Pericytes are widely believed to function as mesenchymal stem cells (MSCs), multipotent tissue-resident progenitors with great potential for regenerative medicine. Cultured pericytes isolated from ...distinct tissues can differentiate into multiple cell types in vitro or following transplantation in vivo. However, the cell fate plasticity of endogenous pericytes in vivo remains unclear. Here, we show that the transcription factor Tbx18 selectively marks pericytes and vascular smooth muscle cells in multiple organs of adult mouse. Fluorescence-activated cell sorting (FACS)-purified Tbx18-expressing cells behaved as MSCs in vitro. However, lineage-tracing experiments using an inducible Tbx18-CreERT2 line revealed that pericytes and vascular smooth muscle cells maintained their identity in aging and diverse pathological settings and did not significantly contribute to other cell lineages. These results challenge the current view of endogenous pericytes as multipotent tissue-resident progenitors and suggest that the plasticity observed in vitro or following transplantation in vivo arises from artificial cell manipulations ex vivo.
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•Tbx18 is selectively expressed by pericytes in multiple adult organs•A Tbx18-CreERT2 line allows highly efficient labeling of pericytes in vivo•In vivo, pericytes do not behave as stem cells during aging•In vivo, pericytes do not behave as stem cells in pathological settings
Guimarães-Camboa et al. permanently labeled pericytes and vascular smooth muscle of multiple organs in vivo and followed the fate of these cells in aging and injury models. Their analyses showed that, in vivo, pericytes did not behave as stem cells, challenging the current view of pericytes as tissue-resident multipotent progenitors.
Circulating subclinical lipopolysaccharide (LPS) occurs in health and disease. Ingesting high fatty meals increases LPS that cause metabolic endotoxemia. Subclinical LPS in periodontal disease may ...impair endothelial function. The heart may be targeted as cardiac cells express TLR4, the LPS receptor. It was hypothesized that recurrent exposure to subclinical LPS increases mortality and causes cardiac fibrosis.
C57Bl/6 mice were injected with intraperitoneal saline (control), low dose LPS (0.1 or 1 mg/kg), or moderate dose LPS (10 or 20 mg/kg), once a week for 3 months. Left ventricular (LV) function (echocardiography), hemodynamics (tail cuff pressure) and electrocardiograms (telemetry) were measured. Cardiac fibrosis was assessed by picrosirius red staining and LV expression of fibrosis related genes (QRT-PCR). Adult cardiac fibroblasts were isolated and exposed to LPS.
LPS injections transiently increased heart rate and blood pressure (<6 hours) and mildly decreased LV function with full recovery by 24 hours. Mice tolerated weekly LPS for 2-3 months with no change in activity, appearance, appetite, weight, blood pressure, LV function, oximetry, or blood chemistries. Mortality increased after 60-90 days with moderate, but not low dose LPS. Arrhythmias occurred a few hours before death. LV collagen fraction area increased dose-dependently from 3.0±0.5% (SEM) in the saline control group, to 5.6±0.5% with low dose LPS and 9.7±0.9% with moderate dose LPS (P<0.05 moderate vs low dose LPS, and each LPS dose vs control). LPS increased LV expression of collagen Iα1, collagen IIIα1, MMP2, MMP9, TIMP1, periostin and IL-6 (P<0.05 moderate vs low dose LPS and vs control). LPS increased α-SMA immunostaining of myofibroblasts. LPS dose-dependently increased IL-6 in isolated adult cardiac fibroblasts.
Recurrent exposure to subclinical LPS increases mortality and induces cardiac fibrosis.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Activation and accumulation of cardiac fibroblasts, which result in excessive extracellular matrix deposition and consequent mechanical stiffness, myocyte uncoupling, and ischemia, are key ...contributors to heart failure progression. Recently, endothelial-to-mesenchymal transition (EndoMT) and the recruitment of circulating hematopoietic progenitors to the heart have been reported to generate substantial numbers of cardiac fibroblasts in response to pressure overload-induced injury; therefore, these processes are widely considered to be promising therapeutic targets. Here, using multiple independent murine Cre lines and a collagen1a1-GFP fusion reporter, which specifically labels fibroblasts, we found that following pressure overload, fibroblasts were not derived from hematopoietic cells, EndoMT, or epicardial epithelial-to-mesenchymal transition. Instead, pressure overload promoted comparable proliferation and activation of two resident fibroblast lineages, including a previously described epicardial population and a population of endothelial origin. Together, these data present a paradigm for the origins of cardiac fibroblasts during development and in fibrosis. Furthermore, these data indicate that therapeutic strategies for reducing pathogenic cardiac fibroblasts should shift from targeting presumptive EndoMT or infiltrating hematopoietically derived fibroblasts, toward common pathways upregulated in two endogenous fibroblast populations.
