Induction of apoptosis in nasal polyp-derived fibroblasts by bleomycin A5 in vitro

Abstract

The present-study aimed to evaluate the pro-apoptotic effects of bleomycin A5 on nasal polyp-derived fibroblasts (NPDFs) and the underlying molecular mechanisms. Nasal polyp tissue was acquired from 10 patients during surgery and NPDFs were isolated from surgical tissues. Fibroblasts were identified using immunohistochemistry. Bleomycin A5 was used to treat NPDFs along a concentration gradient. Cell viability was evaluated using a Cell Counting Kit-8 assay. A flow cytometric Annexin V-fluorescein isothiocyanate/propidium iodide assay was used to determine the percentage of apoptotic NPDFs. The mRNA expression levels of apoptotic genes were determined by reverse transcription-quantitative polymerase chain reaction and levels of proteins associated with apoptosis were determined by western blotting. The results indicated that bleomycin A5 was able to induce apoptosis in NPDFs in a dose-dependent manner. NPDFs treated with bleomycin A5 were identified to contain significantly high amounts of the active forms of caspase-3 and showed considerable cleavage of poly(ADP-ribose) polymerase. The mRNA and protein expression levels of the pro-apoptotic molecule Bcl-2-associated X protein were significantly higher in treated NPDFs than in untreated NPDFs. In contrast, the mRNA and protein expression of the anti-apoptotic molecule B-cell lymphoma 2 (Bcl-2) was significantly lower in treated NPDFs. These results indicated that bleomycin A5 could induce apoptosis in primary NPDFs through activation of the Bcl-2 family and caspase cascades in a time-, and concentration-dependent manner.