mir-150-5p inhibits the osteogenic differentiation of bone marrow-derived mesenchymal stem cells by targeting irisin to regulate the p38/MAPK signaling pathway.

PURPOSE: To study the effect of miR-150-5p on the osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs), and further explore the relationship between its regulatory mechanism and irisin. METHODS: We isolated mouse BMSCs, and induced osteogenic differentiation by osteogenic induction medium. Using qPCR to detect the expression of osteogenic differentiation-related genes, western blot to detect the expression of osteogenic differentiation-related proteins, and luciferase reporter system to verify that FNDC5 is the target of miR-150-5p. Irisin intraperitoneal injection to treat osteoporosis in mice constructed by subcutaneous injection of dexamethasone. RESULTS: Up-regulation of miR-150-5p inhibited the proliferation of BMSCs, and decreased the content of osteocalcin, ALP activity, calcium deposition, the expression of osteogenic differentiation genes (Runx2, OSX, OCN, OPN, ALP and BMP2) and protein (BMP2, OCN, and Runx2). And down-regulation of miR-150-5p plays the opposite role of up-regulation of miR-150-5p on osteogenic differentiation of BMSCs. Results of luciferase reporter gene assay showed that FNDC5 gene was the target gene of miR-150-5p, and miR-150-5p inhibited the expression of FNDC5 in mouse BMSCs. The expression of osteogenic differentiation genes and protein, the content of osteocalcin, ALP activity and calcium deposition in BMSCs co-overexpressed by miR-150-5p and FNDC5 was significantly higher than that of miR-150-5p overexpressed alone. In addition, the overexpression of FNDC5 reversed the blocked of p38/MAPK pathway by the overexpression of miR-150-5p in BMSCs. Irisin, a protein encoded by FNDC5 gene, improved symptoms in osteoporosis mice through intraperitoneal injection, while the inhibitor of p38/MAPK pathway weakened this function of irisin. CONCLUSION: miR-150-5p inhibits the osteogenic differentiation of BMSCs by targeting irisin to regulate the/p38/MAPK signaling pathway, and miR-150-5p/irisin/p38 pathway is a potential target for treating osteoporosis.

CCL2 Knockdown Attenuates Inflammatory Response After Spinal Cord Injury Through the PI3K/Akt Signaling Pathway: Bioinformatics Analysis and Experimental Validation.

Spinal cord injury (SCI) is a common clinical problem in orthopedics with a lack of effective treatments and drug targets. In the present study, we performed bioinformatic analysis of SCI datasets GSE464 and GSE45006 in the Gene Expression Omnibus (GEO) public database and experimentally validated CCL2 expression in an animal model of SCI. This was followed by stimulation of PC-12 cells using hydrogen peroxide to construct a cellular model of SCI. CCL2 expression was knocked down using small interfering RNA (si-CCL2), and PI3K signaling pathway inhibitors and activators were used to validate and observe the changes in downstream inflammation. Through data mining, we found that the inflammatory chemokine CCL2 and PI3K/Akt signaling pathways after SCI expression were significantly increased, and after peroxide stimulation of PC-12 cells with CCL2 knockdown, their downstream cellular inflammatory factor levels were decreased. The PI3K/Akt signaling pathway was blocked by PI3K inhibitors, and the downstream inflammatory response was suppressed. In contrast, when PI3K activators were used, the inflammatory response was enhanced, indicating that the CCL2-PI3K/Akt signaling pathway plays a key role in the regulation of the inflammatory response. This study revealed that the inflammatory chemokine CCL2 can regulate the inflammatory response of PC-12 cells through the PI3K/Akt signaling pathway, and blocking the expression of the inflammatory chemokine CCL2 may be a promising strategy for the treatment of secondary injury after SCI.

From cells to organs: progress and potential in cartilaginous organoids research.

While cartilage tissue engineering has significantly improved the speed and quality of cartilage regeneration, the underlying metabolic mechanisms are complex, making research in this area lengthy and challenging. In the past decade, organoids have evolved rapidly as valuable research tools. Methods to create these advanced human cell models range from simple tissue culture techniques to complex bioengineering approaches. Cartilaginous organoids in part mimic the microphysiology of human cartilage and fill a gap in high-fidelity cartilage disease models to a certain extent. They hold great promise to elucidate the pathogenic mechanism of a diversity of cartilage diseases and prove crucial in the development of new drugs. This review will focus on the research progress of cartilaginous organoids and propose strategies for cartilaginous organoid construction, study directions, and future perspectives.

