Macrophage mannose receptor CD206 targeting of fluoride-18 labeled mannosylated dextran: A validation study in mice.

PURPOSE: Aluminum fluoride-18-labeled 1,4,7-triazacyclononane-1,4,7-triacetic acid-conjugated mannosylated dextran derivative (Al[18F]F-NOTA-D10CM) is a new tracer for PET imaging. We report here on in vitro and in vivo validation of the tracer's ability to target the macrophage mannose receptor CD206. METHODS: First, the uptake of intravenously (i.v.) administered Al[18F]F-NOTA-D10CM was compared between wild-type (WT) and CD206-/- knockout (KO) mice. C57BL/6N mice were injected with complete Freund's adjuvant (CFA) in the left hind leg and the uptake of Al[18F]F-NOTA-D10CM after i.v. or intradermal (i.d.) injection was studied at 5 and 14 days after CFA induction of inflammation. Healthy C57BL/6N mice were studied as controls. Mice underwent PET/CT on consecutive days with [18F]FDG, i.v. Al[18F]F-NOTA-D10CM, and i.d. Al[18F]F-NOTA-D10CM. After the last imaging, Al[18F]F-NOTA-D10CM was i.v. injected for an ex vivo biodistribution study and autoradiography of inflamed tissues. Blood plasma samples were analyzed using high-performance liquid chromatography. To evaluate the specificity of Al[18F]F-NOTA-D10CM binding, an in vitro competitive displacement study was performed on inflamed tissue sections using autoradiography. CD206 expression was assessed by immunohistochemical staining. RESULTS: Compared with WT mice, the uptake of Al[18F]F-NOTA-D10CM was significantly lower in several CD206-/- KO mice tissues, including liver (SUV 8.21 ± 2.51 vs. 1.06 ± 0.16, P < 0.001) and bone marrow (SUV 1.63 ± 0.37 vs. 0.22 ± 0.05, P < 0.0001). The uptake of i.v. injected Al[18F]F-NOTA-D10CM was significantly higher in inflamed ankle joint (SUV 0.48 ± 0.13 vs. 0.18 ± 0.05, P < 0.0001) and inflamed foot pad skin (SUV 0.41 ± 0.10 vs. 0.04 ± 0.01, P < 0.0001) than in the corresponding tissues in healthy mice. The i.d.-injected Al[18F]F-NOTA-D10CM revealed differences between CFA-induced lymph node activation and lymph nodes in healthy mice. Ex vivo γ-counting, autoradiography, and immunohistochemistry supported the results, and a decrease of ~ 80% in the binding of Al[18F]F-NOTA-D10CM in the displacement study with excess NOTA-D10CM confirmed that tracer binding was specific. At 60 min after i.v. injection, an average 96.70% of plasma radioactivity was derived from intact Al[18F]F-NOTA-D10CM, indicating good in vivo stability. The uptake of Al[18F]F-NOTA-D10CM into inflamed tissues was positively associated with the area percentage of CD206-positive staining. CONCLUSION: The uptake of mannosylated dextran derivative Al[18F]F-NOTA-D10CM correlated with CD206 expression and the tracer appears promising for inflammation imaging.

Characterization of Immune Cell Populations of Cutaneous Neurofibromas in Neurofibromatosis 1.

Cutaneous neurofibromas (cNFs) are characteristic of neurofibromatosis 1 (NF1), yet their immune microenvironment is incompletely known. A total of 61 cNFs from 10 patients with NF1 were immunolabeled for different types of T cells and macrophages, and the cell densities were correlated with clinical characteristics. Eight cNFs and their overlying skin were analyzed for T cell receptor CDR domain sequences, and mass spectrometry of 15 cNFs and the overlying skin was performed to study immune-related processes. Intratumoral T cells were detected in all cNFs. Tumors from individuals younger than the median age of the study participants (33 years), growing tumors, and tumors smaller than the data set median showed increased T cell density. Most samples displayed intratumoral or peritumoral aggregations of CD3-positive cells. T cell receptor sequencing demonstrated that the skin and cNFs host distinct T cell populations, whereas no dominant cNF-specific T cell clones were detected. Unique T cell clones were fewer in cNFs than in skin, and mass spectrometry suggested lower expression of proteins related to T cell-mediated immunity in cNFs than in skin. CD163-positive cells, suggestive of M2 macrophages, were abundant in cNFs. Human cNFs have substantial T cell and macrophage populations that may be tumor-specific.

