Standardization of flow cytometry and cell sorting to enable a transcriptomic analysis in a multi-site sarcoidosis study.

The contribution and regulation of various CD4+ T cell lineages that occur with remitting vs progressive courses in sarcoidosis are poorly understood. We developed a multiparameter flow cytometry panel to sort these CD4+ T cell lineages followed by measurement of their functional potential using RNA-sequencing analysis at six-month intervals across multiple study sites. To obtain good quality RNA for sequencing, we relied on chemokine receptor expression to identify and sort lineages.

Autoantibody profile in sarcoidosis, analysis from the GRADS sarcoidosis cohort.

Background: Sarcoidosis, a multi-systemic granulomatous disease, is a predominantly T-cell disease but evidence for a role for humoral immunity in disease pathogenesis is growing. Utilizing samples from the Genomic Research in Alpha-1 anti-trypsin Deficiency and Sarcoidosis (GRADS) study, we examined the prevalence of autoantibodies in sarcoidosis patients with pulmonary-only and extra-pulmonary organ involvement compared to normal controls.

Study Design And Methods: We analyzed serum samples from sarcoidosis patients who participated in the GRADS study utilizing an autoantigen microarray platform for both IgM and IgG antibodies.

Defining CD4 T helper and T regulatory cell endotypes of progressive and remitting pulmonary sarcoidosis (BRITE): protocol for a US-based, multicentre, longitudinal observational bronchoscopy study.

Introduction: Sarcoidosis is a multiorgan granulomatous disorder thought to be triggered and influenced by gene-environment interactions. Sarcoidosis affects 45-300/100 000 individuals in the USA and has an increasing mortality rate. The greatest gap in knowledge about sarcoidosis pathobiology is a lack of understanding about the underlying immunological mechanisms driving progressive pulmonary disease.

Circulating exosomal microRNA expression patterns distinguish cardiac sarcoidosis from myocardial ischemia.

Objective: Cardiac sarcoidosis is difficult to diagnose, often requiring expensive and inconvenient advanced imaging techniques. Circulating exosomes contain genetic material, such as microRNA (miRNA), that are derived from diseased tissues and may serve as potential disease-specific biomarkers. We thus sought to determine whether circulating exosome-derived miRNA expression patterns would distinguish cardiac sarcoidosis (CS) from acute myocardial infarction (AMI).

DNA Methylation Changes in Lung Immune Cells Are Associated with Granulomatous Lung Disease.

Epigenetic marks are likely to explain variability of response to antigen in granulomatous lung disease. The objective of this study was to identify DNA methylation and gene expression changes associated with chronic beryllium disease (CBD) and sarcoidosis in lung cells obtained by BAL. BAL cells from CBD (n = 8), beryllium-sensitized (n = 8), sarcoidosis (n = 8), and additional progressive sarcoidosis (n = 9) and remitting (n = 15) sarcoidosis were profiled on the Illumina 450k methylation and Affymetrix/Agilent gene expression microarrays.

5-Aminosalicylic Acid Modulates the Immune Response in Chronic Beryllium Disease Subjects.

Introduction: Chronic beryllium disease (CBD) is characterized by accumulation of macrophages and beryllium-specific CD4 T cells that proliferate and produce Th1 cytokines. 5-Amino salicylic acid (5-ASA) is currently used to treat inflammatory bowel disease and has both antioxidant and anti-inflammatory actions. We hypothesized that 5-ASA may be a beneficial therapeutic in CBD.

Beryllium-induced lung disease exhibits expression profiles similar to sarcoidosis.

A subset of beryllium-exposed workers develop beryllium sensitisation (BeS) which precedes chronic beryllium disease (CBD). We conducted an in-depth analysis of differentially expressed candidate genes in CBD.We performed Affymetrix GeneChip 1.

The effect of an oral anti-oxidant, N-Acetyl-cysteine, on inflammatory and oxidative markers in pulmonary sarcoidosis.

