Svein Ivar Mellgren

European federation of neurological societies/peripheral nerve society guideline on the use of skin biopsy in the diagnosis of small fiber neuropathy. report of a joint task force of the european fe-deration of neurological societies and the peripheral nerve society

Background:  Revision of the guidelines on the use of skin biopsy in the diagnosis of peripheral neuropathy, published in 2005, has become appropriate owing to publication of more relevant articles. Most of the new studies focused on small fiber neuropathy (SFN), a subtype of neuropathy for which the diagnosis was first developed through skin biopsy examination. This revision focuses on the use of this technique to diagnose SFN.

EFNS guidelines on the use of skin biopsy in the diagnosis of peripheral neuropathy

Skin biopsy has become a widely used tool to investigate small calibre sensory nerves including somatic unmyelinated intraepidermal nerve fibres (IENF), dermal myelinated nerve fibres, and autonomic nerve fibres in peripheral neuropathies and other conditions. Different techniques for tissue processing and nerve fibre evaluation have been used. In March 2004, a Task Force was set up under the auspices of the European Federation of Neurological Societies (EFNS) with the aim of developing guidelines on the use of skin biopsy in the diagnosis of peripheral neuropathies. We searched the Medline database from 1989, the year of the first publication describing the innervation of human skin using immunostaining with anti‐protein‐gene‐product 9.5 (PGP 9.5) antibodies, to 31 March 2005. All pertinent papers were rated according to the EFNS guidance. The final version of the guidelines was elaborated after consensus amongst members of the Task Force was reached. For diagnostic purposes in peripheral neuropathies, we recommend performing a 3‐mm punch skin biopsy at the distal leg and quantifying the linear density of IENF in at least three 50‐μm thick sections per biopsy, fixed in 2% PLP or Zambonis solution, by bright‐field immunohistochemistry or immunofluorescence with anti‐PGP 9.5 antibodies (level A recommendation). Quantification of IENF density closely correlated with warm and heat‐pain threshold, and appeared more sensitive than sensory nerve conduction study and sural nerve biopsy in diagnosing small‐fibre sensory neuropathy. Diagnostic efficiency and predictive values of this technique were very high (level A recommendation). Confocal microscopy may be particularly useful to investigate myelinated nerve fibres, dermal receptors and dermal annex innervation. In future, the diagnostic yield of dermal myelinated nerve fibre quantification and of sweat gland innervation should be addressed. Longitudinal studies of IENF density and regeneration rate are warranted to correlate neuropathological changes with progression of neuropathy and to assess the potential usefulness of skin biopsy as an outcome measure in peripheral neuropathy trials (level B recommendation). In conclusion, punch skin biopsy is a safe and reliable technique (level A recommendation). Training in an established cutaneous nerve laboratory is recommended before using skin biopsy as a diagnostic tool in peripheral neuropathies. Quality control at all levels is mandatory.

Intraepidermal nerve fiber density at the distal leg: a worldwide normative reference study

The diagnostic reliability of skin biopsy in small fiber neuropathy depends on the availability of normative reference values. We performed a multicenter study to assess the normative values of intraepidermal nerve fiber (IENF) density at distal leg stratified by age deciles. Eight skin biopsy laboratories from Europe, USA, and Asia submitted eligible data. Inclusion criteria of raw data were healthy subjects 18 years or older; known age and gender; 3‐mm skin biopsy performed 10‐cm above the lateral malleolus; bright‐field immunohistochemistry protocol, and quantification of linear IENF density in three 50‐µm sections according to published guidelines. Data on height and weight were recorded, and body mass index (BMI) was calculated in subjects with both available data. Normative IENF density reference values were calculated through quantile regression analysis; influence of height, weight, or BMI was determined by regression analyses. IENF densities from 550 participants (285 women, 265 men) were pooled. We found a significant age‐dependent decrease of IENF density in both genders (women p < 0.001; men p = 0.002). Height, weight, or BMI did not influence the calculated 5th percentile IENF normative densities in both genders. Our study provides IENF density normative reference values at the distal leg to be used in clinical practice.

Peripheral neuropathy in primary Sjögren's syndrome

Sjögrens syndrome (dryness of eyes, mouth, and other mucous membranes) may be associated with disease of joints, blood, internal organs, skin, and central and peripheral nervous systems. We reviewed 33 cases of primary Sjögrens syndrome and peripheral neuropathy evaluated by neurologic examinations and EMG at the Mayo Clinic from 1976 to 1988, and studied sural nerve biopsy specimens in 11 of them. Symmetric sensorimotor polyneuropathy occurred most frequently, followed by symmetric sensory neuropathy. Autonomic neuropathy, mononeuropathy, or cranial neuropathy (especially trigeminal neuropathy) was superimposed on generalized neuropathy in approximately one-fourth of patients. The course generally was slowly progressive, except for a few patients who may have improved with prednisone therapy. Although spinal ganglion involvement might have accounted for some of the clinical and neurophysiologic findings, we found evidence that necrotizing vasculitis was involved in fiber degeneration. All nerve biopsies revealed perivascular inflammatory infiltrates and other vessel abnormalities, which were diagnostic in two cases and strongly suggestive of necrotizing vasculitis in six cases. Axonal degeneration predominated over demyelination and sometimes was focal or multifocal. In neuropathy of unknown cause, particularly if it is sensory, autonomic, or involves trigeminal nerve, consider Sjögrens syndrome.

