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Lens deviations in rabbit offspring after short-term prenatal exposure to dexamethasone: A light-microscopic study
195
lhp. Eye Res. (1984) 39, 195206
Lens Deviations in Rabbit Offspring after Short-term Prenatal Exposure to Dexamethasone : A Light-Microscopic Study JENNEKE
M. A. KOKSMA, PHILIP AND BEN W. WILLEKENS
F. J. HOYXG
The Netherlands Ophthalmic Research Institute, P.O. Box 12141, 1100 AC Amsterdam, The Netherlands (Received 12 October 1983 and accepted 21 March 1984. London) Female New Zealand White rabbits were treated subcutaneously with 90375 or 915 mg kg-’ dexamethasone or physiological saline solution from the 8th through to the 1 lth day of gestation. Short-term administration of 915 mg kg-’ dexamethasone induced two types of morphological lens deviations in the offspring (lenses were studied by means of sagittal serial sections). In the first type of deviation a progressive series of changes in the lens bow region was observed. Early changes consisted of an enlargement of some lens bow nuclei, an increase in the number of lens bow nuclei and a disturbance in the shape of the lens bow. Later changes consisted of the degeneration and vacuolization of a peripheral zone of cortex fibre cells. The overall picture showed that the size of up to 8 y0 of the nuclei in the deviating lens bows was enlarged and that the average number of lens bow nuclei in the deviating lenses w&s 28 o/0higher (P < 001) than in the lens bows without deviations. The second type of lens deviation consisted of isolated bundles of disintegrated cortex fibre cells, which extend downwards (anterior-posterior) from subepithelial vesicles. In these bundles all nuclei were smaller and more basophilic than normally. Both types of lens deviations occurred independently in the superficial cortex. Control offspring treated with physiological saline solution showed neither the first nor the second type of lens deviation. Of the offspring exposed to 00375 mg kg-’ dexamethasone one animal (8 %) showed lens changes of the first type only, whereas 59 y/o of the offspring exposed to 915 mg kg-’ dexamethasone displayed either one or both types of lens deviations, Key words: light microscopy; teratogenic lens deviations: prenatal dexamethasone; lens bow region ; cortex fibre cell bundles.
1. Introduction There is ample evidence that systemic as well as topical treatment with glucocorticoids can induce lens opacities in man (Black, Oglesby, Von Sallmann and Bunim, 1960; Spencer and Andelman, 1965; Younessian, 1970; Polak, 1980; Adhikary, Sells and Basu, 1982). However in animal experiments this effect is not clear. Some investigators were not successful in raising lens opacities in rats, rabbits and chickens after administration of glucocorticoids (Von Sallmann, Caravaggio, Collins and Weaver, 1960; Cotlier and Becker, 1965; Bettman, Fung, Webster, Noyes and Vincent, 1968), while others reported lens opacities (Tarkkanen, Esilii and Liesmaa, 1966; Wood, Contaxis, Sweet and Van Dolah, 1967). The cataractogenic effect of some drugs, insufficient diets or certain diseases is potentiated by additional treatment with glucocorticoids (Bettman, Fung and Noyes, 1964; Bettman et al., 1968; Cotlier et al., 1965; Koch, 1976). There is only scarce evidence that prenatal treatment with glucocorticoids may result in cataract development in man (Wells, 1953 ; Kraus, 1975). In mice however, the development of cataracts after prenatal administration of glucocorticoids has been Please address reprint requests to: Jenneke M. A. Koksma at the above address 00144835/84/080195+12
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01984 Academic Press Inc. (London) Limited
196
.I. >I. A. KtbKSM.4
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demonstrated (Rogoyski and Trzcinska-Dahrowska, 1969). Experiment,s with rats support this result (Haumont, 1958). Recently the application of hydrocortisone was shown to induce reversible cataracts in chicken embryos (Nishigori, Lee and Iwatsuru. 1983). The aim of the present study is t’o evaluate the teratogenic effect of dexamethasone on the rabbit lens after short-term prenatal exposure.
2. Materials
and Methods
Female New Zealand White rabbits, weighing 3-4 kg, after mating with male rabbits of the same strain and weight, (one male per two randomly-chosen females), were treated with various doses of dexamethasone (Oradexon: Organon, 0~s). From the 8th to the 1lth day of supposed gestation (the first day of gestation is regarded as day 0) all females received either @0375 mg kg-l dexamethasone, @15 mg kg-’ dexamethasone, or physiological saline solution subcutaneously once daily between 0900 hr and 1000 hr. (The females were randomly distributed in the three dose groups; the doses of dexamethasone corresponded to therapeutic doses given to man.) The offspring were killed at the age of 12 weeks. Both eyes were enucleated and immediately fixed in a 10% formalin solution. One eye per rabbit was selected at random. After dehydration and embedding in celloidin, 10pm saggital serial sections were cut with a Reichert sledge microtome. Every 10th section was stained with haematoxylin and eosin. Two independent light-microscopic examinations of all the specimens were carried out. Morphometry In the central lens sections of both the control offspring and the low and the higher dose dexamethasone-exposed offspring the lens bow nuclei were counted by means of photographs (magnification 63 x ). As each central lens section contained two lens bows, both numbers of nuclei were averaged. Statistical methods The following statistical tests were performed: the X-square test and the exact test of Fisher (to compare percentages), the two sample test of Wilcoxon (to compare two groups) and the test of Kruskal-Wallis (to compare more than two groups). Results were considered significant at the 5 y0 level (two-sided tail probability P < @05).
3. Results In Table I the mean number of neonati and of 1%week-old offspring per group of females is reported. A comparison of both numbers shows that in this study the higher death rate of the offspring before birth on account of the dexamethasone treatment is compensated by a higher degree of survival after birth. The young rabbits never showed gross abnormalities such as palatoschizis, hare lip, club foot or polydactyly. Morphology In the superficial cortex of the lenses of the offspring exposed to the high dose of dexamethasone two types of lens deviations can be demonstrated. (a) Deviation in the lens bow region. The high dose-exposed offspring showed changes in the lens bow region, which can be seen most distinctly in the central lens sections. These changes consisted of a series of progressive alterations, Firstly, in a total of 13 offspring, round enlarged nuclei (up to 8 o/Oof the total number) were observed in the lens bow. The enlarged nuclei are as basophilic as the normal ones. This stage is considered pre-deviant. Secondly, the configuration of the lens bow was disturbed
TERATOGEKIC’CHASGES
IN THE
RABBIT
LEKS
197
TABLE I
Number of newborn and 12-week-old off.yring dexeamethasone
Drxamethasone dose (mg kg-‘) Control 00375 0.15
born to rabbits treated with
Females (n)
Neon&i per female (Mean n)
12-week-old offspring per female (Mean n)
8 7 15
1.2 64 57
2.0 (n = 16) 1.9 (n = 13) 1.9 (n = 29)
F‘IQ. 1. Normal lens bow; control group. The lens bow is formed by the nuclei of the superficial corl ;ex fibr e cellr 1. Central lens section, x 86. Fk3. 2. Disturbed lens bow region, stage 1; @15 mg dexamethasone. the lens 1low are displaced posterially. Central lens section. x X0.
