Tissue-specific progenitor cells contribute to local cellular regeneration and maintain organ function. Recently, we have determined that cardiac side-population (CSP) cells represent a distinct cardiac progenitor cell population, capable of in vitro differentiation into functional cardiomyocytes. The response of endogenous CSP to myocardial injury, however, and the cellular mechanisms that maintain this cardiac progenitor cell pool in vivo remain unknown. In this report we demonstrate that local progenitor cell proliferation maintains CSP under physiologic conditions, with little contribution from extracardiac stem cell sources. Following myocardial infarction in adult mice, however, CSP cells are acutely depleted, both within the infarct and noninfarct areas. CSP pools are subsequently reconstituted to baseline levels within 7 days after myocardial infarction, through both proliferation of resident CSP cells, as well as through homing of bone marrow-derived stem cells (BMC) to specific areas of myocardial injury and immunophenotypic conversion of BMC to adopt a CSP phenotype. We, therefore, conclude that following myocardial injury, cardiac progenitor cell populations are acutely depleted and are reconstituted to normal levels by both self-proliferation and selective homing of BMC. Understanding and enhancing such processes hold enormous potential for therapeutic myocardial regeneration.T issue-specific progenitor cell populations maintain the regenerative capacity of terminally differentiated organs, both under basal conditions and following local tissue injury. Side population (SP) cells, characterized by their intrinsic capacity to efflux Hoechst dye through ATP-binding cassette transporters, contribute to the long-term regenerative potential of hematopoietic and extrahematopoietic tissues. 1 Recently, we have demonstrated that cardiac SP (CSP) cells, immunophenotypically distinct from bone marrow (BM)-derived stem cells (BMC), are present in the adult heart and are capable of both biochemical and functional cardiomyogenic differentiation into mature cardiomyocytes, thereby identifying CSP as a distinct cardiac progenitor cell population. 2 Supplementation of cardiac progenitor cell pools after myocardial infarction (MI) with exogenous cells has been shown to improve ventricular function by regenerating myocardium and cardiac vasculature. [3][4][5] The response of endogenous CSP cells to myocardial injury, however, and the cellular mechanisms that maintain this endogenous cardiac progenitor cell pool under basal and after injury conditions remain unknown. We, therefore, serially assessed CSP pools in hearts following MI and determined the role of selfproliferation and BMC in reconstituting cardiac progenitor pools. Methods and MaterialsCSP were isolated from mouse hearts as described previously. 2 MI was performed in mice via permanent coronary ligation. 6 BM transplantation was performed in lethally irradiated mice using marrow isolated from C57bl/6-Tg(ACTbEGFP) mice. 7 All animals were obtained from The ...

The advent of human induced pluripotent stem cells (iPSCs) presents unprecedented opportunities to model human diseases. Differentiated cells derived from iPSCs in two-dimensional (2D) monolayers have proven to be a relatively simple tool for exploring disease pathogenesis and underlying mechanisms. In this Spotlight article, we discuss the progress and limitations of the current 2D iPSC disease-modeling platform, as well as recent advancements in the development of human iPSC models that mimic tissues and organs at the three-dimensional (3D) level. Recent bioengineering approaches have begun to combine different 3D organoid types into a single '4D multi-organ system'. We summarize the advantages of this approach and speculate on the future role of 4D multi-organ systems in human disease modeling.

Due to their immunosuppressive properties, mesenchymal stem cells (MSC) have been evaluated for the treatment of immunological diseases. However, the animal-derived growth supplements utilized for MSC manufacturing may lead to clinical complications. Characterization of alternative media formulations is imperative for MSC therapeutic application. Human BMMSC and AdMSC were expanded in media supplemented with either human platelet lysates (HPL), serum-free media/xeno-free FDA-approved culture medium (SFM/XF), or fetal bovine serum (FBS) and the effects on their properties were investigated. The immunophenotype of resting and IFN-γ primed BMMSC and AdMSC remained unaltered in all media. Both HPL and SFM/XF increased the proliferation of BMMSC and AdMSC. Expansion of BMMSC and AdMSC in HPL increased their differentiation, compared to SFM/XF and FBS. Resting BMMSC and AdMSC, expanded in FBS or SFM/XF, demonstrated potent immunosuppressive properties in both non-primed and IFN-γ primed conditions, whereas HPL-expanded MSC exhibited diminished immunosuppressive properties. Finally, IFN-γ primed BMMSC and AdMSC expanded in SFM/XF and HPL expressed attenuated levels of IDO-1 compared to FBS. Herein, we provide strong evidence supporting the use of the FDA-approved SFM/XF medium, in contrast to the HPL medium, for the expansion of MSC towards therapeutic applications.

