Studies for mechanisms of bovine marbling and developments of genetic markers

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To investigate genes involved in ruminants intramuscular adipogenesis, sixteen target genes(8 up and 8 down regulated genes) with dramatically changed expression(P<10^<-12>) were selected between differentiation and proliferation phases libraries in our previous SAGE studies of a clonal bovine intramuscular preadipocyte(BIP) cell line(Mizoguchi, 2010), with reference to 3T3-L1 cell lines as monogastric animals. We harvested the BIP and 3T3-L1 cells 0, 3, 6, 9 and 12 days after adipogenic stimulation containing all-trans retinoic acid(RA), which inhibits preadipocyte differentiation in non-ruminants and measured the gene expression levels by quantitative real-time PCR and the accumulation levels of triglyceride(TG). Fourteen genes expression pattern of the 16 genes during adipogenesis were confirmed the SAGE studies. The expression pattern of eleven genes clearly differed between 3T3-L1 and BIP cells. Simultaneously, we investigated the profiles of target genes by RA. TG accumulation was significantly induced(P<0.05) in BIP cells whereas completely inhibited(P<0.05) in 3T3-L1 cells by RA at day 6 after adipogenic stimulation. RA induced the expression levels of six genes and suppressed these of eight genes at day 3 after adipogenesis in BIPs(P<0.05 ; respectively). The effect of ADFP down-regulation on the accumulation of TG was examined by transfecting the BIP cells with ADFP siRNA. The expression level of ADFP in the ADFP-siRNA transfected cells was 21.4% of that in the scr-siRNA transfected cells in BIP 3 days after induction(P<0.05). Also, 6 days after induction, the accumulation of TG in the ADFP-siRNA transfected cells was 20% less than in the control cells(P<0.001). Our findings of gene expression profiles by RA will cause a favorable and detailed understanding in the molecular mechanisms of bovine intramuscular adipogenesis.