[Correlation between combined deflection angle classification adduction typing and complications after internal fixation of adduction femoral neck fracture].

Objective: To analyze the correlation between postoperative complications and combined deflection angle classification adduction type (CDAC-ADT) of femoral neck fractures after cannulated screw internal fixation. Methods: The clinical data of 121 patients with CDAC-ADT femoral neck fracture admitted between January 2018 and December 2021 and met the selected criteria were retrospectively analyzed. There were 69 males and 52 females, the age ranged from 19 to 79 years (mean, 48.1 years). The causes of injury included 52 cases of traffic accident, 24 cases of falling from height, and 45 cases of fall. The time from injury to operation ranged from 2 to 12 days, with an average of 6.0 days. Among them, there were 18 cases of CDAC-ADT type Ⅰ, 46 cases of type Ⅱ, and 57 cases of type Ⅲ; 6 cases of Garden type Ⅱ, 103 cases of type Ⅲ, and 12 cases of type Ⅳ; and according to the location of the fracture line, there were 26 cases of subcapitate type, 88 cases of transcervical type, and 7 cases of basal type. All patients were treated with cannulated screw internal fixation. The occurrence of complications (including internal fixation failure, fracture nonunion, and osteonecrosis of the femoral head) was recorded, and the correlation between complications and CDAC-ADT typing, Garden typing, and fracture line location were analyzed. Results: The patients were followed up 8-44 months, with a mean of 24.9 months. There were 10 cases of internal fixation failure, 7 cases of fracture nonunion, and 30 cases of osteonecrosis of the femoral head after operation. Correlation analysis showed that patients' CDAC-ADT typing was significantly correlated with the overall incidence of complication and the incidence of internal fixation failure, fracture nonunion, and osteonecrosis of the femoral head ( P<0.05), and the Pearson coefficient of contingency were 0.435, 0.251, 0.254, and 0.241, respectively. Garden typing did not correlate with the overall incidence of complication and the incidence of internal fixation failure and fracture nonunion ( P>0.05), but correlated with the incidence of osteonecrosis of the femoral head ( P<0.05), and the Pearson coefficient of contingency was 0.251. Fracture line position typing had no correlation with the overall incidence of complication and the incidence of internal fixation failure, fracture nonunion, and osteonecrosis of the femoral head ( P>0.05). Conclusion: CDAC-ADT typing has obvious correlation with postoperative complications of femoral neck fracture and can be used to predict complications of femoral neck fracture.

An evidence-based guideline on treating lumbar disc herniation with traditional Chinese medicine.

BACKGROUND: Lumbar disc herniation (LDH), as one of the most common causes of lower back pain, imposes a heavy economic burden on patients and society. Conservative management is the first-line choice for the majority of LDH patients. Traditional Chinese medicine (TCM) is an important part of conservative treatment and has attracted more and more international attention. STUDY DESIGN: Evidence-based guideline. METHODS: We formed a guideline panel of multidisciplinary experts. The clinical questions were identified on the basis of a systematic literature search and a consensus meeting. We searched the literature for direct evidence on the management of LDH and assessed its certainty-generated recommendations using the grading of recommendations, assessment, development, and evaluation (GRADE) approach. RESULTS: The guideline panel made 20 recommendations, which covered the use of Shentong Zhuyu decoction, Shenzhuo decoction, Simiao San decoction, Duhuo Jisheng decoction, Yaobitong capsule, Yaotongning capsule, Osteoking, manual therapy, needle knife, manual acupuncture, electroacupuncture, Chinese exercise techniques (Tai Chi, Baduanjin, or Yijinjing), and integrative medicine, such as combined non-steroidal anti-inflammatory drugs, neural nutrition, and traction. Recommendations were either strong or weak, or in the form of ungraded consensus-based statement. CONCLUSION: This is the first LDH treatment guideline for TCM and integrative medicine with a systematic search, synthesis of evidence, and using the GRADE method to rate the quality of evidence. We hope these recommendations can help support healthcare workers caring for LDH patients.

Study of the curative effect of Zhang's Xibi formula and its underlying mechanism involving inhibition of inflammatory responses and delay of knee osteoarthritis.

