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 ([Ca^2+]i) as well as the blocking...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 ([Ca^2+]i) as well as the blocking effect of verapamil on ET-l-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 [Ca^2+]i inwardHSCs.RESULTS: ET-1 at the concentration of 5×10^-8 mol/L,caused significant increase both in HSC DNA synthesis (2 247+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 vscontrol group). Besides, inward HSC [Ca^2+]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 [Ca^2+li inward HSCs compared with control group(P<0.05). Fura-2/AM loaded HSC was suspended in no Ca^2+ buffer containing 1 mol/L EGTA, 5 rain later, 10^-8 mol/L of ET-1 was added, [Ca^2+]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 [Ca^2+]i inward HSCs even without Ca^2+ 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 wholecell calcium.展开更多
基金Supported by the Grant for Nature and Science from Shanghai,No.03ZR 14097
文摘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 ([Ca^2+]i) as well as the blocking effect of verapamil on ET-l-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 [Ca^2+]i inwardHSCs.RESULTS: ET-1 at the concentration of 5×10^-8 mol/L,caused significant increase both in HSC DNA synthesis (2 247+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 vscontrol group). Besides, inward HSC [Ca^2+]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 [Ca^2+li inward HSCs compared with control group(P<0.05). Fura-2/AM loaded HSC was suspended in no Ca^2+ buffer containing 1 mol/L EGTA, 5 rain later, 10^-8 mol/L of ET-1 was added, [Ca^2+]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 [Ca^2+]i inward HSCs even without Ca^2+ 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 wholecell calcium.
