Muscle biopsy in anti-neutrophil cytoplasmic antibody-associated vasculitis: diagnostic yield depends on anti-neutrophil cytoplasmic antibody type, sex and neutrophil count.

OBJECTIVES: This study aimed to examine the sensitivity of muscle biopsy (MB) in ANCA-associated vasculitis (AAV), identify factors predicting MB positivity and assess the prognostic value of a positive MB. METHODS: We conducted a single-centre retrospective study of AAV with an MB performed at diagnosis. AAV classification [granulomatosis with polyangiitis (GPA), microscopic polyangiitis (MPA), eosinophilic granulomatosis with polyangiitis (EGPA)] followed the European Medicines Agency algorithm. A logistic regression model was used to identify the factors associated with MB positivity. Survival curves were generated using the Kaplan-Meier method. RESULTS: Among 276 AAV patients (1995-2018), 101 had an MB. Seventy-eight patients were included: 33 with GPA, 25 with MPA and 20 with EGPA. MB samples were positive in 45 cases (58%): 17 GPA, 16 MPA and 12 EGPA. Univariate analysis focussed on GPA and MPA, revealed that the MB yield was higher in females [22/31 (71%) vs 11/27 (41%); P = 0.02] and in anti-MPO patients [25/37 (68%) vs 6/19 (32%) for anti-PR3; P = 0.01]. By multivariate analysis, three factors predicted MB positivity: anti-MPO ANCA [odds ratio (OR) 10.67 (CI 2.09, 81.68)], female sex [OR 5.3 (CI 1.16, 32.35)] and neutrophil count [OR 1.33 (CI 1.07, 1.8)]. MB positivity had no impact on relapse, death or end-stage renal disease-free survival. CONCLUSIONS: MB is a safe and efficient diagnostic tool for AAV. Predictors of MB yield include ANCA type, sex and neutrophil count. MB cannot substitute for kidney biopsy when indicated, but should be considered in other cases.

Two novel variants in CNTNAP1 in two siblings presenting with congenital hypotonia and hypomyelinating neuropathy.

Homozygous frameshift variants in CNTNAP1 have recently been reported in patients with arthrogryposis and abnormal axon myelination. In two brothers with severe congenital hypotonia and foot deformities, we identified compound heterozygous variants in CNTNAP1, reporting the first causative missense variant, p.(Cys323Arg). Motor nerve conductions were markedly decreased. Nerve microscopical lesions confirmed a severe hypomyelinating process and showed loss of attachment sites of the myelin loops on the axons, which could be a characteristic of Caspr loss-of-function. We discuss the pathophysiology of the myelination process and we propose to consider this disorder as a congenital hypomyelinating neuropathy.

Sjögren's syndrome-associated myositis with germinal centre-like structures.

OBJECTIVE: Muscular impairment is a rare systemic manifestation of SS that is rarely described in the literature and classically non-specific, both clinically and histologically. We reviewed the cases of 4 patients with primary SS presenting with myositis and a common histologic pattern on muscular biopsy with germinal centre-like structures resembling that which occurs in salivary glands. METHODS: We analysed the data files of patients with SS who had muscular manifestations and underwent a muscular biopsy. Among 23 patients with SS who had muscle biopsies, 13 had non-specific myositis and 10 (4 primary and 6 secondary SS) had a common histologic pattern consisting of germinal centre-like structures. We analysed the data files of the 4 patients with primary SS presenting with myositis with muscular germinal-centre like structures. RESULTS: The 4 patients had an unspecific clinical presentation, with myalgias, muscular weakness and normal or elevated values of CPK. In the four patients, SS-associated myositis had common histologic characteristics, with endomysial and perimysial inflammatory infiltrate. The cellular infiltrate was composed predominantly of CD4+ T lymphocytes and B lymphocytes. The B and T CD4+ cells infiltrates may gather into masses, even forming lymphoid follicles. Three patients were treated with corticosteroids and/or hydroxychloroquine with improvement of myositis and 1 patient was lost to follow-up. CONCLUSIONS: We describe four patients with a common histologic appearance of myositis with lymphoid follicles associated with primary SS. The clinical presentation was non-specific and non-severe, with favorable outcome with corticosteroids and/or hydroxycholoroquine. The discovery of this particular histologic appearance in a muscle biopsy independent of the final diagnosis should indicate the possibility of SS.

