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关键词： 膀胱癌   FBXL6   TNF-α   增殖   迁移   RNA-seq  

摘要： 目的：探讨FBXL6在膀胱癌中的作用，特别是其通过调控肿瘤坏死因子（TNF）-α信号通路对膀胱癌细胞增殖和迁移的影响，评估FBXL6作为膀胱癌潜在预后标志物的价值。方法：首先通过TCGA数据库分析了FBXL6在膀胱癌组织中的表达水平，并与临床预后数据进行了相关性分析。接着，采用小干扰RNA(siRNA)技术在5637细胞和T24细胞中敲低FBXL6表达，通过Western Blot(WB)和qRT-PCR验证其在蛋白质和mRNA水平的抑制效果。进一步通过MTT实验评估细胞增殖能力，并进行Transwell以及Wound healing实验评估细胞迁移能力。最后采用RNA测序（RNA-seq）分析FBXL6下游调控的分子通路，其中TNF-α信号通路最为显著，我们进一步选取其中关键蛋白，通过WB实验加以验证。结果：TCGA数据分析显示，FBXL6在膀胱癌组织中的表达显著高于正常膀胱组织，且其高表达与膀胱癌患者的不良预后密切相关。细胞实验结果表明，敲低FBXL6后，膀胱癌细胞的增殖和迁移能力显著降低，过表达FBXL6则升高。RNA-seq分析进一步揭示，FBXL6通过调控TNF-α信号通路，促进了膀胱癌的发生和进展。结论：本研究证明FBXL6在膀胱癌中的关键作用，特别是在促进细胞增殖和迁移方面。通过调节TNF-α信号通路，FBXL6可能促进膀胱癌的进展，提示FBXL6可能成为膀胱癌的预后标志物及潜在治疗靶点。

关键词： Neuromyelitis optica   demyelinating diseases   genetic ancestry   immunogenetics  

摘要： Background: Internationally, specific human leukocyte antigen (HLA) class II alleles have been associated with neuromyelitis optica spectrum disorder (NMOSD) risk. Objective: To establish the association between HLA class II alleles and NMOSD in a Colombian population, while considering genetic ancestry. Methods: A multicenter case-control study was conducted with NMOSD patients diagnosed with the 2015 Wingerchuk criteria. HLA-DRB1 and HLA-DQB1 alleles were identified using sequence-specific primer polymerase chain reaction (SSP-PCR). Mitochondrial hypervariable region 1 was amplified, and haplogroups were determined with HaploGrep software. Genomic ancestry was assessed with ancestry-informative markers. Associations between HLA polymorphisms and NMOSD were analyzed using logistic regression models. Results: In total, 82 patients with NMOSD (mean age 43.8 +/- 13.3 years;83% females) and 164 controls (mean age 36.4 +/- 11.5 years;85% females) were enrolled. Mitochondrial haplogroup frequencies were similar between groups. Ancestry analysis revealed distinct population structures. In multivariable logistic regression, the HLA-DQB1*04 allele was significantly associated with increased NMOSD risk (odds ratio (OR) = 3.16, confidence interval (CI): 1.58-6.34, p < 0.0023), while the HLA-DQB1*03 allele was associated with a protective effect (OR = 0.23, CI: 0.12-0.46, p < 0.0023). Conclusion: HLA-DQB1*04 allele may increase susceptibility to NMOSD, while DQB1*03 allele could exert a protective effect in our population.

关键词： multiple sclerosis pathogenesis   blood   IL-6   IL-6 signaling   CD4 Tregs   CD8 Tregs  

