Objective To investigate the effects of amplitude dependent morphological and proliferative changes in human bladder smooth muscle cells (hBSMCs) undergoing physiological stretch in vitro.Methods The hBSMCs were cuhured on silicone membrane and stretched similarly to a bladder cycle at range of stretches and time.The elongation would increase up to 2.5％ every 3 h and 5％ (or 10％,15％,20％ and 25％ depending on the experiment design) in the next 1 h,followed by a rapid decrease,cyclically maintained for a total of 16 h.In subsequent 8 h (24 h cycles) the membrane was maintained in relaxed position.Immunofluorescence and confocal laser scanning microscope were employed to assess the morphological changes.Cell counting kit-8 (CCK-8) and flow cytometry were used to assess the cell proliferation.Results The hBSMCs showed contractile phenotype after application of mechanical strain.Compared with control,the strains at 5％,10％,15％,20％ and 25％ induced most of the cells to change from a more spread-out and stellate state with large cell surface contact areas to a typical spindle-like morphology.The orientation angle of BSMCs remarkably differed depending on the applied strain's magnitude.Absorbance value,which reflects the proliferation activity,analyzed by CCK-8 was improved from 0.471 ± 0.027 (control) to 1.320 ± 0.094 (5％ elongation group,P ＜ 0.0001),1.001 ± 0.029 (10％ elongation group,P ＜0.0001),0.821 ±0.032 (15％ elongation group,P＜0.0001),0.621 ±0.032 (20％ elongation group,P =0.0004) and 0.591 ± 0.056 (25％ elongation group,P =0.0268),respectively.Cell proliferation index increased from (29.35 ±0.55)％ (control) to (55.55 ± 1.05)％ (5％ elongation group,P ＜0.0001),(47.70 ±0.20)％ (10％ elongation group,P＜0.0001),(35.40 ±2.10)％ (15％ elongation group,P ＜0.0001),respectively.However,no significant difference was found in either 20％ elongation group (34.85 ±0.55)％ (P=0.1372) or 25％ elongation group (30.35 ±0.45)％ (P=0.5234).Conclusions Proliferative and morphological changes could be observed in hBSMCs in vitro.Maximal proliferative potential could be seen at 5％ of stretch.