scholarly journals An Autoimmune Disease-Associated Risk Variant in theTNFAIP3Gene Plays a Protective Role in Brucellosis That Is Mediated by the NF-κB Signaling Pathway

2018 ◽  
Vol 56 (4) ◽  
Author(s):  
Lixin Lou ◽  
Wanguo Bao ◽  
Xianjun Liu ◽  
Hongxiao Song ◽  
Yang Wang ◽  
...  
Keyword(s):  
Autoimmune Disease ◽  
Mononuclear Cells ◽  
Protective Role ◽  
Negative Regulator ◽  
Functional Study ◽  
Mrna Levels ◽  
Peripheral Blood Mononuclear ◽  
Necrosis Factor Alpha ◽  
Tnfaip3 Gene ◽  
Functional Variants

ABSTRACTNaturally occurring functional variants (rs148314165 and rs200820567, collectively referred to as TT>A) reduce the expression of the tumor necrosis factor alpha-induced protein 3 (TNFAIP3) gene, a negative regulator of NF-κB signaling, and predispose individuals to autoimmune disease. In this analysis, we conducted a genetic association study of the TT>A variants in 1,209 controls and 150 patients with brucellosis, an infectious disease, and further assessed the role of the variants in brucellosis. Our data demonstrated that the TT>A variants were correlated with cases of brucellosis (P= 0.002; odds ratio [OR] = 0.34) and with individuals who had a positive serum agglutination test (SAT) result (titer of >1/160) (P= 4.2 × 10−6; OR = 0.23). A functional study demonstrated that brucellosis patients carrying the protective allele (A) showed significantly lower expression levels of the TNFAIP3 gene in their peripheral blood mononuclear cells and showed increased NF-κB signaling. Monocytes from individuals carrying the A allele that were stimulated withBrucella abortushad lower mRNA levels of TNFAIP3 and produced more interleukin-10 (IL-10), IL-6, and IL-1β than those from TT allele carriers. These data showed that autoimmune disease-associated risk variants, TT>A, of the TNFAIP3 locus play a protective role in the pathogenesis of brucellosis. Our findings suggest that a disruption of the normal function of the TNFAIP3 gene might serve as a therapeutic target for the treatment of brucellosis.

scholarly journals Blood functional assay for rapid clinical interpretation of germline TP53 variants

2020 ◽  
pp. jmedgenet-2020-107059 ◽  
Author(s):  
Sabine Raad ◽  
Marion Rolain ◽  
Sophie Coutant ◽  
Céline Derambure ◽  
Raphael Lanos ◽  
...  
Keyword(s):  
Mononuclear Cells ◽  
Transcriptional Response ◽  
Functional Assay ◽  
Mrna Levels ◽  
Peripheral Blood Mononuclear ◽  
Patients With Cancer ◽  
Pathogenic Variants ◽  
Functional Variants ◽  
Multiplex Ligation Probe Amplification ◽  
P53 Mrna

BackgroundThe interpretation of germline TP53 variants is critical to ensure appropriate medical management of patients with cancer and follow-up of variant carriers. This interpretation remains complex and is becoming a growing challenge considering the exponential increase in TP53 tests. We developed a functional assay directly performed on patients’ blood.MethodsPeripheral blood mononuclear cells were cultured, activated, exposed to doxorubicin and the p53-mediated transcriptional response was quantified using reverse transcription–multiplex ligation probe amplification and RT-QMPSF assays, including 10 p53 targets selected from transcriptome analysis, and two amplicons to measure p53 mRNA levels. We applied this blood functional assay to 77 patients addressed for TP53 analysis.ResultsIn 51 wild-type TP53 individuals, the mean p53 functionality score was 12.7 (range 7.5–22.8). Among eight individuals harbouring likely pathogenic or pathogenic variants, the scores were reduced (mean 4.8, range 3.1–7.1), and p53 mRNA levels were reduced in patients harbouring truncating variants. We tested 14 rare unclassified variants (p.(Pro72His), p.(Gly105Asp), p.(Arg110His), p.(Phe134Leu), p.(Arg158Cys), p.(Pro191Arg), p.(Pro278Arg), p.(Arg283Cys), p.(Leu348Ser), p.(Asp352Tyr), p.(Gly108_Phe109delinsVal), p.(Asn131del), p.(Leu265del), c.-117G>T) and 12 yielded functionally abnormal scores. Remarkably, the assay revealed that the c.*1175A>C polymorphic variant within TP53 poly-adenylation site can impact p53 function with the same magnitude as a null variant, when present on both alleles, and may act as a modifying factor in pathogenic variant carriers.ConclusionThis blood p53 assay should therefore be a useful tool for the rapid clinical classification of germline TP53 variants and detection of non-coding functional variants.

