Eyes Of Diabetic Rats Research Articles

Prospective insulin-based ophthalmic delivery systems for the treatment of dry eye syndrome and corneal injuries

The presence of insulin (INS) receptors on the ocular surface (OS) and lacrimal gland (LG), and the high prevalence of dry eye syndrome (DES) and corneal lesions in diabetic patients suggest that INS is relevant for OS homeostasis and wound healing. The study aims at developing delivery systems for the topical administration of INS to the OS in order to improve INS local bioavailability and evaluate the influence of the delivery systems on DES in diabetic rats (DM) (n = 05/group). Chitosan microparticles (MP), chitosan/poloxamer gel (GEL) and MP-loaded GEL (GELMP), with or without INS were developed. Formulations were instilled into the eyes of diabetic rats (DM) for 15 days and the tear fluid volume, corneal cells morphology and cornea thickness were assessed and compared with an aqueous dispersion of INS (DISP-INS). All delivery systems had pH of about 6, osmolality suitable for topical application and positive zeta potential. The MPs with or without INS had sizes close to 4 μm, spherical morphology and INS encapsulation efficiency of 77 ± 6%. DISP-INS and GELMP-INS formulations produced tear secretion amounts significantly higher than those receiving formulations containing no INS and similar to healthy animals. Cornea surface impression cytology showed that treatment with INS-delivery systems and not DISP-INS almost normalized cells morphology. Treatment with GELMP-INS increased INS by 2.5 in the LG and eyeball as compared to the groups treated with GEL-INS and MP-INS, while treatment with DISP-INS left no traces of drug in the eye after treatment termination. GEL and GELMP containing INS were also able to normalize the thickness of the corneal epithelia. In conclusion, GELMP-INS normalized tear fluid volume, corneal thickness, protected corneal cells morphology and increased ocular bioavailability of INS, making it a promising treatment strategy for DES and corneal lesions.

P38-MAPK pathway is activated in retinopathy of microvascular disease of STZ-induced diabetic rat model.

To investigate the role of corticotropin releasing hormone (CRH) in diabetic retinopathy of microvascular disease and the potential mechanism. The diabetic rat model was constructed by a single intraperitoneal injection of streptozotocin (STZ). The expression of CRH in the retina of diabetic rats and wild-type rats was detected by Real-Time Polymerase Chain Reaction (RT-PCR). CRH shRNA or Scr shRNA adenovirus was injected into the eyes of diabetic rats and wild-type rats, respectively. The effect of down-regulated CRH on visual electrophysiology in rats was evaluated. Protein expressions of vascular endothelial growth factor (VEGF) and inflammatory factors that were related to the microvascular lesion after CRH downregulation were detected by Western blot. Furthermore, p38 expression was detected by Western blot to explore whether mitogen-activated protein kinase (MAPK) signaling pathway was involved in the function of retinal endothelial cells regulated by CRH. The expression of CRH was significantly up-regulated in the retina of diabetic rats. RT-PCR results showed that the mRNA level of CRH in the retina of diabetic rats injected with CRH shRNA was decreased. However, no significant change in CRH level was observed in rats injected with Scr shRNA adenovirus. The down-regulated CRH could improve the diabetes-induced visual impairment and retinal inflammatory response. Moreover, the down-regulated CRH led to a decreased phosphorylation level of p38. CRH improves the diabetic retinopathy of microvascular disease via the p38-MAPK pathway, which is expected to be a new target for the treatment of diabetic microangiopathy.

The Expression of Calcium Sensing Receptor in Normal and Diabetic Rat Eyes.

