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Abnormal unmyelinated fiber density in skin and nerve biopsies. (A) Immunostaining for Protein Gene Product 9.5 of a skin biopsy examined by optical microscopy. There is an absence of unmyelinated fibers. White arrow, dermo- epidermal junction. (B) Electron microscopy shows absence of normal unmyelinated fibers. RF, regressive unmyelinated fiber; N, Schwann cell nucleus; C, stacks of Schwann cytoplasm without any axon; black arrow, collagen pocket.
Source publication
Peripheral polyneuropathies are common and their diagnosis may be challenging. We compared the results from sural-nerve and skin biopsies in 33 patients with a polyneuropathy and neuropathic pain examined in our hospital over a 6-year period. The biopsies were all from the same lower limb of each patient. Intraepidermal nerve fiber (IENF) densities...
Citations
... Consequently, there is a high risk of ulceration with subsequent distal amputation in lower limbs [6,7]. During the last decade, diagnostic tests such as skin biopsy and corneal confocal microscopy have confirmed its usefulness in the diagnosis of peripheral neuropathies through quantitative analysis of Aδ and C fine intraepithelial nerve fibers [3,[8][9][10][11]. Thus, more severe small fiber damage in the skin of patients with painful diabetic neuropathy compared with painless diabetic neuropathy has been observed, and the density of intraepithelial nerve fibers was lower in subjects with painful compared with painless neuropathy [11,12]. ...
Distal diabetic sensorimotor polyneuropathy (DDSP) is the most prevalent form of diabetic neuropathy, and some of the patients develop gradual pain. Specialized sensory structures present in the skin encode different modalities of somatosensitivity such as temperature, touch, and pain. The cutaneous sensory structures responsible for the qualities of mechanosensitivity (fine touch, vibration) are collectively known as cutaneous mechanoreceptors (Meissner corpuscles, Pacinian corpuscles, and Merkel cell-axonal complexes), which results are altered during diabetes. Here, we used immunohistochemistry to analyze the density, localization within the dermis, arrangement of corpuscular components (axons and Schwann-like cells), and expression of putative mechanoproteins (PIEZO2, ASIC2, and TRPV4) in cutaneous mechanoreceptors of subjects suffering clinically diagnosed non-painful and painful distal diabetic sensorimotor polyneuropathy. The number of Meissner corpuscles, Pacinian corpuscles, and Merkel cells was found to be severely decreased in the non-painful presentation of the disease, and almost disappeared in the painful presentation. Furthermore, there was a marked reduction in the expression of axonal and Schwann-like cell markers (with are characteristics of corpuscular denervation) as well as of all investigated mechanoproteins in the non-painful distal diabetic sensorimotor polyneuropathy, and these were absent in the painful form. Taken together, these alterations might explain, at least partly, the impairment of mechanosensitivity system associated with distal diabetic sensorimotor polyneuropathy. Furthermore, our results support that an increasing severity of DDSP may increase the risk of developing painful neuropathic symptoms. However, why the absence of cutaneous mechanoreceptors is associated with pain remains to be elucidated.
... Consequently, there is a high risk of ulceration with subsequent distal amputation in lower limbs [6,7]. During the last decade, diagnostic tests such as skin biopsy and corneal confocal microscopy have confirmed its usefulness in the diagnosis of peripheral neuropathies through quantitative analysis of Aδ and C fine intraepithelial nerve fibers [3,[8][9][10][11]. Thus, more severe small fiber damage in the skin of patients with painful diabetic neuropathy compared with painless diabetic neuropathy has been observed, and the density of intraepithelial nerve fibers was lower in subjects with painful compared with painless neuropathy [11,12]. ...
Pannexins are channel proteins displaying functional similarities to gap junctions in vertebrates and are regarded as transmembrane ATP-releasing channels. A member of this family, denominate pannexin1, has been detected in the epidermis and cutaneous adnexal structures. Here we used immunohistochemistry to investigate whether human digital Merkel cells express this protein since ATP is postulated as a neurotransmitter in the Merkel cell-axon complexes low-threshold mecahoreceptors. Pannexin1 immunoreactivity was found in cytokeratine 20-, chromogranin A- and synaptophysin-positive cells placed at the basal layer of the epidermis. Cell displaying pannexin1 immunoreactivities were thus identified as Merkel cells and showed close contact with nerve profiles. Light pannexin1 immunoreactivity in dermal blood vessels was also verified. Present results demonstrate for the first time the expression of pannexin1 in human digital Merkel cells supporting the idea that ATP can be involved directly or indirectly in the mechanotransductional process at Merkel-axon complexes.