The senescent secretome drives PLVAP expression in cultured human hepatic endothelial cells to promote monocyte transmigration.

Liver sinusoidal endothelial cells (LSEC) undergo significant phenotypic change in chronic liver disease (CLD), and yet the factors that drive this process and the impact on their function as a vascular barrier and gatekeeper for immune cell recruitment are poorly understood. Plasmalemma-vesicle-associated protein (PLVAP) has been characterized as a marker of LSEC in CLD; notably we found that PLVAP upregulation strongly correlated with markers of tissue senescence. Furthermore, exposure of human LSEC to the senescence-associated secretory phenotype (SASP) led to a significant upregulation of PLVAP. Flow-based assays demonstrated that SASP-driven leukocyte recruitment was characterized by paracellular transmigration of monocytes while the majority of lymphocytes migrated transcellularly. Knockdown studies confirmed that PLVAP selectively supported monocyte transmigration mediated through PLVAP's impact on LSEC permeability by regulating phospho-VE-cadherin expression and endothelial gap formation. PLVAP may therefore represent an endothelial target that selectively shapes the senescence-mediated immune microenvironment in liver disease.

Prognostic Values of Tissue and Serum Angiogenic Growth Factors Depend on the Phenotypic Subtypes of Colorectal Cancer.

We classified colorectal cancer (CRC) patients into four phenotypic subgroups and investigated the prognostic value of angiogenic growth factors across subgroups. Preoperative serum concentrations and tissue expressions of VEGF, bFGF, and PDGF-bb were determined among 322 CRC patients. We classified patients into phenotypic subgroups (immune, canonical, metabolic, and mesenchymal) according to a method described in our earlier work. Among the metabolic subgroup, patients with high serum concentrations of VEGF, bFGF, or PDGF-bb exhibited a significantly improved prognosis. Moreover, those with high VEGF tissue expressions exhibited a significantly improved prognosis among patients in the metabolic subgroup. Among immune patients, a high VEGF serum expression is associated with a worse prognosis. A high serum bFGF concentration is associated with a favorable prognostic factor among patients with a canonical tumor phenotype. A high PDGF-bb tissue expression is associated with non-metastasized disease and with the immune, canonical, and metabolic subtypes. To our knowledge, this is the first study to show that the prognostic value of angiogenic growth factors differs between phenotypic subtypes.

Serum cytokine profiles in patients with pancreatic cancer and chronic pancreatitis.

BACKGROUND: Chronic pancreatitis (CP) may cause tumor-like lesions, creating a challenge in distinguishing between CP and pancreatic ductal adenocarcinoma (PDAC) in a patient. Given that invasive surgery is a standard cancer treatment, we aimed to examine whether a noninvasive diagnostic tool utilizing serum cytokines could safely differentiate between PDAC and CP. METHODS: A pre-operative serum panel comprising 48 inflammatory cytokines, CA19-9, and C-reactive protein (CRP) was analyzed, consisting of 231 patients, 186 with stage I-III PDAC and 45 with CP. We excluded PDAC patients who underwent neoadjuvant therapy and those CP patients with other active malignancies. The laboratory variables most associated with PDAC diagnosis were assessed using logistic regression and selected using the lasso method. RESULTS: The cytokines CTACK, GRO-α, and ß-NGF were selected alongside CA19-9 and CRP for our differential diagnostic model. The area under the curve (AUC) for our differential diagnostic model was 0.809 (95% confidence interval [CI] 0.738-0.880), compared with 0.791 (95% CI 0.728-0.854) for CA19-9 alone (not significant). CONCLUSIONS: We found that inflammatory cytokines CTACK, GRO-α, and ß-NGF alongside CA19-9 and CRP may help distinguish PDAC from CP.

The interplay between inflammatory cytokines and cardiometabolic disease: bi-directional mendelian randomisation study.

