The biological behavior optimization of human periodontal ligament stem cells via preconditioning by the combined application of fibroblast growth factor-2 and A83-01 in in vitro culture expansion

Abstract

Background As the optimal source of seed cells in periodontal tissue engineering, periodontal ligament stem cells (PDLSCs) have always been researched to improve cell expansion due to their limited resource and spontaneous differentiation in vitro cultivation. Fibroblast growth factor-2 (FGF-2) has been proven to stimulate bone marrow mesenchymal stem cells (BMMSCs) proliferation and maintain their pluripotency when being added to the culture medium. As a small molecule inhibitor of transforming growth factor-beta receptors (TGF-βRs), A83-01 can also promote cell proliferation. Therefore, the aim of this study was to verify whether the combined application of FGF-2 and A83-01 could augment cell quantity and quality during in vitro culture. Methods PDLSCs were preconditioned with A83-01, FGF-2, or their combination. A cell counting kit-8 (CCK8) assay, cell apoptosis assay, ALP activity assay, Alizarin Red S staining assay, RT-PCR assay, Western blot assay and ELISA were used to determine the sustained effects of different preconditioning strategies on the proliferation, apoptosis, stemness, osteogenic differentiation and paracrine action of PDLSCs. Results The combined application of FGF-2 and A83-01 significantly augmented cell expansion, reduced cell apoptosis, magnified stemness expression, promoted later osteogenic differentiation and mineralization and increased paracrine action of PDLSCs compared with the control. Moreover, the combination presented significant advantages in enhancing proliferation, stemness expression and paracrine action over FGF-2 alone. Conclusions The combined application of A83-01 and FGF-2 may be an improved strategy for PDLSCs biological behavior optimization in culture expansion and advantageous for reinforcing proliferation, stemness expression and cytokine secretion over FGF-2 alone.

Cellular Mechanisms of FGF-Stimulated Tissue Repair

“…A number of in vitro studies have demonstrated the importance of FGFs for the maintenance of cell stemness, i.e., the ability of stem cells to maintain the non-differentiated status and to give rise to differentiated cells under proper conditions. Thus, FGF2 efficiently maintained the stemness of rabbit [ 90 ] and human [ 91 ] embryonic stem cells in culture, as well as the stemness of neural stem cells [ 92 ], trophoblast stem cells [ 93 ], and periodontal ligament stem cells [ 94 ] in mice. In addition, FGF2 and FGF4 supported the stemness of bone marrow-derived mesenchymal stem cells in vitro [ 95 ].…”

Expansion of murine and human olfactory epithelium/mucosa colonies and generation of mature olfactory sensory neurons under chemically defined conditions

“…Visible colonies with both filled and cystic appearance were present on Day 7 and typical cystic cultures were observed on Day 14 (Figure 7 C). Ttreatment with A83-01 (a TGF-β kinase/activin receptor-like kinase inhibitor to promote cell proliferation in in vitro culture expansion 55 ) and SB431542 (TGF-β RI Kinase Inhibitor VI) increased the colony size by 11 ± 3% on Day 10, but the size was not apparently different on Day 14. Treatment with A83-01 and SB431542 did not significantly change morphology of human colonies ( Figure S7 A).…”

The influence of an electromagnetic field on adipose-derived stem/stromal cells’ growth factor secretion: Modulation of FGF-2 production by in vitro exposure

“…It was shown that in human periodontal ligament stem cells, supplementation of FGF-2 into the culture medium causes a significant increase in proliferation as well as the expression of a marker determining the stemness of the cells and a decrease in apoptosis after 5-7 days. Similarly, for BM-MSCs, FGF-2 supplementation helps maintain pluripotency [38].…”

“…Basic fibroblast growth factor (FGF-2 protein) is a GF that increases stem cell proliferation and supports them by maintaining their functions, such as pluripotency, self-renewal and regeneration [33,[36][37][38][39]. This GF can also increase the differentiation potential of MSCs and affect osteogenesis and chondrogenesis [19,33], and it can stimulate neuropoiesis and angiogenesis [40,41].…”