A diagnostic approach integrated multimodal radiomics with machine learning models based on lumbar spine CT and X-ray for osteoporosis.

INTRODUCTION: The aim of this analysis is to construct a combined model that integrates radiomics, clinical risk factors, and machine learning algorithms to diagnose osteoporosis in patients and explore its potential in clinical applications. MATERIALS AND METHODS: A retrospective analysis was conducted on 616 lumbar spine. Radiomics features were extracted from the computed tomography (CT) scans and anteroposterior and lateral X-ray images of the lumbar spine. Logistic regression (LR), support vector machine (SVM), and random forest (RF) algorithms were used to construct radiomics models. The receiver operating characteristic curve (ROC) was employed to select the best-performing model. Clinical risk factors were identified through univariate logistic regression analysis (ULRA) and multivariate logistic regression analysis (MLRA) and utilized to develop a clinical model. A combined model was then created by merging radiomics and clinical risk factors. The performance of the models was evaluated using ROC curve analysis, and the clinical value of the models was assessed using decision curve analysis (DCA). RESULTS: A total of 4858 radiomics features were extracted. Among the radiomics models, the SVM model demonstrated the optimal diagnostic capabilities and accuracy, with an area under the curve (AUC) of 0.958 (0.9405-0.9762) in the training cohort and 0.907 (0.8648-0.9492) in the test cohort. Furthermore, the combined model exhibited an AUC of 0.959 (0.9412-0.9763) in the training cohort and 0.910 (0.8690-0.9506) in the test cohort. CONCLUSION: The combined model displayed outstanding ability in diagnosing osteoporosis, providing a safe and efficient method for clinical decision-making.

Conditional knockout of the PDK-1 gene in osteoblasts affects osteoblast differentiation and bone formation.

Osteoblasts are the main functional cells of bone formation, and they are responsible for the synthesis, secretion, and mineralization of the bone matrix. Phosphatidylinositol-3-kinase/Akt is an important signaling pathway involved in the regulation of cell proliferation, death, and survival. Some studies have shown that 3-phosphoinositide-dependent protein kinase-1 (PDK-1) plays an important role in the phosphorylation of Akt. In the present study, an osteocalcin (OCN) promoter-driven Cre-LoxP system was established to specifically delete the PDK-1 gene in osteoblasts. It was found that the size and weight of PDK-1 conditional gene knockout (cKO) mice were significantly reduced. von Kossa staining and microcomputed tomography showed that the trabecular thickness, trabecular number, and bone volume were significantly decreased, whereas trabecular separation was increased, as compared with wide-type littermates, which were characterized by a decreased bone mass. A model of distal femoral defect was established, and it was found that cKO mice delayed bone defect repair. In osteoblasts derived from PDK-1 cKO mice, the alkaline phosphatase (ALP) secretion and ability of calcium mineralization were significantly decreased, and the expressions of osteoblast-related proteins, runt-related transcription factor 2, OCN, and ALP were also clearly decreased. Moreover, the phosphorylation level of Akt and downstream factor GSK3ß and their response to insulin-like growth factor-1 (IGF-1) decreased clearly. Therefore, we believe that PDK-1 plays a very important role in osteoblast differentiation and bone formation by regulating the PDK-1/Akt/GSK3ß signaling pathway.

PDK1 is important lipid kinase for RANKL-induced osteoclast formation and function via the regulation of the Akt-GSK3ß-NFATc1 signaling cascade.

