The Effect And Mechanisms Of Intestinal And Hepatic Bile Acid Sensing In The Remission Of Type 2 Diabetes After Duodenal-jejunal Bypass Surgery

Part Ⅰ The construction and evaluation of T2DM rat model and duodenal-jejunal bypass modelBackgroundDiabetes mellitus is a most common chronic disease, and more than patients are type 2 diabetes mellitus (T2DM). According to the international diabetes federation (IDF) collected from 91 countries diabetes prevalence data, the results show that the estimated prevalence of diabetes in 2010 was 6.4%(285 million), and in 2030 will rise to 7.7%(439 million), in which developing countries diabetes will increase by 69%, while developed countries will increase by 20%. Although by changing the way of life, including diet, exercise, weight loss, smoking and comply with certain medications, which has some effect for prevention and control of diabetes. Due to patients need lifelong medication or insulin injections, it is hard to guide adherence, meanwhile, maintain long-term stable blood sugar effect is not ideal. Weight loss surgery can improve obesity patients with systemic symptom such as overweight, high blood pressure, high blood lipids, diabetes and other symptoms. Studies have shown that weight-loss surgery is not purely depend on weight loss effect to relieve the T2DM, there were other anti-diabetic effect depends on the weight loss of the path. There are a large number of clinical studies and animal experiments confirmed that the gastric bypass surgery for the treatment of T2DM have obvious effect, but its the proximal small intestine hypothesis, the distal small intestine hypothesis theory for ease of T2DM molecular mechanism remains unclear.ObjectiveIn our study, we develop a new T2DM rat model, Duodenal-jejunal bypass (DJB) will be performed, we will evaluate the glycemic control and the associated mechanisms. The study was conducted to investigate:(1) Evaluation of T2DM rats model, to guarantee the model consistent with the characteristic of T2DM close; (2) Monitoring the DJB operation T2DM rats sugar metabolism and the effect of this model to prove its effect on glucose metabolism can improve; (3) To evaluate the lipid parameters of DJB; (4) To detect the gastrointestinal hormones level associated with glucose metabolism. (5) To detect the change of concentration of serum bile acids after DJB, and analyze its correlation with glucose metabolism.MethodsThe SD rats were feed by high fat diet for one month. Then low dose of streptozotocin (STZ,35 mg/kg) was injected intraperitoneally to induce hyperglycemia. The SD rats were divided into DJB, sham DJB (S-DJB) and the control, the following parameters are detected (1) The fasting blood glucose:The fasting blood glucose were detected once every week; (2) The glucose tolerance:The oral glucose tolerance test (OGTT) was evaluated after surgery; (3) The insulin sensitivity:insulin tolerance test (ITT) was evaluated after surgery; (4) The function of pancreatic β cells:The glucose stimulated insulin secretion was determined after surgery; (5) The gastrointestinal hormones level:The glucose stimulated glucagon-like peptide-1(GLP-1), peptide YY (PYY),Glucose dependent insulinotropic peptide, (GIP) were detected after surgery; (6) The lipid parameters: triglycerides、FFAs、total cholesterol levels and liver function index were determined after surgery. (7) The change of concentration of serum bile acids and its correlation with blood glucose、insulin and intestinal hormones. Results (1) The T2DM rat model was successfully induced by HFD and low dose of STZ. (2) There is no significant differences between the DJB and S-DJB groups in terms ofbody weight and caloric intake (*P> 0.05). (3) Compared with S-DJB group, The fasting blood glucose was decreasedsignificantly in the DJB group, a significant difference was observed after surgery(*P<0.05). (4) Compared with S-DJB group,the glucose tolerance was significantly improvedafter DJB.(5) Compared with S-DJB group,the insulin sensitivity was significantly improved 2 weeks after DJB, and more marked improvement was observed at 4 weeks after surgery, while more drastic difference was observed in the S-DJB group.(6) Compared with S-DJB group,the GLP-1 and PYY levels in the DJB groups were much higher at 2、16 weeks.(7) Compared to the S-DJB group, triglycerides and FFAs were significantly lower after DJB.(8) Compared to the S-DJB group, serum bile acids was remarkably increased after DJB and there was significant relevance between the change of serum bile acids and the change of blood glucose、insulin and intestinal hormones.Conclusions(1) The T2DM rat model was successfully induced by HFD and low dose of STZ.