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1.
Eur Heart J ; 2019 Apr 19.
Artigo em Inglês | MEDLINE | ID: mdl-31004144

RESUMO

AIMS: Heart failure with preserved ejection fraction (HFpEF) and pathological cardiac aging share a complex pathophysiology, including extracellular matrix remodelling (EMR). Protease-activated receptor 2 (PAR2) deficiency is associated with EMR. The roles of PAR1 and PAR2 have not been studied in HFpEF, age-dependent cardiac fibrosis, or diastolic dysfunction (DD). METHODS AND RESULTS: Evaluation of endomyocardial biopsies from patients with HFpEF (n = 14) revealed that a reduced cardiac PAR2 expression was associated with aggravated DD and increased myocardial fibrosis (r = -0.7336, P = 0.0028). In line, 1-year-old PAR2-knockout (PAR2ko) mice suffered from DD with preserved systolic function, associated with an increased age-dependent α-smooth muscle actin expression, collagen deposition (1.7-fold increase, P = 0.0003), lysyl oxidase activity, collagen cross-linking (2.2-fold increase, P = 0.0008), endothelial activation, and inflammation. In the absence of PAR2, the receptor-regulating protein caveolin-1 was down-regulated, contributing to an augmented profibrotic PAR1 and transforming growth factor beta (TGF-ß)-dependent signalling. This enhanced TGF-ß/PAR1 signalling caused N-proteinase (ADAMTS3) and C-proteinase (BMP1)-related increased collagen I production from cardiac fibroblasts (CFs). PAR2 overexpression in PAR2ko CFs reversed these effects. The treatment with the PAR1 antagonist, vorapaxar, reduced cardiac fibrosis by 44% (P = 0.03) and reduced inflammation in a metabolic disease model (apolipoprotein E-ko mice). Patients with HFpEF with upstream PAR inhibition via FXa inhibitors (n = 40) also exhibited reduced circulating markers of fibrosis and DD compared with patients treated with vitamin K antagonists (n = 20). CONCLUSIONS: Protease-activated receptor 2 is an important regulator of profibrotic PAR1 and TGF-ß signalling in the heart. Modulation of the FXa/FIIa-PAR1/PAR2/TGF-ß-axis might be a promising therapeutic approach to reduce HFpEF.

2.
Cardiovasc Res ; 2018 Aug 08.
Artigo em Inglês | MEDLINE | ID: mdl-30101304

RESUMO

Background: The immune system is considered a key driver of atherosclerosis, and beyond proteins and microRNAs (miRs), long noncoding RNAs (lncRNAs) are implicated in immune control. We previously described that lncRNA MALAT1 is involved in cardiac innate immunity in a myocarditis model. Here, we investigated the impact of MALAT1 deficiency upon atherosclerosis development. Methods and Results: Heterozygous MALAT1-deficient ApoE-/- mice displayed massive immune system dysregulation and atherosclerosis within two months even when kept on normal diet. Aortic plaque area (p < 0.05) and aortic root plaque size (p < 0.001) were increased in MALAT1-deficient vs. MALAT1-wildtype ApoE-/- mice. Serum levels of interferon-γ (IFN-γ), tumor necrosis factor (TNF), and interleukin 6 (IL6) were elevated (p < 0.001) in MALAT1-deficient animals. MALAT1-deficient bone marrow derived macrophages showed enhanced expression of TNF (p = 0.001) and inducible NO synthase (NOS2) (p = 0.002), suppressed MMP9 (p < 0.001), and impaired phagocytic activity (p < 0.001) upon LPS stimulation. RNA-sequencing revealed grossly altered transcriptomes of MALAT1-deficient splenocytes already at baseline, with massive induction of IFN-γ, TNF, NOS2, and granzyme B; CC and CXC chemokines and CCR8; and innate immunity genes IFIT1/3, IFITM1/3, ISG15. Multiple miRs were up to 45-fold upregulated. Further, selective ablation of the cytosolic part of the MALAT1 system only, the enzymatically MALAT1-derived mascRNA, resulted in massive induction of TNF (p = 0.004) and IL6 (p = 0.028) in macrophages. Northern analysis of post-MI patient vs. control PBMCs showed reduced (p = 0.005) mascRNA in the patients. CHART-enriched RNA-sequencing reads at the genomic loci of MALAT1 and neighbouring NEAT1 documented direct interaction between these lncRNA transcripts. Conclusion: The data suggest a molecular circuit involving the MALAT1-mascRNA system, interactions between MALAT1 and NEAT1, and key immune effector molecules, cumulatively impacting upon the development of atherosclerosis. It appears reasonable to look for therapeutic targets in this circuit and to screen for anomalies in the NEAT1-MALAT1 region in humans, too, as possible novel disease risk factors.

