l-Theanine and NEP1-40 promote nerve regeneration and functional recovery after brachial plexus root avulsion.

Brachial plexus root avulsion causes severe sequelae Treatments and prognosis face many problems, including inflammatory reaction, oxidative damage, and myelin related inhibitory effect. l-Theanine has anti-inflammatory, anti-oxidative, and neuroprotective effects. NEP1-40 competitively inhibits Nogo-66 receptor (NgR1) promotes axonal regeneration. Forty-eight Sprague-Dawley rats were randomly assigned into four groups to establish an animal model of brachial plexus root avulsion. Inflammation and oxidative damage were evaluated by spectrophotometry and motor function of the upper limbs was assessed via Terzis grooming test after modeling. Immunofluorescence and hematoxylin and eosin staining were utilized to determine the content of reactive oxygen species, activation of microglial cells, neuroprotection, and nerve regeneration. Compared with the control group, the L-Theanine + NEP1-40 group had significantly decreased myeloperoxidase, malondialdehyde, interleukin-6, reactive oxygen species, and microglial cells, significantly increased score on the Terzis grooming test, increased motor neuron content, and thickened muscle fibers, increased area, and appearance of large and clear motor endplate structures. The results of this study suggest that l-Theanine combined with NEP1-40significantly promoted nerve regeneration after brachial plexus root avulsion, and may be a potential treatment for promoting nerve regeneration. Possible mechanisms underlying these results are alleviation of oxidative damage and inflammatory responses in the injured area and antagonism of myelin inhibition.

Inhibition of LncRNA Vof-16 expression promotes nerve regeneration and functional recovery after spinal cord injury.

Our previous RNA sequencing study showed that the long non-coding RNA ischemia-related factor Vof-16 (lncRNA Vof-16) was upregulated after spinal cord injury, but its precise role in spinal cord injury remains unclear. Bioinformatics predictions have indicated that lncRNA Vof-16 may participate in the pathophysiological processes of inflammation and apoptosis. PC12 cells were transfected with a pHBLV-U6-MCS-CMV-ZsGreen-PGK-PURO vector to express an lncRNA Vof-16 knockdown lentivirus and a pHLV-CMVIE-ZsGree-Puro vector to express an lncRNA Vof-16 overexpression lentivirus. The overexpression of lncRNA Vof-16 inhibited PC12 cell survival, proliferation, migration, and neurite extension, whereas lncRNA Vof-16 knockdown lentiviral vector resulted in the opposite effects in PC12 cells. Western blot assay results showed that the overexpression of lncRNA Vof-16 increased the protein expression levels of interleukin 6, tumor necrosis factor-α, and Caspase-3 and decreased Bcl-2 expression levels in PC12 cells. Furthermore, we established rat models of spinal cord injury using the complete transection at T10. Spinal cord injury model rats were injected with the lncRNA Vof-16 knockdown or overexpression lentiviral vectors immediately after injury. At 7 days after spinal cord injury, rats treated with lncRNA Vof-16 knockdown displayed increased neuronal survival and enhanced axonal extension. At 8 weeks after spinal cord injury, rats treated with the lncRNA Vof-16 knockdown lentiviral vector displayed improved neurological function in the hind limb. Notably, lncRNA Vof-16 knockdown injection increased Bcl-2 expression and decreased tumor necrosis factor-α and Caspase-3 expression in treated animals. Rats treated with the lncRNA Vof-16 overexpression lentiviral vector displayed opposite trends. These findings suggested that lncRNA Vof-16 is associated with the regulation of inflammation and apoptosis. The inhibition of lncRNA Vof-16 may be useful for promoting nerve regeneration and functional recovery after spinal cord injury. The experiments were approved by the Institutional Animal Care and Use Committee of Guangdong Medical University, China.

Effect of lentiviral vector-mediated overexpression of hypoxia-inducible factor 1 alpha delivered by pluronic F-127 hydrogel on brachial plexus avulsion in rats.

Brachial plexus avulsion often results in massive motor neuron death and severe functional deficits of target muscles. However, no satisfactory treatment is currently available. Hypoxia-inducible factor 1α is a critical molecule targeting several genes associated with ischemia-hypoxia damage and angiogenesis. In this study, a rat model of brachial plexus avulsion-reimplantation was established, in which C5-7 ventral nerve roots were avulsed and only the C6 root reimplanted. Different implants were immediately injected using a microsyringe into the avulsion-reimplantation site of the C6 root post-brachial plexus avulsion. Rats were randomly divided into five groups: phosphate-buffered saline, negative control of lentivirus, hypoxia-inducible factor 1α (hypoxia-inducible factor 1α overexpression lentivirus), gel (pluronic F-127 hydrogel), and gel + hypoxia-inducible factor 1α (pluronic F-127 hydrogel + hypoxia-inducible factor 1α overexpression lentivirus). The Terzis grooming test was performed to assess recovery of motor function. Scores were higher in the hypoxia-inducible factor 1α and gel + hypoxia-inducible factor 1α groups (in particular the gel + hypoxia-inducible factor 1α group) compared with the phosphate-buffered saline group. Electrophysiology, fluorogold retrograde tracing, and immunofluorescent staining were further performed to investigate neural pathway reconstruction and changes of neurons, motor endplates, and angiogenesis. Compared with the phosphate-buffered saline group, action potential latency of musculocutaneous nerves was markedly shortened in the hypoxia-inducible factor 1α and gel + hypoxia-inducible factor 1α groups. Meanwhile, the number of fluorogold-positive cells and ChAT-positive neurons, neovascular area (labeled by CD31 around avulsed sites in ipsilateral spinal cord segments), and the number of motor endplates in biceps brachii (identified by α-bungarotoxin) were all visibly increased, as well as the morphology of motor endplate in biceps brachil was clear in the hypoxia-inducible factor 1α and gel + hypoxia-inducible factor 1α groups. Taken together, delivery of hypoxia-inducible factor 1α overexpression lentiviral vectors mediated by pluronic F-127 effectively promotes spinal root regeneration and functional recovery post-brachial plexus avulsion. All animal procedures were approved by the Institutional Animal Care and Use Committee of Guangdong Medical University, China.