Studying serum neurofilament light chain levels as a potential new biomarker for small fiber neuropathy.

BACKGROUND AND PURPOSE: Diagnosing small fiber neuropathies can be challenging. To address this issue, whether serum neurofilament light chain (sNfL) could serve as a potential biomarker of damage to epidermal Aδ- and C-fibers was tested. METHODS: Serum NfL levels were assessed in 30 patients diagnosed with small fiber neuropathy and were compared to a control group of 19 healthy individuals. Electrophysiological studies, quantitative sensory testing and quantification of intraepidermal nerve fiber density after skin biopsy were performed in both the proximal and distal leg. RESULTS: Serum NfL levels were not increased in patients with small fiber neuropathy compared to healthy controls (9.1 ± 3.9 and 9.4 ± 3.8, p = 0.83) and did not correlate with intraepidermal nerve fiber density at the lateral calf or lateral thigh or with other parameters of small fiber impairment. CONCLUSION: Serum NfL levels cannot serve as a biomarker for small fiber damage.

Cutaneous biomarkers of therapeutic efficacy in early treatment of hereditary ATTR amyloid polyneuropathy with tafamidis.

BACKGROUND: ATTR (ATTRv) amyloidosis neuropathy is characterized by progressive sensorimotor and autonomic nerve degeneration secondary to amyloid deposition caused by a misfolded transthyretin protein (TTR). Small nerve fiber neuropathy is an early clinical manifestation of this disease resulting from the dysfunction of the Aδ and C small nerve fibers. Tafamidis, a selective TTR stabilizer, has proven its efficacy in the earlier stages of hATTR. OBJECTIVES: To evaluate the clinical course and utility of cutaneous pathological biomarkers in patients with ATTR amyloidosis treated with tafamidis compared to control patients. METHODS: Forty patients diagnosed with early stages of ATTRv amyloidosis (polyneuropathy disability [PND] scores 0-II) underwent small and large nerve fiber neurological evaluations, and annual skin biopsies for intraepidermal nerve fiber density (IENFD) and amyloid deposition index (ADI) estimation. Thirty patients were allocated to receive tafamidis, and 10 patients served as controls. Tafamidis pharmacokinetics analysis was performed in patients who received the treatment. RESULTS: At baseline, 12% of patients in stage PND 0 and 28% in PND I displayed small nerve fiber denervation in the distal thigh, whereas 23% and 38%, respectively, in the distal leg. Similarly, 72% and 84% had amyloid deposition in the distal thigh and 56% and 69% in the distal leg. Following 1 year of treatment, the tafamidis group showed significant clinical improvement compared to the control group, revealed by the following mean differences (1) -9.3 versus -4 points (p = <.00) in the patient's neuropathy total symptom score 6 (NTSS-6) questionnaire, (2) -2.5 versus +2.8 points (p = <.00) in the Utah Early Neuropathy Score (UENS), and (3) +1.2°C versus -0.6 (p = .01) in cold detection thresholds. Among the patients who received tafamidis, 65% had stable or increased IENFD in their distal thigh and 27% in the distal leg. In contrast, all patients in the control group underwent denervation. The ADI either decreased or remained constant in 31% of the biopsies in the distal thigh and in 24% of the biopsies in the distal leg of the tafamidis-treated patients, whereas it rose across all the biopsies in the control group. At the 4-year follow-up, the tafamidis group continued to display less denervation in the distal thigh (mean difference [MD] of -3.0 vs. -9.3 fibers/mm) and the distal leg (mean difference [MD] -4.9 vs. -8.6 fibers/mm). ADI in tafamidis-treated patients was also lower in the distal thigh (10 vs. 30 amyloid/mm2) and the distal leg (23 vs. 40 amyloid/mm2) compared to control patients. Plasma tafamidis concentrations were higher in patients with IENFD improvement and in patients with reduced amyloid deposition. Patients without amyloid deposition in the distal leg at baseline displayed delayed disease progression at 4 years. CONCLUSIONS: Cutaneous IENFD and amyloid deposition assessments in the skin of the distal thigh and distal leg are valuable biomarkers for early diagnosis of ATTR amyloidosis and for measuring the progression of small nerve fiber neuropathy. Early treatment with tafamidis slows the clinical progression of the disease, skin denervation, and amyloid deposition in the skin. Higher plasma concentrations of tafamidis are associated with better disease outcomes, suggesting that increasing the drug dose could achieve better plasma concentrations and response rates. This study describes the longest small nerve fiber neuropathy therapeutic trial with tafamidis and is the first to report small fiber symptoms, function, and structural assessments as outcomes.

