scholarly journals Novel mutations in the WFS1 gene are associated with Wolfram syndrome and systemic inflammation

2021 ◽  
Author(s):  
Eleonora Panfili ◽  
Giada Mondanelli ◽  
Ciriana Orabona ◽  
Maria L Belladonna ◽  
Marco Gargaro ◽  
...  
Keyword(s):  
Er Stress ◽  
Mononuclear Cells ◽  
Autosomal Recessive Disorder ◽  
Wolfram Syndrome ◽  
Therapeutic Interventions ◽  
T Helper 17 ◽  
Gene Encoding ◽  
Peripheral Blood Mononuclear ◽  
Inflammatory State ◽  
Wfs1 Gene

Abstract Mutations in the WFS1 gene, encoding wolframin (WFS1), cause endoplasmic reticulum (ER) stress and are associated with a rare autosomal-recessive disorder known as Wolfram syndrome (WS). WS is clinically characterized by childhood-onset diabetes mellitus, optic atrophy, deafness, diabetes insipidus and neurological signs. We identified two novel WFS1 mutations in a patient with WS, namely, c.316-1G > A (in intron 3) and c.757A > T (in exon 7). Both mutations, located in the N-terminal region of the protein, were predicted to generate a truncated and inactive form of WFS1. We found that although the WFS1 protein was not expressed in peripheral blood mononuclear cells (PBMCs) of the proband, no constitutive ER stress activation could be detected in those cells. In contrast, WS proband’s PBMCs produced very high levels of proinflammatory cytokines (i.e. TNF-α, IL-1β, and IL-6) in the absence of any stimulus. WFS1 silencing in PBMCs from control subjects by means of small RNA interference also induced a pronounced proinflammatory cytokine profile. The same cytokines were also significantly higher in sera from the WS patient as compared to matched healthy controls. Moreover, the chronic inflammatory state was associated with a dominance of proinflammatory T helper 17 (Th17)-type cells over regulatory T (Treg) lymphocytes in the WS PBMCs. The identification of a state of systemic chronic inflammation associated with WFS1 deficiency may pave the way to innovative and personalized therapeutic interventions in WS.

scholarly journals Development of a Clinical Assay To Evaluate Toll-Like Receptor Function

2006 ◽  
Vol 13 (1) ◽  
pp. 68-76 ◽  
Author(s):  
Raquel P. Deering ◽  
Jordan S. Orange
Keyword(s):  
Mononuclear Cells ◽  
Genetic Defect ◽  
Recurrent Infection ◽  
Severe Infection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Gene Encoding ◽  
Peripheral Blood Mononuclear ◽  
Healthy Donors ◽  
Healthy Control ◽  
Molecular Patterns

ABSTRACT Toll-like receptors (TLRS) recognize pathogen-associated molecular patterns to enable innate immune responses. A number of genetic defects influencing the function of these receptors have been identified and are associated with recurrent and/or severe infection. Our goal was to develop a reproducible assay of TLR function in order to evaluate patients with recurrent infection who would be suspected of having a genetic defect affecting TLR signaling. We chose to study peripheral blood mononuclear cells (PBMCS) to avoid potential influences of soluble factors contained in whole blood, and we utilized ligands for TLRS 1/2, 2/6, 3, 4, 5, 6, 7, and 9. Tumor necrosis factor (TNF) production in PBMC supernatants was measured by an enzyme-linked immunosorbent assay after TLR ligand stimulation and was dependent on gene transcription and NF-κB activation. Some variables affecting the assay were assessed, including the effects of: blood anticoagulant, serum-containing media, incubation time, ligand storage, blood storage time, and cell cryopreservation. By using optimized assay conditions, effective concentrations of individual ligands and mean responses to those ligands were established for healthy control donors. Finally, three patients with a mutation in the IKBKG gene, encoding the NF-κB essential modulator (NEMO) protein, were evaluated as disease controls and were almost uniformly below the standard deviation of healthy donors for all ligands tested. Although a number of variables influence TLR ligand-induced TNF responses, this assay can be optimized for potential clinical use to screen patients with primary immunodeficiencies affecting TLR function.

