATIFIED COLUMNAR
squamous nonkeratinized tissue. In contrast, the conjunctiva contains many mucous goblet cells and is two-to-three layers thick, with cuboidal basal cells. If the cornea is denuded the epithelium is replaced from the conjunctiva by migration in about seven days. Over a period ofabout three weeks the epithelium undergoes a metaplasia from conjunctiva to corneal epithelium.1.2 The factors which produce this metaplasia are unknown. If there are superficial blood vessels in the corneal stroma, the migrating cells retain their conjunctival architecture, including the goblet cells.3 The mysteries of morphogenesis, organ formation, and histogenesis (tissue stratification) have not been solved, but some interesting observations have been made.4 Many of these findings are based on studies of embryonic tissues, especially those of the chicken. An excellent example is the regulation of epidermal differentiation and the formation ofkeratinized cells into the stratum corneum. The architecture of the epidermis varies in thickness of layers and degree of keratinization from one area of the body to another, but is relatively constant for a given region. The degree of keratinization probably does not depend on direct stimulation or initiation of keratin synthesis, but rather on maintenance of a regularly organized basal layer and of a harmonious stratification. The stimuli may be divided into dermal and external factors. THE EXTERNAL FACTORS
In 1953 Fell and Mellanby5 showed that exoplants of skin from six-toseven-day-old chick embryos in tissue culture could be converted from keratinized epithelium containing mucous cells and, in some instances, ciliated cells, by the addition of excessive amounts ofvitamin A to the tissue culture media. When these cultures were returned to media without the addition of vitamin A the tissues converted back to keratinized epithelium *From The
Wilmer Institute, The Johns Hopkins Hospital, Baltimore, Maryland 21205.
TR. AM. OPHTH. Soc. vol. LXXVII, 1979
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Maumenee
in four to five days. Lasnitzkif in 1958 showed that vitamin A prevents keratinization of human fetal skin cultured in vitro. True mucoid metaplasia, however, does not occur although some mucous is produced in discrete locations by the epithelial cells. Metaplasia has also been produced on the ear skin of adult guinea pigs by direct repeated applications of vitamin A.7 This transformation, however, is not stable, for after eight days the skin strata return to normal keratinogenic activity even with continuous daily treatment. In contrast it has been found that hydrocortisone, epidermal growth factor, prostaglandins, and thyroid hormone may enhance keratinization. 4 THE DERMAL INFLUENCE
The dermal influence on the epidermal layer has been studied extensively in vitro using chick embryos, exoplants, and tissue culture. Skin pieces have been split into epidermal and dermal constituents by treating them with trypsin solution. The epidermis is then recombined with the dermis in various tissue culture media. Isolated epidermal grafts to chick chorioallantoic membrane degenerate within five days by pyknosis of the malpighian layer and disappearance of the granular layer. However, when the epidermis is recombined with either its own dermis or that of another species, differentiation occurs with the formation of keratinized cells and a stratum corneum. This is true for chicken, human, and other mammalian tissues. The dermal factor for keratinogenesis decreases with increasing age of the embryo. Adult dermis is much less efficient in supporting epidermal histogenesis than is embryonic tissue. However, the dermis obtained in the region of basal cell epitheliomas, epidermoid carcinomas, and cicatricial lesions more closely resembles embryonic dermis in its keratinizing potency. The dermal factor is active when the epidermis is recombined either to the right-side-up or the inverted dermis, which means that the dermal factor is diffusible and present throughout the thickness of the dermis and that its mode of action is not polarized. In contrast, it is not effective on the epidermis when its keratinized surface, rather than its base, is placed on the dermis. The keratinizing factor of the dermis is destroyed by heat and collagenase, but not when the tissue is freeze-killed. In spite of the fact that the dermal factor does not appear to be species specific, Billingham and Silvers8 have shown that it does exert a regionspecific influence. Thus, the epidermis of the sole of the foot, the ear, and the trunk of the guinea pig contain architecturally different epidermis.