Expanding the clinical spectrum of hereditary fibrosing poikiloderma with tendon contractures, myopathy and pulmonary fibrosis due to FAM111B mutations.

BACKGROUND: Hereditary Fibrosing Poikiloderma (HFP) with tendon contractures, myopathy and pulmonary fibrosis (POIKTMP [MIM 615704]) is a very recently described entity of syndromic inherited poikiloderma. Previously by using whole exome sequencing in five families, we identified the causative gene, FAM111B (NM_198947.3), the function of which is still unknown. Our objective in this study was to better define the specific features of POIKTMP through a larger series of patients. METHODS: Clinical and molecular data of two families and eight independent sporadic cases, including six new cases, were collected. RESULTS: Key features consist of: (i) early-onset poikiloderma, hypotrichosis and hypohidrosis; (ii) multiple contractures, in particular triceps surae muscle contractures; (iii) diffuse progressive muscular weakness; (iv) pulmonary fibrosis in adulthood and (v) other features including exocrine pancreatic insufficiency, liver impairment and growth retardation. Muscle magnetic resonance imaging was informative and showed muscle atrophy and fatty infiltration. Histological examination of skeletal muscle revealed extensive fibroadipose tissue infiltration. Microscopy of the skin showed a scleroderma-like aspect with fibrosis and alterations of the elastic network. FAM111B gene analysis identified five different missense variants (two recurrent mutations were found respectively in three and four independent families). All the mutations were predicted to localize in the trypsin-like cysteine/serine peptidase domain of the protein. We suggest gain-of-function or dominant-negative mutations resulting in FAM111B enzymatic activity changes. CONCLUSIONS: HFP with tendon contractures, myopathy and pulmonary fibrosis, is a multisystemic disorder due to autosomal dominant FAM111B mutations. Future functional studies will help in understanding the specific pathological process of this fibrosing disorder.

Atypical nuclear abnormalities in a patient with Brody disease.

Brody disease was first described as a benign pseudo-myotonic disorder with muscular stiffness, which increased with exercise. Biochemical and genetic studies have pointed out its close relationship to a functional defect of the fast-twitch sarcoplasmic reticulum Ca(++) ATPase pump (SERCA1) encoded by the ATP2A1 gene located on chromosome 16. The histopathological features in this form of myopathy were generally described as non-specific, i.e. moderate degree of type 2 fibre atrophy and excess of internal nuclei. We here present the clinical and histopathological features of a patient with Brody disease over a 19-year follow-up period. This patient had two heterozygous ATP2A1 mutations and complained about muscle stiffness immediately after effort. He had suffered from this since early childhood and exhibited clinical symptoms mimicking myotonia. Histological, ultrastructural and cytogenetic analyses revealed morphologically abnormal nuclei with polyploidy. In this report, we discuss the possible links between the consequences of the genetic abnormality and the peculiar aspect of the nuclei.

Muscle magnetic resonance imaging abnormalities in X-linked myopathy with excessive autophagy.

INTRODUCTION: X-linked myopathy with excessive autophagy (XMEA) is an X-linked recessive myopathy due to recently reported mutations in the VMA21 gene. METHODS: Four men from 2 separate families were studied. The clinical presentation, genetic data, muscle biopsy, and muscle MRI were analyzed. RESULTS: A known VMA21 mutation, c.163+4A>G, and a new mutation, c.163+3A>G, respectively, were found in the 2 families. The clinical course was characterized by onset in childhood and progressive muscle weakness with a limb-girdle pattern. Muscle biopsy revealed a mild myopathy with an increased number of giant autophagic vacuoles. Whole-body muscle MRI showed that pelvic girdle and proximal thighs were the most and earliest affected territories, with sparing of rectus femoris muscles. Muscle changes essentially consisted of degenerative fatty replacement. CONCLUSIONS: This study highlights a distinctive MRI pattern of muscle involvement, which can be helpful for diagnosis of XMEA, even before muscle biopsy or genetic analysis.

Crushed and injected buprenorphine tablets: characteristics of princeps and generic solutions.

