AIMS: Triacylglycerol hydrolase (TGH) plays an important role in intracellular lipid metabolism. However, most previous studies on TGH were focused on rodent animals. Pig is considered as one of the best models for studying obesity, diabetes, and other lipid metabolism related diseases. So far, we do not know whether TGH is to play a role in lipolysis in porcine primary adipocyte through stimulation by hormones as Hormone-sensitive lipase (HSL). The objective of this study is to explore the mechanisms that regulate TGH expression in porcine adipocyte and its role in fasting-induced and Isoproterenol-stimulated lipolysis in porcine primary adipocytes. MATERIALS AND METHODS: Stromal-vascular cells containing preadipocytes were isolated from cervical and dorsal subcutaneous adipose tissues of approximately 3-day-old Chinese male piglets. After confluence, the differentiation was induced by Insulin, hydrocortisone, and transferrin. The primary adipocytes were cultured in various concentration (0.1 to 200 micromol/l) Isoproterenol for 0-12 h or serum-free medium after differentiation. The glycerol release and triacylglycerol (TG) contents were detected when the cells were collected. Then, expression of TGH and HSL mRNA and their protein were determined by RT-PCR, Western Blot, and lipolytic analysis. RESULTS: Both TGH mRNA and protein were not detected on day 0, but as differentiation was induced, their expression displayed a great increase. The expression of TGH mRNA showed the highest level on day 4, but its protein reached the highest level on day 6 and began to fall down from day 8. The expression of TGH mRNA and proteins was increased in serum-free media, and mRNA expression was decreased while changed into complete media again. The glycerol release increased significantly when the cells were cultured with serum-free media. The lower concentration of Isoproterenol (0.1 and 1 micromol/l) did not affect the expression of TGH and HSL, but higher concentration (10, 100, and 200 micromol/l) could greatly up-regulate HSL expression but did not affect TGH level. Also, the higher concentration of Isoproterenol could increase the glycerol release and decrease TG content in dose-dependent manner. CONCLUSIONS: These results suggested that TGH expression is differentiation-dependent in porcine primary adipocytes and TGH plays a role in fasting-induced lipolysis not hormone-stimulated lipolysis.
AIMS: Triacylglycerol hydrolase (TGH) plays an important role in intracellular lipid metabolism. However, most previous studies on TGH were focused on rodent animals. Pig is considered as one of the best models for studying obesity, diabetes, and other lipid metabolism related diseases. So far, we do not know whether TGH is to play a role in lipolysis in porcine primary adipocyte through stimulation by hormones as Hormone-sensitive lipase (HSL). The objective of this study is to explore the mechanisms that regulate TGH expression in porcine adipocyte and its role in fasting-induced and Isoproterenol-stimulated lipolysis in porcine primary adipocytes. MATERIALS AND METHODS: Stromal-vascular cells containing preadipocytes were isolated from cervical and dorsal subcutaneous adipose tissues of approximately 3-day-old Chinese male piglets. After confluence, the differentiation was induced by Insulin, hydrocortisone, and transferrin. The primary adipocytes were cultured in various concentration (0.1 to 200 micromol/l) Isoproterenol for 0-12 h or serum-free medium after differentiation. The glycerol release and triacylglycerol (TG) contents were detected when the cells were collected. Then, expression of TGH and HSL mRNA and their protein were determined by RT-PCR, Western Blot, and lipolytic analysis. RESULTS: Both TGH mRNA and protein were not detected on day 0, but as differentiation was induced, their expression displayed a great increase. The expression of TGH mRNA showed the highest level on day 4, but its protein reached the highest level on day 6 and began to fall down from day 8. The expression of TGH mRNA and proteins was increased in serum-free media, and mRNA expression was decreased while changed into complete media again. The glycerol release increased significantly when the cells were cultured with serum-free media. The lower concentration of Isoproterenol (0.1 and 1 micromol/l) did not affect the expression of TGH and HSL, but higher concentration (10, 100, and 200 micromol/l) could greatly up-regulate HSL expression but did not affect TGH level. Also, the higher concentration of Isoproterenol could increase the glycerol release and decrease TG content in dose-dependent manner. CONCLUSIONS: These results suggested that TGH expression is differentiation-dependent in porcine primary adipocytes and TGH plays a role in fasting-induced lipolysis not hormone-stimulated lipolysis.
Authors: Christopher M Jenkins; David J Mancuso; Wei Yan; Harold F Sims; Beverly Gibson; Richard W Gross Journal: J Biol Chem Date: 2004-09-10 Impact factor: 5.157
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