BackgroundCalcium sensing receptor (CaSR) is widely expressed in many tissues of the body, but it is rarely reported to be expressed in the eyes. This research explored the expression and distribution of CaSR in eye tissues of normal and diabetic rats.Material/MethodsThirty male Sprague Dawley (SD) rats were randomly divided into a diabetic and a normal group. Diabetes mellitus (DM) models were successfully established by intraperitoneal injection of streptozotocin. The expression and distribution of CaSR in the rat eyeballs were detected by immunohistochemistry. Quantitative RT-PCR and western blotting were used to detect the presence of CaSR in normal and diabetic rats.ResultsCaSR was detected in the cornea, lens epithelium, and retina. CaSR was expressed the most in the cornea, followed by the lens epithelium, and the retina (p<0.05). The expression of CaSR was decreased in the eye tissue of diabetic rats (p<0.05).ConclusionsIn this study, CaSR was detected in rat cornea, lens, and retina. It was significantly decreased in the eyes of diabetic rats. This indicated that the downregulated expression of CaSR was associated with diabetic oculopathy.

Impaired Rhodopsin Generation in the Rat Model of Diabetic Retinopathy

Diabetic retinopathy is a common complication of diabetes mellitus. Diabetic patients experience functional deficits in dark adaptation, contrast sensitivity, and color perception before microvascular pathologies become apparent. Herein, we evaluated early changes in neural retinal function and in retinoid metabolism in the eye in diabetes. Streptozotocin-induced diabetic rats showed decreased a- and b-wave amplitudes of scotopic and photopic electroretinography responses 4 months after diabetes induction compared to nondiabetic controls. Although Western blot analysis revealed no difference in opsin expression, rhodopsin content was decreased in diabetic retinas, as shown by a difference in absorbance. Consistently, levels of 11-cis-retinal, the chromophore for visual pigments, were significantly lower in diabetic retinas compared to those in controls, suggesting a retinoid deficiency. Among visual cycle proteins, interphotoreceptor retinoid-binding protein and stimulated by retinoic acid 6 protein showed significantly lower levels in diabetic rats than those in nondiabetic controls. Similarly, serum levels of retinol-binding protein 4 and retinoids were significantly lower in diabetic rats. Overall, these results suggest that retinoid metabolism in the eye is impaired in type 1 diabetes, which leads to deficient generation of visual pigments and neural retinal dysfunction in early diabetes.

Light Modulates Ocular Complications in an Albino Rat Model of Type 1 Diabetes Mellitus.

PurposeThe purpose of the study was to assess potential interactions of light exposure and hyperglycemia upon ocular complications in diabetic rats.MethodsStreptozotocin-induced (STZ-induced) diabetic rats (N = 39) and non-diabetic rats (N = 9) were distributed into eight groups according to the irradiance and color of the light phase during the 12/12-hour light/dark regime. Follow-up lasted 90 days and included assessment of cataract development and electroretinogram (ERG) recordings. Stress to the retina was also assessed by glial fibrillary acidic protein immunocytochemistry.ResultsCataract development was fast in diabetic rats that were exposed to unattenuated white light or to bright colored lights during the light phase. Diabetic rats that were kept under attenuated brown or yellow light during the light phase exhibited slower rate of cataract development. Electroretinogram responses indicated very severe retinal damage in diabetic rats kept under bright colored lights in the blue-yellow range or bright white light during the light phase. Electroretinogram damage was milder in rats kept under bright red light or attenuated yellow or brown light during the light phase. Glial fibrillary acidic protein expression in retinal Müller cells was consistent with ERG assessment of retinal damage.ConclusionsAttenuating white light and filtering out short wavelengths have a protective effect on the eyes of diabetic rats as evident by slower rate of cataract formation and a smaller degree of retinal damage.Translational RelevanceOur findings suggest that special glasses attenuating light exposure and filtering out short wavelengths (400–530 nm) may be beneficial for diabetic patients.

Enzyme-Induced Vitreolysis Can Alleviate the Progression of Diabetic Retinopathy Through the HIF-1α Pathway