scholarly journals Expression of Tumor Necrosis Factor Alpha-Induced Protein 3 mRNA in Peripheral Blood Mononuclear Cells Negatively Correlates with Disease Severity in Psoriasis Vulgaris

2012 ◽  
Vol 19 (12) ◽  
pp. 1938-1942 ◽  
Author(s):  
Xuebing Jiang ◽  
Hongqing Tian ◽  
Yuchen Fan ◽  
Jie Chen ◽  
Yonghong Song ◽  
...  
Keyword(s):  
Tumor Necrosis ◽  
Psoriasis Vulgaris ◽  
Mononuclear Cells ◽  
Healthy Controls ◽  
Peripheral Blood Mononuclear ◽  
Necrosis Factor Alpha ◽  
Tnfaip3 Gene ◽  
Severe Group ◽  
Factor Alpha ◽  
Necrosis Factor

ABSTRACTPsoriasis vulgaris is considered a chronic inflammatory disease, but its immunopathogenesis has not been well understood. The tumor necrosis factor alpha-induced protein 3 (TNFAIP3) gene functions in negative-feedback regulation of inflammation, and its single nucleotide polymorphism is associated with psoriasis. However, the relationship between the expression level of the TNFAIP3 gene in immune cells and psoriasis is not known so far. In the present study, TNFAIP3 mRNA expression levels in peripheral blood mononuclear cells from 44 patients with psoriasis vulgaris and 30 healthy controls were determined using real-time reverse transcription-PCR analysis. We found that expression of TNFAIP3 mRNA in all patients negatively correlated with the psoriatic area and severity index (PASI) (r= −0.5126;P= 0.0004) as well as with the percentage of body surface area affected by psoriasis (r= −0.5013;P= 0.0005). Patients were divided into mild and severe groups based on the mean PASI score. Expression of TNFAIP3 mRNA in the mild group was higher than that in the severe group (P= 0.0064). Moreover, compared with that in healthy controls, the expression of TNFAIP3 mRNA in the mild group was significantly upregulated (P= 0.0004), but the expression of TNFAIP3 mRNA in the severe group was not. These results suggest that the expression level of TNFAIP3 plays an important role in the pathology of psoriasis vulgaris and that the loss of upregulation of TNFAIP3 expression may contribute to the severity of psoriasis vulgaris.

scholarly journals Identification of Promiscuous African Swine Fever Virus T-Cell Determinants Using a Multiple Technical Approach

Vaccines ◽  
2021 ◽  
Vol 9 (1) ◽  
pp. 29
Author(s):  
Laia Bosch-Camós ◽  
Elisabet López ◽  
María Jesús Navas ◽  
Sonia Pina-Pedrero ◽  
Francesc Accensi ◽  
...  
Keyword(s):  
T Cell ◽  
Mononuclear Cells ◽  
African Swine Fever Virus ◽  
African Swine Fever ◽  
Cd8 T Cell ◽  
Protective Role ◽  
Full Length ◽  
T Cell Epitopes ◽  
Peripheral Blood Mononuclear

The development of subunit vaccines against African swine fever (ASF) is mainly hindered by the lack of knowledge regarding the specific ASF virus (ASFV) antigens involved in protection. As a good example, the identity of ASFV-specific CD8+ T-cell determinants remains largely unknown, despite their protective role being established a long time ago. Aiming to identify them, we implemented the IFNγ ELISpot as readout assay, using as effector cells peripheral blood mononuclear cells (PBMCs) from pigs surviving experimental challenge with Georgia2007/1. As stimuli for the ELISpot, ASFV-specific peptides or full-length proteins identified by three complementary strategies were used. In silico prediction of specific CD8+ T-cell epitopes allowed identifying a 19-mer peptide from MGF100-1L, as frequently recognized by surviving pigs. Complementarily, the repertoire of SLA I-bound peptides identified in ASFV-infected porcine alveolar macrophages (PAMs), allowed the characterization of five additional SLA I-restricted ASFV-specific epitopes. Finally, in vitro stimulation studies using fibroblasts transfected with plasmids encoding full-length ASFV proteins, led to the identification of MGF505-7R, A238L and MGF100-1L as promiscuously recognized antigens. Interestingly, each one of these proteins contain individual peptides recognized by surviving pigs. Identification of the same ASFV determinants by means of such different approaches reinforce the results presented here.

scholarly journals Lipopolysaccharides prime whole human blood and isolated neutrophils for the increased synthesis of 5-lipoxygenase products by enhancing arachidonic acid availability: involvement of the CD14 antigen.