Self-injection of high-dose buprenorphine is responsible for well-described complications. In 2011, we have been alerted by unusual but serious cutaneous complication among injection buprenorphine users. A prospective data collection identified 30 cases of necrotic cutaneous lesions after injection of filtered buprenorphine solution, among which 25 cases occurred following injection of buprenorphine generics. The main goal of our study was to put forward particularities that could explain the cutaneous complications, by qualitatively and quantitatively confronting particles present in Subutex and generics solutions. We used the same protocol that injected-buprenorphine users: generic or subutex tablets were crushed in sterile water and filtered through 2 filters commonly used (cotton-pad and sterifilt). Solutions were analyzed by laser granulometry, flow cytometry and scanning electron microscopy. We have highlighted the wide variation of the quantity and the size of the particles present in solution between the two drugs after cotton-pad filtration. The proportion of particles <10 µm is systematically higher in the generic solutions than with Subutex. All of the insoluble particles found in generic solutions contain silica, whereas non- organic element was to be identified in the insoluble particles of Subutex. One skin biopsy obtained from one patient who developed a necrotic lesion after intravenous injection of filtrated solution of buprenorphine generic, shows non-organic elements. Identification of particles in situ enables us to confirm the presence of silica in the biopsy. Actually the monitoring of patient receiving generic of buprenorphine must be strengthened.

The spectrum of renal involvement in patients with inflammatory myopathies.

Data regarding the incidence and outcome of renal involvement in patients with inflammatory myopathies (IM) remain scarce. We assessed the incidence and causes of acute kidney injury (AKI) and chronic kidney disease (CKD) in 150 patients with dermatomyositis, polymyositis, and antisynthetase syndrome followed in 3 French referral centers. Renal involvement occurred in 35 (23.3%) patients: AKI in 16 (10.7%), and CKD in 31 (20.7%) patients. The main cause of AKI was drug or myoglobinuria-induced acute tubular necrosis. Male sex, cardiovascular risk factors, cardiac involvement, and initial proteinuria >0.3 g/d were associated with the occurrence of AKI. The outcome of patients with AKI was poor: 13 (81%) progressed to CKD and 2 (12.5%) reached end-stage renal disease. In multivariate survival analysis, age at IM onset, male sex, a history of cardiovascular events, and a previous episode of AKI were associated with the risk of CKD. We also identified 14 IM patients who underwent a kidney biopsy in 10 nephrology centers. Renal pathology disclosed a wide range of renal disorders, mainly immune-complex glomerulonephritis. We identified in 5 patients a peculiar pattern of severe acute renal vascular damage consisting mainly of edematous thickening of the intima of arterioles. We found that AKI and CKD are frequent in patients with IM. Prevention of AKI is crucial in these patients, as AKI is a major contributor to their relatively high risk of CKD. A peculiar pattern of acute vascular damage is part of the spectrum of renal diseases associated with IM.

Mutations in CNTNAP1 and ADCY6 are responsible for severe arthrogryposis multiplex congenita with axoglial defects.

Non-syndromic arthrogryposis multiplex congenita (AMC) is characterized by multiple congenital contractures resulting from reduced fetal mobility. Genetic mapping and whole exome sequencing (WES) were performed in 31 multiplex and/or consanguineous undiagnosed AMC families. Although this approach identified known AMC genes, we here report pathogenic mutations in two new genes. Homozygous frameshift mutations in CNTNAP1 were found in four unrelated families. Patients showed a marked reduction in motor nerve conduction velocity (<10 m/s) and transmission electron microscopy (TEM) of sciatic nerve in the index cases revealed severe abnormalities of both nodes of Ranvier width and myelinated axons. CNTNAP1 encodes CASPR, an essential component of node of Ranvier domains which underlies saltatory conduction of action potentials along the myelinated axons, an important process for neuronal function. A homozygous missense mutation in adenylate cyclase 6 gene (ADCY6) was found in another family characterized by a lack of myelin in the peripheral nervous system (PNS) as determined by TEM. Morpholino knockdown of the zebrafish orthologs led to severe and specific defects in peripheral myelin in spite of the presence of Schwann cells. ADCY6 encodes a protein that belongs to the adenylate cyclase family responsible for the synthesis of cAMP. Elevation of cAMP can mimic axonal contact in vitro and upregulates myelinating signals. Our data indicate an essential and so far unknown role of ADCY6 in PNS myelination likely through the cAMP pathway. Mutations of genes encoding proteins of Ranvier domains or involved in myelination of Schwann cells are responsible for novel and severe human axoglial diseases.

