Effect of β-Casomorphin-5 on Blood Glucose and Oxidative Stress of Diabetic Rats
|School||Nanjing Agricultural College|
|Course||Basic Veterinary Science|
|Keywords||β-CM5 blood glucose diabetes mellitus glucose transport oxidative stress C2C12|
We will explore the effect ofβ-casomorphin-5 on blood glucose and oxidative strss of diabetic rats. Studies include the following three parts:Experiment 1. Inhibition ofβ-casomorphin-5 on the absorption of glucose in intestine in vitro and its mechanismWe will explore the influence of theβ-CM5 on the absorption of glucose and its mechanism in intestinal by using of reverted sacs of adult rats in vitro. We will test glucose concentration and the transport velocity of glucose different concentrations of P-CM5 group within 60 minutes;In addition,we will examine the activity of a-glucosidase enzyme and Na+-K+-ATPase; Last,we will analyze the mRNA expression of SGLT-1 and GLUT-2.The results showed that(1)glucose contents in intestinal capsule volume and transfer rate of (3-CM5 groups were significantly lower than that of the control group; (2)the activity of a-glucosidase ofβ-CM5 group was significantly lower than the control group;(3) Compared with the control group, small intestinal glucose transporter protein, SGLT-1, GLUT-2 mRNA expression of (3-CM5 group were significantly reduced, and ignificantly inhibited the activity of Na+-K+-ATPase.Conclusion:β-CM5 can inhibite the activity of enzyme related to glucose uptake and reduced the mRNA expression of glucose transporter, so P-CM5 can inhibite intestinal absorption of glucose,β-CM5 may have a hypoglycemic effect.Experiment 2. Effect ofβ-CM5 on blood glucose and the capacity of antioxidant in diabetic ratsTo explore the effect ofβ-CM5 on streptozotocin (STZ)-induced diabetes mellitus (DM) rats blood glucose and the capacity of anti-oxidative.60 male SD rats were selected randomly, and 50 rats of them were used for the experimental model of DM rats, and the other 10 rats as normal control group. The rats which used for making DM model were injected STZ solution according to the dose of 60mg/kg(rats were fasted overnight before administration).Normal rats in control group were injected intraperitoneally in the same way with an equal volume of normal saline (NS).27 hours later, blood glucose was measured for all rats (fasting 8 hour), blood glucose levels of DM rats had to more than 11.1 mmol/L. Modeling 40 rats from the DM rats were chosen and were divided into high dose(H) group (7.5×10-5mol/Lβ-CM5), middle dose(M) group(7.5×10-6mol/L (3-CM5) and low dose(L) group(7.5×10-7mol/Lβ-CM5). Rats ofβ-CM5 groups were given with 1mL ofβ-CM5 solution every 12 hours, the rats of DM group and normal control group were given saline with the same volume. Drinking, feeding and so on were observed every day.The blood glucose of all rats were measured every five days.25 days later, all rats were killed,their serum、liver and muscle were taken for the following test:1)blood glucose levels will be tested;2)we will test the glycogen of liver and muscle,insulin and glucagon in blood serum.3)we will test the content of MDA and the activity of SOD AND GSH-Px in blood serum.the result shoued that:l) blood glucose levels were decreased in rats which fed withβ-CM5. blood glucose levels were significantly lower than DM rats after given 20 days; 2) compared with DM group, the contents of liver glycogen and muscle glycogen of different dose P-CM5 groups were increased; levels of insulin and glucagon were no significant difference; 3)β-CM5 can significantly increase the activity of SOD, it also can lower the content of MDA. Conclusion:β-CM5 can reduce the level of blood glucose in diabetic rats and increase the content of liver glycogen, muscle glycogen. In addition, It also can enhance the body’s ability to anti-oxidative stress.Experiment 3. Cytological evidence ofβ-CM5 on anti-oxidative damageC2C12 cells are the most common cell model which is used for searching oxidative stress damage, our results which in the previous chapter indicate thatβ-CM5 can increase the capacity of anti-oxidative damage in DM rats. In this experiment we intend to explore the mechanism ofβ-CM5’s anti-oxidative damage on the cellular level, and explore the protective effect ofβ-CM5 and its possible mechanism by the establishment of hydrogen peroxide (H2O2)oxidative damage model. Method:1) We will research the effect ofβ-CM5 on the activity of cells by MTT within 72h and select the safe dose ofβ-CM5 acting on muscle cells; 2)We will test the effect that different concentrations of H2O2 on the activity of cells by MTT.So that we can select the optimum dose of H2O2; 3)We will test the effect ofβ-CM5 on the viability of cells after injuried by MTT; 4)We will test the activity of SOD and LDH in medium. The results showed that:1)Compared with control group, the values of MTT in various concentrations ofβ-CM5 have no difference; 2)0.25 mmol/L H2O2 acting on C2C12 one hour later, MTT values were significantly lower than the control group(0.31±0.01, P<0.01); 3)MTT values in pre-protection ofβ-CM5 group were more significantly higher than the injury group (0.53±0.01,P<0.05),MTT value in the treatment of (3-CM5 group was no significant difference between the injury groups (0.71±0.01,P<0.05); 4)compared with the injury group, the activity of SOD in (3-CM5 Protection Unit was significantly increased(10.18±0.35,P<0.05),the activity of LDH was significantly decreased(397.34±15.96,P<0.05).Conclusion,β-CM5 have the pre-protective effects on H2O2 injury muscle cells, the mechanism may be related to increased activity of antioxidant enzymes to maintain the integrity of the cell,howerer,the mechanism was needed to be studied further.