Perturbations in the balanced process of osteoblast-mediated bone formation and osteoclast-mediated bone resorption leading to excessive osteoclast formation and/or activity is the cause of many pathological bone conditions such as osteoporosis. The osteoclast is the only cell in the body capable of resorbing and degrading the mineralized bone matrix. Osteoclast formation from monocytic precursors is governed by the actions of two key cytokines macrophage-colony-stimulating factor and receptor activator of nuclear factor-κB ligand (RANKL). Binding of RANKL binding to receptor RANK initiates a series of downstream signaling responses leading to monocytic cell differentiation and fusion, and subsequent mature osteoclast bone resorption and survival. The phosphoinositide-3-kinase (PI3K)-protein kinase B (Akt) signaling cascade is one such pathway activated in response to RANKL. The 3-phosphoinositide-dependent protein kinase 1 (PDK1), is considered the master upstream lipid kinase of the PI3K-Akt cascade. PDK1 functions to phosphorylate and partially activate Akt, triggering the activation of downstream effectors. However, the role of PDK1 in osteoclasts has yet to be clearly defined. In this study, we specifically deleted the PDK1 gene in osteoclasts using the cathepsin-K promoter driven Cre-LoxP system. We found that the specific genetic ablation of PDK1 in osteoclasts leads to an osteoclast-poor osteopetrotic phenotype in mice. In vitro cellular assays further confirmed the impairment of osteoclast formation in response to RANKL by PDK1-deficient bone marrow macrophage (BMM) precursor cells. PDK1-deficient BMMs exhibited reduced ability to reorganize actin cytoskeleton to form a podosomal actin belt as a result of diminished capacity to fuse into giant multinucleated osteoclasts. Notably, biochemical analyses showed that PDK1 deficiency attenuated the phosphorylation of Akt and downstream effector GSK3ß, and reduced induction of NFATc1. GSK3ß is a reported negative regulator of NFATc1. GSK3ß activity is inhibited by Akt-dependent phosphorylation. Thus, our data provide clear genetic and mechanistic insights into the important role for PDK1 in osteoclasts.

Salidroside promotes rat spinal cord injury recovery by inhibiting inflammatory cytokine expression and NF-κB and MAPK signaling pathways.

Spinal cord injury (SCI) is a public health problem in the world. The SCI usually triggers an excessive inflammatory response that brings about a secondary tissue wreck leading to further cellular and organ dysfunction. Hence, there is great potential of reducing inflammation for therapeutic strategies of SCI. In this study, we aim to investigate if Salidroside (SAD) exerts an anti-inflammatory effect and promotes recovery of motor function on SCI through suppressing nuclear factor-κB (NF-κB) and the mitogen-activated protein kinase (MAPK) pathways. In vitro, real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) were used to examine the inhibitory effect of SAD on the expression and release of interleukin-1ß (IL-1ß), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) activated by lipopolysaccharide (LPS) in astrocytes. In addition, SAD was found to inhibit NF-κB, p38 and extracellular-regulated protein kinases (ERK) signaling pathways by western blot analysis. Further, in vivo study showed that SAD was able to improve hind limb motor function and reduce tissue damage accompanied by the suppressed expression of inflammatory cytokines IL-1ß, IL-6, and TNF-α. Overall, SAD could reduce the inflammatory response and promote motor function recovery in rats after SCI by inhibiting NF-κB, p38, and ERK signaling pathways.

Artesunate inhibits RANKL-induced osteoclastogenesis and bone resorption in vitro and prevents LPS-induced bone loss in vivo.

Osteoclasts are multinuclear giant cells responsible for bone resorption in lytic bone diseases such as osteoporosis, arthritis, periodontitis, and bone tumors. Due to the severe side-effects caused by the currently available drugs, a continuous search for novel bone-protective therapies is essential. Artesunate (Art), the water-soluble derivative of artemisinin has been investigated owing to its anti-malarial properties. However, its effects in osteoclastogenesis have not yet been reported. In this study, Art was shown to inhibit the nuclear factor-κB ligand (RANKL)-induced osteoclastogenesis, the mRNA expression of osteoclastic-specific genes, and resorption pit formation in a dose-dependent manner in primary bone marrow-derived macrophages cells (BMMs). Furthermore, Art markedly blocked the RANKL-induced osteoclastogenesis by attenuating the degradation of IκB and phosphorylation of NF-κB p65. Consistent with the in vitro results, Art inhibited lipopolysaccharide (LPS)-induced bone resorption by suppressing the osteoclastogenesis. Together our data demonstrated that Art inhibits RANKL-induced osteoclastogenesis by suppressing the NF-κB signaling pathway and that it is a promising agent for the treatment of osteolytic diseases.

miR-136-5p Regulates the Inflammatory Response by Targeting the IKKß/NF-κB/A20 Pathway After Spinal Cord Injury.

