ABSTRACT
Objective To investigate the influence of three extracellular matrix (ECM) proteins, namely, laminin (LN), collagen type I (Col I), fibronectin (FN) on the morphology and contractility changes in airway smooth muscle cells (ASMCs) induced by platelet-derived growth factor-BB (PDGF-BB). Methods ASMCs were seeded on the culture dish coated with LN, Col I, or FN, respectively, and divided into two groups to be cultured either in the absence or presence of PDGF-BB (10 mg/L) for 0~5 d. Subsequently, cell morphology was examined by the optical microscopy and quantified as the ratio of cell width to length, and the KCl/histamine-induced contractile responses of the cell were measured by optical magnetic twisting cytometry (OMTC). Results ASMCs cultured in the presence of PDGF-BB generally appeared in longer and thinner cell shapes, namely, a smaller ratio of cell width to length, but the cell width/length ratio for ASMCs adhered on LN was relatively bigger than that on Col I or FN. In the absence of PDGF-BB, contractility of ASMCs to KCl increased with the duration of culture, which was independent of the ECM proteins. In contrast, in the presence of PDGF-BB, contractility of ASMCs to KCl or histamine decreased in all situations, but degree of the decrease was smaller for ASMCs adhered on LN than those on Col I or FN. Conclusions The morphology and contractility changes in ASMCs induced by PDGF-BB are influenced by ECM proteins on which cells are grown. For ASMCs adhered on LN, the morphology and contractility changes are relatively smaller than those on Col I and FN. The differential effect of ECM proteins on PDGF-BB induced changes in morphology and contractility of ASMCs is important to fully understand the interactions between ECM proteins, inflammatory factors, ASMCs, and their relation to the pathophysiological mechanism of asthma.
ABSTRACT
Objective To investigate the effect from different pore sizes of co culture inserts on the permeability of biomacromolecules through polyethylene terephthalate (PET) membrane so as to solve the key technology problem in mechanobiology experiment on vascular cells. Methods Inserts with 0.4 μm and 1.0 μm pores on the PET membrane were studied using flow chamber system. Low shear stress was subjected to the co-cultured system of endothelial cell (EC)/vascular smooth muscle cell (VSMC) and the concentration of platelet-derived growth factor BB (PDGF-BB) was detected by ELISA. Under the static condition, vascular cells were cultured on the plate (with no cell on PET membrane), on the outer side of PET membrane, and on the both sides of PET membrane, respectively. Then the recombinants PDGF-BB (rPDGF-BB) were added on the different sides of PET membrane. Western blotting was used to detect the change in expressions of p-ERK1/2, p-Akt and Lamin after cells were stimulated by rPGDF BB. Results After low shear stress subjection for 12 h, the concentration of PDGF-BB in the medium from VSMC side was significantly higher than that from EC-side. rPDGF-BB passed through 0.4 μm and 1.0 μm pores on the PET membrane and modulated expressions of p-ERK1/2, p-Akt and Lamin A in cells cultured on the opposite side of PET membrane and cells cultured on the plate separately. When cells were cultured on the both sides of PET membrane, rPDGF-BB only stimulated cells cultured on the same side of 0.4 μm pores on PET membrane, but had no specific effect on cells cultured on the opposite side. Conclusions PET membrane with both 0.4 μm and 1.0 μm pores was permeable to PDGF-BB, and cells cultured on the membrane could affect the permeability. The efficiency of PDGF BB passing through 0.4 μm pores was significantly repressed with cells cultured on the both sides, which was more similar to that in vivo.