Calcium Homeostasis in Parvalbumin DRG Neurons is Altered After Sciatic Nerve Crush and Sciatic Nerve Transection Injuries

Reflex abnormalities mediated by proprioceptive sensory neurons after peripheral nerve injury (PNI) can limit functional improvement, leaving patients with disability that affects their quality of life. We examined post-injury calcium transients in a subpopulation of DRG neurons consisting primarily of proprioceptors to determine whether alterations in calcium homeostasis are present in proprioceptors, as has been documented in other DRG neurons after PNI. Using transgenic mice, we restricted expression of the calcium indicator GCaMP6s to DRG neurons containing parvalbumin (PV). Mice of both sexes were randomly assigned to sham, sciatic nerve crush, or sciatic nerve transection and resuture conditions. Calcium transients were recorded from ex-vivo preparations of animals at one of three post-surgery time points: 1-3 days, 7-11 days, and after 60 days of recovery. Results demonstrated that the post-PNI calcium transients of PV DRG neurons are significantly different than sham. Abnormalities were not present during the acute response to injury (1-3 days), but transients were significantly different than sham at the recovery stage where axon regeneration is thought to be underway (7-11 days). During late-stage recovery (60 days post-injury), disturbances in the decay time course of calcium transients in transection animals persisted, whereas parameters of transients from crush animals returned to normal. These findings identify a deficit in calcium homeostasis in proprioceptive neurons, which may contribute to the failure to fully recover proprioceptive reflexes after PNI. Significant differences in the calcium transients of crush versus transection animals after reinnervation illustrate calcium homeostasis alterations are distinctive to injury type.

7,8-Dihydroxiflavone Maintains Retinal Functionality and Protects Various Types of RGCs in Adult Rats with Optic Nerve Transection

To analyze the neuroprotective effects of 7,8-Dihydroxyflavone (DHF) in vivo and ex vivo, adult albino Sprague-Dawley rats were given a left intraorbital optic nerve transection (IONT) and were divided in two groups: One was treated daily with intraperitoneal (ip) DHF (5 mg/kg) (n = 24) and the other (n = 18) received ip vehicle (1% DMSO in 0.9% NaCl) from one day before IONT until processing. At 5, 7, 10, 12, 14, and 21 days (d) after IONT, full field electroretinograms (ERG) were recorded from both experimental and one additional naïve-control group (n = 6). Treated rats were analyzed 7 (n = 14), 14 (n = 14) or 21 d (n = 14) after IONT, and the retinas immune stained against Brn3a, Osteopontin (OPN) and the T-box transcription factor T-brain 2 (Tbr2) to identify surviving retinal ganglion cells (RGCs) (Brn3a+), α-like (OPN+), α-OFF like (OPN+Brn3a+) or M4-like/α-ON sustained RGCs (OPN+Tbr+). Naïve and right treated retinas showed normal ERG recordings. Left vehicle-treated retinas showed decreased amplitudes of the scotopic threshold response (pSTR) (as early as 5 d), the rod b-wave, the mixed response and the cone response (as early as 10 d), which did not recover with time. In these retinas, by day 7 the total numbers of Brn3a+RGCs, OPN+RGCs and OPN+Tbr2+RGCs decreased to less than one half and OPN+Brn3a+RGCs decreased to approximately 0.5%, and Brn3a+RGCs showed a progressive loss with time, while OPN+RGCs and OPN+Tbr2+RGCs did not diminish after seven days. Compared to vehicle-treated, the left DHF-treated retinas showed significantly greater amplitudes of the pSTR, normal b-wave values and significantly greater numbers of OPN+RGCs and OPN+Tbr2+RGCs for up to 14 d and of Brn3a+RGCs for up to 21 days. DHF affords significant rescue of Brn3a+RGCs, OPN+RGCs and OPN+Tbr2+RGCs, but not OPN+Brn3a+RGCs, and preserves functional ERG responses after IONT.

Sodium phenylbutyrate inhibits Schwann cell inflammation via HDAC and NFκB to promote axonal regeneration and remyelination

