[Monocrotaline pyrrole induces A549 cells and activates TGF-β1/SMAD2/SMAD3 pathway to promote proliferation and migration of human pulmonary artery smooth muscle cells].

Objective To explore the effects of A549 cells on the proliferation and migration of human pulmonary arterial smooth muscle cells (HPASMCs) and its mechanism. Methods A549 cells and HPASMCs were cultured in vitro. The A549 cells were randomly divided into four groups: control group, dimethylformamide (DMF) solvent group, monocrotaline pyrrole (MCTP) group, MCTP combined with SB431542 group. The cells were assigned into four groups: HPASMC group, A549 and HPASMC co-culture group, MCTP-stimulated A549 and HPASMC co-culture group, MCTP and SB431542-stimulated A549 and HPASMC co-culture group, and IL-6-stimulated HPASMC group. A549 cell viability was detected by CCK-8 assay. The level of IL-6 in the A549 cell culture supernatant was tested by ELISA. The mRNA levels of SMAD2 and SMAD3 in the A549 cells were detected by real-time PCR. The protein levels of TGF-β1, SMAD2, SMAD3 and p-SMAD2, p-SMAD3 in the A549 cells were detected by Western blot analysis. The protein levels of TGF-β1, SMAD2, SMAD3 and p-SMAD2, p-SMAD3 in the A549 cells were examined by Western blot analysis. The protein levels of osteopontin (OPN) and proliferating nuclear antigen (PCNA) in the HPASMCs were determined by Western blot analysis. The migration ability of HPASMCs was measured by wound healing and TranswellTM assay. Results In the A549 cells, compared with the control group, the cell proliferation ability decreased, the production of IL-6 increased, the mRNA levels of SMAD2 and SMAD3, and the expression of TGF-β1, SMAD2, SMAD3, p-SMAD2, p-SMAD3 proteins significantly increased in the MCTP group. Compared with the MCTP group, the cell proliferation ability increased, the production of IL-6 decreased, the mRNA levels of SMAD 2 and SMAD3, and the expression of TGF-β1, SMAD2, SMAD3, p-SMAD2, p-SMAD3 proteins significantly decreased in the MCTP and SB431542-stimulated group. In the co-culture system, compared with the HPASMC group, the expression of PCNA and OPN proteins and migration ability did not change significantly in the A549 and HPASMC co-cultured group. The expression of PCNA and OPN proteins significantly increased in the MCTP-stimulated A549 and HPASMC co-culture group, and the cell migration ability increased. Compared with the MCTP-stimulated A549 and HPASMC co-culture group, the expression of PCNA and OPN proteins significantly decreased in MCTP and SB431542-stimulated A549 and HPASMC co-culture group, and the cell migration ability decreased. Compared with the HPASMC group, the migration ability of HPASMCs increased and the expression of PCNA and OPN proteins increased in the IL-6 control group. Conclusion Activation of the TGF-β1/SMAD2/SMAD3 signaling pathway in A549 cells induced by MCTP increases IL-6 secretion, thus promoting the proliferation and migration of HPASMCs.