Department of Physiology, University of Arizona, Tucson, Arizona, USA.
Department of Ophthalmology and Vision Science, University of Arizona, Tucson, Arizona, USA.
J Cell Physiol. 2024 Nov;239(11):e31369. doi: 10.1002/jcp.31369. Epub 2024 Jul 16.
Previously we showed hyperosmotic solution caused TRPV1-dependent NKCC1 activation in the lens by a mechanism that involved ERK1/2 signaling. In various tissues, integrins and the cytoskeletal network play a role in responses to osmotic stress. Here, we examined the association between integrins and TRPV1-dependent activation of NKCC1 in mouse lens epithelium. Wild-type (WT) lenses exposed to the integrin agonist leukadherin-1 (LA-1) for 10 min displayed a ~33% increase in the bumetanide-sensitive rate of Rb uptake indicating NKCC activation. Paclitaxel, a microtubule stabilizing agent, abolished the Rb uptake response. In primary cultured lens epithelium LA-1 caused a robust ERK1/2 activation response that was almost fully suppressed by paclitaxel. The TRPV1 agonist capsaicin caused a similar ERK1/2 activation response. Consistent with an association between integrins and TRPV1, the TRPV1 antagonist A889425 prevented the Rb uptake response to LA-1 as did the ERK inhibitor U0126. LA-1 did not increase Rb uptake by lenses from TRPV1 knockout mice. In cells exposed to a hyperosmotic stimulus, both the ERK1/2 activation and Rb uptake responses were prevented by paclitaxel. Taken together, the findings suggest TRPV1 activation is associated with integrins and the tubulin cytoskeleton. This aligned with the observation that LA-1 elicited a robust cytoplasmic calcium rise in cells from WT lenses but failed to increase calcium in cells from TRPV1 knockout lenses. The results are consistent with the notion that integrin activation by LA-1, or a hyperosmotic stimulus, causes TRPV1 channel opening and the consequent downstream activation of the ERK1/2 and NKCC1 responses.
先前我们表明，渗透压溶液通过涉及 ERK1/2 信号转导的机制引起晶状体中 TRPV1 依赖性 NKCC1 的激活。在各种组织中，整合素和细胞骨架网络在应对渗透胁迫中发挥作用。在这里，我们检查了整合素与 TRPV1 依赖性 NKCC1 在小鼠晶状体上皮细胞中的激活之间的关联。用整合素激动剂白细胞粘附素-1（LA-1）处理 10 分钟的野生型（WT）晶状体显示出布美他尼敏感的 Rb 摄取率增加约 33%，表明 NKCC 激活。紫杉醇，一种微管稳定剂，消除了 Rb 摄取反应。在原代培养的晶状体上皮细胞中，LA-1 引起强烈的 ERK1/2 激活反应，该反应几乎被紫杉醇完全抑制。TRPV1 激动剂辣椒素引起类似的 ERK1/2 激活反应。与整合素和 TRPV1 之间的关联一致，TRPV1 拮抗剂 A889425 以及 ERK 抑制剂 U0126 均可防止 LA-1 引起的 Rb 摄取反应。LA-1 不会增加 TRPV1 敲除小鼠晶状体的 Rb 摄取。在暴露于高渗刺激的细胞中，紫杉醇可防止 ERK1/2 激活和 Rb 摄取反应。综上所述，这些发现表明 TRPV1 的激活与整合素和微管细胞骨架有关。这与 LA-1 引发 WT 晶状体细胞中强烈的细胞质钙升高而未能增加 TRPV1 敲除细胞中钙的观察结果一致。这些结果与以下观点一致，即 LA-1 的整合素激活或高渗刺激引起 TRPV1 通道开放，从而导致 ERK1/2 和 NKCC1 反应的下游激活。
