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BACKGROUND The combination of serum $\beta$2-microglubulin and albumin levels is highly prognostic in multiple myeloma (MM),defined as the International Staging System (ISS). Recurrent genomic abnormalities present in myeloma cells also have a strong prognostic power. This study aimed to assess,in a routine diagnostic setting,whether genomic aberrations can be used to identify sub-groups in ISS staging,as this system does not incorporate intrinsic myeloma cell variability at the molecular level. MATERIALS AND METHODS A prospective population-based study of 123 patients newly diagnosed with MM with ISS staging were included for karyotyping,interphase nuclei fluorescence in situ hybridization (iFISH) and oligo-based array comparative genomic hybridization (oaCGH) analyses. RESULTS Clonal abnormalities were identified in 27{\%} of analyses by karyotyping,in 83{\%} by iFISH,and in 99{\%} by oaCGH analysis. ISS staging combined with oaCGH aberrations identified ISS sub-groups. CONCLUSION oaCGH analysis is a valuable asset in detecting prognostically relevant genomic abnormalities. The combination of oaCGH data with ISS staging might help define new sub-groups in MM. View Publication

In addition to antigen-driven signals,T cells need co-stimulatory signals for robust activation. Several receptors,including members of the tumor necrosis factor receptor superfamily (TNFRSF),can deliver co-stimulatory signals to T cells. Thioredoxin interacting protein (TXNIP) is an important inhibitor of glucose uptake and cell proliferation,but it is unknown how TXNIP is regulated in T cells. The aim of this study was to determine expression levels and regulation of TXNIP in human T cells. We found that na{\{i}}ve T cells express high levels of TXNIP and that treatment of blood samples with TNF results in rapid down-regulation of TXNIP in the T cells. TNF-induced TXNIP down-regulation correlated with increased glucose uptake. Furthermore we found that density gradient centrifugation (DGC) induced down-regulation of TXNIP. We demonstrate that DGC induced TNF production that paralleled the TXNIP down-regulation. Treatment of blood with toll-like receptor (TLR) ligands induced TNF production and TXNIP down-regulation suggesting that damage-associated molecular patterns (DAMPs) such as endogenous TLR ligands released during DGC play a role in DGC-induced TXNIP down-regulation. Finally we demonstrate that TNF-induced TXNIP down-regulation is dependent on caspase activity and is caused by caspase-mediated cleavage of TXNIP." View Publication

MicroRNAs (miRNA) are small noncoding RNAs that regulate gene expression by targeting mRNAs in a sequence specific manner,thereby determining their degradation or inhibiting translation. They are involved in processes such as proliferation,differentiation and apoptosis by fine-tuning the expression of genes underlying such events. The expression of specific miRNAs is involved in hematopoietic differentiation and their deregulation contributes to the development of hematopoietic malignancies such as acute myeloid leukemia (AML). miR-130a is over-expressed in AML. Here we show that miR-130a is physiologically expressed in myeloblasts and down-regulated during monocyte differentiation. Gain- and loss-of-function experiments performed on CD34+ human hematopoietic stem cells confirmed that expression of miR-130a inhibits monocyte differentiation by interfering with the expression of key transcription factors HOXA10,IRF8,KLF4,MAFB and PU-1. The data obtained in this study highlight that the correct modulation of miR-130a is necessary for normal differentiation to occur and confirming that deregulation of this miRNA might underlie the differentiation block occurring in AML. View Publication

