Number Of Epidermal Nerve Fibers Research Articles

Statistical modeling of diabetic neuropathy: Exploring the dynamics of nerve mortality.

Diabetic neuropathy is a disorder characterized by impaired nerve function and reduction of the number of epidermal nerve fibers per epidermal surface. Additionally, as neuropathy related nerve fiber loss and regrowth progresses over time, the two-dimensional spatial arrangement of the nerves becomes more clustered. These observations suggest that with development of neuropathy, the spatial pattern of diminished skin innervation is defined by a thinning process which remains incompletely characterized. We regard samples obtained from healthy controls and subjects suffering from diabetic neuropathy as realisations of planar point processes consisting of nerve entry points and nerve endings, and propose point process models based on spatial thinning to describe the change as neuropathy advances. Initially, the hypothesis that the nerve removal occurs completely at random is tested using independent random thinning of healthy patterns. Then, a dependent parametric thinning model that favors the removal of isolated nerve trees is proposed. Approximate Bayesian computation is used to infer the distribution of the model parameters, and the goodness-of-fit of the models is evaluated using both non-spatial and spatial summary statistics. Our findings suggest that the nerve mortality process changes as neuropathy advances.

Role of the skin expression of neuropeptides, neurotrophins and their receptors in the pathogenesis of dermatoses

to define the role of neurotransmitters and their receptors in the development of itch and in the maintenance of a skin inflammatory response in patients with psoriasis and atopic dermatitis. Skin biopsy specimens from 30 patients with psoriasis and 30 patients with atopic dermatitis were investigated by histological, immunoperoxidase, and indirect immunofluorescence assays. The investigators determined the expression of protein gene product 9.5 (PGP9.5), amphiregulin, semaphorin 3A, calcitonin gene-related peptide (CGRP) and its receptor (CGRP-R), nerve growth factor (NGF) and its receptor TrkA, and substance P (SP) and its receptor SP-R. The indirect immunofluorescence assay was used for quantitative analysis. The findings were statistically analyzed using a Statistica 10 program. Immunoperoxidase examination of the skin biopsy specimens from patients with atopic dermatitis and psoriasis revealed enhanced expression of amphiregulin, NGF, and PGP9.5, appearance of positively stained epidermal nerve fibers, and decreased expression of the nerve reduction factor semaphorin 3A in all cases. Some patients with atopic dermatitis and psoriasis showed increased expression of CGRP and CGRP-R, SP, SP-R, and TrkA. A pronounced inflammatory response was generally observed in these cases. The investigation performed suggests that atopic dermatitis and psoriasis are characterized by a larger number of epidermal nerve fibers and by a direct correlation between this indicator, disease severity, and itch intensity. The production of neuropeptides and neurotrophins is closely related to the development of a skin inflammatory response irrespective of its cause and dysregulation of these processes is likely to favor the body's sensitization and the chronic pattern the course of diseases.

Psoralen-ultraviolet A therapy alters epidermal Sema3A and NGF levels and modulates epidermal innervation in atopic dermatitis

Epidermal nerve densities are increased in patients with atopic dermatitis (AD), suggesting that it is partly responsible for the intense itching in the skin. Epidermal hyperinnervation in AD patients is decreased by ultraviolet (UV) phototherapy, although the underlying mechanisms are poorly understood. Interestingly, abnormal expression of axonal guidance molecules, such as nerve growth factor (NGF) and semaphorin 3A (Sema3A), is found in the epidermis of AD patients. Therefore, UV phototherapy may alter levels of axonal guidance molecule expression in atopic skin. This study was performed to investigate whether epidermal Sema3A and NGF levels in AD are influenced by psoralen-UVA (PUVA) therapy. Skin biopsies obtained from chronic AD patients before and after PUVA therapy were used. Both Sema3A and NGF in the skin were examined at mRNA and protein levels by quantitative RT-PCR and immunohistochemistry, respectively. Nerve fibers in the skin were stained with anti-PGP9.5 antibody, and the number of epidermal nerve fibers was counted. PUVA therapy decreased epidermal nerve densities in AD patients, concomitant with decreases in both visual analog scale (VAS) scores for pruritus and clinical severity scores. Increased fluorescence intensity of Sema3A and decreased fluorescence intensity of NGF were observed in the epidermis of PUVA-treated group. Moreover, Sema3A mRNA levels were upregulated in the PUVA-treated skins compared with untreated controls, while NGF mRNA levels in the skin were downregulated by the treatment. PUVA therapy may reduce epidermal hyperinnervation of AD by normalization of abnormal Sema3A and NGF expression in the epidermis.