Keratinization of Conjuctiva
135
When the relatively thin superficial epidermis of the trunk is recombined with the dermis from the sole, a thick, fully stratified epidermis characteristic of the footpad differentiates. Other combinations of these tissues confirm the regional specificity of the dermal factor. Further evidence of the multi-potential of the epithelial cells, which would indicate that they are not genetically coded for a specific region, are the findings of Sun9 and his coworkers. They have shown by the indirect immunofluorescent test that stratified squamous epithelia (cornea, conjunctiva, tongue, esophagus, vagina and anus) and the epidermal appendages (hair follicles, sebaceous glands, and myoepithelial cells ofthe sweat gland) contain abundant keratins. No staining by the antisera was observed in fibroblasts, muscle cells ofany type, cartilage, blood vessels, nerve tissue, iris or lens epithelia, or glomerular or tubular cells of the kidney. In contrast, the antisera stained the cells of the intestinal tract, urinary tract, female genital tract, and respiratory tract. Likewise, when corneal epithelial cells and dermal epithelial cells were cultured in vitro in identical media, their keratin content was almost the same when observed by electron-microscopy and tested by immunofluorescence. Electrophoresis also showed a similar banding of the keratinoproteins. Billingham and Silvers8 noted that the epithelia of noncutaneous origin, such as from the tongue, esophagus, and cheek pouch ofthe hamster, could not be transformed to produce a stratum corneum by contact with skin dermis, but maintained their specific histological structure. These studies would appear to be somewhat in contrast to the observations of Sun and Green9 and also to the fact that the chorionic epithelium of four-to-six-day chick embryos when combined with nondermal tissue, such as lung, gizzard, or heart, produced no keratin but undergoes a metaplastic change towards construction of cilia and synthesis of mucus. 10 METAPLASIA OF OPHTHALMIC EPITHELIA
It was previously mentioned that the mucous membrane structure of the conjunctival epithelium undergoes a metaplasia to stratified squamous columnar epithelium when it grows over normal corneal stroma, but that superficial vascularization of the stroma prevented this conversion. It is also interesting to note that even a few subconjunctival collagen fibrils under a thin conjunctival flap through which a patient might see will prevent the conjunctival tissue from undergoing metaplasia into corneal epithelia (Figs 1 and 2). A further example of the influence of the subepithelial tissue is the prolonged localized production of keratinized epithelium when a piece of
Maumenee
136
.....
.::
FIGURE 1
Thin conjunctival flap covering a cornea which had been opacified by chemical burn.
skin is transplanted to the globe to prevent the recurrence of pterygium (Fig 3). Similar localized keratinization occurs when skin is grafted into conjunctival mucous membrane to reconstruct a socket after enucleation. KERATINIZATION OF BULBAR AND TARSAL CONJUNCTIVAL EPITHELIUM
It was cited previously that the comeal and conjunctival epithelium had the potential for developing keratin fibrils comparable to those found in the epithelia of the skin in suitable tissue culture. There are a number of clinical instances where tarsal and bulbar conjunctiva may produce keratinized cells. The most frequent of these is exposure or drying, as in ectropion of the lid, staphyloma of the globe, or lagophthalmos. In some instances this is merely a drying of the superficial epithelial cells and not a true metaplasia of the epithelial layer. More interesting are those conditions in which there are adequate tears, but the tissue undergoes a true metaplasia to form a stratum granulosum and corneum. The most common example of this is vitamin A deficiency
Keratinization of Conjuctiva
137
.. ...
.... ...
FIGURE 2
Histopathology of surface conjunctival epithelium removed from eye in Fig 1 at the time of a centrally placed penetrating keratoplasty.
FIGURE 3
Skin graft to conjunctiva performed by plastic surgeon to prevent recurrence of pterygium.
138
Maumenee
and the development of xerophthalmia. Although such cases are seldom seen in the United States today, it is still one of the principal causes of blindness in developing countries. Another frequent cause of true metaplasia is scar formation after trachoma, Stevens-Johnson disease, irradiation ofthe lids, and infections in patients with atopic dermatitis. In some otherwise normal individuals, chronic irritation from topical medications may produce keratinizing metaplasia ofthe tarsal conjunctiva. This may occur either from an allergy to the drug from such medications as sulfonamides or penicillin or from drug irritation without obvious evidence of allergy. The worst offender of this latter type of reaction was furtrethonium iodide (Furmethide), which was used during the 1940s and 1950s to lower intraocular pressure (Fig 4). A high percentage of patients who used this drug over a period of a year or more developed stenosis of the lacrimal canaliculi and keratinization of the tarsal conjunctiva. Other offenders of this latter type have been pilocarpine and epinephrine. The conjunctiva overlying tumors of both the lid and the globe may
FIGURE
4
Keratinization and stenosis of lacrimal puncta following the use of furtrethonium iodide (Furmethide) for the treatment of glaucoma.
Keratinization of Conjuctiva
139
FIGURE 5
Keratinization of tarsal conjunctiva overlying sebaceous cell carcinoma of lid.
