Department of Pharmacology and Toxicology, Virginia Commonwealth University, School of Medicine, Richmond, VA 23298, United States of America.
Department of Pharmacology and Toxicology, Virginia Commonwealth University, School of Medicine, Richmond, VA 23298, United States of America.
Biochim Biophys Acta Mol Cell Biol Lipids. 2018 Oct;1863(10):1246-1256. doi: 10.1016/j.bbalip.2018.07.016. Epub 2018 Aug 1.
The pannexin-1 (Panx1) channel has been reported to mediate the release of ATP that is involved in local tissue inflammation, obesity, and many chronic degenerative diseases. It remains unknown whether Panx1 is present in podocytes and whether this channel in podocytes mediates ATP release leading to glomerular inflammation or fibrosis. To answer these questions, we first characterized the expression of Panx channels in podocytes. Among the three known pannexins, Panx1 was the most enriched in podocytes, either cultured or native in mouse glomeruli. Using a Port-a-Patch planar patch-clamp system, we recorded a large voltage-gated outward current through podocyte membrane under the Csin/Naout gradient. Substitution of gluconate or aspartate for chloride in the bath solution blocked voltage-gated outward currents and shifted the reversal potential of Panx1 currents to the right, indicating the anion permeability of this channel. Pharmacologically, the recorded voltage-gated outward currents were substantially attenuated by specific Panx1 channel inhibitors. Given the anti-inflammatory and intracellular ATP restorative effects of adiponectin, we tested whether this adipokine inhibits Panx1 channel activity to block ATP release. Adiponectin blocked Panx1 channel activity in podocytes. Mechanistically, inhibition of acid ceramidase (AC) remarkably enhanced Panx1 channel activity under control conditions and prevented the inhibition of Panx1 channel by adiponectin. Correspondingly, intracellular addition of AC products, sphingosine or sphingosine-1-phosphate (S1P), blocked Panx1 channel activity, while elevation of intracellular ceramide had no effect on Panx1 channel activity. These results suggest that adiponectin inhibits Panx1 channel activity in podocytes through activation of AC and associated elevation of intracellular S1P.
缝隙连接蛋白-1 (Panx1) 通道已被报道介导 ATP 的释放，ATP 参与局部组织炎症、肥胖和许多慢性退行性疾病。目前尚不清楚 Panx1 是否存在于足细胞中，以及该通道是否在足细胞中介导 ATP 的释放，从而导致肾小球炎症或纤维化。为了回答这些问题，我们首先鉴定了 Panx 通道在足细胞中的表达。在三种已知的缝隙连接蛋白中，Panx1 在培养的或天然的小鼠肾小球中的足细胞中最为丰富。我们使用 Port-a-Patch 平面膜片钳系统，在 Cs+/Na+外梯度下记录到通过足细胞膜的大电压门控外向电流。在浴液中用葡糖酸盐或天冬氨酸替代氯离子可阻断电压门控外向电流，并将 Panx1 电流的反转电位向右移动，表明该通道的阴离子通透性。药理学研究表明，记录到的电压门控外向电流被特定的 Panx1 通道抑制剂显著减弱。鉴于脂联素具有抗炎和恢复细胞内 ATP 的作用，我们测试了这种脂肪因子是否通过抑制 Panx1 通道活性来阻断 ATP 的释放。脂联素抑制了足细胞中的 Panx1 通道活性。机制上，在对照条件下，酸 ceramidase (AC) 的抑制显著增强了 Panx1 通道的活性，并阻止了脂联素对 Panx1 通道的抑制。相应地，细胞内添加 AC 产物神经酰胺、鞘氨醇或鞘氨醇-1-磷酸 (S1P) 可阻断 Panx1 通道活性，而细胞内神经酰胺的升高对 Panx1 通道活性没有影响。这些结果表明，脂联素通过激活 AC 并伴随细胞内 S1P 的升高来抑制足细胞中的 Panx1 通道活性。