Identification of the CCL2 PI3K/Akt axis involved in autophagy and apoptosis after spinal cord injury.

Spinal cord injury (SCI) is a devastating neurological disease with no cure that usually results in irreversible loss of sensory and voluntary motor functions below the injury site. We conducted an in-depth bioinformatics analysis combining the gene expression omnibus spinal cord injury database and the autophagy database and found that the expression of the autophagy gene CCL2 was significantly upregulated and the PI3K/Akt/mTOR signaling pathway was activated after SCI. The results of the bioinformatics analysis were verified by constructing animal and cellular models of SCI. We then used small interfering RNA to inhibit the expression of CCL2 and PI3K to inhibit and activate the PI3K/Akt/mTOR signaling pathway; western blot, immunofluorescence, monodansylcadaverine, and cell flow techniques were used to detect the expression of key proteins involved in downstream autophagy and apoptosis. We found that when PI3K inhibitors were activated, apoptosis decreased, the levels of autophagy-positive proteins LC3-I/LC3-II and Bcl-1 increased, the levels of autophagy-negative protein P62 decreased, the levels of pro-apoptotic proteins Bax and caspase-3 decreased, the levels of the apoptosis-inhibiting protein Bcl-2 increased. In contrast, when a PI3K activator was used, autophagy was inhibited, and apoptosis was increased. This study revealed the effect of CCL2 on autophagy and apoptosis after SCI through the PI3K/Akt/mTOR signaling pathway. By blocking the expression of the autophagy-related gene CCL2, the autophagic protective response can be activated, and apoptosis can be inhibited, which may be a promising strategy for the treatment of SCI.

Erythropoietin inhibits ferroptosis and ameliorates neurological function after spinal cord injury.

Ferroptosis is one of the critical pathological events in spinal cord injury. Erythropoietin has been reported to improve the recovery of spinal cord injury. However, whether ferroptosis is involved in the neuroprotective effects of erythropoietin on spinal cord injury has not been examined. In this study, we established rat models of spinal cord injury by modified Allen's method and intraperitoneally administered 1000 and 5000 IU/kg erythropoietin once a week for 2 successive weeks. Both low and high doses of erythropoietin promoted recovery of hindlimb function, and the high dose of erythropoietin led to better outcome. High dose of erythropoietin exhibited a stronger suppressive effect on ferroptosis relative to the low dose of erythropoietin. The effects of erythropoietin on inhibiting ferroptosis-related protein expression and restoring mitochondrial morphology were similar to those of Fer-1 (a ferroptosis suppressor), and the effects of erythropoietin were largely diminished by RSL3 (ferroptosis activator). In vitro experiments showed that erythropoietin inhibited RSL3-induced ferroptosis in PC12 cells and increased the expression of xCT and Gpx4. This suggests that xCT and Gpx4 are involved in the neuroprotective effects of erythropoietin on spinal cord injury. Our findings reveal the underlying anti-ferroptosis role of erythropoietin and provide a potential therapeutic strategy for treating spinal cord injury.

hsa_circ_0058122 knockdown prevents steroid-induced osteonecrosis of the femoral head by inhibiting human umbilical vein endothelial cells apoptosis via the miR-7974/IGFBP5 axis.