develop true keratinizing epithelium. Some examples of these are intraepithelial epithelioma (Bowen's-like changes in the conjunctiva), squamous cell carcinoma, epithelial hyperplasia of the limbus, and sebaceous cell carcinomas of the lid (Fig 5). It is interesting that the bulbar conjunctiva appears to be affected more in vitamin A deficiency. In contrast, drug irritation, trachoma, and Stevens-Johnson disease are more likely to involve primarily the tarsal conjunctiva. A very interesting type of keratinization occurs in superior limbic keratoconjunctivitis. Light and electron-microscopic examinations of biopsies ofthis lesion have shown keratinization ofthe superficial epithelial cells and keratohyaline granules in the deeper layers of the cells."1 CLINICAL SIGNIFICANCE
I have been particularly interested in isolated areas of keratinization of the tarsal conjunctiva.12 This is a relatively rare condition that occurs after
140
Maumenee
Stevens-Johnson syndrome, drug reactions, and irradiation of the lids. When the lesion comes into contact with the corneal epithelium, it produces a superficial punctate keratitis and if allowed to persist over a long period of time, superficial vascularization and scarring of the cornea may occur. I have previously reported that the lesions recurred if the keratinized tissue was superficially removed. However, deep excision of the lesion and repair of the area with a mucous membrane graft from the mouth results in a permanent cure with relief of irritating symptoms. I have followed some patients treated in this manner for a period of 20 years without evidence of recurrence of the lesion. More recently, I have observed that the keratinized plaque may disappear spontaneously if the topical medication which has caused the lesion is discontinued. Approximately three years ago I was attempting to destroy the lashes with cryotherapy, according to the technique described by Beard,13 in a patient who had trichiasis and keratinization of the tarsal conjunctiva as a result ofan allergy to drug therapy. The cryoprobe was applied to the area of keratinization and when the tissue healed the superficial tissue was no longer keratinized. Since that time, five other eyes have been treated with a light application of the cryoprobe to areas of long-standing keratinization of the lid, with either a resultant total obliteration of the keratinized tissue or marked improvement of the lesion. Four of these patients had had Stevens-Johnson syndrome, and one patient had atopic dermatitis and secondary corneal ulcers. The gratifying response of the keratinized tissue of the lids to cryotherapy makes it interesting to speculate whether a similar result would occur if superficial limbic keratoconjunctivitis was treated by freezing after the conjunctiva had been lifted from the globe by the injection of fluid. SUMMARY AND CONCLUSIONS
There are many examples that one may cite where the eye or its adnexa may be used as a "test tube" to study biological phenomena. The interesting morphological and biochemical metaplasia of the conjunctiva which occurs when it migrates across the cornea is an excellent example of this. A number of years ago Billingham and Medawar"4 reported that the corneal epithelium did not alter its histologic structure when transplanted to the abdomen of an experimental animal. The observation of Sun9 and his coworkers that conjunctival and corneal epithelium when grown in tissue culture will produce keratin fibrils comparable to those found in
Keratinization of Conjuctiva
141
skin epithelium, and the observation that under certain pathologic conditions both the bulbar and tarsal conjunctiva may undergo true keratinization, strongly suggests that these cells have the potential for multiple differentiation, rather than being terminal elements in development. The shedding of keratinized cells onto the globe, or the rubbing of such cells onto the cornea when the keratinization is localized on the lids, can produce chronic irritation. Excision of the lesions and replacement with mucous membrane, or destruction of the keratinized tissue by freezing, will relieve these symptoms. SUMMARY
Keratinizaton of the tarsal conjunctiva in an eye with adequate tears occurs following a number of conditions, including irritation and sensitivity to topical medications, Stevens-Johnson syndrome, radiation to the lid, and occasionally from unknown causes. The keratinized cells produce an epithelial keratitis with subsequent vascularization of the cornea. The morphogenesis of this condition is discussed, along with therapy including mucous membrane grafts and freezing of the tissue. REFERENCES
1. Friedenwald JS: Growth pressure and metaplasia of conjunctival and corneal epithelium. Doc Ophthalmol V-VI:184-192, 1951. 2. Thoft R, Friend J: Biochemical transformation of regenerating ocular surface epithelium. Invest Ophthalmol 16:14-20, 1977. 3. Maumenee AE, Scholz RO: The histopathology of ocular lesions produced by sulphur and nitrogen mustards. Bull Johns Hopkins Hosp 82:121-147, 1948. 4. Sengel P: Morphogenesis of the Skin. Cambridge Univ Press, Cambridge, England, 1976. 5. Fell HB, Mellanby E: Metaplasia produced in cultures of chick ectoderm by high vitamin A. J Physiol 119:470-488, 1953. 6. Lasnitzki I: Effect of carcinogens, hormones, and vitamins on organ cultures. Inter Rev Cytol 70:79-121, 1958. 7. Wolff HH, Christophers E, Braun-Falco 0: Beeinflussung der epidermalen Ausdifferenzierung durch Vitamin A Saure. Eine electronenmikroskopische Untersuchung. Arch Klin Exp Derm 237:774-795, 1970. 8. Billingham RE, Silvers WK: Studies on the conservation of epidermal specificities of the skin and certain mucosas in adult mammals. J Exp Med 125:429-446, 1967. 9. Sun TT, Green H: Immunofluorescent staining of keratin fibers in cultured cells. Cell 14:469-476, 1978. 10. McLoughlin CB: The importance of mesenchymal factors in the differentiation of chick epidermis. II. Modification of epidermal differentiation by contact with different types of mesenchyme. J Embryol Exp Morphol 9:385409, 1961. 11. Donshik PC, Collin HB, Foster CS, et al: Conjunctival resection treatment and ultrastructural histopathology of superior limbic kerato-conjunctivitis. Am J Ophthalmol 85:101-110, 1978.