OBJECTIVE: To verify the clinical efficacy of Zhang's Xibi formula (ZSXBF) and explain the mechanism underlying its therapeutic effect. METHODS: Preliminary elucidation of the clinical efficacy of ZSXBF in treating KOA in self-control studies, exploration of its mechanism of action with network pharmacology methods, and validation in animal experiments. RESULTS: In clinical studies, ZSXBF administration effectively improved patient quality of life and reduce pain. Network pharmacology was used to explore the possible mechanisms underlying its treatment effect, and after verification in clinical experience and animal experiments, it was found that ZSXBF regulated the expression of immune-related proteins such as IL-17, ERK1, and TP53 in mouse knee joints. CONCLUSION: ZSXBF, which is a traditional Chinese medicine compound that is used to clear heat and detoxify, can effectively improve the clinical symptoms of KOA patients, and its underlying mechanism includes the regulation of human immune-related proteins.

miR-126 mitigates the osteogenic differentiation of human bone marrow-derived mesenchymal stem cells by targeting the ERK1/2 and Bcl-2 pathways.

Human bone marrow mesenchymal stem cells (hBMMSCs) are a promising cell source for bone engineering owing to their high potential to differentiate into osteoblasts. The objective of the present study is to assess microRNA-126 (miR-126) and examine its effects on the osteogenic differentiation of hBMMSCs. In this study, we investigate the role of miR-126 in the progression of osteogenic differentiation (OD) as well as the apoptosis and inflammation of hBMMSCs during OD induction. OD is induced in hBMMSCs, and matrix mineralization along with other OD-associated markers are evaluated by Alizarin Red S (AR) staining and quantitative PCR (qPCR). Gain- and loss-of-function studies are performed to demonstrate the role of miR-126 in the OD of hBMMSCs. Flow cytometry and qPCR-based cytokine expression studies are performed to investigate the effect of miR-126 on the apoptosis and inflammation of hBMMSCs. The results indicate that miR-126 expression is downregulated during the OD of hBMMSCs. Gain- and loss-of function assays reveal that miR-126 upregulation inhibits the differentiation of hBMMSCs into osteoblasts, whereas the downregulation of miR-126 promotes hBMMSC differentiation, as assessed by the determination of osteogenic genes and alkaline phosphatase activity. Furthermore, the miR-126 level is positively correlated with the production of inflammatory cytokines and apoptotic cell death. Additionally, our results suggest that miR-126 negatively regulates not only B-cell lymphoma 2 (Bcl-2) expression but also the phosphorylation of extracellular signal­regulated protein kinase (ERK) 1/2. Moreover, restoring ERK1/2 activity and upregulating Bcl-2 expression counteract the miR-126-mediated suppression of OD in hBMMSCs by promoting inflammation and apoptosis, respectively. Overall, our findings suggest a novel molecular mechanism relevant to the differentiation of hBMMSCs into osteoblasts, which can potentially facilitate bone formation by counteracting miR-126-mediated suppression of ERK1/2 activity and Bcl-2 expression.

Combined Deflection Angle Classification: A Novel Typing System of Adult Femoral Neck Fracture.

OBJECTIVE: Femoral neck fracture (FNF) is a common clinical trauma with high mortality and disability rates. Furthermore, its incidence increases exponentially with increasing age. Existing classifications have some disadvantages. Thus, this study aimed to establish a novel typing system for FNF. METHODS: We retrospectively analyzed all adult patients with FNF admitted to our hospital between December 2015 and November 2017 for cannulated screw internal fixation. The study population was divided into the femoral varus offset group (VAR) and the valgus offset group (VAL). The data collected included sex, age, affected side, injury mode, body mass index, complications, pelvic incidence (PI), hip deflection angle (HDA), combined deflection angle (CDA), and neck shaft angle. Statistical analysis was conducted to determine the correlation between complications and deviation angles. A novel typing system was developed and compared with the Garden classification to detect its superiority. RESULTS: A total of 108 patients were recruited, with 59 patients in the VAR and 49 patients in the VAL groups. The incidence of complications in the VAR group was significantly higher than that in the VAL group (P < 0.05). Moreover, there were more male participants in the VAR group. Compared with the VAL group, the VAR group had significantly higher PI, HDA, and CDA (P < 0.05). The CDA classification (CDAC) was defined, with CDA as the main criterion and HDA as the supplementary criterion. Furthermore, there was a hierarchical correlation between the actual incidence of complications and the typing level, which was increased in CDAC but not in the Garden classification. This showed that CDAC was more accurate. CONCLUSION: A novel typing system, CDAC, for FNF was established, which was more accurate than the Garden classification. We suggest combining CDAC and Garden classifications for the preoperative diagnosis, treatment selection, and prognostic evaluation for patients with FNF.

