S100A9 exacerbates sepsis-induced acute lung injury via the IL17-NFκB-caspase-3 signaling pathway.

BACKGROUND: Sepsis-induced acute lung injury (ALI) is associated with considerable morbidity and mortality in critically ill patients. S100A9, a key endothelial injury factor, is markedly upregulated in sepsis-induced ALI; however, its specific mechanism of action has not been fully elucidated. METHODS: The Gene Expression Omnibus database transcriptome data for sepsis-induced ALI were used to screen for key differentially expressed genes (DEGs). Using bioinformatics analysis methods such as Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, and protein-protein interaction network analyses, the pathogenesis of sepsis-induced ALI was revealed. Intratracheal infusion of lipopolysaccharide (LPS, 10 mg/kg) induced ALI in wild-type (WT) and S100A9 knockout mice. Multiomics analyses (transcriptomics and proteomics) were performed to investigate the potential mechanisms by which S100A9 exacerbates acute lung damage. Hematoxylin-eosin, Giemsa, and TUNEL staining were used to evaluate lung injury and cell apoptosis. LPS (10 µg/mL)-induced murine lung epithelial MLE-12 cells were utilized to mimic ALI and were modulated by S100A9 lentiviral transfection. The impact of S100A9 on cell apoptosis and inflammatory responses were identified using flow cytometry and PCR. The expression of interleukin (IL)-17-nuclear factor kappa B (NFκB)-caspase-3 signaling components was identified using western blotting. RESULTS: Six common DEGs (S100A9, S100A8, IFITM6, SAA3, CD177, and MMP9) were identified in the six datasets related to ALI in sepsis. Compared to WT sepsis mice, S100A9 knockout significantly alleviated LPS-induced ALI in mice, with reduced lung structural damage and inflammatory exudation, decreased exfoliated cell and protein content in the lung lavage fluid, and reduced apoptosis and necrosis of pulmonary epithelial cells. Transcriptomic analysis revealed that knocking out S100A9 significantly affected 123 DEGs, which were enriched in immune responses, defense responses against bacteria or lipopolysaccharides, cytokine-cytokine receptor interactions, and the IL-17 signaling pathway. Proteomic analysis revealed that S100A9 knockout alleviated muscle contraction dysfunction and structural remodeling in sepsis-induced ALI. Multiomics analysis revealed that S100A9 may be closely related to interferon-induced proteins with tetratricopeptide repeats and oligoadenylate synthase-like proteins. LPS decreased MLE12 cell activity, accompanied by high expression of S100A9. The expression of IL-17RA, pNFκB, and cleaved-caspase-3 were increased by S100A9 overexpression and reduced by S100A9 knockdown in LPS-stimulated MLE12 cells. S100A9 knockdown decreases transcription of apoptosis-related markers Bax, Bcl and caspase-3, alleviating LPS-induced apoptosis. CONCLUSIONS: S100A9 as a key biomarker of sepsis-induced acute lung injury, and exacerbates lung damage and epithelial cell apoptosis induced by LPS via the IL-17-NFκB-caspase-3 signaling pathway.

S100A8/9 modulates perturbation and glycolysis of macrophages in allergic asthma mice.

Background: Allergic asthma is the most prevalent asthma phenotype and is associated with the disorders of immune cells and glycolysis. Macrophages are the most common type of immune cells in the lungs. Calprotectin (S100A8 and S100A9) are two pro-inflammatory molecules that target the Toll-like receptor 4 (TLR4) and are substantially increased in the serum of patients with severe asthma. This study aimed to determine the effects of S100A8/A9 on macrophage polarization and glycolysis associated with allergic asthma. Methods: To better understand the roles of S100A8 and S100A9 in the pathogenesis of allergic asthma, we used ovalbumin (OVA)-induced MH-S cells, and OVA-sensitized and challenged mouse models (wild-type male BALB/c mice). Enzyme-linked immunosorbent assay, quantitative real-time polymerase chain reaction, flow cytometry, hematoxylin-eosin staining, and western blotting were performed. The glycolysis inhibitor 3-bromopyruvate (3-BP) was used to observe changes in glycolysis in mice. Results: We found knockdown of S100A8 or S100A9 in OVA-induced MH-S cells inhibited inflammatory cytokines, macrophage polarization biomarker expression, and pyroptosis cell proportion, but increased anti-inflammatory cytokine interleukin (IL)-10 mRNA; also, glycolysis was inhibited, as evidenced by decreased lactate and key enzyme expression; especially, knockdown of S100A8 or S100A9 inhibited the activity of TLR4/myeloid differentiation primary response gene 88 (MyD88)/Nuclear factor kappa-B (NF-κB) signaling pathway. Intervention with lipopolysaccharides (LPS) abolished the beneficial effects of S100A8 and S100A9 knockdown. The observation of OVA-sensitized and challenged mice showed that S100A8 or S100A9 knockdown promoted respiratory function, improved lung injury, and inhibited inflammation; knockdown of S100A8 or S100A9 also suppressed macrophage polarization, glycolysis levels, and activation of the TLR4/MyD88/NF-κB signaling pathway in the lung. Conversely, S100A9 overexpression exacerbated lung injury and inflammation, promoting macrophage polarization and glycolysis, which were antagonized by the glycolysis inhibitor 3-BP. Conclusion: S100A8 and S100A9 play critical roles in allergic asthma pathogenesis by promoting macrophage perturbation and glycolysis through the TLR4/MyD88/NF-κB signaling pathway. Inhibition of S100A8 and S100A9 may be a potential therapeutic strategy for allergic asthma.

