Leukocyte toll-like receptor expression in pathergy positive and negative Behçet's disease patients

OBJECTIVES: To investigate whether the auto-inflammatory nature and the pathergic reaction in Behçet's disease (BD) are driven by a disturbed toll-like receptor (TLR) response. METHODS: We compared both TLR expression by flow-cytometry and TLR response by stimulation assay in 18 BD patients (both pathergy positive and pathergy negative) with 15 healthy controls. RESULTS: Expression of TLR1 and 2 was significantly elevated in B-lymphocytes of BD patients compared with healthy controls. TLR1, 2 and 4 were significantly more highly expressed in both CD4+ and CD8+ T-lymphocytes of BD patients. Granulocytes of BD patients displayed significantly higher expression of TLR1, 2, 4 and 6. TLR2, 4 and 5 expression was significantly increased on classical monocytes of BD patients. Intermediate monocytes of BD patients showed an increase in expression of TLR2. Furthermore, TLR2 and 5 were significantly more highly expressed in non-classical monocytes of BD patients. In pathergy positive patients, TLR5 was even more highly expressed compared with pathergy negative patients on B- and T-lymphocytes and granulocytes. Furthermore, TLR2 and 5 showed an elevated TNF-α response to stimulation with their cognate ligands. CONCLUSION: Immune cells of BD patients overexpress TLR1, 2, 4, 5 and 6. Furthermore, after stimulation of TLR2 and 5, BD patients demonstrate a more potent TNF-α response. Although this is a small cohort, in the pathergy positive patients, TLR5 expression is even further augmented, suggesting that a microbial (flagellin) or damage (HMGB1) associated signal may trigger the exaggerated immune response that is characteristic for the pathergy phenomenon in BD. In conclusion, these results point to an exaggerated TLR response in the auto-inflammatory nature of BD

B-cell dysregulation in Crohn's disease is partially restored with infliximab therapy

Background: B-cell depletion can improve a variety of chronic inflammatory diseases, but does not appear beneficial for patients with Crohn's disease. Objective: To elucidate the involvement of B cells in Crohn's disease, we here performed an 'in depth' analysis of intestinal and blood B-cells in this chronic inflammatory disease. Methods: Patients with Crohn's disease were recruited to study B-cell infiltrates in intestinal biopsies (n = 5), serum immunoglobulin levels and the phenotype and molecular characteristics of blood B-cell subsets (n = 21). The effects of infliximab treatment were studied in 9 patients. Results: Granulomatous tissue showed infiltrates of B lymphocytes rather than Ig-secreting plasma cells. Circulating transitional B cells and CD21low B cells were elevated. IgM memory B cells were reduced and natural effector cells showed decreased replication histories and somatic hypermutation (SHM) levels. In contrast, IgG and IgA memory B cells were normally present and their Ig gene transcripts carried increased SHM levels. The numbers of transitional and natural effector cells were normal in patients who responded clinically well to infliximab. Conclusions: B cells in patients with Crohn's disease showed signs of chronic stimulation with localization to granulomatous tissue and increased molecular maturation of IgA and IgG. Therapy with TNFα-blockers restored the defect in IgM memory B-cell generation and normalized transitional B-cell levels, making these subsets candidate markers for treatment monitoring. Together, these results suggest a chronic, aberrant B-cell response in patients with Crohn's disease, which could be targeted with new therapeutics that specifically regulate B-cell function

Characteristics of COVID-19 infection and antibody formation in patients known at a tertiary immunology department

Background: Knowledge about COVID-19 infections is expanding, although knowledge about the disease course and antibody formation in patients with an auto-immune disease or immunodeficiency is not fully unraveled yet. It could be hypothesized that immunodeficient patients, due to immunosuppressive drugs or their disease, have a more severe disease course due to their immunocompromised state. However, it could also be hypothesized that some of the immunosuppressive drugs protect against a hyperinflammatory state. Methods: We collected data on the incidence of COVID-19, disease course and SARS-CoV-2 antibody formation in COVID-19 positive patients in a cohort of patients (n ​= ​4497) known at the Clinical Immunology outpatient clinic in a tertiary care hospital in the Netherlands. Results: In the first six months of the pandemic, 16 patients were identified with COVID-19, 14 by nasal swab PCR, and 2 pa

B-Cell Dysregulation in Crohn's Disease Is Partially Restored with Infliximab Therapy