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12. Maumenee AE: Keratitis secondary to keratinization of the tarsal conjunctiva. Am J Ophthalmol 41:477-487, 1956. 13. Beard C: Ptosis, ed 2. St Louis, CV Mosby, 1976. 14. Billingham RE, Medawar PB: A note on the specificity of corneal epithelium. J Anat 84:50-56, 1950.
DISCUSSION
DR ARNALL PATZ. In reference to the blood vessels emanating from the limbus, there is new work in terms of growth factors in serum. I wonder if in Doctor Maumenee's review of this work, if there have been any specific studies on the role of serum or epidermal transformation that might occur in metaplasia of the epithelium. DR HAROLD GIFFORD. I would like to ask a question about pemphigoid. Where does pemphigoid fit into this picture? I had a patient with ocular pemphigoid. The lids were treated with cryotherapy to get rid of the lashes which were irritating the cornea. The freezing removed the lashes but made the conjunctival scarring worse. One eye lost the fornix completely and the other one was mostly epidermalized with early scarring of the cornea. I just wonder what is the connection with pemphigoid and if there is anything you know that can be done for this terrible disease? DR ALBERT RUEDEMANN, JR. I think this is a very interesting paper for a number of reasons, and I think this business of squamous metaplasia going over the lid margins is very remarkable but it is reversible. We reported at this meeting some years ago a ten-year study using scleral lenses with adherent lids, squamous metaplasia and Stevens-Johnson's disease and pemphigus. We treated these patients with scleral lenses by merely cutting the lids free from the globe and inserting the lens on the table. We allowed the tissue to regrow and actually got return of normal conjunctival and tear flow. DR MYRON YANOFF. Doctor Maumenee and Doctor Patz' comments bring up the question of what really is keratinization, specifically Doctor Patz' question about serum. Some years ago we did organ cultures of corneal buttons and showed that if serum was not used in the culture medium, most of the time, after about four or five days, the epithelium became keratinized. (Yanoff M: In vitro biology of corneal epitheliun and endothelium. Trans Am Ophthalmol Soc 73:571-620, 1975.) If serum was used, the epithelium did not become keratinized, even after a month in organ culture.
DR RALPH HAMILTON. I think this is a very useful thing, and I would like to ask Doctor Maumenee exactly what light freezing is. Did you use a thermocouple in this and how much freezing do you require for this? DR A EDWARD MAUMENEE. I am very sorry that Doctor Crowell Beard was not able to attend the meeting because of illness, and I wish him a rapid recovery. I would like to thank those discussants who spoke extemporaneously. In answer to the first
Keratinization of Conjuctiva
143
question, I did not find anything in the literature specifically relating serum to keratinization. It was mentioned in the experiments on tissue culture ofembryonic skin that the nutrients in the tissue culture were extremely important in carrying out these experiments. As far as ocular pemphigus or pemphigoid is concerned, it has been my impression that keratinization of the tarsal conjunctiva was extremely rare in this condition during the early stage of the disease. It is only in the end stage when there is marked drying of the conjunctiva and exposure of the tissue that keratinization occurs. It has also been my experience that anything that is done to the tissue to traumatize it, such as freezing, incising it, or injuring it in any way, tends to cause the disease to progress and become worse. In answer to Doctor Ruedemann's comments, I did mention in the written paper that I had several patients in whom there was a spontaneous regression of the keratinization of the tarsal conjunctiva. I did not use a contact lens on these patients, nor had they used a contact lens prior to the development of their keratinization. Certainly chronic irritation from Furmethide or pilocarpine or all9rgy to various sulfonamides will produce this metaplasia of the tissue. All of the patients I treated with the freezing technique had had long-standing alterations of the tissue that had not cleared spontaneously. Doctor Hamilton's question was a good one. I did not use a thermocouple. I merely applied the cryoprobe to produce a superficial freezing of the tissue and as soon as it turned white, the tissue was allowed to thaw. The cryoprobes that I used were those that we routinely use at Wilmer for cataract extraction or retinal detachment surgery. These were the mechanical cryoprobes and not the disposable ones. As far as the tissue culture experiments that Doctor -Yanoff mentioned, I had not been aware that the addition of serum alone would cause a metaplasia of the cultured material. I wonder if in his tissue cultures there was an absence or deficiency in vitamin A content of the tissue culture material before the serum was added.