摘要： Background: Peripheral immune cells participate in the pathogenesis and progression of central nervous system diseases including relapsing-remitting multiple sclerosis (RRMS), which is an immune-mediated demyelinating disorder. The association between IL-6 and RRMS pathogenesis is clear but there is some uncertainty about the role of IL-6 and IL-6 pathway components in blood and the molecular mechanisms through which T regulatorys (Tregs) contribute to MS pathogenesis. The purpose of this study was to identify markers of IL-6 pathways in serum and regulatory CD8+ and CD4+ T cells in the blood of RRMS patients and to analyze their associations with multiple sclerosis, with each other, and with ***: Peripheral blood was collected from female RRMS patients (16), and healthy controls (18) recruited between December, 2019 and July, 2022. The serum levels of IL-6, TGF-beta 1, IL-6R alpha, IL-6/IL-6R alpha complex, and soluble glycoprotein-130 (sgp-130) were measured by ELISA. Flow cytometry was used to quantify the surface expression of IL-6R (CD126), membrane glycoprotein 130 (gp130, CD130), and phospho-STAT3 (pSTAT3) and pSTAT5 in CD4+CD25+FoxP3+, CD8+CD25+FoxP3+, and CD8+CD122+ Tregs. Differences were compared using the Student's t-test or Welch's t-test. Pearson product-moment correlations were used to detect correlations. A p-value <= 0.05 was considered statistically ***: The CD8+CD122+ Treg subset in RRMS patients exhibited an increased level of surface CD126 and CD130 associated with classical IL-6R signaling without STAT3 phosphorylation. For CD4+CD25+FoxP3+ and CD8+CD25+FoxP3+ Tregs, no changes in classical IL-6R surface markers were observed in RRMS, but there was an increased percentage of pSTAT3 in these cells. Age-related changes in pSTAT5 expression across Treg subsets in healthy controls were absent in RRMS ***: Our findings underscore the complex interplay between IL-6 signaling and Tregs as well as age-related imm

关键词： COVID-19   pneumonia   proinflammatory cytokines  

摘要： Background/Objectives: Proinflammatory cytokines have been associated with poor prognosis in community-acquired and COVID-19 pneumonia. There is a paucity of reports on the cytokine release syndrome, also called cytokine storm in the Mexican population with pneumonia and COVID-19;therefore, our objective was to compare proinflammatory cytokine levels in Mexican patients without COVID-19 (non-COVID-19) and those with moderate, severe, and critical COVID-19 pneumonia. Methods: This study included 30 patients with non-COVID-19 pneumonia and 57 with COVID-19 pneumonia. Disease diagnosis and severity were determined using the radiographic pulmonary edema assessment (RALE) score. Quantification of IL-6, IL-10, and TNF-alpha was performed using multiplex immunoassays. A receiver operating characteristic curve was constructed to classify subjects with elevated cytokine levels. Logistic regression was used to find associations between elevated cytokine levels and the presence of COVID-19 pneumonia. Results: The severity classification of patients with COVID-19 pneumonia was as follows: moderate (n = 20), severe (n = 19), and critical (n = 18). The proinflammatory cytokines IL-6 and IL-10 were significantly increased in COVID-19 patients compared to non-COVID-19 patients (p < 0.005), while TNF-alpha levels were lower in critically ill patients with COVID-19 pneumonia. High levels of IL-6 and IL-10, adjusted for age, sex, the presence of comorbidities, and the neutrophil-to-lymphocyte ratio (NLR), showed an elevated risk (OR IL-6 = 4.02;OR IL-10 = 9.36) of presenting pneumonia and COVID-19 compared to pneumonia without COVID-19 in patients. Likewise, 61% of COVID-19 patients with elevated proinflammatory cytokines (IL-6 and IL-10) had a fatal outcome. Conclusions: Elevated levels of both IL-6 and IL-10 are a differential risk factor for developing COVID-19 pneumonia. These elevated levels were more frequently observed in Mexican COVID-19 pneumonia patients who died at the onse

关键词： IL-33   ST2 knockdown   Breast cancer stem cells   Wnt pathway   Immunotherapy  

摘要： Interleukin-33 (IL-33) plays multifaceted roles in tumor progression, but its autocrine regulation of breast cancer stemness and metastasis via the Wnt pathway remains unclear. Here, we investigated the IL-33/ST2 axis in breast cancer using CRISPR/Cas9, single-cell RNA sequencing, and murine models (orthotopic 4T1 and spontaneous MMTV-PyMT). Elevated IL-33 levels correlated with aggressive subtypes and poor prognosis. IL-33 overexpression enhanced proliferation, migration, and cancer stem cell (CSC) marker expression (CD44, ALDH1) in 4T1 and MDA-MB-231 cells, whereas ST2 knockdown via CRISPR or adeno-associated virus (AAV) attenuated tumor growth and metastasis in vivo, reducing CSC frequency. Mechanistically, IL-33 activated Wnt/beta-catenin signaling to promote stemness, which was reversed by the Wnt inhibitor XAV-939. Single-cell analysis revealed that IL-33 overexpression skewed the immune microenvironment toward immunosuppression, while ST2 knockdown restored antitumor immunity. Our findings establish an IL-33-Wnt axis as a critical driver of breast cancer aggressiveness and propose AAV-mediated ST2 silencing as a novel therapeutic strategy. Targeting this axis may offer dual benefits by suppressing stemness and enhancing immune surveillance, warranting clinical exploration for advanced breast cancer.

