Dermal Nerve Research Articles

Abstracts of the 8th Meeting of the Italian Peripheral Nerve Study Group: 70

The availability of antibodies against the pan‐axonal marker protein‐gene‐protein 9.5 (PGP) allowed to definitely demonstrate the existence of small‐diameter nerve fibers within the vital layers of the epidermis and prompted to their quantification in sensory neuropathies. However, morphological features of intra‐epidermal nerve fibers (IENF) have not been fully characterized. We performed immunohistochemical and confocal immunofluorescence studies in skin biopsies taken in 15 healthy subjects at the proximal thigh or the distal leg using anti‐PGP, anti‐unique b‐tubulin (TuJ1), anti‐non‐phosphorylated microtubule‐associated protein‐1B (MAP‐1B), anti‐70 and 200 kDa neurofilament (NF), anti‐phosphorylated neurofilament‐312 (SMI), anti‐peripheral myelin protein‐22 (PMP22), anti‐basic myelin protein (MBP), and anti‐myelin associated glycoprotein (MAG) antibodies. IENF density was quantified under light microscope using PGP, TuJ1, MAP‐1B, NF, and SMI immunostaining. In each site, the density of TuJ1‐ and MAP‐1B‐positive IENF was similar to that of PGP‐positive IENF, whereas that of NF‐positive IENF was significantly lower. Only sporadic SMI‐positive IENF were found. Double confocal staining studies confirmed that TuJ1 and MAP‐1B diffusely co‐localized with PGP in IENF. These findings suggest that IENF cytoskeleton is predominantly composed by tubules, whereas neurofilaments are less represented. No IENF showed PMP22, MBP, or MAG labeling. All dermal nerve fibers presented PMP22 immunoreactivity, which labeled Schwann cell membrane, whereas only a few of them were MBP‐ or MAG‐positive, confirming that most fibers are unmyelinated. Double staining confocal microscope studies revealed that PMP22 labeling in dermal nerves stopped at the dermal‐epidermal junction, whereas TuJ1‐positive axons crossed it and run between keratinocytes. This definitely demonstrates that IENF are naked axons, arising from sub‐papillary unmyelinated nerves which loose the Schwann cell at the dermal‐epidermal junction. The presence of naked nerve fibers within the epidermis suggests that functional interactions with the resident cells, even in sensory transduction, might exist.

Cutaneous innervation in Guillain-Barre syndrome: pathology and clinical correlations

Guillain-Barré syndrome (GBS) is traditionally considered to be a large-fibre neuropathy. However, the presence of hypo-aesthesia, dysaesthesia and dysautonomia in GBS patients raises the possibility that small-diameter sensory and autonomic nerves may also be affected. To investigate small-fibre neuropathy in GBS, we performed a skin biopsy from the distal leg of 20 patients with the demyelinating form of GBS. Skin sections were immunohistochemically stained with antiserum against protein gene product 9.5 (PGP 9.5), a ubiquitin C-terminal hydrolase. Cutaneous innervation was evaluated by measuring epidermal nerve density (END), and END was further correlated with various clinical and electrophysiological parameters. In GBS patients, END values were much lower than in age- and gender-matched control subjects (5.03 +/- 1.18 versus 10.16 +/- 0.87 fibres/mm, P < 0.001). Eleven patients (55%) had reduced epidermal innervation with pathological evidence of active nerve degeneration in the dermis: fragmentation of subepidermal nerve plexuses and a beaded appearance of dermal nerves. GBS patients had significantly elevated thermal thresholds with higher warm threshold temperatures (44.54 +/- 1.04 versus 39.00 +/- 0.35 degrees C, P < 0.001) and lower cold threshold temperatures (25.57 +/- 1.11 versus 29.05 +/- 0.21 degrees C, P = 0.032). Reduced END values were associated with an elevated warm threshold (P = 0.027), ventilatory distress (P = 0.037) and dysautonomia (P = 0.001). END values were negatively correlated with disability grade on a scale of 1-6 (slope -0.134 +/- 0.038, P = 0.0018). Patients with reduced END values tended to have a slower recovery than those with normal END values (P = 0.013, median time 12 versus 2 weeks). Patho logically, sudomotor innervation of the skin was reduced in five of 17 (29.4%) GBS patients in whom sweat glands could be recognized. These findings suggest that small-fibre sensory and autonomic neuropathies exist in a significant proportion of GBS patients, and that END values are correlated with functional disabilities. In summary, GBS should be considered a global neuropathy instead of a pure large-fibre neuropathy.

