Laboratory of Veterinary Pathology, College of Veterinary Medicine and Research Institute for Veterinary Science, Seoul National University, Seoul, Korea.
Acta Pharmacol Sin. 2013 Jan;34(1):129-36. doi: 10.1038/aps.2012.142. Epub 2012 Dec 10.
To determine the presence of voltage-gated K(+) (Kv) channels in bone marrow-derived human mesenchymal stem cells (hMSCs) and their impact on differentiation of hMSCs into adipocytes.
For adipogenic differentiation, hMSCs were cultured in adipogenic medium for 22 d. The degrees of adipogenic differentiation were examined using Western blot, Oil Red O staining and Alamar assay. The expression levels of Kv channel subunits Kv1.1, Kv1.2, Kv1.3, Kv1.4, Kv2.1, Kv3.1, Kv3.3, Kv4.2, Kv4.3, and Kv9.3 in the cells were detected using RT-PCR and Western blot analysis.
The expression levels of Kv2.1 and Kv3.3 subunits were markedly increased on d 16 and 22. In contrast, the expression levels of other Kv channel subunits, including Kv1.1, Kv1.2, Kv1.3, Kv1.4, Kv4.2, Kv4.3, and Kv9.3, were decreased as undifferentiated hMSCs differentiated into adipocytes. Addition of the Kv channel blocker tetraethylammonium (TEA, 10 mmol/L) into the adipogenic medium for 6 or 12 d caused a significant decrease, although not complete, in lipid droplet formation and adipocyte fatty acid-binding protein 2 (aP(2)) expressions. Addition of the selective Kv2.1 channel blocker guangxitoxin (GxTX-1, 40 nmol/L) into the adipogenic medium for 21 d also suppressed adipogenic differentiation of the cells.
The results demonstrate that subsets of Kv channels including Kv2.1 and Kv3.3 may play an important role in the differentiation of hMSCs into adipocytes.
确定电压门控钾通道(Kv)在骨髓间充质干细胞(hMSCs)中的存在及其对 hMSCs 向脂肪细胞分化的影响。
为了进行成脂分化,hMSCs 在成脂培养基中培养 22 天。使用 Western blot、油红 O 染色和 Alamar 测定法检查成脂分化程度。使用 RT-PCR 和 Western blot 分析检测细胞中 Kv 通道亚基 Kv1.1、Kv1.2、Kv1.3、Kv1.4、Kv2.1、Kv3.1、Kv3.3、Kv4.2、Kv4.3 和 Kv9.3 的表达水平。
Kv2.1 和 Kv3.3 亚基的表达水平在第 16 天和第 22 天显著增加。相比之下,在未分化的 hMSCs 分化为脂肪细胞的过程中,其他 Kv 通道亚基(包括 Kv1.1、Kv1.2、Kv1.3、Kv1.4、Kv4.2、Kv4.3 和 Kv9.3)的表达水平降低。将 Kv 通道阻断剂四乙铵(TEA,10 mmol/L)添加到成脂培养基中 6 或 12 天会导致脂滴形成和脂肪细胞脂肪酸结合蛋白 2(aP(2))表达显著减少,但不完全。将选择性 Kv2.1 通道阻断剂 Guangxitoxin(GxTX-1,40 nmol/L)添加到成脂培养基中 21 天也会抑制细胞的成脂分化。
结果表明,包括 Kv2.1 和 Kv3.3 在内的 Kv 通道亚基可能在 hMSCs 向脂肪细胞分化中发挥重要作用。