Inhibition Of Cataract In Diabetic Rats By Carnosine Eyedrop

Objective:To establish an animal model for suitable diabetic cataract study with long-term, stable hyperglycemia.To investigate the progression of lens opacification and some of biological chemistry changes during diabetic cataract formation.To investigate the effect of carnosine, inhibitors of advanced glycation, on the development of diabetic cataract.Methods:1. Fifty male Sprague-Dawley rats weighing 120±20 g received a single intraperitoneal injection of streptozotocin (STZ, 65 mg/kg) dissolved in 0.02 mol/L sodium citrate buffer. In addition, 5 healthy rats of the same age were taken out randomly as non-diabetic animal group (NonDb), received 0.02 mol/L citrate buffer (pH 4.5) alone. Animals with blood glucose levels greater than that of 14 mmol/L on 72 h after STZ injection were treated as diabetic rats.2. Rats were randomly divided into lower concentration of carnosine treated group, higher concentration of carnosine treated group and diabetic group. Received eye drops consisting of a freshly 5 g/L, 10 g/L solution of carnosine (25 mmol/L phosphate buffered solution, pH 7.4) or 25 mmol/L phosphate buffered solution (pH 7.4) administered as twice daily ,respectively. Every 4 weeks during the experiment, the blood glucose was reconfirmed and rats with the blood glucose levels higher than 14 mmol/L all the time were selected for further study. Those rats with blood glucose levels dropped to less 14 mmol/L or died before the end of study were excluded. The body weight of rats was measured weekly.3. After dilation of the pupils with a drop of 1% tropicamide, the rats were anaesthetized by ether inhalation. Progression of cataract formation in both eye lenses from all rats was monitored once a week by slit lamp (Haag-Streit BQ 900). The grading of lens opacification was performed according to the Oxford University system.4. At the 13 weeks after the injection, the rats were sacrificed and the lenses were dissected out, then placed into pre-weighed Eppendorf tubes and frozen at -70℃until analyzed. The levels of glutathione and advanced glycation end products, and the activites of glutathione reductase and catalase were measured in both of the left and right lenses.Results:1. Blood glucose was significantly greater in all of the diabetic rats 72 h after injection of STZ (33.64±1.76 mmol/L, P<0.01) compared with the normal group (approx. 6.8±0.62 mmol/L). Throughout the experiment peroid, elevated blood glucose and decreased body weight were observed in Db group. However, both carnosine eye-drops failed to prevent these changes.2. The opacity of lenses was not observed until the 4th week after STZ injection. Then the lenses opacification progressed in a biphasic manner in the diabetic rats, an initial slow increase during the first 4 to 8 weeks of diabetes followed by a steep increase in next 5 weeks. Lens opacities were consistently greater in the diabetic rats compared to the normal rats, in fact, the lenses were retained transparency in the control rats throughout experiment peroid. Carnosine eye-drops treatment delayed the progression of cataract formation in diabetic rats and the delay was statistically significant on the 4th week of diabetes (p<0.05, when compared with untreated moderately diabetic rats).3. Compared with the normal rats at 13 weeks after injection, the activities of catalase and glutathione reductase were decreased by 52% (31.3μmol/g lens protein, P<0.05) and 51% (5.7μmol/g lens protein, P<0.05) in the lens of untreated diabetic rats, respectively. The level of glutathione was decreased by 56% (57.5 mg/g lens protein), while the level of advanced glycation end products was increased by 29% (1.2 mmoles HMF/mole lens protein, P<0.05). However, the changes of GR, CAT, GSH, AGEs were delayed in carnosine (5mg/ml and 10mg/ml) treated diabetic rats. Compared with the diabetic rats, the activities of catalase and glutathione reductase were increased by 30% (18.0μmol/g lens protein, P<0.05) and 19% (2.1μmol/g lens protein, P<0.05) in the lens of the lower concentration of carnosine (5mg/ml) treated rats at 13 weeks after injection, respectively. The content of glutathione was increased by 21% (21.1 mg/g lens protein, P<0.05), the content of advanced glycation end products was decreased 34% (1.4 mmoles HMF/mole lens protein, P<0.05). Moreover, the activities of catalase and glutathione reductase were increased 37% (22.4μmol/g lens protein, P<0.05) and 44% (4.9μmol/g lens protein, P<0.05) in the lens of the higher concentration of carnosine (10mg/ml) treated rats at 13 weeks after injection, respectively. The content of glutathione was increased by 31% (31.1 mg/g lens protein, P<0.05), the content of advanced glycation end products was decreased 48% (2.0 mmoles HMF/mole lens protein, P<0.05). It indicated that carnosine can prevent the inactivation of catalase and glutathione reductase, and the decrease of GSH and the increase of AGEs induced by diabetes.Conclusions:A single intraperitoneal injection of streptozotocin (65 mg/kg) can induce overt diabetes with a long-term, stable hyperglycemia in adult rats. It can induce diabetic cataract.Carnosine eye-drop can delay the progression of lens opacification in the diabetic rats during the earlier stages. This delay may associate with the protective effect of carnosine against inactivation of antioxidation enzymes and the formation of advanced glycation end products.