Environmental factors contribute to Type 1 diabetes (T1D) susceptibility. The gut microbiome,which includes bacteria,viruses,and fungi,contributes to this environmental influence,and can induce immunological changes. The gut viral component of the microbiome,related to T1D has mostly focused on coxsackieviruses and rotavirus. The role of norovirus,another common enteric virus,in susceptibility to T1D was hitherto unknown. Norovirus is highly infectious and encountered by many children. We studied the mouse norovirus 4 (MNV4),related to human noroviruses,in the Non-obese diabetic (NOD) mouse model,to determine its role in influencing susceptibility to T1D. We infected MNV-free NOD mice with MNV4 by exposing the mice to MNV4-positive bedding from an endemically-infected mouse colony to mimic a natural infection. Control MNV-free NOD mice were exposed to MNV-free bedding from the same colony. Interestingly,MNV4 infection protected NOD mice from the development of T1D and was associated with an expansion of Tregs and reduced proinflammatory T cells. We also found MNV4 significantly modified the gut commensal bacteria composition,promoting increased $\alpha$-diversity and Firmicutes/Bacteroidetes ratio. To elucidate whether T1D protection was directly related to MNV4,or indirectly through modulating gut microbiota,we colonized germ-free (GF) NOD mice with the MNV4-containing or non-MNV4-containing viral filtrate,isolated from filtered fecal material. We found that MNV4 induced significant changes in mucosal immunity,including altered Tuft cell markers,cytokine secretion,antiviral immune signaling markers,and the concentration of mucosal antibodies. Systemically,MNV4-infection altered the immune cells including B cell subsets,macrophages and T cells,and especially induced an increase in Treg number and function. Furthermore,in vitro primary exposure of the norovirus filtrate to na{\{i}}ve splenocytes identified significant increases in the proportion of activated and CTLA4-expressing Tregs. Our data provide novel knowledge that norovirus can protect NOD mice from T1D development by inducing the expansion of Tregs and reducing inflammatory T cells. Our study also highlights the importance of distinguishing the mucosal immunity mediated by bacteria from that by enteric viruses." View Publication

As the prognosis of invasive aspergillosis remains unacceptably poor in patients undergoing hematopoietic stem cell transplantation (HSCT),there is a growing interest in the adoptive transfer of antifungal effector cells,such as Natural Killer (NK) cells. Because immunosuppressive agents are required in most HSCT recipients,knowledge of the impact of these compounds on the antifungal activity of NK cells is a prerequisite for clinical trials. We,therefore,assessed the effect of methylprednisolone (mPRED),cyclosporin A (CsA) and mycophenolic acid (MPA) at different concentrations on proliferation,apoptosis/necrosis,and the direct and indirect anti-Aspergillus activity of human NK cells. Methylprednisolone decreased proliferation and increased apoptosis of NK cells in a significant manner. After seven days,a reduction of viable NK cells was seen for all three immunosuppressants,which was significant for MPA only. Cyclosporin A significantly inhibited the direct hyphal damage by NK cells in a dose-dependent manner. None of the immunosuppressive compounds had a major impact on the measured levels of interferon-$\gamma$,granulocyte-macrophage colony-stimulating factor and RANTES (regulated on activation,normal T cell expressed and secreted; CCL5). Our data demonstrate that commonly used immunosuppressive compounds have distinct effects on proliferation,viability and antifungal activity of human NK cells,which should be considered in designing studies on the use of NK cells for adoptive antifungal immunotherapy. View Publication

Immunosuppression increases the risk of cancers that are associated with viral infection1. In particular,the risk of squamous cell carcinoma of the skin-which has been associated with beta human papillomavirus ($\beta$-HPV) infection-is increased by more than 100-fold in immunosuppressed patients2-4. Previous studies have not established a causative role for HPVs in driving the development of skin cancer. Here we show that T cell immunity against commensal papillomaviruses suppresses skin cancer in immunocompetent hosts,and the loss of this immunity-rather than the oncogenic effect of HPVs-causes the markedly increased risk of skin cancer in immunosuppressed patients. To investigate the effects of papillomavirus on carcinogen-driven skin cancer,we colonized several strains of immunocompetent mice with mouse papillomavirus type 1 (MmuPV1)5. Mice with natural immunity against MmuPV1 after colonization and acquired immunity through the transfer of T cells from immune mice or by MmuPV1 vaccination were protected against skin carcinogenesis induced by chemicals or by ultraviolet radiation in a manner dependent on CD8+ T cells. RNA and DNA in situ hybridization probes for 25 commensal $\beta$-HPVs revealed a significant reduction in viral activity and load in human skin cancer compared with the adjacent healthy skin,suggesting a strong immune selection against virus-positive malignant cells. Consistently,E7 peptides from $\beta$-HPVs activated CD8+ T cells from unaffected human skin. Our findings reveal a beneficial role for commensal viruses and establish a foundation for immune-based approaches that could block the development of skin cancer by boosting immunity against the commensal HPVs present in all of our skin. View Publication

