Circulating bioactive bacterial DNA is associated with immune activation and complications in common variable immunodeficiency.

Common variable immunodeficiency (CVID) is characterized by profound primary antibody defects and frequent infections, yet autoimmune/inflammatory complications of unclear origin occur in 50% of individuals and lead to increased mortality. Here, we show that circulating bacterial 16S rDNA belonging to gut commensals was significantly increased in CVID serum (P < 0.0001), especially in patients with inflammatory manifestations (P = 0.0007). Levels of serum bacterial DNA were associated with parameters of systemic immune activation, increased serum IFN-γ, and the lowest numbers of isotype-switched memory B cells. Bacterial DNA was bioactive in vitro and induced robust host IFN-γ responses, especially among patients with CVID with inflammatory manifestations. Patients with X-linked agammaglobulinemia (Bruton tyrosine kinase [BTK] deficiency) also had increased circulating bacterial 16S rDNA but did not exhibit prominent immune activation, suggesting that BTK may be a host modifier, dampening immune responses to microbial translocation. These data reveal a mechanism for chronic immune activation in CVID and potential therapeutic strategies to modify the clinical outcomes of this disease.

[Measurement of angiotensin-I-converting enzyme in the blood: help for method validation]. / Dosage de l'enzyme de conversion de l'angiotensine-I sanguine : aide à la validation de méthode.

Blood angiotensin-converting enzyme (ACE) assay is now realized by the determination of enzyme activity on synthetic substrate, mostly furylacryloyl-phenylalanyl-L-glycyl-L-glycine (FAPGG). The matrix can be serum or heparin-plasma, with or without a separator; the assay developed on serum or plasma is not adapted to other matrix such as cerebrospinal fluid where the ACE activity is much lower. This assay has been adapted on a number of automated biochemistry analyzers with the specifications of the supplier of reagents, sometimes with modification of volumes or times for analysis. Samples can be stored at +4̊C for at least for one week, freezing at -20̊C is possible but refreezing is not advised. The assay is linear from 10 to 200 UI/L. Fidelity is excellent after calibration of the assay. Accuracy can be calculated from IQA and EQA results, and the analytical uncertainty is between 2% and 5% in function of the serum ACE value. Usual values will be soon available from studies on age brackets and sex, because ACE activity seems to be more elevated in boys during adolescence. At signature, it is interesting to have medical information on the diagnosis of sarcoidosis or its treatment including ACE inhibitors as a proof of intake; we can give a commentary on elevation of serum ACE activity from other causes than sarcoidosis and the causes for low activities.

Derangement of Metabolic and Lysosomal Gene Profiles in Response to Dexamethasone Treatment in Sarcoidosis.

Glucocorticoids (GCs) play a central role in modulation of inflammation in various diseases, including respiratory diseases such as sarcoidosis. Surprisingly, the specific anti-inflammatory effects of GCs on different myeloid cells especially in macrophages remain poorly understood. Sarcoidosis is a systemic granulomatous disease of unknown etiology that occurs worldwide and is characterized by granuloma formation in different organs. Alveolar macrophages play a role in sarcoidosis granuloma formation and progressive lung disease. The goal of the present study is to identify the effect of GCs on transcriptomic profiles and the cellular pathways in sarcoidosis alveolar macrophages and their corresponding blood myeloid cells. We determined and compared the whole transcriptional signatures of alveolar macrophages from sarcoidosis patients and blood CD14+ monocytes of the same subjects in response to in vitro treatment with dexamethasone (DEX) via RNA-sequencing. In response to DEX, we identified 2,834 genes that were differentially expressed in AM. Predominant pathways affected were as following: metabolic pathway (FDR = 4.1 × 10-10), lysosome (FDR = 6.3 × 10-9), phagosome (FDR = 3.9 × 10-5). The DEX effect on AMs is associated with metabolic derangements involving glycolysis, oxidative phosphorylation and lipid metabolisms. In contrast, the top impacted pathways in response to DEX treatment in blood CD14+ monocytes were as following; cytokine-cytokine receptor interaction (FDR = 6 × 10-6) and transcriptional misregulation in cancer (FDR = 1 × 10-4). Pathways similarly affected in both cell types were genes involved in lysosomes, cytoskeleton and transcriptional misregulation in cancer. These data suggest that the different effects of DEX on AMs and peripheral blood monocytes are partly dictated by lineage specific transcriptional programs and their physiological functions.