Background Epigenetic regulation by histone deacetylases (HDACs) in Schwann cells (SCs) after injury facilitates them to undergo de- and redifferentiation processes necessary to support various stages of nerve repair. Although de-differentiation activates the synthesis and secretion of inflammatory cytokines by SCs to initiate an immune response during nerve repair, changes in either the timing or duration of prolonged inflammation mediated by SCs can affect later processes associated with repair and regeneration. Limited studies have investigated the regulatory processes through which HDACs in SCs control inflammatory cytokines to provide a favorable environment for peripheral nerve regeneration. Methods We employed the HDAC inhibitor (HDACi) sodium phenylbutyrate (PBA) to address this question in an in vitro RT4 SC inflammation model and an in vivo sciatic nerve transection injury model to examine the effects of HDAC inhibition on the expression of pro-inflammatory cytokines. Furthermore, we assessed the outcomes of suppression of extended inflammation on the regenerative potential of nerves by assessing axonal regeneration, remyelination, and reinnervation. Results Significant reductions in lipopolysaccharide (LPS)-induced pro-inflammatory cytokine (tumor necrosis factor-α [TNFα]) expression and secretion were observed in vitro following PBA treatment. PBA treatment also affected the transient changes in nuclear factor κB (NFκB)-p65 phosphorylation and translocation in response to LPS induction in RT4 SCs. Similarly, PBA mediated long-term suppressive effects on HDAC3 expression and activity. PBA administration resulted in marked inhibition of pro-inflammatory cytokine secretion at the site of transection injury when compared with that in the hydrogel control group at 6-week post-injury. A conducive microenvironment for axonal regrowth and remyelination was generated by increasing expression levels of protein gene product 9.5 (PGP9.5) and myelin basic protein (MBP) in regenerating nerve tissues. PBA administration increased the relative gastrocnemius muscle weight percentage and maintained the intactness of muscle bundles when compared with those in the hydrogel control group. Conclusions Suppressing the lengthened state of inflammation using PBA treatment favors axonal regrowth and remyelination following nerve transection injury. PBA treatment also regulates pro-inflammatory cytokine expression by inhibiting the transcriptional activation of NFκB-p65 and HDAC3 in SCs in vitro.

Stem cells from human exfoliated deciduous teeth attenuate mechanical allodynia in mice through distinct from the siglec-9/MCP-1-mediated tissue-repairing mechanism

The effects of stem cells from human exfoliated deciduous teeth (SHED) on mechanical allodynia were examined in mice. A single intravenous injection of SHED and conditioned medium from SHED (SHED-CM) through the left external jugular vein significantly reversed the established mechanical allodynia induced by spinal nerve transection at 6 days after injection. SHED or SHED-CM significantly decreased the mean numbers of activating transcription factor 3-positive neurons and macrophages in the ipsilateral side of the dorsal root ganglion (DRG) at 20 days after spinal nerve transection. SHED or SHED-CM also suppressed activation of microglia and astrocytes in the ipsilateral side of the dorsal spinal cord. A single intravenous injection of secreted ectodomain of sialic acid-binding Ig-like lectin-9 and monocyte chemoattractant protein-1 had no effect on the established mechanical allodynia, whereas a single intravenous injection of protein component(s) contained in SHED-CM with molecular weight of between 30 and 50 kDa reversed the pain. Therefore, it may be concluded that protein component(s) with molecular mass of 30–50 kDa secreted by SHED could protect and/or repair DRG neurons damaged by nerve transection, thereby ameliorating mechanical allodynia.

Teneligliptin Exerts Antinociceptive Effects in Rat Model of Partial Sciatic Nerve Transection Induced Neuropathic Pain

Neuropathic pain (NP), is a chronic pain resulting from nerve injury, with limited treatment options. Teneligliptin (TEN) is a dipeptidyl peptidase-4 inhibitor (DPP-4i) approved to treat type 2 diabetes. DPP-4is prevent the degradation of the incretin hormone glucagon-like peptide 1 (GLP-1) and prolong its circulation. Apart from glycemic control, GLP-1 is known to have antinociceptive and anti-inflammatory effects. Herein, we investigated the antinociceptive properties of TEN on acute pain, and partial sciatic nerve transection (PSNT)-induced NP in Wistar rats. Seven days post PSNT, allodynia and hyperalgesia were confirmed as NP, and intrathecal (i.t) catheters were implanted and connected to an osmotic pump for the vehicle (1 μL/h) or TEN (5 μg/1 μL/h) or TEN (5 μg) + GLP-1R antagonist Exendin-3 (9–39) amide (EXE) 0.1 μg/1 μL/h infusion. The tail-flick response, mechanical allodynia, and thermal hyperalgesia were measured for 7 more days. On day 14, the dorsal horn was harvested and used for Western blotting and immunofluorescence assays. The results showed that TEN had mild antinociceptive effects against acute pain but remarkable analgesic effects against NP. Furthermore, co-infusion of GLP-1R antagonist EXE with TEN partially reversed allodynia but not tail-flick latency. Immunofluorescence examination of the spinal cord revealed that TEN decreased the immunoreactivity of glial fibrillary acidic protein (GFAP). Taken together, our findings suggest that TEN is efficient in attenuation of PSNT-induced NP. Hence, the pleiotropic effects of TEN open a new avenue for NP management.