Notalgia Paresthetica with a Significant Increase in the Number of Intradermal Nerves

Notalgia paresthetica is an isolated mononeuropathy involving the skin over or near the scapula. The cause remains unknown. We histologically observed a significant increase in the number of dermal nerves in a case of notalgia paresthetica. Immunohistochemical examination using a neural marker, S-100, positively stained the nerves. Interestingly, a biopsy from perilesional skin also showed an abnormal nerve proliferation.

Localization of μ-opioid receptor 1A on sensory nerve fibers in human skin

Opioid peptides are endogenous neuromodulators that play a major role in the nociceptive pathway by interacting with opioid receptors. So far, four opioid receptors (μ-, δ-, κ-, orphan-receptor) have been cloned with a wide distribution in the central and peripheral nervous system. In the present study, we give first evidence for the presence of the μ-opioid receptor (MOR) isoform 1A in nerve fibers of human skin. Immunohistochemical analysis revealed MOR immunoreactivity to be present in dermal and epidermal nerve fibers. Double-immunofluorescence staining revealed that MOR is present on calcitonin gene-related protein (CGRP)-positive sensory nerve fibers, while autonomic nerves of blood vessels, hair follicles, or skin glands were negative. In diseased skin such as psoriasis vulgaris, atopic dermatitis, and prurigo nodularis, distribution of MOR 1A immunoreactivity was similar to that of normal skin. These findings expand our knowledge about a direct regulatory role of cutaneous opioid receptors in the skin. Thus, peripheral cutaneous opioid receptors may be involved in the transmission of pain and pruritus, respectively. This is supported by previous observation that opioid receptor antagonists may significantly diminish experimentally evoked histamine-induced itch of the skin. Together, our findings contribute to the understanding of the high antipruritic potency of opioid receptor antagonists in various skin and systemic diseases.

Neuropathology of Skin Denervation in Acrylamide-Induced Neuropathy

Previous studies have established the neurotoxicity and pathology of acrylamide to large-diameter nerves. It remains unclear (1) whether small-diameter sensory nerves are vulnerable to acrylamide and (2) if so, how the pathology evolves during intoxication. We investigated the influence of acrylamide on small-diameter sensory nerves by studying the pathology of sensory nerve terminals in the skin. The neurotoxic effects of acrylamide (400 ppm in drinking water) on mice were assessed by immunostaining the skin with protein gene product 9.5, a ubiquitin C-terminal hydrolase, particularly useful for demonstrating cutaneous nerve terminals. Within 5 days of acrylamide administration (the initial stage), epidermal nerves showed two major changes: (1) terminal swelling and (2) increased branching. There was a progressive reduction in epidermal nerve density (END) thereafter. Fifteen days after acrylamide intoxication (the late stage), reduction in END became evident (25.22 ± 2.19 fibers/mm vs 41.74 ± 2.60 fibers/mm in control mice, P < 0.003). At this stage, there was significant dermal nerve degeneration with ultrastructural demonstrations of vacuolar changes. These findings establish the pathological consequences of acrylamide neurotoxicity in cutaneous sensory nerves with far-reaching implications: (1) providing an animal system to study “dying-back” pathology of nociceptive nerves and (2) forming the ultrastructural foundation for interpreting the pathology of cutaneous nerve degeneration in skin biopsies.