BACKGROUND/AIMS: miR-136-5p participates in recovery after spinal cord injury (SCI) via an unknown mechanism. We investigated the mechanism underlying the involvement of miR-136-5p in the inflammatory response in a rat model of SCI. METHODS: Sprague-Dawley rat astrocytes were cultured in vitro to construct a reporter plasmid. Luciferase assays were used to detect the ability of miR-136-5p to target the IKKß and A20 genes. Next, recombinant lentiviral vectors were constructed, which either overexpressed miR-136-5p or inhibited its expression. The influence of miR-136-5p overexpression and miR-136-5p silencing on inflammation was observed in vivo in an SCI rat model. The expression of IL-1ß, IL-6, TNF-α, IFN-α, and related proteins (A20, IKKß, and NF-κB) was detected. RESULTS: In vitro studies showed that luciferase activity was significantly activated in the presence of the 3' untranslated region (UTR) region of the IKKß gene after stimulation of cells with miR-136-5p. However, luciferase activity was significantly inhibited in the presence of the 3'UTR region of the A20 gene. Thus, miR-136-5p may act directly on the 3'UTR regions of the IKKß and A20 genes to regulate their expression. miR-136-5p overexpression promoted the production of related cytokines and NF-κB in SCI rats and inhibited the expression of A20 protein. CONCLUSION: Overexpression of miR-136-5p promotes the generation of IL-1ß, IL-6, TNF-α, IFN-α, IKKß, and NF-κB in SCI rats but inhibits the expression of A20. Under these conditions, inflammatory cell infiltration into the rat spinal cord increases and injury is significantly aggravated. Silencing of miR-136-5p significantly reduces the protein expression results described after miR-136-5p overexpression and ameliorates the inflammatory cell infiltration and damage to the spinal cord. Therefore, miR-136-5p might be a new target for the treatment of SCI.

Monocrotaline Suppresses RANKL-Induced Osteoclastogenesis In Vitro and Prevents LPS-Induced Bone Loss In Vivo.

BACKGROUND/AIMS: Extensive osteoclast formation plays a critical role in bone diseases, including rheumatoid arthritis, periodontitis and the aseptic loosening of orthopedic implants. Thus, identification of agents that can suppress osteoclast formation and bone resorption is important for the treatment of these diseases. Monocrotaline (Mon), the major bioactive component of crotalaria sessiliflora has been investigated for its anti-cancer activities. However, the effect of Mon on osteoclast formation and osteolysis is not known. METHODS: The bone marrow macrophages (BMMs) were cultured with M-CSF and RANKL followed by Mon treatment. Then the effects of Mon on osteoclast differentiation were evaluated by counting TRAP (+) multinucleated cells. Moreover, effects of Mon on hydroxyapatite resorption activity of mature osteoclast were studied through resorption areas measurement. The involved potential signaling pathways were analyzed by performed Western blotting and quantitative real-time PCR examination. Further, we established a mouse calvarial osteolysis model to measure the osteolysis suppressing effect of Mon in vivo. RESULTS: In this study, we show that Mon can inhibit RANKL-induced osteoclast formation and function in a dose-dependent manner. Mon inhibits the expression of osteoclast marker genes such as tartrate-resistant acid phosphatase (TRAP) and cathepsin K. Furthermore, Mon inhibits RANKL-induced the activation of p38 and JNK. Consistent with in vitro results, Mon exhibits protective effects in an in vivo mouse model of LPS-induced calvarial osteolysis. CONCLUSION: Taken together our data demonstrate that Mon may be a potential prophylactic anti-osteoclastic agent for the treatment of osteolytic diseases caused by excessive osteoclast formation and function.

Molecular Mechanism of MiR-136-5p Targeting NF-κB/A20 in the IL-17-Mediated Inflammatory Response after Spinal Cord Injury.

BACKGROUND/AIMS: The pathophysiology of spinal cord injury (SCI) results in serious damage to the human body via an increase in the secondary biological processes imposed by activated astrocytes. Abnormal expression of microRNAs after SCI has become a potential research focus. However, the underlying mechanisms are poorly understood. METHODS: SCI models were established in rats using Allen's method, and the BBB scoring method was employed to assess locomotor function. Lentivirus was used to infect rat astrocytes and SCI rats. Real-time PCR and antibody chip were used to measure gene expression and cytokine secretion. Western blot analysis was employed to detect protein expression. HE staining was used to assess the histological changes in SCI. The immunohistochemical staining of A20 and p-NF-κB in SCI was also analyzed. RESULTS: The in vitro experiment showed that miR-136-5p up-regulated the expression of p-NF-κB by down-regulating the expression of A20 so that astrocytes produced inflammatory factors and chemokines. The in vivo experiment indicated that overexpressed miR-136-5p promoted the production of inflammatory factors, chemokines and p-NF-κB in SCI rats, whereas it inhibited the expression of A20 protein and increased inflammatory cell infiltration and injuries in the spinal cord. CONCLUSION: The current findings indicate that silencing miR-136-5p effectively decreased inflammatory factors and chemokines and protected the spinal cord via NF-κB/A20 signaling in vivo and in vitro. In contrast, overexpression of miR-136-5p had the opposite effect.