Expression in skin biopsies supports genetic evidence linking CAMKK2, P2X7R and P2X4R with HIV-associated sensory neuropathy.

HIV-associated sensory neuropathy (HIV-SN) affects 14-38% of HIV+ individuals stable on therapy with no neurotoxic drugs. Polymorphisms in CAMKK2, P2X7R and P2X4R associated with altered risk of HIV-SN in Indonesian and South African patients. The role of CaMKK2 in neuronal repair makes this an attractive candidate, but a direct role for any protein is predicated on expression in affected tissues. Here, we describe expression of CaMKK2, P2X7R and P2X4R proteins in skin biopsies from the lower legs of HIV+ Indonesians with and without HIV-SN, and healthy controls (HC). HIV-SN was diagnosed using the Brief Peripheral Neuropathy Screen. Biopsies were stained to detect protein gene product 9.5 on nerve fibres and CaMKK2, P2X7R or P2X4R, and were examined using 3-colour sequential scanning confocal microscopy. Intraepidermal nerve fibre densities (IENFD) were lower in HIV+ donors than HC and correlated directly with nadir CD4 T-cell counts (r = 0.69, p = 0.004). However, IENFD counts were similar in HIV-SN+ and HIV-SN- donors (p = 0.19) and so did not define neuropathy. CaMKK2+ cells were located close to dermal and epidermal nerve fibres and were rare in HC and HIV-SN- donors, consistent with a role for the protein in nerve damage and/or repair. P2X7R was expressed by cells in blood vessels of HIV-SN- donors, but rarely in HC or HIV-SN+ donors. P2X4R expression by cells in the epidermal basal layer appeared greatest in HIV-SN+ donors. Overall, the differential expression of CaMKK2, P2X7R and P2X4R supports the genetic evidence of a role for these proteins in HIV-SN.

Dorsal Root Ganglion Stimulation in Chronic Painful Polyneuropathy: A Potential Modulator for Small Nerve Fiber Regeneration.

OBJECTIVES: Neuromodulatory treatments like spinal cord stimulation and dorsal root ganglion stimulation (DRGS) have emerged as effective treatments to relieve pain in painful polyneuropathy. Animal studies have demonstrated that neurostimulation can enhance nerve regeneration. This study aimed to investigate if DRGS may impact intraepidermal nerve fiber regeneration and sensory nerve function. MATERIALS AND METHODS: Nine patients with chronic, intractable painful polyneuropathy were recruited. Intraepidermal nerve fiber density (IENFD) quantification in 3 mm punch skin biopsy was performed 1 month before DRGS (placed at the level of the L5 and S1 dorsal root ganglion) and after 12- and 24-month follow-up. Quantitative sensory testing, nerve conduction studies, and a clinical scale score were also performed at the same time points. RESULTS: In 7 of 9 patients, DRGS was successful (defined as a reduction of ≥ 50% in daytime and/or night-time pain intensity), allowing a definitive implantable pulse generator implantation. The median baseline IENFD among these 7 patients was 1.6 fibers/mm (first and third quartile: 1.2; 4.3) and increased to 2.6 fibers/mm (2.5; 2.9) and 1.9 fibers/mm (1.6; 2.4) at 1- and 2-years follow-up, respectively. These changes were not statistically significant (p = 1.000 and 0.375). Sensory nerve tests did not show substantial changes. CONCLUSIONS: Although not significant, the results of this study showed that in most of the patients with implants, there was a slight increase of the IENFD at the 1- and 2-year follow-up. Larger-scale clinical trials are warranted to explore the possible role of DRGS in reversing the progressive neurodegeneration over time. CLINICAL TRIAL REGISTRATION: The Clinicaltrials.gov registration number for the study is NCT02435004; Swiss National Clinical Trials Portal: SNCTP000001376.

Temporal trend of small nerve fibre degeneration in people with and without type 2 diabetes mellitus.

