Axillary web syndrome assessment using a self-assessment questionnaire: a prospective cohort study.

BACKGROUND: Surgical procedure for breast cancer is not without its side effects and one such side effect is axillary web syndrome (AWS), characterized by palpable fibrotic-like cords in the operated arm. As physical evaluation is the only gold standard method used, our study aims to assess the incidence and early detection of AWS with a self-assessment questionnaire. METHODS: From July 2013 to July 2014, 370 breast cancer patients were enrolled. AWS incidence was 51.1%, with 94.1% onset in the first 4 weeks after surgery; 43.5% of the patients did not recover in the first 8 weeks. Univariate analysis showed that BMI (P < 0.001), age (P < 0.001), educational level (P = 0.01), and exercise frequency in the eighth week of follow-up (P < 0.001) were significantly associated with the AWS detection, and multivariate analyses confirmed that younger patients (age < 50) have significantly higher AWS detection (OR = 2.38 (95%CI 1.53, 3.71) and that BMI is associated with AWS, with normal weight patients (BMI ≤ 25) having a significantly greater AWS detection with an odds ratio of 2.11 (95%CI 1.33, 3.36). CONCLUSION: Our findings indicated that the incidence of AWS is high in breast cancer patients, particularly in the first month after surgery. Not all patients achieved recovery during our 8 week follow-up, suggesting that evaluation and treatment should be longer. Double AWS detection was found for patients who were younger (age < 50) and with normal weight.

Management of pregnant women infected with Ebola virus in a treatment centre in Guinea, June 2014.

We report two cases of confirmed Ebola virus disease in pregnant women, who presented at the Médecins Sans Frontières Ebola treatment centre in Guéckédou. Despite the very high risk of death, both pregnant women survived. In both cases the critical decision was made to induce vaginal delivery. We raise a number of considerations regarding the management of Ebola virus-infected pregnant women, including the place of amniocentesis and induced delivery, and whether certain invasive medical acts are justified.

Analysis of T cell receptor repertoire of muscle-infiltrating T lymphocytes in polymyositis. Restricted V alpha/beta rearrangements may indicate antigen-driven selection.

Polymyositis is an inflammatory myopathy characterized by mononuclear cell infiltration of muscle tissue. Myocytotoxic T lymphocytes have been recognized in the infiltrates, but the muscle antigen, target of the immune attack, has not been identified. Molecular characterization of the variable regions of T cell receptors (TCRs) on the infiltrating lymphocytes can be expected to provide insights into the pathogenic process. The V alpha/beta TCR repertoire was investigated by RNA-PCR in muscle biopsies from 15 polymyositis patients and 16 controls (6 Duchenne muscular dystrophy and 10 with no inflammatory or dystrophic myopathy). A variety of rearranged variable TCR genes was found in polymyositis, V alpha 1, V alpha 5, V beta 1, and V beta 15 being the most common (present in 60-100% of patients). In Duchenne muscular dystrophy patients TCR V alpha or beta rearrangements were found although no restriction was observed; no rearrangements were found in muscles from the other controls. Sequence analysis revealed the presence of the J beta 2.1 region in 90% of the V beta 15 clones studied, no random N additions in the diversity region, and a common motif within the CDR3 region. These results suggest that selection of muscle-infiltrating T lymphocytes is antigen driven in polymyositis.

Oral administration of an immunodominant T-cell epitope downregulates Th1/Th2 cytokines and prevents experimental myasthenia gravis.

The mucosal administration of the native antigen or peptide fragments corresponding to immunodominant regions is effective in preventing or treating several T cell-dependent models of autoimmune disease. No data are yet available on oral tolerance with immunodominant T-cell peptides in experimental autoimmune myasthenia gravis (EAMG), an animal model of B cell-dependent disease. We report that oral administration of the T-cell epitope alpha146-162 of the Torpedo californica acetylcholine receptor (TAChR) alpha-subunit suppressed T-cell responses to AChR and ameliorated the disease in C57Bl/6 (B6) mice. Protection from EAMG was associated with reduced serum Ab's to mouse AChR and reduced AChR loss in muscle. The effect of Talpha146-162 feeding was specific; treatment with a control peptide did not affect EAMG manifestations. The protective effect induced by peptide Talpha146-162 was mediated by reduced production of IFN-gamma, IL-2, and IL-10 by TAChR-reactive cells, suggesting T-cell anergy. TGF-beta-secreting Th3 cells did not seem to be involved in tolerance induction. We therefore demonstrate that feeding a single immunodominant epitope can prevent an Ab-mediated experimental model of autoimmune disease.

