Objective: To observe the inhibitory effect and potential mechanism of mitomycin C and paclitaxel on the proliferation of Human Pulmonary Fibroblast in vitro. So as to providing an experimental reference for the design of drug eluting airway stents.
Methods: Cell viability was measured by MTT assay after different concentrations of mitomycin C or paclitaxel varying from 10(-1)1 mol/L to 10(-4) mol/L had been applied to the fibroblasts for 24, 48 or 72 h, respectively. Cell apoptosis was assessed by flow cytometry using dual staining with annexin V-FITC and propidium iodide 48 h after administering mitomycin C or paclitaxel at a concentration of 5×10(-6), 10(-5), 5×10(-5), 10(-4), 2×10(-4) mol/L, respectively. And the morphological character of cell apoptosis was observed by Hoechst 33342 fluorescent staining.
Results: The results of MTT revealed that cell proliferation were inhibited by mitomycin C and paclitaxel at all concentrations and exposure times. Among them, the inhibitory effect of mitomycin C were weak when the concentrations were between 10(-1)1 mol/L to 10(-8) mol/L. And within this context, the inhibitory ratio didn't correspond to the elevation of the concentration or the prolongation of the exposure times.However, when the concentration were between 10(-7) mol/L to 10(-4) mol/L, the inhibitory ratio rise progressively as the elevation of the concentration at all exposure times. The inhibitory ratio were 53.52%, 60.23%, 89.81% and 96.47% respectively when cells were treated by 10(-7), 10(-6), 10(-5) mol/L and 10(-4) mol/L mitomycin C for 72 h. An apparent "threshold dose effect" was observed in the paclitaxel treated groups.It's worth noting that the inhibitory ratio was only 48.22% when the cells had already been treated by 10(-5) mol/L paclitaxel for 72 h.However, when the concentration had reached 10(-4) mol/L, the inhibitory ratio sharply climbed to 93.38% even the cells had only been treated for 24 h. And the inhibitory ratio continued to rise as time prolonged. The results of cell apoptosis were consistent with MTT.When a significant inhibitory effect were detected by MTT, remarkable cell apoptosis could be observed by flow cytometry, and typical apoptotic cell could be identified by Hoechst 33342 fluorescent staining.
Conclusions: A certain concentration of mitomycin C or paclitaxel can inhibit Human Pulmonary Fibroblast proliferation in vitro. Both of these two drugs have potential value for the preparation of drug eluting airway stents. In order to ensure the inhibitory effect, the eluting concentration of mitomycin C and paclitaxel should not be less than 10(-7) mol/L and 10(-5) mol/L. But the eluting concentration of these two drugs should not exceed 10(-4) mol/L when both of the inhibitory ratio of these two drugs were higher than 95%.On this basis, elevating the drug concentration has little significance for improving the inhibitory effect, but increase the risk of systemic toxicity. Inducing cell apoptosis is one of the potential mechanisms of mitomycin C and paclitaxel in inhibiting cell proliferation.