Biodegradable nanofiber has become a popular candidate as tissue engineering scaffolds due to its biomimic structure as natural extracellular matrix (ECM). Certain tissue regeneration may require prolonged in vitro culture time for cellular reorganization and tissue remodeling. Therefore, long term understanding of cellular effects on scaffold degradation is needed. Although there are some degradation studies on nanofiber, degradation study of nanofibers with cell culture is rare. In our study, polyglycolide (PGA), poly(DL-lactide-co-glycolide) (PLGA), poly(L-lactide-co-ε- caprolactone) [P(LLA-CL)] were electrospun into nanofiber scaffolds. The scaffolds were cultured with porcine smooth muscle cells (PSMC) for up to 3 months to evaluate their degradation behavior and cellular response. The results showed that their degradation rates were in the order of PGA>>PLGA>P(LLA-CL). PGA nanofiber degraded in 3 weeks and only supported cell growth in the first few days. Cell culture accelerated the surface erosion of PLGA and P(LLA-CL) nanofiber while the bulk degradation remained unaffected. Furthermore, the cell culture did not significantly reduce the mechanical strength of PLGA and P(LLA-CL) during degradation.