... A 5-cm nerve sample was surgically removed under local anesthesia from a clinically or electrophysiologically affected sural or superficial peroneal nerve. One part of the nerve sample was frozen and cut to 7-lm-thickness and another part of the nerve sample was fixed in 2.5% glutaraldehyde to make semithin section (1 mm) as described elsewhere (19). ...
Polyneuropathy, organomegaly, endocrinopathy, monoclonal protein, skin changes (POEMS) syndrome is a rare systemic disorder linked to plasma cell dyscrasia and is related to elevation of vascular endothelial growth factor (VEGF). Diagnosis is still challenging and pathophysiology unclear. Because VEGF drives neovascularization, we investigated skin and nerve vascularization in 6 patients with POEMS syndrome compared with 5 control groups of polyneuropathies and healthy subjects (n = 104) from the University Hospital of Limoges between 2009 and 2018. We evaluated loss of small and large fibers in these patients. Skin and nerve vascularization were quantified manually on immunofluorescence using vessel staining (anti-α-SMA antibody). Dermal vascularization was significantly higher in POEMS syndrome than in other groups, but unrelated to loss of small fibers and VEGF. Perineurial vascularization was higher in POEMS syndrome than in healthy controls, and was related to loss of large fibers and VEGF level. Our study highlights the existence of neovascularization in skin of patients with this rare disorder. These data suggest that skin neovascularization could be an additional biomarker to help in the diagnosis and understanding of POEMS syndrome. Moreover, nerve neovascularization, driven by VEGF overexpression, may play a role in the pathophysiology of large fiber loss in this condition.
... In turn, this implies that small-diameter axons will have lower thresholds for responding to such treatments than will large-diameter axons. Since extracellular electrical currents, a major modality for affecting neural activity, preferentially affect large-diameter axons before affecting small-diameter axons (Rattay 1986), the results presented in this study open up a range of new approaches to analyzing nervous system function, and new ways to alter physiology or to treat diseases that selectively affect small-diameter unmyelinated axons (Dubin and Patapoutian 2010, Duchesne et al 2016, Pekala et al 2016. ...
Objective:
Thermal block of action potential conduction using infrared lasers is a new modality for manipulating neural activity. It could be used for analysis of the nervous system and for therapeutic applications. We sought to understand the mechanisms of thermal block.
Approach:
To analyze the mechanisms of thermal block, we studied both the original Hodgkin/Huxley model, and a version modified to more accurately match experimental data on thermal responses in the squid giant axon.
Main results:
Both the original and modified models suggested that thermal block, especially at higher temperatures, is primarily due to a depolarization-activated hyperpolarization as increased temperature leads to faster activation of voltage-gated potassium ion channels. The minimum length needed to block an axon scaled with the square root of the axon's diameter.
Significance:
The results suggest that voltage-dependent potassium ion channels play a major role in thermal block, and that relatively short lengths of axon could be thermally manipulated to selectively block fine, unmyelinated axons, such as C fibers, that carry pain and other sensory information.
... A single skin biopsy was performed under local anaesthesia with 1% xylocaine, with a 4-mm punch on 7-mm depth, on the lower limb (10 cm above the lateral malleolus) on the side predominantly affected by PD. Immunofluorescence was performed using protein gene product 9.5 immunostaining and intraepidermal nerve fibre (IENF) density was quantified at high magnification (×400) with a light microscope by two blinded operators as previously described [28]. ...