Dendritic cells pulsed with glioma lysates induce immunity against syngeneic intracranial gliomas and increase survival of tumor-bearing mice.

In recent years, the use of dendritic cells (DC), the most powerful antigen presenting cells, has been proposed for the creation of vaccines against gliomas. This approach has been demonstrated to be safe and non-toxic in phase I or I-II trials (2, 3). Immunotherapy plays a central role in the search for new treatments for glioblastoma multiforme (GBM). In particular, several phase I studies have been performed using DC pulsed by GBM proteins as a vaccine for patients with relapsing GBM. The studies demonstrated that DC vaccination is safe and may produce a significant increase in overall survival. As the first step in the preparation of appropriate conditions for a clinical evaluation in Italy, we have performed pre-clinical experiments on immune-competent mice injected intra-cerebrally with syngeneic GL261GBM cells and treated subcutaneously and intra-tumorally with DC loaded with a GL261 homogenate. These results show that vaccination with DC pulsed with a tumor lysate increases considerably survival in mice bearing intracranial glioblastomas and supports the development of DC-based clinical trials for patients with glioblastomas that do not respond to standard therapies.

Species identification of non-tuberculous mycobacteria from humans and cattle of Chad.

In Chad, during a study on tuberculosis in humans and cattle, 52 non-tuberculous mycobacteria (NTM) strains were isolated. By means of INNO-LiPA, PRA-hsp65 amplification and sequencing of 16S rDNA, NTM species of 25/52 isolates were identified. M. fortuitum complex (8) was the most frequent species, followed by M. nonchromogenicum (4) and M. avium complex (4). PRA method could identify M. fortuitum 3rd variant among isolates derived from cattle specimens. This finding could confirm the existence of farcy in the Chadian cattle population as M. fortuitum 3rd variant and putitative pathogen M. farcinogenes can't be distinguished by the methods used in this study. Half of the NTM isolates could not be specified and we considered them as contaminants from the environment.

Rapid detection of diarrheagenic E. coli by real-time PCR.

Enterovirulent Escherichia coli are among the most important causes of acute diarrhea in developing as well as in developed countries. We have adapted classical PCR to detect these organisms in stool specimens to real-time PCR using the LightCycler (LC) SYBR Green format followed by melting curve analysis. With only two different cycling protocols we could detect enteropathogenic E. coli (EPEC) and verocytotoxin-producing E. coli (VTEC) (duplex assay for both Verotoxin 1 (VT1) and Verotoxin 2 (VT2)) in one run and enteroaggregative E. coli (EAEC), enteroinvasive E. coli (EIEC) and enterotoxigenic E. coli (ETEC) (duplex assay detecting both heat-stable enterotoxin (ST) and heat-labile enterotoxin (LT)) in another run. Using serial dilutions of control strains, the LC proved to be clearly more sensitive than conventional PCR for five out of seven investigated targets: VTEC (VT1 and VT2), ETEC (ST and LT) and EIEC. For EPEC and EAEC, LC and conventional PCR had identical sensitivities. With stool samples, we found an optimal agreement between LC-PCR and the conventional PCR when samples were tested in a 1:10 dilution. Only one specimen was discrepant, being repetitively positive for VT by LightCycler but not by conventional PCR. Given the significantly higher sensitivity of the LC-PCR for the VT target (up to a 10(-4) dilution factor by melting curve analysis and up to a 10(-6) dilution factor following gel electrophoresis), this is probably a false negative result by conventional PCR. We conclude that LightCycler PCR is more rapid, easier than and at least as sensitive as our conventional PCR for the detection of enterovirulent E. coli in stool specimens after culture on MacConkey.

