Effect Of Hoxa5 On Development And Remodeling In Mice Adipose Tissue And The Underlying Machinism

Animal adipose tissue is an important organ of the body,mainly responsible for energy storage and metabolism,and secreting a variety of adipocytokines involved in the regulation of the body's physiological functions.The development and remodeling of adipose tissue is not only closely related to human obesity,diabetes,and metabolic syndrome,but also is the key events to affecting the lean meat percentage and meat quality of livestock.Homeobox genes,belonging to the developmental transcription factor,play a part in the determination of adipose tissue expandability and body fat distribution.Hoxa5,one of the Hox genes,has been identified differentially expressed in adipose tissue depots.It is reported that the Hoxa5 expression is decreased with a high fat diet and increased in human adipose tissue after fat loss surgery.There is a strong correlation between brown adipocytes adipogenesis and Hox genes expression.Lipid accumulation decreased in the Hoxa5-siRNA treated 3T3-L1 adipocytes.In addition,Hox5 mutant mice have increased lung inflammation and HOXA5 can be applicated to vascular inflammation.However,the regulatory function and molecular mechanisms of Hoxa5 on adipose tissue still needs to be illustrated.In this study,Hoxa5 was overexpressed/interfered both in adipocytes or macrophages and in vivo,and the mechanism of Hoxa5 on regulating adipose tissue differentiation,trans-differentiation and chronic inflammation were systematically studied.The main results are listed below:1.Effect of Hoxa5 on adipocytes differentiation and mitochondrial biogenesis.Quantitative real-time PCR(qPCR)and western blot were performed to determine Hoxa5 expression in primary adipocytes and in adipose tissues from mice.Lipid accumulation was evaluated by bodipy staining.Dual luciferase assay was applied to explore the transcription factor of Hoxa5 and the transcriptional target gene modulated by Hoxa5.Results:(1)A significant reduction of Hoxa5 expression was observed in adipose tissue of High Fat Diet(HFD)induced obesity mice.(2)This study determined Hoxa5 increased adipocytes differentiation and mitochondrial biogenesis in adipocytes in vitro.(3)CEBP? was determined a factor of Hoxa5 and inhibited methylation level of Hoxa5 by combining on the promoter of Hoxa5.(4)Fabp4 was transcriptional activation by Hoxa5.(5)Hoxa5 promotes adipocytes differentiation by inhibiting PKA/HSL pathway.2.Regulatory mechanism of Hoxa5 on white adipose tissue browning.To explore the role of Hoxa5 in inflammation-mediated mice adipose tissue browning.Transcriptional and methylation regulation was determined by RNA-seq,luciferase assay,electrophoretic mobility shift assay(EMSA)and bisulphite conversion experiment.Result:(1)Hoxa5 and Tenascin C(TNC)were involved in WAT inflammation and browning in mice with LPS injection.(2)Hoxa5 inhibited TNC involved activation of TLR4/NF?B signal pathway and promoted WAT browning.(3)The BMP4/Smad1 signal,closely related to browning,was activated by Hoxa5.(4)Hoxa5 relieved adipocyte inflammation by decreasing TNC-mediated TLR4 transducer and activator of the NF?B/NLRP3 pathway.(5)Descended methylation level increased Hoxa5 expression in cold exposure.3.Effect of Hoxa5 on obesity-induced chronic inflammation of adipose tissue.Obesity represents the chronic inflammatory state promoted by the infiltration of pro-inflammatory M1-macrophage into white adipose tissue(WAT).To elucidate the regulation of Hoxa5 on adipose tissue during obesity,this study overexpressed/interfered Hoxa5 in fat cells and macrophages,and performed the co-culture experiments to study the regulatory mechanism of Hoxa5 on chronic inflammation.This study further confirmed the effect of Hoxa5 on adipose tissue by injection of Hoxa5 over-expressing/interfering adenovirus vectors in high-fat-induced obese mice.The results are as follows:(1)Hoxa5 inhibited inflammatory cytokines by alleviating ER stress in adipocytes.(2)Hoxa5 decreases ERS via PERK/eIF2? signal pathway in mice adipocytes.(3)Hoxa5 alleviates adipose tissue inflammation by inhibiting M1 macrophage infiltration and promoting M2 macrophages polarization.(4)Hoxa5 increases M2-polarized macrophages by transcriptional activating PPAR? signal pathway.(5)Hoxa5 inhibits obesity by promoting M2 polarization macrophages in HFD mice.In summary,Hoxa5 has an important physiological role on development,thermogenesis and inflammation of adipose tissue.Hoxa5 promotes differentiation of pre-adipocytes and mitochondrial biogenesis.Hoxa5 also promotes browning of white adipose tissue by inhibiting LPS-induced inflammation and activating BMP4/Smad1 pathway.In obesity state,Hoxa5 alleviates chronic inflammation of adipose tissue by reducing ER stress in adipocyte and promoting M2 macrophage polarization.This research reveals the regulatory function and mechanism of Hoxa5 on mice adipocytes as well as adipose tissue,providing new clues for elucidating the regulation mechanism of animal adipose tissue development and remodeling,and providing theoretical basis for improving lean meat rate and meat quality.In addition,the study open up new target of treatment for obesity and type 2 diabetes.