Mutations in FAM111B cause hereditary fibrosing poikiloderma with tendon contracture, myopathy, and pulmonary fibrosis.

Congenital poikiloderma is characterized by a combination of mottled pigmentation, telangiectasia, and epidermal atrophy in the first few months of life. We have previously described a South African European-descent family affected by a rare autosomal-dominant form of hereditary fibrosing poikiloderma accompanied by tendon contracture, myopathy, and pulmonary fibrosis. Here, we report the identification of causative mutations in FAM111B by whole-exome sequencing. In total, three FAM111B missense mutations were identified in five kindreds of different ethnic backgrounds. The mutation segregated with the disease in one large pedigree, and mutations were de novo in two other pedigrees. All three mutations were absent from public databases and were not observed on Sanger sequencing of 388 ethnically matched control subjects. The three single-nucleotide mutations code for amino acid changes that are clustered within a putative trypsin-like cysteine/serine peptidase domain of FAM111B. These findings provide evidence of the involvement of FAM111B in congenital poikiloderma and multisystem fibrosis.

A monoclonal antibody to O-acetyl-GD2 ganglioside and not to GD2 shows potent anti-tumor activity without peripheral nervous system cross-reactivity.

BACKGROUND: Monoclonal antibodies (mAb) against GD2 ganglioside have been shown to be effective for the treatment of neuroblastoma. Beneficial actions are, however, associated with generalized pain due to the binding of anti- GD2 mAbs to peripheral nerve fibers followed by complement activation. Neuroblastoma cells that express GD2 also express its O-acetyl derivative, O-acetyl- GD2 ganglioside (OAcGD2). Hence, we investigated the distribution of OAcGD2 in human tissues using mAb 8B6 to study the cross-reactivity of mAb 8B6 with human tissues. METHODOLOGY/PRINCIPAL FINDINGS: The distribution of OAcGD2 was performed in normal and malignant tissues using an immunoperoxydase technique. Anti-tumor properties of mAb 8B6 were studied in vitro and in vivo in a transplanted tumor model in mice. We found that OAcGD2 is not expressed by peripheral nerve fibers. Furthermore, we demonstrated that mAb 8B6 was very effective in the in vitro and in vivo suppression of the growth of tumor cells. Importantly, mAb 8B6 anti-tumor efficacy was comparable to that of mAb 14G2a specific to GD2. CONCLUSION/SIGNIFICANCE: Development of therapeutic antibodies specific to OAcGD2 may offer treatment options with reduced adverse side effects, thereby allowing dose escalation of antibodies.

Use of muscle biopsies for diagnosis of systemic vasculitides.

OBJECTIVE: Few studies have investigated the use of muscle biopsies (MB) for the diagnosis of systemic vasculitides (SV). We aimed to evaluate the diagnostic use of MB in this condition. METHODS: We reviewed 310 consecutive MB performed in our center between 2000 and 2008 and correlated them with clinical data from the corresponding patients. Thirty-one of the patients, representing a total of 33 MB, were diagnosed with active SV. MB were considered positive when they demonstrated either necrotizing vasculitis or nonnecrotizing vasculitis. RESULTS: Twenty-two of the 33 MB were positive (sensitivity of 66.7%), with necrotizing vasculitis and nonnecrotizing vasculitis being equally frequent. The SV were antineutrophil cytoplasmic antibody (ANCA)-associated in 22 patients (71%), and ANCA-negative in 9 cases (29%). Neither the type nor the clinical spectrum of the SV was predictive of MB positivity. None of the muscle symptoms (myalgias or biological rhabdomyolysis) were correlated with MB positivity. All the biopsies were performed uneventfully. CONCLUSION: The feasibility and positive predictive value of MB make it a valuable tool for ruling out a diagnosis of SV. Since no clinical signs could predict its positivity, MB should be considered in all suspected cases of SV. Unlike other biopsies, including kidney biopsy, MB had no prognostic value.

A new MPZ mutation associated with a mild CMT1 phenotype presenting with recurrent nerve compression.