Identified S100A9 as a target for diagnosis and treatment of ulcerative colitis by bioinformatics analysis.

Ulcerative colitis (UC) is a chronic, recurrent inflammatory bowel disease. UC confronts with severe challenges including the unclear pathogenesis and lack of specific diagnostic markers, demanding for identifying predictive biomarkers for UC diagnosis and treatment. We perform immune infiltration and weighted gene co-expression network analysis on gene expression profiles of active UC, inactive UC, and normal controls to identify UC related immune cell and hub genes. Neutrophils, M1 macrophages, activated dendritic cells, and activated mast cells are significantly enriched in active UC. MMP-9, CHI3L1, CXCL9, CXCL10, CXCR2 and S100A9 are identified as hub genes in active UC. Specifically, S100A9 is significantly overexpressed in mice with colitis. The receiver operating characteristic curve demonstrates the excellent performance of S100A9 expression in diagnosing active UC. Inhibition of S100A9 expression reduces DSS-induced colonic inflammation. These identified biomarkers associated with activity in UC patients enlighten the new insights of UC diagnosis and treatment.

S100A9-/- alleviates LPS-induced acute lung injury by regulating M1 macrophage polarization and inhibiting pyroptosis via the TLR4/MyD88/NFκB signaling axis.

Acute lung injury (ALI) is characterized by pulmonary diffusion abnormalities that may progress to multiple-organ failure in severe cases. There are limited effective treatments for ALI, which makes the search for new therapeutic avenues critically important. Macrophages play a pivotal role in the pathogenesis of ALI. The degree of macrophage polarization is closely related to the severity and prognosis of ALI, and S100A9 promotes M1 polarization of macrophages. The present study assessed the effects of S100A9-gene deficiency on macrophage polarization and acute lung injury. Our cohort study showed that plasma S100A8/A9 levels had significant diagnostic value for pediatric pneumonia and primarily correlated with monocyte-macrophages and neutrophils. We established a lipopolysaccharide (LPS)-induced mouse model of acute lung injury and demonstrated that knockout of the S100A9 gene mitigated inflammation by suppressing the secretion of pro-inflammatory cytokines, reducing the number of inflammatory cells in the bronchoalveolar lavage fluid, and inhibiting cell apoptosis, which ameliorated acute lung injury in mice. The in vitro and in vivo mechanistic studies demonstrated that S100A9-gene deficiency inhibited macrophage M1 polarization and reduced the levels of pulmonary macrophage chemotactic factors and inflammatory cytokines by suppressing the TLR4/MyD88/NF-κB signaling pathway and reversing the expression of the NLRP3 pyroptosis pathway, which reduced cell death. In conclusion, S100A9-gene deficiency alleviated LPS-induced acute lung injury by inhibiting macrophage M1 polarization and pyroptosis via the TLR4/MyD88/NFκB pathway, which suggests a potential therapeutic strategy for the treatment of ALI.

[Single-cell RNA sequencing combined experimental verifies the core genes of dendritic cells in chronic obstructive pulmonary disease].