MiRNA-320a-5p contributes to the homeostasis of osteogenesis and adipogenesis in bone marrow mesenchymal stem cell.

Objective: A number of miRNAs and their targets were dragged in the differentiation of bone marrow mesenchymal stem cells (BMSCs). We aimed to elaborate the underlying molecular mechanisms of miRNA-320a in the osteoblast and adipocyte differentiation. Methods: Trauma-induced osteonecrosis of the femoral head (TIONFH) and normal control samples (n = 10 for each group) were collected, followed by miRNA chip analysis to identify the differentially expressed miRNAs. H&E staining was used to observe the pathological development of TIONFH. Lentiviral vector was used for overexpression and inhibition of miRNA-320a in vitro. Quantitative real-time PCR (qPCR), Western blotting and immunohistochemistry staining were employed to determine the expression of interested genes at mRNA or protein level. Luciferase report assay was employed to determine the binding of miRNA-320a and RUNX2. Alkaline phosphatase (ALP) and Alizarin red staining were performed to observe the osteogenesis and Oil red O staining were conducted to visualize the adipogenesis. Results: Expression of miRNA-320a was up-regulated while RUNX2 expression was down-regulated in TIONFH than Normal control. Luciferase report assay confirmed that miRNA-320a directly targeted to the 3'UTR of RUNX2. miRNA-320a overexpression significantly declined the expressions of osteogenesis-related markers: RUNX2, OSTERIX, Collagen I, Osteocalcin and Osteopontin. ALP and Alizarin red staining confirmed the inhibition function of miRNA-320a in osteogenesis of BMSCs. miRNA-320a inhibition significantly decreased the expression of adipogenesis-related markers: AP2, C/EBPα, FABP4 and PPARγ. Oil Red O staining confirmed the miRNA-320a inhibition reduced adipogenesis of BMSCs. Conclusions: miRNA-320a inhibits osteoblast differentiation via targeting RUNX2 and promotes adipocyte differentiation of BMSCs.

SOCS1, the feedback regulator of STAT1/3, inhibits the osteogenic differentiation of rat bone marrow mesenchymal stem cells.

Our study showed that Signal transducer and activator of transcription (STAT)1 and STAT3 phosphorylation was firstly upregulated in the early stage of osteogenic differentiation (OD), and quickly eliminated in hours. Following with phosphorylation of STAT1/3, its downstream feedback regulator Suppressor of cytokine signaling 1 (SOCS1) protein also underwent a quick elevation. Further activation and deactivation of STAT1/3, by administrated with Colivelin and Nifuroxazide in Bone mesenchymal stem cells (BMSCs), increased and decreased SOCS1 expression, inhibited and promoted OD of BMSCs, respectively, as evidenced by Alizarin staining, alkaline phosphatase (ALP) activity, and determination of Run-related transcription factor 2 (RUNX2), Osteocalcin (OCN), ALP, and Bone sialoprotein (BSP). In addition, administration of Colivelin and Nifuroxazide caused and blocked inflammation and apoptosis of BMSCs. To further elucidate the role of STAT1/3-SOCS1 regulatory loop on OD of BMSCs, we overexpressed or silenced SOCS1 in BMSCs during OD. WB data showed that overexpression of SOCS1 repressed STAT1/3 phosphorylation, and knockdown of SOCS1 increased the phosphorylated STAT1/3. Further mechanism study showed that OD of BMSCs was elevated or reduced by SOCS1 overexpression or knockdown, respectively. The findings presenting indicated that the STAT1/3-SOCS1 axis may be exploited as an innovative strategy to enhance osteogenesis in regenerative medicine.

Long noncoding RNA XIST modulates microRNA-135/CREB1 axis to influence osteogenic differentiation of osteoblast-like cells in mice with tibial fracture healing.