We studied the mechanism by which complete posterior vitreous detachment by enzyme-induced vitreolysis alleviates the progression of diabetic retinopathy. We enrolled 50 diabetic rats and 20 normal control rats in this study. The right eyes of these diabetic rats were treated with a hyaluronidase (5 U) plus plasmin (0.25 U) by intravitreous injection (ET group), while the left eyes of diabetic rats (DR group) and eyes of the normal rats (NC group) were injected with balanced saline. Eight months after intravitreous injection, the oxygen concentration in the vitreous was measured, and the expression levels of hypoxia inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF), pigment epithelium-derived factor (PEDF), and basic fibroblast growth factor (bFGF) in retinas were determined by real-time PCR and Western blot. Clinical observation, visual electrophysiology tests, and scanning electron microscopy (SEM) also were performed on the rats. SEM results showed that the surface of retinas in the ET group had little vitreous cortex and the inner limiting membrane could be observed. The oxygen concentration of the midvitreous was higher in the ET group than other groups. A significantly high expression of HIF-1α, VEGF, and bFGF was detected in the retinas of the DR group compared to the ET and NC groups. In visual electrophysiology tests, amplitude decline and peak time latency were found in diabetic rats, and changes in the DR group were more obvious than in the ET group. Enzyme-induced vitreolysis can increase oxygen concentration in the vitreous, with reduced expression of HIF-1α, VEGF, and bFGF, and increased expression of PEDF in the retinas, thus alleviating the progression of diabetic retinopathy.

Prevention of VEGF-induced growth and tube formation in human retinal endothelial cells by aldose reductase inhibition

ObjectiveSince diabetes-induced vascular endothelial growth factor (VEGF) is implicated in retinal angiogenesis, we aimed to examine the role of aldose reductase (AR) in VEGF-induced human retinal endothelial cells (HREC) growth and tube formation. Materials and MethodsHRECs were stimulated with VEGF and cell-growth was determined by MTT assay. AR inhibitor, fidarestat, to block the enzyme activity and AR siRNA to ablate AR gene expression in HREC were used to investigate the role of AR in neovascularization using cell-migration and tube formation assays. Various signaling intermediates and angiogenesis markers were assessed by Western blot analysis. Immuno-histochemical analysis of diabetic rat eyes was performed to examine VEGF expression in the retinal layer. ResultsStimulation of primary HREC with VEGF caused increased cell growth and migration, and AR inhibition with fidarestat or ablation with siRNA significantly prevented it. VEGF-induced tube formation in HREC was also significantly prevented by fidarestat. Treatment of HREC with VEGF also increased the expression of VCAM, AR, and phosphorylation and activation of Akt and p38-MAP kinase, which were prevented by fidarestat. VEGF-induced expression of VEGFRII in HREC was also prevented by AR inhibition or ablation. ConclusionsOur results indicate that inhibition of AR in HREC prevents tube formation by inhibiting the VEGF-induced activation of the Akt and p38-MAPK pathway and suggest a mediatory role of AR in ocular neovascularization generally implicated in retinopathy and AMD.

Effects of intravitreal injection of KH902, a vascular endothelial growth factor receptor decoy, on the retinas of streptozotocin‐induced diabetic rats

KH902 is a fusion protein that can bind vascular endothelial growth factor (VEGF) and placental growth factor (PlGF) through its binding ligand taken from the domains of VEGF receptor 1 and VEGF receptor 2 (VEGFR2). This study was to investigate the effects of intravitreal injection of KH902 on the retinas of streptozotocin-induced diabetic rats. Two weeks after induction of diabetes, the left eyes of diabetic rats in each group received an intravitreal injection of phosphate-buffered saline (PBS), Avastin or KH902 solution, respectively. Four weeks after intravitreal injection, retinal electrophysiological function and the integrity of inner blood retinal barrier (iBRB) were measured by electroretinogram and Evans blue perfusion. The protein levels of VEGF signal pathway were assayed by western blot. The expression and distribution of claudin-5 and occludin were analysed by double immunofluorescent staining under confocal microscope. The expression of VEGFR2 and PlGF was measured by immunohistochemistry. Four weeks after intravitreal injection, KH902-treated rats had better retinal electrophysiological function, less retinal vessel leakage and lower levels of VEGFR2, PI3K, AKT, p-AKT, p-ERK and p-SRC than PBS or Avastin-treated rats. The distribution of claudin-5 and occludin in the retinal vessels of diabetic rats treated by KH902 was smoother and more uniform than those of diabetic rats treated by PBS or Avastin. The expression of PlGF and VEGFR2 in KH902-treated rats was decreased compared with those in PBS or Avastin-treated rats. KH902 could improve retinal electrophysiological function and inhibit the breakdown of iBRB by inhibiting the expression of VEGFR2, PlGF and PI3K, and the activation of SRC, AKT and ERK.

