The Pathogenesis Of Type 2 Diabetes Induced By Oxidative Stress In A Long-term High-fat High-fructose Diet

Background:Long-term consumption of a western diet is a major cause of type 2 diabetes mellitus(T2DM).However,the effects of western diets on the pancreas remain unclear.This study aimed to explore pancreatic morphological,functional and transcriptomic changes among islet sub-populations of rats fed with a high-fat high-fructose(HFHF)diet.Methods:Fifty T2DM patients and fifty healthy candidates were recruited,and their plasma insulin,C-peptide,proinsulin and intact proinsulin levels were measured to evaluate the impact of T2DM on protein process and secretion function of islet cells.An 18-month HFHF diet was used to trigger the onset of T2DM in rats.Tissues and blood were collected from rats at the beginning,3 and 18 months of treatments.Plasma biochemical parameters and inflammatory factors were determined to reflect the levels of metabolism and inflammation in rats.The effect of HFIIF diet on oxidation was evaluated through plasma oxidoreductases activity and the level of nucleic acid oxidation markers(8oxo-Gsn 和 8-oxo-dGsn)in urine.The structure and function of the pancreas were determined by a histological analysis and hormone examinations.Changes in pancreatic cell composition,function,and the transcription levels of oxidoreductase gene induced by long-term HFHF diet and aging were further explored using a 10 x genomics single cell RNA-sequencing analysis(scRNA-seq).Results:①Disorders of glucose regulation in islets of T2DM patients:The levels of insulin,C-peptide,proinsulin,and intact proinsulin in T2DM patients were significantly higher than those in healthy subjects.Besides verifying previous studies,we confirmed that proinsulin and intact proinsulin could be used as markers of hormone-processing ability of islet cells in subsequent rat experiments.②A HFHF diet could cause metabolism disorders and structural changes in tissues:After 3 months of a HFHF diet,the level of plasma glycosylated serum protein,pyruvic acid,total cholesterol and low-density lipoprotein cholesterol in HFHF group were significantly higher than those in control group.At the same time,severe vacuolar degeneration occurred in liver and islet volume in pancreas increased significantly in the HFHF group.After 18 months of a HFHF diet,vacuolar degeneration in livers was aggravated,and the inflammation and fibrosis of the liver and kidney exacerbated.Meanwhile,islet volume decreased and the shape changed from oval to irregular.Fasting insulin,C-peptide,proinsulin and intact proinsulin were significantly increased in the HFHF group compared with the age-matched controls,suggesting a long-term HFHF diet could induce islets dysfunctions in blood glucose regulation and hormone processing.③A HFHF diet could increase the levels of oxidation and inflammation in rats:Plasma oxidative parameters(such as superoxide dismutase,xanthine oxidase and glutathione reductase)and nucleic acid oxidation markers(8-oxoGsn)elevated after 3 months of HFHF diet.At the same time,the expression levels of oxidative nucleotides scavenger proteins MTH1 and NUDT5 increased in liver.When rats were fed a HFHF diet for 18 months,the oxidative parameters levels in plasma were further increased,the expression levels of oxidative nucleotides scavenger proteins in kidney and pancreas were changed.Moreover,the level of inflammatory factors,such as IL-1β and TNF-α,were significantly increased,suggesting that HFHF diet increased the degree of oxidative stress before inducing inflammation.④A long-term HFHF diet induced changes in pancreatic cell composition and transcriptome of rats:The results of scRNA-seq analysis also verified that the transcriptional level of oxidoreductases changed differently in islet sub-populations with aging and the long-term HFHF diet.Oxidative stress and inflammatory-related pathways were also enriched in the HFHF group.The expression levels of major glucose-regulating hormone genes in each islet subpopulation cells increased with age but were significantly down-regulated by a long-term HFHF diet.GO analysis showed that hormone secretion related pathways were upregulated in islet alpha,beta,and PP cells of the long-term HFHF group.In the HFHF group,alpha and PP cells count decreased,while immune cells count increased.Conclusions:We demonstrated the long-term HFHF diet associated decrease in pancreatic islet endocrine cells numbers,down-regulation of functional genes expressions,and transcriptomic changes of oxidoreductase genes which underlie pancreatic islet functional decay,suggesting a possible underlying mechanism of the T2DM and might provide new insights to prevent the development of the diet-induced T2DM.