Background: Molecular targeted chemotherapies have been shown to significantly improve cancer patient outcomes, but often cause cardiovascular side effects that limit their use and impair patients' quality of life. Cardiac dysfunction induced by these therapies, especially trastuzumab, shows a distinct cardiotoxic clinical phenotype compared to cardiotoxicity induced by conventional chemotherapies. Methods: We employed the human induced pluripotent stem cell-derived cardiomyocyte (iPSC-CM) platform to determine the underlying cellular mechanisms in trastuzumab-induced cardiac dysfunction. We assessed the effects of trastuzumab on structural and functional properties in iPSC-CMs from healthy individuals and performed RNA-sequencing (RNA-seq) to further examine the effect of trastuzumab on iPSC-CMs. We also generated iPSCs from patients receiving trastuzumab and examined whether patients' phenotype could be recapitulated in vitro using patient-specific iPSC-CMs.

Abstract-Recently, the side population (SP) phenotype has been introduced as a reliable marker to identify subpopulations of cells with stem/progenitor cell properties in various tissues. We and others have identified SP cells from postmitotic tissues, including adult myocardium, in which they have been suggested to contribute to cellular regeneration following injury. SP cells are identified and characterized by a unique efflux of Hoechst 33342 dye. Key Words: Abcg2 Ⅲ Mdr1 Ⅲ progenitor cells Ⅲ proliferation Ⅲ SP cells R ecently, the side population (SP) phenotype has been introduced as a reliable marker to identify subpopulations of cells with stem/progenitor cell properties in various tissues including the heart. 1 On the molecular level, the SP phenotype is linked to the presence of ATP-binding cassette (ABC) transporters with the ability to efficiently efflux the DNA binding dye Hoechst 33342. 2 This ABC transporterdependent Hoechst efflux phenomenon confers the characteristic fluorescent-activated cell sorting (FACS) profile of SP cells as a Hoechst-low "side population" located to the periphery of the Hoechst-high main population. 2 Among the various members of the ABC transporter superfamily, Abcg2 (also referred to as breast cancer resistance protein 1 [Bcrp1]) and Mdr1 (also referred to as P-glycoprotein [p-gp] or Abcb1) have been shown to efficiently efflux Hoechst 33342 and thereby confer the SP phenotype. 3 Although both transporters are highly expressed in bone marrow (BM)SP cells, studies performed in mice with targeted disruption of the Mdr1a and Mdr1b genes, the murine homologs of the human Abcb1/Mdr1 gene, demonstrated that Abcg2 is the sole molecular determinant of the SP phenotype in hematopoietic stem cells. 4 Moreover, Abcg2 expression is conserved in SP cells from a wide range of tissues including blood, gonad, lung, skeletal muscle and the retina, suggesting an important role of Abcg2 in stem cells. 4 -7 We and others have characterized SP cells isolated from adult myocardium. 8 -11 These cardiac (c)SP cells are phenotypically distinct from BMSP cells, in that they are not hematopoietic but exhibit the potential to differentiate into functional cardiomyocytes. 10 As in SP cells from the bone marrow, Abcg2 is expressed in SP cells from the heart. 9 The contribution of Abcg2 to the cSP phenotype and its biological significance in cSP progenitor cells, however, remain unknown. In this study, we find that the contribution of Abcg2 to the SP phenotype in the heart exists in an age-dependent manner, with Abcg2 as the molecular determinant of the SP phenotype in the neonatal heart and Mdr1 as the basis for the SP phenotype in the adult heart. In addition, we demonstrate Original