scholarly journals Modulation of Wolframin Expression in Human Placenta during Pregnancy: Comparison among Physiological and Pathological States

2014 ◽  
Vol 2014 ◽  
pp. 1-7 ◽  
Author(s):  
Angela Lucariello ◽  
Angelica Perna ◽  
Carmine Sellitto ◽  
Alfonso Baldi ◽  
Alessandro Iannaccone ◽  
...  
Keyword(s):  
Human Placenta ◽  
Autosomal Recessive Disorder ◽  
Cell Loss ◽  
Wolfram Syndrome ◽  
First Trimester ◽  
Third Trimester ◽  
Gene Encoding ◽  
Cytotrophoblast Cell ◽  
The Third ◽  
Normal Placenta

TheWFS1gene, encoding a transmembrane glycoprotein of the endoplasmic reticulum called wolframin, is mutated in Wolfram syndrome, an autosomal recessive disorder defined by the association of diabetes mellitus, optic atrophy, and further organ abnormalities. Disruption of theWFS1gene in mice causes progressiveβ-cell loss in the pancreas and impaired stimulus-secretion coupling in insulin secretion. However, little is known about the physiological functions of this protein. We investigated the immunohistochemical expression of wolframin in human placenta throughout pregnancy in normal women and diabetic pregnant women. In normal placenta, there was a modulation of wolframin throughout pregnancy with a strong level of expression during the first trimester and a moderate level in the third trimester of gestation. In diabetic women, wolframin expression was strongly reduced in the third trimester of gestation. The pattern of expression of wolframin in normal placenta suggests that this protein may be required to sustain normal rates of cytotrophoblast cell proliferation during the first trimester of gestation. The decrease in wolframin expression in diabetic placenta suggests that this protein may participate in maintaining the physiologic glucose homeostasis in this organ.

scholarly journals The Increased RNase Activity of IRE1α in PBMCs from Patients with Rheumatoid Arthritis

2019 ◽  
Vol 9 (3) ◽  
pp. 505-509 ◽  
Author(s):  
Mahdieh Ahmadiany ◽  
Mahshid Alavi-Samani ◽  
Zahra Hashemi ◽  
Mohammad Amin Moosavi ◽  
Marveh Rahmati
Keyword(s):  
Rheumatoid Arthritis ◽  
Er Stress ◽  
Mononuclear Cells ◽  
Gradient Centrifugation ◽  
Rnase Activity ◽  
Healthy Controls ◽  
Peripheral Blood Mononuclear ◽  
Micro Rnas ◽  
Density Gradient Centrifugation Method ◽  
Gene Expression Levels

Purpose: Despite recent advances in the diagnosis and treatment of rheumatoid arthritis (RA), this inflammatory disease remains a challenge to patients and physicians. Recent evidence highlights the contribution of endoplasmic reticulum (ER) stress in the pathogenesis and treatment of RA. Herein, we study the expression of the ER stress sensor inositol-requiring enzyme 1α (IRE1α), as well as XBP1 splicing and the regulated IRE1-dependent decay (RIDD), in peripheral blood mononuclear cells (PBMCs) from patients with RA compared with healthy controls. Methods: The PBMCs from blood samples of RA patients and healthy volunteers were isolated by a density gradient centrifugation method using Ficoll. The gene expression levels of GRP78/ Bip, IRE1, XBP1s, micro-RNAs (miRNAs) were evaluated by real-time PCR. Results: The expression of GRP78, IRE1, and XBP1s were increased in PBMCs of RA patients compared with healthy controls. We further show that the RIDD targets (miRNA-17, -34a, -96, and -125b) were downregulated in RA samples. Conclusion: This study can expand our knowledge on the importance of RNase activity of IRE1α in RA and may offer new potentials for developing novel diagnostic and/or therapeutic biomarkers.

scholarly journals The gene encoding the Wiskott-Aldrich syndrome (WAS) protein is transcriptionally repressed following stimulation of peripheral blood mononuclear cells with muramyl dipeptide.