Objective Single-cell RNA sequencing (scRNA-Seq) and experimental verifies core genes of dendritic cells in chronic obstructive pulmonary disease (COPD). Methods scRNA-seq data GSE173896 and chip data GSE38974 were extracted from the Gene Expression Omnibus (GEO) database. GSE173896 was used to perform quality control, batch correction, dimensionality reduction clustering, cell type annotation and dendritic cell differentially expressed genes (DC-DEGs) identification. DEGs from the analysis of GSE38974 were intersected with DC-DEGs to obtain the common DC-DEGs. The diagnostic efficacy of the common DC-DEGs for COPD and their enrichment analysis were conducted. The correlation of the common DC-DEGs with activated dendritic cell (DCs), plasmacytoid dendritic cell (pDCs) and type 17 T helper(Th17) cells were analyzed. The mRNA expression level of the common DC-DEGs in the lung tissue of emphysema mice was verified. Results From GSE173896, 18 DC-DEGs were obtained between groups and from GSE38974, 646 DEGs were obtained. The intersection of the two resulted in 3 common DC-DEGs, including interleukin 1 receptor antagonist 1 (IL1RN), S100 calcicum-binding protein A8 (S100A8) and S100A9. Their respective area under curve (AUC) values were 0.841, 0.804 and 0.966. The GO and KEGG enrichment analysis mainly concentrated on chronic inflammatory response, collagen-containing extracellular matrix, receptor for advanced glycation end products (RAGE) binding, Toll-like receptor (TLR) binding and interleukin 17 (IL-17) signaling pathway. IL1RN, S100A8 and S100A9 were positively correlated with activated DCs, pDCs and Th17 cells. The results showed that the mRNA relative expression levels of IL1RN, S100A8 and S100A9 were up-regulated in the lung tissue of emphysema mice. Conclusion IL1RN, S100A8 and S100A9 may be the core genes of DCs in the pathogenesis of COPD, which potentially provide targets and a theoretical basis for subsequent COPD immunotherapy.

Association Between Clinicopathological Parameters and S100A8/A9 Expression According to Smoking History in Patients With Non-small Cell Lung Cancer.

BACKGROUND/AIM: Lung cancer is a major cause of cancer-related deaths worldwide, and chronic inflammation caused by cigarette smoke plays a crucial role in the development and progression of this disease. S100A8/9 and RAGE are associated with chronic inflammatory diseases and cancer. This study aimed to investigate the expression of S100A8/9, HMBG1, and other related pro-inflammatory molecules and clinical characteristics in patients with non-small cell lung cancer (NSCLC). PATIENTS AND METHODS: We obtained serum and bronchoalveolar lavage (BAL) fluid samples from 107 patients and categorized them as never or ever-smokers. We measured the levels of S100A8/9, RAGE, and HMGB1 in the collected samples using enzyme-linked immunosorbent kits. Immunohistochemical staining was also performed to assess the expression of S100A8/9, CD11b, and CD8 in lung cancer tissues. The correlation between the expression of these proteins and the clinical characteristics of patients with NSCLC was also explored. RESULTS: The expression of S100A8/A9, RAGE, and HMGB was significantly correlated with smoking status and was higher in people with a history of smoking or who were currently smoking. There was a positive correlation between serum and BAL fluid S100A8/9 levels. The expression of S100A8/A9 and CD8 in lung tumor tissues was significantly correlated with smoking history in patients with NSCLC. Ever-smokers, non-adenocarcinoma histology, and high PD-L1 expression were significant factors predicting high serum S100A8/9 levels in multivariate analysis. CONCLUSION: The S100A8/9-RAGE pathway and CD8 expression were increased in smoking-related NSCLC patients. The S100A8/9-RAGE pathway could be a promising biomarker for chronic airway inflammation and carcinogenesis in smoking-related lung diseases.

Targeting TAM-secreted S100A9 effectively enhances the tumor-suppressive effect of metformin in treating lung adenocarcinoma.

Metformin's effect on tumor treatment was complex, because it significantly reduced cancer cell proliferation in vitro, but made no difference in prognosis in several clinical cohorts. Our transcriptome sequencing results revealed that tumor-associated macrophage (TAM) infiltration significantly increased in active lung adenocarcinoma (LUAD) patients with long-term metformin use. We further identified that the tumor suppressive effect of metformin was more significant in mice after the depletion of macrophages, suggesting that TAMs might play an important role in metformin's effects in LUAD. Combining 10X Genomics single-cell sequencing of tumor samples, transcriptome sequencing of metformin-treated TAMs, and the ChIP-Seq data of the Encode database, we identified and validated that metformin significantly increased the expression and secretion of S100A9 of TAMs through AMPK-CEBP/ß pathway. For the downstream, S100A9 binds to RAGE receptors on the surface of LUAD cells, and then activates the NF-κB pathway to promote EMT and progression of LUAD, counteracting the inhibitory effect of metformin on LUAD cells. In cell-derived xenograft models (CDX) and patient-derived xenograft models (PDX) models, our results showed that neutralizing antibodies targeting TAM-secreted S100A9 effectively enhanced the tumor suppressive effect of metformin in treating LUAD. Our results will enable us to better comprehend the complex role of metformin in LUAD, and advance its clinical application in cancer treatment.