AIMS: We investigated the long-term temporal trend of intraepidermal nerve fibre density (IENFD) and the association between changes in IENFD and metabolic factors in individuals with and without type 2 diabetes. METHODS: A total of 66 participants were enrolled in this longitudinal population-based study, at baseline consisting of 35 individuals (median 61 years) without diabetes and 31 individuals with type 2 diabetes mellitus. Participants underwent clinical and electrophysiological examinations, as well as a skin biopsy both at baseline and at the follow-up visit (mean 8.1 ± 0.5 years). IENFD was assessed in thin sections of 5 µm, stained with the protein gene product 9.5-antibody and compared between the groups. RESULTS: IENFD decreased during the period in both groups, with a greater decline in the group without diabetes than in type 2 diabetes (-2.3 and -0.6 fibres/mm respectively; p < 0.001). While IENFD at baseline was significantly reduced in type 2 diabetes relative to people without (p < 0.001), no difference in IENFD was found between groups at the follow-up (p = 0.183). Linear mixed model analysis indicated that age, weight and HbA1c were associated with decrease in IENFD in the total population (p < 0.007). IENFD also decreased with increasing age and weight, but not with HbA1c , in the separate groups (p < 0.049). CONCLUSIONS: Despite lower IENFD levels at baseline in type 2 diabetes, IENFD was equal between the groups at follow-up. A decrease in IENFD is to a limited extent affected by body weight, and HbA1c , but age seems to be the long-term determinant of IENFD in an elderly population.

Subepidermal Schwann cell counts correlate with skin innervation - an exploratory study.

INTRODUCTION/AIMS: Schwann cell clusters have been described at the murine dermis-epidermis border. We quantified dermal Schwann cells in the skin of patients with small-fiber neuropathy (SFN) compared with healthy controls to correlate with the clinical phenotype. METHODS: Skin punch biopsies from the lower legs of 28 patients with SFN (11 men, 17 women; median age, 54 [range, 19-73] years) and 9 healthy controls (five men, four women, median age, 34 [range, 25-69] years) were immunoreacted for S100 calcium-binding protein B as a Schwann cell marker, protein-gene product 9.5 as a pan-neuronal marker, and CD207 as a Langerhans cell marker. Intraepidermal nerve fiber density (IENFD) and subepidermal Schwann cell counts were determined. RESULTS: Skin samples of patients with SFN showed lower IENFD (P < .05), fewer Schwann cells per millimeter (P < .01), and fewer Schwann cell clusters per millimeter (P < .05) than controls. When comparing SFN patients with reduced (n = 13; median age, 53 [range, 19-73] years) and normal distal (n = 15, median age, 54 [range, 43-68] years) IENFD, the number of solitary Schwann cells per millimeter (p < .01) and subepidermal nerve fibers associated with Schwann cell branches (P < .05) were lower in patients with reduced IENFD. All three parameters correlated positively with distal IENFD (P < .05 to P < .01), whereas no correlation was found between Schwann cell counts and clinical pain characteristics. DISCUSSION: Our data raise questions about the mechanisms underlying the interdependence of dermal Schwann cells and skin innervation in SFN. The temporal course and functional impact of Schwann cell presence and kinetics need further investigation.

Progression and regression of nerve fibre pathology and dysfunction early in diabetes over 5 years.

It has traditionally been suggested that the early development of diabetic sensorimotor polyneuropathy (DSPN) is characterized by predominant and progressive injury to small nerve fibres followed by large fibre impairment. We alternatively hypothesized that small and large fibre damage due to DSPN in type 1 and type 2 diabetes could develop in parallel and may not only be progressive but also reversible. Participants from the German Diabetes Study baseline cohort with recent-onset type 1/type 2 diabetes (n = 350/570) and age-matched glucose-tolerant control individuals (Control 1/Control 2: n = 114/190) were assessed using nerve conduction studies, thermal detection thresholds, vibration perception thresholds, neuropathy symptom scores, neuropathy disability scores and intraepidermal nerve fibre density (IENFD) in skin biopsies (type 1/type 2 diabetes: n = 102/226; Control 1/Control 2: n = 109/208). Subsets of participants with type 1/type 2 diabetes were followed for 5 years (n = 184/307; IENFD subset: n = 18/69). DSPN was defined by the Toronto Consensus criteria. At baseline, DSPN was present in 8.1% and 13.3% of the type 1 and type 2 diabetes groups, respectively. The most frequently abnormal tests in the lower limbs below or above the 2.5th and 97.5th centiles of the controls were the IENFD (13.7%) and individual nerve conduction studies (up to 9.4%) in type 1 diabetes participants and IENFD (21.8%), malleolar vibration perception thresholds (17.5%), and individual nerve conduction studies (up to 11.8%) in those with type 2 diabetes, whereas thermal detection threshold abnormalities did not differ between the control and diabetes groups. After 5 years, the highest progression rates from the normal to the abnormal range in type 2 diabetes participants were found for IENFD (18.8%) by -4.1 ± 2.8 fibres/mm, malleolar vibration perception threshold (18.6%) by 9.1 ± 20.2 µm and nerve conduction studies (15.0%) by 3.7 ± 1.5 points, while vice versa the highest regression rates were observed for neuropathy disability scores (11.2%) by -3.1 ± 1.3 points, sural nerve amplitudes (9.1%) by 4.7 ± 3.0 µV, IENFD (8.7%) by 1.4 ± 1.3 fibres/mm, and neuropathy symptom scores (8.2%) by -5.8 ± 1.6 points. In type 1 diabetes participants, no major progression was seen after 5 years, but subclinical DSPN regressed in 10.3%. These findings point to early parallel damage to both small and large nerve fibres in well-controlled recent-onset type 2 and, to a lesser extent, type 1 diabetes. After 5 years, peripheral nerve morphology and function and clinical measures progress to the abnormal range in type 2 diabetes, but initial nerve alterations are also reversible to a meaningful degree.

