Excessive peptidergic sensory innervation of cutaneous arteriole-venule shunts (AVS) in the palmar glabrous skin of fibromyalgia patients: implications for widespread deep tissue pain and fatigue.

OBJECTIVE: To determine if peripheral neuropathology exists among the innervation of cutaneous arterioles and arteriole-venule shunts (AVS) in fibromyalgia (FM) patients. SETTING: Cutaneous arterioles and AVS receive a convergence of vasoconstrictive sympathetic innervation, and vasodilatory small-fiber sensory innervation. Given our previous findings of peripheral pathologies in chronic pain conditions, we hypothesized that this vascular location may be a potential site of pathology and/or serotonergic and norepinephrine reuptake inhibitors (SNRI) drug action. SUBJECTS: Twenty-four female FM patients and nine female healthy control subjects were enrolled for study, with 14 additional female control subjects included from previous studies. AVS were identified in hypothenar skin biopsies from 18/24 FM patient and 14/23 control subjects. METHODS: Multimolecular immunocytochemistry to assess different types of cutaneous innervation in 3 mm skin biopsies from glabrous hypothenar and trapezius regions. RESULTS: AVS had significantly increased innervation among FM patients. The excessive innervation consisted of a greater proportion of vasodilatory sensory fibers, compared with vasoconstrictive sympathetic fibers. In contrast, sensory and sympathetic innervation to arterioles remained normal. Importantly, the sensory fibers express α2C receptors, indicating that the sympathetic innervation exerts an inhibitory modulation of sensory activity. CONCLUSIONS: 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.

Air-stimulated ATP release from keratinocytes occurs through connexin hemichannels.

Cutaneous ATP release plays an important role in both epidermal stratification and chronic pain, but little is known about ATP release mechanisms in keratinocytes that comprise the epidermis. In this study, we analyzed ATP release from cultured human neonatal keratinocytes briefly exposed to air, a process previously demonstrated to trigger ATP release from these cells. We show that exposing keratinocytes to air by removing media for 15 seconds causes a robust, long-lasting ATP release. This air-stimulated ATP release was increased in calcium differentiated cultures which showed a corresponding increase in connexin 43 mRNA, a major component of keratinocyte hemichannels. The known connexin hemichannel inhibitors 1-octanol and carbenoxolone both significantly reduced air-stimulated ATP release, as did two drugs traditionally used as ABC transporter inhibitors (glibenclamide and verapamil). These same 4 inhibitors also prevented an increase in the uptake of a connexin permeable dye induced by air exposure, confirming that connexin hemichannels are open during air-stimulated ATP release. In contrast, activity of the MDR1 ABC transporter was reduced by air exposure and the drugs that inhibited air-stimulated ATP release had differential effects on this transporter. These results indicate that air exposure elicits non-vesicular release of ATP from keratinocytes through connexin hemichannels and that drugs used to target connexin hemichannels and ABC transporters may cross-inhibit. Connexins represent a novel, peripheral target for the treatment of chronic pain and dermatological disease.

Keratinocyte expression of calcitonin gene-related peptide ß: implications for neuropathic and inflammatory pain mechanisms.

Calcitonin gene-related peptide (CGRP) is a vasodilatory peptide that has been detected at high levels in the skin, blood, and cerebrospinal fluid (CSF) 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, quantitative Polymerase Chain Reaction (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, Calcitonin receptor-like receptor (CRLR), Receptor activity-modifying protein 1 (RAMP1), CGRP-receptor component protein (RCP) consistent with known observations that CGRP promotes several functional changes in keratinocytes, including proliferation and cytokine production. Our results indicate that keratinocyte-derived CGRPß may modulate epidermal homeostasis through autocrine/paracrine signaling and may contribute to chronic pain under pathological conditions.

Voltage-gated sodium channel expression in rat and human epidermal keratinocytes: evidence for a role in pain.

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.

Sensory perception and neuroanatomical structures in normal and grafted skin of burn survivors.

BACKGROUND: This study compared the neural structures found in grafted skin of burn survivors with neural structures found in site-matched normal skin and correlated these structures with psychophysical measures of sensation. METHODS: Fifteen skin-grafted male burn survivors (47.7+/-10.4 years old) with deep partial- or full-thickness thermal burn injuries covering an average of 11+/-5.6% of their total body surface and with normal skin at a matching, unburned, contralateral site were recruited into this study. Threshold determinations and magnitude estimations for touch, cold, warmth and heat-pain were performed at sites with grafted and normal skin, using Semmes-Weinstein monofilaments and the Medoc TSA 2001 thermal stimulator. Skin biopsies from both the grafted and normal sites were stained with antibodies for protein gene product 9.5 (PGP) and neurofilament 200 kDa. Nerve fibers in the epidermis and nerve fibers or bundles of nerve fibers in the superficial and deep dermis as well as innervated blood vessels, hair follicles and sweat glands were counted. RESULTS: On average, the data were collected 43.1+/-10.4 months after grafting. When thresholds on grafted skin were compared to thresholds on normal skin, they showed elevated sensory thresholds [touch (p<0.003), cold (p<0.031), warmth (p<0.009)]. Magnitude estimates of touch, cold and warmth differed on the two sides with sensations elicited from grafts being smaller than those from normal skin. Heat-pain thresholds and heat-pain magnitude estimations were not statistically different on the two sites. By comparison to the normal side, and consistent with the attenuated sensory functions of the grafts, counts of neural structures showed a reduction in innervation density; PGP-immunoreactive nerve fibers/bundles were reduced in grafted epidermis (p<0.026) and superficial dermis (p<0.001). The numbers of sweat glands (p<0.006) and hair follicles (p<0.001) were also reduced. The number of innervated blood vessels did not differ significantly on the two sides. There were significant correlations between sensory thresholds and the neuroanatomical variables: thresholds of cold and touch were correlated with the number of sweat glands in both grafted and normal skin (r2=0.56 and 0.50, respectively; p<0.001), while warmth thresholds were significantly correlated with the number of innervated blood vessels in grafted skin (r2=0.62, p<0.001). Encapsulated mechanoreceptors were not encountered in this study of hairy skin. CONCLUSIONS: Touch, cold and warmth thresholds and magnitude estimations do not return to normal levels after skin grafting in burn survivors. The elevation of thresholds and reduction of sensory intensity is accompanied by a general decrease in the density of nerve terminals. The lack, or numerical reduction, of sweat glands and innervated blood vessels was also indicative of diminished sensation on grafted skin.

[Surgical management of bilateral mandibular angle prominence associated with microgenia].

OBJECTIVE: A surgical procedure was developed to improve the operative results for bilateral mandibular prominence with microgenia. METHODS: Through an oral approach the bilateral prominent mandibular angles were resected using the technique of continuous curve-line ostectomy with the masseteric muscle intact. Meanwhile, the chin was elongated and moved forward by horizontal osteotomy. The distal part of the chin was rigid fixed with miniplates and screws, and the bone gaps were filled with autogenous bone of the resected mandibular angle. RESULTS: From November 1996 to August 1999, 20 cases of bilateral mandibular angle prominence with microgenia were corrected with this procedure. The postoperative appearance was improved greatly after 3-6 months. The lower face was not only narrowed but elongated, and consequently in harmony with the upper and middle face. CONCLUSION: Bilateral continuous curve-line mandibular angle ostectomy together with chin osteotomy and autogenous bone graft is a good procedure for the management of the above-mentioned deformities.