摘要
AIM: Many growth factors, such as epidermal growth factor(EGF), are associated with the carcinogenesis. EGF plays itsrole in the proliferation of hepatoma cells through bindingwith EGF receptor (EGFR) and a series of signal transduction.But the postreceptor pathway is still not clear. In the presentexperiment, we studied the effect of tyrosine kinase, proteinkinase C, Na+/H+ exchange, calmodulin and voltage-dependent Ca2+ channel on EGF-induced hepatoma cellproliferation.METHODS: Hepatoma cell line SMMC7721 was cultured inRPMI1640 serum-free medium. In order to study the effectof thyrosine kinase, protein kinase C, Na+/H+ exchange,calmodulin and voltage-dependent Ca2+ channel on humanheptoma cell proliferation induced by epidermal growth factor(EGF), DNA synthesis rate of hepatoma cells was measuredby the method of 3H-TdR incorporation.RESULTS: EGF (10-9 M) stimulated the proliferation of heptomacells significantly (3H-TdR incorporation was 1 880+281 cpm/well, P＜0.05), and this effect was significantly inhibited bytyrosine kinase inhibitor genistein (3H-TdR incorporation was808±209 cpm/well, P＜0.001). Calmedulin inhibitor W-7, proteinkinase C inhibitor H-7 and Na+/H+ exchange inhibitor amilorideindividually had significant inhibiting effect on EGF-inducedproliferation of hepatoma cells (3H-TdR incorporation was978±87.3 cpm/well, 1 241+147 cpm/well, 1 380+189 cpm/well, respectivly, P＜0.001, P＜0.01, P＜0.05), but they allhad no effect on the basal level proliferation of culturedhepatoma cells (3H-TdR incorporation was 1 284+260 cpm/well, 1 179+150 cpm/well, 1 392+152 cpm/well, respectivly,3H-TdR incorporation of the control was 1353+175 cpm/well, P＞0.05). Voltage-dependent Ca2+ channel inhibitorverapamil had no inhibition on EGF-induced proliferation ofhepatoma cells (3H-TdR incorporation was 1 637+133 cpm/well, P＞0.05), it also had no effect on the basal levelproliferation of cultured hepatoma cells (3H-TdR incorporationwas 1196+112 cpm/well,P＞0.05).CONCLUSION: Our data suggest that tyrosine kinase, Ca2+-calmodulin-dependent pathway, protein kinase C and Na+/H+ exchange play a critical role in EGF-induced proliferationof hepatoma cells and that the effect of EGF is independentof voltage-dependent Ca2+ channel.
AIM:Many growth factors,such as epidermal growth factor (EGF),are associated with the carcinogenesis.EGF plays its role in the proliferation of hepatoma ceils through binding with EGF receptor(EGFR)and a series of signal transduction. But the postreceptor pathway is still not clear.In the present experiment,we studied the effect of tyrosine kinase,protein kinase C,Na^+/H^+ exchange,calmodulin and voltage- dependent Ca^(2+)channel on EGF-induced hepatoma cell proliferation. METHODS:Hepatoma cell line SMMC7721 was cultured in RPMI1640 serum-free medium.In order to study the effect of thyrosine kinase,protein kinase C,Na^+/H^+ exchange, calmodulin and voltage-dependent Ca^(2+)channel on human heptoma cell proliferation induced by epidermal growth factor (EGF),DNA synthesis rate of hepatoma cells was measured by the method of ~3H-TdR incorporation. RESULTS:EGF(10^(-9)M)stimulated the proliferation of heptoma cells significantly(~3H-TdR incorporation was 1 880±281 cpm/ well,P<0.05),and this effect was significantly inhibited by tyrosine kinase inhibitor genistein(~3H-TdR incorporation was 808±209 cpm/well,P<0.001).Calmodulin inhibitor W-7,protein kinase C inhibitor H-7 and Na^+/H^+ exchange inhibitor amiloride individually had significant inhibiting effect on EGF-induced proliferation of hepatoma cells(~3H-TdR incorporation was 978±87.3 cpm/well,1 241±147 cpm/well,1 380±189 cpm/ well,respectivly,P<0.001,P<0.01,P<0.05),but they all had no effect on the basal level proliferation of cultured hepatoma cells(~3H-TdR incorporation was 1 284±260 cpm/ well,1 179±150 cpm/well,1 392±152 cpm/well,respectivly, ~3H-TdR incorporation of the control was 1 353±175 cpm/ well,P>0.05).Voltage-dependent Ca^(2+)channel inhibitor verapamil had no inhibition on EGF-induced proliferation of hepatoma cells(~3H-TdR incorporation was 1 637±133 cpm/ well,P>0.05),it also had no effect on the basal level proliferation of cultured hepatoma cells(~3H-TdR incorporation was 1 196±112 cpm/well,P>0.05). CONCLUSION:Our data suggest that tyrosine kinase,Ca^(2+)- calmodulin-dependent pathway,protein kinase C and Na^+/ H^+ exchange play a critical role in EGF-induced proliferation of hepatoma cells and that the effect of EGF is independent of voltage-dependent Ca^(2+)channel.