The analysis of epidermal nerve fibre spatial distribution improves the diagnostic yield of skin biopsy.

AIM: Small fibre neuropathy (SFN) diagnosis represents a challenge for neurologists. The diagnostic gold standard is intraepidermal nerve fibre (IENF) density, but in about 10-20% of patients with symptoms/signs and abnormalities on functional tests, it remains within normal range. We propose an adjunctive parameter to improve the efficiency of skin biopsy diagnosis. METHODS: We recruited 31 patients with SFN symptoms/signs, normal nerve conduction study, abnormal quantitative sensory testing and normal IENF density. We also included 31 healthy controls and 31 SFN patients with reduced IENF density as control groups. RESULTS: We measured the distance between consecutive IENFs in the three groups. Mean inter-fibre distances did not differ between patients with normal counts and healthy controls (66.7 ± 14.5 µm vs. 76.7 ± 13.4 µm; P = 0.052), while the relative standard deviation was significantly (P < 0.001) higher in patients (79.3 ± 29.9) compared to controls (51.6 ± 12.2). Using ROC analysis, we identified an inter-fibre distance of 350 µm as the measure that better differentiated patients from controls (AUC = 0.85, sensitivity: 74%, specificity: 94%). At least one such segment was also observed in all patients with reduced IENF count. CONCLUSION: Irregular spatial distribution is an SFN intrinsic feature preceding actual nerve loss. The presence of a stretch of denervated epidermis longer than 350 µm is a parameter able to increase the diagnostic efficiency of skin biopsy.

Early changes of nerve integrity in preclinical carriers of hereditary transthyretin Ala117Ser amyloidosis with polyneuropathy.

BACKGROUND AND PURPOSE: Disease-modifying therapies provide new horizons for hereditary transthyretin amyloidosis with polyneuropathy (ATTRv-PN) to slow neuropathic progression. Initiating treatment at the earliest time requires biomarkers reflecting both small- and large-fiber degeneration in carriers. METHODS: This study included examinations of pathology (intraepidermal nerve fiber [IENF] density), physiology (nerve conduction studies, autonomic function test, and nerve excitability), and psychophysics (thermal thresholds) in carriers to compare to healthy controls and asymptomatic diabetic patients. RESULTS: There were 43 carriers (44.2 ± 11.4 years, p.Ala117Ser in 42 carriers), 43 controls (43.4 ± 12.7 years) including 26 noncarrier families, and 50 asymptomatic diabetic patients (58.1 ± 9.5 years). Carriers had lower IENF densities than controls and similar densities as diabetic patients. Median nerve conduction parameters, especially distal motor latency, were the most frequent neurophysiological abnormality in carriers, could differentiate carriers from controls and diabetic patients, were correlated with IENF densities in carriers but not in controls and diabetic patients, and were correlated with nerve excitability parameters in carriers but not in controls. Fifteen carriers (34.9%) with electrophysiological evidence of median nerve entrapment at the wrist had lower IENF densities and more abnormal conduction parameters than carriers without. We defined nerve dysfunction index-the ratio of median distal motor latency to IENF density-which differentiated carriers from controls. CONCLUSIONS: In late-onset ATTRv-PN carriers with predominant p.Ala117Ser, median conduction parameters were the most common neurophysiological abnormalities and served as surrogate signatures of small- and large-fiber impairment. Combination of median distal motor latency and IENF density can reflect early neuropathy in carriers.