Two isoforms of the muscle acetylcholine receptor alpha-subunit are translated in the human cell line TE671.

We have previously reported the existence of 2 forms of mRNA for the human muscle acetylcholine receptor (AChR) alpha-subunit, thought to be generated by alternate splicing of a primary transcript and to encode 2 alpha-subunit protein isoforms. The 2 predicted alpha-subunit isoforms, differing by the insertion of 25 amino acids at position 58/59, have been synthesized from cRNA transcripts using rabbit reticulocyte lysates; these protein isoforms could be differentiated by immunoprecipitation using antibodies raised against synthetic peptides. The antibodies were used to demonstrate translation of both AChR alpha-subunit isoforms in the rhabdomyosarcoma (muscle) cell line TE671, in an approximate 1:1 ratio.

Presentation of endogenous acetylcholine receptor epitope by an MHC class II-transfected human muscle cell line to a specific CD4+ T cell clone from a myasthenia gravis patient.

Muscle or thymic myoid cells, if induced to express MHC class II in addition to endogenous acetylcholine receptor (AChR), might present epitopes derived from the AChR to specific CD4+ T cells. These T cells could in turn initiate or maintain the anti-AChR response that is responsible for AChR loss in myasthenia gravis (MG). We transfected the AChR+ TE671 (rhabdomyosarcoma) cells with HLA-DR4 and co-cultured them with the DR4-restricted, CD4+ T cell clone (PM-A1; raised from a hyperplastic thymus of an MG patient and previously shown to recognise all forms of the AChR that contain the sequence alpha 144-156). Significant T cell activation, demonstrated both by 3H-thymidine incorporation and by lysis of the TE671 cells, was found in the presence of added alpha 144-156 and, more importantly, in the absence of exogenous antigen. These results show that MHC class II-expressing muscle or other AChR-expressing cells could present endogenous AChR to pathogenic T cells. This process may be important in the aetiology of MG.

Major histocompatibility complex class II molecule expression on muscle cells is regulated by differentiation: implications for the immunopathogenesis of muscle autoimmune diseases.

Major histocompatibility complex (MHC) class II molecules are expressed on myoblasts after interferon-gamma (IFN-gamma) treatment, suggesting a muscle cell involvement in antigen presentation in inflammatory myopathies. However, they were not observed on normal or pathological myofibers. This discrepancy might be related to different responsiveness of developmentally differentiated muscle cells to IFN-gamma. Myoblasts expressed class II transcripts and proteins after IFN-gamma, while myotubes and innervated contracting muscle cells did not show staining for class II molecules. At all cell stages no loss of IFN-gamma receptor was detected indicating that myofiber maturation blocks their capacity to express MHC class II molecules. This suggests that completely differentiated myofibers cannot participate in class II restricted immunological reactions.

The expression of co-stimulatory and accessory molecules on cultured human muscle cells is not dependent on stimulus by pro-inflammatory cytokines: relevance for the pathogenesis of inflammatory myopathy.

Specific activation of naive T cells requires TCR engagement plus interaction of CD28 on T cells with co-stimulatory B7-1/B7-2 on APCs. Since muscle cells may be directly involved in activating muscle-infiltrating T lymphocytes in polymyositis and inclusion body myositis, we analyzed B7 expression on myoblasts before and after treatment with pro-inflammatory cytokines. We found no expression of B7-1/B7-2, either constitutively or after stimulus with cytokines. Furthermore, myoblasts failed to stimulate alloreactive peripheral blood lymphocytes in mixed lymphocyte reactions. Lack of B7 expression was confirmed by immunostaining of polymyositis patients' muscle: only T and the few B lymphocytes present in inflammation areas expressed B7-1.

Persistence of viral pathogens and bacteriophages during sewage treatment: lack of correlation with indicator bacteria.

The effects of different sewage treatments on the viral contamination in rivers which receive water from treatment plants without a final sand filtration step were investigated. They were all heavily contaminated with bacteriophages and human enteric viruses (detected by single step reverse transcription amplification followed by a nested polymerase chain reaction). Bacteriophages, but not faecal indicator organisms, were correlated with viral contamination.

