[Prefeeding of aldose reductase inhibitor inhibits galactose cataract].

To examine the effect of prefeeding the aldose reductase inhibitor (ARI) on the cataract induced by the subsequent feeding of galactose diet, rats were first fed rat chow with or without ARI for 2 weeks. The animals were then transferred to a 25% galactose diet for different periods of time. Lenses were enucleated and examined by light microscopy. The amount of galactitol in the lenses was examined biochemically. The cataract was less severe and there was less galactitol in the lenses in the rats first fed with ARI than in those without ARI. These results indicate that the feeding the ARI prior to galactose feeding partly inhibits galactose cataract.

Proliferative changes of lens epithelial cells in rat and mouse galactose cataracts--examination using whole-mount preparations.

The proliferative activity of the lens epithelium in the early stages of cataract crisis was investigated in rats and mice using the 3H-thymidine autoradiographic method with the whole-mount preparations of total lens epithelial cells. Three-week-old Sprague-Dawley (SD) rats were given a diet which included galactose in three different concentrations (15%, 25%, 50%) to produce sugar cataracts of three different degrees. Seven-week-old ICR mice were given a 50% galactose-diet. In the lenses of control rats and mice, 3H-thymidine labeled cells were observed mainly in the germinative zone at the lens equator; a few labeled cells were detected in the anterior subcapsular central zone. In the lenses of the SD rats on the 4th day of the diet, labeled cells increased remarkably in the central zone. However, labeled cells decreased as the cataract progressed. The peak in the number of labeled cells was observed on the 4th day of the diet regardless of the galactose concentrations, and was not proportional to the degree of the cataracts. In the galactose-fed ICR mice, the blood galactose level was high, but there was no increase in the number of labeled cells or the development of galactose cataract. The marked increase of labeled cells in the central zone in the SD rat lenses had probably occurred because of the accumulation of galactitol.

[The localization of aldose reductase mRNA in the rat lens].

The localization of aldose reductase (AR), which is thought to be involved in the pathogenesis of sugar and galactosemic cataracts, has been studied using various methods. The AR exists in the lens and its presence is considered to be important at the site of the cataractogenesis. In this study, the localization of AR messenger RNA (ARmRNA) in the lenses of 3-week old and fetal rats was studied by in situ hybridization using a complementary DNA to the rat AR gene. In the lens of 3-week-old rats, ARmRNA was present in the lens-epithelium and the equatorial superficial cortex. The distribution profile was similar to that of AR-immunoreactivities revealed by an immunocytochemical study. In the fetal lens, however ARmRNA was present in the epithelium and the equatorial region, while AR-immunoreactivity was observed mainly in the center of the lens.

Localization of vasoactive intestinal peptide (VIP) messenger RNA (mRNA) in amacrine cells of rat retina.

We detected vasoactive intestinal peptide (VIP) messenger RNA (mRNA) in the rat retina using an in situ hybridization technique and a 35S-labelled cDNA probe. VIP mRNA was present in the cells of the inner nuclear layer (INL). The VIP mRNA-positive cells showed a distribution similar to that of the VIP-like immunoreactive amacrine cells. This observation suggests that VIP mRNA undergoes transcription in the VIP-immunoreactive amacrine cells.

[Ciliary body changes associate with aldose reductase in galactosemic rats (2)].

Morphological changes of the ciliary body epithelium in rats fed a 50% galactose diet for 18 months with or without aldose reductase inhibitor (ARI: FR74366, Fujisawa) were investigated on coronal sections by both the light microscopic observation and the measurement of the area of non-pigmented epithelium (non-PE) as the ratio of non-PE/PE. The measurement was performed with color image analyzer. Structural change (hypertrophy) was observed only in non-pigmented epithelial cells of galactosemic rats. In contrast, no hypertrophic changes were seen in galactosemic rats treated with FR74366. These results strongly suggest that aldose reductase is associated with hypertrophic change of non-pigmented epithelial cells of the ciliary body as one of the diabetic complications.

[Differences in cataractogenesis according to neodymium-YAG laser injury site].

The author examined morphological changes of the lens induced by mechanical damage with a Q-switch Nd-YAG laser (anterior capsule, anterior subcapsular deep cortex, posterior capsule). In the ruptured anterior capsule group, epithelial cell proliferation covered the ruptured capsule, and separation of posterior subcapsular sutures and swelling of the posterior subcapsular end of lens fiber cells were observed. In the ruptured subcapsular deep cortex group, swollen lens fiber cells were observed in both the anterior subcapsular and the posterior subcapsular cortex. In the ruptured posterior capsule group, the ruptured cortex was not repaired. Swollen lens fiber cells were first recognized in areas surrounding the ruptured capsule, then at the posterior side of the equator, and finally at the anterior subcapsular cortex. The continuity of the lens capsule and lens fiber cells themselves is very important part to maintain lens clarity. The destruction of this feature induced swelling of fiber cells on the opposite side of the injury area.