3.
Cardiovasc Diabetol ; 17(1): 34, 2018 02 24.
Artigo em Inglês | MEDLINE | ID: mdl-29477147

RESUMO

BACKGROUND: Diabetes mellitus is characterized by chronic vascular disorder and presents a main risk factor for cardiovascular mortality. In particular, hyperglycaemia and inflammatory cytokines induce vascular circulating tissue factor (TF) that promotes pro-thrombotic conditions in diabetes. It has recently become evident that alterations of the post-transcriptional regulation of TF via specific microRNA(miR)s, such as miR-126, contribute to the pathogenesis of diabetes and its complications. The endothelial miR-19a is involved in vascular homeostasis and atheroprotection. However, its role in diabetes-related thrombogenicity is unknown. Understanding miR-networks regulating procoagulability in diabetes may help to develop new treatment options preventing vascular complications. METHODS AND RESULTS: Plasma of 44 patients with known diabetes was assessed for the expression of miR-19a, TF protein, TF activity, and markers for vascular inflammation. High miR-19a expression was associated with reduced TF protein, TF-mediated procoagulability, and vascular inflammation based on expression of vascular adhesion molecule-1 and leukocyte count. We found plasma expression of miR-19a to strongly correlate with miR-126. miR-19a reduced the TF expression on mRNA and protein level in human microvascular endothelial cells (HMEC) as well as TF activity in human monocytes (THP-1), while anti-miR-19a increased the TF expression. Interestingly, miR-19a induced VCAM expression in HMEC. However, miR-19a and miR-126 co-transfection reduced total endothelial VCAM expression and exhibited additive inhibition of a luciferase reporter construct containing the F3 3'UTR. CONCLUSIONS: While both miRs have differential functions on endothelial VCAM expression, miR-19a and miR-126 cooperate to exhibit anti-thrombotic properties via regulating vascular TF expression. Modulating the post-transcriptional control of TF in diabetes may provide a future anti-thrombotic and anti-inflammatory therapy.

6.
Infect Immun ; 83(2): 482-91, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25385798

RESUMO

Accumulation of Tropheryma whipplei-stuffed macrophages in the duodenum, impaired T. whipplei-specific Th1 responses, and weak secretion of interleukin-12 (IL-12) are hallmarks of classical Whipple's disease (CWD). This study addresses dendritic cell (DC) functionality during CWD. We documented composition, distribution, and functionality of DC ex vivo or after in vitro maturation by fluorescence-activated cell sorting (FACS) and by immunohistochemistry in situ. A decrease in peripheral DC of untreated CWD patients compared to healthy donors was due to reduced CD11c(high) myeloid DC (M-DC). Decreased maturation markers CD83, CD86, and CCR7, as well as low IL-12 production in response to stimulation, disclosed an immature M-DC phenotype. In vitro-generated monocyte-derived DC from CWD patients showed normal maturation and T cell-stimulatory capacity under proinflammatory conditions but produced less IL-12 and failed to activate T. whipplei-specific Th1 cells. In duodenal and lymphoid tissues, T. whipplei was found within immature DC-SIGN(+) DC. DC and proliferating lymphocytes were reduced in lymph nodes of CWD patients compared to levels in controls. Our results indicate that dysfunctional IL-12 production by DC provides suboptimal conditions for priming of T. whipplei-specific T cells during CWD and that immature DC carrying T. whipplei contribute to the dissemination of the bacterium.