EPITHELIUM IS A FIVE-TO-SIX-LAYERED STRATIFIED COLUMNAR
squamous nonkeratinized tissue. In contrast, the conjunctiva contains many mucous goblet cells and is two-to-three layers thick, with cuboidal basal cells. If the cornea is denuded the epithelium is replaced from the conjunctiva by migration in about seven days. Over a period ofabout three weeks the epithelium undergoes a metaplasia from conjunctiva to corneal epithelium.1.2 The factors which produce this metaplasia are unknown. If there are superficial blood vessels in the corneal stroma, the migrating cells retain their conjunctival architecture, including the goblet cells.3 The mysteries of morphogenesis, organ formation, and histogenesis (tissue stratification) have not been solved, but some interesting observations have been made.4 Many of these findings are based on studies of embryonic tissues, especially those of the chicken. An excellent example is the regulation of epidermal differentiation and the formation ofkeratinized cells into the stratum corneum. The architecture of the epidermis varies in thickness of layers and degree of keratinization from one area of the body to another, but is relatively constant for a given region. The degree of keratinization probably does not depend on direct stimulation or initiation of keratin synthesis, but rather on maintenance of a regularly organized basal layer and of a harmonious stratification. The stimuli may be divided into dermal and external factors. THE EXTERNAL FACTORS
In 1953 Fell and Mellanby5 showed that exoplants of skin from six-toseven-day-old chick embryos in tissue culture could be converted from keratinized epithelium containing mucous cells and, in some instances, ciliated cells, by the addition of excessive amounts ofvitamin A to the tissue culture media. When these cultures were returned to media without the addition of vitamin A the tissues converted back to keratinized epithelium *From The
Wilmer Institute, The Johns Hopkins Hospital, Baltimore, Maryland 21205.
TR. AM. OPHTH. Soc. vol. LXXVII, 1979
134
Maumenee
in four to five days. Lasnitzkif in 1958 showed that vitamin A prevents keratinization of human fetal skin cultured in vitro. True mucoid metaplasia, however, does not occur although some mucous is produced in discrete locations by the epithelial cells. Metaplasia has also been produced on the ear skin of adult guinea pigs by direct repeated applications of vitamin A.7 This transformation, however, is not stable, for after eight days the skin strata return to normal keratinogenic activity even with continuous daily treatment. In contrast it has been found that hydrocortisone, epidermal growth factor, prostaglandins, and thyroid hormone may enhance keratinization. 4 THE DERMAL INFLUENCE
The dermal influence on the epidermal layer has been studied extensively in vitro using chick embryos, exoplants, and tissue culture. Skin pieces have been split into epidermal and dermal constituents by treating them with trypsin solution. The epidermis is then recombined with the dermis in various tissue culture media. Isolated epidermal grafts to chick chorioallantoic membrane degenerate within five days by pyknosis of the malpighian layer and disappearance of the granular layer. However, when the epidermis is recombined with either its own dermis or that of another species, differentiation occurs with the formation of keratinized cells and a stratum corneum. This is true for chicken, human, and other mammalian tissues. The dermal factor for keratinogenesis decreases with increasing age of the embryo. Adult dermis is much less efficient in supporting epidermal histogenesis than is embryonic tissue. However, the dermis obtained in the region of basal cell epitheliomas, epidermoid carcinomas, and cicatricial lesions more closely resembles embryonic dermis in its keratinizing potency. The dermal factor is active when the epidermis is recombined either to the right-side-up or the inverted dermis, which means that the dermal factor is diffusible and present throughout the thickness of the dermis and that its mode of action is not polarized. In contrast, it is not effective on the epidermis when its keratinized surface, rather than its base, is placed on the dermis. The keratinizing factor of the dermis is destroyed by heat and collagenase, but not when the tissue is freeze-killed. In spite of the fact that the dermal factor does not appear to be species specific, Billingham and Silvers8 have shown that it does exert a regionspecific influence. Thus, the epidermis of the sole of the foot, the ear, and the trunk of the guinea pig contain architecturally different epidermis.
