Becker Daniel, Blase Christopher, Bereiter-Hahn Juergen, Jendrach Marina
Kinematic Cell Research Group, Johann Wolfgang Goethe University, Marie-Curie-Str. 9, 60439 Frankfurt, Germany.
J Cell Sci. 2005 Jun 1;118(Pt 11):2435-40. doi: 10.1242/jcs.02372.
Tight regulation of the cell volume is important for the maintenance of cellular homeostasis. In a hypotonic environment, cells swell owing to osmosis. With many vertebrate cells, swelling is followed by an active reduction of volume, a process called regulatory volume decrease (RVD). A possible participant in RVD is the non-selective cation channel TRPV4, a member of the TRP superfamily that has been shown to react to hypotonic stimuli with a conductance for Ca2+. As a model for cell-volume regulation, we used a human keratinocyte cell line (HaCaT) that produces TRPV4 endogenously. When HaCaT cells were exposed to a hypotonic solution (200 mOsm) maximal swelling was followed by RVD. During swelling and volume regulation, a strong Ca2+ influx was measured. Gd3+, an inhibitor of TRPV4, blocked RVD of HaCaT cells and the accompanying rise of cytosolic Ca2+. To define the role of TRPV4 in volume regulation, a TRPV4-EGFP fusion protein was produced in CHO cells. CHO cells are unable to undergo RVD under hypotonic conditions and do not produce TRPV4 endogenously. Fluorescence imaging revealed that recombinant TRPV4 was localized to the cell membrane. Production of TRPV4 enabled CHO cells to undergo typical RVD after hypo-osmolarity-induced cell swelling. RVD of TRPV4-transfected CHO cells was significantly reduced by Gd3+ treatment or in Ca2+-free solution. Taken together, these results show a direct participation of TRPV4 in RVD.
细胞体积的严格调控对于维持细胞内环境稳定至关重要。在低渗环境中，细胞因渗透作用而膨胀。对于许多脊椎动物细胞而言，膨胀之后会出现主动的体积减小，这一过程称为调节性容积减小（RVD）。RVD的一个可能参与者是TRP超家族成员、非选择性阳离子通道TRPV4，该通道已被证明可通过对Ca2+的电导对低渗刺激做出反应。作为细胞体积调节的模型，我们使用了一种内源性产生TRPV4的人角质形成细胞系（HaCaT）。当HaCaT细胞暴露于低渗溶液（200 mOsm）时，最大程度的膨胀之后会出现RVD。在膨胀和体积调节过程中，检测到强烈的Ca2+内流。TRPV4抑制剂Gd3+可阻断HaCaT细胞的RVD以及伴随的胞质Ca2+升高。为了确定TRPV4在体积调节中的作用，在CHO细胞中产生了TRPV4-EGFP融合蛋白。CHO细胞在低渗条件下无法进行RVD，且内源性不产生TRPV4。荧光成像显示重组TRPV4定位于细胞膜。TRPV4的产生使CHO细胞在低渗诱导的细胞肿胀后能够进行典型的RVD。用Gd3+处理或在无Ca2+溶液中，TRPV4转染CHO细胞的RVD显著降低。综上所述，这些结果表明TRPV4直接参与了RVD。
