BMP and activin membrane-bound inhibitor (BAMBI) inhibits the adipogenesis of porcine preadipocytes through Wnt/ß-catenin signaling pathway.

The process of differentiation from preadipocytes to adipocytes contributes to adipose tissue expansion in obesity. Blocking adipogenesis may be conducive to the etiology of obesity-related diseases. BMP and activin membrane-bound inhibitor (BAMBI) is a transmembrane protein, which was identified as a target of ß-catenin in colorectal and hepatocellular tumor cells. However, whether BAMBI affects adipogenesis by Wnt/ß-catenin signaling remains to be explored. In this study, we distinguish BAMBI as an inhibitor of preadipocytes differentiation. We found that BAMBI was downregulated during preadipocytes differentiation. Knockdown of BAMBI increased adipogenesis and blocked Wnt/ß-catenin signaling by repressing ß-catenin accumulation. In BAMBI overexpression cells, lipid accumulation was reduced by promoting nuclear translocation of ß-catenin. Lithium chloride (LiCl) is an activator of Wnt/ß-catenin signaling, which is an inhibitor of glycogen synthetase kinase-3 (GSK-3), maintaining the stability of ß-catenin in cytosolic. We showed BAMBI strengthened the anti-adipogenic effects of LiCl. In addition, the results indicated that BAMBI was upregulated by ß-catenin. These observations illuminated that BAMBI inhibits adipogenesis by a feedback loop (BAMBI→ß-catenin nuclear translocation→BAMBI), which forms with Wnt/ß-catenin signaling.

CTSB promotes porcine preadipocytes differentiation by degrading fibronectin and attenuating the Wnt/ß-catenin signaling pathway.

The process of preadipocytes differentiation plays a vital role in adipose tissue expansion and many factors are involved in this event. Cathepsin B (CTSB), secreted from lysosome, has been reported in regulating a variety of physiological processes. In this study, we demonstrated CTSB promotes lipid accumulation and adipogenic genes expression in porcine primary preadipocytes by degrading fibronectin (Fn), a key component of extracellular matrix. Lithium chloride (LiCl) is an activator of Wnt/ß-catenin signaling through stabilizing ß-catenin. We found that CTSB can relieve the anti-adipogenic effects of LiCl, indicating that CTSB could impact Wnt/ß-catenin signaling pathway. Interestingly, Fn is an important target gene of Wnt/ß-catenin. So we considered that CTSB promote preadipocytes differentiation by suppressing these two pathways.

MiR-15a/b promote adipogenesis in porcine pre-adipocyte via repressing FoxO1.

Diabetes and many other metabolism syndromes are closely related to obesity. To reveal the underlying mechanism of fat deposition, an increasing number of studies are focusing on the functions of miRNAs during adipocytes development. Previous studies have proved that miR-15a/b play important roles in multiple physiological processes; however, their functions during adipogenesis remain unclear. To reveal this, we detected the expression profiles of miR-15a/b during adipogenesis in porcine pre-adipocyte, and found that their expression levels increased in the early stage of adipocyte differentiation and dropped after day 4. Moreover, over-expression of miR-15a/b in porcine pre-adipocytes promoted adipocyte differentiation and lipid accumulation. Target genes of miR-15a/b were predicted and examined, which revealed that Forkhead box protein O1 (FoxO1) is the target gene of miR-15a/b. The inhibition of FoxO1 expression level caused by miR-15a/b over-expression had a positive effect on adipogenesis. Thus, we conclude that miR-15a/b promote adipogenesis in porcine pre-adipocyte via repressing FoxO1.

[BAMBI inhibits porcine preadipocyte differentiation by facilitating ERK1/2 phosphorylation].

To study the role of BAMBI in adipogenesis, we constructed lentivirus interfering vector targeting on porcine BAMBI, packaged and infected the porcine preadipocyte. The differentiation state of preadipocyte was detected by Oil Red O staining and Oil Red O extraction assay and the expression levels of adipogenic marker genes were detected by Real-time qPCR and Werstern bloting. Results show that BAMBI expression was significant decreased after lentivirus infection, which was repressed more than 60% by shRNA2. Moreover, knockdown BAMBI increased the lipid accumulation of porcine preadipocyte and improved the expression of PPARγ (peroxisome proliferator-activated receptorγ) and ap2 (adipocyte protein 2). In summary, these data indicated that BAMBI inhibited adipocyte differentiation by facilitating the phosphorylation of ERK1/2.

[Over-expressed MiR-103 promotes porcine adipocyte differentiation].

To clarify the function of miR-103 in the differentiation of porcine preadipocyte, we carried out real-time PCR to detect the expression pattern of miR-103 during adipogenesis, and clarified its expression tendency through cell differentiation. Then we used adenovirus that overexpressed miR-103 to infect porcine preadipocyte. Subsequently, mRNA and protein expression of adipogenesis marker--PPARgamma and aP2 was analyzed by real-time PCR and Western blotting. At last, Oil-Red O staining was used to detect lipids accumulation in the 8th day after adipogenic inducement. The expression of miR-103 increased during adipocyte differentiation; compared with the control, the preadipocyte infected by pAd-miR-103 had an elevated expression level of adipocyte marker gene PPARgamma, aP2, and obvious lipid droplet was seen in the 8th day after adipogenic inducement. These results showed that miR-103 can enhance adipogenesis in primary cultured porcine adipocytes.