Keratinization of Conjuctiva
135
When the relatively thin superficial epidermis of the trunk is recombined with the dermis from the sole, a thick, fully stratified epidermis characteristic of the footpad differentiates. Other combinations of these tissues confirm the regional specificity of the dermal factor. Further evidence of the multi-potential of the epithelial cells, which would indicate that they are not genetically coded for a specific region, are the findings of Sun9 and his coworkers. They have shown by the indirect immunofluorescent test that stratified squamous epithelia (cornea, conjunctiva, tongue, esophagus, vagina and anus) and the epidermal appendages (hair follicles, sebaceous glands, and myoepithelial cells ofthe sweat gland) contain abundant keratins. No staining by the antisera was observed in fibroblasts, muscle cells ofany type, cartilage, blood vessels, nerve tissue, iris or lens epithelia, or glomerular or tubular cells of the kidney. In contrast, the antisera stained the cells of the intestinal tract, urinary tract, female genital tract, and respiratory tract. Likewise, when corneal epithelial cells and dermal epithelial cells were cultured in vitro in identical media, their keratin content was almost the same when observed by electron-microscopy and tested by immunofluorescence. Electrophoresis also showed a similar banding of the keratinoproteins. Billingham and Silvers8 noted that the epithelia of noncutaneous origin, such as from the tongue, esophagus, and cheek pouch ofthe hamster, could not be transformed to produce a stratum corneum by contact with skin dermis, but maintained their specific histological structure. These studies would appear to be somewhat in contrast to the observations of Sun and Green9 and also to the fact that the chorionic epithelium of four-to-six-day chick embryos when combined with nondermal tissue, such as lung, gizzard, or heart, produced no keratin but undergoes a metaplastic change towards construction of cilia and synthesis of mucus. 10 METAPLASIA OF OPHTHALMIC EPITHELIA
It was previously mentioned that the mucous membrane structure of the conjunctival epithelium undergoes a metaplasia to stratified squamous columnar epithelium when it grows over normal corneal stroma, but that superficial vascularization of the stroma prevented this conversion. It is also interesting to note that even a few subconjunctival collagen fibrils under a thin conjunctival flap through which a patient might see will prevent the conjunctival tissue from undergoing metaplasia into corneal epithelia (Figs 1 and 2). A further example of the influence of the subepithelial tissue is the prolonged localized production of keratinized epithelium when a piece of
Maumenee
136
.....
.::
FIGURE 1
Thin conjunctival flap covering a cornea which had been opacified by chemical burn.
skin is transplanted to the globe to prevent the recurrence of pterygium (Fig 3). Similar localized keratinization occurs when skin is grafted into conjunctival mucous membrane to reconstruct a socket after enucleation. KERATINIZATION OF BULBAR AND TARSAL CONJUNCTIVAL EPITHELIUM
It was cited previously that the comeal and conjunctival epithelium had the potential for developing keratin fibrils comparable to those found in the epithelia of the skin in suitable tissue culture. There are a number of clinical instances where tarsal and bulbar conjunctiva may produce keratinized cells. The most frequent of these is exposure or drying, as in ectropion of the lid, staphyloma of the globe, or lagophthalmos. In some instances this is merely a drying of the superficial epithelial cells and not a true metaplasia of the epithelial layer. More interesting are those conditions in which there are adequate tears, but the tissue undergoes a true metaplasia to form a stratum granulosum and corneum. The most common example of this is vitamin A deficiency
Keratinization of Conjuctiva
137
.. ...
.... ...
FIGURE 2
Histopathology of surface conjunctival epithelium removed from eye in Fig 1 at the time of a centrally placed penetrating keratoplasty.
FIGURE 3
Skin graft to conjunctiva performed by plastic surgeon to prevent recurrence of pterygium.
138
Maumenee
and the development of xerophthalmia. Although such cases are seldom seen in the United States today, it is still one of the principal causes of blindness in developing countries. Another frequent cause of true metaplasia is scar formation after trachoma, Stevens-Johnson disease, irradiation ofthe lids, and infections in patients with atopic dermatitis. In some otherwise normal individuals, chronic irritation from topical medications may produce keratinizing metaplasia ofthe tarsal conjunctiva. This may occur either from an allergy to the drug from such medications as sulfonamides or penicillin or from drug irritation without obvious evidence of allergy. The worst offender of this latter type of reaction was furtrethonium iodide (Furmethide), which was used during the 1940s and 1950s to lower intraocular pressure (Fig 4). A high percentage of patients who used this drug over a period of a year or more developed stenosis of the lacrimal canaliculi and keratinization of the tarsal conjunctiva. Other offenders of this latter type have been pilocarpine and epinephrine. The conjunctiva overlying tumors of both the lid and the globe may
FIGURE
4
Keratinization and stenosis of lacrimal puncta following the use of furtrethonium iodide (Furmethide) for the treatment of glaucoma.
