Regulation Of Porcine Chemerin, ChemR23 And GPR1 In Lipid Metabolism

As a chemoattractant, chemerin takes part in the innate immunity and acquired immunity through recruiting kinds of immunocytes to inflammatory sites. Recently, It elicits people interests that chemerin serves as an adipocytokine. More and more studies uncover that chemerin is closely related to obesity, systemic inflammation and metabolic syndrome. We attempted to discuss the effect and underlying mechanism of chemerin on obesity and insulin resistance. It should be of particular importance to clarify the regulation mechanism of animal lipid metabolism and the pathogenesis of type 2 diabetes.Using Meishan pig as a research object, we cloned the procine chemerin and its receptors ChemR23 and GPR1 and analysed their genomic structures as well as the profiles of their expressions in various tissues for the first time. The similarity in nucleotide sequence and structure between ChemR23 and GPR1 was analyzed by bioinformatics. The expression of chemerin, ChemR23 and GPR1 which is regulated by the two transcription factors in lipogenesis (KLF15 and PPARy) was acquired. The alteration of expressions of genes involved in lipid metabolism in hepatocyte after overexpression chemerin, ChemR23 or GPR1 was detected. The detailed results are as follows:1. The coding region of porcine chemerin, ChemR23 and GPR1and partial non-coding sequence of porcine chemerin were amplified and then bioinformatic analysis was conducted. The coding region of porcine chemerin covered 492bp encoding 163 amino acids. The uninterrupted CDS of ChemR23 covered 1092bp and encoded 363 amino acids. Porcine GPR1 spanned 1068bp and encoded 355 amino acids. Bioinformatic analysis also showed that ChemR23 and GPR1 have the corresponding transmembrane domains and similar hydrophilicity/hydrophobicity distribution in extracellular loop 2 and 3.2. Using Real-time PCR, we detected the expression profiles of chemerin, ChemR23 and GPR1 in heart, liver, spleen, lung, kidney, stomach, skeletal muscle, testis, subcutaneous fat, epididymal fat, brain, small intestine in Meishan pig. The result showed that chemerin and GPR1 both expressed abundantly in liver, implying their association in hepatic metabolism. What is more, all the three genes had the potential abundance in adipose tissue. It implied that chemerin may play a role in lipogenesis.3. In order to reveal the regulation mechanism of chemerin, ChemR23 and GPR1 in lipogenesis, KLF15 or PPARy was overexpressed in IBRS-2 and then realtime PCR was conducted to detect the changes in expression of chemerin, ChemR23 and GPR1. The results manifested that Chemerin, ChemR23 and GPR1 were up-regulate by KLF15. However, PPARy has no effect on the expression of all three genes.4. Chemerin led to lipid metabolism dysfunction and triggered fat deposition. Different forms of chemerin in circulation and its two receptors ChemR23 and GPR1 were constructed on a eukaryotic expression vector pcDNA3.1 and were overexpressed in human hepatoma cell line HepG2, those rate-limiting enzymes and key transcription factors involved in lipid metabolism were assessed by employing realtime PCR. It indicated that those genes participated in the de novo synthesis of FFAs and of triglyceride as well as those took part in fatty acid oxidation were all suppressed in their mRNA significantly. Due to cellular fatty acid oxidation mainly happenes in mitochondrial, the genes related to mitochondrial function were also determined. We found chemerin inhibits the genes related to mitochondrial biogenesis, mitochondrial fatty acid oxidation and anti-oxidation significantly. All these implied that P-oxidation in mitochondrial was impaired. What’s more, the uptake of FFAs was not altered by all these genes. MTP, which is a rate-limit transport protein in VLDL assemble, was repressed by chemerin, ChemR23 and GPR1. It supposed that chemerin signal pathway suppress export of VLDL. We implied that inhibition of fatty acid oxidation and VLDL secretion are the two main causes which led to lipid accululation inversely.5. High-energy status down-regulated chemerin in hepatocyte. The expression of chemerin in mRNA was curbed in the existence of free fatty acid, such as palmitic acid or oleic acid. What’s more, these effects depended on dosage and time. Chemerin was inhibited by insulin but not glucose. When SREBP or ChREBP was overexpressed, the expression of chemerin in mRNA was suppressed significantly. And SREBP, a key transcriptional factor involved in lipogenesis, inhibited the activity of the promoters of ChemR23 and GPR1.