Artificial Sweeteners Regulate ?-cells Endocrine Function Of Pancreatic Islets Through The Sweet Taste Receptor Pathway

Artificial sweeteners,because of its characters of high sweetness and non-calorie,have served as the substitutions of natural sugar and widely used in food industry.Recent years,more and more studies have indicated that artificial sweeteners could destroy blood glucose tolerance,but its specific mechanism is not clear yet.Islets composing of ?,?,?endocrine cells are the vital organ of animals and human in regulation of blood glucose balance.Islets endocrine cells release kinds of hormones like insulin and glucagon to maintain dynamic homeostasis of blood glucose of organism.Sweet recepters play a role in signal reception of sugar and artificial sweeteners.Some of research have showed that sweet receptor subunit T1R2 and downstream signal transduction protein?-gustducin exist in islets endocrine cells.They might mediate the hormone release function and blood glucose regulation function of islets endocrine cells.However,for the detailed distribution of the sweet receptor T1R3/T1R2 and the sweetness signal transduction downstream G protein a-gustducin in islets,and the function of artificial sweeteners in the regulation of hormone release of islet endocrine cells is unclear yet.Therefore,this study used wild-type and sweet receptor T1R3 knockout mice as experimental animal models,and used islet isolation,RT-PCR,immunohistochemistry,HE staining,Elisa,laser confocal and other experimental techniques to demonstrate the detailed distribution of receptor T1R2/T1R3 and downstream signal transduction protein a-gustducin in islets,and to discuss the role of artificial sweeteners in islet ?-cell insulin release and glucose-induced insulin release,as well as to clarify the important role of sweet receptor in the above effects.In addition,the study used sucralose as representative of artificial sweeteners to analyze the phenotypic characteristics,blood glucose,insulin release and islets morphological changes after long-term exposure to artificial sweeteners.The specific experimental results are as follows:1.Expression and distribution of sweet receptors in mouse isletsThe results of RT-PCR and immunohistochemistry showed that the islets specifically express the sweet receptor and sweet-stimulus signal transduction protein a-gustducin;the results of double immunofluorescent indicated that the sweet receptors T1R3 and T1R2 were widely existed in the ?,? and ? cells of mice islets,and the sweet signal transduction protein ?-gustducin was present in ? and ? cells.2.Artificial sweetener synergizes with glucose to induce beta cell insulin releaseStatic incubation combined with dynamic perfusion of isolated mouse islets,ELISA analysis of insulin release,the results showed that artificial sweeteners(sucralose,acesulfame,saccharin)alone stimulating islet had no significant effect on the release of ?-cell insulin,but significant synergy glucose-induced insulin release.The efficacy of artificial sweeteners to induce insulin release was strongly dependent on the presence of glucose.3.Loss of the sweet receptor T1R3 attenuates the effect of sweetener synergistic glucose on ?-cell insulin releaseThe sweet receptor T1R3 gene knockout and littermate wild type mice were used as experimental animals to isolate islets,combined with static islet incubation and ELISA.We found that although knockout of sweet receptor T1R3 had no effect on insulin secretion induced by single artificial sweetener sucralose,acesulfame,saccharin,but significantly attenuated the synergistic effect of glucose-induced insulin release in combination with artificial sweeteners.This result revealed an important role for the sweet receptor T1R3 in islets in regulating the pathway of insulin release.4.Loss of sweet receptor T1R3 reduces islets cells density and number of beta cellsThe sweet receptor T1R3 gene knockout and littermate wild type mice were used as experimental animals.Frozen sections were combined with immunohistochemistry.The results showed that compared with wild type mice,the knockout of sweet receptor T1R3 gene significantly reduced the number of ? cells in islets,and showed down-regulated trend in ? and ? cell numbers,islet area of the mice islet,but there was no significant difference in them.The effect of combining the sweet receptor T1R3 gene knockout on the synergistic effect of artificial sweetener synergistic glucose indicated that the knockdown of the sweet receptor T1R3 gene might trigger the decrease of the number of mouse islets and the number of beta cells in the islets,and thereby attenuated levels of sweetener-induced insulin release.5.Effects of long-term exposure to sucralose on blood glucose physiological and biochemical indexes,glucose tolerance,first-phase insulin release and islet cells densityMale C57BL/6 mice exposed to sucralose for 16-weeks were used as experimental subjects.The body weight and food intake of the exposure process showed that exposure of sucralose had no significant effect on body weight and daily food intake,but because of the sweetness characteristics of the sucralose,daily solution intake was significantly increased compared with the control water group;blood glucose tolerance test during exposure showed that sucralose exposure enhanced glucose tolerance in mice,and this tolerance might come from increasement of insulin release level during the long-term exposure of sucralose.In summary,results of the study demonstrated that the artificial sweetener synergized glucose-induced insulin release,and revealed the important role for the sweet receptor T1R3 in regulating the pathway of insulin release.The above research may provide a new way for blood glucose regulation in future.