Physicochemical predictors of Multi-Walled Carbon Nanotube-induced pulmonary histopathology and toxicity one year after pulmonary deposition of 11 different Multi-Walled Carbon Nanotubes in mice.

Multi-walled carbon nanotubes (MWCNT) are widely used nanomaterials that cause pulmonary toxicity upon inhalation. The physicochemical properties of MWCNT vary greatly, which makes general safety evaluation challenging to conduct. Identification of the toxicity-inducing physicochemical properties of MWCNT is therefore of great importance. We have evaluated histological changes in lung tissue 1 year after a single intratracheal instillation of 11 well-characterized MWCNT in female C57BL/6N BomTac mice. Genotoxicity in liver and spleen was evaluated by the comet assay. The dose of 54 µg MWCNT corresponds to three times the estimated dose accumulated during a work life at a NIOSH recommended exposure limit (0.001 mg/m3 ). Short and thin MWCNT were observed as agglomerates in lung tissue 1 year after exposure, whereas thicker and longer MWCNT were detected as single fibres, suggesting biopersistence of both types of MWCNT. The thin and entangled MWCNT induced varying degree of pulmonary inflammation, in terms of lymphocytic aggregates, granulomas and macrophage infiltration, whereas two thick and straight MWCNT did not. By multiple regression analysis, larger diameter and higher content of iron predicted less histopathological changes, whereas higher cobalt content significantly predicted more histopathological changes. No MWCNT-related fibrosis or tumours in the lungs or pleura was found. One thin and entangled MWCNT induced increased levels of DNA strand breaks in liver; however, no physicochemical properties could be related to genotoxicity. This study reveals physicochemical-dependent difference in MWCNT-induced long-term, pulmonary histopathological changes. Identification of diameter size and cobalt content as important for MWCNT toxicity provides clues for designing MWCNT, which cause reduced human health effects following pulmonary exposure.

Adjunct antibody administration with standard treatment reduces relapse rates in a murine tuberculosis model of necrotic granulomas.

Matrix metalloproteinase (MMP)-9 is a zinc-dependent protease associated with early immune responses to Mycobacterium tuberculosis infection, macrophage recruitment and granuloma formation. We evaluated whether adjunctive inhibition of MMP-9 could improve the response to standard TB treatment in a mouse model that develops necrotic lesions. Six weeks after an aerosol infection with M. tuberculosis, C3HeB/FeJ mice received standard TB treatment (12 weeks) comprising rifampin, isoniazid and pyrazinamide alone or in combination with either anti-MMP-9 antibody, etanercept (positive control) or isotype antibody (negative control) for 6 weeks. Anti-MMP-9 and the isotype control had comparable high serum exposures and expected terminal half-life. The relapse rate in mice receiving standard TB treatment was 46.6%. Compared to the standard TB treatment, relapse rates in animals that received adjunctive treatments with anti-MMP-9 antibody or etanercept were significantly decreased to 25.9% (P = 0.006) and 29.8% (P = 0.019) respectively, but were not different from the arm that received the isotype control antibody (25.9%). Immunostaining demonstrated localization of MMP-9 primarily in macrophages in both murine and human lung tissues infected with M. tuberculosis, suggesting the importance of MMP-9 in TB pathogenesis. These data suggest that the relapse rates in M. tuberculosis-infected mice may be non-specifically improved by administration of antibodies in conjunction with standard TB treatments. Future studies are needed to evaluate the mechanism(s) leading to improved outcomes with adjunctive antibody treatments.