Sp1 and VEGF mRNA expression in the retina micrangium endothelial cells of streptozotocin-induced diabetic rats

The aim of this study is to evaluate the Sp1 and VEGF mRNA expression in the retina micrangium endothelial cells of streptozotocin-induced diabetic rats. At the end of our study, we determined that SP1 positive expression rate and the Sp1, VEGF mRNA and protein expression in the retina micrangium endothelial cells in isolated rat eyes of diabetic rats were significantly increased compared to normal animals. The ERK, Sp1 and VEGF protein expression in the retina micrangium endothelial cells of diabetic rats was significantly decreased in high-surcose (Sp1 specific inhibitor Mithramycin A or ERK specific inhibitor PD98059) culture group in comparison to normal (vechile PBS) culture group. These findings suggest that ERK, Sp1 and VEGF protein expression is an important factor to affect the occurrence and development of vision diseases in diabetic patients. Key words: Sp1, VEGF, PCR, Retina, Immunohistochemistry

Investigation of Ocular Neovascularization–Related Genes and Oxidative Stress in Diabetic Rat Eye Tissues After Resveratrol Treatment

Changes in vascular endothelial growth factor (VEGF), angiotensin-converting enzyme (ACE), matrix metalloproteinase (MMP)-9, and endothelial nitric oxide synthase (eNOS) mRNA expression profiles and oxidative stress in the eye tissue microenviroment may have important roles in ocular neovascularization and permeability in proliferative diabetic retinopathy. The present study investigated the effects of resveratrol (RSV) treatment on the mRNA expression profile of VEGF, ACE, MMP-9, and eNOS, which are associated with vascular neovascularization, and glutathione, protein carbonyl, and nitrite-nitrate levels, which are markers of oxidative stress in eyes of diabetic rats. Twenty-four Wistar albino male rats were divided into four groups. After diabetes induction with streptozotocin (10 mg/kg/day) RSV was administered to the RSV and diabetes mellitus (DM) + RSV groups for 4 weeks. The mRNA levels were measured by quantitative real-time polymerase chain reaction assay, and biochemical estimations were determined with spectrophotometric assays in eye homogenates. The mRNA expression levels of VEGF, ACE, and MMP-9 were increased in the DM group compared with the control group, and RSV treatment decreased their mRNA levels. Expression of eNOS mRNA was increased in the RSV and DM groups and decreased in the DM + RSV group. Nitrite-nitrate levels and protein carbonyl content were increased and glutathione levels were decreased in the DM group compared with controls. Consequently, these data suggest that RSV suppressed the expression of eNOS, which is actively involved in the inflammation and healing process in chronic diabetes. Although oxidative stress was increased in eye tissue from diabetic rats, mRNA levels of VEGF, MMP-9, and ACE genes associated with vascular remodeling did not change in diabetic eyes.

Placental Growth Factor Contributes to Micro-Vascular Abnormalization and Blood-Retinal Barrier Breakdown in Diabetic Retinopathy

ObjectiveThere are controversies regarding the pro-angiogenic activity of placental growth factor (PGF) in diabetic retinopathy (DR). For a better understanding of its role on the retina, we have evaluated the effect of a sustained PGF over-expression in rat ocular media, using ciliary muscle electrotransfer (ET) of a plasmid encoding rat PGF-1 (pVAX2-rPGF-1).Materials and MethodspVAX2-rPGF-1 ET in the ciliary muscle (200 V/cm) was achieved in non diabetic and diabetic rat eyes. Control eyes received saline or naked plasmid ET. Clinical follow up was carried out over three months using slit lamp examination and fluorescein angiography. After the control of rPGF-1 expression, PGF-induced effects on retinal vasculature and on the blood-external barrier were evaluated respectively by lectin and occludin staining on flat-mounts. Ocular structures were visualized through histological analysis.ResultsAfter fifteen days of rPGF-1 over-expression in normal eyes, tortuous and dilated capillaries were observed. At one month, microaneurysms and moderate vascular sprouts were detected in mid retinal periphery in vivo and on retinal flat-mounts. At later stages, retinal pigmented epithelial cells demonstrated morphological abnormalities and junction ruptures. In diabetic retinas, PGF expression rose between 2 and 5 months, and, one month after ET, rPGF-1 over-expression induced glial activation and proliferation.ConclusionThis is the first demonstration that sustained intraocular PGF production induces vascular and retinal changes similar to those observed in the early stages of diabetic retinopathy. PGF and its receptor Flt-1 may therefore be looked upon as a potential regulatory target at this stage of the disease.

