© 2021 Informa UK Limited, trading as Taylor & Francis Group.Purpose/Aim of the study: The ultimate goal of periodontal treatment is to regenerate the lost periodontal tissues. The interest in nanomaterials in dentistry is growing rapidly and has focused on improvements in various biomedical applications, such as periodontal regeneration and periodontal tissue engineering. To enhance periodontal tissue regeneration, hydroxyapatite (HA) was used in conjunction with other scaffold materials, such as Poly lactic-co-glycolic-acid (PLGA) and collagen (C). The main target of this study was to compare the effects of nano and macrostructures of the tissue scaffolds on cell behavior in vitro for periodontal tissue engineering. Materials and Methods: Nanofibrillar and macroporous-spongious composite tissue scaffolds were produced using PLGA/C/HA. Subgroups with BMP-2 signal molecule and without HA were also created. The scaffolds were characterized by FTIR, SEM/EDX techniques, and mechanical tests. The scaffolds were compared in the periodontal ligament (PDL) and MCT3-E1 cell cultures. The cell behaviors; adhesions by SEM, proliferation by WST-1, differentiation by ALP and mineralization with Alizarin Red Tests were determined. Results: Cell adhesion and mineralization were higher in the nanofibrillar scaffolds compared to the macroporous-spongious scaffolds. Macroporous-spongious scaffolds seemed better for the proliferation of PDL cells and differentiation of MC3T3-E1-preosteoblastic cells, while nanofibrillar scaffolds were more convenient for the differentiation of PDL cells and proliferation of MC3T3-E1-preosteoblastic cells. Conclusions: In general, nanofibrillar scaffolds showed more favorable results in cell behaviors, compared to the macroporous-spongious scaffolds, and mostly, BMP-2 and HA promoted the activities of the cells.