The nuclei in the inner zone of
3b
Fro. 3. Disturbed lens bow region. stage 2; 0.15 mg dexamethasone. Presence of a disturbed lens bow and a lateral zone of degenerated cortex fibre cells. (a) Central lens section, x 54. (b) The lateral zbnes of degenerated cortex fibre cells extend throughout the whole lens. Peripheral lens section, x 25.
in 11 out of the 13 above-mentioned offspring. While in Fig. 1 a normal lens bow is shown, Fig. 2 shows a disturbed one wherein the nuclei in the inner zone of the bow are displaced posterially. Thirdly, of the 11 offspring with a disturbed lens bow configuration, seven showed distinct, and two beginning, stages of cortex fibre cell degeneration. In the cases of distinct fibre cell degeneration, bilateral zones of affected fibre ceils were present [Fig. 3(a)]. These zones extended through the lens towards the periphery [Fig. 3(b)]. Finally, four out of the seven lenses with distinct zones of degenerated cortex fibre cells showed subcapsular vacuolization in the anterior region of the lens [Fig. 4(a)]. The vacuoles, which enlarge and possibly confluence, were formed in the anterior parts of the affected lens fibre cells [Fig. 4(b)]. Figure 4(c) shows a detail of the degenerated fibre cells with enlarged nuclei.
TERATOGENIC
CHANGES
IN THE
RABBIT
LEXS
199
FIG. 4. Final stage of changes in the lens bow region; 0.15 mg dexamethasone. In addition vacuoles are formed in the anterior parts of the degenerated cortex fibre cells. (a) Disturbed lens bow, zone of lateral degenerated cortex fibre cells and subcapsular vacuolization. Central lens section, x 62. (b) Detail of the enlarging and maybe confluencing vacuoles. Interference contrast, x 225. (c) Detail of some enlarged (1) and normal (2) nuclei; many nuclei are disorientated (3), x 360. The enlarged nuclei are present throughout the whole series ofprogressive changes, also in small numbers in the prt-deviant stage. The small, dark spots on the right of the lens bow are artifacbs.
In the offspring of the low dose dexamethasone-treated females this type of lens deviation was observed once. The deviation consisted of changes in the lens bow region (round, enlarged nuclei ; disturbed shape of the lens bow : bilateral zones of degenerated cortex fibre cells and subcapsular vacuolization). (b) Degeneration of isolated bundles ofcortex$bre cells. In the deviating lenses bundles of disintegrated cortex fibre cells, which extend downwards from subepithelial vesicles, were present [Fig. 5(a)]. The vesicles were filled with a fluid substance. The affected fibre cells stained scantily with eosin. They had small, round, dark-staining nuclei compared with the surrounding normal fibre cells [Fig. 5(b)]. The cytoplasm also contained many lightly eosin-staining globules. which demonstrates the disinte-
200
.J. 11. A. KOKSMA
ET AL.
FIQ. 5. Degeneration of isolated bundles of cortex fibre cells; 0.15 mg dexamethasone. (a) Bundles of degenerated cortex fibre cells, which extend downwards from subepithelial vesicles and stain scantily with eosin. Peripheral lens section, x53. (1)) Uptail of the helt with nuclei: the small. round, very basophilic nuclei in the degenerated bundles surrounded by the oblong, less basophilic. normal nuclei, x 252. (c) Detail which shows globules (arrow) in the cytoplasm of the degenerated cortex fibre cells. Interference contrst, x 360.
TERATOGENIC
CHANGES
IN THE
RABBIT
201
LENS
Fro. 6. Localization of the two types of lens deviations (scheme; frontal view of the lens). Shaded area: zone with changes in the lens bow region. Black dots: bundles with degenerated cortex fibre cells.
gration of the lens fibres [Fig. 5(c)]. Of interest is that this deviation occurred on several places in the equatorial part of the lens, while the alterations in the lens bow region were present as a continuous zone in a more extended part of the periphery (Fig. 6). Table II reports the number of young rabbits with each type of lens deviation in the three dose groups. In the table the percentages in the dose groups show significant differences [Table II(a) P < O-01; Table II(b) P < O*OOl]. The two types of lens deviations occurred separately as well as together. Table III shows the distribution of both types of changes among the offspring exposed to the higher dose of dexamethasone. A relation between both types of deviations is not found. In total 17 of the 29 young rabbits (59 %) sh owed either one or both types of lens deviations. Morphometry Counting of lens bow nuclei. In Table IV(a) the results are registered of the counts of the nuclei in the central lens bows of the 16 control lenses and of the 13 and 29 lenses originating from the offspring exposed to the low and the higher dose of TABLE
The distribution
II
among the rabbit offspring
of the two types of lens deviations Dexamethesone dose (mg WI
Offspring (nl
Offspring with lens deviation (n)
(a) Changes in the lens bow region* Control 16 00375 13 0.15 29 (b) Degeneration of isolated cortex fibre cell bundles? Control 16 00375 13 0.15 29 * The distribution of the deviation P < 0.01). t Idem (x-square test, P < 0901).
among the dose-groups
0 (0%) 1(8%) 11 (38%) 0 (0%) 0 W%l 13 (45%) differs
significantly
(X-squaretest,,
202
.I. M. A. KOKSMA
Distribution
ET AI,
of the two types of lens deviations twnong &a-e of dexamethasone*
the offspring
exposed
to the high,
Changes in the lens how region Present Degenerated cortex fibre cell bundles Total number * No significant
Absent
Total number
Present
7
6
13
Absent
4 11
1% 18
16 29
relation between both types of deviations
is found (Fisher’s test)
dexamethasone. According to the test of Kruskal-W&s, the levels of the numbers of lens bow nuclei in the three dose-groups differ significantly (P < O-05). Since in the low dose-group only one abnormal lens was observed, the lenses exposed to the higher dose of the drug were divided into four groups depending on the presence or absence of one or both types of lens deviations [see Table IV(b)]. The numbers of lens bow nuclei derived from the two groups of lenses which showed no changes in the lens bow region (groups 1 and 2) did not differ significantly from each other. The same held for the numbers of lens bow nuclei derived from the two groups of lenses which showed changes in the lens bow region (groups 3 and 4). Therefore groups 1 and 2 on the one side, and 3 and 4 on the other side were pooled and both resulting groups compared with the control group. A significant difference in the number of lens bow nuclei was TABLE
Median
IV
number of the lens bow nuclei Offspring (n)
(a) Dose DEX* (mg kg-‘) control @0375 mg kg-’ DEX 0.15 mg kg-’ DEX
in central lens sections Median number (extreme values)
16 13 29
38% (312,540)t 436 (360,500)t 459 (278. 602)t
offspring I2
447 (330. 507)
(2) Degeneration of isolated cortex fibre cell bundles only
6
4244 (278. 482)
(3) Disturbed lens bow region only
4
541 (476. 561)
(4) Both types of deviations
7
490 (398. 602)
(b) Group of @15 mg kg-’ DEX-exposed (1) R’o deviations
43% (278, 507)f
500 (398,602)s * t $ §
DEX = Dexamethasone. Significantly different (test of Kruskal-Wallis. P c @05). Not significantly different from the control value (test of Wilcoxon). Significantly different from the control value (test of Wilcoxon, P < @Ol).