1993 ◽  
Vol 178 (4) ◽  
pp. 1347-1355 ◽  
Author(s):  
M E Surette ◽  
R Palmantier ◽  
J Gosselin ◽  
P Borgeat
Keyword(s):  
Arachidonic Acid ◽  
Mononuclear Cells ◽  
Leukotriene B4 ◽  
Eicosatetraenoic Acid ◽  
Tnf Alpha ◽  
Peripheral Blood Mononuclear ◽  
Necrosis Factor Alpha ◽  
Factor Alpha ◽  
Priming Activity

Stimulation of heparinized blood with 1 microM formyl-methionyl-leucyl-phenylalanine (FMLP) resulted in the formation of < 30 pmol/ml plasma of 5-lipoxygenase (5-LO) products. The preincubation of blood with 1 microgram/ml of lipopolysaccharide (LPS) (Escherichia coli 0111-B4) for 30 min before stimulation with FMLP resulted in the accumulation of 250-300 pmol of 5-LO products per ml plasma. The major products detected were leukotriene B4 and (5S)-hydroxy-6,8,11,14-eicosatetraenoic acid which were produced in equivalent amounts. The priming activity was detectable with as little as 1-10 ng LPS per ml blood and was optimal using 1-10 micrograms LPS/ml blood. The priming for 5-LO product synthesis was optimal after 20-30 min of preincubation with LPS and declined at preincubation times > 30 min. The priming effect of LPS was also observed using the complement fragment C5a or interleukin 8 as agonists. Polymorphonuclear leukocytes (PMN) and peripheral blood mononuclear cells accounted for 80 and 20% of the synthesis of 5-LO products, respectively. The ability of LPS to prime isolated PMN was dependent on the presence of plasma and was inhibited by the anti-CD14 antibody IOM2, indicating a CD14-dependent priming mechanism. The priming of whole blood with tumor necrosis factor alpha (TNF-alpha) and LPS was additive and the presence of mononuclear cells did not enhance the ability of LPS to prime PMN, indicating that the priming activity of LPS is independent of LPS-induced TNF-alpha synthesis. The mechanism by which LPS enhance 5-LO product synthesis in PMN was investigated. Treatment of PMN with LPS strongly enhanced the release of arachidonic acid after stimulation with FMLP. The release of arachidonic acid was optimal 2-3 min after stimulation with FMLP, attaining levels 5-15-fold greater than those observed in unprimed cells stimulated with FMLP. These results demonstrate that LPS dramatically increases the ability of blood to generate 5-LO products, and support the putative role of leukotrienes in pathological states involving LPS.

scholarly journals Decreased levels of miR-28-5p and miR-361-3p and increased levels of insulin-like growth factor 1 mRNA in mononuclear cells from patients with hereditary hemorrhagic telangiectasia

2019 ◽  
Vol 97 (6) ◽  
pp. 562-569 ◽  
Author(s):  
Anthony Cannavicci ◽  
Qiuwang Zhang ◽  
Si-Cheng Dai ◽  
Marie E. Faughnan ◽  
Michael J.B. Kutryk
Keyword(s):  
Growth Factor ◽  
Peripheral Blood ◽  
Hereditary Hemorrhagic Telangiectasia ◽  
Mononuclear Cells ◽  
Quantitative Polymerase Chain Reaction ◽  
Clinical Manifestations ◽  
Mrna Levels ◽  
Insulin Like Growth Factor ◽  
Peripheral Blood Mononuclear ◽  
Micro Rnas

Hereditary hemorrhagic telangiectasia (HHT) is a rare vascular disorder inherited in an autosomal dominant manner. Patients with HHT can develop vascular dysplasias called telangiectasias and arteriovenous malformations (AVMs). Our objective was to profile and characterize micro-RNAs (miRNAs), short noncoding RNAs that regulate gene expression posttranscriptionally, in HHT patient-derived peripheral blood mononuclear cells (PBMCs). PBMCs, comprised mostly of lymphocytes and monocytes, have been reported to be dysfunctional in HHT. A total of 40 clinically confirmed HHT patients and 22 controls were enrolled in this study. PBMCs were isolated from 16 mL of peripheral blood and purified for total RNA. MiRNA expression profiling was conducted with a human miRNA array analysis. Select dysregulated miRNAs and miRNA targets were validated with reverse transcription–quantitative polymerase chain reaction. Of the 377 miRNAs screened, 41 dysregulated miRNAs were identified. Both miR-28-5p and miR-361-3p, known to target insulin-like growth factor 1 (IGF1), a potent angiogenic growth factor, were found to be significantly downregulated in HHT patients. Consequently, IGF1 mRNA levels were found to be significantly elevated. Our research successfully identified miRNA dysregulation and elevated IGF1 mRNA levels in PBMCs from HHT patients. This novel discovery represents a potential pathogenic mechanism that could be targeted to alleviate clinical manifestations of HHT.