BACKGROUND: Steroid-induced osteonecrosis of femoral head (SONFH) is a serious complication of glucocorticoid overused. Recent evidence has demonstrated that circRNAs exert key pathophysiological roles in a variety of disease processes. However, the role of circRNA in SONFH remains largely unknown. The current study sought to evaluate how hsa_circ_0058122 affects SONFH in dexamethasone (DEX) treated human umbilical vein endothelial cells (HUVECs) model. METHODS: RT-PCR was used to demonstrate the hsa_circ_0058122 expression level in Dex-treated HUVECs cells. The effects of hsa_circ_0058122 on HUVECs apoptosis were evaluated via overexpression plasmid and siRNA. Using dual-luciferase and fluorescence in situ hybridization assays, we demonstrated that hsa_circ_0058122 binds to miR-7974 thereby facilitating HUVECs apoptosis. Bioinformatics analysis and western blot were performed to confirm target genes of hsa-miR-7974. RESULTS: In our previous work, we revealed the top 20 elevated circRNAs in SONFH patients were hsa_circ_0010027, hsa_circ_0058115, hsa_circ_0010026, hsa_circ_0058839, hsa_circ_0056886, hsa_circ_0056885, hsa_circ_0058146, hsa_circ_0058105, hsa_circ_0058112, hsa_circ_0058143, hsa_circ_0058102, hsa_circ_0058090, hsa_circ_0075353, hsa_circ_0058126, hsa_circ_0058130, hsa_circ_0058140, hsa_circ_0058122, hsa_circ_0058123, hsa_circ_0058103, and hsa_circ_0058121. Among these, hsa_circ_0058122 was finally selected for further investigation. We found hsa_circ_0058122 expression was markedly elevated in Dex-treated HUVECs cells, and the Dex-mediated HUVEC apoptosis was impaired in hsa_circ_0058122-silenced cells and increased in hsa_circ_0058122-overexpressing cells. hsa_circ_0058122 competitively binds to hsa-miR-7974, which in turn interacts with insulin-like growth factor binding protein 5 (IGFBP5). CONCLUSIONS: hsa_circ_0058122/miR-7974/IGFBP5 was proposed to be a key regulatory pathway for SONFH. DEX treatment upregulated hsa_circ_0058122 expression in HUVECs, which sponged miR-7974, thereby increasing IGFBP5 expression, the hsa_circ_0058122/miR-7974/IGFBP5 axis contributed to the Dex-mediated apoptosis. These findings may identify novel targets for SONFH molecular therapy.

Identification of the Fosl1/AMPK/autophagy axis involved in apoptotic and inflammatory effects following spinal cord injury.

Strategies for reducing spinal cord injury (SCI) have become a research focus because an effective treatment of SCI is unavailable. The objective of this study was to explore the underlying mechanisms of Fosl1 following SCI. Based on the analysis of the Gene Expression Omnibus (GEO) database, Fosl1 was found to be highly enhanced in SCI. This result was confirmed in our animal model, and Fosl1 was found to be obviously expressed in neurons. Next, we treated PC-12 cells with H2O2 to mimic injured neurons and further verified that Fosl1 silencing upregulated AMPK expression, promoted autophagy and inhibited inflammation and apoptosis. Subsequently, a special inhibitor of AMPK was used to examine the role of AMPK, and we learned that the inhibition of AMPK suppressed autophagy and promoted inflammation and apoptosis following Fosl1 silencing. These changes completely reversed the beneficial effects of Fosl1 silencing on injured PC-12 cells. Moreover, treatment with an AMPK activator resulted in effects that were opposite those of the inhibitor. Finally, rats were injected intrathecally with si-Fosl1 to detect its role in vivo. The results showed that si-Fosl1 improved neurological function and decreased apoptosis and inflammation at 14 d postoperation, and the activator further benefited the rats of si-Fosl1 treatment. In conclusion, Fosl1 inhibits autophagy and promotes inflammation and apoptosis through the AMPK signaling pathway following SCI in vivo and in vitro.

Therapeutic effect of neohesperidin on TNF-α-stimulated human rheumatoid arthritis fibroblast-like synoviocytes.

During the pathogensis of rheumatoid arthritis (RA), activated RA fibroblast-like synoviocytes (RA-FLSs) combines similar proliferative features as tumor and inflammatory features as osteoarthritis, which eventually leads to joint erosion. Therefore, it is imperative to research and develop new compounds, which can effectively inhibit abnormal activation of RA-FLSs and retard RA progression. Neohesperidin (Neo) is a major active component of flavonoid compounds with anti-inflammation and anti-oxidant properties. In this study, the anti-inflammation, anti-migration, anti-invasion, anti-oxidant and apoptosis-induced effects of Neo on RA-FLSs were explored to investigate the underlying mechanism. The results suggested that Neo decreased the levels of interleukin IL-1ß, IL-6, IL-8, TNF-α, MMP-3, MMP-9 and MMP-13 in FLSs. Moreover, Neo blocked the activation of the MAPK signaling pathway. Furthermore, treatment with Neo induced the apoptosis of FLSs, and inhibited the migration of FLSs. It was also found that Neo reduced the accumulation of reactive oxygen species (ROS) induced by TNF-α. Taken together, our results highlighted that Neo may act as a potential and promising therapeutic drug for the management of RA.

