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Cardiac mesenchymal progenitors differentiate into adipocytes via Klf4 and c-Myc.

ABSTRACT: Direct reprogramming of differentiated cells to pluripotent stem cells has great potential to improve our understanding of developmental biology and disorders such as cancers, and has implications for regenerative medicine. In general, the effects of transcription factors (TFs) that are transduced into cells can be influenced by pre-existing transcriptional networks and epigenetic modifications. However, previous work has identified four key TFs, Oct4, Sox2, Klf4 and c-Myc, which can reprogram various differentiated cells to generate induced pluripotent stem cells. Here, we show that in the heart, the transduction of cardiac mesenchymal progenitors (CMPs) with Klf4 and c-Myc (KM) was sufficient to drive the differentiation of these cells into adipocytes without the use of adipogenic stimulation cocktail, that is, insulin, 3-isobutyl-1-methylxanthine (IBMX) and dexamethasone. KM-transduced CMPs exhibited a gradually increased expression of adipogenic-related genes, such as C/Ebp?, Ppar? and Fabp4, activation of the peroxisome proliferator-activated receptor (PPAR) signaling pathway, inactivation of the cell cycle-related pathway and formation of cytoplasmic lipid droplets within 10 days. In contrast, NIH3T3 fibroblasts, 3T3-L1 preadipocytes, and bone marrow-derived mesenchymal stem cells transduced with KM did not differentiate into adipocytes. Both in vitro and in vivo cardiac ischemia reperfusion injury models demonstrated that the expression of KM genes sharply increased following a reperfusion insult. These results suggest that ectopic adipose tissue formation in the heart following myocardial infarction results from CMPs that express KM following a stress response.

SUBMITTER: Kami D

PROVIDER: S-EPMC4855651 | biostudies-literature | 2016 Apr

REPOSITORIES: biostudies-literature

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Cardiac mesenchymal progenitors differentiate into adipocytes via Klf4 and c-Myc.

Kami D D Kitani T T Kawasaki T T Gojo S S

Cell death & disease 20160414

Direct reprogramming of differentiated cells to pluripotent stem cells has great potential to improve our understanding of developmental biology and disorders such as cancers, and has implications for regenerative medicine. In general, the effects of transcription factors (TFs) that are transduced into cells can be influenced by pre-existing transcriptional networks and epigenetic modifications. However, previous work has identified four key TFs, Oct4, Sox2, Klf4 and c-Myc, which can reprogram v ...[more]

PMID: 27077806

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Project description:RationaleMutations in desmosome proteins cause arrhythmogenic cardiomyopathy (AC), a disease characterized by excess myocardial fibroadipocytes. Cellular origin(s) of fibroadipocytes in AC is unknown.ObjectiveTo identify the cellular origin of adipocytes in AC.Methods and resultsHuman and mouse cardiac cells were depleted from myocytes and flow sorted to isolate cells expressing platelet-derived growth factor receptor-α and exclude those expressing other lineage and fibroblast markers (CD32, CD11B, CD45, Lys76, Ly(-6c) and Ly(6c), thymocyte differentiation antigen 1, and discoidin domain receptor 2). The PDGFRA(pos):Lin(neg):THY1(neg):DDR2(neg) cells were bipotential as the majority expressed collagen 1 α-1, a fibroblast marker, and a subset CCAAT/enhancer-binding protein α, a major adipogenic transcription factor, and therefore, they were referred to as fibroadipocyte progenitors (FAPs). FAPs expressed desmosome proteins, including desmoplakin, predominantly in the adipogenic but not fibrogenic subsets. Conditional heterozygous deletion of Dsp in mice using Pdgfra-Cre deleter led to increased fibroadipogenesis in the heart and mild cardiac dysfunction. Genetic fate mapping tagged 41.4±4.1% of the cardiac adipocytes in the Pdgfra-Cre:Eyfp:Dsp(W/F) mice, indicating an origin from FAPs. FAPs isolated from the Pdgfra-Cre:Eyfp:Dsp(W/F) mouse hearts showed enhanced differentiation to adipocytes. Mechanistically, deletion of Dsp was associated with suppressed canonical Wnt signaling and enhanced adipogenesis. In contrast, activation of the canonical Wnt signaling rescued adipogenesis in a dose-dependent manner.ConclusionsA subset of cardiac FAPs, identified by the PDGFRA(pos):Lin(neg):THY1(neg):DDR2(neg) signature, expresses desmosome proteins and differentiates to adipocytes in AC through a Wnt-dependent mechanism. The findings expand the cellular spectrum of AC, commonly recognized as a disease of cardiac myocytes, to include nonmyocyte cells in the heart.