Assuntos
Células Dendríticas/imunologia , Subunidade p35 da Interleucina-12/biossíntese , Células Th1/imunologia , Doença de Whipple/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos CD/biossíntese , Antígeno B7-2/biossíntese , Antígeno CD11c/biossíntese , Moléculas de Adesão Celular/biossíntese , Proliferação de Células , Duodeno/imunologia , Duodeno/microbiologia , Feminino , Citometria de Fluxo , Humanos , Imunoglobulinas/biossíntese , Subunidade p35 da Interleucina-12/imunologia , Subunidade p35 da Interleucina-12/metabolismo , Lectinas Tipo C/biossíntese , Linfonodos/imunologia , Ativação Linfocitária/imunologia , Macrófagos/imunologia , Macrófagos/microbiologia , Masculino , Glicoproteínas de Membrana/biossíntese , Pessoa de Meia-Idade , Receptores CCR7/biossíntese , Receptores de Superfície Celular/biossíntese , Tropheryma/imunologia , Tropheryma/patogenicidade , Doença de Whipple/microbiologia , Doença de Whipple/mortalidade
7.
Inflamm Res ; 62(9): 865-9, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23775039

RESUMO

BACKGROUND: Macrophage heterogeneity reflects their plasticity in response to environmental stimuli. Usually human macrophages are characterized by analysis of surface molecules or cytokine expression while functional assays are established in the mouse system but lacking for various human specimens. METHODS: To evaluate the value of analysis of arginine metabolism for characterization of human macrophage differentiation, we analyzed nitrite production and arginase activity in plasma, duodenal biopsies, and in vitro differentiated macrophages of patients with classical Whipple's disease. RESULTS: We demonstrate that it is feasible to determine the content of urea in supernatants of stimulated duodenal biopsies, arginase activity in fresh duodenal biopsies and plasma samples, and arginase activity and nitrite production in lysates and supernatants of in vitro differentiated macrophages. However, only selected tests are appropriate to define macrophage polarization in human specimens. CONCLUSION: Analysis of arginine metabolism is not suitable for the characterization of in vitro differentiated human macrophages. Besides the measurement of nitrite in duodenal biopsy supernatants, the determination of arginase activity in human plasma seems to be a reasonable functional test to detect enhanced M2 macrophage activation and, thus, is of great value for the analysis of macrophage activity with a minimum of material and costs.


Assuntos
Arginina/metabolismo , Testes Diagnósticos de Rotina/métodos , Duodeno/metabolismo , Ativação de Macrófagos/fisiologia , Macrófagos/metabolismo , Doença de Whipple/metabolismo , Adulto , Idoso , Biomarcadores/metabolismo , Biópsia , Estudos de Casos e Controles , Diferenciação Celular , Duodeno/patologia , Duodeno/fisiopatologia , Feminino , Humanos , Técnicas In Vitro , Masculino , Pessoa de Meia-Idade , Nitritos/metabolismo , Valor Preditivo dos Testes , Doença de Whipple/patologia , Doença de Whipple/fisiopatologia
8.
J Immunol ; 190(5): 2354-61, 2013 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-23365082

