Figure 4.

TRPV4 mediates mechanically evoked changes in [Ca²⁺]_i. (a): western blot analyses of HCD cell lysates (5 μg protein/lane) using an antibody against human TRPV4 (120 kDa) confirmed siRNA knock-down of TRPV4 expression in lane 4, as compared to control cells (lane 1, untransfected cells; lane 2, cells transfected with lipid alone; lane 3, cells transfected with scrambled siRNA). Knock-down reduced TRPV4 expression to approximately 60% of control. Results represent mean ± SEM; n = 3; **P < 0.01. In (b), a single cell within a cluster of HCD-cells can be identified following co-transfection with anti-TRPV4 siRNA and Red Fluorescent Protein (RFP) using the siLentGene U6 Cassette RNA Interference System (Promega; (b) C). The same cell cluster is visualised as an overlay image following loading with fura-2/AM ((b) D). In (b) F, a nontransfected cell (cell 1) is stimulated mechanically to evoke a rise in [Ca²⁺]_i ((b) H). The RFP-identified cell 2 ((b) D) fails to evoke a touch-evoked Ca²⁺-signal ((b) G).