2020 ◽  
Author(s):  
Shahan Mamoor
Keyword(s):  
Gene Expression ◽  
Pattern Recognition ◽  
Crohn’S Disease ◽  
Crohn's Disease ◽  
Mononuclear Cells ◽  
Muramyl Dipeptide ◽  
Gene Encoding ◽  
Peripheral Blood Mononuclear ◽  
Wiskott Aldrich Syndrome ◽  
Blood Mononuclear Cells

The nucleotide-binding oligomerization domain protein 2, NOD2, is the pattern recognition receptor for muramyl dipeptide, a component of the cell wall of both gram-positive and gram-negative bacteria (1, 2). Sensing of muramyl dipeptide by NOD2 triggers signal transduction downstream of the RIP2 kinase to transcriptionally activate a diverse innate immune gene expression program (3-5). Single nucleotide variants in NOD2 are the strongest genetic risk factors for developing Crohn’s Disease (6, 7), an inflammatory bowel disease (8). By mining a published dataset (9), we found that among the genes whose expression changed most significantly in peripheral blood mononuclear cells (PBMC) stimulated with muramyl dipeptide, genome-wide, was the gene encoding the Wiskott-Aldrich syndrome (WAS) protein (10), a regulator of actin polymerization in hematopoietic cells (11). The WAS gene is mutated in patients with Wiskott-Aldrich syndrome (12), a genetic disorder characterized by thrombocytopenia, eczema, immunodeficiency and lymphoid malignancies (13), and whose symptoms include bloody diarrhea (14, 15). WAS RNA message was transcriptionally repressed following exposure of PBMC to muramyl dipeptide. These data link together a gene, that when mutated in humans leads to gastrointestinal symptoms not dissimilar to those found in patients with Crohn’s Disease, with the ligand for the gene whose product encodes the single most significant genetic variant conferring risk for development of Crohn’s Disease, and further suggest that the actin cytoskeleton may be a central target of gene expression changes effected by NOD2 pattern recognition of MDP.

scholarly journals Sialylation of IgG inhibits the formation of galactose-deficient IgA1-containing immune complexes and protects mesangial cells from injury in IgA nephropathy

2022 ◽  
Vol 23 (1) ◽  
Author(s):  
Youxia Liu ◽  
Hongfen Li ◽  
Huyan Yu ◽  
Fanghao Wang ◽  
Junya Jia ◽  
...  
Keyword(s):  
Iga Nephropathy ◽  
B Lymphocytes ◽  
Immune Complexes ◽  
Mononuclear Cells ◽  
Mesangial Cells ◽  
Healthy Controls ◽  
Peripheral Blood Mononuclear ◽  
Tnf Α ◽  
Inflammatory State