CD146 deficiency aggravates chronic obstructive pulmonary disease via the increased production of S100A9 and MMP-9 in macrophages.

Chronic obstructive pulmonary disease (COPD) is a leading cause of global death. As a molecule beyond adhesion, CD146 is involved in COPD pathogenesis. However, the mechanisms of CD146 in COPD remain largely elusive. We hypothesized that CD146 regulates the production of matrix metalloproteinase-9 (MMP-9) in macrophages and thereby contributes to COPD. Here, we constructed a murine model of COPD using lipopolysaccharide (LPS) and porcine pancreatic elastase (PPE). In COPD-like mice, LPS and PPE decreased the pulmonary expression of CD146. MMP-9 expression and bioactivity were increased in CD146 knockout COPD-like mice. In vitro, LPS decreased CD146 expression in macrophages. With or without LPS challenge, CD146-defective macrophages produced more MMP-9. Transcriptome analysis based on next-generation sequencing (NGS) revealed that S100A9 regulated MMP-9 production in CD146-defective macrophages. Targeting S100A9 with paquinimod decreased lung inflammation and alleviated alveolar destruction in COPD-like mice. Collectively, our study suggests that CD146 negatively regulates MMP-9 production in macrophages via the S100A9 pathway in COPD.

Targeting S100A9 protein affects mTOR-ER stress signaling and increases venetoclax sensitivity in Acute Myeloid Leukemia.

Acute Myeloid Leukemia (AML) is a heterogeneous disease with limited treatment options and a high demand for novel targeted therapies. Since myeloid-related protein S100A9 is abundantly expressed in AML, we aimed to unravel the therapeutic impact and underlying mechanisms of targeting both intracellular and extracellular S100A9 protein in AML cell lines and primary patient samples. S100A9 silencing in AML cell lines resulted in increased apoptosis and reduced AML cell viability and proliferation. These therapeutic effects were associated with a decrease in mTOR and endoplasmic reticulum stress signaling. Comparable results on AML cell proliferation and mTOR signaling could be observed using the clinically available S100A9 inhibitor tasquinimod. Interestingly, while siRNA-mediated targeting of S100A9 affected both extracellular acidification and mitochondrial metabolism, tasquinimod only affected the mitochondrial function of AML cells. Finally, we found that S100A9-targeting approaches could significantly increase venetoclax sensitivity in AML cells, which was associated with a downregulation of BCL-2 and c-MYC in the combination group compared to single agent therapy. This study identifies S100A9 as a novel molecular target to treat AML and supports the therapeutic evaluation of tasquinimod in venetoclax-based regimens for AML patients.

Effect of Proinflammatory S100A9 Protein on Migration and Proliferation of Microglial Cells.

Alzheimer's disease (AD) is a multifactorial disease affecting aging population worldwide. Neuroinflammation became a focus of research as one of the major pathologic processes relating to the disease onset and progression. Proinflammatory S100A9 is the central culprit in the amyloid-neuroinflammatory cascade implicated in AD and other neurodegenerative diseases. We studied the effect of S100A9 on microglial BV-2 cell proliferation and migration. The responses of BV-2 cells to S100A9 stimulation were monitored in real-time using live cell microscopy, transcriptome sequencing, immunofluorescence staining, western blot analysis, and ELISA. We observed that a low dose of S100A9 promotes migration and proliferation of BV-2 cells. However, acute inflammatory condition (i.e., high S100A9 doses) causes diminished cell viability; it is uncovered that S100A9 activates TLR-4 and TLR-7 signaling pathways, leading to TNF-α and IL-6 expression, which affect BV-2 cell migration and proliferation in a concentration-dependent manner. Interestingly, the effects of S100A9 are not only inhibited by TNF-α and IL-6 antibodies. The addition of amyloid-ß (Aß) 1-40 peptide resumes the capacities of BV-2 cells to the level of low S100A9 concentrations. Based on these results, we conclude that in contrast to the beneficial effects of low S100A9 dose, high S100A9 concentration leads to impaired mobility and proliferation of immune cells, reflecting neurotoxicity at acute inflammatory conditions. However, the formation of Aß plaques may be a natural mechanism that rescues cells from the proinflammatory and cytotoxic effects of S100A9, especially considering that inflammation is one of the primary causes of AD.

Deficiency of S100A8/A9 attenuates pulmonary microvascular leakage in septic mice.