Early changes in tests of peripheral nerve function during oxaliplatin treatment and their correlation with chemotherapy-induced polyneuropathy symptoms and signs.

BACKGROUND AND PURPOSE: Assessment of the severity of chronic peripheral neuropathy during oxaliplatin treatment is based on symptoms. Efforts to adjust the total dose of oxaliplatin to prevent severe neuropathy can be complicated by the worsening of neuropathy symptoms following treatment. Objective measures of the structure and function of peripheral nerves during early phases of treatment may aid in determining the optimal oxaliplatin dose in individual patients. Intraepidermal nerve fibre density (IENFD) has been suggested as an early marker of peripheral neuropathy. METHODS: Sixty patients were examined before treatment and following 25% and 50% of the total planned oxaliplatin dose. Fifty-five of them were also examined at completion of chemotherapy and 6 months later. IENFD in skin biopsies from the distal leg, nerve conduction studies and quantitative sensory testing at the dorsum of the foot were performed. Forty-six healthy subjects were examined at baseline and after 6 and 52 weeks for comparison. RESULTS: Intraepidermal nerve fibre density was not reduced during treatment. Sural nerve amplitude and conduction velocity, vibration detection thresholds, mechanical detection threshold and cold detection threshold were significantly reduced during treatment. Compared to reference values and spontaneous changes in healthy subjects, the largest proportions of patients with deterioration were found for vibration detection thresholds followed by nerve conduction studies, mechanical detection threshold, cold detection threshold and IENFD. CONCLUSIONS: Significant changes were most pronounced for measures of large nerve fibre function, especially vibration sensation. Skin biopsies do not seem to provide a clinically relevant objective measure of peripheral nerve deterioration during oxaliplatin treatment.

Intraepidermal Nerve Fiber Analysis in Human Patients and Animal Models of Peripheral Neuropathy: A Comparative Review.

Analysis of intraepidermal nerve fibers (IENFs) in skin biopsy samples has become a standard clinical tool for diagnosing peripheral neuropathies in human patients. Compared to sural nerve biopsy, skin biopsy is safer, less invasive, and can be performed repeatedly to facilitate longitudinal assessment. Intraepidermal nerve fiber analysis is also more sensitive than conventional nerve histology or electrophysiological tests for detecting damage to small-diameter sensory nerve fibers. The techniques used for IENF analysis in humans have been adapted for large and small animal models and successfully used in studies of diabetic neuropathy, chemotherapy-induced peripheral neuropathy, HIV-associated sensory neuropathy, among others. Although IENF analysis has yet to become a routine end point in nonclinical safety testing, it has the potential to serve as a highly relevant indicator of sensory nerve fiber status in neurotoxicity studies, as well as development of neuroprotective and neuroregenerative therapies. Recently, there is also interest in the evaluation of IENF via skin biopsy as a biomarker of small fiber neuropathy in the regulatory setting. This article provides an overview of the anatomic and pathophysiologic principles behind IENF analysis, its use as a diagnostic tool in humans, and applications in animal models with focus on comparative methodology and considerations for study design.

Evaluation of small nerve fiber dysfunction in type 2 diabetes.

OBJECTIVES: To assess potential correlations between intraepidermal nerve fiber densities (IENFD), graded with light microscopy, and clinical measures of peripheral neuropathy in elderly male subjects with normal glucose tolerance (NGT), impaired glucose tolerance (IGT), and type 2 diabetes (T2DM), respectively. MATERIALS AND METHODS: IENFD was assessed in thin sections of skin biopsies from distal leg in 86 men (71-77 years); 24 NGT, 15 IGT, and 47 T2DM. Biopsies were immunohistochemically stained for protein gene product (PGP) 9.5, and intraepidermal nerve fibers (IENF) were quantified manually by light microscopy. IENFD was compared between groups with different glucose tolerance and related to neurophysiological tests, including nerve conduction study (NCS; sural and peroneal nerve), quantitative sensory testing (QST), and clinical examination (Total Neuropathy Score; Neuropathy Symptom Score and Neuropathy Disability Score). RESULTS: Absent IENF was seen in subjects with T2DM (n = 10; 21%) and IGT (n = 1; 7%) but not in NGT. IENFD correlated weakly negatively with HbA1c (r = -.268, P = .013) and Total Neuropathy Score (r = -.219, P = .042). Positive correlations were found between IENFD and sural nerve amplitude (r = .371, P = .001) as well as conduction velocity of both the sural (r = .241, P = .029) and peroneal nerve (r = .258, P = .018). Proportions of abnormal sural nerve amplitude became significantly higher with decreasing IENFD. No correlation was found with QST. Inter-rater reliability of IENFD assessment was good (ICC = 0.887). CONCLUSIONS: Signs of neuropathy are becoming more prevalent with decreasing IENFD. IENFD can be meaningfully evaluated in thin histopathological sections using the presented technique to detect neuropathy.