Local Skin Lesions in the Rat after Subcutaneous Deposition of Capsaicin

Capsaicin is used to investigate the role of peripheral sensory nerve fibers. In previous studies of rats treated by injection of capsaicin into the skin of the neck, ‘spontaneous’ lesions in the head and neck region were observed. In this study, the course of development over time, the regional distribution and the innervation of capsaicin-induced dermal lesions were assessed in young male Sprague-Dawley rats. In one experiment, capsaicin was administered subcutaneously by injection in the skin of the neck. In a second experiment, capsaicin was injected in the back by a long needle that tunneled under the skin and allowed the capsaicin to be deposited in the subcutaneous fat of the neck. The density and the distribution of dermal nerve fibers were investigated by immunohistochemistry, using antisera against a panneuronal marker, protein gene product 9.5 (PGP), and calcitonin gene-related peptide (CGRP). In the first experiment, rats developed lesions in the neck area 11 days after injection. In the second experiment, lesions appeared in the skin of the back and occasionally in the neck area 10 days after injection. Development of lesions in the afflicted areas was paralleled by local reduction in the density of CGRP-immunoreactive (IR) nerve fibers, 80% in the first experiment and 72% in the second. The number of PGP-IR fibers was likewise reduced, by 39 and 41%, respectively. The density of the CGRP-IR fibers in the wound area was the same as in the adjacent, nonlesioned skin. The healing of the capsaicin-induced lesions was slow compared with surgical wounds in control animals. The wounds healed with hypertrophic scars. The healing process in the skin of the back was associated with the proliferation of CGRP-IR fibers. The study shows cutaneous lesions to appear in the region of the subcutaneous deposition of capsaicin. A uniform depletion of capsaicin-sensitive nerve fibers in the area of deposition suggests that an additional factor is needed to induce lesions. Possibly, impaired nociception in the afflicted area results in more vigorous grooming behavior and this, in turn, in a local skin damage.

Developmental timing of hair follicle and dorsal skin innervation in mice.

The innervation of hair follicles offers an intriguing, yet hardly studied model for the dissection of the stepwise innervation during cutaneous morphogenesis. We have used immunofluorescence and a panel of neuronal markers to characterize the developmental choreography of C57BL/6 mouse backskin innervation. The development of murine skin innervation occurs in successive waves. The first cutaneous nerve fibers appeared before any morphological evidence of hair follicle development at embryonic day 15 (E15). Stage 1 and 2 developing hair follicles were already associated with nerve fibers at E16. These fibers approached a location where later in development the follicular (neural) network A (FNA) is located on fully developed pelage hair follicles. Prior to birth (E18), some nerve fibers had penetrated the epidermis, and an additional set of perifollicular nerve fibers arranged itself around the isthmus and bulge region of stage 5 hair follicles, to develop into the follicular (neural) network B (FNB). By the day of birth (P1), the neuropeptides substance P and calcitonin gene-related peptide became detectable in subcutaneous and dermal nerve fibers first. Newly formed hair follicles on E18 and P1 displayed the same innervation pattern seen in the first wave of hair follicle development. Just prior to epidermal penetration of hair shafts (P5), peptide histidine methionine-IR nerve fibers became detectable and epidermal innervation peaked; such innervation decreased after penetration (P7- P17). Last, tyrosine hydroxylase-IR and neuropeptide Y-IR became readily detectable. This sequence of developing innervation consistently correlates with hair follicle development, indicating a close interdependence of neuronal and epithelial morphogenesis.

Reduction in vesicant-induced cellular inflammation and hyperalgesia by local injection of corticotropin releasing factor in rabbit eyelid

This study tested the ability of corticotropin releasing factor (CRF) to mitigate inflammatory cell infiltration and hyperalgesia after a direct subcutaneous doxorubicin injection, a vesicant drug that causes localized inflammation at the site of subcutaneous exposure. This study used 62 adult New Zealand white rabbits. Doxorubicin (1-2 mg), followed by either CRF (300, 150, 75, 35, 20 or 10 microg in 1 ml saline) or saline only, was injected into pairs of rabbit eyelids. One set of doxorubicin and CRF treated eyelid pairs also received injection of a CRF antagonist into one eyelid. The eyelid tissue was assayed from 1-7 days later for morphometric changes in CD11b-positive inflammatory cell infiltrate. Hyperalgesia was assessed using the blink response to von Frey hair stimulation. Epidermal and dermal nerve fibers were visualized immunochemically using an antibody to PGP 9.5. The cellular localization of corticotropin releasing factor was determined immunohistochemically in eyelid tissue. Doses of CRF from 35 to 150 microg significantly reduced inflammatory cell infiltration at all the post-injury time intervals examined, from 2 h post-injection up until 7 days. Doses of 10 and 20 microg CRF had no effect on local tissue inflammation caused by subcutaneous doxorubicin exposure. Doses of 35-150 microg CRF treatment significantly reduced the development of doxorubicin-induced hyperalgesia; this was maintained up to 4 days. Local CRF injections of 35-150 microg markedly reduced the doxorubicin induction of PGP-positive axons within the epidermis and subepidermis. CRF receptors were localized to the basal layer of the epithelium and to the subcutaneous nerve afferents. Local injection of CRF, after a subcutaneous exposure to the vesicant drug doxorubicin, significantly reduced the infiltration of CD11b-positive cells and reduced doxorubicin-induced hyperalgesia in the treated eyelids. This reduction in hyperalgesia appears to be a local peripheral effect, reducing nerve fibers within the subepidermal treatment area. This demonstrates the potential clinical usefulness of CRF because it could be used post-injury as a locally injected treatment; this potent local anti-inflammatory agent may have wide applications following drug, chemical, physical or surgical injury.

