The Effect Of Aminoguanidine On Diabetic Rat's Surface Changes

Objective:Diabetic mellitus is a common general disease mainly manifestating by glycometabolism disorders whose prevalence is rising year by year. It can cause many complications in eyes. The recent clinical data indicates that the symptom of Dryness of eye, foreign body sensation and burning sensation appeare widespread in DM patients. The experimental model of diabetic rats are induced by intravenous administration of streptozotocin, then we observe the change of ocular surface by Schirmer-Ⅰtest and tear film break up time(BUT) measurement; The histopathological changes and the expressions of tumor necrosis factor alpha(TNF-α), nuclear factor-KBp65(NF-KBp65), receptor for advanced glycation end products(RAGE)of the cornea, conjunctiva and lacrimal gland are observed in the diabetic rats, then we conform diabetic mellitus, the effect of aminoguanidine on it, in order to provide pathological and theoretical basis for treating diabetic ocular surface diseases.Methods:1 Method of grouping30 male Wistar rats that weighted 180-200g,were randomly divided into 3 groups: normal control group (normal group) , diabetic control group (diabetic group) and aminoguanidine treated group (treated group).2 Preparation of diabetic animal modelAfter fasting 12h, three groups'rats were sterilized the site of intraperitoneal injection with Iodophors.Then rats of diabetic and treated group received a single 65mg/kg intraperitonea injection of 10g/L streptozotocin(STZ) dissolved in 20mmol/L sodium citrate buffer (pH 4.5). Detect blood glucose by 72h and per 4 weeks of experiment after STZ injection, then animals that blood levels more than 16.7mmol/L all the time were selected for study. In addition, 10 normal group rats received 20mmol/L sodium citrate buffer (pH 4.5).3 Ways of administration72h after STZ injection, three groups'rats were sterilized the site of intraperitoneal injection with Iodophors. Then treated group received injection of a freshly 80mg/kg.d solution of 0.2% aminoguanidine (25mmol/L phosphate buffered solution, pH7.4) once a day. In addition, diabetic and normal groups received 25mmol/L phosphate buffered solution (pH7.4) alone.4 Indexes to be observedThe volume of drinking, appetite and the urinary volume were observed every day. Body weights of rats were measured weekly. Every 4 weeks during the experiment and 72h after STZ injection, blood glucose of rats were detected. All rats took the Schirmer-Ⅰtest and BUT on the 12th week. Then 10 rats of each group were sacrificed, cornea, conjunctiva and lacrimal gland were obtained and fixed by 10g/L formalin for histopathology and immunohistochemistry.Results:1 Body weight of normal group rats increased gradually. 2-12wk after injection of STZ, the weight was significantly decreased in diabetic and treated groups compared with normal group (p<0.05). Through all facets of the study, the difference of weight beween diabetic and treated groups rats were non-significant(P>0.05) (Fig 1).2 Blood glucose was significantly higher in diabetic and treated groups 72h after injection of STZ,compared with control group (p<0.01); Through all facets of the study, the difference of blood glucose between diabetic and treated groups rats were non-significant(P>0.05)(Fig 2).3 The test values of BUT and Schirmer-Ⅰ: compared with normal group,they were significantly lower in diabetic group(p<0.01),still significantly lower in treated group(p< 0.05); compared with diabetic group, they were significantly greater in treated group(p<0.01) (Table 1).4 Histopathological changes of lacrimal gland, conjunctiva and cornea: Lacrimal glandular follieles of diabetic group shrinked, caryon were pyknosised and anachromasised. Fibers were prolifered, tissues were confused.Goblet cell lost and the basal epithelial cells and the stroma of diabetic groups'cornea were swollen. All of the changes were significantly lightener in treated group than in diabetic group, but still differed from normal group (Fig 3-11).5 Expressions of TNF-α, NF-KBp65 and RAGE in lacrimal gland,conjunctiva and cornea: TNF-α, NF-KBp65 and RAGE are all expressed in the kytoplasm of lacrimal gland,conjunctiva and cornea. Compared with normal group, expressions of TNF-α,NF-KBp65 and RAGE were significantly greater in diabetic and treated groups (p<0.01). But compared with diabetic group, they were significantly lower(p<0.01)(Fig 12-20, Table 2-4). In addition, there were positive correlations in TNF-αand NF-KBp65, TNF-αand RAGE, NF-KBp65 and RAGE (Table 5).Conclusions:1 Diabetic mellitus is related with dry eye.2 Diabetic mellitus ocular disease is concerned with protein glycosylation.3 The effect of AG is significant in treating diabetic mellitus ocular diseases, and its mechanism of action is concerned with protein glycosylation and the decrease of TNF-α, NF-KBp65 and RAGE. In addition, it has no influence on body weight and blood glucose.