Outdoor activities and diet in childhood and adolescence relate to MS risk above the Arctic Circle

BackgroundA relationship between the latituderelated distribution of multiple sclerosis (MS) and exposure to sunlight has long been considered. Higher sun exposure during early life has been associated with decreased risk of MS.ObjectiveSince Norway is an exception to the latitude gradient of MS prevalence, we tested here whether sunlight exposure or vitamin D-related dietary factors in childhood and adolescence are associated with the risk of MS.MethodsRetrospective recall questionnaire data from 152 MS patients and 402 population controls born at and living at latitudes 66–71°N were analysed by means of conditional logistic regression analysis accounting for the matching variables age, sex, and place of birth.ResultsIncreased outdoor activities during summer in early life were associated with a decreased risk of MS, most pronounced at ages 16–20 years (odds ratio (OR) 0.55, 95% CI 0.39–0.78, p = 0.001, adjusted for intake of fish and cod-liver oil). A protective effect of supplementation with cod-liver oil was suggested in the subgroup that reported low summer outdoor activities (OR 0.57, 95% CI 0.31–1.05, p = 0.072). Consumption of fish three or more times a week was also associated with reduced risk of MS (OR 0.55, 95% CI 0.33–0.93, p = 0.024).ConclusionSummer outdoor activities in childhood and adolescence are associated with a reduced risk of MS even north of the Arctic Circle. Supplemental cod-liver oil may be protective when sun exposure is less, suggesting that both climate and diet may interact to influence MS risk at a population level.

The Inflammatory Properties of Contained and Noncontained Lumbar Disc Herniation

Study Design. The inflammatory properties of nucleus pulposus were assessed in biopsy samples from patients who underwent surgery for lumbar disc herniation. Objectives. To investigate the inflammatory properties of the different types of disc herniation. Background Data. High levels of phospholipase A2 previously have been demonstrated in a small number of patients undergoing lumbar disc surgery. Phospholipase A2 is the enzyme responsible for the liberation of arachidonic acid from cell membranes at the site of inflammation and is considered to be the limiting agent in the production of prostaglandins and leukotrienes, which are powerful mediators of inflammation. Cytokines are among the many agonists inducing phospholipase A2 activation. Several reports previously have demonstrated the difference in clinical appearance of different types of lumbar disc herniation. Methods. Thirty‐seven patients undergoing surgery for lumbar disc herniation were investigated. During surgery the disc pathology of each patient was classified into one of three groups: bulging disc, contained herniation, and noncontained disc herniation. Also during surgery, biopsy samples were taken from the nucleus, immediately frozen in liquid nitrogen, and subsequently stored at −70 C until analyzed. Results. No traces of interleukin‐6 or tumor necrosis factor alpha were found in the biopsy samples. There was a significant difference in the levels of leukotriene B4 and thromboxane B2 in contained versus noncontained disc herniation, and the highest concentration was found in the noncontained disc herniation group. Conclusion. The results support the theory that inflammatory mechanisms are involved in sciatica because of lumbar disc herniation and indicate that the different types of disc herniation have different inflammatory properties.

Neuropsychiatric disturbances in SLE are associated with antibodies against NMDA receptors

To determine whether neuropsychiatric manifestations in patients with systemic lupus erythematosus (SLE) are influenced by antibodies against the human N‐methyl‐d‐aspartate (NMDA) receptor types NR2a or NR2b. A decapeptide was synthesized containing a sequence motif present in the extracellular ligand‐binding domain of NMDA receptors NR2a and NR2b, bound by the monoclonal murine anti‐DNA antibody R4A. In an ELISA with the murine monoclonal R4v as positive control, plasma samples of 57 patients with SLE were examined for the anti‐peptide (anti‐NR2) antibody after the patients had been subjected to comprehensive psychological and cognitive testing. Poor performance on the Visual Paired Associates test (immediate), the Grooved Pegboard test, as well as high scores on the Beck Depression Inventory, and scales D‐2 (depression), Pd‐4 (psychopathic deviate), Sc‐8 (schizophrenia), and Ma‐9 (hypomania) of the MMPI‐2 were significantly associated with elevated levels of anti‐NR2 antibodies. The findings in several domains indicate an association between anti‐NR2 antibodies and depressed mood in addition to decreased short‐time memory and learning. Antibodies to NMDA receptors thus may represent one of several mechanisms for cerebral dysfunction in patients with SLE.