TERATOGENIC
CHANGES
IN THE
RABBIT
LEES
203
found for the group of lenses with lens bow deviations (P < O*Ol), but not for the group of lenses without lens bow deviations. In conclusion, the lenses with deviations in the lens bow region contained significantly more nuclei than the control lenses. 4. Discussion In this study the offspring of female rabbits treated with dexamethasone for a short period during early pregnancy showed two types of lens deviations. As in the study of Rogoyski et al. (1969) the lenses were studied morphologically. In the first type of lens deviation an enlargement of a part of the lens bow nuclei, an increase in the number of lens bow nuclei, and a disturbance in the shape of the lens bow, followed by degeneration and vacuolization of a zone of cortex fibre cells were observed (Figs. 2, 3 and 4). These deviations are comparable to the changes found in various forms of hereditary cataract in mice (Hamai, Fukui and Kuwabara, 1974; Burns, Anderson and Feeney-Burns, 1980; Uga, Kador and Kuwabara, 1980; Kuck, Kuwabara and Kuck 1981-82). However, the hereditary cataracts mentioned have different features, i.e. in some studies the number of nuclei in the lens bow has increased (Hamai et al., 1974; Uga et al., 1980), while in others it has decreased (Burns et al., 1980; Kuck et al., 1981-82). Furthermore, the localization of the swollen lens fibres and the vacuoles is mostly in the posterior cortex, in contrast to the anterior-lateral position reported in this study. In the lenses with changes in the lens bow region we found, in comparison with the control lenses, an average increment of 28 o/0 in the number of lens bow nuclei. This increment may be caused by either a delay in denucleation of the superficial cortex cells (Hamai et al., 1974; McAvoy and Van Heyningen, 1976: Uga et al., 1980), or by an increase in the formation of cortex fibre cells by the action of dexamethasone. Since in vitro experiments (Van Venrooy, Groeneveld and Bloemendal, 1974) have demonstrated an increased transition of lens epithelial cells into cortex fibre cells after the addition of dexamethasone, we favour the latter hypothesis. The second type of lens deviation outlined in this study includes the degeneration of isolated cortex fibre cell bundles (Fig. 5). To our knowledge this deviation has not been reported before. Two doses of dexamethasone were applied, namely 90375 mg kg-’ and O-15 mg kg-l, which correspond to about 19 mg and 74 mg prednisone-equivalents in man. Almost all lens changes were found in the offspring exposed to the. higher dose of dexamethasone (see Table II). To establish if the two types of lens deviations have the same origin or not, various characteristics of both types of changes were compared in the lenses exposed to the higher dose of dexamethasone. In the lenses with degenerated cortex cell bundles and without lens bow changes (see Fig. 7) we found the same number of lens bow nuclei as in the control lenses; lenses with deviations in the lens bow region contained on average 28% more nuclei than the control lenses [Table IV(b)]. Furthermore, the nuclei of the degenerated cortex cell bundles were small, round and highly basophilic [Fig. 5(b)], whereas in the lenses with a disturbed lens bow region up to 8 o/0 of the nuclei were enlarged and as basophilic as the control nuclei [Fig. 4(c)]. Finally, the two types of lens deviations, although occurring together as well as separately, did not seem to be related statistically (Table III). Hence, we conclude that the two types of lens deviations reported here have developed independently. A comparison of the peripheral sections [Figs. 3(b) and 5(a)] and of the central
7
FIQ. 7. Lens with degenerated cortex fihre cell bundle and normal lrns how: 0.1.’ mg drxamethasone. Central lens section. X 90.
sections [Figs. 2, 3(a), 4(a) and 71 of lenses with either the first or t,he second type of lens deviation also demonstrates the different character of both types of deviations. Moreover, the fact that in the first type of deviation the lens bow is involved and that the cortex cells whose nuclei compose the lens bow derive from lens epithelium cells (Hanna and O’Brien, 1961; Fine and Yanoff, 1979; Beebe, Compart, Johnson, Feagans and Feinberg, 1982) indicate that the first type of lens deviation may develop after a complicated series of events resulting in the degeneration and vacuolization of a peripheral zone of cortex fibre cells (Kinoshita, 1974). The results of the present study suggest that this type of deviation develops in another way and in another period than the second type of lens deviation. In the second type of lens deviation the bundles of degenerated cortex fibre cells with small, round nuclei may be induced after the formation of the subepithelial vesicles, which cut off groups of underlying cortex cells from the feeding epithelium. The experimental set-up of the present investigation is very similar to that of Haumont (1958) and Rogoyski et al. (1969). Haumont states that some offspring of female rats treated with cortisone during pregnancy show cataract formation. Rogoyski et al. (1969) examined the fetuses of female mice treated daily with a high dose of hydrocortisone during, e.g., the 9th and 10th day of gestation; based on the mean weights of mice and men, the dose of the drug corresponds to about 875 mg prednisone-equivalents in man. After this treatment the fetuses show a high percentage (31%) of morphological cataract. It is not clear whether the degenerative changes, which are most frequently found in the secondary nuclear fibres, consist of swelling or of vacuolization. Changes in the composition or shape of the lens bow are not reported. Nishigori et al. (1983) describe the induction of a reversible cataract in chicken fetuses after injection of a moderately high dose of hydrocortisone corresponding to about 30 mg prednisone-equivalents in man. Since the examination occurred with the slit-lamp, it is not known if concomitant changes in the morphological structure might be demonstrable. However, the cataractous changes found by Rogoyski et al. (1969) and Nishigori’et al. (1983) occur in the fetal stages of the laboratory animals, whereas
TERATOGENIC
CHANGES
IX THE
RABBIT
LENS
20.5