Quantitative pathology of cutaneous nerve terminal degeneration in the human skin.

Pathological diagnosis of neuropathy has traditionally depended on ultrastructural examinations of nerve biopsy specimens, particularly for sensory neuropathies affecting unmyelinated and small-myelinated nociceptive nerves. These sensory nerves terminate in the epidermis of the skin, and the pathology of neuropathy usually begins from nerve terminals. We investigated the feasibility of diagnosing small-fiber sensory neuropathy by evaluating cutaneous innervation. Skin biopsy specimens of 3-mm in diameter were obtained from the distal leg and the distal forearm of 55 healthy controls and 35 patients with sensory neuropathy. In the healthy controls, conventional intraepidermal nerve fiber densities (IENF densities) as measured using the image analysis system in the distal forearm and in the distal leg were correlated (r=0.55, P<0.0001), with significantly higher values in the distal forearm than in the distal leg (17.07+/-6.51 vs 12.92+/-5.33 fibers/mm, P<0.001). Compared to IENF densities of healthy controls, these values of neuropathic patients were significantly reduced in the distal forearm (5.82+/-6.50 fibers/mm, P<0.01) and in the distal leg (2.40+/-2.30, P<0.001). We further explored the possibility of quantifying skin innervation by counting "ocular intraepidermal nerve fiber density" (ocular nerve fiber density) with no aid of an image analysis system. This was based on the fact that the epidermal length on specifically defined sections was very close to the predicted epidermal length of 3 mm, the diameter of skin punches (P=0.14). Ocular nerve fiber densities were significantly correlated with IENF densities as measured by the image analysis system (r=0.99, P<0.0001). Dermal nerve fibers of neuropathic patients either disappeared or became degenerated. These findings were consistent with the notion of early terminal degeneration in neuropathy, and will facilitate quantitative interpretation of epidermal innervation in human neuropathy.

Expression of inducible nitric oxide synthase and nitrotyrosine in borderline leprosy lesions.

In the response to T-helper cell (Th1)-type cytokines and interactions with pathogens, high levels of nitric oxide (NO) are produced by activated macrophages expressing the inducible NO synthase (iNOS). The role and importance of reactive nitrogen intermediates (RNIs) such as NO and peroxynitrite in the host response to diseases caused by intracellular pathogens such as Mycobacterium leprae and M. tuberculosis is unclear. The aim of this study was to investigate the presence of local production of NO and peroxynitrite in borderline leprosy by using antibodies against iNOS and the product of peroxynitrite, nitrotyrosine (NT). We detected the presence of iNOS and NT in skin biopsies from borderline leprosy patients, with and without reversal reaction (RR), by immunohistochemistry (n = 26). In general, the granulomas from borderline leprosy lesions with and without RR showed high and specific expression of iNOS and NT. Moreover, strong immunoreactivity to iNOS and NT was observed in granulomas surrounding and infiltrating dermal nerves. The expression of iNOS and NT was also strong in keratinocytes, fibroblasts and endothelial cells in close relation to the granulomatous reaction. In contrast, normal human skin showed no expression of iNOS and NT in these cells. We conclude that iNOS and NT are expressed in granulomas from borderline leprosy patients with and without RR and propose that RNIs might be involved in the nerve damage following RR in leprosy.

Leishmanial antigens in the diagnosis of active lesions and ancient scars of American tegumentary leishmaniasis patients.

