Cardiac macrophage subsets differentially regulate lymphatic network remodeling during pressure overload.

The lymphatic network of mammalian heart is an important regulator of interstitial fluid compartment and immune cell trafficking. We observed a remodeling of the cardiac lymphatic vessels and a reduced lymphatic efficiency during heart hypertrophy and failure induced by transverse aortic constriction. The lymphatic endothelial cell number of the failing hearts was positively correlated with cardiac function and with a subset of cardiac macrophages. This macrophage population distinguished by LYVE-1 (Lymphatic vessel endothelial hyaluronic acid receptor-1) and by resident macrophage gene expression signature, appeared not replenished by CCR2 mediated monocyte infiltration during pressure overload. Isolation of macrophage subpopulations showed that the LYVE-1 positive subset sustained in vitro and in vivo lymphangiogenesis through the expression of pro-lymphangiogenic factors. In contrast, the LYVE-1 negative macrophage subset strongly expressed MMP12 and decreased the endothelial LYVE-1 receptors in lymphatic endothelial cells, a feature of cardiac lymphatic remodeling in failing hearts. The treatment of mice with a CCR2 antagonist during pressure overload modified the proportion of macrophage subsets within the pathological heart and preserved lymphatic network from remodeling. This study reports unknown and differential functions of macrophage subpopulations in the regulation of cardiac lymphatic during pathological hypertrophy and may constitute a key mechanism underlying the progression of heart failure.

Selective Cardiomyocyte Oxidative Stress Leads to Bystander Senescence of Cardiac Stromal Cells.

Accumulation of senescent cells in tissues during normal or accelerated aging has been shown to be detrimental and to favor the outcomes of age-related diseases such as heart failure (HF). We have previously shown that oxidative stress dependent on monoamine oxidase A (MAOA) activity in cardiomyocytes promotes mitochondrial damage, the formation of telomere-associated foci, senescence markers, and triggers systolic cardiac dysfunction in a model of transgenic mice overexpressing MAOA in cardiomyocytes (Tg MAOA). However, the impact of cardiomyocyte oxidative stress on the cardiac microenvironment in vivo is still unclear. Our results showed that systolic cardiac dysfunction in Tg MAOA mice was strongly correlated with oxidative stress induced premature senescence of cardiac stromal cells favoring the recruitment of CCR2+ monocytes and the installation of cardiac inflammation. Understanding the interplay between oxidative stress induced premature senescence and accelerated cardiac dysfunction will help to define new molecular pathways at the crossroad between cardiac dysfunction and accelerated aging, which could contribute to the increased susceptibility of the elderly to HF.

Kidney inflammaging is promoted by CCR2+ macrophages and tissue-derived micro-environmental factors.

The incidence of disorders associated with low inflammatory state, such as chronic kidney disease, increases in the elderly. The accumulation of senescent cells during aging and the senescence-associated secretory phenotype, which leads to inflammaging, is known to be deleterious and account for progressive organ dysfunction. To date, the cellular actors implicated in chronic inflammation in the kidney during aging are still not well characterized. Using the DECyt method, based on hierarchical clustering of flow cytometry data, we showed that aging was associated with significant changes in stromal cell diversity in the kidney. In particular, we identified two cell populations up-regulated with aging, the mesenchymal stromal cell subset (kMSC) expressing CD73 and the monocyte-derived Ly6C+ CCR2+ macrophage subset expressing pro-inflammatory cytokines. Aged CD73+ kMSCs depicted senescence associated features with low proliferation rate, increased DNA damage foci and Ccl2 expression. Using co-cultures experiments, we showed that aged CD73+ kMSC promoted monocyte activation and secretion of inflammatory cytokines albeit less efficiently than young CD73+ kMSCs. In the context of ageing, increased frequency of CD73+ kMSC subpopulations could provide additional niche factors to newly recruited monocytes favoring a positive regulatory loop in response to local inflammation. Interfering with such partnership during aging could be a valuable approach to regulate kidney inflammaging and to limit the risk of developing chronic kidney disease in the elderly.