Objective: To leverage large scale genetic association data to investigate the interplay between circulating cytokines and cardiometabolic traits, and thus identifying potential therapeutic targets. Design: Bi-directional Mendelian randomisation study. Setting: Genome-wide association studies from three Finnish cohorts (Northern Finland Birth Cohort 1966, Young Finns Study, or FINRISK study), and genetic association summary statistics pooled from observational studies for expression quantitative trait loci and cardiometabolic traits. Participants: Data for 47 circulating cytokines in 13 365 individuals from genome-wide association studies, summary statistic data for up to 21 735 individuals on circulating cytokines, summary statistic gene expression data across 49 tissues in 838 individuals, and summary statistic data for up to 1 320 016 individuals on cardiometabolic traits. Interventions: Relations between circulating cytokines and cardiovascular, anthropometric, lipid, or glycaemic traits (coronary artery disease, stroke, type 2 diabetes mellitus, body mass index, waist circumference, waist to hip ratio, systolic blood pressure, glycated haemoglobin, high density lipoprotein cholesterol, low density lipoprotein cholesterol, total cholesterol, triglycerides, C reactive protein, glucose, fasting insulin, and lifetime smoking). Main outcome methods: Genetic instrumental variables that are biologically plausible for the circulating cytokines were generated. The effects of cardiometabolic risk factors on concentrations of circulating cytokines, circulating cytokines on other circulating cytokines, and circulating cytokines on cardiometabolic outcomes were investigated. Results: Genetic evidence (mendelian randomisation P<0.0011) suggests that higher body mass index, waist circumference, smoking, higher concentrations of lipids, and systolic blood pressure increase circulating concentrations of several inflammatory cytokines and C reactive protein. Evidence for causal relations (mendelian randomisation P<0.0011) were noted between circulating cytokines, including a key role of vascular endothelial growth factor on influencing the concentrations of 10 other cytokines. Both mendelian randomisation (P<0.05) and colocalisation (posterior probability >0.5) suggested that coronary artery disease risk is increased by higher concentrations of circulating tumour necrosis factor related apoptosis-inducing ligand (TRAIL), interleukin-1 receptor antagonist (IL1RA), and macrophage colony-stimulating factor (MCSF). Conclusion: This study offers insight into inflammatory mediators of cardiometabolic risk factors, cytokine signalling cascades, and effects of circulating cytokines on different cardiometabolic outcomes.

Inflammatory proteomics profiling for prediction of incident atrial fibrillation.

OBJECTIVE: Atrial fibrillation (AF) has emerged as a common condition in older adults. Cardiovascular risk factors only explain about 50% of AF cases. Inflammatory biomarkers may help close this gap as inflammation can alter atrial electrophysiology and structure. This study aimed to determine a cytokine biomarker profile for this condition in the community using a proteomics approach. METHODS: This study uses cytokine proteomics in participants of the Finnish population-based FINRISK cohort studies 1997/2002. Risk models for 46 cytokines were developed to predict incident AF using Cox regressions. Furthermore, the association of participants' C reactive protein (CRP) and N-terminal pro B-type natriuretic peptide (NT-proBNP) concentrations with incident AF was examined. RESULTS: In 10 744 participants (mean age of 50.9 years, 51.3% women), 1246 cases of incident AF were observed (40.5% women). The main analyses, adjusted for participants' sex and age, suggested that higher concentrations of macrophage inflammatory protein-1ß (HR=1.11; 95% CI 1.04, 1.17), hepatocyte growth factor (HR=1.12; 95% CI 1.05, 1.19), CRP (HR=1.17; 95% CI 1.10, 1.24) and NT-proBNP (HR=1.58; 95% CI 1.45, 1.71) were associated with increased risk of incident AF. In further clinical variable-adjusted models, only NT-proBNP remained statistically significant. CONCLUSION: Our study confirmed NT-proBNP as a strong predictor for AF. Observed associations of circulating inflammatory cytokines were primarily explained by clinical risk factors and did not improve risk prediction. The potential mechanistic role of inflammatory cytokines measured in a proteomics approach remains to be further elucidated.