TNF-α blockade impairs in vitro tuberculous granuloma formation and down modulate Th1, Th17 and Treg cytokines.

Tuberculosis (TB) is a granulomatous disease that has affected humanity for thousands of years. The production of cytokines, such as IFN-γ and TNF-α, is fundamental in the formation and maintenance of granulomas and in the control of the disease. Recently, the introduction of TNF-α-blocking monoclonal antibodies, such as Infliximab, has brought improvements in the treatment of patients with chronic inflammatory diseases, but this treatment also increases the risk of reactivation of latent tuberculosis. Our objective was to analyze, in an in vitro model, the influence of Infliximab on the granulomatous reactions and on the production of antigen-specific cytokines (TNF-α, IFN-γ, IL-12p40, IL-10 and IL-17) from beads sensitized with soluble Bacillus Calmette-Guérin (BCG) antigens cultured in the presence of peripheral blood mononuclear cells (PBMC) from TB patients. We evaluated 76 individuals, with tuberculosis active, treated and subjects with positive PPD. Granuloma formation was induced in the presence or absence of Infliximab for up to 10 days. The use of Infliximab in cultures significantly blocked TNF-α production (p <0.05), and led to significant changes in granuloma structure, in vitro, only in the treated TB group. On the other hand, there was a significant reduction in the levels of IFN-γ, IL-12p40, IL-10 and IL-17 after TNF-α blockade in the three experimental groups (p <0.05). Taken together, our results demonstrate that TNF-α blockade by Infliximab directly influenced the structure of granuloma only in the treated TB group, but negatively modulated the production of Th1, Th17 and regulatory T cytokines in the three groups analyzed.

TARC expression in the circulation and cutaneous granulomas correlates with disease severity and indicates Th2-mediated progression in patients with sarcoidosis.

BACKGROUND: Sarcoidosis is a systemic disorder characterized by the accumulation of lymphocytes and monocyte/macrophage lineage cells that results in the formation of non-caseating granulomas. Thymus- and activation-regulated chemokine (TARC)/CCL17 is an important chemokine in the amplification of Th2 responses, which are achieved by recruiting CCR4-expressing CD4+ T lymphocytes. TARC concentrations are known to increase in the serum of sarcoidosis patients; however, its role in the assessment of severity and prognosis of sarcoidosis remains unknown. The objective of this study is to elucidate the role of TARC in sarcoidosis by investigating its expression in peripheral blood and at inflammatory sites. We also examined its relationship with clinical features. METHODS: Serum levels of TARC, soluble interleukin 2 receptor, angiotensin-converting enzyme, and lysozyme were measured in 82 sarcoidosis patients. The Th1 and Th2 balance in circulating CD4+ T cells was evaluated by flow cytometry. The immunohistochemical staining of TARC and CCR4 was performed in order to identify the source of TARC in affected skin tissues. RESULTS: TARC serum levels were elevated in 78% of patients and correlated with disease severity. The percentage of CCR4+ cells and the CCR4+/CXCR3+ cell ratios were significantly higher in sarcoidosis patients than in normal subjects (P = 0.002 and P = 0.015, respectively). Moreover, TARC was expressed by monocyte/macrophage lineage cells within granulomas. The abundancy as well as distribution of TARC staining correlated with its serum levels. CONCLUSIONS: The present results suggest that elevations in TARC drive an imbalanced Th2- weighted immune reaction and might facilitate prolonged inflammatory reactions in sarcoidosis.

Anticentromere antibody positive Ackerman's Syndrome with Granulomatous Anterior Uveitis.

Ackerman's Syndrome or Intersticial Granulomatous Dermatitis with Arthritis has been an issue of increasing number of reports in the last decade which had focused its heterogeneous cutaneous and rheumatologic expression besides the initial manifestations reported by Ackerman and his group. Granulomatosis anterior uveitis has not been previously described. Some patients are reported to have positive autoantibodies but association with anticentromere antibodies has not been previously described as well, to our knowledge. We report a new case of Ackerman Syndrome with cutaneous, articular and ocular involvement with positive anticentromere antibodies successfully treated with systemic steroids, methotrexate, hydroxychloroquine and cyclosporine. The ocular involvement and the association of anticentromere antibodies lead us to hypothesize that constellation of symptoms and autoimmune mechanisms of this uncommon multisystemic syndrome are yet to be clarified.

