The New Role of Sirtuin1 in Human Osteoarthritis Chondrocytes by Regulating Autophagy.

OBJECTIVE: The aim of this study is to investigate the role of Sirtuin1 (Sirt1) in the regulation of autophagy for human osteoarthritis (OA) chondrocytes. DESIGN: All cartilage samples were collected from human donors, including young group, aged group, and OA group. Primary chondrocytes were isolated and cultured with Sirt1 activator or inhibitor. Sirt1 expression in cartilage tissue and chondrocytes was evaluated, and the deacetylation activity of Sirt1 was determined. The alteration of autophagy activity after upregulating or downregulating Sirt1 was detected. Chondrocytes were treated with autophagy activator and inhibitor, and then the protein level of Sirt1 was examined. The interactions between Sirt1 and autophagy-related proteins Atg7, microtubule associated protein 1 light chain 3 (LC3), and Beclin-1 were determined by using immunoprecipitation. RESULTS: The assay of articular cartilage revealed that the expression of Sirt1 might be age-related: highly expressed in of younger people, and respectively decreased in the elderly people and OA patients. In vitro study was also validated this result. Further study confirmed that higher levels of Sirt1 significantly increased autophagy in aged chondrocytes, while the lower expression of Sirt1 reduced autophagy in young chondrocytes. Of note, the high levels of Sirt1 reduced autophagy in OA chondrocytes. When the chondrocytes were treated with autophagy activator or inhibitor, we found the expression of Sirt1 was not affected. In addition, we found that Sirt1 could interact with Atg7. CONCLUSION: These results suggest that Sirt1 in human chondrocytes regulates autophagy by interacting with autophagy related Atg7, and Sirt1 may become a more important target in OA treatment.

Knockdown of TBRG4 suppresses proliferation, invasion and promotes apoptosis of osteosarcoma cells by downregulating TGF-ß1 expression and PI3K/AKT signaling pathway.

Transforming growth factor beta regulator 4 (TBRG4) is a novel regulator in tumorigenic progression of several tumors. However, so far, the expression and functions of TBRG4 in osteosarcoma are unknown. The aim of this study was to investigate the potential biological functions of TBRG4 in osteosarcoma. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression of TBRG4 in osteosarcoma tissues and cell lines. The levels of TBRG4 protein in osteosarcoma tissues were assessed by immunohistochemistry. Lentivirus-mediated short hairpin (sh) RNA was employed to knock down TBRG4 in osteosarcoma cells, and the expressions of TBRG4 mRNA and protein were determined by qRT-PCR and Western blot assay, respectively. Subsequently, the proliferation, clonogenic ability, apoptosis and invasion of osteosarcoma cells were measured using high content screening analysis and CCK8 assay, tumor sphere formation assay, flow cytometry and Transwell invasion assays, respectively. Furthermore, the osteosarcoma cells growth and metastasis in vivo were detected, and the effect of TBRG4 on the transforming growth factor ß1 (TGF-ß1) and PI3K/AKT signaling pathway was explored by qRT-PCR and Western blot assay, respectively. The results showed the levels of TBRG4 were overexpressed in osteosarcoma tissues and cell lines, confirming that the high TBRG4 expression was related to advanced tumor stages, large tumor size, and lymph node metastasis. Functional assays showed knockdown of TBRG4 could inhibit proliferation, invasion and induce apoptosis of osteosarcoma cells in vitro, and could also suppress osteosarcoma growth and metastasis in vivo. By examining the expression levels of TGF-ß1, p-PI3K, PI3K, p-AKT and AKT, it showed that the suppression of TBRG4 would reduce TGF-ß1 expression and inactivate the PI3K/AKT signaling pathway. These results showed for the first time that TBRG4 knockdown could suppress osteosarcoma progression, suggesting TBRG4 might be a promising therapeutic target for osteosarcoma treatment.