Project description:The mammalian heart has little capacity to regenerate, and following injury the myocardium is replaced by non-contractile scar tissue. Consequently, increased wall stress and workload on the remaining myocardium leads to chamber dilation, dysfunction, and heart failure. Cell-based therapy with an autologous, epigenetically reprogrammed, and cardiac-committed progenitor cell source could potentially reverse this process by replacing the damaged myocardium with functional tissue. However, it is unclear whether cardiac progenitor cell-derived cardiomyocytes are capable of attaining levels of structural and functional maturity comparable to that of terminally-fated cardiomyocytes. Here, we first describe the derivation of mouse induced pluripotent stem (iPS) cells, which once differentiated allow for the enrichment of Nkx2-5(+) cardiac progenitors, and the cardiomyocyte-specific expression of the red fluorescent protein. We show that the cardiac progenitors are multipotent and capable of differentiating into endothelial cells, smooth muscle cells and cardiomyocytes. Moreover, cardiac progenitor selection corresponds to cKit(+) cell enrichment, while cardiomyocyte cell-lineage commitment is concomitant with dual expression of either cKit/Flk1 or cKit/Sca-1. We proceed to show that the cardiac progenitor-derived cardiomyocytes are capable of forming electrically and mechanically coupled large-scale 2D cell cultures with mature electrophysiological properties. Finally, we examine the cell progenitors' ability to form electromechanically coherent macroscopic tissues, using a physiologically relevant 3D culture model and demonstrate that following long-term culture the cardiomyocytes align, and form robust electromechanical connections throughout the volume of the biosynthetic tissue construct. We conclude that the iPS cell-derived cardiac progenitors are a robust cell source for tissue engineering applications and a 3D culture platform for pharmacological screening and drug development studies.

Project description:Homocysteine (Hcy) was discovered to be an independent risk factor for the development of atherosclerosis (AS). Moreover, endothelial‑mesenchymal transition (EndMT) was found to be one of main mechanisms contributing to the pathogenesis of AS. Salidroside (SAL) has diverse pharmacological activities, including anti‑inflammatory, anti‑cancer, anti‑oxidative and anti‑fibrosis properties. However, whether SAL serves a beneficial role in Hcy‑induced EndMT remains unknown. The present study aimed to investigate whether SAL exerted its effects on Hcy‑induced EndMT via the Kruppel‑like factor 4 (KLF4)/endothelial nitric oxide (NO) synthase (eNOS) signaling pathway. HUVECs were pretreated with high and low doses (10 or 50 µmol/l) of SAL for 2 h, followed by 1 mmol/l Hcy for 48 h to induce EndMT. Western blotting was used to analyze the protein expression levels of the endothelial marker, VE‑cadherin, the mesenchymal cell marker, α‑smooth muscle actin (SMA), and the nuclear transcription factors, KLF4 and eNOS. Wound healing assays were used to determine the cell migratory ability, and the levels of NO in the cell culture supernatants were measured using a nitrate reductase assay. Cellular immunofluorescence was used to analyze the expression and localization of KLF4. Small interfering (si)RNA targeting KLF4 (siKLF4) was used to knock down KLF4 expression in HUVECs. The results of the present study revealed that treatment with SAL upregulated the expression levels of VE‑cadherin, downregulated the expression levels of α‑SMA, reduced cell migration and activated the eNOS/NO signaling axis, as well as downregulated KLF4 expression and translocation to the nucleus. Compared with the SAL + siKLF4 co‑administration group, no significant differences were observed in the expression levels of the phenotypic markers in the SAL or siKLF4 groups. In conclusion, the findings of the present study revealed that SAL may inhibit Hcy‑induced EndMT via regulation of the KLF4/eNOS signaling pathway.

Dataset Information

Cardiac mesenchymal progenitors differentiate into adipocytes via Klf4 and c-Myc.

Publications

Cardiac mesenchymal progenitors differentiate into adipocytes via Klf4 and c-Myc.

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