V-Loc™ 180 suture in total laparoscopic hysterectomy: a retrospective study comparing Polysorb to barbed suture used for vaginal cuff closure.

OBJECTIVE: Sutures in laparoscopy have been extensively developed in recent years. In this study, we assessed differences between Polysorb™ braided absorbable suture (CL-914) and V-Loc™ barbed suture (V-Loc 180) used in vaginal cuff closure during laparoscopic hysterectomy from various aspects. STUDY DESIGN: This paper presented a prospective cohort study of 490 consecutive women underwent total laparoscopic hysterectomy (TLH) performed between January 2013 and September 2015 applying identical procedure technique, with cuff closure approaches selected by surgeons. Data collected included operative time, estimated blood loss, postoperative changes in body temperature, perioperative routine blood parameters changes, total average hospital stay, and postoperative hospital stay. In addition, short-term (at least 6 months) outcomes of vaginal cuff granulomatous (VCG) between the two suture approaches were compared. RESULTS: Between groups, statistical differences were detected in operative duration, estimated blood loss, total and postoperative hospital stay, WBC, neutrophil counts and Hb in postoperative routine blood parameters; while there were no significant differences in other data (all P>0.05). Postoperative routine blood parameters in each group: compared to preoperative baseline, in Group 1, WBC, N increased (P<0.05), while RBC, Hb decreased (P<0.05). In Group 2, same tendency in WBC, N and Hb was indicated, but RBC increased with no significant difference. In both groups, vaginal cuff healing was well, with no dehiscence. VCG occurred more often in women used CL-914 than women applied V-Loc 180. CONCLUSIONS: V-Loc barbed suture can be used for vaginal cuff closure during TLH due to advantages such as less operative duration and blood loss, shorter postoperative and total hospital stay, and reduced VCG formation six months after TLH.

Innate immunity in sarcoidosis pathobiology.

PURPOSE OF REVIEW: The immunopathogenesis of sarcoidosis is considered to involve contributions from both adaptive and innate immune responses. Although the identification of adaptive responses to candidate pathogenic antigens will elucidate mechanisms that regulate inflammation in sarcoidosis, innate mechanisms likely represent the 'missing link' to the initiation, maintenance, and resolution of noncaseating granulomatous inflammation, the hallmark feature of sarcoidosis. Furthermore, environments that expose patients to candidate pathogenic antigens for sarcoidosis also provide opportunities for engagement with innate ligands. RECENT FINDINGS: Several studies have identified enhanced responsiveness via pattern recognition receptor pathways, potentiating the local induction of cytokines relevant to granulomatous inflammation, such as through stimulation of nucleotide-binding oligomerization domain-like receptor and Toll-like receptor pathways. These pathways contribute to the inherent properties of granulomas including, in some cases, persistent localization of pathogens and antigen. Serum amyloid A has been identified to be abundant in sarcoidosis tissues, and this promiscuous host protein can serve as an innate ligand to regulate experimental granulomatous inflammation. Nascent evidence supports a potential role for alternatively activated macrophages to direct histopathological outcomes in sarcoidosis. SUMMARY: Innate pathways deserve further investigation as potential therapeutic targets for inhibiting granuloma formation in sarcoidosis.

Polyacrylate/nanosilica causes pleural and pericardial effusion, and pulmonary fibrosis and granuloma in rats similar to those observed in exposed workers.

