Function And Mechanism Of Hacd2 In Lipid Metabolism

In mammals,fatty acids（FA）with carbon chain lengths up to 16 are synthesized by the de novo FA synthesis pathway and then further elongated by the FA elongation pathway.Todate,the study on FASN has been relatively mature,whereas the fatty acid elongases,especially the Hacd family of elongases,have not been systematically studied.3-hydroxyacyl-Co A dehydratase 2（Hacd2）participates in FA elongation as a member of the Hacds family,is mainly involved in the third step of fatty acid elongation.There are few reports about Hacd2 gene,and the effect of Hacd2 on mammalian body,especially the effect of lipid metabolism,is still unknown.With the development and maturity of transgenic animal technology,it is now possible to use gene knockout technology to achieve global knockout and tissue specific knockout of target genes.In this study,we used CRISPR/Cas9 technology and Cre-lox P system to construct Hacd2 knockout mice and tissue-specific Hacd2 knockout mouse models to explore the role of Hacd2 in lipid metabolism and how it affects the growth and development of the body or regulates metabolic homeostasis.This study has done the following work:（1）After Hacd2^+/-mice model were successfully constructed,Hacd2^+/-mice were mated with Hacd2^+/-/WT mice.Genotypes of the offspring mice were identified and statistically analyzed.It was found that the genotypes of the offspring mice did not conform to Mendelian law of inheritance,and no homozygous mice were born.We hypothesized that global deficiency of Hacd2 in mice resulted in embryonic lethality.We used the method of site-directed mating to preliminarily determine the lethal time of the embryo at about E7.5 days.Subsequently,we analyzed the fatty acid composition of homozygous embryos and wild-type embryos as the control group and found that the content of long-chain fatty acids in homozygous embryos was significantly lower than that in wild-type embryos.（2）As an important site for lipid synthesis,adipose tissue can secrete a variety of hormones and cytokines through a variety of secretion ways,and then regulate the energy metabolism of the body.Therefore,we first constructed adipose tissue-specific Hacd2 knockout mouse model using the Cre-Lox P system.In the study of knockout mice,we found that loss of Hacd2 in adipose tissue had no significant effect on the growth and development of the mice,and there was no significant change in body weight or insulin sensitivity compared with wild-type mice.Metabolic cage monitoring also showed no significant change in energy expenditure in mice.（3）After finding that adipose tissue is not a target organ for Hacd2 action,we knocked out Hacd2 gene in liver tissue,another important site of lipid synthesis.In liver-specific knockout mice and corresponding wild-type mice on a high-fat diet,we found that the loss of Hacd2 in liver protected mice from obesity and insulin resistance induced by a high-fat diet.In our study,we found that the loss of Hacd2 in liver tissue could affect the production of ceramide in the serum of mice,and ceramide has a strong ability of associating water molecules,which maintains skin moisture through the formation of a network structure in the cuticle.The significant decrease of ceramides disrupts the function of the skin barrier in mice,leading to the destruction of heat exchange in mice,and increases the energy consumption of adipose tissue browning,which is dominated by heat production.In the further study,we found that the loss of Hacd2 in liver increased insulin sensitivity in mice.Basal blood glucose was also significantly controlled in the mice.After acute insulin injection,the phosphorylation levels of Akt and GSK3βproteins in liver tissues were significantly up-regulate.These results indicate that:1)Hacd2 totally absence can cause mouse embryos to death,suggesting that Hacd2 is indispensable in living organisms and participates in important processes of body growth and development;2)We demonstrate that loss of Hacd2 expression in the liver but not in adipose tissue reduces the level of very-long-chain fatty acids,alters skin thermal and water barrier,increases mouse energy expenditure and adipose tissue browning,and protects mice against high fat diet induced obesity as well as associated fatty liver disease and diabetes.The results suggest that Hacd2 may be a promising therapeutic target for the management of obesity and associated metabolic diseases,and also provide certain guiding significance for future clinical studies.