Objective: To explore the functional responses of normal rat pulmonary artery smooth muscle cells (PASMCs) from different orders of pulmonary artery to the platelet-derived growth factor (PDGF).
Methods: The pulmonary artery branches were gently isolated from Sprague-Dawley rats (250-350 g) and eventually cut into three groups according to the vascular grading: the proximal pulmonary arteries, including pulmonary trunk and the first order branches of intrapulmonary arteries;the middle pulmonary arteries, including the second to third orders of intrapulmonary arteries;the distal pulmonary arteries, including the forth to fifth orders of intrapulmonary arteries. PASMCs from the three groups of vessels were isolated and cultured in DMEM. Cells were identified by immunofluorescence-stained smooth muscle cell markers α-SMA, S100A4 and Calponin antibody. Cell proliferation was quantified by using 0.5 g/L methyl thiazolyl tetrazolium (MTT) assay. Western blot was used for detecting the protein expression level of proliferating cell nuclear antigen (PCNA) in PASMCs. Wound healing test was employed for cell migration assay.
Results: All of the three groups of cells positively expressed α-SMA, S100A4 and Calponin marker and no obvious differences were found in the appearance among the three groups. Under control condition (cultured in DMEM+2%FBS), three groups of PASMCs showed similar cell proliferation rate (137.0 ± 12.1)%, (166.1 ± 23.6)%, (156.8 ± 2.5)%. The expression level of PCNA by Western blot was (0.42 ± 0.10), (0.55 ± 0.18), (0.48 ± 0.28), respectively. The numbers of cells migrating to blank areas of the three groups were (49 ± 14), (68 ± 13) and (71 ± 7). The above results were not significantly different. While after incubation with PDGF (10 µg/L, 48 h), the proliferation rate of middle and distal groups were (215.7 ± 17.8)% and (195.2 ± 6.6)%, higher than that of the proximal group(167.0 ± 14.4)%, (P < 0.01). In middle and distal groups, the expression level of PCNA were (0.93 ± 0.11) and (1.21 ± 0.14) respectively, which were higher than that in the proximal group (0.62 ± 0.03), (P < 0.001). The number of cells migrating to blank areas of the middle and distal groups were (194 ± 46), (171 ± 16), and exhibited increased migration ability compared to the proximal group (67 ± 21), (P < 0.01).
Conclusion: These results suggest that PASMCs from different orders of pulmonary arteries have different responses to PDGF stimulation. This will help to understand the possible mechanism for that under same pathophysiological stimulus, different orders of pulmonary arteries may have different responses.