in the present investigation changes were observed in the lenses of animals 12 weeks after birth. The present lens changes were found in the superficial cortex only; the embryonal, fetal and early postnatal nucleus had a normal appearance. Various possibilities arise as regards the developmental line of the present lens deviations. The first is that, dependent on the species of the experimental animal and/or the type of glucocorticoid applied, the morphological expression of the lenticular changes is observable in the fetal stage or after a time delay in the postnatal stage. Secondly, exposure to dexamethasone may produce morphological changes in the rabbit lens, starting in the prenatal stage and continuing for some time (see later) after birth, but because of, e.g., natural growth (see Nishigori et al., 1983) changes in the embryonal, fetal and early postnatal nucleus may disappear. Finally, the fetuses, which are reabsorbed by the mother because of the treatment with dexamethasone (21%, see Table I), might have shown severe embryonal and fetal lens damage. The present types of lens deviations are localized in a peripheral zone in the cortex (see Fig. 6). Compared with the size of lenses of 28-day-old-i.e. nearly full-grownfetuses, the morphological expression of the changes observed at the 12-week stage, must have started at some time after birth. Figs 2,3(a) and 4(a) show that the changes in the lens bow region cease before the 12th week after birth, since outside the disturbed lens bow region normal cortical fibres are present. Presumably the lens deviation consisting of the bundles of degenerated cortex fibre cells has a zonular character as well. As mentioned before, the two types of lens deviations probably developed differently. Glucocorticoids influence lens metabolism in several ways. In addition to the already mentioned effect on the lens epithelium (Van Venrooy et al., 1974) which may be involved in the development of the deviation in the lens bow region, they act on the lenticular content of glucose and various cations (Harris, 1966; Apponi Battini, 1968; Benatti and Balli, 1971). Since in this study only histological examinations were performed, it is not relevant to discuss the possible biochemical pathways leading to each type of lens deviation. In conclusion, two independent types of lens deviations were demonstrated in the eyes of rabbit offspring after short-term prenatal exposure to dexamethasone. ACKNOWLEDGMENTS The authors wish to thank Prof. Dr G. M. Bleeker, Prof. Dr J. van Limborgh and Dr H. A. Brouwer for their encouragement and valuable advice during this study; Dr P. D. Bezemer C.E. for his expert statistical assistance; Mr J. Bult for correcting the manuscript and Mrs T. Kierks for secretarial help. REFERENCES Adhikary, H. P., Sells, R. A. and Basu, P. K. (1982). Ocular complications of systemic steroid after renal transplantation and their association with HLA. Br. J. OphthalmoE. 66,29&l. Apponi Battini, G. (1968). Ueber den Glukose- und Glykoproteingehalt im Kammerwasser, im Glaskiirper und in der Linse der Kaninchen naeh Behandlung mit Glukokortikoiden. Min. Mbl. Augenheilk. 152, i’OS-14. Beebe, D. C., Compart, P. J., Johnson, M. C., Feagans, D. E. and Feinberg, R. N. (1982). The mechanism of cell elongation during lens fiber cell differentiation. Dev. Biol. 92, 54-9. Benatti, C. and Balli, F. (1971). Cataract caused by long-term corticosteroid therapy and its prevention. Minerva Pediatr. 23, 1599-l. Bettman, J. W., Fung, W. E. and Noyes, P. P. (1964). Potentiating action of prednisone on galactose cataracts in rats. Invest. Opthalmol. 3, 678-9.
‘06
.l 11. A. KOKSMA
F:T ;il,.
Bettman. J. W., Fung, W. E., W’rbster. 1~. G., Noyes. P. I’. and Vincent, N. J. (1968). The cataractogenic effect, of corticosteroids on animals rim. J. Ophthalmol. 65, 581.-6. Black, R. L.. Oglesby. R. B.. Von Sallmann, I,. and Bunim. .J. .J. (1960). .J. Am. Jfed. A4~.s~~c. 174, 16671. Burns, R. I’., Anderson, R. S. and Feeney-Burns, L. (1980). C’ataract-webbed trait in Peromyscus. TT Riomicroscopy and histology of eyes. Invest. Ophthdmol. Vis. ,Qi. 19, 3142. Cotlier, E. and Becker, B. (1965). Topical corticosteroids and galactose-cataracts. I~~l;e,st. OphthaZmoZ. 4. 80614. Fine, B. S. and Yanoff. M. (1979). Ocular Histology; A Text and Atlas (2nd edn). Pp. 149-53. Harper and Row Pub]. Inc., Hagerstown, MD. Hamai, Y ., Fukui, H. N. and Kuwabara, T. (1974). Morphology of hereditary mouse cataract. Exp. Eye Res. 18, 53746. Hanna, C. and O’Brien, J. E. (1961). Cell production and migration in the epithelial layer of the lens. Arch. Ophthalmol. 66, 103-7. Harris, .J. E. (1966). The temperature-reversible cation shift of the lens. Trans Am. Ophthalmol. Sot. 64, 67.599. Haumont, St. (1958). Effets de la cortisone administree pendant la gestation. Ann. d’EndocGnol. 19, 442-5. Kinoshita, ,J. H. (1974). Mechanisms initiating cataract formation. Invest. Ophthdmol. 13, 713-24. Koch, H. R. (1976). Klinische und experimentelle IJntersuchungen iiber den Einfluss von Corticosteroiden auf die Augenlinse. Pp. 88-93. Ph.D. Thesis, University of Bonn. Kraus, A. M. (1975). Developmental cataracts. N. Y. State J. Med. 75, 1757-8. Kuck, J. F. R., Kuwabara. T. and Kuck, K. D. (1981-82). The Emory mouse cataract--an animal model for human senile cataract. Curr. Eye Res. 1, 643-51. McAvoy, J. W. and Van Heyningen. R. (1976). Changes in the cells of the lens bow and epithelium of tryptophan-deficient rats. INSERM 60, 245-50. Nishigori, H., Lee, J. W. and Iwatsuru. M. (1983). An animal model for cataract research: cataract formation in developing chick embryo by glucocorticoid. Exp. Eye Res. 36, 617-22. Polak, B. C. P. (1980). Ophthalmological Complications of Haemodialysis and Kidney Transplantation. Dr W. Junk (Pub].), The Hague. Rogoyski. A. and Trzcinska-Dabrowska, Z. (1969). Corticosteroid-induced cataract and palatoschisis in the mouse fetus. Am. J. OphdhaZmoZ. 68, 128-33. Spencer, R. W. and Andelman, S. Y. (1965). Steroid cataracts. Posterior subcapsular cataract formation in rheumatoid arthritis patients on long term steroid therapy. Arch Opt?&. 74, 3841. Tarkkanen, A., Esila, R. and Liesmaa, M. (1966). Experimental cataracts following long-term administration of corticosteroids. Acta Ophthalmol. 44, 665-8. IJga, S., Kador, P. F. and Kuwabara, T. (1980). Cytological study of Philly mouse cataract, Exp. Eye Res. 30, 74-92. Van Venrooy, W. J., Groeneveld, A., Bloemendal, H. and Benedetti, E. L. (1974). Cultured calf lens epithelium. II The effect of dexamethasone. Exp. Eye Res. 18, 527-36. Von Sallmann, L., Caravaggio L. L., Collins, E. M. and Weaver, K. (1960). Examination of lenses of steroid-treated rats. Am. J. OphthaZmoE. 50 1147-51. Wells, C. N. (1953). Treatment of hyperemesis gravidarum with cortisone. Am. J. Obst. Gynec. 66, 599601. Wood, D. C., Contaxis, I., Sweet, D., Smith, J. C. and Van Dolah, J. (1967). Response of rabbits to corticosteroids. I Influence on growth, intraocular pressure and lens transparency. Am. J. Ophthalmol. 63, 841-8. Younessian, S. (1970). Glaucome et cataracte apres corticotherapie par voie g&&ale et locale. Adv. Ophthalmol. 23, 74-152.