Effects of altered salt intake and diet on cytokines in humans: A 20-week randomized cross-over intervention study.

High sodium concentration alters leukocyte activation, and in particular T-helper (Th) lymphocyte polarization, and drives the development of autoimmune diseases in mouse studies. Similar results have been obtained with human leukocytes under in vitro settings and in few observational studies. Therefore, salt has been implicated as a risk factor for autoimmune diseases. Here, we examined whether physiologically relevant changes in salt intake or diet alter cytokine concentrations. In a 20-wk double-blinded, placebo-controlled study 106 participants were randomized to Habitual and Healthy Nordic diets, and further to Usual Sodium and Reduced Sodium intake groups using a cross-over setup. Plasma concentrations of 45 cytokines were measured at three different time-points using a multiplex assay. Repeated analyses of covariance revealed that high salt ingestion (or changes in the diet) did not induce significant changes in any of the signature cytokines controlling Th1, Th2 or Th17 polarization. Several other pro-inflammatory interleukins, chemokines and growth factors were also unaffected by the level of salt intake or changes in the diet. We conclude that in humans clinically relevant changes in salt intake or diet do not have reflections on the systemic concentrations of pro-inflammatory cytokines in vivo.

Lymph node lymphatic endothelial cells as multifaceted gatekeepers in the immune system.

Single-cell technologies have recently allowed the identification of multiple lymphatic endothelial cell (LEC) subsets in subcapsular, paracortical, medullary, and other lymph node (LN) sinus systems in mice and humans. New analyses show that LECs serve key immunological functions in the LN stroma during immune responses. We discuss the roles of different LEC types in guiding leukocyte and cancer cell trafficking to and from the LN parenchyma, in capturing microbes, and in transporting, presenting, and storing lymph-borne antigens in distinct types of lymphatic sinuses. We underscore specific adaptations of human LECs and raise unanswered questions concerning LEC functions in human disease. Despite our limited understanding of human lymphatics - hampering clinical translation in inflammation and metastasis - we support the potential of LN LECs as putative targets for boosting/inhibiting immunoreactivity.

Inflammatory Diseases, Inflammatory Biomarkers, and Alzheimer Disease: An Observational Analysis and Mendelian Randomization.

BACKGROUND AND OBJECTIVES: Whether chronic autoimmune inflammatory diseases causally affect the risk of Alzheimer disease (AD) is controversial. We characterized the relationship between inflammatory diseases and risk of AD and explored the role of circulating inflammatory biomarkers in the relationships between inflammatory diseases and AD. METHODS: We performed observational analyses for chronic autoimmune inflammatory diseases and risk of AD using data from 2,047,513 participants identified in the UK Clinical Practice Research Datalink (CPRD). Using data of a total of more than 1,100,000 individuals from 15 large-scale genome-wide association study data sets, we performed 2-sample Mendelian randomizations (MRs) to investigate the relationships between chronic autoimmune inflammatory diseases, circulating inflammatory biomarker levels, and risk of AD. RESULTS: Cox regression models using CPRD data showed that the overall incidence of AD was higher among patients with inflammatory bowel disease (hazard ratio [HR] 1.17; 95% CI 1.15-1.19; p = 2.1 × 10-4), other inflammatory polyarthropathies and systematic connective tissue disorders (HR 1.13; 95% CI 1.12-1.14; p = 8.6 × 10-5), psoriasis (HR 1.13; 95% CI 1.10-1.16; p = 2.6 × 10-4), rheumatoid arthritis (HR 1.08; 95% CI 1.06-1.11; p = 4.0 × 10-4), and multiple sclerosis (HR 1.06; 95% CI 1.04-1.07; p = 2.8 × 10-4) compared with the age (±5 years) and sex-matched comparison groups free from all inflammatory diseases under investigation. Bidirectional MR analysis identified relationships between chronic autoimmune inflammatory diseases and circulating inflammatory biomarkers. Particularly, circulating monokine induced by gamma interferon (MIG) level was suggestively associated with a higher risk of AD (odds ratio from inverse variance weighted [ORIVW] 1.23; 95% CI 1.06-1.42; p IVW = 0.007) and lower risk of Crohn disease (ORIVW 0.73; 95% CI -0.62 to 0.86; p IVW = 1.3 × 10-4). Colocalization supported a common causal single nucleotide polymorphism for MIG and Crohn disease (posterior probability = 0.74), but not AD (posterior probability = 0.03). Using a 2-sample MR approach, genetically predicted risks of inflammatory diseases were not associated with higher AD risk. DISCUSSION: Our data suggest that the association between inflammatory diseases and risk of AD is unlikely to be causal and may be a result of confounding. In support, although inflammatory biomarkers showed evidence for causal associations with inflammatory diseases, evidence was weak that they affected both inflammatory disease and AD.