Keratinization of Conjuctiva
139
FIGURE 5
Keratinization of tarsal conjunctiva overlying sebaceous cell carcinoma of lid.
develop true keratinizing epithelium. Some examples of these are intraepithelial epithelioma (Bowen's-like changes in the conjunctiva), squamous cell carcinoma, epithelial hyperplasia of the limbus, and sebaceous cell carcinomas of the lid (Fig 5). It is interesting that the bulbar conjunctiva appears to be affected more in vitamin A deficiency. In contrast, drug irritation, trachoma, and Stevens-Johnson disease are more likely to involve primarily the tarsal conjunctiva. A very interesting type of keratinization occurs in superior limbic keratoconjunctivitis. Light and electron-microscopic examinations of biopsies ofthis lesion have shown keratinization ofthe superficial epithelial cells and keratohyaline granules in the deeper layers of the cells."1 CLINICAL SIGNIFICANCE
I have been particularly interested in isolated areas of keratinization of the tarsal conjunctiva.12 This is a relatively rare condition that occurs after
140
Maumenee
Stevens-Johnson syndrome, drug reactions, and irradiation of the lids. When the lesion comes into contact with the corneal epithelium, it produces a superficial punctate keratitis and if allowed to persist over a long period of time, superficial vascularization and scarring of the cornea may occur. I have previously reported that the lesions recurred if the keratinized tissue was superficially removed. However, deep excision of the lesion and repair of the area with a mucous membrane graft from the mouth results in a permanent cure with relief of irritating symptoms. I have followed some patients treated in this manner for a period of 20 years without evidence of recurrence of the lesion. More recently, I have observed that the keratinized plaque may disappear spontaneously if the topical medication which has caused the lesion is discontinued. Approximately three years ago I was attempting to destroy the lashes with cryotherapy, according to the technique described by Beard,13 in a patient who had trichiasis and keratinization of the tarsal conjunctiva as a result ofan allergy to drug therapy. The cryoprobe was applied to the area of keratinization and when the tissue healed the superficial tissue was no longer keratinized. Since that time, five other eyes have been treated with a light application of the cryoprobe to areas of long-standing keratinization of the lid, with either a resultant total obliteration of the keratinized tissue or marked improvement of the lesion. Four of these patients had had Stevens-Johnson syndrome, and one patient had atopic dermatitis and secondary corneal ulcers. The gratifying response of the keratinized tissue of the lids to cryotherapy makes it interesting to speculate whether a similar result would occur if superficial limbic keratoconjunctivitis was treated by freezing after the conjunctiva had been lifted from the globe by the injection of fluid. SUMMARY AND CONCLUSIONS
There are many examples that one may cite where the eye or its adnexa may be used as a "test tube" to study biological phenomena. The interesting morphological and biochemical metaplasia of the conjunctiva which occurs when it migrates across the cornea is an excellent example of this. A number of years ago Billingham and Medawar"4 reported that the corneal epithelium did not alter its histologic structure when transplanted to the abdomen of an experimental animal. The observation of Sun9 and his coworkers that conjunctival and corneal epithelium when grown in tissue culture will produce keratin fibrils comparable to those found in
Keratinization of Conjuctiva
141
skin epithelium, and the observation that under certain pathologic conditions both the bulbar and tarsal conjunctiva may undergo true keratinization, strongly suggests that these cells have the potential for multiple differentiation, rather than being terminal elements in development. The shedding of keratinized cells onto the globe, or the rubbing of such cells onto the cornea when the keratinization is localized on the lids, can produce chronic irritation. Excision of the lesions and replacement with mucous membrane, or destruction of the keratinized tissue by freezing, will relieve these symptoms. SUMMARY
Keratinizaton of the tarsal conjunctiva in an eye with adequate tears occurs following a number of conditions, including irritation and sensitivity to topical medications, Stevens-Johnson syndrome, radiation to the lid, and occasionally from unknown causes. The keratinized cells produce an epithelial keratitis with subsequent vascularization of the cornea. The morphogenesis of this condition is discussed, along with therapy including mucous membrane grafts and freezing of the tissue. REFERENCES
1. Friedenwald JS: Growth pressure and metaplasia of conjunctival and corneal epithelium. Doc Ophthalmol V-VI:184-192, 1951. 2. Thoft R, Friend J: Biochemical transformation of regenerating ocular surface epithelium. Invest Ophthalmol 16:14-20, 1977. 3. Maumenee AE, Scholz RO: The histopathology of ocular lesions produced by sulphur and nitrogen mustards. Bull Johns Hopkins Hosp 82:121-147, 1948. 4. Sengel P: Morphogenesis of the Skin. Cambridge Univ Press, Cambridge, England, 1976. 5. Fell HB, Mellanby E: Metaplasia produced in cultures of chick ectoderm by high vitamin A. J Physiol 119:470-488, 1953. 6. Lasnitzki I: Effect of carcinogens, hormones, and vitamins on organ cultures. Inter Rev Cytol 70:79-121, 1958. 7. Wolff HH, Christophers E, Braun-Falco 0: Beeinflussung der epidermalen Ausdifferenzierung durch Vitamin A Saure. Eine electronenmikroskopische Untersuchung. Arch Klin Exp Derm 237:774-795, 1970. 8. Billingham RE, Silvers WK: Studies on the conservation of epidermal specificities of the skin and certain mucosas in adult mammals. J Exp Med 125:429-446, 1967. 9. Sun TT, Green H: Immunofluorescent staining of keratin fibers in cultured cells. Cell 14:469-476, 1978. 10. McLoughlin CB: The importance of mesenchymal factors in the differentiation of chick epidermis. II. Modification of epidermal differentiation by contact with different types of mesenchyme. J Embryol Exp Morphol 9:385409, 1961. 11. Donshik PC, Collin HB, Foster CS, et al: Conjunctival resection treatment and ultrastructural histopathology of superior limbic kerato-conjunctivitis. Am J Ophthalmol 85:101-110, 1978.