Diffuse large B cell lymphoma (DLBCL) is the commonest disorder derived from the B-lymphocytes. Inhibiting the immune checkpoint through naturalizing programmed death-1 (PD-1) and programmed death ligand 1 (PD-L1) is proved to be a successful therapeutic regime for lymphoma. Long non-coding RNAs (lncRNAs) are unceasingly reported to be promising biological targets for the cancer therapies. This study planned to explore the regulation of small nucleolar RNA host gene 14 (SNHG14) on DLBCL. SNHG14 level in DLBCL samples and cell lines was analyzed by GEPIA bioinformatics tool and RT-qPCR. Biological functions of SNHG14 in DLBCL were detected by CCK-8,colony formation,and transwell invasion assays. Molecular interaction was determined by RNA immunoprecipitation (RIP) and luciferase reporter assays. MiR-5590-3p-related pathway was identified through KEGG pathway analysis applying DAVID6.8 online bioinformatics tool. Effect of SNHG14 on CD8+ T cells was detected by flow cytometry. Results depicted that SNHG14 was upregulated in DLBCL and its depletion retarded proliferation,migration and epithelial-to-mesenchymal transition (EMT). Mechanistically,SNHG14 sponged miR-5590-3p to upregulate Zinc finger E-box binding homeobox 1 (ZEB1),and ZEB1 transcriptionally activated SNHG14 and PD-L1 to promote the immune evasion of DLBCL cells. In conclusion,we firstly showed that SNHG14/miR-5590-3p/ZEB1 positive feedback loop promoted diffuse large B cell lymphoma progression and immune evasion through regulating PD-1/PD-L1 checkpoint,indicating that targeting SNHG14 was a potential approach to improve the efficacy of immunotherapy in DLBCL. View Publication

The association between type 2 diabetes (T2D) pathogenesis and immune-mediated tissue damage and insulin resistance suggests that T2D patients might benefit from the suppression of pathogenic inflammation. Foxp3+ Treg cells are crucial suppressors of inflammation,but the differentiation of Foxp3+ Treg cells is not static and is subject to conversion into IL-17-producing Th17-like cells upon receiving external signals. In this study,we examined the production of IL-17 by Treg cells. Compared to non-T2D controls,T2D patients presented significantly higher levels of IL-17-expressing cells in both Foxp3- CD4 T cells and Foxp3+ Treg cells. The frequencies of IL-17-nonexpressing Foxp3+ Treg cells,on the other hand,were not changed. Interestingly,IL-17-expressing Foxp3+ Treg cells were mutually exclusive from IL-10-expressing and TGF-$\beta$-expressing Foxp3+ Treg cells,suggesting that multiple subpopulations exist within the Foxp3+ Treg cells from T2D patients. In T2D patients,the frequencies of IL-17-expressing Foxp3+ Treg cells were positively correlated with the body mass index (BMI) and the HbA1c levels of T2D patients. The frequencies of IL-10-expressing Treg cells,on the other hand,were inversely associated with the BMI of both non-T2D controls and T2D patients. In addition,the suppressive activity of Treg cells was significantly lower in T2D patients than in non-T2D controls. Together,our study uncovered a dysregulation in Foxp3+ Treg cells from T2D patients,characterized by high IL-17 expression and low suppression activity. View Publication

To date,the effects of endurance exercise training on lymphocyte physiology at the kinome level are largely unknown. Therefore,the present study used a highly sensitive peptide-based kinase activity profiling approach to investigate if the basal activity of tyrosine (Tyr) and serine/threonine (Ser/Thr) kinases of human lymphocytes is affected by the aerobic endurance training status. Results revealed that the activity of various tyrosine kinases of the FGFR family and ZAP70 was increased,whereas the activity of multiple Ser/Thr kinases such as IKK$\alpha$,CaMK4,PKA$\alpha$,PKC$\alpha$+$\delta$ (among others) was decreased in lymphocytes of endurance trained athletes (ET). Moreover,functional associations between several differentially regulated kinases in ET-derived lymphocytes were demonstrated by phylogenetic mapping and network analysis. Especially,Ser/Thr kinases of the AGC-kinase (protein kinase A,G,and C) family represent exercise-sensitive key components within the lymphocytes kinase network that may mediate the long-term effects of endurance training. Furthermore,KEGG (Kyoto Encyclopedia of Genes and Genomes) and Reactome pathway analysis indicate that Ras as well as intracellular signaling by second messengers were found to be enriched in the ET individuals. Overall,our data suggest that endurance exercise training improves the adaptive immune competence by modulating the activity of multiple protein kinases in human lymphocytes. View Publication

