No abstract

Keratinocytes are implicated in sensory transduction and can influence nociception, but whether these contribute to chronic pain is not known. In neurons, voltage-gated sodium channels (Na(v)) are involved in neuropathic pain and are activated by depolarization. Since keratinocytes can also show changes in membrane potential, we used RT-PCR, in situ hybridization, and immunohistochemistry to investigate the expression of sodium channels in these cells. Na(v)1.1, Na(v)1.6, and Na(v)1.8 were localized within keratinocytes in rat epidermis. In addition, sodium channels contribute to the release of ATP from rat keratinocytes in response to increased [K(+)](o), implicating sodium channels in keratinocyte ligand release and nociception. To examine whether keratinocytes may contribute to human pain states, we analyzed sodium channel expression in human skin biopsies from subjects with complex regional pain syndrome Type 1 (CRPS) and post-herpetic neuralgia (PHN) using immunohistochemistry. Control skin exhibited immunolabeling for Na(v)1.5, Na(v)1.6 and Na(v)1.7. In contrast, painful skin from CRPS and PHN subjects displayed Na(v)1.1, Na(v)1.2, and Na(v)1.8 immunolabeling, in addition to substantially increased signal for Na(v)1.5, Na(v)1.6, Na(v)1.7. These observations lead us to propose that pathological increases in keratinocyte sodium channel expression may contribute to pain by increasing epidermal ATP release, resulting in excessive activation of P2X receptors on primary sensory axons. Consistent with this hypothesis, animal models of neuropathic pain exhibit increases in subcutaneous ATP release and activity of primary sensory neurons, and peripheral administration of P2X antagonists has been shown to reduce neuropathic pain in humans.

Calcitonin Gene-Related Peptide (CGRP) is a vasodilatory peptide that has been detected at high levels in the skin, blood, and cerebral spinal fluid under a variety of inflammatory and chronic pain conditions, presumably derived from peptidergic C and Aδ innervation. Herein, CGRP immunolabeling (IL) was detected in epidermal keratinocytes at levels that were especially high and widespread in the skin of humans from locations afflicted with postherpetic neuralgia (PHN) and complex region pain syndrome type 1 (CRPS), of monkeys infected with simian immunodeficiency virus, and of rats subjected to L5/L6 spinal nerve ligation, sciatic nerve chronic constriction, and subcutaneous injection of Complete Freund’s Adjuvant. Increased CGRP-IL was also detected in epidermal keratinocytes of transgenic mice with keratin-14 promoter driven overexpression of noggin, an antagonist to BMP-4 signaling. Transcriptome microarray, qPCR, and Western blot analyses using laser captured mouse epidermis from transgenics, monolayer cultures of human and mouse keratinocytes, and multilayer human keratinocyte organotypic cultures, revealed that keratinocytes express predominantly the beta isoform of CGRP. Cutaneous peptidergic innervation has been shown to express predominantly the alpha isoform of CGRP. Keratinocytes also express the cognate CGRP receptor components, CRLR, RAMP1, and RCP, consistent with known observations that CGRP promotes several functional changes in keratinocytes, including proliferation and cytokine production. Our results indicate that keratinocyte derived CGRPβ may modulate epidermal homeostasis through autocrine/paracrine signaling and may contribute to chronic pain under pathological conditions.

Myotonic dystrophy, or dystrophia myotonica (DM), is a highly variable multisystem disease in which the classic adult-onset form displays progressive muscle wasting, cataracts, heart block, gonadal atrophy, insulin resistance and neuropsychiatric impairment. Its genetic basis is an expansion of CTG trinucleotide repeats in the DMPK protein kinase gene. Among the triplet repeat expansion disorders, DM is distinguished by the extended length of the repeat tract (5-13 kb in postmortem tissue) and its location in the 3' untranslated region of the gene that contains it. The pathophysiological mechanism for multisystem degeneration in DM is not understood. In contrast to the profound muscle wasting that characterizes advanced DM, only minor histopathological abnormalities have occurred in DMPK knockout mice or in mice that overexpress a human DMPK transgene, making it unlikely that changes in DMPK activity provide a unitary explanation for the disease. A DNAse hypersensitive site that maps 0.7 kb downstream (centromeric) from the CTG repeats is eliminated on DM chromosomes. This finding indicates that the repeat expansion may alter the adjacent chromatin structure and raises the possibility that it may also affect the expression of flanking genes. An interesting candidate flanking gene is DMAHP, a recently discovered homeodomain-encoding gene. We show here that DMAHP expression in myoblasts, muscle and myocardium is reduced by the DM mutation is cis, and the magnitude of this effect depends on the extent of CTG repeat expansion. These observations support the hypothesis that DMAHP participates in the pathophysiology of DM.