lhp. Eye Res. (1984) 39, 195206
Lens Deviations in Rabbit Offspring after Short-term Prenatal Exposure to Dexamethasone : A Light-Microscopic Study JENNEKE
M. A. KOKSMA, PHILIP AND BEN W. WILLEKENS
F. J. HOYXG
The Netherlands Ophthalmic Research Institute, P.O. Box 12141, 1100 AC Amsterdam, The Netherlands (Received 12 October 1983 and accepted 21 March 1984. London) Female New Zealand White rabbits were treated subcutaneously with 90375 or 915 mg kg-’ dexamethasone or physiological saline solution from the 8th through to the 1 lth day of gestation. Short-term administration of 915 mg kg-’ dexamethasone induced two types of morphological lens deviations in the offspring (lenses were studied by means of sagittal serial sections). In the first type of deviation a progressive series of changes in the lens bow region was observed. Early changes consisted of an enlargement of some lens bow nuclei, an increase in the number of lens bow nuclei and a disturbance in the shape of the lens bow. Later changes consisted of the degeneration and vacuolization of a peripheral zone of cortex fibre cells. The overall picture showed that the size of up to 8 y0 of the nuclei in the deviating lens bows was enlarged and that the average number of lens bow nuclei in the deviating lenses w&s 28 o/0higher (P < 001) than in the lens bows without deviations. The second type of lens deviation consisted of isolated bundles of disintegrated cortex fibre cells, which extend downwards (anterior-posterior) from subepithelial vesicles. In these bundles all nuclei were smaller and more basophilic than normally. Both types of lens deviations occurred independently in the superficial cortex. Control offspring treated with physiological saline solution showed neither the first nor the second type of lens deviation. Of the offspring exposed to 00375 mg kg-’ dexamethasone one animal (8 %) showed lens changes of the first type only, whereas 59 y/o of the offspring exposed to 915 mg kg-’ dexamethasone displayed either one or both types of lens deviations, Key words: light microscopy; teratogenic lens deviations: prenatal dexamethasone; lens bow region ; cortex fibre cell bundles.
1. Introduction There is ample evidence that systemic as well as topical treatment with glucocorticoids can induce lens opacities in man (Black, Oglesby, Von Sallmann and Bunim, 1960; Spencer and Andelman, 1965; Younessian, 1970; Polak, 1980; Adhikary, Sells and Basu, 1982). However in animal experiments this effect is not clear. Some investigators were not successful in raising lens opacities in rats, rabbits and chickens after administration of glucocorticoids (Von Sallmann, Caravaggio, Collins and Weaver, 1960; Cotlier and Becker, 1965; Bettman, Fung, Webster, Noyes and Vincent, 1968), while others reported lens opacities (Tarkkanen, Esilii and Liesmaa, 1966; Wood, Contaxis, Sweet and Van Dolah, 1967). The cataractogenic effect of some drugs, insufficient diets or certain diseases is potentiated by additional treatment with glucocorticoids (Bettman, Fung and Noyes, 1964; Bettman et al., 1968; Cotlier et al., 1965; Koch, 1976). There is only scarce evidence that prenatal treatment with glucocorticoids may result in cataract development in man (Wells, 1953 ; Kraus, 1975). In mice however, the development of cataracts after prenatal administration of glucocorticoids has been Please address reprint requests to: Jenneke M. A. Koksma at the above address 00144835/84/080195+12
$03.00/0
01984 Academic Press Inc. (London) Limited
196
.I. >I. A. KtbKSM.4
IC’I’ AI,
demonstrated (Rogoyski and Trzcinska-Dahrowska, 1969). Experiment,s with rats support this result (Haumont, 1958). Recently the application of hydrocortisone was shown to induce reversible cataracts in chicken embryos (Nishigori, Lee and Iwatsuru. 1983). The aim of the present study is t’o evaluate the teratogenic effect of dexamethasone on the rabbit lens after short-term prenatal exposure.
2. Materials
and Methods
Female New Zealand White rabbits, weighing 3-4 kg, after mating with male rabbits of the same strain and weight, (one male per two randomly-chosen females), were treated with various doses of dexamethasone (Oradexon: Organon, 0~s). From the 8th to the 1lth day of supposed gestation (the first day of gestation is regarded as day 0) all females received either @0375 mg kg-l dexamethasone, @15 mg kg-’ dexamethasone, or physiological saline solution subcutaneously once daily between 0900 hr and 1000 hr. (The females were randomly distributed in the three dose groups; the doses of dexamethasone corresponded to therapeutic doses given to man.) The offspring were killed at the age of 12 weeks. Both eyes were enucleated and immediately fixed in a 10% formalin solution. One eye per rabbit was selected at random. After dehydration and embedding in celloidin, 10pm saggital serial sections were cut with a Reichert sledge microtome. Every 10th section was stained with haematoxylin and eosin. Two independent light-microscopic examinations of all the specimens were carried out. Morphometry In the central lens sections of both the control offspring and the low and the higher dose dexamethasone-exposed offspring the lens bow nuclei were counted by means of photographs (magnification 63 x ). As each central lens section contained two lens bows, both numbers of nuclei were averaged. Statistical methods The following statistical tests were performed: the X-square test and the exact test of Fisher (to compare percentages), the two sample test of Wilcoxon (to compare two groups) and the test of Kruskal-Wallis (to compare more than two groups). Results were considered significant at the 5 y0 level (two-sided tail probability P < @05).
3. Results In Table I the mean number of neonati and of 1%week-old offspring per group of females is reported. A comparison of both numbers shows that in this study the higher death rate of the offspring before birth on account of the dexamethasone treatment is compensated by a higher degree of survival after birth. The young rabbits never showed gross abnormalities such as palatoschizis, hare lip, club foot or polydactyly. Morphology In the superficial cortex of the lenses of the offspring exposed to the high dose of dexamethasone two types of lens deviations can be demonstrated. (a) Deviation in the lens bow region. The high dose-exposed offspring showed changes in the lens bow region, which can be seen most distinctly in the central lens sections. These changes consisted of a series of progressive alterations, Firstly, in a total of 13 offspring, round enlarged nuclei (up to 8 o/Oof the total number) were observed in the lens bow. The enlarged nuclei are as basophilic as the normal ones. This stage is considered pre-deviant. Secondly, the configuration of the lens bow was disturbed
TERATOGEKIC’CHASGES
IN THE
RABBIT
LEKS
197
TABLE I
Number of newborn and 12-week-old off.yring dexeamethasone
Drxamethasone dose (mg kg-‘) Control 00375 0.15
born to rabbits treated with
Females (n)
Neon&i per female (Mean n)
12-week-old offspring per female (Mean n)
8 7 15
1.2 64 57
2.0 (n = 16) 1.9 (n = 13) 1.9 (n = 29)
F‘IQ. 1. Normal lens bow; control group. The lens bow is formed by the nuclei of the superficial corl ;ex fibr e cellr 1. Central lens section, x 86. Fk3. 2. Disturbed lens bow region, stage 1; @15 mg dexamethasone. the lens 1low are displaced posterially. Central lens section. x X0.
