ABSTRACT Objective: The aim of this study was to examine the role of NT-3 as a single neurotrophic factor in the expression of nestin in the neural differentiation of MSCs. Methods: MSCs were isolated from rat bone marrow and induced with NT-3 at concentrations of 20, 25, and 30 ng/ml for 7 and 14 d (the control was no NT-3). Nestin underwent immunocytochemical analysis on days 7 and 14. Five high-power random fields were documented. Results: A post-hoc analysis using LSD after one-way ANOVA test yielded a statistically significant difference in the percentage of nestin-positive cells in MSCs with NT-3 at concentrations of 20, 25, and 30 ng/ml for 7 d compared to the control group (p<0.05). The percentages of nestinpositive cells at concentrations of 20, 25, and 30 ng/ml, and in the control data on day 7 were 14.55±1.26%, 16.20±1.07%, 13.78±1.19%, and 9.81±0.79%, respectively. NT-3 at 25 ng/ml induced the highest MSCs neural differentiation on day 7 and remained constant until day 14. Conclusion: NT-3 plays a role in the early stage of differentiating MSCs from rat bone marrow into neurons, with the optimal concentration being 25 ng/ml. Keywords: Differentiation, Mesenchymal stem cells, Nestin, Neuron, Neurotrophin-3