P0 is a transmembrane protein of the immunoglobulin superfamily that plays a role in myelin structure and function. Myelin protein zero gene (MPZ) mutations usually cause a demyelinating variant of Charcot-Marie-Tooth disease type 1B (CMT1B), but there is a wide spectrum of phenotypic manifestation of these mutations. We describe three patients from one family and one separate patient who presented with a demyelinating neuropathy. Some had recurrent lesions at compression sites mimicking hereditary neuropathy with liability to pressure palsies (HNPP). A heterozygous nonsense mutation (Tyr145Stop) corresponding to a T-to-A transition at nucleotide position 435 in exon 3 of the MPZ gene was identified in all patients. This mutation leads to an extracellular truncated protein, which may explain the mild phenotype. Therefore, such MPZ gene mutations should be searched for in cases of demyelinating neuropathy with acute nerve compression as well as in cases of the HNPP phenotype associated with normal the PMP22 gene.

Distal inflammatory myopathy: unusual presentation of polymyositis or new entity?

New classification of idiopathic inflammatory myopathy (IIM) defined three major entities, polymyositis (PM), dermatomyositis (DM) and sporadic inclusion body myositis (s-IBM). We report the clinical, electrophysiological and pathological characteristics of three patients with a rare form of IIM not fulfilling the diagnostic criteria for any of these three major entities. The three patients presented with a subacute, distal asymmetrical weakness in upper limbs. Muscle biopsy showed an active myositis, with necrosis and regeneration, T cell infiltrates with invasion of non-necrotic fibers, without rimmed vacuoles, and diffuse major histocompatibility complex-I (MHC-I) immunostaining in muscle fibers. All patients responded to immunosuppressive agents. Seven others cases were identified in the literature. It is important to recognize this atypical presentation as it seems to respond to immunosuppressive agents.

The gene encoding adipose triglyceride lipase (PNPLA2) is mutated in neutral lipid storage disease with myopathy.

Neutral lipid storage disease comprises a heterogeneous group of autosomal recessive disorders characterized by systemic accumulation of triglycerides in cytoplasmic droplets. Here we report a neutral lipid storage disease subgroup characterized by mild myopathy, absence of ichthyosis and mutations in both alleles of adipose triglyceride lipase (PNPLA2, also known as ATGL). Three of these mutations are predicted to lead to a truncated ATGL protein with an intact patatin domain containing the active site, but with defects in the hydrophobic domain. The block in triglyceride degradation was mimicked by short interfering RNA directed against ATGL. NLSDM is distinct from Chanarin-Dorfman syndrome, which is characterized by neutral lipid storage disease with ichthyosis, mild myopathy and hepatomegaly due to mutations in ABHD5 (also known as CGI-58).

In situ immunophenotype of the inflammatory infiltrate in eosinophilic fasciitis.

OBJECTIVE: Eosinophilic fasciitis (EF) is histologically characterized by a fibrous and inflammatory thickening of subcutaneous septal-fascial-perimysial collagenous scaffold. This study aims to define the immunophenotype of inflammatory cells of fascia and muscle underlying the in situ immune response in EF. METHODS: In 11 cases of EF, we determined the phenotype of inflammatory cells, expression of MHC class I and class II antigens, and C5b9 membranolytic attack complex (MAC) deposits by immunohistochemistry analysis of fascia tissue. Muscle biopsies from 9 patients with active dermatomyositis and 5 with active polymyositis were used as controls. Results. In all patients but one, the inflammatory infiltrate was mainly composed of macrophages associated with CD8+ T lymphocytes (CD4/CD8 ratio < 1) and few eosinophils. Cytotoxic properties were found in 14% of CD8+ T lymphocytes, as shown by granzyme B expression. MHC Class I antigens were overexpressed (5/7) by muscle fibers, with a paratrabecular reinforcement in 4 cases. MHC class II antigens were not expressed by muscle fibers except in one case. C5b9 MAC deposits were not detected. CONCLUSION: Our in situ characterization of inflammatory infiltrate demonstrates the predominancy of macrophages and CD8+ T lymphocytes. Some of these CD8+ lymphocytes contain granzyme B, thus suggesting a cytotoxic cellular immune response in EF, which could be triggered by infectious or environmental agents.