The Effects of miR-136-5p-Mediated Regulation of A20 in Astrocytes from Cultured Spinal Cord Cultured Cells In Vitro.

BACKGROUND/AIMS: This study focused on investigating the regulatory mechanism of miR-136-5p in mouse astrocytes stimulated with interleukin-17(IL-17). METHODS: C57BL/6 mouse astrocytes were stimulated with IL-17 (100ng/ml) for various periods of time (0-48 hours) and at various doses (0-200 ng), and the expression levels of inflammatory cytokine and chemokine genes (IL-6, TNF-α, MCP-1, MCP-5 and MIP-2) were then detected by real-time PCR. The expression of the A20 gene was measured with real-time PCR in cells that were stimulated with IL-17 (50 ng/ml) for various periods of time (0-48 hours). C57BL/6 mouse astrocytes were transfected with Ctrl-anti-miR-136-5p or LNA -anti-miR-136-5p for 48 h. Thereafter, the cells were stimulated with or without IL-17 (50ng/ml) for 6 h. The level of A20 protein (TNFα-induced protein 3, TNFAIP3) was detected by Western blot analysis. RESULTS: (1) Compared with the DMEM control group, within six hours, IL-17 stimulation significantly increased the expression levels of inflammatory cytokine and chemokine genes and clearly decreased the expression level of the A20 protein. (2) Without IL-17 stimulation, the expression level of the miR-136-5p gene was significantly decreased, whereas in the miR-136-5p-inhibition group, the A20 protein expression was elevated. IL-17 stimulation slightly decreased the expression of the A20 protein in the miR-136-5p-inhibition group, but it was still slightly higher than in the control group. CONCLUSION: This study demonstrated that miR-136-5p affected the expression of A20 in IL-17-stimulated astrocytes.

Vitamin D receptor gene polymorphisms and lumbar disc degeneration: a systematic review and meta-analysis.

PURPOSE: To examine the association between Vitamin D receptor (VDR) gene polymorphisms and lumbar disc degeneration (LDD) predisposition. METHODS: A comprehensive literature search was conducted to identify all the relevant studies. The allele/genotype frequencies were extracted from each study. We calculated the pooled odds ratios (ORs) and 95 % confidence intervals (CI) to assess the strength of the association between the VDR gene polymorphisms and LDD risk. Statistical analysis was performed using RevMan 5.31 software. RESULTS: A total of 23 case-control studies (1835 cases and 1923 controls) were included in this systematic review. For the TaqI (rs731236), FokI (rs2228570) and ApaI (rs7975232) polymorphisms of VDR gene, nine studies, seven studies, and five studies, were eventually included in the meta-analysis, respectively. There was no evidence that the VDR gene polymorphisms (TaqI, FokI, ApaI) had significant associations with LDD risk.(for TaqI allelic comparison, OR = 1.07, 95 % CI 0.81-1.40, p = 0.64; for FokI allelic comparison, OR = 1.23, 95 % CI 0.83-1.82, p = 0.31; for ApaI allelic comparison, OR = 0.79, 95 % CI 0.55-1.14, p = 0.20). For stratified analyses by ethnicity and study design, no significant associations were found in Caucasian population and Asian population, as well as the population-based studies and hospital-based studies under all genetic models. CONCLUSIONS: TaqI, FokI, and ApaI polymorphisms of VDR gene were not significantly associated with the predisposition of LDD. Large-scale and well-designed international studies are needed to further analyze this field.

The Effects of Interleukin-17 (IL-17)-Related Inflammatory Cytokines and A20 Regulatory Proteins on Astrocytes in Spinal Cord Cultured In Vitro.