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12. Maumenee AE: Keratitis secondary to keratinization of the tarsal conjunctiva. Am J Ophthalmol 41:477-487, 1956. 13. Beard C: Ptosis, ed 2. St Louis, CV Mosby, 1976. 14. Billingham RE, Medawar PB: A note on the specificity of corneal epithelium. J Anat 84:50-56, 1950.
DISCUSSION
DR ARNALL PATZ. In reference to the blood vessels emanating from the limbus, there is new work in terms of growth factors in serum. I wonder if in Doctor Maumenee's review of this work, if there have been any specific studies on the role of serum or epidermal transformation that might occur in metaplasia of the epithelium. DR HAROLD GIFFORD. I would like to ask a question about pemphigoid. Where does pemphigoid fit into this picture? I had a patient with ocular pemphigoid. The lids were treated with cryotherapy to get rid of the lashes which were irritating the cornea. The freezing removed the lashes but made the conjunctival scarring worse. One eye lost the fornix completely and the other one was mostly epidermalized with early scarring of the cornea. I just wonder what is the connection with pemphigoid and if there is anything you know that can be done for this terrible disease? DR ALBERT RUEDEMANN, JR. I think this is a very interesting paper for a number of reasons, and I think this business of squamous metaplasia going over the lid margins is very remarkable but it is reversible. We reported at this meeting some years ago a ten-year study using scleral lenses with adherent lids, squamous metaplasia and Stevens-Johnson's disease and pemphigus. We treated these patients with scleral lenses by merely cutting the lids free from the globe and inserting the lens on the table. We allowed the tissue to regrow and actually got return of normal conjunctival and tear flow. DR MYRON YANOFF. Doctor Maumenee and Doctor Patz' comments bring up the question of what really is keratinization, specifically Doctor Patz' question about serum. Some years ago we did organ cultures of corneal buttons and showed that if serum was not used in the culture medium, most of the time, after about four or five days, the epithelium became keratinized. (Yanoff M: In vitro biology of corneal epitheliun and endothelium. Trans Am Ophthalmol Soc 73:571-620, 1975.) If serum was used, the epithelium did not become keratinized, even after a month in organ culture.
DR RALPH HAMILTON. I think this is a very useful thing, and I would like to ask Doctor Maumenee exactly what light freezing is. Did you use a thermocouple in this and how much freezing do you require for this? DR A EDWARD MAUMENEE. I am very sorry that Doctor Crowell Beard was not able to attend the meeting because of illness, and I wish him a rapid recovery. I would like to thank those discussants who spoke extemporaneously. In answer to the first
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question, I did not find anything in the literature specifically relating serum to keratinization. It was mentioned in the experiments on tissue culture ofembryonic skin that the nutrients in the tissue culture were extremely important in carrying out these experiments. As far as ocular pemphigus or pemphigoid is concerned, it has been my impression that keratinization of the tarsal conjunctiva was extremely rare in this condition during the early stage of the disease. It is only in the end stage when there is marked drying of the conjunctiva and exposure of the tissue that keratinization occurs. It has also been my experience that anything that is done to the tissue to traumatize it, such as freezing, incising it, or injuring it in any way, tends to cause the disease to progress and become worse. In answer to Doctor Ruedemann's comments, I did mention in the written paper that I had several patients in whom there was a spontaneous regression of the keratinization of the tarsal conjunctiva. I did not use a contact lens on these patients, nor had they used a contact lens prior to the development of their keratinization. Certainly chronic irritation from Furmethide or pilocarpine or all9rgy to various sulfonamides will produce this metaplasia of the tissue. All of the patients I treated with the freezing technique had had long-standing alterations of the tissue that had not cleared spontaneously. Doctor Hamilton's question was a good one. I did not use a thermocouple. I merely applied the cryoprobe to produce a superficial freezing of the tissue and as soon as it turned white, the tissue was allowed to thaw. The cryoprobes that I used were those that we routinely use at Wilmer for cataract extraction or retinal detachment surgery. These were the mechanical cryoprobes and not the disposable ones. As far as the tissue culture experiments that Doctor -Yanoff mentioned, I had not been aware that the addition of serum alone would cause a metaplasia of the cultured material. I wonder if in his tissue cultures there was an absence or deficiency in vitamin A content of the tissue culture material before the serum was added.