Approaches for studying the pathogenic T cells in autoimmune patients.

Our provisional conclusions from this work are as follows. (1) For screening responses of established lines, native human AChR is not prohibitively scarce, especially if it is concentrated onto beads, and class II-transfected TE671 cells may be useful too; both may give vital evidence of AChR-specificity, but it is still crucial to confirm that with synthetic peptides. (2) For mapping epitopes, panels of full-length and shorter recombinant human polypeptides, and of synthetic peptides, are invaluable complementary material: longer peptides tend to stimulate particularly strongly. (3) Initial selection with pooled synthetic peptides can easily generate interesting lines from both patients and controls, but they may depend on the artificial processing sites that are an inevitable consequence of arbitrarily chosen start and stop points. Of course, these might conceivably be employed in unusual antigen-presenting cells (such as thymic myoid cells), so we cannot totally dismiss such "cryptic" epitopes. This system can sometimes select T cells responding to "natural" epitopes too, as now reported for tetanus toxin. Nevertheless, for these and other reasons, at present, we strongly favor using the longest human recombinant material possible, because it is apparently processed more naturally. This must be combined with rigorous screening for reactivity to E. coli-derived contaminants plus concomitant mapping of epitopes as above. Use of intact AChR for initiating lines may yet become feasible. (4) The T cells thus isolated and characterized so far are proving to be heterogeneous in the epitopes and presenting class II molecules they recognize, and in their T-cell receptor gene usage. It is premature to claim key myasthenogenic epitopes or clonotypes, but HLA-DR3 and the linked -DQw2 do not appear to monopolize presentation. (5) Assessing the disease-relevance of these T cells is a separate problem, highlighted by their apparent similarity in healthy controls. In the meantime, to test their potential pathogenicity, we are assaying their cytokine profiles and ability to help specific antibody production in vitro. In the hope that they do prove to be relevant, we are also using some of them to test possible therapeutic strategies that might prove applicable in the patients.

A short plasma exchange protocol is effective in severe myasthenia gravis.

Plasma exchange has been reported to be a successful therapeutic procedure for the treatment of severely compromised myasthenic patients, but the optimal regimen in terms of costs or clinical benefit has not so far been determined. We have investigated the efficacy of a short plasmapheresis protocol of two exchanges 1 day apart in a series of 70 patients with severe forms of myasthenia gravis. Patients were evaluated before and 7 days after the first exchange. A positive outcome was observed in 70% of the plasma exchange cycles performed. Disease severity did not seem to be a negative prognostic factor for the efficacy of this short protocol, which was well tolerated by patients. In only 1 case were major side-effects observed. In spite of its short duration, the exchange treatment plus concomitant immunosuppressive drug therapy was not followed by early clinical deterioration.

[First isolation of tuberculous mycobacteria in man and animals in Chad]. / Premiers isolements de mycobactéries tuberculeuses chez l'homme et l'animal au Tchad.

This report describes the first successful isolation and identification of mycobacterial infection in humans and animals of Chad. All mycobacterial strains from human specimens were M. tuberculosis and strains from animal specimens (cattle) were M. bovis. None of the 10 of M. tuberculosis strains tested for antibiotic resistance were multidrug resistant. Due to the intrinsic resistance of M. bovis to pyrazinamide and the growing number of tuberculosis cases in HIV-infected people in Africa and elsewhere, more information on the potential of M. bovis for human infection is needed to guide disease control policy.

Motor and functional recovery after neck dissection: comparison of two early physical rehabilitation programmes.

The aim of this prospective, single-centre, non-randomized explorative study is to comparatively assess two-month results of two early rehabilitation programmes in patients receiving neck dissection for head and neck cancer, with the hypothesis that those not receiving therapist-assisted physiotherapy would take an active role in their own rehabilitation to enhance outcomes. At the European Institute of Oncology, Milan (Italy), 97 patients were registered during the pre-hospitalization period and divided into an Autonomous group (living distant from the hospital) and a Physio group (living near). As expected, only 50 patients (25 per group) completed the study. Both groups received a Physical Therapy Brochure with instructions on to how to perform exercises at home. Home physical exercises started five days after surgery and continued for two months. The Autonomous group received a pre-surgery instruction session; the Physio group attended four once-weekly therapist-guided physiotherapy sessions. Two months after surgery, arm mobility and pain had recovered to pre-operative levels. Most endpoints, including the main composite, did not differ between groups. Although longer-follow-up is necessary, early physiotherapy seems to be effective in maintaining arm mobility and reducing pain, even in patients empowered to do exercises autonomously.