(2) DJB could rapidly and sustainably improve the fasting blood glucose in the T2DM rat model.(3) DJB could significantly improve the glucose tolerance and insulin sensitivity of T2DM rats.(4) GLP-1 and PYY,were significantly increased after DJB in the T2DM rat model; but GIP had no change.(5) The FFA and triglyceride level were effectively decreased after DJB surgery.(6) serum bile acids was remarkably increased after DJB and there was significant relevance between the change of serum bile acids and the change of blood glucose、insulin and intestinal hormones.This study include that DJB could rapidly improve glucose homeostasis. The increase of GLP-1 and PYY level may contribute to the glucose homeostasis improvement. We speculated that DJB might influence the changes of GLP-1, PYY and other intestinal hormones in some way associated with bile acid, and ultimately improve the glycometabolism of T2DM rats model.The concrete mechanisms are still indistinct, and we will further research in the following study.Part II The mechanisms of elevated bile acid concentrations after DJB of T2DM ratsBackgroundBile is secreted by most vertebrate liver cells, a kind of green and with an indication of alkaline liquid. Bile composition is complex, its main components is the bile acid. Bile acid is mainly exist in the enterohepatic circulation system and protect liver function by recirculation of the important components of bile. After entering the intestines, most bile acid is reabsorbed into the blood stream in the terminal ileum, and then promote liver glycogen synthesis, inhibit sugar dysplasia through the excited liver TGR-5 and FXR a alpha receptors. The clinical and animal experimental study of DJB surgery found that DJB marked increase in postoperative serum bile acid concentration, ratio of components and bile acid also has had the obvious change, but the bile acid synthesis from the start the key enzyme of cholesterol 7 alpha hydroxylase (CYP7A1), and no obvious change, the main parts in bile acid absorption as the terminal ileum. Recent research suggests that it can also obviously improve the body’s sensitivity to insulin through duodenum inversion technique to bile flow directly to the terminal ileum. This suggests that postoperative bile acid concentration increases may not be a synthetic increase. DJB surgery improve the body’s sensitivity to insulin by speeding up the bile acid contact with the terminal ileum, the bile acid in the terminal ileum of heavy absorption increases, causing bile acid concentrations in the blood increases, further by activating the liver bile acid receptor promote liver glycogen synthesis, inhibition of sugar dysplasia. However, there is no elevated DJB postoperative bile acid in the blood concentration of mechanism. ObjectiveNumerous studies have shown DJB postoperative bile acid concentration increased were positively correlated with the improvement of insulin sensitivity in the body’s blood and bile acid concentration increases. Because the elevated bile acid concentration in the blood can activate the liver bile acid receptor promote liver glycogen synthesis, inhibition of sugar dysplasia, in regulating insulin sensitivity play a crucial role. We study in this part of the terminal ileum bile acid transporter and bile acid synthesis in the liver protein expression of key enzyme for testing, in order to make clear DJB postoperative bile acid concentrations in the blood increases.MethodsApplication of tissue samples obtained in the first part, using Western blotting technique detect the following outcomes:CYP7A1 in liver tissue and CYP27A1 expression changes; With immunohistochemical technique to test the following indicators:ASBT and bile acid binding protein in terminal ileum organization IBABP expression.Results(1) Compared with S-DJB group, DJB group expression of CYP27A1 in rat liver tissue has a downward trend, but did not reach significant difference (P> 0.05), the expression of CYP7A1 quantity there was no significant difference (P> 0.05).