Cutaneous biopsies (n = 94) obtained from 88 patients with American tegumentary leishmaniasis were studied by conventional and immunohistochemical techniques. Specimens were distributed as active lesions of cutaneous leishmaniasis (n = 53) (Group I), cicatricial lesions of cutaneous leishmaniasis (n = 35) (Group II) and suggestive scars of healed mucosal leishmaniasis patients (n = 6) (Group III). In addition, active cutaneous lesions of other etiology (n = 24) (Group C1) and cutaneous scars not related to leishmaniasis (n = 10) (Group C2) were also included in the protocol. Amastigotes in Group I biopsies were detected by routine histopathological exam (30.2%), imprint (28.2%), culture (43.4%), immunofluorescence (41.4%) and immunoperoxidase (58.5%) techniques; and by the five methods together (79.3%). In Group II, 5.7% of cultures were positive. Leishmanial antigen was also seen in the cytoplasm of macrophages and giant cells (cellular pattern), vessel walls (vascular pattern) and dermal nerves (neural pattern). Positive reaction was detected in 49 (92.5%), 20 (57%) and 4 (67%) biopsies of Groups I, II and III, respectively. Antigen persistency in cicatricial tissue may be related to immunoprotection or, on the contrary, to the development of late lesions. We suggest that the cellular, vascular and neural patterns could be applied in the immunodiagnosis of active and cicatricial lesions in which leishmaniasis is suspected.

Expression of neuropeptide-degrading enzymes in alopecia areata: an immunohistochemical study.

Much clinical evidence suggests that the nervous system, including psychological factors, can influence the course of alopecia areata (AA). However, there has been little substantial evidence of specific participation of cutaneous neurogenic factors in the disease process. As previous studies have demonstrated that stress elicits the release of the neuropeptide substance P (SP) from peripheral nerves and that some patients with AA show prominent SP expression in nerves surrounding their hair follicles, we aimed to evaluate the role of SP in AA. We used immunohistochemistry to examine the expression of SP and SP-degrading enzymes in scalp biopsies from patients with AA and from healthy controls. Affected hair follicles in the centre of the areas of hair loss of patients with AA were richly innervated by SP-staining nerve fibres. Strong expression of the SP-degrading enzyme, neutral endopeptidase (NEP), was observed in hair follicles not only in the acute progressive phase of AA but also in the chronic stable phase. Expression of NEP in hair follicles from the margins of areas of hair loss was stronger than in normal controls, but was weaker than in the centre of the areas of hair loss. In addition, endothelial immunoreactivity for angiotensin-converting enzyme (also capable of degrading SP) was not observed in the centre of the areas of hair loss, which was in significant contrast to normal controls as well as to the margins of areas of hair loss where it was expressed. Further, intense expression of endothelial leucocyte adhesion molecule-1 on vessels and many degranulating mast cells was observed adjacent to affected hair follicles in AA, in admixture with dense lymphocytic inflammation. These findings suggest that SP is endogenously released by dermal nerve fibres around hair follicles and that it may play an important part in epithelial-mesenchymal-neuroectodermal interactions in AA. This study reveals that SP and its degrading enzymes are involved in the pathogenesis of AA, which in turn might explain the pathological significance of neurogenic and psychogenic aspects in the disease process.

Regeneration of cutaneous innervation in a human hand allograft

On Sept 23, 1998, a human hand allograft was done in our hospital. We followed up the patients for 24 months to assess the skin structure by immunohistology. Most cutaneous structures (including dermal nerves with their Schwann cells and perineurial fibroblasts) were present immediately after surgery and remained detectable throughout the study; from day 464 onwards, axons became detectable within dermal nerves, and their density increased progressively with time. Merkel cells reappeared in the epidermis 12 months after the operation. The regeneration of cutaneous innervation paralleled the recovery of cutaneous sensitivity.

Cutaneous nerve degeneration induced by acrylamide in mice

Acrylamide is a neurotoxin producing distal axonopathy. Previous studies mainly focused on large-diameter motor and sensory nerves, and the influences of acrylamide neurotoxicity on small-diameter sensory nerves in the skin remained elusive. We investigated skin innervation in mice intoxicated by acrylamide. Small-diameter sensory nerves in the skin degenerated after acrylamide intoxication. Epidermal nerve swelling was the earliest sign of acrylamide intoxication, with 29.5±2.4% of swollen epidermal nerves in the initial stage ( P<0.001). There was a trend of progressive loss of epidermal nerves with a significantly reduced epidermal nerve density in the late stage ( P<0.003). In the mean time, degenerating dermal nerves exhibited a beaded appearance. These results suggest the scenario of small-diameter cutaneous nerve degeneration in acrylamide neurotoxicity: beginning with epidermal nerve terminal swelling in the initial stage and resultant epidermal nerve depletion in the late phase.