The nuclei in the inner zone of
3b
Fro. 3. Disturbed lens bow region. stage 2; 0.15 mg dexamethasone. Presence of a disturbed lens bow and a lateral zone of degenerated cortex fibre cells. (a) Central lens section, x 54. (b) The lateral zbnes of degenerated cortex fibre cells extend throughout the whole lens. Peripheral lens section, x 25.
in 11 out of the 13 above-mentioned offspring. While in Fig. 1 a normal lens bow is shown, Fig. 2 shows a disturbed one wherein the nuclei in the inner zone of the bow are displaced posterially. Thirdly, of the 11 offspring with a disturbed lens bow configuration, seven showed distinct, and two beginning, stages of cortex fibre cell degeneration. In the cases of distinct fibre cell degeneration, bilateral zones of affected fibre ceils were present [Fig. 3(a)]. These zones extended through the lens towards the periphery [Fig. 3(b)]. Finally, four out of the seven lenses with distinct zones of degenerated cortex fibre cells showed subcapsular vacuolization in the anterior region of the lens [Fig. 4(a)]. The vacuoles, which enlarge and possibly confluence, were formed in the anterior parts of the affected lens fibre cells [Fig. 4(b)]. Figure 4(c) shows a detail of the degenerated fibre cells with enlarged nuclei.
TERATOGENIC
CHANGES
IN THE
RABBIT
LEXS
199
FIG. 4. Final stage of changes in the lens bow region; 0.15 mg dexamethasone. In addition vacuoles are formed in the anterior parts of the degenerated cortex fibre cells. (a) Disturbed lens bow, zone of lateral degenerated cortex fibre cells and subcapsular vacuolization. Central lens section, x 62. (b) Detail of the enlarging and maybe confluencing vacuoles. Interference contrast, x 225. (c) Detail of some enlarged (1) and normal (2) nuclei; many nuclei are disorientated (3), x 360. The enlarged nuclei are present throughout the whole series ofprogressive changes, also in small numbers in the prt-deviant stage. The small, dark spots on the right of the lens bow are artifacbs.
In the offspring of the low dose dexamethasone-treated females this type of lens deviation was observed once. The deviation consisted of changes in the lens bow region (round, enlarged nuclei ; disturbed shape of the lens bow : bilateral zones of degenerated cortex fibre cells and subcapsular vacuolization). (b) Degeneration of isolated bundles ofcortex$bre cells. In the deviating lenses bundles of disintegrated cortex fibre cells, which extend downwards from subepithelial vesicles, were present [Fig. 5(a)]. The vesicles were filled with a fluid substance. The affected fibre cells stained scantily with eosin. They had small, round, dark-staining nuclei compared with the surrounding normal fibre cells [Fig. 5(b)]. The cytoplasm also contained many lightly eosin-staining globules. which demonstrates the disinte-
200
.J. 11. A. KOKSMA
ET AL.
FIQ. 5. Degeneration of isolated bundles of cortex fibre cells; 0.15 mg dexamethasone. (a) Bundles of degenerated cortex fibre cells, which extend downwards from subepithelial vesicles and stain scantily with eosin. Peripheral lens section, x53. (1)) Uptail of the helt with nuclei: the small. round, very basophilic nuclei in the degenerated bundles surrounded by the oblong, less basophilic. normal nuclei, x 252. (c) Detail which shows globules (arrow) in the cytoplasm of the degenerated cortex fibre cells. Interference contrst, x 360.
TERATOGENIC
CHANGES
IN THE
RABBIT
201
LENS
Fro. 6. Localization of the two types of lens deviations (scheme; frontal view of the lens). Shaded area: zone with changes in the lens bow region. Black dots: bundles with degenerated cortex fibre cells.
gration of the lens fibres [Fig. 5(c)]. Of interest is that this deviation occurred on several places in the equatorial part of the lens, while the alterations in the lens bow region were present as a continuous zone in a more extended part of the periphery (Fig. 6). Table II reports the number of young rabbits with each type of lens deviation in the three dose groups. In the table the percentages in the dose groups show significant differences [Table II(a) P < O-01; Table II(b) P < O*OOl]. The two types of lens deviations occurred separately as well as together. Table III shows the distribution of both types of changes among the offspring exposed to the higher dose of dexamethasone. A relation between both types of deviations is not found. In total 17 of the 29 young rabbits (59 %) sh owed either one or both types of lens deviations. Morphometry Counting of lens bow nuclei. In Table IV(a) the results are registered of the counts of the nuclei in the central lens bows of the 16 control lenses and of the 13 and 29 lenses originating from the offspring exposed to the low and the higher dose of TABLE
The distribution
II
among the rabbit offspring
of the two types of lens deviations Dexamethesone dose (mg WI
Offspring (nl
Offspring with lens deviation (n)
(a) Changes in the lens bow region* Control 16 00375 13 0.15 29 (b) Degeneration of isolated cortex fibre cell bundles? Control 16 00375 13 0.15 29 * The distribution of the deviation P < 0.01). t Idem (x-square test, P < 0901).
among the dose-groups
0 (0%) 1(8%) 11 (38%) 0 (0%) 0 W%l 13 (45%) differs
significantly
(X-squaretest,,
202
.I. M. A. KOKSMA
Distribution
ET AI,
of the two types of lens deviations twnong &a-e of dexamethasone*
the offspring
exposed
to the high,
Changes in the lens how region Present Degenerated cortex fibre cell bundles Total number * No significant
Absent
Total number
Present
7
6
13
Absent
4 11
1% 18
16 29
relation between both types of deviations
is found (Fisher’s test)
dexamethasone. According to the test of Kruskal-W&s, the levels of the numbers of lens bow nuclei in the three dose-groups differ significantly (P < O-05). Since in the low dose-group only one abnormal lens was observed, the lenses exposed to the higher dose of the drug were divided into four groups depending on the presence or absence of one or both types of lens deviations [see Table IV(b)]. The numbers of lens bow nuclei derived from the two groups of lenses which showed no changes in the lens bow region (groups 1 and 2) did not differ significantly from each other. The same held for the numbers of lens bow nuclei derived from the two groups of lenses which showed changes in the lens bow region (groups 3 and 4). Therefore groups 1 and 2 on the one side, and 3 and 4 on the other side were pooled and both resulting groups compared with the control group. A significant difference in the number of lens bow nuclei was TABLE
Median
IV
number of the lens bow nuclei Offspring (n)
(a) Dose DEX* (mg kg-‘) control @0375 mg kg-’ DEX 0.15 mg kg-’ DEX
in central lens sections Median number (extreme values)
16 13 29
38% (312,540)t 436 (360,500)t 459 (278. 602)t
offspring I2
447 (330. 507)
(2) Degeneration of isolated cortex fibre cell bundles only
6
4244 (278. 482)
(3) Disturbed lens bow region only
4
541 (476. 561)
(4) Both types of deviations
7
490 (398. 602)
(b) Group of @15 mg kg-’ DEX-exposed (1) R’o deviations
43% (278, 507)f
500 (398,602)s * t $ §
DEX = Dexamethasone. Significantly different (test of Kruskal-Wallis. P c @05). Not significantly different from the control value (test of Wilcoxon). Significantly different from the control value (test of Wilcoxon, P < @Ol).