Results in multiple sclerosis patients were similar to those of a previous study in amyotrophic lateral sclerosis, demonstrating that DM/Q may be beneficial in treating potentially disabling pseudobulbar affect in a variety of neurological disorders.

An orally administered, fixed-dose coformulation of sodium phenylbutyratetaurursodiol (PB-TURSO) significantly slowed functional decline in a randomized, placebo-controlled, phase 2 trial in ALS (CENTAUR). Herein we report results of a long-term survival analysis of participants in CENTAUR. In CENTAUR, adults with ALS were randomized 2:1 to PB-TURSO or placebo. Participants completing the 6-month (24-week) randomized phase were eligible to receive PB-TURSO in the open-label extension. An all-cause mortality analysis (35-month maximum follow-up post-randomization) incorporated all randomized participants. Participants and site investigators were blinded to treatment assignments through the duration of followup of this analysis. Vital status was obtained for 135 of 137 participants originally randomized in CENTAUR. Median overall survival was 25.0 months among participants originally randomized to PB-TURSO and 18.5 months among those originally randomized to placebo (hazard ratio, 0.56; 95% confidence interval, 0.34-0.92; P = .023). Initiation of PB-TURSO treatment at baseline resulted in a 6.5-month longer median survival as compared with placebo. Combined with results from CEN-TAUR, these results suggest that PB-TURSO has both functional and survival benefits in ALS.

The excessive sensory innervation to the glabrous skin AVS is a likely source of severe pain and tenderness in the hands of FM patients. Importantly, glabrous AVS regulate blood flow to the skin in humans for thermoregulation and to other tissues such as skeletal muscle during periods of increased metabolic demand. Therefore, blood flow dysregulation as a result of excessive innervation to AVS would likely contribute to the widespread deep pain and fatigue of FM. SNRI compounds may provide partial therapeutic benefit by enhancing the impact of sympathetically mediated inhibitory modulation of the excess sensory innervation.

The amino-terminal domain of the large subunit of herpes simplex virus type 2 (HSV-2) ribonucleotide reductase (ICP 10) has protein kinase (PK) activity and properties similar to those of growth factor receptor kinases which can be activated to transforming potential. DNA sequences that encode the PK domain cause neoplastic transformation of immortalized cells. The studies described in this report used a spontaneous mutant (ts5-152) temperature-sensitive for the synthesis of 1CPI0 and the previously described ICP10 expression vectors to study the role of ICP10 expression in HSV-2 growth and neoplastic potential. The titres of the ts5-152 mutant are 1000-fold lower at 39 °C compared to 34 °C after 12 h post-infection. The efficiency of plaquing is 0-003. The growth defect at 39 °C correlates with decreased ICP10 synthesis. Sequence analysis of the PK domain of the ts5-152 ICP10 gene identified a pair of frameshift mutations resulting in a 19 amino acid residue substitution at positions 275 to 293 and a downstream single base pair mutation causing a substitution at position 309. Cloning of the mutant ICP 10 gene from tsS-152 into a wild-type HSV-2 isolate resulted in a recombinant (859/152) with growth properties and rates of ICP10 synthesis at 39 °C similar to those of ts5-152. Cells transformed with u.v.-inactivated ts5-152, or the recombinant 859/152, have significantly decreased cloning efficiency in agarose at 39 °C, but only during the first 250 post-transfer population doublings. Anchorage-independent growth was observed in cells transfected with expression vectors pJWl 7 or pJW32 that express ICP10 or its PK domain, respectively. Cells transfected with the frameshift mutant pJW21 or the ICP10 carboxy-terminal vector pJW31 did not form clones in agarose.