Published online Sep 15, 2004. doi: 10.3748/wjg.v10.i18.2697
Revised: September 23, 2003
Accepted: October 29, 2003
Published online: September 15, 2004
AIM: To explore the effects of endothelin-1 (ET-1) on hepatic stellate cells (HSCs) DNA uptake, DNA synthesis, collagen synthesis and secretion, inward whole-cell calcium concentration ([Ca2+]i) as well as the blocking effect of verapamil on ET-1-stimulated release of inward calcium (Ca2+) of HSC in vitro.
METHODS: Rat hepatic stellate cells (HSCs) were isolated and cultivated. 3H-TdR and 3H-proline incorporation used for testing DNA uptake and synthesis, collagen synthesis and secretion of HSCs cultured in vitro; Fluorescent calcium indicator Fura-2/AM was used to measure [Ca2+]i inward HSCs.
RESULTS: ET-1 at the concentration of 5 × 10-8 mol/L, caused significant increase both in HSC DNA synthesis (2247 ± 344 cpm, P < 0.05) and DNA uptake (P < 0.05) when compared with the control group. ET-1 could also increase collagen synthesis (P < 0.05 vs control group) and collagen secretion (P < 0.05 vs control group). Besides, inward HSC [Ca2+] i reached a peak concentration (422 ± 98 mol/L, P < 0.001) at 2 min and then went down slowly to165 ± 51 mol/L (P < 0.01) at 25 min from resting state (39 ± 4 mol/L) after treated with ET-1. Verapamil (5 mol/L) blocked ET-1-activated [Ca2+]i inward HSCs compared with control group (P < 0.05). Fura-2/AM loaded HSC was suspended in no Ca2+ buffer containing 1 mol/L EGTA, 5 min later, 10-8 mol/L of ET-1 was added, [Ca2+]i inward HSCs rose from resting state to peak 399 ± 123 mol/L, then began to come down by the time of 20 min. It could also raise [Ca2+]i inward HSCs even without Ca2+ in extracellular fluid, and had a remarkable dose-effect relationship (P < 0.05). Meanwhile, verapamil could restrain the action of ET-1 (P < 0.05).
CONCLUSION: Actions of ET-1 on collagen metabolism of HSCs may depend on the transportation of inward whole-cell calcium.