<div><p>Background</p><p>B-cell depletion can improve a variety of chronic inflammatory diseases, but does not appear beneficial for patients with Crohn’s disease.</p><p>Objective</p><p>To elucidate the involvement of B cells in Crohn’s disease, we here performed an ‘in depth’ analysis of intestinal and blood B-cells in this chronic inflammatory disease.</p><p>Methods</p><p>Patients with Crohn’s disease were recruited to study B-cell infiltrates in intestinal biopsies (n = 5), serum immunoglobulin levels and the phenotype and molecular characteristics of blood B-cell subsets (n = 21). The effects of infliximab treatment were studied in 9 patients.</p><p>Results</p><p>Granulomatous tissue showed infiltrates of B lymphocytes rather than Ig-secreting plasma cells. Circulating transitional B cells and CD21^low B cells were elevated. IgM memory B cells were reduced and natural effector cells showed decreased replication histories and somatic hypermutation (SHM) levels. In contrast, IgG and IgA memory B cells were normally present and their Ig gene transcripts carried increased SHM levels. The numbers of transitional and natural effector cells were normal in patients who responded clinically well to infliximab.</p><p>Conclusions</p><p>B cells in patients with Crohn’s disease showed signs of chronic stimulation with localization to granulomatous tissue and increased molecular maturation of IgA and IgG. Therapy with TNFα-blockers restored the defect in IgM memory B-cell generation and normalized transitional B-cell levels, making these subsets candidate markers for treatment monitoring. Together, these results suggest a chronic, aberrant B-cell response in patients with Crohn’s disease, which could be targeted with new therapeutics that specifically regulate B-cell function.</p></div

IgA and IgG subclass analysis.

<p><b>A,</b> Schematic representation of the constant region of the human IGH locus. <b>B,</b> Distribution of IgA and IgG subclass use in switched transcripts of healthy controls and patients with Crohn’s disease. Total numbers of analyzed sequences are indicated in the middle of the plots. χ² Test was performed to analyze differences in distributions. <b>C,</b> Combined <i>IGHV</i> mutation frequencies in IgA and IgG transcripts in patients and controls (total numbers of sequences indicated between brackets). Grey dots represent unique sequences; red lines represent median values. Statistical analysis was performed with the Mann-Whitney test; *, P<0.05; **, P<0.01; ***, P<0.001.</p

Replication history and SHM levels in <i>IGHV</i> genes of natural effector B cells.

<p><b>A,</b> Replication history of naïve and natural effector B cells as assessed using the KREC assay [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160103#pone.0160103.ref025" target="_blank">25</a>]. <b>B,</b> <i>IGHV</i> mutation frequencies in rearranged <i>IGH</i> genes of natural effector B cells in patients and controls (total numbers of sequences indicated between brackets). Grey dots represent unique sequences; red lines represent median values. <b>C</b>, Selection for replacement mutation in IGHV-CDR (red line) and IGHV-FR regions (blue lines) as determined with the BASELINe program [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160103#pone.0160103.ref028" target="_blank">28</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160103#pone.0160103.ref029" target="_blank">29</a>]. Solid lines represent patients; dashed lines represent healthy controls. Selection Strengths >0 indicate positive selection. <b>D,</b> IGH-CDR3 size distributions. All individual sizes are indicated as grey dots, red lines representing median values. The dashed line represents median values for centroblasts and centrocytes. Sorted cells were analyzed from patients 15, 16, 18 and 19. Controls were published previously [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160103#pone.0160103.ref030" target="_blank">30</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160103#pone.0160103.ref031" target="_blank">31</a>]. Statistical analysis was performed with the Mann-Whitney test; *, P<0.05; **, P<0.01; ***, P<0.001; ****, P<0.0001.</p

Clinical and basic immunological characteristics of patients with Crohn’s disease.

<p>Clinical and basic immunological characteristics of patients with Crohn’s disease.</p

Composition of the blood B-cell compartment in patients with Crohn’s disease.

<p><b>A.</b> Schematic overview of peripheral B-cell subsets. <b>B.</b> Average numbers of blood B cell subsets of 21 patients affected with Crohn’s disease (black bars) and 28 healthy controls (light grey bars). <b>C.</b> Distribution of IgM, IgA and IgG within CD21^low in patients and controls <b>D.</b> Total CD21^low B cells in relation to disease duration. Statistical analyses were performed with the Mann-Whitney test or Spearman correlation; *, P<0.05; **, P<0.01.</p