BACKGROUND/AIMS: This study focused on investigating the expression of several inflammatory cytokines and chemokines, including regulatory proteins in the astrocytes of mice stimulated with IL-17. MATERIALS AND METHODS: The cultured astrocytes from the spinal cords of mice were stimulated with IL-17. The expression of interleukin-6 (IL-6), tumor necrosis factor (TNF), monocyte chemotactic protein-1/5 (MCP-1/5) and macrophage inflammatory protein-2 (MIP-2) stimulated with IL-17 (50 ng/ml) at different time points (3 h, 6 h, 12 h, 24 h and 48 h) were determined using real-time PCR and ELISA. The expressions of A20 (tumor necrosis factor α inducible protein 3, TNFAIP3) and NF-x03BA;B were examined using real-time PCR and western blotting. RESULTS: Compared with the control group, the mRNA expression levels of IL-6, TNF, MCP-1/5 and MIP-2 increased significantly at 6 h after IL-17 stimulation, while the protein expression levels also increased significantly and peaked at 12 h. The mRNA expression level of NF-x03BA;B increased and peaked at 6 h before gradually declining, while the expression of A20 decreased. The protein expression level of NF-x03BA;B increased and peaked at 12 h, while the expression A20 had an opposite response. CONCLUSION: The study showed that NF-x03BA;B may have an effect on the cytokines secreted by astrocytes after IL-17 stimulation. Moreover, both A20 and NF-x03BA;B could regulate the expression and secretion of inflammatory mediators.

The effects of α-zearalanol on the proliferation of bone-marrow-derived mesenchymal stem cells and their differentiation into osteoblasts.

The aim of this study was to explore the effects of α-zearalanol (α-ZAL) on the proliferation of mouse bone-marrow-derived mesenchymal stem cells (BMSCs) and their differentiation into osteoblasts. Six- to eight-week-old BALB/C mice were used either as recipients or as bone marrow donors. BMSCs were isolated and collected using a differential adhesion method, with use of 10 % fetal bovine serum and Iscove's modified Dulbecco's medium. After the third generation, the BMSCs were randomly placed into the following subgroups: a control group, an osteogenic medium (OM) group, a 17ß-estradiol group, an α-ZAL 10(-7) mol/L group, an α-ZAL 10(-6) mol/L group, and an α-ZAL 10(-5) mol/L group. Flow cytometry was used to identify the BMSCs collected from the bone marrow. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide test was performed, and markers of the osteoblasts were measured in the different subgroups. In addition, expression of osteoprotegerin and expression of receptor activator of nuclear factor κB ligand were examined using Western blot. In contrast to the control and OM groups, BMSCs in the α-ZAL groups exhibited long fusiform shapes, and contact inhibition was observed when the cells were closely packed. After induction, the BMSCs grew well and exhibited triangular, star, polygonal, or irregular shapes. Clumps and multiple cells were evident. The trends of the proliferation and differentiation for the control, OM, 17ß-estradiol, and α-ZAL groups were similar. Compared with the control and OM groups, in the α-ZAL groups the expression levels of alkaline phosphatase, procollagen type I N-terminal propeptide, bone morphogenetic protein 2, and osteocalcin were significantly increased (p < 0.05). In addition, α-ZAL inhibited osteoclastogenesis by increasing the expression of osteoprotegerin and decreasing the expression of nuclear factor κB ligand. In conclusion, α-ZAL can increase the proliferation of BMSCs and their differentiation into osteoblasts and can effectively suppress osteoclastogenesis.

Occipito-cervical fusion following gross total resection for the treatment of spinal extramedullary tumors in craniocervical junction: a retrospective case series.

BACKGROUND: Previous studies found that the facet joint of the C1 vertebra were removed (C1 facetectomy) before extirpation from the extramedullary tumor in craniocervical junction, leading to postoperative upper cervical instability or deformity. Occipito-cervical fusion (OCF) is a demanding and morbid surgical procedure, which can be used in such patients. This study is to analyze the clinical manifestation and surgical outcome of patients with craniocervical extramedullary tumor undergoing an extirpation of spinal tumors and OCF by one-stage posterior approach. METHODS: The surgical and clinical databases were searched for operative procedures that had been performed for patients with spinal extramedullary tumors in craniocervical junction at a single institution from January 2008 to July 2011. The following inclusion criteria were applied: (1) initial surgery for craniocervical extramedullary tumor, (2) gross total resection and occipito-cervical fusion had been performed, (3) minimum 2-year follow-up, and (4) no previous cervical spine surgery. Medical records included demographic characteristics, clinical assessment, and radiographic studies. Clinical outcomes before and after the surgery were assessed using Frankel grade and the Japanese Orthopaedic Association (JOA) score. Cervical sagittal alignment was evaluated by C0-2 angle and C2-7 angle based on X-ray. RESULTS: Nine patients were included in the study. Five patients had schwannoma, three patients had meningioma, and only one patient had neurofibroma. All cases were followed up for 24-42 months (average, 34.2 months). At the last follow-up, three patients improved from Frankel grade C to grade D, two patients from Frankel grade C to grade E, and one patient from Frankel grade D to grade E, while two patients remained stationary at the Frankel grade D. The JOA score of the eight patients were 9.0 (range, 6-17) before surgery and were 14.6 (range, 12-17) at the most recent follow-up (p < 0.05). The mean C0-2 angle and the mean C2-7 angle before surgery were 26.2 ± 5.3° and 17.4 ± 13.1°, respectively. At the end of follow-up, the mean C0-2 angle was 25.6 ± 4.8°, and the mean C2-7 angle decreased to 12.7 ± 10.9°. However, this trend did not reach statistical significance (p < 0.05). Two patients suffered from cerebrospinal fluid leaks postoperatively. All patients had a satisfactory fusion and did not exhibit a tumor recurrence during the follow-up period. CONCLUSIONS: OCF following gross total resection appears to be a useful surgical procedure for the craniocervical extramedullary tumors requiring C1 facetectomy and does not cause postoperative kyphosis of the upper cervical spine.