BACKGROUND: We have reported a positive correlation between S100 calcium-binding protein (S100) A8/S100A9 and sepsis-induced lung damage before. However, limited knowledge exists concerning the biological role of S100A8/A9 in pulmonary vascular endothelial barrier dysfunction, as well as the diagnostic value of S100A8/A9 in sepsis. METHODS: Sepsis was induced in C57BL/6J mice and S100A9-knockout (KO) mice through the cecal ligation and puncture (CLP). Pulmonary vascular leakage was determined by measuring extravasated Evans blue (EB). Reverse transcription polymerase chain reaction and the histological score were used to evaluate inflammation and lung injury, respectively. Recombinant S100A8/A9 (rhS100A8/A9) was used to identify the effects of S100A8/A9 on endothelial barrier dysfunction in human umbilical vein endothelial cells (HUVECs). Additionally, the diagnostic value of S100A8/A9 in sepsis was assessed using receiver operating characteristic. RESULTS: S100A8/A9 expression was up-regulated in the lungs of CLP-operated mice. S100A9 KO significantly reversed CLP-induced hypothermia and hypotension, resulting in an improved survival rate. S100A9 KO also decreased the inflammatory response, EB leakage, and histological scores in the lungs of CLP-operated mice. Occludin and VE-cadherin expressions were decreased in the lungs of CLP-operated mice; However, S100A9 KO attenuated this decrease. Moreover, CLP-induced signal transducer and activator of transcription 3 (STAT3) and p38/extracellular signal-regulated kinase (ERK) signalling activation and apoptosis were mitigated by S100A9 KO in lungs. In addition, rhS100A8/A9 administration significantly decreased occludin and VE-cadherin expressions, increased the phosphorylated (p)-ERK/ERK, p-p38/p38, and B-cell leukaemia/lymphoma 2 protein (Bcl-2)-associated X protein/Bcl-2 ratios in HUVECs. CONCLUSION: The present study demonstrated S100A8/A9 aggravated sepsis-induced pulmonary inflammation, vascular permeability, and lung injury. This was achieved, at least partially, by activating the P38/STAT3/ERK signalling pathways. Moreover, S100A8/A9 showed the potential as a biomarker for sepsis diagnosis.

Viaminate Inhibits Propionibacterium Acnes-induced Abnormal Proliferation and Keratinization of HaCat Cells by Regulating the S100A8/S100A9- MAPK Cascade.

BACKGROUND: Viaminate, a vitamin A acid drug developed in China, has been clinically used in acne treatment to regulate epithelial cell differentiation and proliferation, inhibit keratinization, reduce sebum secretion, and control immunological and anti-inflammatory actions; however, the exact method by which it works is unknown. METHODS: In the present study, acne was induced in the ears of rats using Propionibacterium acnes combined with sebum application. RESULTS: After 30 days of treatment with viaminate, the symptoms of epidermal thickening and keratin overproduction in the ears of rats were significantly improved. Transcriptomic analysis of rat skin tissues suggested that viaminate significantly regulated the biological pathways of cellular keratinization. Gene differential analysis revealed that the S100A8 and S100A9 genes were significantly downregulated after viaminate treatment. The results of qPCR and Western blotting confirmed that viaminate inhibited the expression of S100A8 and S100A9 genes and proteins in rat and HaCat cell acne models, while its downstream pathway MAPK (MAPK p38/JNK/ERK1/2) protein expression levels were suppressed. Additional administration of the S100A8 and S100A9 complex protein significantly reversed the inhibitory effect of viaminate on abnormal proliferation and keratinization levels in acne cell models. CONCLUSION: In summary, viaminate can improve acne by modulating S100A8 and S100A9 to inhibit MAPK pathway activation and inhibit keratinocyte proliferation and keratinization levels.

Transcriptomic Analysis of Retinal Gene in Experimental Retinal Detachment Rats and Exploration of S100A9 and TLR4 in Human Vitreous.

