目的：本研究旨在探讨程序性细胞死亡配体1（PD-L1）通过介导CCCTC结合因子（CTCF）表达促进人牙髓干细胞（hDPSCs）增殖和成骨分化的作用及其机制。
方法：通过免疫共沉淀法验证PD-L1与CTCF的相互作用。用脂多糖或成骨诱导培养基处理用PD-L1过表达和CTCF敲低载体转染的hDPSC。检测炎性细胞因子和骨/牙源性分化相关基因。使用碱性磷酸酶(ALP)和茜素红S染色评估hDPSC的骨/牙源性分化。
结果：PD-L1过表达抑制LPS诱导的促炎细胞因子上调，细胞增殖，ALP活性,和钙在hDPSC中的沉积，并提高了骨/牙源性分化相关基因的表达；然而，这种表达模式可以通过CTCF敲低逆转。免疫共沉淀结果证实了PD-L1与CTCF的结合，表明hDPSC中PD-L1过表达增加CTCF表达，从而抑制炎症反应并增加hDPSC的骨/牙源性分化。
结论：PD-L1在hDPSC中的过表达增强了hDPSC的增殖和骨/牙源性分化，并通过上调CTCF表达来抑制炎症反应。
OBJECTIVE: The aim of this study was to explore the effect and mechanism of programmed cell death ligand 1 (PD-L1) in promoting the proliferation and osteo/odontogenic-differentiation of human dental pulp stem cells (hDPSCs) by mediating CCCTC-binding factor (CTCF) expression.
METHODS: The interaction between PD-L1 and CTCF was verified through co-immunoprecipitation. hDPSCs transfected with PD-L1 overexpression and CTCF knockdown vectors were treated with lipopolysaccharide or an osteogenic-inducing medium. Inflammatory cytokines and osteo/odontogenic-differentiation related genes were measured. Osteo/odontogenic-differentiation of hDPSCs was assessed using alkaline phosphatase (ALP) and alizarin red S staining.
RESULTS: Overexpression of PD-L1 inhibited LPS-induced pro-inflammatory cytokine upregulation, cell proliferation, ALP activity, and calcium deposition in hDPSCs and elevated the expression of osteo/odontogenic-differentiation related genes; however, such expression patterns could be reversed by CTCF knockdown. Co-immunoprecipitation results confirmed the binding of PD-L1 to CTCF, indicating that PD-L1 overexpression in hDPSCs increases CTCF expression, thus inhibiting the inflammatory response and increasing osteo/odontogenic-differentiation of hDPSCs.
CONCLUSIONS: PD-L1 overexpression in hDPSCs enhances the proliferation and osteo/odontogenic-differentiation of hDPSCs and inhibit the inflammatory response by upregulating CTCF expression.
