Heat shock proteins, endothelin, and peripheral neuronal injury.

Heat shock proteins (HSP) are important in neuroprotection after a variety of stresses or injuries. Both heat shock proteins and endothelin are upregulated after peripheral nerve injury, but HSP regulation after injury has not been systematically studied in peripheral nerve. The purpose of this study was to examine the regulation of small and large heat shock proteins after injury to rat sciatic nerve. Secondly, using a parallel tissue culture model for the sciatic nerve (PC12 cells), we examined potential regulation of heat shock proteins by endothelin. Western analysis of constricted, mobilized or unperturbed nerve was used to examine HSP abundance after injury. Semiquantitative PCR was used to examine heat shock message levels after nerve injury in the dorsal root ganglia. Cultured undifferentiated and differentiated PC12 cells were treated with endothelin, then western analysis of cytosol- and membrane-enriched fractions of these cells was used to examine heat shock protein regulation after endothelin treatment. Heat shock proteins are expressed at very low levels in unperturbed sciatic nerve. Constriction injury of the nerve results in increased expression of small and large heat shock proteins, but no upregulation of HSP message in corresponding dorsal root ganglia. Endothelin treatment of PC12 does not cause upregulation of heat shock proteins. Together these data show that a broad range of HSP is involved in endogenous response to peripheral nerve injury and deserve further study as potential neuroprotectants. Regulation of heat shock proteins after nerve injury is not likely due to endothelin signaling.

Intravenous mononuclear marrow cells reverse neuropathic pain from experimental mononeuropathy.

BACKGROUND: Stem cells mediate neuroprotection in a variety of nervous system injury models. In this study, we evaluated a potential role for stem cells in pain therapies. Marrow mononuclear cells containing mixed stem cell populations were used because of wide experience with these cells in experimental and clinical transplantation. METHODS: After sciatic nerve chronic constriction injury (CCI), adult male Sprague Dawley rats were treated with freshly isolated marrow mononuclear cells (10(7) cells in 0.5 mL IV) from the same strain, or with carrier. The major end points of analysis were thermal and mechanical hypersensitivity using paw withdrawal latency (PWL) to a calibrated heat source and paw withdrawal response to von Frey filaments, evaluated by a blinded investigator. RESULTS: Marrow transplantation did not prevent pain, and 5 days after CCI all animals were equivalently lesioned. However, 10 days after CCI, rats that received marrow transplants demonstrated paw withdrawal response and PWL patterns indicating recovery from pain, whereas untreated rats continued to have significant pain behavior patterns. For example, PWL values for marrow-treated animals were similar to baseline pre-CCI values (P = 0.54) but significantly shorter latency to withdrawal indicative of continuing pain was seen in untreated rats compared with pre-CCI values (P < 0.001). CONCLUSIONS: These studies suggest that stem or progenitor cell-mediated therapies may be useful for the treatment of pain after nerve injury, and deserve further study to elucidate the mechanisms of analgesia.

A reducing redox environment promotes C2C12 myogenesis: implications for regeneration in aged muscle.

Intracellular redox potential of skeletal muscle becomes progressively more oxidized with aging, negatively impacting regenerative ability. We examined the effects of oxidizing redox potential on terminal differentiation of cultured C2C12 myoblasts. Redox potentials were manipulated by changing the culture O(2) environment, by free radical scavenging, or addition of H(2)O(2.) Intracellular reactive oxygen species (ROS) production was higher in 20% environmental O(2) and in this condition, redox potential became progressively oxidized compared to cultures in 6% O(2.) Treatment with a ROS trapping agent (phenyl-N-tert-butylnitrone, PBN) caused reducing redox potentials and enhanced C2C12 differentiation, while addition of 25 micromolar H(2)O(2) to cells in 20% O(2) dramatically slowed differentiation. Under these most oxidative conditions, quantitative PCR showed a significant decrease in myogenic basic helix-loop-helix transcription factor expression compared to cultures treated with PBN or grown in 6% O(2). Thus, oxidative intracellular environments impair myoblast differentiation, while reducing environments favor myogenesis.

A role for endothelin in neuropathic pain after chronic constriction injury of the sciatic nerve.

The purpose of this study was to explore the role of endothelin in neuropathic pain. Endothelins (ET) are a family (ET-1, ET-2, ET-3) of ubiquitously expressed peptides involved in control of vascular tone. Injected ET-1 causes intense pain via activation of ETA receptors, modulated by analgesic signals initiated by ETB receptor activation. Using a rat model of chronic constriction injury of the sciatic nerve, we found that pharmacologic ETA receptor antagonism acutely and significantly reduced thermal and mechanical hyperalgesic responses 5 days after injury. Furthermore, ET-1 and the ETA receptor are locally upregulated at the site of chronic constriction injury at both the message and the protein levels, suggesting that ET-1 may be involved in establishing pain after the injury. These data point to ET-1 as an important mediator of pain in general and suggest that ETA antagonism deserves study as a potential novel therapy for neuropathic pain.