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BACKGROUND: Intervertebral disc (IVD) degeneration is a condition characterized by a reduction in the water and extracellular matrix content of the nucleus pulposus (NP) and is considered as one of the dominating contributing factors to low back pain. Recent evidence suggests that stromal cell-derived factor 1α (SDF-1α) and its receptor C-X-C chemokine receptor type 4 (CXCR4) direct the migration of stem cells associated with injury repair in different musculoskeletal tissues. AIM: To investigate the effects of SDF-1α on recruitment and chondrogenic differentiation of nucleus pulposus-derived stem cells (NPSCs). METHODS: We performed real-time RT-PCR and enzyme-linked immunosorbent assay to examine the expression of SDF-1α in nucleus pulposus cells after treatment with pro-inflammatory cytokines in vitro. An animal model of IVD degeneration was established using annular fibrosus puncture in rat coccygeal discs. Tissue samples were collected from normal control and degeneration groups. Differences in the expression of SDF-1α between the normal and degenerative IVDs were analyzed by immunohistochemistry. The migration capacity of NPSCs induced by SDF-1α was evaluated using wound healing and transwell migration assays. To determine the effect of SDF-1α on chondrogenic differentiation of NPSCs, we conducted cell micromass culture and examined the expression levels of Sox-9, aggrecan, and collagen II. Moreover, the roles of SDF-1/CXCR4 axis in the migration and chondrogenesis differentiation of NPSCs were analyzed by immunofluorescence, immunoblotting, and real-time RT-PCR. RESULTS: SDF-1α was significantly upregulated in the native IVD cells cultured in vitro with pro-inflammatory cytokines, such as interleukin-1ß and tumor necrosis factor-α, mimicking the degenerative settings. Immunohistochemical staining showed that the level of SDF-1α was also significantly higher in the degenerative group than in the normal group. SDF-1α enhanced the migration capacity of NPSCs in a dose-dependent manner. In addition, SDF-1α induced chondrogenic differentiation of NPSCs, as evidenced by the increased expression of chondrogenic markers using histological and immunoblotting analyses. Real-time RT-PCR, immunoblotting, and immunofluorescence showed that SDF-1α not only increased CXCR4 expression but also stimulated translocation of CXCR4 from the cytoplasm to membrane, accompanied by cytoskeletal rearrangement. Furthermore, blocking CXCR4 with AMD3100 effectively suppressed the SDF-1α-induced migration and differentiation capacities of NPSCs. CONCLUSION: These findings demonstrate that SDF-1α has the potential to enhance recruitment and chondrogenic differentiation of NPSCs via SDF-1/CXCR4 chemotaxis signals that contribute to IVD regeneration.
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Purpose To investigate the effect of grape seed proanthocyanidins on the phenotype-transforming marker protein expression of db/db renal cells in mice model of type 2 diabetes,and to explore the protective mechanism of grape seed extract on diabetic renal injury in db/db mice.Methods Male db/db diabetic mice were randomly divided into two groups:diabetic group (db/db group) and diabetic + grape seed proanthocyanidin extract group (db/db + GSPE).The same week-old male db/m mice was used as normal controls (db/m) and grape seed proanthocyanidin extract gavage treatment group (db/m +grape seed proanthocyanidin extract group,db/m + GSPE).The mice of db/db + GSPE group and db/m + GSPE group were administered daily with grape seed proanthocyanidin extract (5mg/kg) by gavage.Results Renal tissues of db/db diabetic mice showed increased expression of α-SMA,p-p38MAPK,pERK1/2 and 8-OHdG level,and down-regulation in E-cadherin expression compared with db/m group (P ＜ 0.05).However,the alternations of α-SMA,p-p38,p-ERK1/2,E-cadherin protein levels,and 8-OHdG level,in db/db group were reversed by addition of grape seed proanthocyanidin extract (P ＜ 0.05).Conclusion Grape seed proanthocyanidin extract inhibits the epithelial to mesenchymal transition (EMT) associated protein,by decreasing ROS production,and activating p38 MAPK and ERK1/2.These findings suggest that grape seed proanthocyanidin extract provides a treatment option for diabetic nephropathy.