(2) Compared with S-DJB group, DJB group rats ASBT and IBABP expression significantly increased in terminal ileum tissue (* P< 0.05).Conclusions(1) After the DJB surgery, the bile acid synthase expression in T2DM rat liver tissue of active has no obvious change, which prove that the rise of bile acid is not increased bile acid synthesis;(2) After the DJB operation, the end of the T2DM rats ileum bile acid transporter and bile acid binding protein expression quantity increased, bile acid proof in terminal ileum absorb increasePartⅢ The effect and mechanisms of elevated bile acid on glycometabolism after DJBBackgroundBile acid and semisynthetic derivatives can activate the terminal ileum L cells of bile acid receptor on cell membrane-G protein coupled receptor (TGR-5) promote the secretion of glp-1, which can promote insulin secretion, the beta cells proliferation and inhibiting apoptosis; In addition, the bile acid can also by activating the nucleus of bile acid receptor-L method, alcohol X receptor (Farnesoid X receptor alpha, FXR alpha) promoting the Fibroblast growth factor (Fibroblast growth factor, FGF)-19/15 of the secretion. FGF-19/15 not only can regulate bile from scratch, but also has promote the liver glycogen synthesis, lowering blood sugar of insulin. FXR a is an important receptor of the bile acid, mainly expressed in the liver and gut. FXR a widely involved in the regulation of carbohydrate and lipid metabolism. FXR a can form in SHP feeling and inhibition of bile acid in the liver, change bile acid by SHP transporter BSEP and NTCP structure promote the secretion of bile acid and reduce reuptake. FGF-19 diet induced obesity rats energy consumption, weight loss and increased insulin sensitivity. FXR a activation induced in the intestinal FGF19 (FGF15 rat) expression and secretion. Intestinal express FGF19/FGF15 secreted into the blood circulation as promote glycogen synthesis factors after a meal, whch may be an important mechanism of controlling glucose metabolism. TGR-5 is another key of bile acid receptor in liver, intestine, expressed in brown adipose tissue. Study shows that the bile acid can combine TGR-5 stimulate intestinal endocrine T cells release glp-1, promote insulin beta cell proliferation and secretion of insulin. Research found that TGR-5 agonists (TNT-777) in the body can induce glp-1 secretion. The mechanism is increasing the level of intracellular cAMP, changing the ATP/ADP ratio, internal flow caused by calcium. By the above analysis, the first DJB changes lead to increase the reuptake of bile acid in gut anatomical structures, which can stimulate the TGR-5. Then, TGR-5 promotes intestinal T cells to release more glp-1 further control of glucose metabolism. Gastric bypass surgery by speeding up the bile acid contact with the terminal ileum, make L early changes of bile acid in intestinal cells, promote the secretion of glp-1 and FGF19/15 and improve the body’s sensitivity to insulin. DJB postoperative bile through what way, however, affect the body glucolipid metabolic mechanism has not been determined.ObjectiveNumerous studies have shown DJB postoperative bile acid concentration increased in the body’s blood, and the bile acid levels have correlation with glucolipid metabolism. Because elevated bile acid concentration can be activated in the intestine, liver circulation of bile acid receptor promote liver glycogen synthesis, inhibit dysplasia, sugar sugar fat metabolism in regulating insulin sensitivity and play a crucial role. We study in this part of the terminal ileum FXR a, TGR-5 protein expression were detected, in order to make clear DJB postoperative bile acid levels affect glucolipid metabolism mechanism.MethodsApplication of tissue samples obtained in the first part, by immunohistochemical technique to test the following indicators:the FXR a expression changes in the liver tissue and ileum; the TGR-5 expression changes in the liver tissue; With western blot technique to test the following indicators:PEPCK and G6Pase in the liverResults(1) Compared with S-DJB group, DJB group FXR a expression was significantly higher in the ileum, but there was slight increase in liver.(2) Compared with S-DJB group, DJB group of TGR-5 expression in liver tissue increased obviously.(3) Compared with S-DJB group, The DJB group expression levels of PEPCK and G6Pase in the liver increased significantlyConclusions(1)After the DJB surgery, FXR a expression in terminal ileum tissue of T2DM rats increase, the expression level of PEPCK and G6Pase in liver tissue was increased, that indicates that bile acids improving glucolipid metabolism may be related to the regulation of the hepatic gluconeogenesis pathway through the intestinal bile acid-FXR α-FGF-19/15 pathway.(2)After the DJB surgery, TGR-5 expression of T2DM rat increase in liver tissue, proved bile acid improve glucolipid metabolism through the bile acid-TGR-5 pathways.