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Histone‐lysine N‐methyltransferase SETD2 (SETD2), the sole histone methyltransferase that catalyzes trimethylation of lysine 36 on histone H3 (H3K36me3), is often mutated in clear cell renal cell carcinoma (ccRCC). SETD2 mutation and/or loss of H3K36me3 is linked to metastasis and poor outcome in ccRCC patients. Epithelial‐to‐mesenchymal transition (EMT) is a major pathway that drives invasion and metastasis in various cancer types. Here, using novel kidney epithelial cell lines isogenic for SETD2, we discovered that SETD2 inactivation drives EMT and promotes migration, invasion, and stemness in a transforming growth factor‐beta‐independent manner. This newly identified EMT program is triggered in part through secreted factors, including cytokines and growth factors, and through transcriptional reprogramming. RNA‐seq and assay for transposase‐accessible chromatin sequencing uncovered key transcription factors upregulated upon SETD2 loss, including SOX2, POU2F2 (OCT2), and PRRX1, that could individually drive EMT and stemness phenotypes in SETD2 wild‐type (WT) cells. Public expression data from SETD2 WT/mutant ccRCC support the EMT transcriptional signatures derived from cell line models. In summary, our studies reveal that SETD2 is a key regulator of EMT phenotypes through cell‐intrinsic and cell‐extrinsic mechanisms that help explain the association between SETD2 loss and ccRCC metastasis. [ABSTRACT FROM AUTHOR]

Intestinal ischemia underlies several clinical conditions and can result in the loss of the intestinal mucosal barrier. Ischemia‐induced damage to the intestinal epithelium is repaired by stimulation of intestinal stem cells (ISCs), and paracrine signaling from the vascular niche regulates intestinal regeneration. Here, we identify FOXC1 and FOXC2 as essential regulators of paracrine signaling in intestinal regeneration after ischemia–reperfusion (I/R) injury. Vascular endothelial cell (EC)‐ and lymphatic EC (LEC)‐specific deletions of Foxc1, Foxc2, or both in mice worsen I/R‐induced intestinal damage by causing defects in vascular regrowth, expression of chemokine CXCL12 and Wnt activator R‐spondin 3 (RSPO3) in blood ECs (BECs) and LECs, respectively, and activation of Wnt signaling in ISCs. Both FOXC1 and FOXC2 directly bind to regulatory elements of the CXCL12 and RSPO3 loci in BECs and LECs, respectively. Treatment with CXCL12 and RSPO3 rescues the I/R‐induced intestinal damage in EC‐ and LEC‐Foxc mutant mice, respectively. This study provides evidence that FOXC1 and FOXC2 are required for intestinal regeneration by stimulating paracrine CXCL12 and Wnt signaling. Synopsis: The transcriptional activity of FOXC1 and FOXC2 in lymphatic and blood endothelial cells in the small intestine contributes to vascular repair and intestinal regeneration after ischemia‐reperfusion injury by regulating CXCL12 and R‐spondin3 signalling, respectively. Both FOXC1 and FOXC2 directly regulate the expression of RSPO3 and CXCL12 in the lymphatic/blood vessels near the crypts, respectively.After ischemia‐reperfusion injury, RSPO3 secreted by LECs stimulates the Wnt/β‐catenin signaling pathway and promotes intestinal epithelial regeneration and repair. LEC‐derived RSPO3 also promotes angiogenesis and lymphangiogenesis.After ischemia‐reperfusion injury, BEC‐derived CXCL12 not only regulates angiogenesis but also stimulates CXCR4 expressed on LECs to enhance lymphangiogenesis. [ABSTRACT FROM AUTHOR]