scholarly journals A 70-Kilodalton Recombinant Heat Shock Protein ofCandida albicans Is Highly Immunogenic and Enhances Systemic Murine Candidiasis

1998 ◽  
Vol 66 (5) ◽  
pp. 2154-2162 ◽  
Author(s):  
Carla Bromuro ◽  
Roberto La Valle ◽  
Silvia Sandini ◽  
Francesca Urbani ◽  
Clara M. Ausiello ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Mononuclear Cells ◽  
Cell Mediated Immunity ◽  
Healthy Human ◽  
Peripheral Blood Mononuclear ◽  
Necrosis Factor Alpha ◽  
Ifn Γ

ABSTRACT The 70-kDa recombinant Candida albicans heat shock protein (CaHsp70) and its 21-kDa C-terminal and 28-kDa N-terminal fragments (CaHsp70-Cter and CaHsp70-Nter, respectively) were studied for their immunogenicity, including proinflammatory cytokine induction in vitro and in vivo, and protection in a murine model of hematogenous candidiasis. The whole protein and its two fragments were strong inducers of both antibody (Ab; immunoglobulin G1 [IgG1] and IgG2b were the prevalent isotypes) and cell-mediated immunity (CMI) responses in mice. CaHsp70 preparations were also recognized as CMI targets by peripheral blood mononuclear cells of healthy human subjects. Inoculation of CaHsp70 preparations into immunized mice induced rapid production of interleukin-6 (IL-6) and tumor necrosis factor alpha, peaking at 2 to 5 h and declining within 24 h. CaHsp70 and CaHsp70-Cter also induced gamma interferon (IFN-γ), IL-12, and IL-10 but not IL-4 production by CD4+ lymphocytes cocultured with splenic accessory cells from nonimmunized mice. In particular, the production of IFN-γ was equal if not superior to that induced in the same cells by whole, heat-inactivated fungal cells or the mitogenic lectin concanavalin A. In immunized mice, however, IL-4 but not IL-12 was produced in addition to IFN-γ upon in vitro stimulation of CD4+ cells with CaHsp70 and CaHsp70-Cter. These animals showed a decreased median survival time compared to nonimmunized mice, and their mortality was strictly associated with organ invasion by fungal hyphae. Their enhanced susceptibility was attributable to the immunization state, as it did not occur in congenitally athymic nude mice, which were unable to raise either Ab or CMI responses to CaHsp70 preparations. Together, our data demonstrate the elevated immunogenicity of CaHsp70, with which, however, no protection against but rather some enhancement of Candida infection seemed to occur in the mouse model used.

Reduced Expression of Annexin A2 is Associated with Impaired Cell Surface Fibrinolysis and Venous Thromboembolism

Blood ◽  
2021 ◽  
Author(s):  
Hannah Fassel ◽  
Huigen Chen ◽  
Mary Ruisi ◽  
Neha Kumar ◽  
Maria T DeSancho ◽  
...  
Keyword(s):  
Risk Factor ◽  
Venous Thromboembolism ◽  
Cell Surface ◽  
Mononuclear Cells ◽  
Annexin A2 ◽  
Clot Lysis ◽  
Mrna Levels ◽  
Peripheral Blood Mononuclear ◽  
Intravascular Thrombosis

Reduced plasma fibrinolysis has been identified as a potential risk factor for venous thromboembolism (VTE), but the role of cell surface fibrinolysis in VTE is unknown. The annexin A2/S100A10 complex serves as a co-receptor for plasminogen and tissue plasminogen activator (tPA), augmenting plasmin generation by 60-fold on the endothelial cell surface. Several studies in both mice and humans support the concept that A2 regulates fibrin homeostasis and intravascular thrombosis in vivo. Here, we examined A2 protein expression and function in 115 adult subjects with venous thromboembolism (VTE) and 87 healthy controls. Using peripheral blood mononuclear cells (PBMCs) as a surrogate for endothelial cells, we found a 41% mean decrease in cell surface tPA-dependent fibrinolytic activity in subjects who had a positive personal and family history of VTE, but tested negative for known inherited thrombophilias. A2 protein was reduced on average by 70%, and mRNA levels by 30%, but neither decrease correlated with anticoagulant therapy. [Sentence omitted] Neither cell A2 protein nor cell surface plasmin generation correlated with plasma-based clot lysis times, suggesting that the plasma and cell surface fibrinolytic systems operate independently of one another. These data suggest that reduced expression of annexin A2 protein is associated with cell surface hypofibrinolysis and may represent a novel risk factor for inherited thrombophilia.

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