TERATOGENIC
CHANGES
IN THE
RABBIT
LEES
203
found for the group of lenses with lens bow deviations (P < O*Ol), but not for the group of lenses without lens bow deviations. In conclusion, the lenses with deviations in the lens bow region contained significantly more nuclei than the control lenses. 4. Discussion In this study the offspring of female rabbits treated with dexamethasone for a short period during early pregnancy showed two types of lens deviations. As in the study of Rogoyski et al. (1969) the lenses were studied morphologically. In the first type of lens deviation an enlargement of a part of the lens bow nuclei, an increase in the number of lens bow nuclei, and a disturbance in the shape of the lens bow, followed by degeneration and vacuolization of a zone of cortex fibre cells were observed (Figs. 2, 3 and 4). These deviations are comparable to the changes found in various forms of hereditary cataract in mice (Hamai, Fukui and Kuwabara, 1974; Burns, Anderson and Feeney-Burns, 1980; Uga, Kador and Kuwabara, 1980; Kuck, Kuwabara and Kuck 1981-82). However, the hereditary cataracts mentioned have different features, i.e. in some studies the number of nuclei in the lens bow has increased (Hamai et al., 1974; Uga et al., 1980), while in others it has decreased (Burns et al., 1980; Kuck et al., 1981-82). Furthermore, the localization of the swollen lens fibres and the vacuoles is mostly in the posterior cortex, in contrast to the anterior-lateral position reported in this study. In the lenses with changes in the lens bow region we found, in comparison with the control lenses, an average increment of 28 o/0 in the number of lens bow nuclei. This increment may be caused by either a delay in denucleation of the superficial cortex cells (Hamai et al., 1974; McAvoy and Van Heyningen, 1976: Uga et al., 1980), or by an increase in the formation of cortex fibre cells by the action of dexamethasone. Since in vitro experiments (Van Venrooy, Groeneveld and Bloemendal, 1974) have demonstrated an increased transition of lens epithelial cells into cortex fibre cells after the addition of dexamethasone, we favour the latter hypothesis. The second type of lens deviation outlined in this study includes the degeneration of isolated cortex fibre cell bundles (Fig. 5). To our knowledge this deviation has not been reported before. Two doses of dexamethasone were applied, namely 90375 mg kg-’ and O-15 mg kg-l, which correspond to about 19 mg and 74 mg prednisone-equivalents in man. Almost all lens changes were found in the offspring exposed to the. higher dose of dexamethasone (see Table II). To establish if the two types of lens deviations have the same origin or not, various characteristics of both types of changes were compared in the lenses exposed to the higher dose of dexamethasone. In the lenses with degenerated cortex cell bundles and without lens bow changes (see Fig. 7) we found the same number of lens bow nuclei as in the control lenses; lenses with deviations in the lens bow region contained on average 28% more nuclei than the control lenses [Table IV(b)]. Furthermore, the nuclei of the degenerated cortex cell bundles were small, round and highly basophilic [Fig. 5(b)], whereas in the lenses with a disturbed lens bow region up to 8 o/0 of the nuclei were enlarged and as basophilic as the control nuclei [Fig. 4(c)]. Finally, the two types of lens deviations, although occurring together as well as separately, did not seem to be related statistically (Table III). Hence, we conclude that the two types of lens deviations reported here have developed independently. A comparison of the peripheral sections [Figs. 3(b) and 5(a)] and of the central
7
FIQ. 7. Lens with degenerated cortex fihre cell bundle and normal lrns how: 0.1.’ mg drxamethasone. Central lens section. X 90.
sections [Figs. 2, 3(a), 4(a) and 71 of lenses with either the first or t,he second type of lens deviation also demonstrates the different character of both types of deviations. Moreover, the fact that in the first type of deviation the lens bow is involved and that the cortex cells whose nuclei compose the lens bow derive from lens epithelium cells (Hanna and O’Brien, 1961; Fine and Yanoff, 1979; Beebe, Compart, Johnson, Feagans and Feinberg, 1982) indicate that the first type of lens deviation may develop after a complicated series of events resulting in the degeneration and vacuolization of a peripheral zone of cortex fibre cells (Kinoshita, 1974). The results of the present study suggest that this type of deviation develops in another way and in another period than the second type of lens deviation. In the second type of lens deviation the bundles of degenerated cortex fibre cells with small, round nuclei may be induced after the formation of the subepithelial vesicles, which cut off groups of underlying cortex cells from the feeding epithelium. The experimental set-up of the present investigation is very similar to that of Haumont (1958) and Rogoyski et al. (1969). Haumont states that some offspring of female rats treated with cortisone during pregnancy show cataract formation. Rogoyski et al. (1969) examined the fetuses of female mice treated daily with a high dose of hydrocortisone during, e.g., the 9th and 10th day of gestation; based on the mean weights of mice and men, the dose of the drug corresponds to about 875 mg prednisone-equivalents in man. After this treatment the fetuses show a high percentage (31%) of morphological cataract. It is not clear whether the degenerative changes, which are most frequently found in the secondary nuclear fibres, consist of swelling or of vacuolization. Changes in the composition or shape of the lens bow are not reported. Nishigori et al. (1983) describe the induction of a reversible cataract in chicken fetuses after injection of a moderately high dose of hydrocortisone corresponding to about 30 mg prednisone-equivalents in man. Since the examination occurred with the slit-lamp, it is not known if concomitant changes in the morphological structure might be demonstrable. However, the cataractous changes found by Rogoyski et al. (1969) and Nishigori’et al. (1983) occur in the fetal stages of the laboratory animals, whereas
TERATOGENIC
CHANGES
IX THE
RABBIT
LENS
20.5
