Urolithin A mediates p38/MAPK pathway to inhibit osteoclast activity / 中国组织工程研究

BACKGROUND:Overactive osteoclasts disrupt bone homeostasis and play a bad role in the pathological mechanisms of related skeletal diseases,such as osteoporosis,fragility fractures,and osteoarthritis.Studies have confirmed that ellagic acid and ellagtannin have the potential to inhibit osteoclast differentiation.As their natural metabolites,urolithin A has antioxidant,anti-inflammatory,anti-proliferative and anti-cancer effects,but its effect on osteoclast differentiation and its underlying molecular mechanisms remain unclear. OBJECTIVE:To explore the effect of urolithin A on osteoclast differentiation induced by receptor activator for nuclear factor-κB ligand and its mechanism. METHODS:Mouse mononuclear macrophage leukemia cells(RAW264.7)that grew stably were cultured in vitro.Toxicity of urolithin A(0,0.1,0.5,1.5,2.5 μmol/L)to RAW264.7 cells were detected by cytotoxic MTS assay to screen out the safe concentration.Different concentrations of urolithin A were used again to intervene with receptor activator for nuclear factor-κB ligand-induced differentiation of RAW264.7 cells in vitro.Then,tartrate-resistant acid phosphatase staining and F-actin ring and nucleus staining were performed to observe its effect on the formation and function of osteoclasts.Finally,the expressions of urolithin A on upstream and downstream genes and proteins in the MAPK signaling pathway were observed by western blot and RT-qPCR assays. RESULTS AND CONCLUSION:Urolithin A inhibited osteoclast differentiation and F-actin ring formation in a concentration-dependent manner and 2.5 μmol/L had the strongest inhibitory effect.Urolithin A inhibited the mRNA expression of Nfatc1,Ctsk,Mmp9 and Atp6v0d2 and the protein synthesis of Nfatc1 and Ctsk,related to osteoclast formation and bone resorption.Urolithin A inhibited the activity of osteoclasts by downregulating the phosphorylation of p38 protein to inhibit the mitogen-activated protein kinase signaling pathway.

The pathological progression of steroid-induced osteonecrosis of the femoral head caused by oxidative stress-induced osteoblast ferroptosis / 中国组织工程研究

BACKGROUND:Studies have shown that imbalance of bone metabolism during glucocorticoid-induced osteonecrosis of the femoral head necrosis is closely related to oxidative stress. OBJECTIVE:To investigate the pathological mechanism by which oxidative stress-induced ferroptosis promote apoptosis in osteoblasts involved in steroid-induced osteonecrosis of the femoral head. METHODS:General data and serum specimens were collected from 47 patients with steroid-induced osteonecrosis of the femoral head.In addition,six femoral head specimens were collected from these patients.According to the Association Research Circulation Osseous(ARCO)staging system,serum specimens were grouped into ARCO Ⅱ,Ⅲ,and IV,while femoral head specimens were classified into ARCO Ⅲ and IV.Serum levels of malondialdehyde and superoxide dismutase 1 were measured.The protein expression of superoxide dismutase 1,glutathione peroxidase 4,Bcl-2 in the femoral head was detected and verified by Data independent acquisition(DIA)for quantitative sequencing,western blot and alkaline phosphate detection. RESULTS AND CONCLUSION:The ARCO stage of patients with steroid-induced osteonecrosis of the femoral head was independent of age,sex and necrotic side.The serum levels of malondialdehyde and superoxide dismutase 1 were higher in patients with ARCO stage Ⅲ compared with those with ARCO stage Ⅱ and IV.The results of DIA protein quantification showed that the function of differential proteins was mainly related to redox.The levels of superoxide dismutase 1,glutathione peroxidase 4,and Bcl-2 in the necrotic region were lower than in the normal region,as well as lower in ARCO stage IV than in ARCO stage Ⅲ.Western blot verified the results of DIA protein quantification.The alkaline phosphatase activity was lower in the necrotic region than in the normal region,as well as lower in ARCO stage IV than in ARCO stage Ⅲ.In the necrotic and sclerotic regions,the function of differential proteins was also related to redox,and superoxide dismutase 1,glutathione peroxidase 4,Bcl-2 protein expression and alkaline phosphatase activity were lower in the necrotic area than in the sclerotic region,as well as lower in ARCO stage IV than in ARCO stage Ⅲ.To conclude,glucocorticoids can influence the progression of steroid-induced osteonecrosis of the femoral head by upregulating oxidative stress levels,inducing osteoblast ferroptosis,and inhibiting osteogenic function.

A deep transfer learning method using plain radiographs for the differential diagnosis of osteonecrosis of the femoral head with other hip diseases / 中华骨科杂志

Objective:To develop a deep transfer learning method for the differential diagnosis of osteonecrosis of the femoral head (ONFH) with other common hip diseases using anteroposterior hip radiographs.Methods:Patients suffering from ONFH, DDH, and other hip diseases including primary hip osteoarthritis, non-infectious inflammatory hip disease, and femoral neck fracture treated in the First Affiliated Hospital of Guangzhou University of Chinese Medicine from January 2018 to December 2020 were enrolled in the study. A clinical data set containing anteroposterior hip radiographs of the eligible patients was created. Data augmentation by rotating and flipping images was performed to enlarge the data set, then the data set was divided equally into a training data set and a testing data set. The ResNet-152, a deep neural network model, was used in the study, but the original Batch Normalization was replaced with Transferable Normalization to construct a novel deep transfer learning model. The model was trained to distinguish ONFH and DDH from other common hip diseases using anteroposterior hip radiographs on the training data set and its classification performance was evaluated on the testing data set.Results:The clinical data set was comprised of anteroposterior hip radiographs of 1024 hips, including 542 with ONFH, 296 with DDH, and 186 with other common hip diseases (56 hips with primary osteoarthritis, 85 hips with non-infectious inflammatory osteoarthritis, 45 hips with femoral neck fracture). After data augmentation, the size of the data set multiplied to 6144. The model was trained 100 050 times in each task. Accuracy was used as the representative parameter to evaluate the performance of the model. In the binary classification task to identify ONFH, the best accuracy was 95.80%. As for the multi-classification task for classification of ONFH and DDH from other hip diseases, the best accuracy was 91.40%. The plateau of the model was observed in each task after 50 000 times of training. The mean accuracy in plateaus was 95.35% (95% CI: 95.33%, 95.37%), and 90.85% (95% CI: 90.82%, 90.87%), respectively. Conclusion:The present study proves the encouraging performance of a deep transfer learning method for the first-visit classification of ONFH, DDH, and other hip diseases using the convenient and economical anteroposterior hip radiographs.