The Effect Of ANGPTL8 On Cattle Preadipocytes Differentiation

Angiopoietin-like proteins(ANGPTLs)as secretion proteins are not only involved in the regulation of angiogenesis,but also regulated the lipids metabolism,glucose metabolism,and insulin sensitivity.Its family member ANGPTL8 can regulate triglyceride(TG)content in the blood and affect the body's transport and absorption of triglycerides.The content of ANGPTL8 in vivo is closely related to the occurrence of hyperlipidemia,atherosclerosis and diabetes.At present,the molecular mechanism of ANGPTL8 regulating preadipocyte differentiation and lipid deposition is not yet clear.Therefore,in this study,based on the primary culture method of cattle preadipocytes,adenovirus-mediated overexpression and lentivirus-mediated gene silencing were used to study the mechanism of ANGPTL8 affecting lipid deposition in cattle.At the same time,the expression pattern of ANGPTL8 and its effect on lipid differentiation-related genes were tentatively explored at the transcriptional level.The main results were as follows:1.Expression analysis of ANGPTL8 by qRT-PCR showed that ANGPTL8 was expressed in heart,liver,kidney,small intestine,muscle and adipose tissue of adult cattle.The expression of adipose tissue was higher than other tissues.Tissue culture was used to culture cattle preadipocytes and induce differentiation.preadipocytes after induction,the expression levels of PPAR?,FABP4,LPL and C/EBP? were significantly upregulated.The expression level of ANGPTL8 in the process of inducing differentiation of preadipocytes increased significantly with the differentiation and maturation of adipocytes,and the expression level of ANGPTL8 and lipoprotein lipase(LPL)increased at the same time.2.The overexpression of cattle preadipocyte ANGPTL8 showes that lipid accumulation ability of preadiposes was significantly enhanced.The results of qRT-PCR showed that overexpression of ANGPTL8 significantly reduced the expression of SREBP1 and LPL genes during the induction process,but had no significant effect on the adipocyte differentiation marker genes PPAR? and C/EBP?.3.The dual luciferase activity analysis of the promoter deletion fragment showed that the core promoter of ANGPTL8 was-522~-885 regions.Bioinformatics predictions revealed that the binding sites of PPAR?,SREBP1,C/EBP? and Znf423 transcription factor exist in the promoter region of ANGPTL8 is regulated by SREBP1 and PPAR? transcription factors.In summary,the primary culture and induction methods of cattle preadipocytes were successfully established in this study.The ANGPTL8 tissue and cell expression profiles,overexpression and interference virus were constructed.The ANGPTL8 was preliminarily confirmed to promote cattle preadipocyte lipid deposition.The expression status of which is regulated by the SREBP1 and PPAR? transcription factors,provides a basis for further investigation of the mechanism of ANGPTL8 regulating preadipocytes differentiation and lipid deposition.