Institute of Biomedical and Clinical Science, Peninsula College of Medicine and Dentistry, University of Plymouth, Plymouth, Devon, UK.
Islets. 2010 Nov-Dec;2(6):374-82. doi: 10.4161/isl.2.6.13917. Epub 2010 Nov 1.
This study has assessed the expression and functional significance of cGMP-dependent signalling components in BRIN-BD11 cells. RT-PCR analysis revealed the expression of two subunits of soluble guanylate cyclase (sGC) suggesting the presence of an α2/β1 heterodimer. The expression of three particulate guanylate cyclases (pGC) was also detected (GC-A, GC-B and GC-C), as well as two cGMP-selective PDE isoforms (PDE5A and PDE9). Stimulation of BRIN-BD11 cells with agonists selective for sGC (NO, YC-1 and BAY 41-2272), GC-A (atrial natriuretic peptide (ANP) or GC-C (guanylin) caused an elevation in cGMP, and in the case of sGC, this was blocked by the selective inhibitor 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ). The stimulatory effects of each activator on cGMP levels were further potentiated by the PDE5A inhibitor, zaprinast. Treatment of cells with sGC activators induced a loss of viability and increased insulin secretion. However these effects were not attenuated by ODQ suggesting that they were independent of a rise in cGMP. A modest increase in β-cell death and insulin secretion were also observed in guanylin and ANP treated cells, although the latter only reduced cell viability in the presence of a PDE5A inhibitor. Taken together, the data reveal that BRIN-BD11 cells express several functionally active enzymes capable of modulating cGMP levels, and they imply that signalling through these proteins may impact upon β-cell viability. The results further suggest that pGC isozymes can also regulate insulin secretion but that the pool of cGMP controlling insulin release is small relative to the global cGMP concentration in the cell.
本研究评估了 cGMP 依赖性信号成分在 BRIN-BD11 细胞中的表达和功能意义。RT-PCR 分析显示两种可溶性鸟苷酸环化酶 (sGC) 的亚基表达，表明存在 α2/β1 异二聚体。还检测到三种颗粒型鸟苷酸环化酶 (pGC) 的表达（GC-A、GC-B 和 GC-C），以及两种 cGMP 选择性 PDE 同工酶（PDE5A 和 PDE9）。BRIN-BD11 细胞用 sGC（NO、YC-1 和 BAY 41-2272）、GC-A（心钠素 (ANP) 或 GC-C（ guanylin）的激动剂刺激，导致 cGMP 升高，而在 sGC 的情况下，这被选择性抑制剂 1H-[1,2,4]恶二唑-[4,3-a]喹喔啉-1-酮 (ODQ) 阻断。PDE5A 抑制剂 zaprinast 进一步增强了每种激活剂对 cGMP 水平的刺激作用。sGC 激活剂处理细胞会导致细胞活力丧失和胰岛素分泌增加。然而，ODQ 并没有减弱这些作用，表明它们与 cGMP 的升高无关。在 guanylin 和 ANP 处理的细胞中也观察到 β 细胞死亡和胰岛素分泌适度增加，尽管后者仅在存在 PDE5A 抑制剂时才降低细胞活力。总之，这些数据表明 BRIN-BD11 细胞表达几种能够调节 cGMP 水平的功能活性酶，并且它们暗示这些蛋白质的信号传导可能会影响 β 细胞活力。结果还表明 pGC 同工酶也可以调节胰岛素分泌，但控制胰岛素释放的 cGMP 池相对于细胞中的总 cGMP 浓度较小。