The effect of age and gender on epidermal nerve fiber density

Objective: Sensory neuropathies often involve small-diameter myelinated and unmyelinated nerve fibers, and neurologic and electrophysiologic findings may be normal unless larger nerve fibers are involved. The small (intra)epidermal nerve fibers (ENFs) now can be visualized with immunohistochemical techniques using the panaxonal marker anti-protein gene product 9.5 (PGP 9.5). Using this technique, the authors have established a reference range for ENF in a healthy white population and evaluated the reliability of the method. Methods: Two punch biopsies, 3 mm in diameter, were taken from the distal part of the leg in 106 healthy volunteers (mean age, 49.0 ± 19.6 years). Fifty-micrometer frozen thick sections were incubated with rabbit polyclonal antibodies to human PGP 9.5. The number of ENF/mm then was reported as the mean of counts in six sections (three sections from each of the two biopsies). Results: The mean number of ENFs was 12.4 ± 4.6 mm. In a multiple regression model, the density of ENF depended on age and gender (Y = 13.92 + 2.25 (gender) − 0.06 × age). The mean difference in ENF by intraobserver analysis was 0.2 ± 1.2 ENF/mm, and by interobserver analysis, it was 0.4 ± 1.5 fibers/mm. Conclusion: Normal means and ranges for the density of epidermal nerve fibers in a reference population have been established. The density of epidermal nerve fibers decreases with age and is lower in men compared with women. Intraobserver and interobserver analysis proves the reliability of the method.

Early diabetic neuropathy: thermal thresholds and intraepidermal nerve fibre density in patients with normal nerve conduction studies

ObjectivesTo determine whether neuropathy in diabetic patients with normal nerve conduction studies could be detected by measurements of thermal thresholds and quantification of intraepidermal nerve fibre (IENF) density, and to evaluate differences in parameters between patients with and without neuropathic symptoms.MethodsA total of 22 patients with and 37 patients without sensory symptoms suggesting distal neuropathy were included. Measurements of warm and cold perception thresholds and skin biopsy for quantification of IENFs were performed distally on the leg. Reference data were used to normalize test results for age and height or gender of individual patients by calculating the Z-scores.ResultsIENF density was significantly reduced in both symptomatic and asymptomatic patients compared to controls (p < 0.001), and in patients with symptoms compared to those without (p = 0.01). Thermal thresholds were significantly elevated (more abnormal) in patients with symptoms compared to controls (p < 0.01), but only for cold perception threshold (CPT) (p < 0.001) in the asymptomatic group. When comparing symptomatic and asymptomatic patients, there was no statistically significant difference in thermal thresholds. Depletion of IENFs in skin biopsy was the most frequent abnormal finding in the subgroup of patients with neuropathic symptoms (36 %) followed by abnormal CPT (27 %).ConclusionPatients with diabetes and normal nerve conduction studies had significantly lower IENF density and higher CPT than controls, whether they had symptoms of polyneuropathy or not. In patients with neuropathic symptoms, abnormal IENF density predominated and seemed thus to be the most sensitive tool of detecting small diameter nerve fibre involvement.

Effect of vitamin D3 supplementation on relapses, disease progression, and measures of function in persons with multiple sclerosis: exploratory outcomes from a double-blind randomised controlled trial

Background: High vitamin D levels may reduce the risk of relapses and disease progression in multiple sclerosis. Methods: This 96-week randomised controlled trial was designed to assess the effect of vitamin D3 supplementation on bone mineral density in persons with multiple sclerosis. Supplementation with 20,000 IU vitamin D3 weekly raised median serum 25-hydroxy vitamin D (25[OH]D) to 121 nmol/L. The modified intention to treat analysis included 35 persons in the vitamin D3 group and 33 in the placebo group. Participants were age 21 to 50 years and fully ambulatory (median Expanded Disability Status Scale (EDSS) 2.5). We studied the effect of supplementing vitamin D3 on the exploratory outcomes annualised relapse rate (ARR), EDSS, multiple sclerosis functional composite (MSFC) components, grip strength, and fatigue. Results: After 96 weeks, there was no significant difference between groups in ARR (absolute difference 0.10, 95% CI -0.07 to 0.27; p = 0.25), EDSS (absolute difference -0.01, 95% CI -0.35 to 0.35; p = 0.97), MSFC components, grip strength, or fatigue. Conclusion: Supplementation with 20,000 IU vitamin D3 weekly did not result in beneficial effects on the measured multiple sclerosis-related outcomes. This study was not powered to address clinical outcomes, but none of the results were suggestive of an effect in this unselected population of fully ambulatory persons with multiple sclerosis.