Evaluation Of Diabetic Neuropathy By Skin Biopsy

Instead of nerve biopsy which is most reliable method for diagnosing diabetic neuropathy but rather harmful, we have examined the usefulness of skin biopsy to evaluate the grade of diabetic neuropathy and therapeutic effects of given compounds. Nerve fibers immunostained by antibodies against protein gene product (PGP)9.5 and labeled with streptavidin fluorescein isothiocyanate and measured under confocal laser scanning microscopy were significantly shorter in the epidermis and dermis and around sweat glands in diabetic patients than in healthy subjects. Sural nerve conduction velocity was significantly correlated with dermal nerve fiber length (NFL) (p &lt; 0.05) in diabetic patients. Patients with higher aldose reductase (AR) level of erythrocytes (&gt;10.8 ng/Hg) showed shorter dermal NFL than those with lower AR level (&lt;10.8)(p &lt; 0.05). Using this technique, we evaluated the therapeutic effect of AR inhibitor (ARI) epalrestat during approximately 13 months in diabetic patients with neuropathy (19, epalrestat‐treated; 12, control). The number of patients whose percent change of dermal NFL exceeded its mean ± 2SD in the control group was significantly higher in the ARI group than in the control group (p &lt; 0.05, 8/9 vs 1/12). These results suggest that quantitation of cutaneous nerves using biopsied skin samples may provide important information about the severity of diabetic neuropathy and that epalrestat can elongate dermal unmyelinated nerve fibers in at least part of diabetic patients.

Pathology of nerve terminal degeneration in the skin.

To characterize the pathology of epidermal nerve degeneration and regeneration, we investigated temporal and spatial changes in skin innervation of the mouse footpad. Within 24 hours after sciatic nerve axotomy, terminals of epidermal nerves appeared swollen and there was a mild reduction in epidermal nerve density (5.7 +/- 2.8 vs 12.7 +/- 2.2 fibers/mm, p < 0.04). Epidermal nerves completely disappeared by 48 hours (0.2 +/- 0.2 vs 14.2 +/- 0.9 fibers/mm, p < 0.001). Concomitant with the disappearance of epidermal nerves, the immunocytochemical pattern of the subepidermal nerve plexus became fragmented. At the electron microscopic level, the axoplasm of degenerating dermal nerves was distended with organelles and later became amorphous. Beginning from day 28 after axotomy, collateral sprouts from the adjacent saphenous nerve territory extended into the denervated area with a beaded appearance. They never penetrated the epidermal-dermal junction to innervate the epidermis. In contrast, 3 months after nerve crushing, the epidermis on the surgery side resumed a normal innervation pattern as the epidermis on the control side (10.3 +/- 3.9 vs 10.6 +/- 1.5 fibers/mm, p = 0.1). This study demonstrates the characteristics of degenerating and regenerating nerves, and suggests that successful reinnervation mainly originates from regenerating nerves of the original nerve trunks. All these findings provide qualitative and quantitative information for interpreting the pathology of cutaneous nerves.

Prevention of death of axotomized hypoglossal neurones and promotion of regeneration by chitin grafting.

1. Chitin is known to promote skin wound healing. In this study, chitin, prepared from Zuwai crab shell, was used as a bridge between the proximal and distal stumps of cut hypoglossal nerves in shrews. We compared the effects of chitin on the regeneration of transected right hypoglossal nerve axons, with those of porcine dermis, bovine dermal aterocollagen, and autologous nerve bundles. 2. To assess the survival of neurones, the size of neuronal cell body, and number of motoneurones were determined in the absence of any bridged material and in the presence of porcine dermis, bovine dermal aterocollagen, chitin, or autologous nerve bundles as a bridge. 3. Our results revealed a significantly better outcome in chitin and autologous nerve bridged groups; the size of neuronal cell body and number of hypoglossal neurones were higher than in the other groups. Chitin also enhanced the regeneration of neurones; the number of horseradish peroxide positive neurones indicative of repaired axonal processes was significantly higher in chitin and autologous nerve-bridged groups than in other groups. 4. Our results demonstrated that the use of chitin sheet or autograft successfully prevented the death of severed neurones and promoted the regeneration of the lesioned nerve. Although the mechanisms underlying the effects of chitin are still unknown, chitin seems to be a potentially useful biocompatible material for nerve repair and regeneration.