Nanomaterials offer great benefit as well as potential damage to humans. Workers exposed to polyacrylate coatings have pleural effusion, pericardial effusion, and pulmonary fibrosis and granuloma, which are thought to be related to the high exposure to nanomaterials in the coatings. The study aimed to determine whether polyacrylate/silica nanoparticles cause similar toxicity in rats, as observed in exposed workers. Ninety male Wistar rats were randomly divided into five groups with 18 rats in each group. The groups included the saline control group, another control group of polyacrylate only, and low-, intermediate-, and high-dose groups of polyacrylate/nanosilica with concentrations of 3.125, 6.25, and 12.5 mg/kg. Seventy-five rats for the 1-week study were terminated for scheduled necropsy at 24 hours, 3 days, and 7 days postintratracheal instillation. The remaining 15 rats (three males/group) had repeated ultrasound and chest computed tomography examinations in a 2-week study to observe the pleural and pericardial effusion and pulmonary toxicity. We found that polyacrylate/nanosilica resulted in pleural and pericardial effusions, where nanosilica was isolated and detected. Effusion occurred on day 3 and day 5 post-administration of nanocomposites in the 6.25 and 12.5 mg/kg groups, it gradually rose to a maximum on days 7-10 and then slowly decreased and disappeared on day 14. With an increase in polyacrylate/nanosilica concentrations, pleural effusion increased, as shown by ultrasonographic qualitative observations. Pulmonary fibrosis and granuloma were also observed in the high-dose polyacrylate/nanosilica group. Our study shows that polyacrylate/nanosilica results in specific toxicity presenting as pleural and pericardial effusion, as well as pulmonary fibrosis and granuloma, which are almost identical to results in reported patients. These results indicate the urgent need and importance of nanosafety and awareness of toxicity of polyacrylate/nanosilica.

Serum IgE Levels: An Effective Diagnostic Tool for Evaluation and Monitoring of Orbital Fungal Granuloma.

OBJECTIVE: To determine the changes in IgE levels in diagnosis and postoperative monitoring of orbital fungal granuloma. STUDY DESIGN: Descriptive analytical study. PLACE AND DURATION OF STUDY: Al-Shifa Trust Eye Hospital, Rawalpindi, Pakistan, from July 2012 to June 2013. METHODOLOGY: Cases with clinically high index of suspicion for orbital fungal granuloma and 50 healthy volunteers were inducted as control subjects. Patients with recurrent cases of orbital fungal granuloma, previous orbital surgery, and those with very low clinical suspicion of fungal granuloma were excluded. Total serum IgE level (IU/ml), eosinophil counts and skin prick test were performed in all subjects. Independent t-test was used for comparison of healthy volunteers and patients with biopsy proven orbital fungal granuloma. Repeated measures ANOVA was used for comparing the preoperative and postoperative total serum IgE level of patients with biopsy proven fungal granuloma. RESULTS: The mean total serum IgE level for the healthy volunteers was 208.82 ±41.43 IU/ml. The mean pre-operative IgE value of histologically confirmed cases of fungal granuloma was 1613.72 ±282.83 IU/ml. The total serum IgE level gradually declined after surgery and anti-fungal treatment. The mean serum IgE level 3, 6 and 9 months postoperatively were 1039.48 ±308.40, 568.77 ±162.01 and 224.92 ±51.55 IU/ml respectively. These tests showed that the drop in IgE level in cases of fungal granuloma with treatment was statistically significant (p < 0.001). CONCLUSION: Total serum IgE level can be used as a reliable diagnostic and postoperative monitoring tool in orbital fungal granuloma.

Serum, liver, and lung levels of the major extracellular matrix components at the early stage of BCG-induced granulomatosis depending on the infection route.

Experiments on the model of mouse BCG-induced granulomatous showed that the content of glycosaminoglycans and proteoglycans in the extracellular matrix of the liver and lungs are changed at the early stages of inflammation (days 3 and 30 postinfection) before cell destruction in the organs begins. This is related to degradation of extracellular matrix structures. Their high content in the blood and interstitium probably contributes to the formation of granulomas, fibroblast proliferation and organ fibrosis. These processes depend on the infection route that determines different conditions for generalization of the inflammation process. Intravenous method of vaccine injection is preferable to use when designing the experiments simulating tuberculosis granulomatosis, especially for the analysis of its early stages.