Background: Oxidative stress (OS) has been shown to play a role in the pathogenesis of sarcoidosis and previous studies have shown that anti-oxidants can reduce markers of oxidative stress and inflammation in the peripheral blood of sarcoidosis subjects. We investigated the effect of N-Acetyl-Cysteine (NAC) on oxidative stress and inflammatory markers in the lungs of sarcoidosis patients.

Methods: We randomized 11 sarcoidosis subjects to active therapy and 3 to placebo for 8 weeks in a double blinded study.

Beryllium increases the CD14(dim)CD16+ subset in the lung of chronic beryllium disease.

CD14dimCD16+ and CD14brightCD16+ cells, which compose a minor population of monocytes in human peripheral blood mononuclear cells (PBMC), have been implicated in several inflammatory diseases. The aim of this study was to investigate whether this phenotype was present as a subset of lung infiltrative alveolar macrophages (AMs) in the granulomatous lung disease, chronic beryllium disease (CBD). The monocytes subsets was determined from PBMC cells and bronchoalveolar lavage (BAL) cells from CBD, beryllium sensitized Non-smoker (BeS-NS) and healthy subjects (HS) using flow cytometry.

Sulfasalazine and mesalamine modulate beryllium-specific lymphocyte proliferation and inflammatory cytokine production.

Occupational exposure to beryllium (Be) results in Be sensitization (BeS) that can progress to pulmonary granulomatous inflammation associated with chronic Be disease (CBD). Be-specific lymphocytes are present in the blood of patients with BeS and in the blood and lungs of patients with CBD. Sulfasalazine and its active metabolite, mesalamine, are clinically used to ameliorate chronic inflammation associated with inflammatory bowel disease.

Modulation of lymphocyte proliferation by antioxidants in chronic beryllium disease.

Rationale: Occupational exposure to beryllium (Be) can result in chronic granulomatous inflammation characterized by the presence of Be-specific CD4+ T cells. Studies show that oxidative stress plays a role in the pathogenesis of chronic inflammatory disorders.

Objectives: We hypothesized that Be-induced oxidative stress modulates the proliferation of Be-specific CD4+ T cells.

TNF polymorphism and bronchoalveolar lavage cell TNF-alpha levels in chronic beryllium disease and beryllium sensitization.

Background: Beryllium stimulates TNF-alpha from chronic beryllium disease (CBD) bronchoalveolar lavage (BAL) cells.

Objective: We sought to relate TNF polymorphisms to beryllium-stimulated TNF-alpha production, to the development of CBD, and to the risk of more severe CBD over time.

Methods: We recruited 147 patients with CBD, 112 beryllium-sensitized subjects, and 323 control subjects; genotyped 5 TNF promoter polymorphisms; and measured beryllium-stimulated and unstimulated BAL cell TNF-alpha production from a subset of subjects.

Beryllium-induced TNF-alpha production is transcription-dependent in chronic beryllium disease.

Beryllium (Be)-antigen presentation to Be-specific CD4(+) T cells from the lungs of patients with chronic beryllium disease (CBD) results in T cell proliferation and TNF-alpha secretion. We tested the hypothesis that Be-induced, CBD bronchoalveolar lavage (BAL) T cell, transcription-dependent, TNF-alpha secretion was accompanied by specific transcription factor upregulation. After 6 h of Be stimulation, CBD BAL cells produced a median of 883 pg/ml TNF-alpha (range, 608-1,275 pg/ml) versus 198 pg/ml (range, 116-245 pg/ml) by unstimulated cells.

Beryllium-induced tumor necrosis factor-alpha production by CD4+ T cells is mediated by HLA-DP.

Beryllium (Be) presentation to CD4+ T cells from patients with chronic beryllium disease (CBD) results in T cell activation, and these Be-specific CD4+ T cells undergo clonal proliferation and T-helper 1-type cytokine production. In exposed workers, genetic susceptibility to this granulomatous disorder is associated with particular HLA-DPB1 alleles. We hypothesized that these HLA-DP molecules could mediate Be-stimulated tumor necrosis factor-alpha (TNF-alpha) messenger RNA (mRNA) and protein production.