The role of IL-17 promotes spinal cord neuroinflammation via activation of the transcription factor STAT3 after spinal cord injury in the rat.

STUDY DESIGN: In this study, we investigated the role of IL-17 via activation of STAT3 in the pathophysiology of SCI. OBJECTIVE: The purpose of the experiments is to study the expression of IL-17 and related cytokines via STAT3 signaling pathways, which is caused by the acute inflammatory response following SCI in different periods via establishing an acute SCI model in rat. METHODS: Basso, Beattie, and Bresnahan hind limb locomotor rating scale was used to assess the rat hind limb motor function. Immunohistochemistry was used to determine the expression levels of IL-17 and p-STAT3 in spinal cord tissues. Western blotting analysis was used to determine the protein expression of p-STAT3 in spinal cord tissue. RT-PCR was used to analyze the mRNA expression of IL-17 and IL-23p19 in the spleen tissue. ELISA was used to determine the peripheral blood serum levels of IL-6, IL-21, and IL-23. RESULTS: Compared to the sham-operated group, the expression levels of IL-17, p-STAT3, IL-6, IL-21, and IL-23 were significantly increased and peaked at 24 h after SCI. The increased levels of cytokines were correlated with the SCI disease stages. CONCLUSION: IL-17 may play an important role in promoting spinal cord neuroinflammation after SCI via activation of STAT3.

Fecal microbiota transplantation regulates the microbiota-gut-spinal cord axis to promote recovery after spinal cord injury.

Spinal cord injury (SCI) is devastating for patients, and currently lacks effective treatments. Dysbiosis commonly occurs after SCI and has significant immunomodulatory effects, but its impact on recovery remains unclear. The current study investigated the effects and mechanisms of fecal microbiota transplantation (FMT) in SCI. FMT was administered in a rat model of SCI and spinal pathology, inflammatory cytokines, and gut microbiome composition were assessed. Flow cytometry identified a source of interleukin (IL)-17 in spinal cord tissues, and carboxyfluorescein succimidyl ester labeling tracked γδ T cell migration. In vitro coculture was used to analyze the regulatory mechanisms of γδ T cells. Seahorse analysis was used to profile dendritic cell (DC) metabolism. Here we show that FMT improved spinal pathology and dampened post-injury inflammation. It also corrected post-SCI dysbiosis, increasing levels of the beneficial bacterium Akkermansia. The therapeutic effects of FMT were mediated by IL-17 produced by γδ T cells. FMT regulated γδ T cells via DC-T regulatory cell interaction, and induced metabolic reprogramming in DCs. These findings suggest that FMT represents a promising therapeutic approach for SCI, with potential to target IL-17+ γδ T cells. Elucidating the interconnected pathways between microbiota, immunity, and the spinal cord may facilitate novel treatment strategies.

Short-chain fatty acids ameliorate spinal cord injury recovery by regulating the balance of regulatory T cells and effector IL-17+ γδ T cells. / çŸ­é“¾è„‚è‚ªé ¸é€šè¿‡è°ƒèŠ‚è°ƒèŠ‚æ€§T细胞和IL-17+ γδ T细胞的平衡来改善脊髓损伤的恢复.