[Formation of congenital cataract in the offspring of galactosemic rats].

It has been established that lenses of offspring of galactosemic rats form sugar cataracts. In the present study, in order to clarify the critical period in pregnancy for the cataract formation, lenses of offspring of pregnant rats which were fed galactose diets at each of three terms of pregnancy, were examined histologically and biochemically. Severe cataract and a large amount of galactitol in the lenses were found in the offspring of rats fed a galactose diet in the last term of pregnancy. Moreover, focal cataract was found in the offspring of rats fed a galactose diet in the middle term. These results indicate that the critical period in pregnancy for cataract formation of the offspring is the last term of pregnancy and that cataract formation begins in the middle term in offspring of galactosemic rats.

[Cell kinetics of lens epithelial cells on the whole-mount preparation (3). Changes in mouse cataract lens by feeding galactose].

In this study, we used ICR (Institute of Cancer Research) strain mouse because they showed hypergalactosemia but not cataractogenesis. We examined the proliferative ability of the lens epithelial cells by means of a whole-mount preparation of epithelium and 3H-thymidine autoradiography. An increased number of labelled cells was not found in the mouse. The increased proliferative ability of epithelial cells in rat galactose cataract did not seem to be caused by hypergalactosemia.

[The influence of aging on the development of rat galactosemic cataract].

In order to investigate the effect of age on the rat galactosemic cataract, 3, 6, 12 and 24-week-old rats were fed with a 50% galactose diet. The cataractous lenses were observed by light microscopy, and the amount of galactitol measured. All rats fed with 50% galactose developed cataracts in the equatorial region. However, the morphological and biochemical development of the galactose cataract in old rats was slower than in young rats. These results suggested that older diabetic patients are less likely to develop diabetic cataract than younger patients.

[Quantitative study of rat diabetic cataract, by the relaxation times of nuclear magnetic resonance].

The prophylactic effects of a new aldose reductase inhibitor (ARI), FR74366 on streptozotocin-induced rat diabetic cataract were examined by means of proton nuclear magnetic resonance (1H-NMR) relaxation time. We compared the findings with the histological finding, and it was recognized that longitudinal and transverse relaxation times (T1, T2) were prolonged before the histological changes appeared. The ARI, FR74366, prevented histologic changes and had detected by the 1H-NMR method. The results showed that 1H-NMR could be useful in the early detection of human diabetic cataract and the evaluation of the effectiveness of anti-cataract agents, for example, AR inhibitors.

[Changes in cell kinetics of rat lens epithelial cells with aging].

Whole-mount preparations of developing rat lens epithelium were made and 3H-thymidine autoradiography was performed on them. The distribution of 3H-thymidine-labeled (DNA-synthesizing) epithelial cells was examined by the above combined methods. Labeled cells were evenly distributed in the epithelial cell layer (not the definitely germinal zone) until 1 week after birth. In 3- or 7-week-old rat lens, labeled cells were mainly distributed in the germinal zone, and the number of labeled cells localized in the anterior subcapsular epithelial cell layer decreased according to age of the rat. The 24-week-old rat showed localization of labeled cells at the germinal zone, which seemed to be similar to the human adult lens.

[Preventional and therapeutic effects of aldose reductase inhibitor FR74366 on rat galactose cataract].

Preventional and reversal effects of a new aldose reductase inhibitor FR74366 on the rat galactose cataract were examined biochemically and light microscopically. A high concentration (0.075%) FR74366 showed sufficient effects both to prevent and reverse cataract, but low concentrations (0.025%) FR74366 was not effective as 0.075% FR74366. These results indicate that FR74366 can be useful drug for cataract.

[Movement of regenerated lens epithelial cells in 50% galactose cataract and an aldose reductase inhibitor].

In experimental galactose cataract of rats, lens fiber cells were gradually destroyed by swelling and liquefaction occurring due to the accumulation of galactitol. In addition to the destruction of lens fiber cells, marked regeneration of lens epithelial cells was universally observed. In this study, we used the technique of 3H-thymidine autoradiography and examined the movement of regenerated (DNA synthesis) lens epithelial cells. On the fourth day from the starting of 50% galactose chow, 3H-thymidine was injected into the anterior chamber. After one week, 3H-thymidine-labelled epithelial cells was observed at the bow region of the equatorial area. After 2 to 4 weeks, labelled cells were found in the regenerated lens fibers of the cortex. After 3 weeks, labelled cells about to be destroyed by swelling and liquefaction were recognized. However, a few labelled cells were still observed in the epithelial cell layer. The same experiments were performed in two groups of normal chow-fed one and 50% galactose + an aldose reductase inhibitor (Statil)-fed one. The movements of 3H-thymidine labelled epithelial cells in the above two groups were almost identical. In addition, the movement of labelled cells was normalized by an aldose reductase inhibitor.