BACKGROUND Lymphotoxin $\beta$ receptor (LT$\beta$R) plays important roles in the development of the immune system and immune response. At the cellular level,ligand-bound LT$\beta$R activates the pro-inflammatory NF-$\kappa$B pathway but the detailed mechanisms regulating its signaling remain unknown. Understanding them is of high importance since LT$\beta$R and its ligands are promising therapeutic targets. Here,we studied the consequences of perturbed cellular cholesterol content on LT$\beta$R-induced NF-$\kappa$B signaling. METHODS To modulate cholesterol availability and/or level in lung carcinoma A549 and H2228,and endothelial HUVEC cells different treatment regimens with filipin,methyl-$\beta$-cyclodextrin and simvastatin were applied. LT$\beta$R localization was studied by confocal microscopy. The activity of LT$\beta$R-induced NF-$\kappa$B pathway was assessed by measuring the levels of NF-$\kappa$B pathway inhibitor I$\kappa$B$\alpha$ and phosphorylation of RelA transcription factor by Western blotting. The NF-$\kappa$B transcriptional response,production of chemokines and adhesion molecules were examined by qRT-PCR,ELISA,and Western blotting,respectively. Adherence of different types of primary immune cells to epithelial A549 cells and endothelial HUVECs was measured fluorometrically. Interactions of LT$\beta$R with its protein partners were investigated by immunoprecipitation. RESULTS We showed that filipin-mediated sequestration of cholesterol or its depletion from the plasma membrane with methyl-$\beta$-cyclodextrin impaired LT$\beta$R internalization and potentiated LT$\beta$R-dependent activation of the canonical branch of the NF-$\kappa$B pathway. The latter was manifested by enhanced degradation of I$\kappa$B$\alpha$ inhibitor,elevated RelA phosphorylation,substantial increase in the expression of NF-$\kappa$B target genes encoding,among others,cytokines and adhesion molecules known to play important roles in immune response. It was followed by robust secretion of CXCL8 and upregulation of ICAM1,that favored the adhesion of immune cells (NK and T cells,neutrophils) to A549 cells and HUVECs. Mechanistically,we showed that cholesterol depletion stabilized interactions of ligand-stimulated LT$\beta$R with modified forms of TRAF2 and NEMO proteins. CONCLUSIONS Our results showed that the reduction of the plasma membrane content of cholesterol or its sequestration strongly potentiated signaling outcome initiated by LT$\beta$R. Thus,drugs modulating cholesterol levels could potentially improve efficacy of LT$\beta$R-based therapies. Video abstract. View Publication

Scleroderma (SSc) is an autoimmune connective tissue disease characterized by immune dysregulation,vasculopathy,and fibrosis. We have previously demonstrated that low Fli1 expression in SSc fibroblasts and endothelial cells plays an important role in SSc pathogenesis. Cells of myeloid and lymphoid origin also express Fli1 and are dysregulated in patients with SSc,playing key roles in disease pathogenesis. However,the role for immune Fli1 in SSc is not yet clear. Our aim was to elucidate whether Fli1 contributes to the immune dysregulation seen in SSc. Comparison of the expression of Fli1 in monocytes,B- and T-cell fractions of PBMCs isolated from SSc patients and healthy controls (HC),showed an increase in Fli1 levels in monocytes. We used siRNA transfected human myeloid cells and mouse peritoneal macrophages obtained from Fli1 flox/flox LysMCre+/+ mice,and found that markers of alternative macrophage activation were increased with Fli1 deletion. Coculture of Fli1-deficient myeloid cells and primary human or mouse fibroblasts resulted in a potent induction of collagen type I,independent of TGF$\beta$ upregulation. We next analyzed global gene expression profile in response to Fli1 downregulation,to gain further insight into the molecular mechanisms of this process and to identify differentially expressed genes in myeloid cells. Of relevance to SSc,the top most upregulated pathways were hallmark IFN-$\gamma$ and IFN-$\alpha$ response. Additionally,several genes previously linked to SSc pathogenesis and fibrosis in general were also induced,including CCL2,CCL7,MMP12,and CXCL10. ANKRD1,a profibrotic transcription co-regulator was the top upregulated gene in our array. Our results show that Fli1-deficient myeloid cells share key features with cells from SSc patients,with higher expression of profibrotic markers and activation of interferon responsive genes,thus suggesting that dysregulation of Fli1 in myeloid cells may contribute to SSc pathogenesis. View Publication