PURPOSE: To screen for the differentially expressed genes in experimental retinal detachment rats, and to explore the expression of S100 calcium-binding protein A9 and Toll-like receptor 4 in the vitreous of rhegmatogenous retinal detachment patients. METHODS: Three rats of experimental retinal detachment and three normal rats were enrolled in the study. Transcriptomics (RNAseq) sequencing technology was used to screen differentially expressed genes in the retinas of the experimental retinal detachment group and the normal group. The selected differentially expressed genes for gene ontology and Kyoto Encyclopedia of Genes and Genomes functional enrichment analysis were performed. In addition, the vitreous of 15 patients with rhegmatogenous retinal detachment and six patients with the control group were collected. The expressions of S100 calcium-binding protein A9 and Toll-like receptor 4 were detected by Elisa, and the differences in expression levels were analyzed statistically. RESULTS: A total of 198 differentially expressed genes were screened by RNAseq sequencing, including 118 upregulated genes and 80 downregulated genes. Kyoto Encyclopedia of Genes and Genomes analysis confirmed that the most enriched pathway was the mitogen-activated protein kinase signaling pathway. Compared to the normal group, the expressions of suppressor of cytokine signaling-3, Storkhead box-2, S100 calcium-binding protein A9, Spi-1 proto-oncogene, phosphodiesterase 1B, and kinesin-light chain 1 mRNA in the retinas of the experimental retinal detachment rats were up-regulated, and the expressions of Max interacting protein 1 and the voltage-gated sodium 1 were down-regulated. Compared to the control group, the expressions of S100 calcium-binding protein A9 and Toll-like receptor 4 were upregulated by Elisa in the vitreous humor of rhegmatogenous retinal detachment patients with a statistically significant difference (p all <.05). CONCLUSION: The differentially expressed genes of experimental retinal detachment rats were suppressor of cytokine signaling-3, Storkhead box-2, S100 calcium-binding protein A9, Spi-1 proto-oncogene, phosphodiesterase 1B, kinesin-light chain 1, Max interacting protein 1, voltage-gated sodium 1, etc. The differences of S100 calcium-binding protein A9 and Toll-like receptor 4 expressions between the rhegmatogenous retinal detachment patients and the control group were statistically significant, indicating that they may play a potential role in the inflammatory process of rhegmatogenous retinal detachment.

Prognostic significance of peripheral blood S100A12, S100A8, and S100A9 concentrations in idiopathic pulmonary fibrosis.

BACKGROUND: S100A12, S100A8, and S100A9 are inflammatory disease biomarkers whose functional significance in idiopathic pulmonary fibrosis (IPF) remains unclear. We evaluated the significance of S100A12, S100A8, and S100A9 levels in IPF development and prognosis. METHODS: The dataset was collected from the Gene Expression Omnibus (GEO) database and differentially expressed genes were screened using GEO2R. We conducted a retrospective study of 106 patients with IPF to explore the relationships between different biomarkers and poor outcomes. Pearson's correlation coefficient, Kaplan-Meier, Cox regression, and functional enrichment analyses were used to evaluate relationships between these biomarkers' levels and clinical parameters or prognosis. RESULTS: Serum levels of S100A12, S100A8, and S100A9 were significantly elevated in patients with IPF. The two most significant co-expression genes of S100A12 were S100A8 and S100A9. Patients with levels of S100A12 (median 231.21 ng/mL), S100A9 (median 57.09 ng/mL) or S100A8 (median 52.20 ng/mL), as well as combined elevated S100A12, S100A9, and S100A8 levels, exhibited shorter progression-free survival and overall survival. Serum S100A12 and S100A8, S100A12 and S100A9, S100A9 and S100A8 concentrations also displayed a strong positive correlation (rs2 = 0.4558, rs2 = 0.4558, rs2 = 0.6373; P < 0.001). S100A12 and S100A8/9 concentrations were independent of FVC%, DLCO%, and other clinical parameters (age, laboratory test data, and smoking habit). Finally, in multivariate analysis, the serum levels of S100A12, S100A8, and S100A9 were significant prognostic factors (hazard ratio 1.002, P = 0.032, hazard ratio 1.039, P = 0.001, and hazard ratio 1.048, P = 0.003). CONCLUSIONS: S100A12, S100A8, and S100A9 are promising circulating biomarkers that may aid in determining IPF patient prognosis. Multicenter clinical trials are needed to confirm their clinical value.

Therapeutic S100A8/A9 blockade inhibits myocardial and systemic inflammation and mitigates sepsis-induced myocardial dysfunction.

