Liao Hui, Pan Yiqin, Liu Yiming, Li Yuxiao, Huang Shiyi, Ding Shan, Xiang Qi
State Key Laboratory of Bioactive Molecules and Drug Gability Assessment, Jinan University, No. 855 East Xingye Avenue, Guangzhou 510632, China.
Institute of Biomedicine, Jinan University, No. 601 Huangpu Avenue West, Guangzhou 510632, China.
Bioengineering (Basel). 2024 Nov 28;11(12):1206. doi: 10.3390/bioengineering11121206.
While TGF-β3 promoted defect healing in a primate baboon skull defect model and patients, it remains unclear whether TGF-β3 affects the formation of osteoclasts and bone resorption between osteogenesis and osteolysis. Analysis of the full transcriptome of hPDLSCs (human periodontal ligament stem cells) revealed that the expression of RANKL was significantly up-regulated after TGF-β3 treatment during osteogenesis, which suggests its involvement in clock-controlled autophagy in bone metabolism. TRAP staining and bone resorption lacunae were used to assess the osteoclasts formed from RANKL-induced differentiated BMMs. During osteoclast differentiation, the characteristics of autophagy regulated by TGF-β3 were observed in BMMs through MDC staining, transmission electron microscopy, and LC3 immunofluorescence. The expression of related genes and proteins were detected on the sixth day in mCherry-EGFP-LC3B lentivirus-transfected BMMs using RT-qPCR and WB. Finally, a trans-well co-culture system was used to evaluate the effects of osteogenic differentiated hPDLSCs treated with TGF-β3 on the osteoclastic differentiation of BMMs. The results showed 10 ng/mL of TGF-β3 significantly suppressed osteoclastic differentiation and bone resorption in BMMs ( < 0.05 vs. RANKL). In particular, TGF-β3 augments the expression of LC3-II to stimulate autophagy, consequently restraining osteoclastic resorption. These findings provide a molecular basis and are beneficial to illustrate the potential druggability of TGF-β3 in osteoporotic diseases.
虽然转化生长因子β3（TGF-β3）在灵长类狒狒颅骨缺损模型和患者中促进了缺损愈合，但TGF-β3是否影响破骨细胞的形成以及成骨与骨吸收之间的骨溶解尚不清楚。对人牙周膜干细胞（hPDLSCs）的全转录组分析表明，在成骨过程中，TGF-β3处理后核因子κB受体活化因子配体（RANKL）的表达显著上调，这表明其参与了骨代谢中生物钟控制的自噬。采用抗酒石酸酸性磷酸酶（TRAP）染色和骨吸收陷窝来评估由RANKL诱导分化的骨髓巨噬细胞（BMMs）形成的破骨细胞。在破骨细胞分化过程中，通过单丹磺酰尸胺（MDC）染色、透射电子显微镜和微管相关蛋白1轻链3（LC3）免疫荧光观察BMMs中TGF-β3调节的自噬特征。在mCherry-EGFP-LC3B慢病毒转染的BMMs中，于第6天使用逆转录定量聚合酶链反应（RT-qPCR）和蛋白质免疫印迹法（WB）检测相关基因和蛋白质的表达。最后，采用Transwell共培养系统评估经TGF-β3处理的成骨分化hPDLSCs对BMMs破骨细胞分化的影响。结果显示，10 ng/mL的TGF-β3显著抑制了BMMs的破骨细胞分化和骨吸收（与RANKL相比，P<0.05）。特别是，TGF-β3增强了LC3-II的表达以刺激自噬，从而抑制破骨细胞吸收。这些发现提供了分子基础，有助于阐明TGF-β3在骨质疏松疾病中的潜在可药用性。