关键词： CD4(+) T cells   IL-12p35   immune thrombocytopenia   JAK2/STAT3/HIF-1 alpha   regulatory B cells  

摘要： Primary immune thrombocytopenia (ITP) is a condition characterized by immune-mediated platelet loss due to excessive destruction and/or insufficient production. IL-12p35 is involved in autoimmune uveitis;however, its role in ITP remains unknown. In the study, CD19(+) B and CD4(+) T cells were isolated from peripheral blood mononuclear cells (PBMCs) of patients with ITP and healthy individuals. CD4(+)CD25(-) T cells were cocultured with the CD19(+) B cells treated with anti-IL-12p35 antibody or IL-12p35 recombinant protein. The impact of IL-12p35 on CD19(+) B cells was assessed by enzyme-linked immunosorbent assay, Western blotting, quantitative real-time PCR and flow cytometry. PBMCs from patients with ITP showed lower proportions of IL-10(+)CD19(+) B cells and decreased mRNA levels of IL-12p35-coding gene IL12A than those from healthy individuals. Anti-IL-12p35 antibody-treated CD19(+) B cells could reduce the population of IL-10(+) in CD19(+) B cells and regulate the differentiation of CD4(+)CD25(-) T cells, while recombinant IL-12p35 demonstrated the opposite effects. Moreover, the increased IL-10(+) B-cell population and HIF-1 alpha expression in CD19(+) B cells induced by recombinant IL-12p35 were reversed by HIF-1 alpha and JAK2/STAT3 inhibitors. In conclusion, CD19(+)IL-10(+) B cells induced by IL-12p35 regulate CD4(+) T-cell subsets in patients with ITP via the JAK2/STAT3/HIF-1 alpha signalling pathway.

关键词： TNFA polymorphisms   TNF-alpha mRNA expression   sTNF-alpha   SLE  

摘要： Background/Objectives: A case-control study was conducted to determine the association of the -238 G>A and -308 G>A TNF-alpha (TNFA) promoter polymorphisms with mRNA and protein expression in 180 Systemic Lupus Erythematosus (SLE) patients and 186 control subjects (CS) from western Mexico. Methods: Genotyping was performed using the PCR-RFLP method. TNFA mRNA expression was assessed by real-time quantitative PCR, and soluble TNF-alpha (sTNF-alpha) levels were quantified by ELISA. For comparison groups, Chi-square, Mann-Whitney U, or Kruskal-Wallis tests were used. Spearman's rank correlation coefficient determined the correlation between variables. The Area Under the Curve was used to determine the diagnosing performance of sTNF-alpha. Results: No differences were found in the genotype distribution of -238 G>A and -308 G>A TNFA polymorphisms between SLE patients and CS. However, the -238A allele was associated with increased SLE susceptibility (OR 1.18 CI 95% 1.02-3.50, p = 0.037). Also, logistic regression analysis showed that LN risk was significantly higher in carriers of the -308A allele (OR 3.11 IC95% 1.15-6.43;p = 0.002). On the other hand, the TNFA mRNA expression was 3.3-fold higher in SLE compared to CS. SLE patients with -308 GG genotype showed higher TNFA mRNA expression compared to GA+AA genotype carriers (p < 0.01). Regarding sTNFa levels, SLE patients showed higher concentration than CS, mainly in lupus nephritis (LN), with a weak negative correlation with estimated Glomerular Filtration Rate and an acceptable accuracy for diagnosing SLE and LN, with areas under the curve of 0.61 and 0.65, respectively. Conclusions: The -238 A allele and -308 A allele of the TNFA gene are linked to a higher risk of susceptibility to SLE and LN in the western Mexican population. Additionally, SLE patients exhibited increased TNF-alpha gene expression and sTNF-alpha, particularly in LN, demonstrating acceptable diagnostic performance.