Bioinformatics-based study to identify immune infiltration and inflammatory-related hub genes as biomarkers for the treatment of rheumatoid arthritis.

Rheumatoid arthritis (RA) is a systemic autoimmune disease whose principal pathological change is aggressive chronic synovial inflammation; however, the specific etiology and pathogenesis have not been fully elucidated. We downloaded the synovial tissue gene expression profiles of four human knees from the Gene Expression Omnibus database, analyzed the differentially expressed genes in the normal and RA groups, and assessed their enrichment in functions and pathways using bioinformatics methods and the STRING online database to establish protein-protein interaction networks. Cytoscape software was used to obtain 10 hub genes; receiver operating characteristic (ROC) curves were calculated for each hub gene and differential expression analysis of the two groups of hub genes. The CIBERSORT algorithm was used to impute immune infiltration. We identified the signaling pathways that play important roles in RA and 10 hub genes: Ccr1, Ccr2, Ccr5, Ccr7, Cxcl5, Cxcl6, Cxcl13, Ccl13, Adcy2, and Pnoc. The diagnostic value of these 10 hub genes for RA was confirmed using ROC curves and expression analysis. Adcy2, Cxcl13, and Ccr5 are strongly associated with RA development. The study also revealed that the differential infiltration profile of different inflammatory immune cells in the synovial tissue of RA is an extremely critical factor in RA progression. This study may contribute to the understanding of signaling pathways and biological processes associated with RA and the role of inflammatory immune infiltration in the pathogenesis of RA. In addition, this study shows that Adcy2, Cxcl13, and Ccr5 have the potential to be biomarkers for RA treatment.

Identification of Regeneration and Hub Genes and Pathways at Different Time Points after Spinal Cord Injury.

Spinal cord injury (SCI) is a neurological injury that can cause neuronal loss around the lesion site and leads to locomotive and sensory deficits. However, the underlying molecular mechanisms remain unclear. This study aimed to verify differential gene time-course expression in SCI and provide new insights for gene-level studies. We downloaded two rat expression profiles (GSE464 and GSE45006) from the Gene Expression Omnibus database, including 1 day, 3 days, 7 days, and 14 days post-SCI, along with thoracic spinal cord data for analysis. At each time point, gene integration was performed using "batch normalization." The raw data were standardized, and differentially expressed genes at the different time points versus the control were analyzed by Gene Ontology enrichment analysis, the Kyoto Encyclopedia of Genes and Genomes pathway analysis, and gene set enrichment analysis. A protein-protein interaction network was then built and visualized. In addition, ten hub genes were identified at each time point. Among them, Gnb5, Gng8, Agt, Gnai1, and Psap lack correlation studies in SCI and deserve further investigation. Finally, we screened and analyzed genes for tissue repair, reconstruction, and regeneration and found that Anxa1, Snap25, and Spp1 were closely related to repair and regeneration after SCI. In conclusion, hub genes, signaling pathways, and regeneration genes involved in secondary SCI were identified in our study. These results may be useful for understanding SCI-related biological processes and the development of targeted intervention strategies.

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.

Utilizing benchmarked dataset and gene regulatory network to investigate hub genes in postmenopausal osteoporosis.

OBJECTIVE: The objective of this paper was to investigate hub genes of postmenopausal osteoporosis (PO) utilizing benchmarked dataset and gene regulatory network (GRN). MATERIALS AND METHODS: To achieve this goal, the first step was to benchmark the dataset downloaded from the ArrayExpress database by adding local noise and global noise. Second, differentially expressed genes (DEGs) between PO and normal controls were identified using the Linear Models for Microarray Data package based on benchmarked dataset. Third, five kinds of GRN inference methods, which comprised Zscore, GeneNet, context likelihood of relatedness (CLR) algorithm, Partial Correlation coefficient with Information Theory (PCIT), and GEne Network Inference with Ensemble of trees (Genie3), were described and evaluated by receiver operating characteristic (ROC) and precision and recall (PR) curves. Finally, GRN constructed according to the method with best performance was implemented to conduct topological centrality (closeness) for the purpose of investigate hub genes of PO. RESULTS: A total of 236 DEGs were obtained based on benchmarked dataset of 20,554 genes. By assessing Zscore, GeneNet, CLR, PCIT, and Genie3 on the basis of ROC and PR curves, Genie3 had a clear advantage than others and was applied to construct the GRN which was composed of 236 nodes and 27,730 edges. Closeness centrality analysis of GRN was carried out, and we identified 14 hub genes (such as TTN, ACTA1, and MYBPC1) for PO. CONCLUSION: In conclusion, we have identified 14 hub genes (such as TN, ACTA1, and MYBPC1) based on benchmarked dataset and GRN. These genes might be potential biomarkers and give insights for diagnose and treatment of PO.