Abstract Background The addition of sialic acid alters IgG from a pro-inflammatory state to an anti-inflammatory state. However, there is a lack of research on the changes of IgG sialylation in IgA nephropathy (IgAN). Methods This study included a total of 184 IgAN patients. The sialylated IgG (SA-IgG), IgG-galactose-deficient IgA1 complex (IgG-Gd-IgA1-IC), IL-6, TNF-α, and TGF-β were detected using commercial ELISA kits. SA-IgG, non-sialylated IgG (NSA-IgG), sialylated IgG-IgA1 complex (SA-IgG-IgA1), and non-sialylated IgG-IgA1 complex (NSA-IgG-IgA1) were purified from IgAN patients and healthy controls (HCs). Results The mean SA-IgG levels in plasma and B lymphocytes in IgAN patients were significantly higher than those of healthy controls. A positive correlation was found between SA-IgG levels in plasma and B lymphocytes. In vitro, the results showed that the release of IgG-Gd-IgA1-IC was significantly decreased in peripheral blood mononuclear cells (PBMCs) cultured with SA-IgG from both IgAN patients and healthy controls. The proliferation ability and the release of IL-6, TNF-α, and TGF-β in human mesangial cells (HMCs) were measured after stimulating with SA-IgG-IgA1-IC and NSA-IgG-IgA1-IC. The mesangial cell proliferation levels induced by NSA-IgG-IgA1-IC derived from IgAN patients were significantly higher than those caused by SA-IgG-IgA1-IC derived from IgAN patients and healthy controls. Compared with NSA-IgG-IgA1 from healthy controls, IgAN-NSA-IgG-IgA1 could significantly upregulate the expression of IL-6 and TNF-α in mesangial cells. The data showed that there weren’t any significant differences in the levels of IL-6, TNF-α, and TGF-β when treated with IgAN-SA-IgG-IgA1 and HC-NSA-IgG-IgA1. Conclusions The present study demonstrated that the sialylation of IgG increased in patients with IgA nephropathy. It exerted an inhibitory effect on the formation of Gd-IgA1-containing immune complexes in PBMCs and the proliferation and inflammation activation in mesangial cells.

scholarly journals Soluble Epoxide Hydrolase 2 Expression Is Elevated in Obese Humans and Decreased by Physical Activity

2020 ◽  
Vol 21 (6) ◽  
pp. 2056 ◽  
Author(s):  
Abdelkrim Khadir ◽  
Sina Kavalakatt ◽  
Dhanya Madhu ◽  
Preethi Cherian ◽  
Fahd Al-Mulla ◽  
...  
Keyword(s):  
Physical Activity ◽  
Er Stress ◽  
Epoxide Hydrolase ◽  
Mononuclear Cells ◽  
Metabolic Stress ◽  
Mrna Levels ◽  
Peripheral Blood Mononuclear ◽  
Before And After ◽  
Activating Transcription Factor ◽  
Factor Α

Epoxide hydrolase 2 (EPHX2) is an emerging therapeutic target in several immunometabolic disorders. EPHX2 metabolizes anti-inflammatory epoxyeicosatrienoic acids into pro-inflammatory diols. The contribution of EPHX2 activity to human obesity remains unexplored. We compared the expression of EPHX2 between lean and obese humans (n = 20 each) in subcutaneous adipose tissue (SAT) and peripheral blood mononuclear cells (PBMCs) using RT-PCR, Western Blot analysis, immunohistochemistry, and confocal microscopy before and after a 3-month physical activity regimen. We also assessed EPHX2 levels during preadipocyte differentiation in humans and mice. EPHX2 mRNA and protein expression were significantly elevated in obese subjects, with concomitant elevated endoplasmic reticulum (ER) stress components (the 78-kDa glucose-regulated protein; GRP78, and the Activating transcription factor 6; ATF6) and inflammatory markers (Tumor necrosis factor-α; TNFα, and Interleukin 6; IL6) as compared to controls (p < 0.05). EPHX2 mRNA levels strongly correlated with adiposity markers. In obese individuals, physical activity attenuated EPHX2 expression levels in both the SAT and PBMCs, with a parallel decrease in ER stress and inflammation markers. EPHX2 expression was also elevated during differentiation of both human primary and 3T3-L1 mouse preadipocytes. Mediators of cellular stress (palmitate, homocysteine, and macrophage culture medium) also increased EPHX2 expression in 3T3-L1 preadipocytes. Our findings suggest that EPHX2 upregulation is linked to ER stress in adiposity and that physical activity may attenuate metabolic stress by reducing EPHX2 expression.

scholarly journals The Anti-Inflammatory Role of Mannich Curcuminoids; Special Focus on Colitis