A plasma metabolite score of three eicosanoids predicts incident type 2 diabetes: a prospective study in three independent cohorts.

INTRODUCTION: Peptide markers of inflammation have been associated with the development of type 2 diabetes. The role of upstream, lipid-derived mediators of inflammation such as eicosanoids, remains less clear. The aim of this study was to examine whether eicosanoids are associated with incident type 2 diabetes. RESEARCH DESIGN & METHODS: In the FINRISK (Finnish Cardiovascular Risk Study) 2002 study, a population-based sample of Finnish men and women aged 25-74 years, we used directed, non-targeted liquid chromatography-mass spectrometry to identify 545 eicosanoids and related oxylipins in the participants' plasma samples (n=8292). We used multivariable-adjusted Cox regression to examine associations between eicosanoids and incident type 2 diabetes. The significant independent findings were replicated in the Framingham Heart Study (FHS, n=2886) and DIetary, Lifestyle and Genetic determinants of Obesity and Metabolic syndrome (DILGOM) 2007 (n=3905). Together, these three cohorts had 1070 cases of incident type 2 diabetes. RESULTS: In the FINRISK 2002 cohort, 76 eicosanoids were associated individually with incident type 2 diabetes. We identified three eicosanoids independently associated with incident type 2 diabetes using stepwise Cox regression with forward selection and a Bonferroni-corrected inclusion threshold. A three-eicosanoid risk score produced an HR of 1.56 (95% CI 1.41 to 1.72) per 1 SD increment for risk of incident diabetes. The HR for comparing the top quartile with the lowest was 2.80 (95% CI 2.53 to 3.07). In the replication analyses, the three-eicosanoid risk score was significant in FHS (HR 1.24 (95% CI 1.10 to 1.39, p<0.001)) and directionally consistent in DILGOM (HR 1.12 (95% CI 0.99 to 1.27, p=0.07)). Meta-analysis of the three cohorts yielded a pooled HR of 1.31 (95% CI 1.05 to 1.56). CONCLUSIONS: Plasma eicosanoid profiles predict incident type 2 diabetes and the clearest signals replicate in three independent cohorts. Our findings give new information on the biology underlying type 2 diabetes and suggest opportunities for early identification of people at risk.

Single-cell characterization of leukemic and non-leukemic immune repertoires in CD8+ T-cell large granular lymphocytic leukemia.

T cell large granular lymphocytic leukemia (T-LGLL) is a rare lymphoproliferative disorder of mature, clonally expanded T cells, where somatic-activating STAT3 mutations are common. Although T-LGLL has been described as a chronic T cell response to an antigen, the function of the non-leukemic immune system in this response is largely uncharacterized. Here, by utilizing single-cell RNA and T cell receptor profiling (scRNA+TCRαß-seq), we show that irrespective of STAT3 mutation status, T-LGLL clonotypes are more cytotoxic and exhausted than healthy reactive clonotypes. In addition, T-LGLL clonotypes show more active cell communication than reactive clones with non-leukemic immune cells via costimulatory cell-cell interactions, monocyte-secreted proinflammatory cytokines, and T-LGLL-clone-secreted IFNγ. Besides the leukemic repertoire, the non-leukemic T cell repertoire in T-LGLL is also more mature, cytotoxic, and clonally restricted than in other cancers and autoimmune disorders. Finally, 72% of the leukemic T-LGLL clonotypes share T cell receptor similarities with their non-leukemic repertoire, linking the leukemic and non-leukemic repertoires together via possible common target antigens. Our results provide a rationale to prioritize therapies that target the entire immune repertoire and not only the T-LGLL clonotype.

