microRNA-365 attenuated intervertebral disc degeneration through modulating nucleus pulposus cell apoptosis and extracellular matrix degradation by targeting EFNA3.

This present study is aimed to investigate the role of microRNA-365 (miR-365) in the development of intervertebral disc degeneration (IDD). Nucleus pulposus (NP) cells were transfected by miR-365 mimic and miR-365 inhibitor, respectively. Concomitantly, the transfection efficiency and the expression level of miRNA were detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Meanwhile, NP cells apoptosis was measured through propidium iodide (PI)-AnnexinV-fluorescein isothiocyanate (FITC) apoptosis detection kit. Subsequently, immunofluorescence (IF) staining was performed to assess the expression of collagen II, aggrecan and matrix metalloproteinase 13 (MMP-13). In addition, bioinformatic prediction and Luciferase reporter assay were used to reveal the target gene of miR-365. Finally, we isolated the primary NP cells from rats and injected NP-miR-365 in rat IDD models. The results showed that overexpression of miR-365 could effectively inhibit NP cells apoptosis and MMP-13 expression and upregulate the expression of collagen II and aggrecan. Conversely, suppression of miR-365 enhanced NP cell apoptosis and elevated MMP-13 expression, but decreased the expression of collagen II and aggrecan. Moreover, the further data demonstrated that miR-365 mediated NP cell degradation through targeting ephrin-A3 (EFNA3). In addition, the cells apoptosis and catabolic markers were increased in NP cells when EFNA3 upregulated. More importantly, the vivo data supported that miR-365-NP cells injection ameliorated IDD in rats models. miR-365 could alleviate the development of IDD by regulating NP cell apoptosis and ECM degradation, which is likely mediated by targeting EFNA3. Therefore, miR-365 may be a promising therapeutic avenue for treatment IDD through EFNA3.

Full-Endoscopic Posterior Lumbar Interbody Fusion with Epidural Anesthesia: Technical Note and Initial Clinical Experience with One-Year Follow-Up.

OBJECTIVE: The purpose of this study was to introduce and evaluate the early clinical outcomes of the full-endoscopic posterior lumbar interbody fusion (Endo-PLIF) technique with epidural anesthesia (EA) for single-segment lumbar degenerative diseases. METHODS: In this retrospective case series study, we explored the feasibility and effectiveness of the Endo-PLIF with EA for single-segment lumbar degenerative diseases. Between March 2018 and January 2019, a series of 24 patients with single-segment lumbar degenerative diseases underwent Endo-PLIF surgery and were followed up for a minimum of 12 months (15.21±2.27 months). Clinical outcomes including visual analog scale (VAS) scores for back and leg pain, Oswestry Disability Index (ODI) scores, and the Short Form-36 health survey questionnaire (SF-36) were evaluated preoperatively, and postoperatively at 3 days and at 3, 6, and 12-months. RESULTS: All patients underwent successful single-segment Endo-PLIF surgery. The mean operation time was 209.17±39.49 min, and average amount of bleeding was 43.33±14.87 mL. The VAS for lower extremity pain and back pain significantly improved at 3 days, and at 3, 6, 12 months compared with preoperative, respectively. The ODI scores decreased from 42.04±3.96 to 12.75±2.71 (P<0.001) at preoperative and 12 months postoperatively, respectively. The SF-36 Physical Component Scores (PCS) improved from 34.96±4.63 preoperatively to 52.08±6.05 (P<0.001) at 12 months postoperatively. Additionally, the SF-36 Mental Component Scores (MCS) improved from 39.38±5.70 at preoperative to 53.13±5.97 (P<0.001) at 12 months postoperatively. Two patients experienced dysesthesia, and one patient had a wound infection. CONCLUSION: Endo-PLIF with EA is a feasible and valuable technique for the treatment of single-segment lumbar degenerative diseases in selected patients.

Inhibition of both endplate nutritional pathways results in intervertebral disc degeneration in a goat model.

BACKGROUND: The vertebral endplate route was demonstrated to be the main pathway for nutrition to the intervertebral disc. However, it is still a controversial issue on whether the blocking of the endplate nutritional pathway could result in intervertebral disc degeneration (IDD) in animal models. The aim was therefore to investigate the effect of the inhibition of both endplate nutritional pathways by bone cement injection on the IDD in a goat model. METHODS: Two lumbar intervertebral discs (L2-3 and L3-4) in eight 24-month-old goats were blocked in both endplate nutritional pathways by cement injection, and the other two lumbar intervertebral discs (L1-2 and L4-5) remained intact as normal controls. Effective blocking area percentage in nucleus pulposus (NP) was calculated, and X-rays, magnetic resonance imaging (MRI), and histology studies were performed at 4, 12, 24, and 48 weeks after operation. RESULTS: The mean effective blocking area percentage was 60.7 ± 5.3%. Imaging examinations at the time of 48 weeks after blocking the endplate nutritional pathways showed obvious IDD, with larger disc height reduction and higher degrees of disc degeneration grading compared with the normal controls. Histological examinations including HE, Masson's trichrome, Sirius Red, and proteoglycan stainings also confirmed the degenerative changes of the blocked discs. CONCLUSIONS: The endplate nutritional route could be inhibited by blocking both endplate pathways with cement injection in a goat model. The severe inhibition in the endplate nutritional pathways may result in IDD.