Erythropoietin-Induced Autophagy Protects Against Spinal Cord Injury and Improves Neurological Function via the Extracellular-Regulated Protein Kinase Signaling Pathway.

The objective of this study was to explore the neuroprotective molecular mechanisms of erythropoietin (EPO) in rats following spinal cord injury (SCI). First, a standard SCI model was established. After drug or saline treatment was administered, locomotor function was evaluated in rats using the Basso, Beattie, and Bresnahan (BBB) locomotor rating scale. H&E, Nissl, and TUNEL staining were performed to assess the ratio of cavities, number of motor neurons, and apoptotic cells in the damaged area. The relative protein and mRNA expressions were examined using western blot and qRT-PCR analyses, and the inflammatory markers, axon special protein, and neuromuscular junctions (NMJs) were detected by immunofluorescence. Both doses of EPO notably improved locomotor function, but high-dose EPO was more effective than low-dose EPO. Moreover, EPO reduced the cavity ratio, cell apoptosis, and motor neuron loss in the damaged area, but enhanced the autophagy level and extracellular-regulated protein kinase (ERK) activity. Treatment with an ERK inhibitor significantly prevented the effect of EPO on SCI, and an activator mimicked the benefits of EPO. Further investigation revealed that EPO promoted SCI-induced autophagy via the ERK signaling pathway. EPO activates autophagy to promote locomotor function recovery in rats with SCI via the ERK signaling pathway.

Microarray profiling of circular RNAs in steroid-associated osteonecrosis of the femoral head: Observational study.

The aim of this study was to elucidate the molecular mechanisms and to identify the differential expression of circular RNAs (circRNAs) for steroid-associated osteonecrosis of the femoral head (SONFH) using bioinformatics analysis.circRNA microarray was performed with 3 SONFH tissues and the adjacent normal tissues, and differentially expressed circRNA were identified by limma package in R. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed using the Database for Annotation, Visualization and Integrated Discovery database. In addition, a differentially expressed genes (DEG)-associated circRNA/microRNA (miRNA) interaction was predicted by combination of TargetScan and miRanda, and the circRNA/miRNA interaction network generated by the cytoscape software.A total of 647 differentially expressed circRNAs, including 433 upregulated and 214 downregulated circRNA were identified. The most enriched GO terms for upregulated and downregulated circRNA were extracellular matrix organization and leukocyte activation in biological process; extracellular matrix and spindle pole in cellular component; integrin binding and ATP binding in molecular function, and KEGG pathway enrichment analyses showed that the upregulated and downregulated circRNA were strongly associated with Protein digestion and absorption and Cell cycle. Moreover, a total of 212 differentially expressed messenger RNAs (mRNAs), including 113 upregulated and 99 downregulated genes were identified. In addition, from the analysis of miRNA, long noncoding RNAs, mRNA, and circRNA networks, we found that hsa_circ_0008136 and hsa_circ_0074758 were respectively the upregulated and downregulated circRNA with highest degrees.The identified circRNA and mRNA could be implicated in the progression of human SONFH. The findings could lead to a better understanding of SONFH pathogenesis.

Evodiamine attenuates adjuvant-induced arthritis in rats by inhibiting synovial inflammation and restoring the Th17/Treg balance.