Defective protein quality control (PQC) systems are implicated in multiple diseases. Molecular chaperones and co-chaperones play a central role in functioning PQC. Constant mechanical and metabolic ...stress in cardiomyocytes places great demand on the PQC system. Mutation and downregulation of the co-chaperone protein BCL-2-associated athanogene 3 (BAG3) are associated with cardiac myopathy and heart failure, and a BAG3 E455K mutation leads to dilated cardiomyopathy (DCM). However, the role of BAG3 in the heart and the mechanisms by which the E455K mutation leads to DCM remain obscure. Here, we found that cardiac-specific Bag3-KO and E455K-knockin mice developed DCM. Comparable phenotypes in the 2 mutants demonstrated that the E455K mutation resulted in loss of function. Further experiments revealed that the E455K mutation disrupted the interaction between BAG3 and HSP70. In both mutants, decreased levels of small heat shock proteins (sHSPs) were observed, and a subset of proteins required for cardiomyocyte function was enriched in the insoluble fraction. Together, these observations suggest that interaction between BAG3 and HSP70 is essential for BAG3 to stabilize sHSPs and maintain cardiomyocyte protein homeostasis. Our results provide insight into heart failure caused by defects in BAG3 pathways and suggest that increasing BAG3 protein levels may be of therapeutic benefit in heart failure.
Several injectable materials have been shown to preserve or improve cardiac function as well as prevent or slow left ventricular (LV) remodeling post-myocardial infarction (MI). However, it is ...unclear as to whether it is the structural support or the bioactivity of these polymers that lead to beneficial effects. Herein, we examine how passive structural enhancement of the LV wall by an increase in wall thickness affects cardiac function post-MI using a bio-inert, non-degradable synthetic polymer in an effort to better understand the mechanisms by which injectable materials affect LV remodeling.
Poly(ethylene glycol) (PEG) gels of storage modulus G' = 0.5±0.1 kPa were injected and polymerized in situ one week after total occlusion of the left coronary artery in female Sprague Dawley rats. The animals were imaged using magnetic resonance imaging (MRI) at 7±1 day(s) post-MI as a baseline and again post-injection 49±4 days after MI. Infarct wall thickness was statistically increased in PEG gel injected vs. control animals (p<0.01). However, animals in the polymer and control groups showed decreases in cardiac function in terms of end diastolic volume, end systolic volume and ejection fraction compared to baseline (p<0.01). The cellular response to injection was also similar in both groups.
The results of this study demonstrate that passive structural reinforcement alone was insufficient to prevent post-MI remodeling, suggesting that bioactivity and/or cell infiltration due to degradation of injectable materials are likely playing a key role in the preservation of cardiac function, thus providing a deeper understanding of the influencing properties of biomaterials necessary to prevent post-MI negative remodeling.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Ca2+/calmodulin-dependent kinase II (CaMKII) has been implicated in cardiac hypertrophy and heart failure. We generated mice in which the predominant cardiac isoform, CaMKIIdelta, was genetically ...deleted (KO mice), and found that these mice showed no gross baseline changes in ventricular structure or function. In WT and KO mice, transverse aortic constriction (TAC) induced comparable increases in relative heart weight, cell size, HDAC5 phosphorylation, and hypertrophic gene expression. Strikingly, while KO mice showed preserved hypertrophy after 6-week TAC, CaMKIIdelta deficiency significantly ameliorated phenotypic changes associated with the transition to heart failure, such as chamber dilation, ventricular dysfunction, lung edema, cardiac fibrosis, and apoptosis. The ratio of IP3R2 to ryanodine receptor 2 (RyR2) and the fraction of RyR2 phosphorylated at the CaMKII site increased significantly during development of heart failure in WT mice, but not KO mice, and this was associated with enhanced Ca2+ spark frequency only in WT mice. We suggest that CaMKIIdelta contributes to cardiac decompensation by enhancing RyR2-mediated sarcoplasmic reticulum Ca2+ leak and that attenuating CaMKIIdelta activation can limit the progression to heart failure.