关键词： Immune related adverse event - irAE   Immune Checkpoint Inhibitor   Cardiotoxicity  

摘要： Background Immune checkpoint blockers (ICBs) have revolutionized cancer therapy, yet they remain largely ineffective in treating pancreatic ductal adenocarcinoma (PDAC). Moreover, ICBs can cause severe immune-related adverse events (irAEs), including fatal cardiac toxicity. Finally, obesity is a risk factor in PDAC that may differentially modulate ICB efficacy in a malignancy-dependent *** We investigated the mechanisms underlying irAEs induced by dual ICB therapy and sought to identify strategies to mitigate them while improving ICB efficacy in the obese setting. To this end, we used a clinically relevant mouse model that integrated key features of human PDAC: (1) high-fat diet-induced obesity, (2) an orthotopic PDAC, and (3) a therapeutic regimen combining chemotherapy (FOLFIRINOX) with ICBs (alpha-programmed cell death protein-1 + alpha-cytotoxic T-lymphocyte associated protein-4 antibodies).Results Obese mice developed cardiac irAEs and had elevated serum interleukin (IL)-1 beta levels after chemoimmunotherapy. IL-1 beta blockade not only prevented myocarditis and reduced cardiac fibrosis but also enhanced the antitumor efficacy of the combination of chemotherapy plus dual ICB therapy and significantly improved the overall survival of PDAC-bearing obese *** Our findings provide the rationale and compelling data to test a Food and Drug Administration-approved anti-IL-1 beta antibody in combination with chemotherapy and dual ICB therapy in patients with pancreatic cancer with obesity.

关键词： IL-33   ST2   Superoxide anion   Glial cells   Inflammation   And pain  

摘要： Reactive oxygen species such as superoxide anion have varied roles in inflammation and pain, which can be mimicked by potassium superoxide (KO2), the superoxide anion donor. Interleukin (IL)-33 has pleiotropic functions by activating its receptor suppression of tumorigenicity 2 (ST2). However, the role of IL-33/ST2 signaling in inflammatory pain initiated by reactive oxygen species (ROS) such as superoxide anion has not been investigated, which was the aim of the present study. IL-33 levels were assessed by enzyme-linked immunosorbent assay (ELISA). Mechanical and thermal hyperalgesia and overt pain were evaluated by electronic von Frey, hot plate, and abdominal writhing/paw flinching/licking, respectively. Edema and leukocyte recruitment (myeloperoxidase assay and total/differential cell count), antioxidant capacity, superoxide anion production and lipid peroxidation were assessed. Paw skin and spinal cord messenger ribonucleic acid (mRNA) expression of pro-inflammatory mediators and glial markers in the spinal cord were evaluated. Immunofluorescence was used to detect spinal glial and neuronal c-Fos activation. KO2 injection triggered IL-33 production in the paw skin and spinal cord of mice, induced hyperalgesia, edema, neutrophil recruitment to the paw tissue, overt pain-like behavior, and leukocyte recruitment to the peritoneum that were reduced in ST2 deficient mice. In the paw skin and spinal cord, KO2 triggered IL-33/ST2-dependent oxidative stress, and mRNA expression of inflammatory molecules, which were reduced by ST2 deficiency. KO2 induced spinal cord glial (at mRNA/protein levels) and neuronal activation in IL-33/ST2-dependent manner. IL-33/ST2 signaling mediates, at least in part, superoxide anion-induced inflammatory pain by modulating local and spinal inflammatory events.

关键词： organoids   precision medicine   air-liquid interface (ALI) culture   pancreatic ductal adenocarcinoma (PDAC)   cancer-associated fibroblasts (CAFs)  

摘要： Cancer organoids have emerged as transformative models for studying tumor biology and therapeutic responses due to the ability to replicate the complexity of the tumor microenvironment (TME). Tumor organoids recapitulate the genetic and phenotypic diversity of cancers, making them invaluable for investigating mechanisms of resistance and identifying novel therapeutic targets. Patient-derived organoids (PDOs) allow specific treatment methods to be designed based on the properties of each individual tumor in vitro. Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with an immunosuppressive nature. PDAC has a poor prognosis, with the survival rates of metastatic PDAC being improved only minimally over the last few decades. In this study, we demonstrate the antitumor effects of an IL-1 receptor antagonist (IL-1RA) in murine and human PDAC organoids. By reducing the burden of suppressive tumor elements like CAFs, IL-1RA treatment facilitates better immune cell access and response.