OBJECTIVES: Evodiamine (Evo) possesses strong anti-inflammatory activity. In this study, we determine the antiarthritic effect of Evo. METHODS: Evo was administered to rats with adjuvant-induced arthritis (AA). We evaluated arthritis symptoms & histopathological changes and measured inflammatory cell infiltration, pro-inflammatory cytokine production and Th17 & Treg percentages in arthritic rats. KEY FINDINGS: Evo significantly improved the clinical signs of AA in rats, including decreases in paw swelling, the polyarthritis index and the number of swollen paw joints. Based on the histopathological analysis, Evo improved synovial inflammation and bone injury by inhibiting inflammatory cell infiltration, synoviocyte proliferation, pannus formation and cartilage erosion. Furthermore, the numbers of synovial CD3+ or CD68+ inflammatory cells were reduced, and the elevated levels of tumour necrosis factor-α, interleukin-1ß (IL-1ß) and IL-6 were restored to control levels by the Evo treatment. In addition, Evo therapy regulated the abnormal differentiation of Treg and Th17 cells, decreasing IL-17 production and increasing IL-10 levels. Finally, Evo inhibited Stat3 phosphorylation and induced Stat5 phosphorylation in rats with AA. CONCLUSIONS: Based on our results, Evo is a promising antiarthritic agent, potentially due to its inhibitory effect on synovial inflammation and regulatory effects on Treg and Th17 differentiation.

Knockdown of SGK1 alleviates the IL-1ß-induced chondrocyte anabolic and catabolic imbalance by activating FoxO1-mediated autophagy in human chondrocytes.

Osteoarthritis (OA) is a common joint disease characterized by the progressive degeneration of articular cartilage with no effective treatment methods available. Cartilage degeneration is closely related to an anabolic and catabolic imbalance in chondrocytes, and accumulating evidence has revealed that autophagy is a crucial protective mechanism that maintains the balance of anabolic and catabolic activities. Therefore, studies aiming to identify additional genes that regulate autophagy as a promising therapeutic strategy for OA are needed. In this study, we analyzed the GSE113825 datasets from Gene Expression Omnibus and validated that serum- and glucocorticoid-regulated kinase 1 (SGK1) was upregulated in OA cartilage. Based on the results from loss-of-function studies, SGK1 silencing promoted the deposition of glycosaminoglycans in interleukin 1 beta (IL-1ß)-treated chondrocytes, and significantly alleviated IL-1ß-induced downregulation of Collagen II and Aggrecan, as well as the upregulation of a disintegrin and metalloproteinase with thrombospondin motifs 5 and matrix metalloproteinase-13. Furthermore, SGK1 knockdown reversed the IL-1ß-induced chondrocyte anabolic and catabolic imbalance by activating autophagy. Moreover, SGK1 directly bound to forkhead box protein O1 (FoxO1) and increased its phosphorylation, which in turn resulted in its translocation from the nucleus. The decreased FoxO1 levels led to a decrease in LC3-I/LC3-II conversion and Beclin-1 levels, subsequently inhibiting autophagosome formation and increasing P62 levels, thus indicating a downregulation of autophagy. Taken together, we identified a critical role of SGK1 in the IL-1ß-induced chondrocyte anabolic and catabolic imbalance, which may represent a potential novel therapeutic target for OA.

Proximal humeral internal locking plate combined with a custom neutral-position shoulder and elbow sling for proximal humerus fractures: A randomized study.

OBJECTIVES: The aim of this study was to investigate the effectiveness of the proximal humeral internal locking system (PHILOS) plate combined with a custom neutral-position shoulder and elbow sling for proximal humerus fractures. METHODS: A total of 112 patients with proximal humerus fractures were assigned randomly into 2 groups. Group A (n = 56) was treated by open reduction and internal fixation (ORIF) with a PHILOS plate; group B (n = 56) was treated by ORIF with a PHILOS plate in combination with the use of a custom neutral-position shoulder and elbow sling for 30 days after surgery. The incidence of internal fixation failure, the Constant-Murley shoulder assessment, and Visual Analogue Scale (VAS) score were recorded and analyzed. RESULTS: Patients included were followed up for an average of 15 months (range, 6-24 months). No significant differences were observed in mean VAS scores and mean Constant-Murley shoulder assessment scores at 1-day preoperative and postoperative day 3 between groups A and B. However, mean VAS scores and mean Constant-Murley shoulder assessment in group B were significantly improved when compared with group A at postoperative day 30 and the final follow-up. No cases of postoperative infection, loss of reduction, PHILOS break, or vascular nerve injury occurred in either group. CONCLUSIONS: Proximal humerus fractures treated with the combination of the PHILOS and custom neutral-position shoulder and elbow sling for 30 days after operation was associated with a lower incidence of internal fixation failure. There was no increase in adverse events compared with open reduction and internal fixation with a PHILOS plate alone.

Endothelial progenitor cell-conditioned medium promotes angiogenesis and is neuroprotective after spinal cord injury.