Circulating inflammatory cytokines and risk of five cancers: a Mendelian randomization analysis.

BACKGROUND: Epidemiological and experimental evidence has linked chronic inflammation to cancer aetiology. It is unclear whether associations for specific inflammatory biomarkers are causal or due to bias. In order to examine whether altered genetically predicted concentration of circulating cytokines are associated with cancer development, we performed a two-sample Mendelian randomisation (MR) analysis. METHODS: Up to 31,112 individuals of European descent were included in genome-wide association study (GWAS) meta-analyses of 47 circulating cytokines. Single nucleotide polymorphisms (SNPs) robustly associated with the cytokines, located in or close to their coding gene (cis), were used as instrumental variables. Inverse-variance weighted MR was used as the primary analysis, and the MR assumptions were evaluated in sensitivity and colocalization analyses and a false discovery rate (FDR) correction for multiple comparisons was applied. Corresponding germline GWAS summary data for five cancer outcomes (breast, endometrial, lung, ovarian, and prostate), and their subtypes were selected from the largest cancer-specific GWASs available (cases ranging from 12,906 for endometrial to 133,384 for breast cancer). RESULTS: There was evidence of inverse associations of macrophage migration inhibitory factor with breast cancer (OR per SD = 0.88, 95% CI 0.83 to 0.94), interleukin-1 receptor antagonist with endometrial cancer (0.86, 0.80 to 0.93), interleukin-18 with lung cancer (0.87, 0.81 to 0.93), and beta-chemokine-RANTES with ovarian cancer (0.70, 0.57 to 0.85) and positive associations of monokine induced by gamma interferon with endometrial cancer (3.73, 1.86 to 7.47) and cutaneous T-cell attracting chemokine with lung cancer (1.51, 1.22 to 1.87). These associations were similar in sensitivity analyses and supported in colocalization analyses. CONCLUSIONS: Our study adds to current knowledge on the role of specific inflammatory biomarker pathways in cancer aetiology. Further validation is needed to assess the potential of these cytokines as pharmacological or lifestyle targets for cancer prevention.

Pancreatic cancer survival prediction via inflammatory serum markers.

BACKGROUND: For prognostic evaluation of pancreatic ductal adenocarcinoma (PDAC), the only well-established serum marker is carbohydrate antigen CA19-9. To improve the accuracy of survival prediction, we tested the efficacy of inflammatory serum markers. METHODS: A preoperative serum panel comprising 48 cytokines plus high-sensitivity CRP (hs-CRP) was analyzed in 173 stage I-III PDAC patients. Analysis of the effect of serum markers on survival utilized the Cox regression model, with the most promising cytokines chosen with the aid of the lasso method. We formed a reference model comprising age, gender, tumor stage, adjuvant chemotherapy status, and CA19-9 level. Our prognostic study model incorporated these data plus hs-CRP and the cytokines. We constructed time-dependent ROC curves and calculated an integrated time-averaged area under the curve (iAUC) for both models from 1 to 10 years after surgery. RESULTS: Hs-CRP and the cytokines CTACK, MIF, IL-1ß, IL-3, GRO-α, M-CSF, and SCF, were our choices for the prognostic study model, in which the iAUC was 0.837 (95% CI 0.796-0.902), compared to the reference model's 0.759 (95% CI 0.691-0.836, NS). These models divided the patients into two groups based on the maximum value of Youden's index at 7.5 years. In our study model, 60th percentile survival times were 4.5 (95% CI 3.7-NA) years (predicted high-survival group, n = 34) and 1.3 (95% CI 1.0-1.7) years (predicted low-survival group, n = 128), log rank p < 0.001. By the reference model, the 60th percentile survival times were 2.8 (95% CI 2.1-4.4) years (predicted high-survival group, n = 44) and 1.3 (95% CI 1.0-1.7) years (predicted low-survival group, n = 118), log rank p < 0.001. CONCLUSION: Hs-CRP and the seven cytokines added to the reference model including CA19-9 are potential prognostic factors for improved survival prediction for PDAC patients.

