OBJECTIVE: To observe the effect of Qibai Pingfei capsule(QBPF) medicated serum on the proliferation of rat pulmonary arterial smooth muscle cells(PASMCs) under hypoxia conditions and to investigate its key molecular ...OBJECTIVE: To observe the effect of Qibai Pingfei capsule(QBPF) medicated serum on the proliferation of rat pulmonary arterial smooth muscle cells(PASMCs) under hypoxia conditions and to investigate its key molecular effects on the Ca^(2+)/calcineurin/nuclear factor of activated T-cells 3(NFATc3) signaling pathway.METHODS: QBPF was provided to rats via continuous gavage for 10 days. Primary rat PASMCs were cultured using the direct adherent culture method.Methyl thiazolyl tetrazolium assay was used to eval-uate the effect of QBPF on PASMCs proliferation under hypoxia conditions. Laser scanning confocal microscopy was used to detect changes in intracellular free calcium([Ca^(2+)]_i) in PASMC-loaded Fluo-3-AM.Real-time quantitative polymerase chain reaction and western blot were used to detect the transcription and protein expression levels of calcineurin and NFATc3 genes in PASMCs.RESULTS: Compared with normoxia conditions,PASMCs proliferated at 12, 24, 48, and 72 h under hypoxia conditions. QBPF at concentrations of 5%,10%, and 20% could inhibit hypoxia-induced PASMC proliferation to different degrees. The inhibitory effect was most significant in the 20% QBPF group under 24 h hypoxia conditions. The concentration of [Ca^(2+)]_iin PASMCs under hypoxia was increased and [Ca^(2+)]_iwas significantly decreased when co-incubated with QBPF at 5%, 10%, and 20%. Compared with normoxia conditions, the m RNA and protein expression levels of calcineurin and NFATc3 in PASMCs induced by hypoxia were up-regulated.QBPF application significantly down-regulated m RNA and protein expression levels of calcineurin and NFATc3 in PASMCs.CONCLUSION: QBPF can effectively inhibit hypoxia-induced proliferation of PASMCs through down-regulation of key molecular expression via the Ca^(2+)/calcineurin/NFATc3 pathway.展开更多
基金Supported by the National Natural Science Foundation of China(Intervention Mechanism of Yiqi Huatan Quyu Recipe on Pulmonary Vascular Remodeling of Chronic Obstructive Pulmonary Disease Based on Ca~(2+)-Calcineurin-NFAT Signaling Pathway,No.81403308)the National Natural Science Foundation of China(Based on"Lung Qi Deficiency and Missing Management and Regulation"Discuss the Mechanism of Chronic Obstructive Pulmonary Disease Pulmonary Vasoconstriction and KATP Channel,No.81373598)
文摘OBJECTIVE: To observe the effect of Qibai Pingfei capsule(QBPF) medicated serum on the proliferation of rat pulmonary arterial smooth muscle cells(PASMCs) under hypoxia conditions and to investigate its key molecular effects on the Ca^(2+)/calcineurin/nuclear factor of activated T-cells 3(NFATc3) signaling pathway.METHODS: QBPF was provided to rats via continuous gavage for 10 days. Primary rat PASMCs were cultured using the direct adherent culture method.Methyl thiazolyl tetrazolium assay was used to eval-uate the effect of QBPF on PASMCs proliferation under hypoxia conditions. Laser scanning confocal microscopy was used to detect changes in intracellular free calcium([Ca^(2+)]_i) in PASMC-loaded Fluo-3-AM.Real-time quantitative polymerase chain reaction and western blot were used to detect the transcription and protein expression levels of calcineurin and NFATc3 genes in PASMCs.RESULTS: Compared with normoxia conditions,PASMCs proliferated at 12, 24, 48, and 72 h under hypoxia conditions. QBPF at concentrations of 5%,10%, and 20% could inhibit hypoxia-induced PASMC proliferation to different degrees. The inhibitory effect was most significant in the 20% QBPF group under 24 h hypoxia conditions. The concentration of [Ca^(2+)]_iin PASMCs under hypoxia was increased and [Ca^(2+)]_iwas significantly decreased when co-incubated with QBPF at 5%, 10%, and 20%. Compared with normoxia conditions, the m RNA and protein expression levels of calcineurin and NFATc3 in PASMCs induced by hypoxia were up-regulated.QBPF application significantly down-regulated m RNA and protein expression levels of calcineurin and NFATc3 in PASMCs.CONCLUSION: QBPF can effectively inhibit hypoxia-induced proliferation of PASMCs through down-regulation of key molecular expression via the Ca^(2+)/calcineurin/NFATc3 pathway.
