| Objectives and significanceWith the improvement of living standards, the incidence of diabetes is increasing year by year, has been the development of chronic non-communicable diseases ranked third following cancer, cardiovascular and cerebrovascular diseases, and still can produce many complications, not only to the human body and mind health harm, but also cause serious economic burden, diabetes appear younger, hidden trend, diabetes pandemic is alsocontinue to develop.Type II diabetes (T2DM) accounts for more than90%of the total diabetic population in recent years that insulin resistance (insulin resistance, IR) is one of the important mechanisms of T2DM onset, IR refers to the biological effects of insulin to reduce blood insulinpromote glucose uptake impaired, resulting in compensatory increase in insulin secretion, the specific performance of the peripheral tissue insulin sensitivity, as well as barriers to the utilization of glucose.IR Strongly indicates the possibility of Type II diabetes, and IR will run through type II diabetes occurred in the whole development process. IR has been more attention, so far, however, there is no ideal for IR in Western medicine, For the treatment of IR use more insulin sensitizers, such as the TZD as pioglitazone, biguanide as metformin; or use can glucose-lowering, improve IR and metabolic disorder drugs; or insulin degradation inhibitors, etc. Although there are better clinical curative effect, but the western medicine in the treatment of side effects is larger.Traditional Chinese medicine to pay attention to the syndrome differentiation and treatment, through the whole adjustment to improve the pathological state. Although in oral glucose-lowering strength and work time less than western medicine, but its have side effect is small, can long-term aspirin and other characteristics, and its function is often many link, the adjustment of the target, Chinese traditional medicine in promoting effect of insulin secretion, improve insulin use, increases insulin sensitivity, and to prevent complications in on certain advantages of Chinese medicine of diabetes drug screening became the new research hot spot.T2DM belong to "thirst disease" category in traditional chinese medicine, traditional chinese medicine has accumulated rich experience in the treatment in the long-term clinical practice. Mei Lian Xiao Ke capsules is commonly used formula of the civil treatment of T2DM, the composition of the berberine, ebony, aconite, ginger, Asarum, Shujiao, cassia twig, Cork. Coptis and ebony is king medicine in the formula. Coptis nature is bitter and cold, the main ingredient is berberine, it has to improve the role of IR, hypoglycemic and correct lipid metabolism disorders, but also reduce the activity of key enzymes of hepatic gluconeogenesis.Commonly used side of Mei Lian Xiao Ke capsules civil treatment of type II diabetes, the clinical effect is significant. In the early pre-experiment can permit Mingmei with diabetes capsules good hypoglycemic improve the role of the clinical symptoms of patients with diabetes detected Mei Lian Xiao Ke capsules influential on the content of insulin in diabetic rat serum, which speculate that its also improve the efficacy of the IR. The purpose of this study Mei Lian Xiao Ke capsules is to investigate whether the regulation of insulin secretion, improve insulin resistance, and the establishment of three kinds of diabetes in rats, mice, invitro model to study to the one for the regulation of insulin secretion to improve the IR role of diabetes drugs. Experiment1. Effect of Mei lian xiao ke capsule on insulin on diabetic ratsMethods:Male SD rats were divided into control group(10) and model group(110), blank control group given ordinary feed-made modules to give high-fat high-sugar diet, feeding five weeks, intraperitoneal injection (ip),dose streptozotocin (STZ,30mg·kg-1) model after do not give food but water for12h, and normal control group was injected with same volume of saline, select72h fasting blood glucose between7.0to25.0mmol/L animals for the modeling as experimental animals. To drug by Irrigation stomach For4weeks, the normal control group and diabetic model group was given same volume of saline. Observed A general status of animals during the administration. Determine fasting blood glucose (FBG) by blood glucose meter with cut tail method at the end of the experiment; abdominal aorta after the end of the experiment, blood serum copper ion colorimetric determination of serum free fatty acid (FFA) content; by radioimmunoassay of serum insulin (FINS) levels, insulin sensitivity index (ISI), insulin resistance index (HMOA-IR); liver tissue for biopsy, HE staining, and evaluation of the degree of pathological damage of the islet tissue; each group to select the six cases of rats pancreatic tissue sections using the aldehyde fuchsin staining of pancreatic islet cell alpha, beta cells were stained to use IPP6.0software to calculate the proportion of β-cells in the islet tissue.Results:Gompared with the normal control group, model control group rat body weight, food intake, water quantity, urine increase, FBG were significantly increased(P=0.000, P<0.01), free FFA were significantly increased(P=0.000, P<0.01), FINS were significantly increased (P=0.000, P<0.01), ISI were significantly reduced (P=0.000, P<0.01), HOMA-IR were significantly increased (P=0.000, P<0.01), islet tissue pathology damaged significantly (P=0.000, P<0.01), β cells significantly lower proportion (P=0.000, P<0.01.After drugs for four weeks, compare with Treatment group and model rats diabetes, Mei Lian Xiao Ke capsules high(P=0.001, P<0.01), medium(P=0.024, P<0.05) dose group could significantly reduce diabetic rats FBG; Mei Lian Xiao Ke capsules