Effect of an intravitreal injection of bevacizumab on the expression of VEGF and CD34 in the retina of diabetic rats

To evaluate the efficacy of intravitreal injection of bevacizumab as a preventive intervention of vascular endothelial cell proliferation in the retina of diabetic rats. Fifty-four streptozotocin-induced diabetic Wistar rats were injected intravitreally with 3 µL (25 mg/mL) of bevacizumab into left eyes and 3 µL of saline into the right eyes. Immunohistochemistry and enzyme-linked immunosorbent assays for CD34 and VEGF were used to assess retinal angiogenesis, and transmission electron microscopy was used to evaluate the ultrastructure of retinal capillaries. Retinal expression of VEGF was inhibited 1 week and 1 month after injection (P < 0.01, paired t-test), and the expression of CD34 was not obviously inhibited until 2 months after injection (P < 0.05, paired t-test). Using multiple comparisons between the left eyes of bevacizumab-treated rats, the VEGF expression before injection was higher than at 1 week or 1 month after injection (P < 0.05, Dunnett's t-test), and similar to 2 months after injection (P > 0.05, Dunnett's t-test). The amount of VEGF expression was higher 2 months after injection than 1 week or 1 month after injection, and also higher 1 week after injection compared with 1 month after injection (P < 0.05, Student-Newman-Keuls test). CD34 expression decreased more significantly 2 months after injection compared with before injection, 1 week or 1 month after injection (P < 0.05, Dunnett's t-test). A single intraocular injection of bevacizumab may be beneficial as a therapy for preventing retinal vascular endothelial cell growth in the eyes of diabetic rats.

Topical instillation of ciliary neurotrophic factor inhibits retinal degeneration in streptozotocin-induced diabetic rats.

Intraocular injections of ciliary neurotrophic factor (CNTF) or intraocular adenovirus-mediated CNTF gene transfer have been reported to inhibit retinal alterations in inherited retinal degeneration in mice strains. To investigate whether or not CNTF administered by eye drops prevents retinal degeneration in streptozotocin (STZ)-induced diabetic rats, recombinant CNTF was administered to eyes of diabetic rats (n=20) twice daily for 1 month after the onset of diabetes. The b-wave amplitude of electroretinogram in CNTF-administered diabetic rats was significantly larger than that of diabetic rats, and approached that of the controls. Atrophy of the inner plexiform layer, and cavity formation in the pigment epithelium, which were observed in diabetic rats, were prevented in CNTF-administered diabetic rats. These results indicate that CNTF administration by eye drops prevents retinal degeneration in STZ-induced diabetic rats.

Thrombospondin-1, a Natural Inhibitor of Angiogenesis, Is Present in Vitreous and Aqueous Humor and Is Modulated by Hyperglycemia

Negative regulators of angiogenesis play a major role in maintaining vascular homeostasis. Thrombospondin-1 (TSP1) is a natural inhibitor of angiogenesis. This report examines the presence of TSP1 in ocular samples and determines whether its production is altered in diabetes. Western blot analysis detected a 140 kDa antiangiogenic fragment of TSP1(gp140) in vitreous samples prepared from normal human and rat eyes. Intact TSP1 was detected in aqueous humor samples prepared from normal rat and bovine eyes. In contrast, TSP1 was virtually absent in vitreous and aqueous humor samples prepared from diabetic rat eyes. Furthermore, production of TSP1 by microvascular endothelial cells in culture was sensitive to high concentrations of glucose. Retinal blood vessels appeared nonuniform and dilated in diabetic animals when compared to control animals. These results demonstrate that TSP1 and its antiangiogenic fragment are present in aqueous humor and vitreous of normal rat eyes and are dramatically reduced in diabetes. Thus, TSP1 may play a role in ocular vascular homeostasis and its absence may contribute to vascular dysfunctions associated with diabetes.