Fracture healing is a complex event with the involvement of many cell systems, cytokines, as well as mRNAs. Herein, we report the interactions among long noncoding RNA X-inactive specific transcript (XIST)/microRNA-135 (miR-135)/cAMP response element-binding protein 1 (CREB1) axis during fracture healing. We observed increased expression of XIST in patients with long-term unhealed fracture by microarray analysis. Subsequently, a mouse model with tibial fracture and a cell model using osteoblast-like MC3T3-E1 cells were generated. The XIST overexpression during fracture healing decreased proliferation and differentiation of MC3T3-E1 cells, while silencing of XIST facilitated MC3T3-E1 cell growth. Furthermore, miR-135 targeted CREB1 and negatively regulated its expression. XIST acted as a sponge for miR-135, thereby upregulating CREB1 and promoting the activity of the TNF-α/RANKL pathway. Transfection of miR-135 inhibitor or CREB1 overexpression blocked the stimulating effects of XIST knockdown on MC3T3-E1 cell growth. Besides, specific inhibitors of the TNF-α/RANKL pathway reversed the repressive role of XIST in cell osteogenic differentiation. All in all, these findings suggest that XIST knockdown induces the differentiation of osteoblast-like cells via regulation of the miR-135/CREB1/TNF-α/RANKL axis. XIST, as a consequence, represents an attractive therapeutic strategy to accelerate fracture healing.

Aucubin prevents steroid-induced osteoblast apoptosis by enhancing autophagy via AMPK activation.

Steroid-induced osteoblast apoptosis is a crucial pathological process in steroid-induced osteonecrosis of the femoral head (SONFH). Autophagy can resist apoptosis and AMPK plays an important role in autophagy regulation. Aucubin from the small tree Eucommia ulmoides Oliv., which has a long history of use in orthopaedics and traumatology in Asian medicine, can promote bone formation, but whether it can slow or prevent steroid-osteoblast apoptosis is unclear. Therefore, we investigated the pathogenesis of SONFH and how the osteoblast responds to aucubin under the dexamethasone stimulation. In human femoral head osteonecrosis specimens, we found that the autophage and apoptosis level were increased, and the AMPK signalling was crucial to autophagy. We observed that aucubin could prevent dexamethasone-induced apoptosis in osteoblasts by enhancing the level of autophagy. Further, we confirmed that the regulatory effect of aucubin on autophagy and apoptosis was achieved by activating AMPK signalling. We have demonstrated a mechanism of disease progression and shown that aucubin could enhance autophagy through AMPK signalling to prevent osteoblast apoptosis. These findings provide a basis for the further investigation of the potential therapeutic role of aucubin in the SONFH.

Fascin1 mediated release of pro-inflammatory cytokines and invasion/migration in rheumatoid arthritis via the STAT3 pathway.

Rheumatoid arthritis (RA) is a chronic, multi-factorial disease characterized by Synovial hyperplasia, chronic inflammation, and autoimmune reaction. Fascin1 overexpression has been implicated in cancer, immune, and inflammatory diseases. However, the relationship between Fascin1 and rheumatoid arthritis (RA) has not yet been determined. We investigated whether Fascin1 could modulate pro-inflammatory cytokine secretion and the proliferation, apoptosis, and invasion/migration of fibroblast-like synoviocytes (RA-FLSs). Fascin 1 was suppressed with a short interfering (si)RNA approach. Functional analysis contained MTT assay, flow cytometry,Transwell™ assays, wound healing, Quantitative polymerase chain reaction and western blotting were used to detect cell proliferation,apoptosis ratio, invasion/ migration, the mRNA and protein expression of the realted markers, respectively. Overexpression of fascin1 was observed in RA-FLSs group compared with control group. Fascin1 expression positively correlated with changes in the expression of RA disease activity markers (RF, CRP, and DAB28, respectively). We also observed a significant positive correlation between Fascin1 and STAT3 mRNA levels in RA- FLSs.Fascin1 silencing attenuated the expression of pro-inflammatory cytokines; reduced FLS proliferation in vitro; and increased apoptosis ratio and bax, cleaved PARP, and caspase-3 expression. si- Fascin1 transfection delayed RA-FLS invasion/migration and reversed the epithelial- mesenchymal transition. These data suggest that Fascin1 exerts positive effects on the proliferation, cell cycle, and invasion/migration of RA-FLSs by activating signal transducer and activator of transcription 3 signaling.After all, Fascin1 contributed to RA development.