Aging induces cardiac mesenchymal stromal cell senescence and promotes endothelial cell fate of the CD90 + subset.

Aging is a major risk factor in the development of chronic diseases, especially cardiovascular diseases. Age-related organ dysfunction is strongly associated with the accumulation of senescent cells. Cardiac mesenchymal stromal cells (cMSCs), deemed part of the microenvironment, modulate cardiac homeostasis through their vascular differentiation potential and paracrine activity. Transcriptomic analysis of cMSCs identified age-dependent biological pathways regulating immune responses and angiogenesis. Aged cMSCs displayed a senescence program characterized by Cdkn2a expression, decreased proliferation and clonogenicity, and acquisition of a senescence-associated secretory phenotype (SASP). Increased CCR2-dependent monocyte recruitment by aged cMSCs was associated with increased IL-1ß production by inflammatory macrophages in the aging heart. In turn, IL-1ß induced senescence in cMSCs and mimicked age-related phenotypic changes such as decreased CD90 expression. The CD90+ and CD90- cMSC subsets had biased vascular differentiation potentials, and CD90+ cMSCs were more prone to acquire markers of the endothelial lineage with aging. These features were related to the emergence of a new cMSC subset in the aging heart, expressing CD31 and endothelial genes. These results demonstrate that cMSC senescence and SASP production are supported by the installation of an inflammatory amplification loop, which could sustain cMSC senescence and interfere with their vascular differentiation potentials.

Local production of tenascin-C acts as a trigger for monocyte/macrophage recruitment that provokes cardiac dysfunction.

Aims: Tenascin-C (TNC) is an endogenous danger signal molecule strongly associated with inflammatory diseases and with poor outcome in patients with cardiomyopathies. Its function within pathological cardiac tissue during pressure overload remains poorly understood. Methods and results: We showed that TNC accumulates after 1 week of transverse aortic constriction (TAC) in the heart of 12-week-old male mice. By cross bone marrow transplantation experiments, we determined that TNC deposition relied on cardiac cells and not on haematopoietic cells. The expression of TNC induced by TAC, or by administration of a recombinant lentivector coding for TNC, triggered a pro-inflammatory cardiac microenvironment, monocyte/macrophage (MO/MΦ) accumulation, and systolic dysfunction. TNC modified macrophage polarization towards the pro-inflammatory phenotype and stimulated RhoA/Rho-associated protein kinase (ROCK) pathways to promote mesenchymal to amoeboid transition that enhanced macrophage migration into fibrillar collagen matrices. The amplification of inflammation and MO/MΦ recruitment by TNC was abrogated by genetic invalidation of TNC in knockout mice. These mice showed less ventricular remodelling and an improved cardiac function after TAC as compared with wild-type mice. Conclusions: By promoting a pro-inflammatory microenvironment and macrophage migration, TNC appears to be a key factor to enable the MO/MΦ accumulation within fibrotic hearts leading to cardiac dysfunction. As TNC is highly expressed during inflammation and sparsely during the steady state, its inhibition could be a promising therapeutic strategy to control inflammation and immune cell infiltration in heart disease.

Oxidative Stress by Monoamine Oxidase-A Impairs Transcription Factor EB Activation and Autophagosome Clearance, Leading to Cardiomyocyte Necrosis and Heart Failure.