Immunological detection of fructated proteins in vitro and in vivo.

An antibody has been raised against fructated lysine in proteins by immunizing fructated lysine-conjugated ovalbumin in rabbits. The affinity-purified antibody specifically recognized proteins incubated with fructose but not with other reducing sugars such as glucose, galactose or ribose, as judged by immunoblotting and ELISA techniques. Competitive binding to this antibody was observed specifically by fructated lysine but not by glucated lysine, glucose, fructose or lysine. The antibody binds specifically to fructated lysine residues in the protein but not to borohydride-reduced material or advanced glycation end products, indicating that the antibody recognizes only the reducing, carbonyl-containing forms produced in the early stage of the fructation reaction. When BSA was incubated with various concentrations of fructose, the reactivity of the antibody increased in a dose- and time-dependent manner. When soluble proteins prepared from either normal or streptozotocin-induced diabetic rat eyes were analysed by ELISA with this antibody, an increase in the reactive components was observed as a function of aging as well as under diabetic conditions. Western blotting analysis showed that lens crystallin reacted highly with this antibody. Because fructose is biosynthesized largely through the polyol pathway, which is enhanced under diabetic conditions, and lens is known to have a high activity of enzymes in this pathway, this antibody is capable of recognizing fructated proteins in vivo. Thus it is a potentially useful tool for investigating two major issues that seem to be involved in diabetic complications, namely the glycation reaction and the polyol pathway.

Altered vasoactivity in the early diabetic eye: Measured in the isolated perfused rat eye

The effect of 4 weeks streptozotocin-induced diabetes on ocular vascular resistance responses to noradrenalin (NA), adrenalin (A), phenylephrine (PHE), isoproterenol (ISOP), prostaglandin F2 alpha (PGF2 alpha). 5-hydroxytryptamine (5-HT) and angiotensin II (ANG II), was determined using a newly-developed, isolated, arterially-perfused rat eye preparation, by comparing responses from control and diabetic eyes. After extensive preliminary experiments to establish optimum parameters, the ophthalmic artery of enucleated control and diabetic rat eyes was cannulated and the retinal and uveal vasculature perfused at a constant flow with Na(+)-Krebs solution after streptozotocin-induced diabetes had been established for 4 weeks. The eyes were maintained in an environment-controlled organ bath. Perfusion pressure was monitored as increasing log M concentrations of agonists were added to the perfusate. Total ocular resistance could be calculated from knowledge of flow and pressure. In control eyes, NA, A, PHE, PGF2 alpha, and 5-HT all produced dose-dependent increases in total vascular resistance, with the following order of potency: NA = A > 5-HT > PHE = PGF2 alpha at 10(-4) M. The ocular circulation was not sensitive to isoproterenol and angiotensin II. In diabetic eyes responses to NA, A, PGF2 alpha and 5-HT were altered. Diabetic responses to NA and A had lower thresholds with larger resistance increases at low concentrations. However, the rate of increase in resistance with concentration was more gradual in diabetic eyes so that at 10(-4) M control responses were larger. Diabetic resistance responses to PGF2 alpha had the same threshold as in control eyes, but were greater in magnitude with an earlier peak at 10(-4) M. In contrast diabetic resistance responses to 5-HT were reduced, peaked at a lower resistance at 10(-4) M, but had the same threshold as those in the control eye. Basal vascular resistances in control: 3.14 +/- 0.32 mmHg min microliter-1 (n = 28), and diabetic eyes: 3.44 +/- 0.19, mmHg min microliter-1 (n = 36), were not significantly different. Vasoactivity in the early diabetic eye is disturbed with the effective balance between different agonists altered in favour of catecholamines at physiological concentrations. This may be related to the early changes in blood flow and oxygen distribution already reported in the rat eye, as well as changes to autonomic function. The isolated perfused rat eye is a valuable technique for investigating such vascular reactivity in animal models of retinal disease.