Rationale Multiple progenitors derived from the heart and bone marrow have been utilized for cardiac repair. Despite this, not much is known about the molecular identity and relationship among these progenitors. To develop a robust stem cell therapy for the heart, it is critical to understand the molecular identity of the multiple ‘cardiogenic progenitor cells’ (CPCs). This study is the first report of high throughput transcriptional profiling of CPCs carried out on an identical platform. Method and Results Microarray based transcriptional profiling was carried out for three cardiac (ckit+, Sca1+, side population) and two bone marrow (ckit+ , mesenchymal stem cell) progenitors, obtained from age- and sex-matched wild type C57BL/6 mice. Analysis indicated that cardiac-derived ckit+ population was very distinct from Sca1+ and SP cells in the downregulation of genes encoding for cell-cell and matrix adhesion proteins, and in the upregulation of developmental genes. Significant enrichment of transcripts involved in DNA replication and repair was observed in bone marrow (BM)-derived progenitors. The BM ckit+ cells appeared to have the least correlation with the other progenitors, with enrichment of immature neutrophil specific molecules. Conclusion Our study indicates that cardiac ckit+ cells represent the most primitive population in the rodent heart. Primitive cells of cardiac versus BM origin differ significantly with respect to stemness and cardiac lineage-specific genes, and molecules involved in DNA replication and repair. The detailed molecular profile of progenitors reported here will serve as a useful reference to determine the molecular identity of progenitors used in future preclinical and clinical studies

Background & Aims Persistent activation of the inflammatory response contributes to development of inflammatory bowel diseases, which increase the risk of colorectal cancer. We aimed to identify microRNAs that regulate inflammation during development of ulcerative colitis (UC) and progression to colitis-associated colon cancer (CAC). Methods We performed quantititave PCR analysis to measure microRNAs in 401 colon specimens from patients with UC, Crohn's disease, irritable bowel syndrome, sporadic colorectal cancer, or CAC, as well as subjects without these disorders (controls); levels were correlated with clinical features and disease activity of patients. Colitis was induced in mice by administration of dextran sodium sulfate (DSS), and carcinogenesis was induced by addition of azoxymethane; some mice were also given inhibitor of microRNA214 (miR214). Results A high-throughput functional screen of the human microRNAome found that miR214 regulated the activity of nuclear factor κB (NFκB). Higher levels of miR214 were detected in colon tissues from patients with active UC or CAC than patients with other disorders or controls and correlated with disease progression. Bioinformatic and genome-wide profile analyses revealed that miR214 activates an inflammatory response and is amplified through a feedback loop circuit mediated by phosphatase and tensin homolog (PTEN) and PDZ and LIM domain 2 (PDLIM2). Interleukin-6 induced STAT3-mediated transcription of miR214. A miR214 chemical inhibitor blocked this circuit and reduced the severity of DSS-induced colitis in mice, as well as the number and size of tumors that formed in mice given azoxymethane and DSS. In fresh colonic biopsies from patients with active UC, the miR214 inhibitor reduced inflammation by increasing levels of PDLIM2 and PTEN. Conclusions Interleukin-6 upregulates STAT3-mediated transcription of miR214 in colon tissues, which reduces levels of PDLIM2 and PTEN, increases phosphorylation of AKT, and activates NFκB. The activity of this circuit correlates with disease activity in patients with UC and progression to colorectal cancer.

CD4 T cells are tightly regulated by microbiota in the intestine, but whether intestinal T cells interface with host-derived metabolites is less clear. Here, we show that CD4 T effector (Teff) cells upregulated the xenobiotic transporter, Mdr1, in the ileum to maintain homeostasis in the presence of bile acids. Whereas wild-type Teff cells upregulated Mdr1 in the ileum, those lacking Mdr1 displayed mucosal dysfunction and induced Crohn's disease-like ileitis following transfer into Rag1 hosts. Mdr1 mitigated oxidative stress and enforced homeostasis in Teff cells exposed to conjugated bile acids (CBAs), a class of liver-derived emulsifying agents that actively circulate through the ileal mucosa. Blocking ileal CBA reabsorption in transferred Rag1 mice restored Mdr1-deficient Teff cell homeostasis and attenuated ileitis. Further, a subset of ileal Crohn's disease patients displayed MDR1 loss of function. Together, these results suggest that coordinated interaction between mucosal Teff cells and CBAs in the ileum regulate intestinal immune homeostasis.