Evidence of small-fiber neuropathy in neurofibromatosis type 1.

INTRODUCTION: Large-fiber neuropathy is rare in neurofibromatosis type 1, but small-fiber neuropathy has not been studied. METHODS: Patients with neurofibromatosis type 1 underwent nerve conduction studies for large-fiber assessment. Small-fiber tests included quantitative thermal thresholds, laser Doppler flare imaging, intraepidermal nerve fiber density, and corneal nerve fiber length. RESULTS: Of the 52 patients enrolled, 31 (60%) were female and the mean age was 33.0 ± 12.3 years. Four (8%) patients had abnormal nerve conduction studies. Small-fiber tests were frequently abnormal: thermal thresholds in 7 (13%); laser Doppler flare imaging in 10 (19%); intraepidermal nerve fiber density in 11 (22%); and corneal nerve fiber length in 27 (52%). The mean corneal nerve fiber length was below normative level (10.1 ± 2.7 mm/mm3 ). DISCUSSION: Small-fiber neuropathy may be common in neurofibromatosis type 1, and should be investigated in symptomatic patients.

Longitudinal follow-up of biopsy-proven small fiber neuropathy.

INTRODUCTION: Little is published on the prognosis of small fiber neuropathy (SFN). METHODS: A retrospective analysis of 101 patients with biopsy proven SFN. RESULTS: Study participants included 87 patients with length-dependent SFN and 14 patients with non-length-dependent SFN. The average duration of symptoms was 3.2 years prior to SFN diagnosis, and the average follow-up duration after diagnosis was 6.2 years. Neuropathic pain was present in 98% of patients and in 84.2% of patients at the final visit. The average total number of pain medications ever used was 4.4 per patient. Signs of autonomic dysfunction were initially present in 24.8% of patients, but improved in most. Large fiber involvement was seen in 11.9% of patients. Small fiber neuropathy affected employment and ambulation status in 5.3% and 6.3% of patients, respectively. DISCUSSION: Small fiber neuropathy tends to be stable and rarely affects ambulation and employment status. Effective pain control remains a challenge.

Expression of macrophage migration inhibitory factor in footpad skin lesions with diabetic neuropathy.

Background Diabetic neuropathy originating in distal lower extremities is associated with pain early in the disease course, overwhelming in the feet. However, the pathogenesis of diabetic neuropathy remains unclear. Macrophage migration inhibitory factor has been implicated in the onset of neuropathic pain and the development of diabetes. Objective of this study was to observe pain syndromes elicited in the footpad of diabetic neuropathy rat model and to assess the contributory role of migration inhibitory factor in the pathogenesis of diabetic neuropathy. Methods Diabetic neuropathy was made in Sprague Dawley rats by streptozotocin. Pain threshold was evaluated using von Frey monofilaments for 24 weeks. On comparable experiment time after streptozotocin injection, all footpads were prepared for following procedures; glutathione assay, terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling staining, immunohistochemistry staining, real-time reverse transcription polymerase chain reaction, and Western blot. Additionally, human HaCaT skin keratinocytes were treated with methylglyoxal, transfected with migration inhibitory factor/control small interfering RNA, and prepared for real-time reverse transcription polymerase chain reaction and Western blot. Results As compared to sham group, pain threshold was significantly reduced in diabetic neuropathy group, and glutathione was decreased in footpad skin, simultaneously, cell death was increased. Over-expression of migration inhibitory factor, accompanied by low expression of glyoxalase-I and intraepidermal nerve fibers, was shown on the footpad skin lesions of diabetic neuropathy. But, there was no significance in expression of neurotransmitters and inflammatory mediators such as transient receptor potential vanilloid 1, mas-related G protein coupled receptor D, nuclear factor kappa B, tumor necrosis factor-alpha, and interleukin-6 between diabetic neuropathy group and sham group. Intriguingly, small interfering RNA-transfected knockdown of the migration inhibitory factor gene in methylglyoxal-treated skin keratinocytes increased expression of glyoxalase-I and intraepidermal nerve fibers in comparison with control small interfering RNA-transfected cells, which was decreased by induction of methylglyoxal. Conclusions Our findings suggest that migration inhibitory factor can aggravate diabetic neuropathy by suppressing glyoxalase-I and intraepidermal nerve fibers on the footpad skin lesions and provoke pain. Taken together, migration inhibitory factor might offer a pharmacological approach to alleviate pain syndromes in diabetic neuropathy.

