RESUMEN
This paper is to report the exploration of the activation of Rho/ROCK signal pathway in 5-HT-induced proliferation of rat pulmonary artery smooth muscle cells (PASMCs) and the inhibitory effect of m-Nis on this pathway. PASMCs were cultured with the explant technique. MTT assay was used to explore the proliferation of PASMCs after 5-HT treated for different time and the intervening effect of m-Nis. RT-PCR and Western blot were used respectively to explore the mRNA expression of RhoA, ROCK1 and the protein expression of p-MYPT1 in 5-HT-treated PASMCs and intervening effect of m-Nis. The results of MTT assay suggested that 5-HT (1 µmol · L(-1)) treatment for 12-72 h significantly induced the proliferation of rat PASMCs (P<0.05 or P < 0.01), which were inhibited by m-Nis (1 x 10(-5), 1 x 10(-6), l x 10(-7), 1 x10(-8) mol · L(-1)) in dose-dependent manners (P < 0.05 or P < 0.01). Similarly, the mRNA expression of RhoA, ROCK1 and the protein expression of p-MYPT1 were also inhibited by m-Nis in different degrees (P < 0.05 or P < 0.01). Thus, the results of this study suggested that Rho/ROCK pathway played an important role in 5-HT-induced proliferation of rat PASMCs, m-Nis inhibited 5-HT-induced proliferation obviously, which may be related to the blockage of Rho/ROCK signal pathway.
RESUMEN
Aim To explore the changes of Apelin/APJ system in LPS-induced injury of rat pulmonary mi-crovascular endothelial cells( PMVECs) , and the effect and mechanism of Apelin. Methods PMVECs were cultured with the explant technique, and the identifica-tion of rat PMVECs was carried out by immunocyto-chemical staining of factorⅧrelated antigen. MTT as-say was used to evaluate the viability of PMVECs. The mRNA expression of Apelin and APJ was detected by RT-PCR. The protein expression of PCNA and the phosphorylation of Akt was analyzed by Western blot. Results The mRNA expression of Apelin and APJ showed a compensatory increase after LPS treatment for a short period of time ( P<0. 01 ) , but with the exten-sion of time, which was significantly inhibited, even lower than the control group ( P<0. 05 or P<0. 01 ) , suggesting that Apelin/ APJ system might be involved in LPS-induced PMVECs injury. MTT results showed that 10 -6 ~10 -9 mol · L-1 Apelin obviously promoted the proliferation of rat PMVECs ( P <0. 05 or P <0. 01 ) , and with certain concentration and time de-pendence. Moreover, Apelin also improved the LPS-induced PMVECs injury in different degrees ( P<0. 05 or P < 0. 01 ) . In addition, Western blot analysis showed that Apelin significantly reversed the decrease of the protein expression of PCNA and the Akt phos-phorylation level induced by LPS ( P <0. 05 or P <0. 01 ) . Conclusions The Apelin/APJ system is in-volved in LPS-induced PMVECs injury. Apelin plays an important role in protecting the pulmonary microvas-cular endothelial function and reversing the LPS-in-duced PMVECs injury, which might be related to the activation of Akt phosphorylation pathway.
RESUMEN
This study is to explore the activation of the Ca2+/CaM/CaN signal pathway in 5-HT-induced proliferation of rat pulmonary artery smooth muscle cells (PASMCs) and the inhibitory effect of m-nisoldipine (m-Nis) on this pathway. PASMCs were cultured with the explant technique. The proliferation of PASMCs was evaluated by MTT assay. Confocal microscopy was used to measure the change of [Ca2+]i. The mRNA expression of CaM and CaN was evaluated by RT-PCR and the activity of CaN was measured according to the instruction of kits. The results of MTT assay suggested that 5-HT (1 micromol x L(-1)) significantly induced the proliferation of rat PASMCs (P < 0.01), which was inhibited obviously by m-Nis (P < 0.05 or P < 0.01). Similarly, m-Nis inhibited 5-HT-induced elevation of [Ca2+]i (P < 0.01). The mRNA expression of CaM, CaN and the activation of CaN were also inhibited by m-Nis at different degrees (P < 0.05 or P < 0.01). Thus, the results of this study suggested that Ca2+/CaM/CaN signal pathway played an important role in 5-HT-induced proliferation of rat PASMCs, the inhibition of m-Nis on proliferation of rat PASMCs may be related to the blockage of Ca2+/CaM/CaN signal pathway by inhibiting the elevation of [Ca2+]i.
RESUMEN
This study is to explore the activation of the Ca2+/CaM/CaN signal pathway in 5-HT-induced proliferation of rat pulmonary artery smooth muscle cells (PASMCs) and the inhibitory effect of m-nisoldipine (m-Nis) on this pathway. PASMCs were cultured with the explant technique. The proliferation of PASMCs was evaluated by MTT assay. Confocal microscopy was used to measure the change of [Ca2+]i. The mRNA expression of CaM and CaN was evaluated by RT-PCR and the activity of CaN was measured according to the instruction of kits. The results of MTT assay suggested that 5-HT (1 ?mol?L-1) significantly induced the proliferation of rat PASMCs (P
RESUMEN
Aim To explore the effect of m-Nisoldipine(m-Nis) on 5-HT-induced proliferation,migration of rat PASMCs and to study the mechanisms.Methods PASMCs were cultured with the explant technique,and were divided into 6 groups:control group,5-HT(1 μmol·L~(-1)) group and m-Nis(10~(-5),10~(-6),10~(-7),10~(-8) mol·L~(-1))group.MTT assay was used to evaluate the proliferation of PASMCs,and transwell chambers were used to detect the migration of PASMCs.In addition,the expression of PCNA and the phosphorylation of ERK1/2 were evaluated by Western blot analysis.Results m-Nis inhibited the proliferation(P<0.05 or P<0.01)and migration(P<0.01)of rat PASMCs induced by 5-HT obviously.Similarly,Western blot analysis of PCNA indicated that the expression of PCNA was significantly higher in 5-HT group than that in control group(P<0.01).Whereas,in four m-Nis treated groups,the level of PCNA was markedly decreased(P<0.05 or P<0.01).Meanwhile,m-Nis 10~(-5),10~(-6) and 10~(-7) mol·L~(-1) pretreatment also reduced 5-HT-induced phosphorylation of ERK1/2 obviously(P<0.05 or P<0.01).Conclusion m-Nis inhibits 5-HT-induced proliferation and migration of rat PASMCs obviously,which may be related to the inhibition of PCNA expression and the blockage of ERK1/2/MAPK signal pathway.