Spinal cord injury (SCI) causes motor, sensory, and autonomic dysfunctions. The gut microbiome has an important role in SCI, while short-chain fatty acids (SCFAs) are one of the main bioactive mediators of microbiota. In the present study, we explored the effects of oral administration of exogenous SCFAs on the recovery of locomotor function and tissue repair in SCI. Allen's method was utilized to establish an SCI model in Sprague-Dawley (SD) rats. The animals received water containing a mixture of 150 mmol/L SCFAs after SCI. After 21 d of treatment, the Basso, Beattie, and Bresnahan (BBB) score increased, the regularity index improved, and the base of support (BOS) value declined. Spinal cord tissue inflammatory infiltration was alleviated, the spinal cord necrosis cavity was reduced, and the numbers of motor neurons and Nissl bodies were elevated. Enzyme-linked immunosorbent assay (ELISA), real-time quantitative polymerase chain reaction (qPCR), and immunohistochemistry assay revealed that the expression of interleukin (IL)|-10 increased and that of IL-17 decreased in the spinal cord. SCFAs promoted gut homeostasis, induced intestinal T cells to shift toward an anti-inflammatory phenotype, and promoted regulatory T (Treg) cells to secrete IL-10, affecting Treg cells and IL-17+ γδ T cells in the spinal cord. Furthermore, we observed that Treg cells migrated from the gut to the spinal cord region after SCI. The above findings confirm that SCFAs can regulate Treg cells in the gut and affect the balance of Treg and IL-17+ γδ T cells in the spinal cord, which inhibits the inflammatory response and promotes the motor function in SCI rats. Our findings suggest that there is a relationship among gut, spinal cord, and immune cells, and the "gut-spinal cord-immune" axis may be one of the mechanisms regulating neural repair after SCI.

Conditional knockout of PDK1 in osteoclasts suppressed osteoclastogenesis and ameliorated prostate cancer-induced osteolysis in murine model.

BACKGROUND: The development and maintenance of normal bone tissue is maintained by balanced communication between osteoblasts and osteoclasts. The invasion of cancer cells disrupts this balance, leading to osteolysis. As the only bone resorbing cells in vivo, osteoclasts play important roles in cancer-induced osteolysis. However, the role of 3-phosphoinositide-dependent protein kinase-1 (PDK1) in osteoclast resorption remains unclear. METHODS: In our study, we used a receptor activator of nuclear factor-kappa B (RANK) promoter-driven Cre-LoxP system to conditionally delete the PDK1 gene in osteoclasts in mice. We observed the effect of osteoclast-specific knockout of PDK1 on prostate cancer-induced osteolysis. Bone marrow-derived macrophage cells (BMMs) were extracted and induced to differentiate osteoclasts in vitro to explore the role of PDK1 in osteoclasts. RESULTS: In this study, we found that PDK1 conditional knockout (cKO) mice exhibited smaller body sizes when compared to the wild-type (WT) mice. Moreover, deletion of PDK1 in osteoclasts ameliorated osteolysis and rPDK1educed bone resorption markers in the murine model of prostate cancer-induced osteolysis. In vivo, we discovered that osteoclast-specific knockout of suppressed RANKL-induced osteoclastogenesis, bone resorption function, and osteoclast-specific gene expression (Ctsk, TRAP, MMP-9, NFATc1). Western blot analyses of RANKL-induced signaling pathways showed that conditional knockout of PDK1 in osteoclasts inhibited the early nuclear factor κB (NF-κB) activation, which consequently suppressed the downstream induction of NFATc1. CONCLUSION: These findings demonstrated that PDK1 performs an important role in osteoclastogenesis and prostate cancer-induced osteolysis by modulating the PDK1/AKT/NF-κB signaling pathway.

The role of α-zearalanol in reversing bone loss induced by ovarian hormone deficiency in rats.

To assess the ability of α-zearalanol (α-ZAL) to prevent bone loss in an ovariectomized (OVX) rat model of osteoporosis, α-ZAL was administered intragastrically to rats. After 35 days, the total body bone mineral density (BMD) was assessed in all rats. All sections were processed for immunohistochemistry and hematoxylin and eosin staining. One-way ANOVA and an LSD multiple-range test were used to determine the significant differences between groups. BMD was lower in the OVX and OVX + α-ZAL high-dose (OVX + High) groups compared to the sham-operated (Sham), OVX + 17ß-ethinylestradiol (OVX + E(2)), OVX + α-ZAL medium-dose (OVX + Medium) and OVX + α-ZAL low-dose (OVX + Low) groups (P < 0.05). Clear bone trabeculae arrangements were observed in the OVX + E(2,) OVX + Medium and OVX + Low groups. The expressions of bone morphogenetic proteins and basic fibroblast growth factor were up-regulated in the OVX + E(2), OVX + Medium and OVX + Low groups compared to the OVX and OVX + High groups (P < 0.05). The OVX + E(2), OVX + Medium and OVX + Low groups showed lower levels of bone Gla protein, bone alkaline phosphatase, tartrate-resistant acid phosphatase and tumor necrosis factor α expressions than the OVX and OVX + High groups (P < 0.05). The administration of α-ZAL to ovariectomized rats reverses bone loss and prevents osteoporosis.