Sensory nerve degeneration in a mouse model mimicking early manifestations of familial amyloid polyneuropathy due to transthyretin Ala97Ser.

AIMS: Sensory nerve degeneration and consequent abnormal sensations are the earliest and most prevalent manifestations of familial amyloid polyneuropathy (FAP) due to amyloidogenic transthyretin (TTR). FAP is a relentlessly progressive degenerative disease of the peripheral nervous system. However, there is a lack of mouse models to replicate the early neuropathic manifestations of FAP. METHODS: We established human TTR knock-in mice by replacing one allele of the mouse Ttr locus with human wild-type TTR (hTTRwt ) or human TTR with the A97S mutation (hTTRA97S ). Given the late onset of neuropathic manifestations in A97S-FAP, we investigated nerve pathology, physiology, and behavioural tests in these mice at two age points: the adult group (8 - 56 weeks) and the ageing group (> 104 weeks). RESULTS: In the adult group, nerve profiles, neurophysiology and behaviour were similar between hTTRwt and hTTRA97S mice. By contrast, ageing hTTRA97S mice showed small fibre neuropathy with decreased intraepidermal nerve fibre density and behavioural signs of mechanical allodynia. Furthermore, significant reductions in sural nerve myelinated nerve fibre density and sensory nerve action potential amplitudes in these mice indicated degeneration of large sensory fibres. The unaffected motor nerve physiology replicated the early symptoms of FAP patients, that is, sensory nerves were more vulnerable to mutant TTR than motor nerves. CONCLUSIONS: These results demonstrate that the hTTRA97S mouse model develops sensory nerve pathology and corresponding physiology mimicking A97S-FAP and provides a platform to develop new therapies for the early stage of A97S-FAP.

The role of skin biopsy in differentiating small-fiber neuropathy from ganglionopathy.

BACKGROUND AND PURPOSE: We aimed to test the clinical utility of the leg:thigh intraepidermal nerve-fiber (IENF) density ratio as a parameter to discriminate between length-dependent small-fiber neuropathy (SFN) and small-fiber sensory ganglionopathy (SFSG) in subjects with signs and symptoms of small-fiber pathology. METHODS: We retrospectively evaluated thigh and leg IENF density in 314 subjects with small-fiber pathology (173 with distal symmetrical length-dependent SFN and 141 with non-length-dependent SFSG). A group of 288 healthy subjects was included as a control group. The leg:thigh IENF density ratio was calculated for all subjects. We used receiver operating characteristic curve analyses to assess the ability of this parameter to discriminate between length-dependent SFN and SFSG, and the decision curve analysis to estimate its net clinical benefit. RESULTS: In patients with neuropathy, the mean IENF density was 14.8 ± 6.8/mm at the thigh (14.0 ± 6.9/mm in length-dependent SFN and 15.9 ± 6.7/mm in patients with SFSG) and 7.5 ± 4.5/mm at the distal leg (5.4 ± 3.2/mm in patients with length-dependent SFN and 10.1 ± 4.6/mm in patients with SFSG). The leg:thigh IENF density ratio was significantly (P < 0.01) lower in patients with length-dependent SFN (0.44 ± 0.23) compared with patients with SFSG (0.68 ± 0.28). The area under the curve of the receiver operating characteristic analysis to discriminate between patients with length-dependent SFN and SFSG was 0.79. The decision curve analysis demonstrated the clinical utility of this parameter. CONCLUSIONS: The leg:thigh IENF ratio represents a valuable tool in the differential diagnosis between SFSG and length-dependent SFN.

Changes in intraepidermal nerve fiber and Langerhans cell densities in the plantar skin of rats after mercuric chloride exposure.