Vascular adhesion protein-1 defines a unique subpopulation of human hematopoietic stem cells and regulates their proliferation.

Although the development of hematopoietic stem cells (HSC) has been studied in great detail, their heterogeneity and relationships to different cell lineages remain incompletely understood. Moreover, the role of Vascular Adhesion Protein-1 in bone marrow hematopoiesis has remained unknown. Here we show that VAP-1, an adhesin and a primary amine oxidase producing hydrogen peroxide, is expressed on a subset of human HSC and bone marrow vasculature forming a hematogenic niche. Bulk and single-cell RNAseq analyses reveal that VAP-1+ HSC represent a transcriptionally unique small subset of differentiated and proliferating HSC, while VAP-1- HSC are the most primitive HSC. VAP-1 generated hydrogen peroxide acts via the p53 signaling pathway to regulate HSC proliferation. HSC expansion and differentiation into colony-forming units are enhanced by inhibition of VAP-1. Contribution of VAP-1 to HSC proliferation was confirmed with mice deficient of VAP-1, mice expressing mutated VAP-1 and using an enzyme inhibitor. In conclusion, VAP-1 expression allows the characterization and prospective isolation of a new subset of human HSC. Since VAP-1 serves as a check point-like inhibitor in HSC differentiation, the use of VAP-1 inhibitors enables the expansion of HSC.

Head and neck squamous cell carcinoma cell lines have an immunomodulatory effect on macrophages independent of hypoxia and toll-like receptor 9.

BACKGROUND: A low tissue oxygen level, < 1% O2, is a typical characteristic inside of solid tumors in head and neck cancer (HNSCC) affecting a wide array of cell populations, such as macrophages. However, the mechanisms of how hypoxia influences macrophages are not yet fully elucidated. Our research aimed to study the effect of soluble mediators produced by hypoxic cancer cells on macrophage polarization. Furthermore, we studied the effect of a hypoxic microenvironment on the expression of tumorigenic toll-like receptor 9 (TLR9) and the consecutive macrophage polarization. METHODS: Conditioned media (CMNOX or CMHOX) from cell lines UT-SCC-8, UT-SCC-74A, FaDu, MDA-MB-231 and HaCat cultured under normoxic (21% O2) and hypoxic (1% O2) conditions were used to polarize human monocyte-derived macrophages. Macrophage polarization was measured by flow cytometry and the production of cytokine mRNA using Taqman qPCR. To study the role of TLR9 in macrophage polarization, the lentiviral CRISPR/Cas9 method was used to establish a stable FaDuTLR9def clone. RESULTS: Our results demonstrate that the soluble mediators produced by the cancer cells under normoxia polarize macrophages towards a hybridized M1/M2a/M2c phenotype. Furthermore, the results suggest that hypoxia has a limited role in altering the array of cancer-produced soluble factors affecting macrophage polarization and cytokine production. Our data also indicates that increased expression of TLR9 due to hypoxia in malignant cells does not markedly influence the polarization of macrophages. TLR9 transcriptional response to hypoxia is dissimilar to a HIF1-α-regulated LDH-A. This may indicate a context-dependent expression of TLR9 under hypoxia. CONCLUSIONS: HNSCC cell lines affect both macrophage activity (polarization) and functionality (cytokines), but with exception to iNOS expression, the effects appear independent of hypoxia and TLR9.

Single-Cell Proteomics Reveals the Defined Heterogeneity of Resident Macrophages in White Adipose Tissue.

Adipose tissue macrophages (ATMs) regulate homeostasis and contribute to the metabolically harmful chronic inflammation in obese individuals. While evident heterogeneity of resident ATMs has been described previously, their phenotype, developmental origin, and functionality remain inconsistent. We analyzed white adipose tissue (WAT) during homeostasis and diet interventions using comprehensive and unbiased single-cell mass cytometry and genetic lineage tracking models. We now provide a uniform definition of individual subsets of resident ATMs. We show that in lean mice, WAT co-harbors eight kinetically evolving CD206+ macrophage subpopulations (defined by TIM4, CD163, and MHC II) and two CD206- macrophage subpopulations. TIM4-CD163+, TIM4-CD163- and CD206- macrophage populations are largely bone marrow-derived, while the proliferating TIM4+CD163+ subpopulation is of embryonic origin. All macrophage subtypes are active in phagocytosis, endocytosis, and antigen processing in vitro, whereas TIM4+CD163+ cells are superior in scavenging in vivo. A high-fat diet induces massive infiltration of CD206- macrophages and selective down-regulation of MHC II on TIM4+ macrophages. These changes are reversed by dietary intervention. Thus, the developmental origin and environment jointly regulate the functional malleability of resident ATMs.

