Studies On The Preparation And Anti-type 2 Diabetes Mechanisms Of Marine Oligosaccharide Derivatives

Diabetes mellitus is the most common metabolic disease and its prevalence is increasing in both developed and developing countries year by year. China has become the country with the largest number of diabetes prevalence all over the world. Diabetes mellitus can mainly be divided into type 1 diabetes (insulin dependent, IDDM) and type 2 diabetes (non insulin dependent, NIDDM). More than 90% of diabetes patients suffer from non-insulin-dependent diabetes mellitus (NIDDM, type 2 diabetes) which is a typical polygenic disease. Type 2 diabetes is mainly associated with two principal physiological defects:resistance to the action of insulin and deficiency in insulin secretion. Insulin resistance mainly occurs in fat tissue, liver and skeletal muscles, where cells contain many insulin receptors that can bind insulin to regulate the steady-state of glucose metabolism. Resistance to the actions of insulin in skeletal muscle is a major pathogenic factor in diabetes mellitus. So the pharmaceutical researches on the prevention of metabolic disorder caused by insulin resistance and improvement of insulin sensitivity are very important for the development of anti-type 2 diabetes drugs. However, current drugs for type 2 diabetes therapies often have many defects such as secondary failure, side effects and risk of hypoglycemia, so it is important to develop low toxic and long acting anti-diabetes drugs which can improve insulin resistance. The unique environment of ocean creates many active substances with special structures and functions, especially for the marine polysaccharides which attract much attention for their wide variety of sources, low toxicity and broad activity. Many researches indicated that different kinds of organic chromium complexes can improve glucose metabolism and regulate blood lipid. Based on the former research, series of marine oligosaccharide-chromium derivatives were prepared by complexing oligosaccharides with chromium ion in this research, and one of them named OM2 was found to be able to effectively increase the insulin sensitivity, and its molecular mechanisms of anti-type 2 diabetes were also investigated.Alginate oligosaccharides and carrageenan oligosaccharides with different degrees of polymerization were firstly prepared by using acid degradation of alginate and carrageenan, and then series of marine oligosaccharide-chromium derivatives were prepared by complexing oligosaccharides with chromium (â…¢) ion. Moreover, the orthogonal experiments were used to optimize the preparation conditions of oligomannuronate-chromium complexes and carrageenan oligosaccharide-chromium complexes, and the optimal reaction conditions were determined in this research. The structure characterization of prepared marine oligosaccharide-chromium derivatives was performed by using full wavelength UV scanning and IR analysisSkeletal muscle C2C12 cell model and Thioflavin T fluorescence assay were then used to perform activity screening of different kinds of marine oligosaccharide derivatives. The results indicated that the marine acidic oligosaccharide-chromium complex OM2 could not only inhibit amylin fibrosis in vitro but also activate AMPK signal pathway, which suggested OM2 has potential activity to improve insulin resistance. Based on these results, the hereditary diabetes transgenic db/db mice were used to evaluate the anti-diabetes effect of marine acidic oligosaccharide-chromium complex OM2 in vivo. The results showed that OM2 has no acute hypoglycemic effect and no risk of transient lower blood sugar. OM2 could not only effectively decrease the blood sugar; improve blood lipid metabolism and attenuate insulin resistance in db/db mice, but also have the activity to increase insulin sensitivity to some extent in vivo.Based on the results that OM2 has good anti-type 2 diabetes effects, the mechanisms of improving insulin sensitivity by marine acidic oligosaccharide OM2 were then systematically investigated in vitro and in vivo by using skeletal muscle C2C12 cells and db/db mice. The results indicated that the marine acidic oligosaccharide-chromium complex OM2 could significantly increase insulin induced glucose transport, and the actions are more effective than metformin, the positive control drug for type 2 diabetes therapy. Moreover, OM2 was found to be able to activate the phosphorylation of key protein IR, Akt and PI3K in insulin signal pathway to increase the production of IR and GLUT4 by using ELISA and real time quantitative RT-PCR analysis. OM2 could also increase the phosphorylation of AMPK and ACC in AMPK signal pathway to regulate lipid metabolism. So OM2 can activate both the insulin signal pathway and AMPK signal pathway to increase insulin sensitivity. Furthermore, by using western blot assay, marine oligosaccharide OM2 was also found to be able to activate the insulin signal pathway to regulate the glycogen synthesis and gluconeogenesis in liver, and can attenuate the hyperglycemic symptom in diabetes mice. Moreover, fluorescence labeled marine acidic oligosaccharide OM2 was found to be able to localize to the mitochondria after its internalization into skeletal muscle C2C12 cells by using living cell imaging, which suggested that the anti-diabetes effect of marine acidic oligosaccharide OM2 might be related to its improving the functions of mitochondria in skeletal muscle cells.In conclusion, marine oligosaccharide chromium complex OM2 which having good anti-diabetes effects in vivo and in vitro were successfully prepared in this research. The marine oligosaccharide OM2 could activate both the insulin signal and AMPK signal pathways to regulate the glucose and lipid metabolism, and increase insulin sensitivity to improve the insulin resistance. Moreover, the researches about improving insulin resistance mechanisms of marine oligosaccharide OM2 can provide the theoretical basis for developing it into new type anti-type 2 diabetes marine drugs.