Molecules ◽  
2019 ◽  
Vol 24 (8) ◽  
pp. 1546 ◽  
Author(s):  
Gábor J. Szebeni ◽  
Lajos I. Nagy ◽  
Anikó Berkó ◽  
Alexandra Hoffmann ◽  
Liliána Z. Fehér ◽  
...  
Keyword(s):  
Mononuclear Cells ◽  
Neutrophil Infiltration ◽  
Therapeutic Interventions ◽  
Inflammatory Activity ◽  
Interleukin 4 ◽  
Special Focus ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Peripheral Blood Mononuclear ◽  
Anti Inflammatory

The incidence of inflammatory bowel disease (IBD) increases gradually in Western countries with high need for novel therapeutic interventions. Mannich curcuminoids, C142 or C150 synthetized in our laboratory, have been tested for anti-inflammatory activity in a rat model of TNBS (2,4,6-trinitrobenzenesulphonic acid) induced colitis. Treatment with C142 or C150 reduced leukocyte infiltration to the submucosa and muscular propria of the inflamed gut. C142 or C150 rescued the loss of body weight and C150 decreased the weight of standard colon preparations proportional with 20% less tissue oedema. Both C142 and C150 curcumin analogues caused 25% decrease in the severity of colonic inflammation and haemorrhagic lesion size. Colonic MPO (myeloperoxidase) enzyme activity as an indicator of intense neutrophil infiltration was 50% decreased either by C142 or C150 Mannich curcuminoids. Lipopolysaccharide (LPS) co-treatment with Mannich curcuminoids inhibited NF-κB (nuclear factor kappa B) activity on a concentration-dependent manner in an NF-κB-driven luciferase expressing reporter cell line. Co-treatment with LPS and curcuminoids, C142 or C150, resulted in NF-κB inhibition with 3.57 μM or 1.6 μM half maximal effective concentration (EC50) values, respectively. C150 exerted a profound inhibition of the expression of inflammatory cytokines, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-4 (IL-4) in human PBMCs (peripheral blood mononuclear cells) upon LPS stimulus. Mannich curcuminoids reported herein possess a powerful anti-inflammatory activity.

scholarly journals Adjuvant Activity of Synthetic Lipid A of Alcaligenes, a Gut-Associated Lymphoid Tissue-Resident Commensal Bacterium, to Augment Antigen-Specific IgG and Th17 Responses in Systemic Vaccine

Vaccines ◽  
2020 ◽  
Vol 8 (3) ◽  
pp. 395 ◽  
Author(s):  
Yunru Wang ◽  
Koji Hosomi ◽  
Atsushi Shimoyama ◽  
Ken Yoshii ◽  
Haruki Yamaura ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Mhc Class Ii ◽  
Lipid A ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
T Helper ◽  
T Helper 17 ◽  
Adjuvant Activity ◽  
Peripheral Blood Mononuclear ◽  
Specific Igg

Alcaligenes spp. are identified as commensal bacteria and have been found to inhabit Peyer’s patches in the gut. We previously reported that Alcaligenes-derived lipopolysaccharides (LPS) exerted adjuvant activity in systemic vaccination, without excessive inflammation. Lipid A is one of the components responsible for the biological effect of LPS and has previously been applied as an adjuvant. Here, we examined the adjuvant activity and safety of chemically synthesized Alcaligenes lipid A. We found that levels of OVA-specific serum IgG antibodies increased in mice that were subcutaneously immunized with ovalbumin (OVA) plus Alcaligenes lipid A relative to those that were immunized with OVA alone. In addition, Alcaligenes lipid A promoted antigen-specific T helper 17 (Th17) responses in the spleen; upregulated the expression of MHC class II, CD40, CD80, and CD86 on bone marrow-derived dendritic cells (BMDCs); enhanced the production of Th17-inducing cytokines IL-6 and IL-23 from BMDCs. Stimulation with Alcaligenes lipid A also induced the production of IL-6 and IL-1β in human peripheral blood mononuclear cells. Moreover, Alcaligenes lipid A caused minor side effects, such as lymphopenia and thrombocytopenia. These findings suggest that Alcaligenes lipid A is a safe and effective Th17-type adjuvant by directly stimulating dendritic cells in systemic vaccination.

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