Embryonic macrophages function during early life to determine invariant natural killer T cell levels at barrier surfaces.

It is increasingly recognized that immune development within mucosal tissues is under the control of environmental factors during early life. However, the cellular mechanisms that underlie such temporally and regionally restrictive governance of these processes are unclear. Here, we uncover an extrathymic pathway of immune development within the colon that is controlled by embryonic but not bone marrow-derived macrophages, which determines the ability of these organs to receive invariant natural killer T (iNKT) cells and allow them to establish local residency. Consequently, early-life perturbations of fetal-derived macrophages result in persistent decreases of mucosal iNKT cells and is associated with later-life susceptibility or resistance to iNKT cell-associated mucosal disorders. These studies uncover a host developmental program orchestrated by ontogenically distinct macrophages that is regulated by microbiota, and they reveal an important postnatal function of macrophages that emerge in fetal life.

A prognostic model for colorectal cancer based on CEA and a 48-multiplex serum biomarker panel.

Mortality in colorectal cancer (CRC) remains high, resulting in 860,000 deaths annually. Carcinoembryonic antigen is widely used in clinics for CRC patient follow-up, despite carrying a limited prognostic value. Thus, an obvious need exists for multivariate prognostic models. We analyzed 48 biomarkers using a multiplex immunoassay panel in preoperative serum samples from 328 CRC patients who underwent surgery at Helsinki University Hospital between 1998 and 2003. We performed a multivariate prognostic forward-stepping background model based on basic clinicopathological data, and a multivariate machine-learned prognostic model based on clinicopathological data and biomarker variables, calculating the disease-free survival using the value of importance score. From the 48 analyzed biomarkers, only IL-8 emerged as a significant prognostic factor for CRC patients in univariate analysis (HR 4.88; 95% CI 2.00-11.92; p = 0.024) after correcting for multiple comparisons. We also developed a multivariate model based on all 48 biomarkers using a random survival forest analysis. Variable selection based on a minimal depth and the value of importance yielded two tentative candidate CRC prognostic markers: IL-2Ra and IL-8. A multivariate prognostic model using machine-learning technologies improves the prognostic assessment of survival among surgically treated CRC patients.

Risky emotional family environment in childhood and depression-related cytokines in adulthood: The protective role of compassion.

BACKGROUND: Previously, compassion has been found to protect against depressive symptoms, while emotional adversities in childhood are suggested to increase inflammatory responses. The current study investigated (a) whether emotional family environment in childhood predicts levels of such cytokines in adulthood that are previously found to be elevated in depression (interleukin [IL]-2, IL-6, IL-1b, monocyte chemoattractant protein-1, interferon-gamma [IFN-γ], and tumor necrosis factor alpha [TNF-α]) and (b) whether these associations are modified by compassion in adulthood. METHODS: The participants (N = 1,198-1,523) came from the prospective population-based Young Finns data. Emotional family environment and parental socioeconomic factors were evaluated in 1980; participants' compassion in 2001; and participants' cytokine levels and adulthood covariates in 2007. RESULTS: Risky emotional family environment in childhood predicted higher levels of IL-2, IL-6, IFN-γ, and TNF-α in adulthood. Additionally, there were significant interaction effects between compassion and emotional risk in childhood, when predicting IL-2, IL-6, and TNF-α. Specifically, individuals who grew up in a risky emotional family environment had on average higher levels of IL-2, IL-6, and TNF-α in adulthood when combined with low compassion. CONCLUSIONS: In individuals coming from risky emotional family environments, high compassion for others may protect against elevated levels of cytokines previously linked with depression.