BACKGROUND AND AIMS: The triggering factors of sepsis-induced myocardial dysfunction (SIMD) are poorly understood and are not addressed by current treatments. S100A8/A9 is a pro-inflammatory alarmin abundantly secreted by activated neutrophils during infection and inflammation. We investigated the efficacy of S100A8/A9 blockade as a potential new treatment in SIMD. METHODS: The relationship between plasma S100A8/A9 and cardiac dysfunction was assessed in a cohort of 62 patients with severe sepsis admitted to the intensive care unit of Linköping University Hospital, Sweden. We used S100A8/A9 blockade with the small-molecule inhibitor ABR-238901 and S100A9-/- mice for therapeutic and mechanistic studies on endotoxemia-induced cardiac dysfunction in mice. RESULTS: In sepsis patients, elevated plasma S100A8/A9 was associated with left-ventricular (LV) systolic dysfunction and increased SOFA score. In wild-type mice, 5 mg/kg of bacterial lipopolysaccharide (LPS) induced rapid plasma S100A8/A9 increase and acute LV dysfunction. Two ABR-238901 doses (30 mg/kg) administered intraperitoneally with a 6 h interval, starting directly after LPS or at a later time-point when LV dysfunction is fully established, efficiently prevented and reversed the phenotype, respectively. In contrast, dexamethasone did not improve cardiac function compared to PBS-treated endotoxemic controls. S100A8/A9 inhibition potently reduced systemic levels of inflammatory mediators, prevented upregulation of inflammatory genes and restored mitochondrial function in the myocardium. The S100A9-/- mice were protected against LPS-induced LV dysfunction to an extent comparable with pharmacologic S100A8/A9 blockade. The ABR-238901 treatment did not induce an additional improvement of LV function in the S100A9-/- mice, confirming target specificity. CONCLUSION: Elevated S100A8/A9 is associated with the development of LV dysfunction in severe sepsis patients and in a mouse model of endotoxemia. Pharmacological blockade of S100A8/A9 with ABR-238901 has potent anti-inflammatory effects, mitigates myocardial dysfunction and might represent a novel therapeutic strategy for patients with severe sepsis.

Activation of Toll-like receptor 4 by thyroid hormone triggers abnormal B-cell activation.

OBJECTIVE: Breakdown of tolerance and abnormal activation of B cells is an important mechanism in the pathogenesis of Graves' disease (GD). High levels of thyroid hormones (THs) play important roles in GD progression. However, the interactions between THs and abnormal activation of B cells remain elusive. This study aimed to explore the effect of high levels of THs on TLR4 expression and abnormal B cell differentiation. MATERIALS AND METHODS: Blood samples were collected from patients with GD and healthy controls (HCs) to evaluate the frequency of B cells, their subsets, and TLR4 expression in B cells. A high-level T3 mouse model was used to study the interaction between THs and the TLR4 signalling pathway. RESULTS: We found that the frequencies of CD19+ , CD19+ TLR4+ , CD19+ CD86+ , and CD19+ CD138+ B cells were significantly higher, as were the expression levels of MRP8/MRP14 and MRP6 and MRP8, MRP14, and MRP6 messenger RNA (mRNA) in peripheral blood mononuclear cells in patients with GD. In high-level T3 mice models, the serum MRP8/MRP14 and MRP6 levels and the TLR4 mRNA expression in PBMCs were significantly higher. TLR4 mRNA, protein expression, and cytokines downstream of TLR4, such as myeloid differentiation factor 88 (MyD88) and nuclear transcription factor-κB, were also increased in mouse spleen mononuclear cells. CONCLUSION: The present study indicated that high levels of T3 can induce abnormal differentiation and activation of B cells by promoting TLR4 overexpression and provide novel insights into the roles of THs in the pathogenesis of GD.

CD74 Promotes a Pro-Inflammatory Tumor Microenvironment by Inducing S100A8 and S100A9 Secretion in Pancreatic Cancer.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive form of pancreatic cancer with a poor prognosis and low survival rates. The prognostic and predictive biomarkers of PDAC are still largely unknown. The receptor CD74 was recently identified as a regulator of oncogenic properties in various cancers. However, the precise molecular mechanism of CD74 action in PDAC remains little understood. We investigated the role of CD74 by silencing CD74 in the pancreatic cancer cell line Capan-1. CD74 knockdown led to reductions in cell proliferation, migration, and invasion and increased apoptosis. Moreover, silencing CD74 resulted in the decreased expression and secretion of S100A8 and S100A9. An indirect co-culture of fibroblasts and tumor cells revealed that fibroblasts exposed to conditioned media from CD74 knockdown cells exhibited a reduced expression of inflammatory cytokines, suggesting a role of CD74 in influencing cytokine secretion in the tumor microenvironment. Overall, our study provides valuable insights into the critical role of CD74 in regulating the oncogenic properties of pancreatic cancer cells and its influence on the expression and secretion of S100A8 and S100A9. Taken together, these findings indicate CD74 as a potential diagnostic biomarker and therapeutic target for pancreatic cancer.

Nuclear proinflammatory cytokine S100A9 enhances expression of human papillomavirus oncogenes via transcription factor TEAD1.

