Anti-VEGF polyclonal antibody [Alexa Fluor® 647]

Gpr126/Adgrg6 is an adhesion G protein-coupled receptor essential for Schwann cell (SC) myelination with important contributions to repair after nerve crush injury. Despite critical functions in myelinating SCs, the role of Gpr126 within nonmyelinating terminal Schwann cells (tSCs) at the neuromuscular junction (NMJ), is not known. tSCs have important functions in synaptic maintenance and reinnervation, and after injury tSCs extend cytoplasmic processes to guide regenerating axons to the denervated NMJ. In this study, we show that Gpr126 is expressed in tSCs, and that absence of Gpr126 in SCs (SC-specific Gpr126 knockout, cGpr126) results in a NMJ maintenance defect in the hindlimbs of aged mice, but not in young adult mice. After nerve transection and repair, cGpr126 mice display delayed NMJ reinnervation, altered tSC morphology with decreased S100 beta expression, and reduced tSC cytoplasmic process extensions. The immune response promoting reinnervation at the NMJ following nerve injury is also altered with decreased macrophage infiltration, Tnf alpha, and anomalous cytokine expression compared to NMJs of control mice. In addition, Vegfa expression is decreased in muscle, suggesting that cGpr126 non-cell autonomously modulates angiogenesis after nerve injury. In sum, cGpr126 mice demonstrated delayed NMJ reinnervation and decreased muscle mass following nerve transection and repair compared to control littermates. The integral function of Gpr126 in tSCs at the NMJ provides the framework for new therapeutic targets for neuromuscular disease.

Objectives We aimed to investigate whether the mesenchymal stem cell-endothelial cell crosstalk enhances angiogenic factor expression via nuclear factor-kappa B (NF-kappa B)-dependent mechanisms. Materials and Methods Human dermal microvascular endothelial cells (HDMECs) and stem cells from human exfoliated deciduous teeth (SHEDs) were cocultured for 96 hr, in the presence of NF-kappa B decoy oligodeoxynucleotides (ODNs) or scramble (control). Vascular endothelial cell growth factor (VEGF) and phospho-NF-kappa B p65 were measured with enzyme-linked immunosorbent assay. Angiogenesis-related gene expression was analyzed with microarray analysis followed by real-time polymerase chain reaction. Tube formation assay was conducted in the presence of NF-kappa B decoy. Results The VEGF and phospho-NF-kappa B p65 levels were significantly higher in the coculture with NF-kappa B decoy scramble than in single culture and coculture with NF-kappa B decoy ODN. Microarray analysis of SHEDs and HDMECs with NF-kappa B decoy scramble showed higher expression of proangiogenic genes, Bcl-2, NF-kappa B1, VEGFA, CXCL8, and CXCR1, and lower expression of proapoptotic genes, Bax and Caspase 9, compared to cells with NF-kappa B decoy ODN. Real-time PCR results for Bcl-2 and CXCL8 showed a similar trend. Tube formation assay showed more tube development in the presence of NF-kappa B decoy scramble. Conclusion The SHED-HDMEC crosstalk enhanced proangiogenic factor expression via NF-kappa B-dependent pathways.