Endothelial progenitor cells secrete a variety of growth factors that inhibit inflammation, promote angiogenesis and exert neuroprotective effects. Therefore, in this study, we investigated whether endothelial progenitor cell-conditioned medium might have therapeutic effectiveness for the treatment of spinal cord injury using both in vitro and in vivo experiments. After primary culture of bone marrow-derived macrophages, lipopolysaccharide stimulation was used to classically activate macrophages to their proinflammatory phenotype. These cells were then treated with endothelial progenitor cell-conditioned medium or control medium. Polymerase chain reaction was used to determine mRNA expression levels of related inflammatory factors. Afterwards, primary cultures of rat spinal cord neuronal cells were prepared and treated with H2O2 and either endothelial progenitor cell-conditioned medium or control medium. Hoechst 33258 and propidium iodide staining were used to calculate the proportion of neurons undergoing apoptosis. Aortic ring assay was performed to assess the effect of endothelial progenitor cell-conditioned medium on angiogenesis. Compared with control medium, endothelial progenitor cell-conditioned medium mitigated the macrophage inflammatory response at the spinal cord injury site, suppressed apoptosis, and promoted angiogenesis. Next, we used a rat model of spinal cord injury to examine the effects of the endothelial progenitor cell-conditioned medium in vivo. The rats were randomly administered intraperitoneal injection of PBS, control medium or endothelial progenitor cell-conditioned medium, once a day, for 6 consecutive weeks. Immunohistochemistry was used to observe neuronal morphology. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay was performed to detect the proportion of apoptotic neurons in the gray matter. The Basso, Beattie and Bresnahan Locomotor Rating Scale was used to evaluate the recovery of motor function of the bilateral hind limbs after spinal cord injury. Compared with the other two groups, the number of axons was increased, cavities in the spinal cord were decreased, the proportion of apoptotic neurons in the gray matter was reduced, and the Basso, Beattie and Bresnahan score was higher in the endothelial progenitor cell-conditioned medium group. Taken together, the in vivo and in vitro results suggest that endothelial progenitor cell-conditioned medium suppresses inflammation, promotes angiogenesis, provides neuroprotection, and promotes functional recovery after spinal cord injury.

CTHRC1 mediates IL­1ß­induced apoptosis in chondrocytes via JNK1/2 signaling.

Osteoarthritis (OA), also known as degenerative joint disease or degenerative arthritis, is characterized by chondrocyte apoptosis. The aim of the present study was to investigate the effects of collagen triple helix repeat containing 1 (CTHRC1) and the c­Jun N­terminal kinase (JNK) 1/2 inhibitor SP600125 on rat chondrocytes cultured in vitro with interleukin (IL)­1ß. Chondrocytes were treated with different doses of IL­1ß and cell viability and CTHRC1 expression were assessed using Cell Counting Kit­8 and western blot assays, respectively. In separate experiments, chondrocytes were treated with CTHRC1­expressing constructs (pLVX­Puro­CTHRC1) and/or SP600125, or IL­1ß with either CTHRC1 short hairpin (sh)RNA constructs (shNRA­CTHRC1) or SP600125. The expression of CTHRC1, B­cell lymphoma (Bcl)­2, Bcl­2­associated X protein (Bax), cleaved caspase­3, poly ADP ribose polymerase (PARP)­1 and matrix metalloproteinase (MMP)­13 was measured using reverse transcription­quantitative polymerase chain reaction and western blotting assays. A Cell Counting Kit­8 assay was performed to examine cell viability. Annexin V/propidium iodide staining and flow cytometry assays were used to detect chondrocyte apoptosis. The expression of JNK1/2 and phosphorylated JNK1/2 was measured using western blotting. CTHRC1 was highly expressed in patients with OA compared with normal controls. IL­1ß treatment (5, 10 and 20 ng/ml) increased the protein expression of CTHRC1 in a dose­dependent manner and decreased the viability of chondrocytes in a time­dependent manner. pLVX­Puro­CTHRC1 mimics the effect of IL­1ß on chondrocyte apoptosis and JNK1/2 activity, and this is reversed by SP600125 treatment. However, transfection with shRNA­CTHRC1 or treatment with SP600125 inhibited IL­1ß­induced cell apoptosis and JNK1/2 activation. These results indicate that CTHRC1 downregulation may protect chondrocytes from IL­1ß­induced apoptosis by inactivating the JNK1/2 pathway.