Figure 4.

Effects of suramin on ATP release under several different conditions. (A,C,E) NRK (A,C) or EGFP-Cx43 LLC-PK1 cells (E) were pretreated with 300 μM suramin, 3 mM heptanol or 100 μM lindane for 20 min, and exposed to HBSS (A), glucose-deprived medium (C), or 50 μM Cd²⁺ (E) for an additional 30 min (A,B) or 3 h (E). Cell supernatants were collected and quantitated for ATP concentration. Results are expressed as relative light unit (RLU; mean ± SE, n = 3). *P < 0.05, #P < 0.01 compared with respective control. (B,D) Down-regulation of Cx43 with specific siRNA on HBSS and glucose deprivation-triggered ATP release. NRK cells were treated with either control siRNA or siRNA against Cx43 for 48 h. After that, they were exposed to HBSS or glucose-free medium for 30 min. The cellular lysates were also subjected to Western blot analysis of Cx43 to verify the effectiveness of Cx43 siRNA in down-regulation of Cx43 (Figure 4B, insert). Note the obvious reduced level of Cx43 in Cx43 siRNA-treated cells. Results are expressed as RLU (mean ± SE, n = 3). (F) Effect of suramin on basal ATP release triggered by medium exchange. NRK cells cultured in normal Ca²⁺ medium were incubated with or without 300 μM suramin for 20 min. Thereafter, culture medium were either left untouched (control) or changed to the same normal Ca²⁺ medium or Ca²⁺-free medium for additional 5 min. Cell supernatants were collected and quantitated for ATP concentration. Results are expressed as RLU (mean ± SE, n = 6). #P < 0.01.