SDF-1α promotes subchondral bone sclerosis and aggravates osteoarthritis by regulating the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells.

BACKGROUND: Subchondral bone sclerosis is a major feature of osteoarthritis (OA), and bone marrow mesenchymal stem cells (BMSCs) are presumed to play an important role in subchondral bone sclerosis. Accumulating evidence has shown that stromal cell-derived factor-1α (SDF-1α) plays a key role in bone metabolism-related diseases, but its role in OA pathogenesis remains largely unknown. The purpose of this study was to explore the role of SDF-1α expressed on BMSCs in subchondral bone sclerosis in an OA model. METHODS: In the present study, C57BL/6J mice were divided into the following three groups: the sham control, destabilization of the medial meniscus (DMM), and AMD3100-treated DMM (DMM + AMD3100) groups. The mice were sacrificed after 2 or 8 weeks, and samples were collected for histological and immunohistochemical analyses. OA severity was assessed by performing hematoxylin and eosin (HE) and safranin O-fast green staining. SDF-1α expression in the OA model was measured using an enzyme-linked immunosorbent assay (ELISA), quantitative real-time polymerase chain reaction (q-PCR), and immunohistochemistry. Micro-CT was used to observe changes in subchondral bone in the OA model. CD44, CD90, RUNX2, and OCN expression in subchondral bone were measured using q-PCR and immunohistochemistry. In vitro, BMSCs were transfected with a recombinant lentivirus expressing SDF-1α, an empty vector (EV), or siRNA-SDF-1α. Western blot analysis, q-PCR, and immunofluorescence staining were used to confirm the successful transfection of BMSCs. The effect of SDF-1α on BMSC proliferation was evaluated by performing a CCK-8 assay and cell cycle analysis. The effect of SDF-1α on the osteogenic differentiation of BMSCs was assessed by performing alkaline phosphatase (ALP) and alizarin red S (ARS) staining. Cyclin D1, RUNX2 and OCN expression were measured using Western blot analysis, q-PCR, and immunofluorescence staining. RESULTS: SDF-1α expression in the DMM-induced OA model increased. In the DMM + AMD3100 group, subchondral bone sclerosis was alleviated, OA was effectively relieved, and CD44, CD90, RUNX2, and OCN expression in subchondral bone was decreased. In vitro, high levels of SDF-1α promoted BMSC proliferation and increased osteogenic differentiation. Cyclin D1, RUNX2, and OCN expression increased. CONCLUSION: The results of this study reveal a new molecular mechanism underlying the pathogenesis of OA. The targeted regulation of SDF-1α may be clinically effective in suppressing OA progression.

Dysregulated circulating microRNA-126 in chronic obstructive pulmonary disease: linkage with acute exacerbation risk, severity degree, and inflammatory cytokines.

BACKGROUND: MicroRNA-126 (miR-126) is engaged in respiratory diseases via regulating airway tissue injury and pulmonary inflammation, while its relation with chronic obstructive pulmonary disease (COPD) is not reported. The study aimed to evaluate the value of miR-126 for estimating COPD acute exacerbation risk and its relation to disease severity and inflammatory cytokines in COPD patients. METHODS: This study was a case-control study. Seventy acute exacerbation COPD (AECOPD) patients, 70 stable COPD (SCOPD) patients, and 70 healthy controls (HCs) were consecutively recruited. Plasma miR-126 expression was detected by reverse transcription quantitative polymerase chain reaction. Plasma tumor necrosis factor α (TNF-α), interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and interleukin-17 (IL-17) in COPD patients were further determined by enzyme-linked immunosorbent assay. RESULTS: MiR-126 was higher in AECOPD patients compared to SCOPD patients and HCs (both Padj  < 0.001). Receiver operating characteristic curves revealed miR-126 distinguished AECOPD patients from SCOPD patients (area under curve (AUC): 0.805, 95%CI: 0.733-0.877) and HCs (AUC: 0.884, 95%CI: 0.829-0.939) and also distinguished SCOPD from HCs (AUC = 0.656, 95%CI: 0.566-0.747). MiR-126 positively related to GOLD stage in both AECOPD patients (p < 0.001) and SCOPD patients (p < 0.001). Furthermore, miR-126 positively linked with TNF-α (p < 0.001), IL-1ß (p = 0.002), IL-6 (p = 0.009), and IL-17 (p < 0.001) levels in AECOPD patients; but miR-126 only positively related to TNF-α and IL-17 levels (all p < 0.050), instead of IL-1ß or IL-6 level (all p > 0.050) in SCOPD patients and HCs. CONCLUSION: Dysregulated circulating miR-126 not only relates to COPD susceptibility and its acute exacerbation risk but also links with disease severity and inflammatory cytokines in COPD patients.

Enhancement of angiogenic effect of co-transfection human NGF and VEGF genes in rat bone marrow mesenchymal stem cells.

The current study explored the feasibility and efficacy of co-transfection of the human nerve growth factor (NGF) and vascular endothelial growth factor 165 (VEGF165) genes in rat bone marrow mesenchymal stem cells (BMSCs). The obtained hNGF and vascular endothelial growth factor (VEGF) cDNAs were cloned into the pEGFP-C1 expression vector to construct the recombinant vectors. Co-transfection in rat BMSCs was performed and the expressions of both genes were detected by RT-PCR, Western blot, and enzyme-linked immunospecific assay. The biological activity of recombinant NGF and VEGF proteins was confirmed using the Chick Chorioallantoic Membrane (CAM) assay. NGF and VEGF genes could be expressed successfully in rat BMSCs. The recombinant NGF and VEGF from the rat BMSCs showed a more significant synergetic biological activity compared with single recombinant NGF or VEGF. These findings demonstrate that the co-transfection of hNGF+VEGF genes can enhance the angiogenic effect in vivo.

Recombinant expression of human nerve growth factor beta in rabbit bone marrow mesenchymal stem cells.

Nerve growth factor (NGF) is required for the differentiation and maintenance of sympathetic and sensory neurons. In the present study, the recombinant expression of human nerve growth factor beta (hNGF-ß) gene in rabbit bone marrow mesenchymal stem cells (rMSCs) was undertaken. Recombinant vector containing hNGF-ß was constructed and transferred into rMSCs, the expressions of the exogenous in rMSCs were determined by reverse transcriptase PCR (RT-PCR), ELISA and Western blot, whereas the biological activity of recombinant hNGF-ß was confirmed using PC12 cells and cultures of dorsal root ganglion neurons from chicken embryos. The results showed that the hNGF-ß gene expressed successfully in the rMSCs, a polypeptide with a molecular weight of 13.2 kDa was detected. The maximal expression level of recombinant hNGF-ß in rMSCs reached 126.8012 pg/10(6) cells, the mean concentration was 96.4473 pg/10(6) cells. The recombinant hNGF-ß in the rMSCs showed full biological activity when compared to commercial recombinant hNGF-ß.