Objective: Nowadays, bone constructs elaborated according to tissue engineering principles are being regarded as an ideal choice for the reconstruction of segmental bone defects. In this study, proliferation and bone differentiation of marrow-derived mesenchymal stem cells (MSCs) were compared in different composite scaffolds containing varying morphologies of nano hydroxyapatite (nHAP). Materials and Methods: Needle nHAP/PLLA (poly (L-lactide acid)), spherical nHAP/ PLLA and rod nHAP/PLLA scaffolds were prepared and 3D cultures of passaged-3 rat MSCs were established using the scaffolds. The loading of the cells onto the scaffold internal spaces was confirmed with microscopy and their proliferation was determined by MTT assay. To compare the osteogenic differentiation of the cells on the scaffold surfaces, osteogenic 3D cultures were established and kept for 21 days. At the end of this period culture mineralization and relative bone-related gene expression were quantified using the alizarin red quantification assay and semi quantitative RT-PCR analysis respectively. ANOVA was used to compare the data. Results: According to the MTT assays, cells adhered to all the studied scaffold surfaces tended to proliferate. In this respect the microenvironment provided by the needle nHAP/ PLLA appeared much better than that of either the spherical or rod nHAP/PLLA scaffolds (P<0.05). Similarly, mineralization was observed to be heavier for the needle nHAP/PLLA scaffold compared to the two other composite scaffolds. In addition, the relative expression of coll I, osteocalcin, runx2 and ALP genes all appeared to be significantly higher in the cells cultivated on needle nHAP/PLLA scaffolds versus their spherical and rod counterparts. Conclusion: Overall, needle nHAP/PLLA scaffolds appear to provide the most appropriate matrix for producing bone construct using MSCs.