Regulation of growth factor mRNA levels in the eyes of diabetic rats

The underlying etiology of diabetic microvascular disease remains unknown. To examine the potential contribution of basic fibroblact growth factor (bFGF), which is an angiogenic factor, and insulin-like growth factor-I (IGF-I) to the development of diabetic microvascular disease, bFGF and IGF-I mRNA levels were measured in tissues of control, diabetic, and insulin-treated diabetic rats. Diabetes was induced in rats by intravenous injection of streptozotocin (STZ) 65 mg/kg, and the rats were maintained for 21 days. bFGF mRNA levels increased threefold in the eyes of diabetic versus control rats, whereas a consistent change in bFGF mRNA levels was not observed in other tissues. In contrast, IGF-I mRNA levels decreased in the eyes and other tissues, including kidney, lung, and skeletal muscle, of diabetic as compared with control rats. Insulin treatment prevented the diabetes-induced increase in bFGF and decrease in IGF-I mRNA levels. Acidic FGF (aFGF) mRNA levels were unchanged in eyes from diabetic versus control rats. In partially purified retinas, diabetes increased bFGF mRNA levels twofold as compared with levels in control retinas, whereas IGF-I mRNA levels decreased to 58% of control levels in retinas from diabetic rats. Insulin treatment again prevented the diabetes-induced increase in IGF-I mRNA levels in the retina but had no effect on the diabetes-induced increase in bFGF mRNA levels, bFGF peptide levels were minimally increased in diabetic versus control retinas. Treatment of diabetic rats with the aldose reductase inhibitor sorbinil prevented the diabetes-induced increase in sorbitol accumulation and myo-inositol depletion in the lens, but it did not affect the diabetes-induced increase in bFGF and decrease in IGF-I mFINA levels in the retina. Induction of hypoinsulinemia by fasting the animals did decrease IGF-I mRNA levels but did not reproduce the diabetes-induced increase in bFGF mRNA levels in the eye. In conclusion, these data demonstrate that the effect of diabetes on growth factor mRNA levels in the eye is gene-specific, suggest that different metabolic abnormalities are responsible for diabetes-induced alterations in the production of different growth factors in the eye, and are consistent with a role for bFGF in the development of diabetic retinopathy.

Corneal epithelial changes in diabetic rats.

Electron microscopy of streptozotocin diabetic rat eyes showed increased intracellular levels of tonofilaments and glycogen, thickenings and infoldings of subepithelial basement membrane and basal cell degeneration after 8 months. Glycogen, glucose, sorbitol, and fructose were measured in corneal epithelium from short- and long-term diabetic rats. The small increase in sorbitol pathway products which were found after 8 months of diabetes (less than 1.0 mosm/1 tissue water) confirmed similar findings in rabbits and humans. Thus, the morphologic changes occur in the absence of significant accumulation of sorbitol pathway products. Osmotic damage secondary to corneal epithelial cell accumulation of sorbitol is probably not a significant factor in corneal epithelial diabetic disease.

Vascular changes in eyes of diabetic rats; alterations in the anterior segment.

In 1952 Janes, Bounds, and Leinfelder 1,2 reported on cataract formation in alloxandiabetic rats. At that time these authors observed intraocular hemorrhages in the form of hyphemas (with subsequent corneal vascularization) to have occurred in some of their animals. These studies have been continued, and to date 30 rats have been noted to have developed hyphemas with subsequent corneal changes. The animals have been killed at various times after the development of these complications. Studies of the ocular vascular system have been made following intravascular injections of neoprene latex * or India ink. This paper describes the gross and histological changes observed. Technique Long-Evans strain adult male rats were fasted for 48 hours and then given a subcutaneous injection of 115 mg. of alloxan per kilogram of body weight. As previously, these rats were followed with regard to blood sugar levels, urine volumes, food intake, and body weight. Feedings consisted of