high(P=0.001, P<0.01), middle(P=0.045, P<0.05) and low(P=0.011, P<0.05) dose group can significantly reduce the FFA; May even high dose group of quench capsule FINS significantly (P=0.000, P<0.01), homa-ir significantly reduced (P=0.003, P<0.01) and the remarkable increase of ISI (P=0.000, P<0.01); Dose group of significant fall in FINS (P=0.000, P<0.01), a significant increase in ISI (P=0.033, P<0.05), homa-ir significantly reduced (P=0.029, P<0.05); Low dose group of FINS significantly (P=0.005, P<0.01); Mei Lian Xiao Ke capsules high dose group (P=0.007, P<0.0125) were significantly improve islet tissue pathology damage; Mei Lian Xiao Ke capsules high dose group make (3cells ratio increased significantly (P=0.000, P<0.01), medium, low dose had increased beta cells of the proportion of the trend, but was not statistically significant. Metformin can significantly reduce diabetic rats FBG (P=0.000, P<0.01), and improve tissue pathology (P=0.003, P<0.0125), but FFA, FINS, β cells has no significant proportion improved.Conclusion:Mei Lian Xiao Ke capsules improve Feed more, drink more, urine more and weight loss clinical symptoms improved in experimental type II diabetic rats, Significantly improve significantly reduce rat model of FBG and FINS, apparent protection organization and repair islet beta cell function, improvement IR state the significantly of the model rats.Experiment2. Effect of Mei lian xiao ke capsule on FINS of diabetic mice induced by ALXMethods:Male KM mice, adaptive feeding3d, were divided into control group and model group. Modeling the model group of mice tail vein (iv) injection of alloxan55mg· kg-1after do not give food but water for12h,normal control group was injected with the same volume of saline. Fasting glucose levels after72h, select the model was successful in10-23mmol/L mice as experimental animals, To drug by Irrigation stomach For4weeks, the normal control group and diabetic model group was given same volume of saline. Observed A general status of animals during the administration. During the experiment, every2weeks docked take blood, blood sugar analyzer to measure FBG during the Experiment2,4weeks; At the end of the experiment, anesthesia to pick the eyeball take blood, using radioimmunoassay determination FINS level.Methods:Compare with the normal control group, diabetes model of mice into appetite, water quantity, urine increase, FFB level increased significantly (P=0.000, P<0.01), and has remained at a stable level, FINS were significantly reduced (P=0.043, P<0.05). The drug after around the treatment group and diabetes model is of mice, the treatment group all can improve general symptoms of diabetic mice, Mei Lian Xiao Ke capsules group FFB reduce thirst, but due to excessive standard deviation was not statistically significant; Xiao Ke Pills group mice reduce the FFA, but this was not statistically significant; Metformin group of mice in two weeks FFB reduce has statistical significance. Mei Lian Xiao Ke capsule group a greater increases level in FINS, but due to excessive standard deviation was not statistically significant; Xiao Ke Pills group can increases FINS level rise, but this was not statistically significant.Conclusion:Mei Lian Xiao Ke capsules improve Feed more, drink more, urine more and weight loss clinical symptoms improved in alloxan mouse, can raise production of insulin of diabetic mice induced by ALX, improve the model of insulin sensitivity in model mice.Experiment3. Effect of Mei lian xiao ke capsule on HepG2-IR cell glucose ConsumptionMethods:HepG2cells train with DMEM in37℃and5%CO2in the environment, passage as1:3. Take the cell growth index, adjust the cell concentration spread96orifice plate, divided into normal control group and made module, made module contains high levels of insulin cells in the medium induced24h, according to the glucose determination in full automatic kit on biochemical analyzer test cell culture of glucose in consumption, when made module cells consumption significantly lower than normal glucose control group and cells when the number is not significant difference, think HepG2-IR model establishment, and investigation HepG2-in48h IR model in stability.The MTT experimental detection cells to determine the survival rate of berberine, and Mei Lian Xiao Ke capsules of cells to quench liquids drug concentration, HepG2cells as a normal control group, HepG2-IR cells as a model control group to join the normal culture medium, in treatment group HepG2-IR cells containing Mei Lian Xiao Ke capsules solution and Mei Lian Xiao Ke capsules drug serum capsule culture medium,24h after for growing the role of serum does not contain the basis of24h culture medium training, according to glucose determination in full automatic kit on biochemical analyzer test cells in the medium of glucose intake ability, evaluation of the HepG2-IR model the influence of glucose consumption.Methods:Compare with the normal control group HepG2cells, HepG2-IR cells have not significantly different in form, HepG2-IR cells of glucose consumption significantly reduced, HepG2-IR cell model can stable in48h. Mei Lian Xiao Ke capsules solution high (P=0.005, P<0.01), medium group (P=0.025, P<0.05) can significantly increase the dose group of HepG2-IR cells the consumption of glucose; Mei Lian Xiao Ke capsules concentration15%serum capsule (P=0.003, P<0.01),10%concentration (P=0.033, P<0.05) can significantly increase the HepG2-IR cells the consumption of glucose; Berberine can significant increase he consumption of glucose in HepG2-IR cells.Conclusion:Mei Lian Xiao Ke capsules solution and Mei Lian Xiao Ke capsules serum can significantly increase HepG2-IR cells of glucose consumption, improve IR state of the HepG2-IR cells. |
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