Transcription of the human papillomavirus (HPV) oncogenes, E6 and E7, is regulated by the long control region (LCR) of the viral genome. Although various transcription factors have been reported to bind to the LCR, little is known about the transcriptional cofactors that modulate HPV oncogene expression in association with these transcription factors. Here, we performed in vitro DNA-pulldown purification of nuclear proteins in cervical cancer cells, followed by proteomic analyses to identify transcriptional cofactors that bind to the HPV16 LCR via the transcription factor TEAD1. We detected the proinflammatory cytokine S100A9 that localized to the nucleus of cervical cancer cells and associated with the LCR via direct interaction with TEAD1. Nuclear S100A9 levels and its association with the LCR were increased in cervical cancer cells by treatment with a proinflammatory phorbol ester. Knockdown of S100A9 decreased HPV oncogene expression and reduced the growth of cervical cancer cells and their susceptibility to cisplatin, whereas forced nuclear expression of S100A9 using nuclear localization signals exerted opposite effects. Thus, we conclude that nuclear S100A9 binds to the HPV LCR via TEAD1 and enhances viral oncogene expression by acting as a transcriptional coactivator. IMPORTANCE Human papillomavirus (HPV) infection is the primary cause of cervical cancer, and the viral oncogenes E6 and E7 play crucial roles in carcinogenesis. Although cervical inflammation contributes to the development of cervical cancer, the molecular mechanisms underlying the role of these inflammatory responses in HPV carcinogenesis are not fully understood. Our study shows that S100A9, a proinflammatory cytokine, is induced in the nucleus of cervical cancer cells by inflammatory stimuli, and it enhances HPV oncogene expression by acting as a transcriptional coactivator of TEAD1. These findings provide new molecular insights into the relationship between inflammation and viral carcinogenesis.

S100A9 Regulated M1/M2 Macrophage Polarization in Interleukin-10-Induced Promotion of Malignant Pleural Effusion.

Interleukin-10 (IL-10) promotes the formation and development of malignant pleural effusion (MPE). Previous studies have elucidated the pathogenesis from the view of the immune-regulation function of CD4+ T-cells. However, the underlying mechanism is still not fully understood. In this study, our results showed that IL-10 deficiency reduced the percentage of macrophages in mouse MPE and regulated M1/M2 polarization in vivo and in vitro. The migration capacity of tumor cells was suppressed, and apoptosis was promoted when tumor cells were cocultured with MPE macrophages in the absence of IL-10. Messenger RNA sequencing of MPE macrophages showed that S100A9 was downregulated in IL-10-/- mice. Bone marrow-derived macrophages obtained from wild-type mice transfected with S100A9-specific small interfering RNAs (siRNAs) also showed less M2 and more M1 polarization than those from the siRNA control group. Furthermore, downregulation of S100A9 using S100A9-specific siRNA suppressed MPE development, decreased macrophages, and modulated macrophage polarization in MPE in vivo. In conclusion, S100A9 plays a vital role in the process of IL-10 deficiency-mediated MPE suppression by regulating M1/M2 polarization, thus influencing the tumor-migration capacity and apoptosis. This could result in clinically applicable strategies to inhibit the formation of MPE by regulating the polarization of MPE macrophages.

S100A9 is indispensable for survival of pneumococcal pneumonia in mice.

S100A8/A9 has important immunomodulatory roles in antibacterial defense, but its relevance in focal pneumonia caused by Streptococcus pneumoniae (S. pneumoniae) is understudied. We show that S100A9 was significantly increased in BAL fluids of patients with bacterial but not viral pneumonia and correlated with procalcitonin and sequential organ failure assessment scores. Mice deficient in S100A9 exhibited drastically elevated Zn2+ levels in lungs, which led to bacterial outgrowth and significantly reduced survival. In addition, reduced survival of S100A9 KO mice was characterized by excessive release of neutrophil elastase, which resulted in degradation of opsonophagocytically important collectins surfactant proteins A and D. All of these features were attenuated in S. pneumoniae-challenged chimeric WT→S100A9 KO mice. Similarly, therapy of S. pneumoniae-infected S100A9 KO mice with a mutant S100A8/A9 protein showing increased half-life significantly decreased lung bacterial loads and lung injury. Collectively, S100A9 controls central antibacterial immune mechanisms of the lung with essential relevance to survival of pneumococcal pneumonia. Moreover, S100A9 appears to be a promising biomarker to distinguish patients with bacterial from those with viral pneumonia. Trial registration: Clinical Trials register (DRKS00000620).