Localization of aldose and aldehyde reductase in the kidney.

The distribution of NADPH-dependent reductase activity in the rat cortex, outer medulla and inner medulla was investigated through biochemical and histochemical methods. Biochemical studies revealed reductase activity to be present in all three regions of the kidney with the highest specific activity observed in the inner medulla, followed by the cortex and the outer medulla. Activity in all three regions was inhibited by the aldose reductase inhibitors sorbinil, tolrestat and 7-hydroxychromone-2-carboxylic acid. Based on substrate utilization and response to sulfate on the inhibitors, the inner medulla contains primarily aldose reductase (EC 1.1.1.21) while the cortex contains primarily aldehyde reductase (EC 1.1.1.2). The outer medulla contains a mixture of both enzymes. This distribution was confirmed by a radioimmunoassay for aldose reductase. Immunohistochemical investigations of the rat kidney with antibodies against rat lens aldose reductase and rat kidney aldehyde reductase revealed a similar distribution of these enzymes. Aldehyde reductase was immunohistochemically detected only in the cortex where it was localized in the proximal convoluted tubules. Immunoreactive aldose reductase was detected in Henle's loop at both the inner stripe of the outer medulla and in the inner medulla, and in the collecting tubules and the epithelial cell lining the pelvis of the inner medulla near the papilla. No specific immunohistochemical staining for aldose reductase was observed in the cortex. A similar immunohistochemical distribution of aldose reductase was also observed in the human kidney with antibodies against human placental aldose reductase.

[Ciliary body changes associate with aldose reductase in galactosemic rats].

Morphological changes of the ciliary body epithelium in rats fed a 50% galactose diet for 10 months with or without aldose reductase inhibitor (ARI) were investigated by both light microscopic observation and measurement of cell size. Aldose reductase was immunohistochemically observed to be present in non-pigmented epithelial cells of ciliary body and structural change (hypertrophy) was observed only in non-pigmented epithelial cells of galactosemic rats. In contrast, no hypertrophic changes were seen in rats fed with ARI. These results strongly suggest that aldose reductase is associated with hypertrophic change of non-pigmented epithelial cells of the ciliary body.

[Morphological classification of the prevention by aldose reductase inhibitor of rat galactosemic cataract].

A morphological classification indicating the stage of the prevention of aldose reductase inhibitor (ARI) on the rat galactosemic cataract was investigated. Three week old rats (body weight 50 g) were fed with a 25% galactose diet. Two kinds of ARIs (ADN-138, FR-74366) were used with a different dose 25% galactose fed each drug. The rats on the 25% galactose diet rapidly developed cataracts in the equatorial region of the lens followed by total cataracts. The morphological processes of the galactose cataracts receiving preventive treatment of ARI were divided into 5 types according to their intensity.

Vasoactive intestinal peptide (VIP)-like immunoreactive neurons located in the rat suprachiasmatic nucleus receive a direct retinal projection.

The existence of a direct projection from retinal ganglion cells to vasoactive intestinal peptide (VIP)-like immunoreactive neuronal elements in the rat suprachiasmatic nucleus (SCN) was revealed by combining analysis of degenerating axons following enucleation and electron microscopic immunocytochemistry. Degenerating axons appeared to make synaptic contact with VIP-like immunoreactive dendrite and neuronal perikarya in the ventral part of the SCN. The possibility of neuronal input from retinal ganglion cells to axons of VIP-like immunoreactive neurons was also suspected since axo-axonic synapses were detected between degenerating axons and axons with VIP-like immunoreactivity. Thus, VIP-like immunoreactive neurons in the SCN receive several neuronal inputs, including those from the retina, and may play a significant role in circadian entrainment.

The effect of aldose reductase inhibitor, statil, on hyperpermeability of iridial vessels in experimental galactosemic rats.

Thirty young male Sprague-Dawley rats were divided into three groups: the control group, given a normal diet; the galactose group, given a 25% galactose diet; and the aldose reductase-galactose group, given a 25% galactose diet containing 0.046% Statil, an aldose reductase inhibitor. The results suggested that Statil could prevent hyperpermeability of iridial vessels in galactose-fed rats.