RESUMO

During antimicrobial treatment of classic Whipple's disease (CWD), the chronic systemic infection with Tropheryma whipplei, immune reconstitution inflammatory syndrome (IRIS), is a serious complication. The aim of our study was to characterize the immunological processes underlying IRIS in CWD. Following the definition of IRIS, we describe histological features of IRIS and immunological parameters of 24 CWD IRIS patients, 189 CWD patients without IRIS, and 89 healthy individuals. T cell reconstitution, Th1 reactivity, and the phenotype of T cells were described in the peripheral blood, and infiltration of CD4(+) T cells and regulatory T cells in the duodenal mucosa was determined. During IRIS, tissues were heavily infiltrated by CD3(+), predominantly CD45RO(+)CD4(+) T cells. In the periphery, initial reduction of CD4(+) cell counts and their reconstitution on treatment was more pronounced in CWD patients with IRIS than in those without IRIS. The ratio of activated and regulatory CD4(+) T cells, nonspecific Th1 reactivity, and the proportion of naive among CD4(+) T cells was high, whereas serum IL-10 was low during IRIS. T. whipplei-specific Th1 reactivity remained suppressed before and after emergence of IRIS. The findings that IRIS in CWD mainly are mediated by nonspecific activation of CD4(+) T cells and that it is not sufficiently counterbalanced by regulatory T cells indicate that flare-up of pathogen-specific immunoreactivity is not instrumental in the pathogenesis of IRIS in CWD.


Assuntos
Síndrome Inflamatória da Reconstituição Imune/patologia , Mucosa Intestinal/patologia , Linfócitos T Reguladores/patologia , Células Th1/patologia , Tropheryma/imunologia , Doença de Whipple/patologia , Adulto , Idoso , Antibacterianos/uso terapêutico , Biópsia , Contagem de Linfócito CD4 , Estudos de Casos e Controles , Feminino , Humanos , Síndrome Inflamatória da Reconstituição Imune/complicações , Síndrome Inflamatória da Reconstituição Imune/tratamento farmacológico , Síndrome Inflamatória da Reconstituição Imune/imunologia , Interleucina-10/sangue , Interleucina-10/imunologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/imunologia , Ativação Linfocitária/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Células Th1/efeitos dos fármacos , Células Th1/imunologia , Tropheryma/efeitos dos fármacos , Doença de Whipple/complicações , Doença de Whipple/tratamento farmacológico , Doença de Whipple/imunologia
9.
J Immunol ; 187(8): 4061-7, 2011 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-21918190

RESUMO

Classical Whipple's disease (CWD) is caused by chronic infection with Tropheryma whipplei that seems to be associated with an underlying immune defect. The pathognomonic hallmark of CWD is a massive infiltration of the duodenal mucosa with T. whipplei-infected macrophages that disperse systemically to many other organ systems. An alleviated inflammatory reaction and the absence of T. whipplei-specific Th1 reactivity support persistence and systemic spread of the pathogen. In this article, we hypothesized that regulatory T cells (T(reg)) are involved in immunomodulation in CWD, and we asked for the distribution, activation, and regulatory capacity of T(reg) in CWD patients. Whereas in the lamina propria of CWD patients before treatment numbers of T(reg) were increased, percentages in the peripheral blood were similar in CWD patients and healthy controls. However, peripheral T(reg) of CWD patients were more activated than those of controls. Elevated secretion of IL-10 and TGF-ß in the duodenal mucosa of CWD patients indicated locally enhanced T(reg) activity. Enhanced CD95 expression on peripheral memory CD4(+) T cells combined with reduced expression of IFN-γ and IL-17A upon polyclonal stimulation by CD4(+) cells from untreated CWD patients further hinted to T(reg) activity-related exhaustion of effector CD4(+) T cells. In conclusion, increased numbers of T(reg) can be detected within the duodenal mucosa in untreated CWD, where huge numbers of T. whipplei-infected macrophages are present. Thus, T(reg) might contribute to the chronic infection and systemic spread of T. whipplei in CWD but in contrast prevent mucosal barrier defect by reducing local inflammation.


Assuntos
Mucosa Intestinal/imunologia , Linfócitos T Reguladores/imunologia , Doença de Whipple/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Separação Celular , Citocinas/imunologia , Feminino , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Imunofenotipagem , Mucosa Intestinal/citologia , Mucosa Intestinal/microbiologia , Masculino , Pessoa de Meia-Idade , Tropheryma/imunologia , Doença de Whipple/microbiologia
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