in the present investigation changes were observed in the lenses of animals 12 weeks after birth. The present lens changes were found in the superficial cortex only; the embryonal, fetal and early postnatal nucleus had a normal appearance. Various possibilities arise as regards the developmental line of the present lens deviations. The first is that, dependent on the species of the experimental animal and/or the type of glucocorticoid applied, the morphological expression of the lenticular changes is observable in the fetal stage or after a time delay in the postnatal stage. Secondly, exposure to dexamethasone may produce morphological changes in the rabbit lens, starting in the prenatal stage and continuing for some time (see later) after birth, but because of, e.g., natural growth (see Nishigori et al., 1983) changes in the embryonal, fetal and early postnatal nucleus may disappear. Finally, the fetuses, which are reabsorbed by the mother because of the treatment with dexamethasone (21%, see Table I), might have shown severe embryonal and fetal lens damage. The present types of lens deviations are localized in a peripheral zone in the cortex (see Fig. 6). Compared with the size of lenses of 28-day-old-i.e. nearly full-grownfetuses, the morphological expression of the changes observed at the 12-week stage, must have started at some time after birth. Figs 2,3(a) and 4(a) show that the changes in the lens bow region cease before the 12th week after birth, since outside the disturbed lens bow region normal cortical fibres are present. Presumably the lens deviation consisting of the bundles of degenerated cortex fibre cells has a zonular character as well. As mentioned before, the two types of lens deviations probably developed differently. Glucocorticoids influence lens metabolism in several ways. In addition to the already mentioned effect on the lens epithelium (Van Venrooy et al., 1974) which may be involved in the development of the deviation in the lens bow region, they act on the lenticular content of glucose and various cations (Harris, 1966; Apponi Battini, 1968; Benatti and Balli, 1971). Since in this study only histological examinations were performed, it is not relevant to discuss the possible biochemical pathways leading to each type of lens deviation. In conclusion, two independent types of lens deviations were demonstrated in the eyes of rabbit offspring after short-term prenatal exposure to dexamethasone. ACKNOWLEDGMENTS The authors wish to thank Prof. Dr G. M. Bleeker, Prof. Dr J. van Limborgh and Dr H. A. Brouwer for their encouragement and valuable advice during this study; Dr P. D. Bezemer C.E. for his expert statistical assistance; Mr J. Bult for correcting the manuscript and Mrs T. Kierks for secretarial help. REFERENCES Adhikary, H. P., Sells, R. A. and Basu, P. K. (1982). Ocular complications of systemic steroid after renal transplantation and their association with HLA. Br. J. OphthalmoE. 66,29&l. Apponi Battini, G. (1968). Ueber den Glukose- und Glykoproteingehalt im Kammerwasser, im Glaskiirper und in der Linse der Kaninchen naeh Behandlung mit Glukokortikoiden. Min. Mbl. Augenheilk. 152, i’OS-14. Beebe, D. C., Compart, P. J., Johnson, M. C., Feagans, D. E. and Feinberg, R. N. (1982). The mechanism of cell elongation during lens fiber cell differentiation. Dev. Biol. 92, 54-9. Benatti, C. and Balli, F. (1971). Cataract caused by long-term corticosteroid therapy and its prevention. Minerva Pediatr. 23, 1599-l. Bettman, J. W., Fung, W. E. and Noyes, P. P. (1964). Potentiating action of prednisone on galactose cataracts in rats. Invest. Opthalmol. 3, 678-9.
‘06
.l 11. A. KOKSMA
F:T ;il,.
Bettman. J. W., Fung, W. E., W’rbster. 1~. G., Noyes. P. I’. and Vincent, N. J. (1968). The cataractogenic effect, of corticosteroids on animals rim. J. Ophthalmol. 65, 581.-6. Black, R. L.. Oglesby. R. B.. Von Sallmann, I,. and Bunim. .J. .J. (1960). .J. Am. Jfed. A4~.s~~c. 174, 16671. Burns, R. I’., Anderson, R. S. and Feeney-Burns, L. (1980). C’ataract-webbed trait in Peromyscus. TT Riomicroscopy and histology of eyes. Invest. Ophthdmol. Vis. ,Qi. 19, 3142. Cotlier, E. and Becker, B. (1965). Topical corticosteroids and galactose-cataracts. I~~l;e,st. OphthaZmoZ. 4. 80614. Fine, B. S. and Yanoff. M. (1979). Ocular Histology; A Text and Atlas (2nd edn). Pp. 149-53. Harper and Row Pub]. Inc., Hagerstown, MD. Hamai, Y ., Fukui, H. N. and Kuwabara, T. (1974). Morphology of hereditary mouse cataract. Exp. Eye Res. 18, 53746. Hanna, C. and O’Brien, J. E. (1961). Cell production and migration in the epithelial layer of the lens. Arch. Ophthalmol. 66, 103-7. Harris, .J. E. (1966). The temperature-reversible cation shift of the lens. Trans Am. Ophthalmol. Sot. 64, 67.599. Haumont, St. (1958). Effets de la cortisone administree pendant la gestation. Ann. d’EndocGnol. 19, 442-5. Kinoshita, ,J. H. (1974). Mechanisms initiating cataract formation. Invest. Ophthdmol. 13, 713-24. Koch, H. R. (1976). Klinische und experimentelle IJntersuchungen iiber den Einfluss von Corticosteroiden auf die Augenlinse. Pp. 88-93. Ph.D. Thesis, University of Bonn. Kraus, A. M. (1975). Developmental cataracts. N. Y. State J. Med. 75, 1757-8. Kuck, J. F. R., Kuwabara. T. and Kuck, K. D. (1981-82). The Emory mouse cataract--an animal model for human senile cataract. Curr. Eye Res. 1, 643-51. McAvoy, J. W. and Van Heyningen. R. (1976). Changes in the cells of the lens bow and epithelium of tryptophan-deficient rats. INSERM 60, 245-50. Nishigori, H., Lee, J. W. and Iwatsuru. M. (1983). An animal model for cataract research: cataract formation in developing chick embryo by glucocorticoid. Exp. Eye Res. 36, 617-22. Polak, B. C. P. (1980). Ophthalmological Complications of Haemodialysis and Kidney Transplantation. Dr W. Junk (Pub].), The Hague. Rogoyski. A. and Trzcinska-Dabrowska, Z. (1969). Corticosteroid-induced cataract and palatoschisis in the mouse fetus. Am. J. OphdhaZmoZ. 68, 128-33. Spencer, R. W. and Andelman, S. Y. (1965). Steroid cataracts. Posterior subcapsular cataract formation in rheumatoid arthritis patients on long term steroid therapy. Arch Opt?&. 74, 3841. Tarkkanen, A., Esila, R. and Liesmaa, M. (1966). Experimental cataracts following long-term administration of corticosteroids. Acta Ophthalmol. 44, 665-8. IJga, S., Kador, P. F. and Kuwabara, T. (1980). Cytological study of Philly mouse cataract, Exp. Eye Res. 30, 74-92. Van Venrooy, W. J., Groeneveld, A., Bloemendal, H. and Benedetti, E. L. (1974). Cultured calf lens epithelium. II The effect of dexamethasone. Exp. Eye Res. 18, 527-36. Von Sallmann, L., Caravaggio L. L., Collins, E. M. and Weaver, K. (1960). Examination of lenses of steroid-treated rats. Am. J. OphthaZmoE. 50 1147-51. Wells, C. N. (1953). Treatment of hyperemesis gravidarum with cortisone. Am. J. Obst. Gynec. 66, 599601. Wood, D. C., Contaxis, I., Sweet, D., Smith, J. C. and Van Dolah, J. (1967). Response of rabbits to corticosteroids. I Influence on growth, intraocular pressure and lens transparency. Am. J. Ophthalmol. 63, 841-8. Younessian, S. (1970). Glaucome et cataracte apres corticotherapie par voie g&&ale et locale. Adv. Ophthalmol. 23, 74-152.