Inhibition of miR-93-5p promotes osteogenic differentiation in a rabbit model of trauma-induced osteonecrosis of the femoral head.

Trauma-induced osteonecrosis of the femoral head (TIONFH) is characterized by femoral head collapse accompanied by degenerative changes of the hip. We previously reported that miR-93-5p expression is abnormally high in patients with TIONFH, but the role of miR-93-5p in the TIONFH process remains unclear. Herein, we investigated the role of miR-93-5p in TIONFH in a rabbit model. Bone marrow mesenchymal stem cells (BMSCs) were used for both in vivo and in vitro experiments. A rabbit model of TIONFH was injected with BMSCs transfected with miR-93-5p inhibitor. In addition, both an miR-93-5p mimic and negative control were transfected into BMSCs. Expression of miR-93-5p was significantly increased in the model group compared with control samples. An miR-93-5p inhibitor induced the expression of bone morphogenetic protein 2 (BMP-2) and alkaline phosphatase. Furthermore, expression of osteogenesis-related markers (BMP-2, secreted phosphoprotein 1, RUNX family transcription factor 2 and Osterix) was higher in the miR-93-5p inhibitor group, as revealed by quantitative PCR and western blotting. In addition, in vitro experimentation revealed that an miR-93-5p mimic decreased BMP-2 and TNF receptor superfamily member 11b expression, but increased receptor activator of nuclear factor-kappaB ligand expression. In summary, the miR-93-5p inhibitor could promote osteogenic differentiation by increasing BMP-2 expression during the development of TIONFH. Thus, miR-93-5p may have potential as a therapeutic target for TIONF treatment.

Tumor-Associated Antigens (TAAs) for the Serological Diagnosis of Osteosarcoma.

Osteosarcoma (OS) is the most common form of malignant bone tumor found in childhood and adolescence. Although its incidence rate is low among cancers, the prognosis of OS is usually poor. Although some biomarkers, such as p53, have been identified in OS, the association between the biomarkers and clinical outcome is not well understood. Thus, it is necessary to establish a method to identify patients diagnosed with OS at an early stage. It is becoming obvious that anti-tumor-associated antigens (TAAs) autoantibodies (TAAbs) in sera could be used as serological biomarkers in the detection of many different types of cancers. This notion indicates that TAAbs are considered as immunological "sentinels" associated with tumorigenesis underlying molecular events. It provides new insights into the molecular and cellular biology of the differential diagnosis of cancers. What's more, it is reported that a customized TAA array could significantly increase the sensitivity/specificity. TAA arrays also have great application prospects in detecting cancer at an early stage, monitoring cancer progression, discovering new therapeutic targets, and designing personalized treatment. In this review, we provide an overview of the TAAs identified in OS as well as the possibility that TAAs and TAAbs system be used as biomarkers in the immunodiagnosis and prognosis of OS.

Glaucocalyxin A suppresses inflammatory responses and induces apoptosis in TNF-a-induced human rheumatoid arthritis via modulation of the STAT3 pathway.

The pathogenesis of rheumatoid arthritis (RA) is characterized by synoviocyte hyperplasia and proinflammatory cytokine secretion, as well as the destruction of cartilage and bone. Glaucocalyxin A (GLA) is an alkaloid derived from a Chinese medicinal plant that exhibits anti-inflammatory, anti-tumor and neuroprotective properties. We investigated the effects of GLA on RA-fibroblast-like synoviocytes (FLS cells), and collagen-induced arthritis (CIA), and further explored the underlying mechanisms. GLA inhibited TNF-a-induced RA-FLS proliferation, increased apoptotic ratios and upregulated levels of caspase-3, cleaved PARP, and Bax. GLA also inhibited the expression of IL-10, IL-1ß, and IL-6 in vitro. Levels of p-STAT3 were downregulated in a dose-dependent manner. Over-expression of STAT3 partly neutralized the GLA-mediated elevation of caspase-3 and cleaved PARP levels as well as the downregulation of IL-10, IL-1B and IL-6 expression levels. This suggests that GLA inactivated the STAT3 pathway. Furthermore, the production of inflammatory cytokines in RA-FLS and a CIA rat model were inhibited effectively by GLA. Taken together, our data suggest that GLA is a potential long-term therapeutic agent for patients with RA.

microRNA-935-modified bone marrow mesenchymal stem cells-derived exosomes enhance osteoblast proliferation and differentiation in osteoporotic rats.

