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Follicular dendritic cell secreted protein (FDC‐SP), has been found to inhibit osteogenic differentiation of human periodontal ligament cells (hPDLCs) in recent studies. Based on these findings, we further investigate its effect on phenotype expression of hPDLCs in the present study, aiming to contribute to a better understanding of the biological functions governing FDC‐SP‐induced hPDLC differentiation. hPDLCs were firstly identified with immunocytochemical staining, followed by transfection with FDC‐SP lentiviral vector. Western blot analysis was used to confirm the expression of FDC‐SP. Then the influence of FDC‐SP transfection on hPDLC proliferation, osteogenic and fibrogenic phenotype expression was evaluated at the mRNA and protein level. Procollagen type I c‐peptide production was measured and alizarin red staining was then conducted to demonstrate effect of FDC‐SP on functional differentiation. We found that hPDLCs could be successfully transfected with FDC‐SP. Cell proliferation and cell cycle tests indicated that transfection with FDC‐SP did not affect hPDLC proliferation.Moreover, according to real‐time PCR andWestern blot results, expression levels of type 1 collagen alpha 1, type 1 collagen alpha 2 and type 3 collagenwere upregulated while that of osteocalcin, osteopontin, and bone sialoprotein were downregulated in FDC‐SP transfected cells. In addition, hPDLCs overexpressing FDC‐SP exhibited higher PIP production than the controls. Our findings demonstrate that transfection with FDC‐SP has negligible adverse effect on proliferation of hPDLCs and imply the biological function of FDC‐SP as a fibroblastic phenotype stabilizer by inhibiting hPDLCs differentiation into mineralized tissue‐forming cells, thus regulating regeneration in periodontal tissue engineering. J. Cell. Biochem. 9999: 1–9, 2013. 2013 Wiley Periodicals, Inc.