Ankrd1/CARP, Ankrd2/Arpp, and Ankrd23/DARP belong to a family of stress inducible ankyrin repeat proteins expressed in striated muscle (MARPs). The MARPs are homologous in structure and localized in ...the nucleus where they negatively regulate gene expression as well as in the sarcomeric I-band, where they are thought to be involved in mechanosensing. Together with their strong induction during cardiac disease and the identification of causative Ankrd1 gene mutations in cardiomyopathy patients, this suggests their important roles in cardiac development, function, and disease. To determine the functional role of MARPs in vivo, we studied knockout (KO) mice of each of the three family members. Single KO mice were viable and had no apparent cardiac phenotype. We therefore hypothesized that the three highly homologous MARP proteins may have redundant functions in the heart and studied double and triple MARP KO mice. Unexpectedly, MARP triple KO mice were viable and had normal cardiac function both at basal levels and in response to mechanical pressure overload induced by transverse aortic constriction as assessed by echocardiography and hemodynamic studies. Thus, CARP, Ankrd2, and DARP are not essential for normal cardiac development and function at basal conditions and in response to mechanical pressure overload.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Adipose tissue is a key endocrine organ that governs systemic homeostasis. PPARγ is a master regulator of adipose tissue signaling that plays an essential role in insulin sensitivity, making it an ...important therapeutic target. The selective PPARγ agonist rosiglitazone (RSG) has been used to treat diabetes. However, adverse cardiovascular effects have seriously hindered its clinical application. Experimental models have revealed that PPARγ activation increases cardiac hypertrophy. RSG stimulates cardiac hypertrophy and oxidative stress in cardiomyocyte-specific PPARγ knockout mice, implying that RSG might stimulate cardiac hypertrophy independently of cardiomyocyte PPARγ. However, candidate cell types responsible for RSG-induced cardiomyocyte hypertrophy remain unexplored. Utilizing cocultures of adipocytes and cardiomyocytes, we found that stimulation of PPARγ signaling in adipocytes increased miR-200a expression and secretion. Delivery of miR-200a in adipocyte-derived exosomes to cardiomyocytes resulted in decreased TSC1 and subsequent mTOR activation, leading to cardiomyocyte hypertrophy. Treatment with an antagomir to miR-200a blunted this hypertrophic response in cardiomyocytes. In vivo, specific ablation of PPARγ in adipocytes was sufficient to blunt hypertrophy induced by RSG treatment. By delineating mechanisms by which RSG elicits cardiac hypertrophy, we have identified pathways that mediate the crosstalk between adipocytes and cardiomyocytes to regulate cardiac remodeling.
Pompe disease, an autosomal recessive disorder caused by deficient lysosomal acid α-glucosidase (GAA), is characterized by accumulation of intra-lysosomal glycogen in skeletal and oftentimes cardiac ...muscle. The c.1935C>A (p.Asp645Glu) variant, the most frequent GAA pathogenic mutation in people of Southern Han Chinese ancestry, causes infantile-onset Pompe disease (IOPD), presenting neonatally with severe hypertrophic cardiomyopathy, profound muscle hypotonia, respiratory failure, and infantile mortality. We applied CRISPR-Cas9 homology-directed repair (HDR) using a novel dual sgRNA approach flanking the target site to generate a Gaa
knock-in mouse model and a myoblast cell line carrying the Gaa c.1935C>A mutation. Herein we describe the molecular, biochemical, histological, physiological, and behavioral characterization of 3-month-old homozygous Gaa
mice. Homozygous Gaa
knock-in mice exhibited normal Gaa mRNA expression levels relative to wild-type mice, had near-abolished GAA enzymatic activity, markedly increased tissue glycogen storage, and concomitantly impaired autophagy. Three-month-old mice demonstrated skeletal muscle weakness and hypertrophic cardiomyopathy but no premature mortality. The Gaa
knock-in mouse model recapitulates multiple salient aspects of human IOPD caused by the GAA c.1935C>A pathogenic variant. It is an ideal model to assess innovative therapies to treat IOPD, including personalized therapeutic strategies that correct pathogenic variants, restore GAA activity and produce functional phenotypes.
Abstract
People heterozygous for an activating mutation in protein kinase G1 (
PRKG1
, p.Arg177Gln) develop thoracic aortic aneurysms and dissections (TAAD) as young adults. Here we report that mice ...heterozygous for the mutation have a three-fold increase in basal protein kinase G (PKG) activity, and develop age-dependent aortic dilation.
Prkg1
R177Q/+
aortas show increased smooth muscle cell apoptosis, elastin fiber breaks, and oxidative stress compared to aortas from wild type littermates. Transverse aortic constriction (TAC)—to increase wall stress in the ascending aorta—induces severe aortic pathology and mortality from aortic rupture in young mutant mice. The free radical-neutralizing vitamin B
12
-analog cobinamide completely prevents age-related aortic wall degeneration, and the unrelated anti-oxidant N-acetylcysteine ameliorates TAC-induced pathology. Thus, increased basal PKG activity induces oxidative stress in the aorta, raising concern about the widespread clinical use of PKG-activating drugs. Cobinamide could be a treatment for aortic aneurysms where oxidative stress contributes to the disease, including Marfan syndrome.
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