AIMS: The involvement of several microRNAs (miRNAs) in osteogenic differentiation has been indicated recently. Also, exosomes, derived from different cells, could shuttle specific miRNAs to other cell systems. Nevertheless, the effect and mechanism of microRNA-935 (miR-935)-containing exosomes in osteoblasts remain basically unclear. The current work was set to inspect the relevance of bone marrow mesenchymal stem cells (BMSCs)-derived exosomes (BMSC-exo) carrying miR-935 to osteoporotic rats. METHODS: The extracted BMSCs and purchased osteoblasts were cultured, followed by exosome isolation and identification. After cell grouping, osteoblasts were co-cultured with BMSCs. CCK-8, alizarin red staining as well as ALP staining were performed to detect osteoblast proliferation and activity. The binding connection between miR-935 and signal transducer and activator of transcription 1 (STAT1) was measured by dual-luciferase reporter gene assays. The expression profiles of miR-935, STAT1 and osteoblast-related proteins were assessed by RT-qPCR and Western blot. A rat model with osteoporosis was induced, and the BMD, BV/TV, Tb.N, Tb.Th and Tb.Sp values in rat bone tissues were observed by Micro-CT. RESULTS: BMSC-exo inhibited STAT1 levels by the delivery of miR-935 into osteoblasts, while STAT1 silencing promoted ALP activity in osteoblasts and mineralized nodules. STAT1 was identified as a target gene of miR-935. Moreover, in vivo experiments showed that in ovariectomized rats, silencing of miR-935 significantly reduced BMD, BV/TV, Tb.N, Tb.Th and increased Tb.Sp. CONCLUSION: BMSC-exo carry miR-935 to promote osteoblast proliferation and differentiation through targeting STAT1.

CircRNA_25487 inhibits bone repair in trauma-induced osteonecrosis of femoral head by sponging miR-134-3p through p21.

We aimed to identify specific circular RNAs (circRNAs) involved in bone repair of trauma-induced osteonecrosis of femoral head (TIONFH) and to explore the potential mechanism. CircRNA sequencing on the blood sample collected from patients with and without TIONFH was performed to select cirRNAs that were significantly differentially expressed, followed by qRT-PCR confirmation. Furthermore, the functions of one selected circRNA and the potential mechanisms in bone repair of TIONFH were validated based on the bone marrow mesenchymal stem cells (BMSCs) and osteoclast-like cells (OLCs) through CCK-8, flow cytometry, transwell assay, luciferase reporter assay, and western blot. A total of 234 upregulated and 148 downregulated differentially expressed circRNAs were identified, and qRT-PCR showed that circRNA_25487 was significantly upregulated in the peripheral blood of TIONFH patients. Luciferase reporter assay confirmed the binding effect between miR-134-3p and circRNA_25487. CircRNA_25487 suppression and miR-134-3p overexpression could promote cell proliferation and invasion while inhibited apoptosis of BMSCs and OLCs. miR-134-3p could target p21. CircRNA_25487 inhibited bone repair in TIONFH by sponging miR-134-3p to upregulate the expression of p21.

Extracellular vesicle-associated repetitive element DNAs as candidate osteosarcoma biomarkers.

Osteosarcoma (OS) is the most common malignant bone tumor in children and young adults. Despite that high-risk factors have been identified, no test for early detection is available. This study aimed to identify circulating nucleic acid sequences associated with serum extracellular vesicle (EV) preparations at the time of OS diagnosis, as a step towards an OS early detection assay. Sequencing of small nucleic acids extracted from serum EV preparations revealed increased representation of diverse repetitive element sequences in OS patient versus control sera. Analysis of a validation cohort using qPCR of PEG-precipitated EV preparations revealed the over-representation of HSATI, HSATII, LINE1-P1, and Charlie 3 at the DNA but not RNA level, with receiver operating characteristic (ROC) area under the curve (AUC) ≥ 0.90. HSATI and HSATII DNAs co-purified with EVs prepared by precipitation and size exclusion chromatography but not by exosome immunocapture, indicative of packaging in a non-exosomal complex. The consistent over-representation of EV-associated repetitive element DNA sequences suggests their potential utility as biomarkers for OS and perhaps other cancers.