Dysautonomic symptoms are frequent non-motor complaints in patients with Parkinson's disease. Numerous neuropathological studies have shown that Lewy bodies and neurites, the pathological hallmarks of Parkinson's disease, are widely distributed throughout the peripheral autonomic nervous systems and across end organs. However, few investigations integrally explored the symptoms and physiology of dysautonomia in Parkinson's disease. We, therefore, performed a comprehensive evaluation of the autonomic function in a prospective group of 45 patients with idiopathic Parkinson's disease. Autonomic components (pupillomotor, tear, salivary, cardiovascular, digestive, urinary, sexual, sudomotor functions and skin sensitivity) were evaluated using questionnaires and functional tests. Skin biopsy was performed for intraepidermal nerve fibre density quantification. In addition, all patients underwent polysomnography and a complete neuropsychological and neurological assessment. The analysis association of autonomic components showed that dysautonomic signs and symptoms were heterogeneously distributed among patients. Skin denervation as assessed by intraepidermal nerve fibre density quantification was only associated with quantitative thermal sensory testing (OR = 12.0, p = 0.02), constipation (OR = 5.5, p = 0.01) and ocular dryness symptoms (OR = 8.29, p = 0.04). Cognitive alteration was associated with cardiovascular symptoms (OR = 4.33, p = 0.03) and dysfunction (OR = 5.83, p = 0.02) as well as with constipation (OR = 5.38, p = 0.02). Axial motor impairment and rapid eye movement (REM) sleep behaviour disorder were not related to any of the autonomic complaint or dysfunction. Our results show that autonomic functions are affected in a heterogeneous pattern in Parkinson's disease, thereby suggesting that the progression of autonomic dysfunction follows an erratic rather than a stepwise progression.
... Contact heat-evoked potential explores the electrophysiology mediated peripherally by Ad nerve fibers, 14 and skin innervation demonstrated by skin biopsies represent the free nerve endings of C nerve fibers in the majority and Ad nerve fibers in the minority. 13,31 In this study, we investigated the relationship between CHEPs and skin innervation in both patients with neuropathy and age-and sex-matched controls. Contact heatevoked potential amplitudes were highly correlated with the skin innervation and were the most significant predictor of skin denervation compared with other parameters in patients with neuropathy. ...
Contact heat-evoked potentials (CHEPs) have become an established method of assessing small-fiber sensory nerves; however, their potential as a physiological signature of neuropathic pain symptoms has not been fully explored. To investigate the diagnostic efficacy in examining small-fiber sensory nerve degeneration, the relationship with skin innervations, and clinical correlates with sensory symptoms, we recruited 188 patients (115 men) with length-dependent sensory symptoms and reduced intraepidermal nerve fiber (IENF) density at the distal leg to perform CHEP, quantitative sensory testing, and nerve conduction study. Fifty-seven age- and sex-matched controls were enrolled for comparison of CHEP and skin innervation. Among patients with neuropathy, 144 patients had neuropathic pain and 64 cases had evoked pain. Compared with quantitative sensory testing and nerve conduction study parameters, CHEP amplitudes showed the highest sensitivity for diagnosing small-fiber sensory nerve degeneration and exhibited the strongest correlation with IENF density in multiple linear regression. Contact heat-evoked potential amplitudes were strongly correlated with the degree of skin innervation in both patients with neuropathy and controls, and the slope of the regression line between CHEP amplitude and IENF density was higher in patients with neuropathy than in controls. Patients with evoked pain had higher CHEP amplitude than those without evoked pain, independent of IENF density. Receiver operating characteristic analysis showed that CHEP had better performance in diagnosing small-fiber sensory nerve degeneration than thermal thresholds. Furthermore, CHEPs showed superior classification accuracy with respect to evoked pain. In conclusion, CHEP is a sensitive tool to evaluate pathophysiology of small-fiber sensory nerve and serves as a physiological signature of neuropathic pain symptoms.
Résumé
L’étude de la densité des fibres nerveuses intra-épidermiques sur la biopsie cutanée est l’un des outils diagnostiques clé pour valider le diagnostic de la neuropathie à petites fibres. Elle doit être réalisée selon un protocole strict lors de toutes les étapes de sa prise en charge. La densité en fibres nerveuses intra-épidermiques obtenue est comparée à des tables de normes en fonction de l’âge et du sexe. Une densité en fibres nerveuses intra-épidermiques est considérée comme pathologique si elle est inférieure au cinquième percentile de ces normes et devient alors un critère diagnostique de neuropathie à petites fibres.
Background
Currently, assessment of unmyelinated axon regeneration is limited to electron microscopy (EM), which is expensive, time consuming and not universally available. This study presents a protocol to estimate the number of unmyelinated axons in a regenerating peripheral nerve without the need for electron microscopy.
New method
The common peroneal nerve of Sprague-Dawley rats was transected, repaired and regenerated for 4 weeks. Two distal adjacent segments of the regenerating nerve were then processed for either conventional histomorphometry using toluidine blue or immunolabeling of neurofilament protein. Myelinated axon and total axon counts were obtained, respectively, to generate estimates of unmyelinated axon numbers, which were then compared to unmyelinated axon counts using EM from the same specimens. For comparison, unmyelinated axons were counted in an uninjured rat laryngeal nerve.