Influence of cutaneous nerves on keratinocyte proliferation and epidermal thickness in mice

We evaluated the influence of skin innervation on the epidermis in mice. The rich innervation of skin was demonstrated by immunocytochemistry with protein gene product 9.5, a ubiquitin carboxy hydrolase. Protein gene product-immunoreactive nerve fibers were in the epidermis, subepidermal plexus, dermal nerve trunks, and nerve terminals around sweat glands. Effects of denervation on the plantar surface of the hind foot was assessed by comparing the thickness of the epidermis, which was innervated by the sciatic nerve. Within 48 h after sectioning of the sciatic nerve, protein gene product (+)-nerves in the territory of the sciatic nerve were completely degenerated. There was a significant thinning of the denervated epidermis 72 h post-transection (30.5±1.1 vs 41.4±2.9 μm, 74±4% of the control side). The reduction in epidermal thickness persisted when skin remained denervated (69–75% of the control side). Incorporation of bromodeoxyuridine was reduced 24 h after denervation (71±6% of the control side). Reduction in bromodeoxyuridine-incorporation was most pronounced within 48 h after denervation (19±6% of the control side). Therefore, the reduction in bromodeoxyuridine-labeling followed a similar temporal course as the thinning of the epidermis (25–50%). Both epidermal thinning and reduced bromodeoxyuridine-labeling were reversed by epidermal reinnervation three months after denervation. Patterns of keratinocyte differentiation and programmed cell death were unaffected by skin denervation. These findings are consistent with the notion that skin innervation exerts influence on the proliferation of keratinocytes and the thickness of the epidermis, and offers a new look at the interaction between nociceptive nerves and their innervated targets.

Superficial angiomyxoma: clinicopathologic analysis of a series of distinctive but poorly recognized cutaneous tumors with tendency for recurrence.

Despite being first described in 1988, superficial angiomyxoma is still a poorly recognized cutaneous tumor. Although its histologic features are distinctive, its existence seems not to be widely accepted. We analyzed the clinicopathologic and immunohistochemical features in a series of 39 cases. Twenty-five patients were males; age range was birth to 82 years (median, 45.5 years). Most cases presented as cutaneous papules, nodules, or polypoid lesions. Seventeen tumors arose on the trunk, 14 on the head and neck, and seven on the lower limbs. All cases were treated by local excision, and eight recurred locally. In four of the latter cases, there were two recurrences. Histologically, the lesions were dermal with variable involvement of the subcutis. Tumors were poorly circumscribed, but a focal lobular outline was always identified. Distinctive histologic features included extensive myxoid stroma, numerous small blood vessels, varying cellularity, acellular mucin pools, stellate or bipolar fibroblastic cells, muciphages, a sparse, mixed inflammatory cell infiltrate with notable neutrophils, and occasional plumper cells with eosinophilic cytoplasm. Cytologic atypia was mild at most, and mitotic figures were rare. In approximately 20% of cases, the primary lesion or its recurrence contained epithelial structures, including epidermoid cysts, thin strands of squamous epithelium, and small buds of basaloid cells. Immunohistochemically, tumor cells were negative for S-100 protein, smooth muscle actin, and pan-keratin. We support the concept of superficial angiomyxoma as a distinctive clinicopathologic entity that should be included in the differential diagnosis of other myxoid cutaneous tumors, including dermal nerve sheath myxoma, trichodiscoma and trichofolliculoma, and low-grade myxofibrosarcoma.

Identification of pituitary adenylate cyclase activating polypeptide (PACAP) and PACAP type 1 receptor in human skin: expression of PACAP-38 is increased in patients with psoriasis

Morphological and biochemical evidence is presented for the presence of pituitary adenylate cyclase activating peptide (PACAP) and the high-affinity PACAP-1 receptor subtype in human skin. Immunohistochemical analysis revealed PACAP-immunoreactivity (IR) to be present predominantly in dermal nerve fibers close to the dermal–epidermal border, hair follicles, blood vessels and sweat glands. Radioimmunoassay, chromatographic analysis and Western blotting revealed this PACAP-IR to be PACAP-38 whereas the second molecular form, PACAP-27, is absent. In tissue of psoriasis patients significantly more PACAP-38 protein was detected as compared to normal skin. Using RT-PCR, the expression of a high-affinity PACAP-1 receptor in human skin was observed. These results indicate a possible role for PACAP-38 in inflammatory processes of psoriasis.