Total flavonoids of hawthorn leaves protect spinal motor neurons via promotion of autophagy after spinal cord injury.

The death of spinal motor neurons (SMNs) after spinal cord injury (SCI) is a crucial cause, contributing to a permanent neurological deficit. Total flavonoids of hawthorn leaves (TFHL) have been confirmed to have potentially therapeutic for SCI. Nonetheless, the roles and mechanisms of TFHL in recovering neuromotor function and regenerating axons of SMNs have not been fully elucidated. In this study, TFHL was applied to treat rats with SCI and injured SMNs for 7 days. In vivo experiment, rats with SCI were evaluated by a BBB (Basso-Beattie-Bresnahan) score to assess their motor functional recovery. The morphology, microstructure, apoptosis, Nissl bodies, and autophagy of SMNs in spinal cord tissue were detected by Hematoxylin-eosin (HE) staining, transmission electron microscopy, TUNEL staining, Nissl staining, and immunohistochemistry respectively. In vitro experiment, the co-culture model of SMNs and astrocytes was constructed to simulate the internal environment around SMNs in the spinal cord tissue. The cell morphology, microstructure, axonal regeneration, and autophagy were observed via optical microscope, transmission electron microscopy, and immunofluorescence. The content of neurotrophic factors in the cell culture medium of the co-culture model was detected by ELISA. Moreover, the expression of axon-related and autophagy-related proteins in the spinal cord tissue and SMNs was measured by Western Blot. We demonstrated that TFHL improved the neuromotor function recovery in rats after SCI. We then found that TFHL significantly promoted injured spinal cord tissue repair, reduced apoptosis, and improved the functional status of neurons in spinal cord tissue in vivo. Meanwhile, the cell morphology, microstructure, and axonal regeneration of damaged SMNs also obviously were improved, and the secretion of neurotrophic factors was facilitated after treatment with TFHL in vitro. Further, we revealed that TFHL promoted autophagy and related protein expression in vivo and vitro. Taken together, our study suggested that TFHL might facilitate autophagy and have neuroprotective properties in SMNs to enhance the recovery of neuromotor function of rats with SCI.

Changes in the gut microbiota of osteoporosis patients based on 16S rRNA gene sequencing: a systematic review and meta-analysis.

BACKGROUND: Osteoporosis (OP) has become a major public health issue, threatening the bone health of middle-aged and elderly people from all around the world. Changes in the gut microbiota (GM) are correlated with the maintenance of bone mass and bone quality. However, research results in this field remain highly controversial, and no systematic review or meta-analysis of the relationship between GM and OP has been conducted. This paper addresses this shortcoming, focusing on the difference in the GM abundance between OP patients and healthy controls based on previous 16S ribosomal RNA (rRNA) gene sequencing results, in order to provide new clinical reference information for future customized prevention and treatment options of OP. METHODS: According to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA), we comprehensively searched the databases of PubMed, Web of Science, Embase, Cochrane Library, and China National Knowledge Infrastructure (CNKI). In addition, we applied the R programming language version 4.0.3 and Stata 15.1 software for data analysis. We also implemented the Newcastle-Ottawa Scale (NOS), funnel plot analysis, sensitivity analysis, Egger's test, and Begg's test to assess the risk of bias. RESULTS: This research ultimately considered 12 studies, which included the fecal GM data of 2033 people (604 with OP and 1429 healthy controls). In the included research papers, it was observed that the relative abundance of Lactobacillus and Ruminococcus increased in the OP group, while the relative abundance for Bacteroides of Bacteroidetes increased (except for Ireland). Meanwhile, Firmicutes, Blautia, Alistipes, Megamonas, and Anaerostipes showed reduced relative abundance in Chinese studies. In the linear discriminant analysis Effect Size (LEfSe) analysis, certain bacteria showed statistically significant results consistently across different studies. CONCLUSIONS: This observational meta-analysis revealed that changes in the GM were correlated with OP, and variations in some advantageous GM might involve regional differences.