Second Application of Low-energy Shock Waves Has a Cumulative Effect on Free Nerve Endings

Some physicians recommend treating tendinopathies with multiple sessions of shock waves. Some evidence, however, suggests shock wave application can induce nerve fiber degeneration. We questioned whether repeated shock wave application provides a cumulative effect on nerve fibers compared with the effect of one application. One thousand shock wave impulses of an energy flux density of 0.08 mJ/mm were applied to the foot pad of 32 rats. After 14 days, 16 rats received a second application. The foot pads were resected on Days 7, 14, 28, and 42. Sections were processed immunohistochemically using antibodies for sensory nerve. We compared the number of epidermal nerve fibers in rats that received one application of shock waves with the fibers in rats that received two applications. During the first 4 weeks, there was nearly complete degeneration of epidermal nerve fibers in both groups. By the end of 6 weeks, reinnervation of the epidermis began in the single-treatment group. Reinnervation occurred slower in the repeated-treatment group. These data show that a second application has a cumulative effect on nerve fibers. Our data suggest multiple applications of low-energy shock waves might a provide longer-lasting antinociceptive effect.

The role of calcitonin gene-related peptide in cutaneous immunosuppression induced by repeated subinflammatory ultraviolet irradiation exposure.

Ultraviolet (UV) light is an effective treatment for skin disorders like psoriasis in which the cutaneous neurosensory system may have a pathogenic role. In this study, we examined the possibility that UV modulation of the cutaneous neurosensory system and calcitonin gene-related peptide (CGRP) may contribute to local immunosuppression mediated by repeated subinflammatory UV irradiation. Our results indicated that exposure of hairless mice to subinflammatory UV three times weekly for 4 weeks significantly increased the number of epidermal nerve fibers (ENFs) immunoreactive for CGRP without altering the total number of ENFs. The skin content of CGRP as measured by enzyme-linked immunosorbent assay was also significantly increased after exposure to this dose of UV. These effects were most apparent 1 day after the last UV exposure and declined 1 week after UV. The role of CGRP in UV-induced immunosuppression of contact hypersensitivity was then examined. Our results indicated that UV suppression of epicutaneous 2,4-dinitro-1-fluorobenzene (DNFB) sensitization could be significantly inhibited by a systemically administered CGRP receptor antagonist. A broad-spectrum sunscreen applied before UV exposure inhibited increased cutaneous CGRP and blocked immunosuppression. These findings support a role for CGRP in the local immunosuppression caused by chronic, repeated subinflammatory UV exposure.

Influence of thermode size for detecting heat pain dysfunction in a capsaicin model of epidermal nerve fiber loss

Quantitative sensory testing of heat pain sensation has become an important tool to evaluate small caliber afferent nerve function in peripheral neuropathy. In earlier studies, we found that topical application of capsaicin in humans results in the loss of epidermal nerve fibers (ENFs) with a corresponding decrease in detection of heat pain sensation. Capsaicin may therefore be a useful model for developing optimal psychophysical testing procedures for detection of neuropathy in its early stages. Here we determined the influence of thermal probe (thermode) size in detecting the diminished heat pain sensation following capsaicin application. Twelve healthy volunteers applied 0.075% capsaicin topically to the volar forearm four times daily for 7 days. Psychophysical measures of heat pain, mechanical (sharp) pain, and tactile threshold were obtained daily from untreated control skin and from capsaicin-treated skin during capsaicin application, and once weekly for 5 weeks following discontinuation of capsaicin. Heat pain sensation was assessed using a large (30×30 mm) and small (3×3 mm) thermode and different algorithms to assess pain threshold and suprathreshold heat pain. Skin biopsies were obtained and were processed for immunohistochemical localization of (ENFs) using the pan neuronal marker protein gene product 9.5. Capsaicin produced a rapid decrease in the number of ENFs, with nearly complete disappearance after 3 days of treatment. Heat pain evoked by the small, but not the large, thermode decreased dramatically after capsaicin treatment. The sensation of heat pain returned toward normal after 2–3 weeks following discontinuation of capsaicin treatment concordant with gradual reinnervation of the epidermis. Regression analysis indicated that the sensation of heat pain evoked by the small thermode correlated much better with the number of ENFs than heat pain evoked by the large thermode. The detection of sharp pain decreased moderately after capsaicin treatment. Assessment of heat pain sensation using small thermodes has potential for detecting sensory deficits in early stages of small fiber neuropathy.

EFFECTS OF NEUTRALIZING ANTIBODIES TO TNF‐ALPHA ON PAIN‐RELATED BEHAVIOR AND NERVE REGENERATION IN MICE WITH CHRONIC CONSTRICTION INJURY