Results
After 4 weeks of regeneration, the estimated number of regenerating unmyelinated axons was 4044 ± 232 using this technique, representing 81.3% of the total axonal population. By comparison, the proportion of unmyelinated axons in the uninjured laryngeal nerve was 55% of the total axonal population.
Comparison with existing method
These estimates correlate with electron microscopy measurements, both in terms of the proportion of unmyelinated axons and also by linear regression analysis.
Conclusions
The neurofilament staining method correlates with electron microscopy estimates of the same nerve sections. It is useful for the efficient counting of unmyelinated axons in the regenerating peripheral nerve and can be used by laboratories that do not have access to EM facilities.
Objective
Small‐fiber sensory and autonomic symptoms are early presentations of familial amyloid polyneuropathy (FAP) with transthyretin (TTR) mutations. This study aimed to explore the potential of skin nerve pathologies as early and disease‐progression biomarkers and their relationship with skin amyloid deposits.
Methods
Skin biopsies were performed in patients and carriers to measure intraepidermal nerve fiber (IENF) density, sweat gland innervation index of structural protein gene product 9.5 [SGII(PGP9.5)] and peptidergic vasoactive intestinal peptide [SGII(VIP)], and cutaneous amyloid index. These skin pathologies were analyzed with clinical disability assessed by FAP stage score (stage 0~4) and compared to neurophysiological and psychophysical tests.
Results
There were 70 TTR‐mutant subjects (22 carriers and 48 patients) and 66 cases were TTR‐A97S. Skin nerve pathologies were distinct according to stage. In carriers, both skin denervation and peptidergic sudomotor denervation were evident: (1) IENF density was gradually reduced from stage 0 through 4, and (2) SGII(VIP) was markedly reduced from stage 1 to 2. In contrast, SGII(PGP9.5) was similar between carriers and controls, but it declined in patients from stage 2. Skin amyloids were absent in carriers and became detectable from stage 1. Cutaneous amyloid index was correlated with SGII(PGP9.5) and stage in a multivariable mixed‐effect model. When all tests were compared, only IENF density, SGII(PGP9.5), and cutaneous amyloid index were correlated with stage, and IENF density had the highest abnormal rate in carriers.
Interpretation
Biomarkers of sensory and sudomotor innervation exhibited a stage‐dependent progression pattern, with sensory nerve degeneration as the early skin nerve pathology.
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Background
The study aimed to investigate the physiology, psychophysics, pathology and their relationship in reversible nociceptive nerve degeneration, and the physiology of acute hyperalgesia.
Methods
We enrolled 15 normal subjects to investigate intraepidermal nerve fibre (IENF) density, contact heat‐evoked potential (CHEP) and thermal thresholds during the capsaicin‐induced skin nerve degeneration–regeneration; and CHEP and thermal thresholds at capsaicin‐induced acute hyperalgesia.
Results
After 2‐week capsaicin treatment, IENF density of skin was markedly reduced with reduced amplitude and prolonged latency of CHEP, and increased warm and heat pain thresholds. The time courses of skin nerve regeneration and reversal of physiology and psychophysics were different: IENF density was still lower at 10 weeks after capsaicin treatment than that at baseline, whereas CHEP amplitude and warm threshold became normalized within 3 weeks after capsaicin treatment. Although CHEP amplitude and IENF density were best correlated in a multiple linear regression model, a one‐phase exponential association model showed better fit than a simple linear one, that is in the regeneration phase, the slope of the regression line between CHEP amplitude and IENF density was steeper in the subgroup with lower IENF densities than in the one with higher IENF densities. During capsaicin‐induced hyperalgesia, recordable rate of CHEP to 43 °C heat stimulation was higher with enhanced CHEP amplitude and pain perception compared to baseline.
Conclusions
There were differential restoration of IENF density, CHEP and thermal thresholds, and changed CHEP–IENF relationships during skin reinnervation. CHEP can be a physiological signature of acute hyperalgesia.
Significance
These observations suggested the relationship between nociceptive nerve terminals and brain responses to thermal stimuli changed during different degree of skin denervation, and CHEP to low‐intensity heat stimulus can reflect the physiology of hyperalgesia.