TIGIT overexpression diminishes the function of CD4 T cells and ameliorates the severity of rheumatoid arthritis in mouse models.

Rheumatoid arthritis (RA) is an immune-mediated disease with a pathogenesis that involves CD4 T cell activation. Multiple immune regulatory molecules expressed on CD4(+) T cells were involved in RA pathogenesis. In this study, we investigated the role of T cell immunoglobulin and ITIM (immunoreceptor tyrosine-based inhibition motif) domain (TIGIT) in RA. The frequency of TIGIT-positive CD4(+) T cells in the synovial fluid (SF) of active RA patients was lower than that of inactive RA patients. And a negative correlation between RA disease activity and TIGIT expression was found. In CD4(+) T cells isolated from SF of active RA patients, TIGIT upregulation significantly decreased cell proliferation, as shown by MTT assay. TIGIT overexpression also significantly decreased the production of IFN-γ and IL-17, and increased that of IL-10, as determined by ELISA and qRT-PCR. In CD4(+) T cells isolated from SF of inactive RA patients, TIGIT was silenced by siRNA transfection. As expected, TIGIT knockdown resulted in an opposite effect on cell proliferation and the production of cytokines, including IFN-γ, IL-17 and IL-10. A RA mouse model was established using type II collagen induction. TIGIT was upregulated in RA mouse by lentivector infection. As expected, TIGIT overexpression in vivo significantly alleviated the disease severity and deceased the levels of anti-collagen II antibodies. TIGIT upregulation in the early stage was more effective to alleviate disease severity. Our data suggested the potential therapeutic role of TIGIT in RA patients.

MicroRNA-24 Regulates Osteogenic Differentiation via Targeting T-Cell Factor-1.

MicroRNAs (miRNAs) have been reported to have diverse biological roles in regulating many biological processes, including osteogenic differentiation. In the present study, we identified that miR-24 was a critical regulator during osteogenic differentiation. We found that overexpression of miR-24 significantly inhibited osteogenic differentiation, which decreased alkaline phosphatase activity, matrix mineralization and the expression of osteogenic differentiation markers. In contrast, inhibition of miR-24 exhibited an opposite effect. Furthermore, we delineated that miR-24 regulates post-transcriptionals of T-cell factor-1 (Tcf-1) via targeting the 3'-untranslated region (UTR) of Tcf-1 mRNA. MiR-24 was further found to regulate the protein expression of Tcf-1 in the murine osteoprogenitors cells and bone mesenchymal stem cells. Additionally, the positive effect of miR-24 suppression on osteoblast differentiation was apparently abrogated by Tcf-1 silencing. Taken together, our data suggest that miR-24 participates in osteogenic differentiation by targeting and regulating Tcf-1 expression in osteoblastic cells.

MiR-21 overexpression improves osteoporosis by targeting RECK.

Osteoporosis is a kind of metabolic bone disorder. MicroRNA-21 (miR-21) has been proven to play an important role in bone formation, whereas its role in osteoporosis is unclear. In the present study, miR-21 expression was inhibited by TNF-α in mesenchymal stem cells (MSCs). TNF-α induced cell apoptosis, and inhibited cell proliferation and differentiation of MSCs. Whereas the effect was reversed by miR-21 mimics. Expression of reversion-inducing cysteine-rich protein with Kazal motifs (RECK) which is a predicted target of miR-21 was inhibited by miR-21 mimics. A luciferase reporter gene assay showed that miR-21 directly bound to RECK 3'-UTR. The effect of TNF-α on MSCs was reversed by RECK siRNA which was consistent with miR-21 mimics. The expression of MT1-MMP was inhibited by TNF-α and enhanced by RECK siRNA and miR-21 mimics. For the in vivo study, an osteoporosis model (OVX) was established by bilateral oophorectomy in mice. The expression of miR-21 decreased and RECK increased in the OVX mice. When treated with lentiviral RECK shRNA, the osteocalcin concentration and alkaline phosphate activity of the OVX mice decreased. The bone mineral density of the right femur mid-diaphysis was improved by RECK shRNA. Collectively, miR-21 modulated the osteoporosis by targeting RECK. These results emphasize the role of miR-21 during osteoporosis and suggest RECK might be a new medical target for osteoporosis.