Mercury and its compounds possess strong neurotoxicity and patients with mercury poisoning often report pain and numbness in the distal extremities that conform to the "stocking-glove" pattern. However, no study has investigated whether damage to small nerve fibers is associated with mercury poisoning. The aims of the present study were to evaluate the effects of different doses of mercury chloride (HgCl2 ) on intraepidermal nerve fibers density (IENFD) and Langerhans cells (LCs) in the plantar skin of rats and to assess the possible relationship between changes in IENFD and sensory testing. Male Sprague-Dawley rats were divided into three experimental groups and administered HgCl2 solutions via gavage at three different doses (4.25, 8.5, and 17 mg/kg/day) for 21 days. Subsequently, behavioral tests and pathological changes in IENFD and LCs were assessed at three different time points (1, 2, and 3 weeks). Rats in all three HgCl2 groups exhibited varying degrees of weight and hair loss. Thermal hypersensitivity was evident in all the HgCl2 groups (for middle-2w subgroup, p < 0.05). Mechanical sensitivity tests revealed hyposensitivity in all the HgCl2 groups except the high-1w subgroup. Significant decreases in IENFD (for the high-1w, middle-1w, low-2w, and low-3w subgroups, p < 0.05) and significant increases in the density of LCs (except for the low-1w and high-2w subgroups, all p < 0.05) were found in all groups after HgCl2 exposure. An association analysis revealed a significant correlation between the decrease in IENFD and the increase in LCs densities (r = -0.573, p < 0.01). The present study demonstrated a decrease in IENFD and an increase in LCs density in the plantar skin of rats after HgCl2 poisoning, indicating that damage of the small nerve fibers occurs after mercury poisoning.

Long-Time Course of Idiopathic Small Fiber Neuropathy.

BACKGROUND: Small fiber neuropathy (SFN) is a challenging subtype of peripheral neuropathies. Once the diagnosis has been established, there is an uncertainty how SFN may progress, whether larger fibers will become involved over time, whether quality of life may be compromised, or whether repeated diagnostic workup in patients with unknown underlying cause may increase the yield of treatable causes of SFN. METHODS: We evaluated 16 patients with documented long-time course of idiopathic SFN. RESULTS: Clinical and electrophysiological course remained stable in 75% of the patients, while 25% SFN-patients developed large fiber neuropathies. CONCLUSIONS: Our data suggest that SFN represents a benign disease course in the majority of patients without severely limiting the quality of life.

SFN-SIQ, SFNSL and skin biopsy of 55 cases with small fibre involvement.

Purpose/aim of the study: To date, there are no validated screening scales for small fibre neuropathy. This study investigated the small-fibre neuropathy and the symptom inventory questionnaire as well as the small fibre neuropathy screening list for small fibre neuropathy diagnosis. METHODS: Fifty-five patients were divided into small fibre neuropathy and mixed fibre damage groups. Relevant scales, nerve conduction studies and skin biopsies were performed. Relationships between the intraepidermal nerve fibre density and different scales as well as the diagnostic and cut-off values (score at which Youden's index is largest) were determined. RESULTS: Compared with healthy Chinese participants, 20 patients were diagnosed with small fibre neuropathy. Intraepidermal nerve fibre density was moderately and highly correlated with the small fibre neuropathy-symptom inventory questionnaire and small fibre neuropathy screening list, respectively. The diagnostic values were moderate and high for the small fibre neuropathy-symptom inventory questionnaire (cut-off value = 5, sensitivity = 80%, specificity = 81.8%) and small fibre neuropathy screening list (cut-off value = 8, sensitivity = 94.1%, specificity = 90.9%), respectively. There were no significant differences in the visual analogue scale between the small fibre neuropathy group, mixed small and large fibre neuropathy group, pure large fibre neuropathy group and the normal group. CONCLUSION: Small fibre neuropathy-symptom inventory questionnaire and small fibre neuropathy screening list represent potential small fibre neuropathy screening tools. Abbreviations EMG electromyography ENA anti-extractable nuclear antigens ESR erythrocyte sedimentation rate IENFD intraepidermal nerve fibre density IGT impaired glucose tolerance NCS nerve conduction studies NDS neuropathy disability score OGTT oral glucose tolerance test PGP protein gene product PN peripheral neuropathy ROC receiver operating characteristic curve ROC-AUC area under the ROC curve SFN small fibre neuropathy SFN-SIQ small-fibre neuropathy and symptom inventory questionnaire SFNSL small fibre neuropathy screening list VAS visual analogue scale WHO World Health Organization.