| Insulin Receptor(IR)belongs to the family of receptor tyrosine kinases and is expressed in many different tissues and organs.It is a binding target for insulin to play a biological role at the cellular level.The insulin receptor gene can produce two functionally related and distinct subtypes through alternative splicing;the difference between the two is due to the involvement of exon 11.In particular,the specific function of IR-B in adipose tissue remains unclear.At present,there are few studies on the changes in the ratio of the two subtypes of IR in the steady state regulation of glycolipid metabolism,but the specific functions of the two insulin receptor isoforms in adipose tissue have not been known so far.In order to verify the function of IR-B in adipose tissue,a novel adipose tissue-specific knockout was constructed using Cre-LoxP technology using the pre-built IR-B-Flox mice and Adipoq-Cre mice.The IR-B gene mouse model(AdIR-BKO)was analyzed for its physiological metabolic phenotype to investigate the effect of deletion of IR-B on adipose glycolipid metabolism in adipose tissue.The comparison of physiological metabolic phenotypes of the model under different dietary conditions,under the specific conditions of IR-B gene deletion and environmental factors,IR-B affects the insulin signaling pathway and its possible mechanism of regulating glycolipid metabolism.The AdIR-BKO mice were constructed using Cre-LoxP technology,and the knockout efficiency was verified from the DNA and RNA levels by PCR and RT-qPCR,respectively.The normal diet or high-fat diet was fed,and the body weight data were monitored weekly.The mice were subjected to glucose tolerance test(GTT)and insulin tolerance test(ITT)at 8 weeks and 12 weeks old.The mice were dissected at 12 weeks of age,and the organs were weighed.Reanalysis.Using H&E,the effect of IR-BKO on the histomorphology of adipocytes was analyzed.The effect of IR-BKO on the morphology of adipocytes was analyzed by H&E staining.The genes related to lipid metabolism in adipose tissue were analyzed by RT-qPCR(such as Acc and Srebp).(Expression),Western Blot was used to detect the expression of adipose tissue synthase(such as Acc);the expression of lipid metabolism-related proteins and insulin signaling pathway-related proteins.This study successfully constructed the AdIR-BKO mouse model,which is the imbalance of IR-A and IR-B expression in adipose tissue,and IR-A is mainly expressed in adipose tissue of AdIR-BKO mice.The growth curve showed that under the RC condition,the body weight of AdIR-BKO mice and Flox mice were significantly different,but the weight of adipose tissue in AdIR-BKO mice was decreased.GTT and ITT results showed that glucose tolerance and insulin in AdIR-BKO mice were found.There was no significant change in sensitivity.After feeding on a high-fat diet,the body weight of AdIR-BKO mice in AdIR-BKO mice was lighter than that of the control Flox mice,and the weight of adipose tissue was reduced but did not produce statistical significance.After 8 weeks of high-fat feeding,GTT and ITT results showed that the glucose tolerance and insulin sensitivity of AdIR-BKO mice were worse.H&E staining of subcutaneous white fat in mice revealed that the subcutaneous white fat cells of AdIR-BKO mice were smaller than the control group,and RNA level analysis revealed that the lipase synthase of AdIR-BKO mice was significantly lower than that of Flox mice.H&E staining found that the liver of AdIR-BKO mice had obvious fat vacuoles,and the adipose tissue of AdIR-BKO mice was significantly larger than that of Flox.There was no significant change in brown adipose tissue.For the first time,our study established an experimental animal model of specific knockout of IR-B in adipose tissue,which revealed the role of different splicing subtypes of insulin receptor gene in the metabolism of glucose and lipids in the body,such as diabetes and obesity.New ideas for the treatment of related metabolic diseases. |
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