AIMS: In heart failure (HF), mitochondrial quality control and autophagy are progressively impaired, but the role of oxidative stress in this process and its underlying mechanism remain to be defined. By degrading norepinephrine and serotonin, the mitochondrial enzyme, monoamine oxidase-A (MAO-A), is a potent source of reactive oxygen species (ROS) in the heart and its activation leads to the persistence of mitochondrial damage. In this study, we analyzed the consequences of ROS generation by MAO-A on the autophagy-lysosome pathway in the heart. RESULTS: Cardiomyocyte-driven expression of MAO-A in mice led to mitochondrial fission and translocation of Drp1 and Parkin in the mitochondrial compartment. Ventricles from MAO-A transgenic mice displayed accumulation of LC3-positive autophagosomes, together with p62 and ubiquitylated proteins, indicating impairment of autophagy. In vitro adenoviral delivery of MAO-A in cardiomyocytes and the consequent generation of ROS blocked autophagic flux with accumulation of LC3II, p62, and ubiquitylated proteins, leading to mitochondrial fission and cell necrosis. In addition, MAO-A activation induced accumulation of lysosomal proteins, cathepsin D and Lamp1, reduced lysosomal acidification, and blocked the nuclear translocation of transcription factor-EB (TFEB), a master regulator of autophagy and lysosome biogenesis. Most interestingly, overexpression of TFEB attenuated autophagosome buildup, mitochondrial fission, cardiomyocyte death, and HF associated with MAO-A activation. INNOVATION AND CONCLUSION: This study unravels a new link between MAO-dependent H2O2 production and lysosomal dysfunction. Altogether, our findings demonstrate that the MAO-A/H2O2 axis has a negative impact on the elimination and recycling of mitochondria through the autophagy-lysosome pathway, which participates in cardiomyocyte death and HF. Antioxid. Redox Signal. 25, 10-27.

CD4+ T cells promote the transition from hypertrophy to heart failure during chronic pressure overload.

BACKGROUND: The mechanisms by which the heart adapts to chronic pressure overload, producing compensated hypertrophy and eventually heart failure (HF), are still not well defined. We aimed to investigate the involvement of T cells in the progression to HF using a transverse aortic constriction (TAC) model. METHODS AND RESULTS: Chronic HF was associated with accumulation of T lymphocytes and activated/effector CD4(+) T cells within cardiac tissue. After TAC, enlarged heart mediastinal draining lymph nodes showed a high density of both CD4(+) and CD8(+) T-cell subsets. To investigate the role of T cells in HF, TAC was performed on mice deficient for recombination activating gene 2 expression (RAG2KO) lacking B and T lymphocytes. Compared with wild-type TAC mice, RAG2KO mice did not develop cardiac dilation and showed improved contractile function and blunted adverse remodeling. Reconstitution of the T-cell compartment into RAG2KO mice before TAC enhanced contractile dysfunction, fibrosis, collagen accumulation, and cross-linking. To determine the involvement of a specific T-cell subset, we performed TAC on mice lacking CD4(+) (MHCIIKO) and CD8(+) T-cell subsets (CD8KO). In contrast to CD8KO mice, MHCIIKO mice did not develop ventricular dilation and dysfunction. MHCIIKO mice also displayed very low fibrosis, collagen accumulation, and cross-linking within cardiac tissue. Interestingly, mice with transgenic CD4(+) T-cell receptor specific for ovalbumin failed to develop HF and adverse remodeling. CONCLUSIONS: These results demonstrate for the first time a crucial role of CD4(+) T cells and specific antigen recognition in the progression from compensated cardiac hypertrophy to HF.

Study of Cynomolgus monkey (Macaca fascicularis) MhcDRB (Mafa-DRB) polymorphism in two populations.

Cynomolgus monkey is one of the macaque species currently used as an animal model for experimental surgery and medicine, in particular, to experiment new drugs or therapy protocols designed for the prevention of allograft rejection. In this field, it is of utmost importance to select histoincompatible recipient-donor pairs. One way to ensure incompatibility between donor and recipient is to check their major histocompatibility complex (MHC) genotypes at the loci playing a determinant role in histocompatibility. We report in this paper on the cynomolgus monkey DRB polymorphism evidenced by sequencing of amplified exon 2 separated either by denaturing gradient gel electrophoresis (DGGE), or by cloning. By the study of 253 unrelated animals from two populations (Mauritius and The Philippines), we characterized 50 exon 2 sequences among which 28 were identical to sequences already reported in Macaca fascicularis or other macaque species (Macaca mulatta, Macaca nemestrina). By cloning and sequencing DRB cDNA, we revealed two additional DRB alleles. Out of the 20 haplotypes that we defined here, only two were found in both populations. The functional impact of DR incompatibility was studied in vitro by mixed lymphocyte culture.