Raised numbers of IgG4-positive plasma cells are a common histopathological finding in orbital xanthogranulomatous disease.

PURPOSE: To determine the relation of orbital xanthogranuloma with IgG4-related disease. METHODS: Retrospective consecutive case series over a period of 25 years. We searched our charts for histologically confirmed orbital xanthogranuloma. Patient files were reviewed for clinical and follow up data including presence or absence of systemic non-ophthalmic manifestations of IgG4 related disease. Slides were re-examined and histopathological classification was re-assessed. Sixteen cases of orbital xanthogranuloma were evaluated. Immunohistochemical stains for IgG and IgG4 were performed. Positive immunohistochemical staining required increased IgG4-positive plasma cells in the involved tissues scored as >50 per high-power field, with IgG4/IgG ratio >0.40. RESULTS: According to the criteria described above 8/16 (50%) cases showed increased numbers of IgG4-positive plasma cells in the specimens. Two of these patients may have had signs of systemic disease. CONCLUSION: Raised numbers of IgG4-positive plasma cells are a common finding in histopathological specimens of xanthogranulomatous disease of the orbit and are often not indicative for IgG4 related systemic disease.

Granulomatous liver involvement in a child with systemic lupus erythematosus: a case report and review of the literature.

Systemic lupus erythematosus (SLE) is uncommon in young children and unusual in infancy. Although a variety of liver pathologies have been reported in SLE, presentation of this disease with granulomatous liver involvement is very rare. In this article, for the first time, we report an infant girl presenting with unexplained hepatosplenomegaly and non-necrotizing granulomatous liver involvement at the age of six months who later developed pancytopenia and proteinuria and was finally diagnosed with SLE at the age of three years. Therefore, we suggest that SLE could be considered as one of the possible differential diagnoses when infants or children present with unexplained granulomatous liver involvement.

microRNAs derived from circulating exosomes as noninvasive biomarkers for screening and diagnosing lung cancer.

INTRODUCTION: Lung cancer is the highest cause of mortality among tumor pathologies worldwide. There are no validated techniques for an early detection of pulmonary cancer lesions other than low-dose helical computed tomography scan. Unfortunately, this method has some negative effects. Recent studies have laid the basis for development of exosomes-based techniques to screen/diagnose lung cancers. As the isolation of circulating exosomes is a minimally invasive procedure, this technique opens new possibilities for diagnostic applications. METHODS: We used a first set of 30 plasma samples from as many patients, including 10 patients affected by lung adenocarcinomas, 10 with lung granulomas, and 10 healthy smokers matched for age and sex as negative controls. Wide-range microRNAs analysis (742 microRNAs) was performed by quantitative real time polymerase chain reaction. Data were compared on the basis of lesion characteristics, using WEKA software for statistics and modeling. Subsequently, selected microRNAs were evaluated on an independent larger group of samples (105 specimens: 50 lung adenocarcinomas, 30 lung granulomas, and 25 healthy smokers). RESULTS: This analysis led to the selection of four microRNAs to perform a screening test (miR-378a, miR-379, miR-139-5p, and miR-200b-5p), useful to divide population into two groups: nodule (lung adenocarcinomas + carcinomas) and non-nodule (healthy former smokers). Six microRNAs (miR-151a-5p, miR-30a-3p, miR-200b-5p, miR-629, miR-100, and miR-154-3p) were selected for a second test on the nodule population to discriminate between lung adenocarcinoma and granuloma. CONCLUSIONS: The screening test showed 97.5% sensitivity, 72.0% specificity, and area under the curve receiver operating characteristic of 90.8%. The diagnostic test had 96.0% sensitivity, 60.0% specificity, and area under the curve receiver operating characteristic of 76.0%. Further evaluation is needed to confirm the predictive power of these models on larger cohorts of samples.