Type I interferon and Toll-like receptor expression characterizes inflammatory myopathies.

OBJECTIVES: Juvenile dermatomyositis (JDM), adult dermatomyositis, and polymyositis (PM) are idiopathic inflammatory myopathies (IIMs) characterized by muscle infiltration and specific muscle fiber alterations. They are thought to have an autoimmune etiology, but triggering factors, and how immunologic attack induces muscle weakness, remain unknown. Recent evidence suggests a key role for type I interferon (IFN)-mediated innate immunity in dermatomyositis, which we explored in JDM, dermatomyositis, and PM by gene expression profiling, and other methods. METHODS: Ten IIM and 5 control muscle biopsies were assessed for expression of approximately 16,000 genes by microarray; 37 additional IIM, 10 dystrophinopathic, and 14 nonmyopathic control muscles were studied for type I IFN-dependent genes, and Toll-like receptor (TLR) expression by immunochemistry and PCR. RESULTS: Type I IFN-dependent transcripts were significantly upregulated in IIM muscles compared to controls; in JDM the most expressed were ISG15 (408-fold), IFIT3 (261-fold), MX1 (99-fold), and IRF7 (37-fold). IFN-ß (but not IFN-α) transcripts were upregulated in PM as well as dermatomyositis/JDM. TLR3 was upregulated particularly in JDM, being localized on vascular endothelial cells, muscle infiltrating cells (mainly myeloid dendritic cells), and regenerating myofibers; TLR7 and TLR9 proteins were present in IIM (prominently in PM), mainly on cell infiltrates, particularly plasma cells, and on some injured myofibers. CONCLUSIONS: IFN-ß and type I IFN-induced molecules are involved in PM as well as JDM/dermatomyositis. Endosomal TLRs (effectors of innate immunity) are also involved (but differently) in the 3 conditions, further suggesting viral involvement, although TLR activation could be secondary to tissue damage.

Detection of poliovirus-infected macrophages in thymus of patients with myasthenia gravis.

BACKGROUND: Genetic and environmental factors are thought to contribute to the etiology of the autoimmune disease myasthenia gravis (MG). Viral involvement has long been suspected, but direct evidence of involvement has not been found. We recently reported that Toll-like receptor 4 (TLR4)-a key activator of innate immunity-was overexpressed in the thymus of some patients with MG, suggesting that thymic infection by pathogens might be involved in MG pathogenesis. We searched for evidence of intrathymic infection in patients with MG. METHODS: Twenty-seven MG thymuses (6 involuted, 7 hyperplastic, 5 thymitis, and 9 thymoma) previously tested for TLR4 expression, 18 nonpathologic control thymuses, and 10 pathologic control thymuses from patients without MG (8 thymoma and 2 hyperplastic) were analyzed for cytomegalovirus, varicella-zoster virus, herpes simplex virus types 1 and 2, eubacteria, respiratory syncytial virus, and enteroviruses using PCR techniques. Immunohistochemistry and double immunofluorescence were used to detect enterovirus capsid protein VP1 in thymic specimens and analyze TLR4 expression in VP1-positive cells. RESULTS: Poliovirus was detected in 4 MG thymuses (14.8%; 2 thymitis and 2 thymoma). No virus was detected in any control thymus. A linear correlation between plus and minus strand poliovirus RNA levels was observed in all 4 thymuses, suggesting persistent thymic infection. VP1 protein was detected in the cytoplasm of CD68-positive macrophages scattered through thymic medulla in all PV-positive thymuses. VP1 and TLR4 colocalized in infected cells. CONCLUSIONS: Poliovirus-infected macrophages are present in thymus of some patients with myasthenia gravis, suggesting a viral contribution to the intrathymic alterations leading to the disease.