Inhibition of proinflammatory cytokines reduces hyperalgesia in animal models of painful neuropathy. We set out to investigate the consequences of this treatment for nerve regeneration. Here we examined the sequels of epineurial application of neutralizing antibodies to tumor necrosis factor‐alpha (TNF) in chronic constriction injury (CCI) of the sciatic nerve in C57/BL 6 mice. The mice were tested behaviorally for manifestations of thermal hyperalgesia and mechanical allodynia. Nerve regeneration was assessed by morphometry of myelinated nerve fibers in the sciatic nerve and of the epidermal innervation density in the glabrous skin of the hindpaws. Antibodies to TNF reduced thermal hyperalgesia and mechanical allodynia after CCI. Myelinated fiber density in the sciatic nerve was reduced to 30% of normal on day 7 after surgery, and reached 60% on day 45, with no difference between antibody‐treated and untreated animals. Epidermal innervation density as shown by PGP 9.5 and CGRP immunohistochemistry was reduced to 25–47% at both time points after CCI, again without differences between antibody treated and untreated mice. Myelinated fiber density but not epidermal innervation density was correlated to thermal and mechanical withdrawal thresholds. We conclude that neutralization of endoneurial TNF attenuates pain related behavior but has no effect on nerve regeneration. Furthermore, the number of epidermal nerve fibers is not relevant to the magnitude of behavioral hyperalgesia in CCI.

Effects of neutralizing antibodies to TNF-alpha on pain-related behavior and nerve regeneration in mice with chronic constriction injury

Inhibition of proinflammatory cytokines reduces hyperalgesia in animal models of painful neuropathy. We set out to investigate the consequences of this treatment for nerve regeneration. Here we examined the sequels of epineurial application of neutralizing antibodies to tumor necrosis factor-alpha (TNF) in chronic constriction injury (CCI) of the sciatic nerve in C57/BL 6 mice. The mice were tested behaviorally for manifestations of thermal hyperalgesia and mechanical allodynia. Nerve regeneration was assessed by morphometry of myelinated nerve fibers in the sciatic nerve and of the epidermal innervation density in the glabrous skin of the hindpaws. Antibodies to TNF reduced thermal hyperalgesia and mechanical allodynia after CCI. Myelinated fiber density in the sciatic nerve was reduced to 30% of normal on day 7 after surgery, and reached 60% on day 45, with no difference between antibody-treated and untreated animals. Epidermal innervation density as shown by PGP 9.5 and CGRP immunohistochemistry was reduced to 25–47% at both time points after CCI, again without differences between antibody treated and untreated mice. Myelinated fiber density but not epidermal innervation density was correlated to thermal and mechanical withdrawal thresholds. We conclude that neutralization of endoneurial TNF attenuates pain related behavior but has no effect on nerve regeneration. Furthermore, the number of epidermal nerve fibers is not relevant to the magnitude of behavioral hyperalgesia in CCI.

Topical capsaicin in humans: parallel loss of epidermal nerve fibers and pain sensation

Capsaicin applied topically to human skin produces itching, pricking and burning sensations due to excitation of nociceptors. With repeated application, these positive sensory responses are followed by a prolonged period of hypalgesia that is usually referred to as desensitization, or nociceptor inactivation. Consequently, capsaicin has been recommended as a treatment for a variety of painful syndromes. The precise mechanisms that account for nociceptor desensitization and hypalgesia are unclear. The present study was performed to determine if morphological changes of intracutaneous nerve fibers contribute to desensitization and hypalgesia. Capsaicin (0.075%) was applied topically to the volar forearm four times daily for 3 weeks. At various time intervals tactile, cold, mechanical and heat pain sensations were assessed in the treated and in contralateral untreated areas. Skin blisters and skin biopsies were collected and immunostained for protein gene product (PGP) 9.5 to assess the morphology of cutaneous nerves and to quantify the number of epidermal nerve fibers (ENFs). Capsaicin resulted in reduced sensitivity to all cutaneous stimuli, particularly to noxious heat and mechanical stimuli. This hypalgesia was accompanied by degeneration of epidermal nerve fibers as evidenced by loss of PGP 9.5 immunoreactivity. As early as 3 days following capsaicin application, there was a 74% decrease in the number of nerve fibers in blister specimens. After 3 weeks of capsaicin treatment, the reduction was 79% in blisters and 82% in biopsies. Discontinuation of capsaicin was followed by reinnervation of the epidermis over a 6-week period with a return of all sensations, except cold, to normal levels. We conclude that degeneration of epidermal nerve fibers contributes to the analgesia accredited to capsaicin. Furthermore, our data demonstrate that ENFs contribute to the painful sensations evoked by noxious thermal and mechanical stimuli.

Quantitation of epidermal nerves in diabetic neuropathy.

We describe methods to quantify epidermal nerve fibers (ENFs) in skin biopsy specimens from diabetic candidates for pancreas transplantation and control subjects. ENFs and the dermal-epidermal basement membrane were stained by immunohistochemical methods, imaged with a confocal microscope, and quantified using a neuron tracing system. The number of ENFs per surface of epidermis was diminished in diabetic subjects. ENF number and summed length of all ENFs per volume of epidermis examined were also decreased. Length and number of branch points of single surviving ENFs were similar in skin of control and diabetic subjects. The methods and results constitute a basis for continued study of the effects of the euglycemia that attends successful pancreas transplantation and the effects of therapy in patients with various types of polyneuropathy.