The realization that soluble factors secreted by heterotypic cells play an importanta role in paracrine signaling, which facilitates intercellular communication, enabled the development of physiologically relevant co‐culture models for drug screening and the engineering of tissues, such as hepatic tissues. The most crucial issues confronting the use of conventional membrane inserts in segregated co‐culture models that are used to study paracrine signaling between heterotypic cells have been identified as long‐term viability and retention of cell‐specific functions, especially when isolated primary cells are used. Herein, we present an in vitro segregated co‐culture model consisting of a well plate incubated with rat primary hepatocytes and normal human dermal fibroblasts which were segregated using a membrane insert with silica nonwoven fabric (SNF) on it. SNF, which mimics a physiological environment much more effectively than a two‐dimensional (2D) one, promotes cell differentiation and resultant paracrine signaling in a manner that is not possible in a conventional 2D culture, owing to high mechanical strength generated by its inorganic materials and interconnected network structure. In segregated co‐cultures, SNF clearly enhanced the functions of hepatocytes and fibroblasts, thereby showing its potential as a measure of paracrine signaling. These results may advance the understanding of the role played by paracrine signaling in cell‐to‐cell communication and provide novel insights into the applications of drug metabolism, tissue repair, and regeneration. [ABSTRACT FROM AUTHOR]

Adipose tissue secretes an abundance of lipid and protein mediators, and this secretome is depot‐specific, with local and systemic effects on metabolic regulation. Intermuscular adipose tissue (IMAT) accumulates within the skeletal muscle compartment in obesity, and is associated with insulin resistance and metabolic disease. While the human IMAT secretome decreases insulin sensitivity in vitro, its composition is entirely unknown. The current study was conducted to investigate the composition of the human IMAT secretome, compared to that of the subcutaneous (SAT) and visceral adipose tissue (VAT) depots. IMAT, SAT, and VAT explants from individuals with obesity were used to generate conditioned media. Proteomics analysis of conditioned media was performed using multiplex proximity extension assays, and eicosanoid analysis using liquid chromatography–tandem mass spectrometry. Compared to SAT and/or VAT, IMAT secreted significantly more cytokines (IL2, IL5, IL10, IL13, IL27, FGF23, IFNγ and CSF1) and chemokines (MCP1, IL8, CCL11, CCL20, CCL25 and CCL27). Adipokines hepatocyte growth factor and resistin were secreted significantly more by IMAT than SAT or VAT. IMAT secreted significantly more eicosanoids (PGE2, TXB2, 5‐HETE, and 12‐HETE) compared to SAT and/or VAT. In the context of obesity, IMAT is a distinct adipose tissue with a highly immunogenic and inflammatory secretome, and given its proximity to skeletal muscle, may be critical to glucose regulation and insulin resistance. [ABSTRACT FROM AUTHOR]

Cardiac fibroblasts constitute a major cell population in the heart. They secrete extracellular matrix components and various other factors shaping the microenvironment of the heart. In silico analysis of intercellular communication based on single-cell RNA sequencing revealed that fibroblasts are the source of the majority of outgoing signals to other cell types. This observation suggests that fibroblasts play key roles in orchestrating cellular interactions that maintain organ homeostasis but that can also contribute to disease states. Here, we will review the current knowledge of fibroblast interactions in the healthy, diseased, and aging heart. We focus on the interactions that fibroblasts establish with other cells of the heart, specifically cardiomyocytes, endothelial cells and immune cells, and particularly those relying on paracrine, electrical, and exosomal communication modes. [ABSTRACT FROM AUTHOR]

Background: The mammalian yolk sac provides nutrients for the growing fetus during critical early developmental processes such as neural tube closure, which precedes the functional maturation of the placenta. In contrast, oviparous species such as the chicken rely solely on the yolk sac for transfer of nutrients from the yolk to the developing embryo. However, the molecular mechanisms that provide the yolk sac with nutrient transfer competence remain poorly understood. Results: We demonstrate that the chicken endodermal epithelial cells (EEC), which are in close contact with the yolk, gain their nutrient‐transport competence by a paracrine crosstalk with the blood‐vessel forming mesodermal cell layer. Bone morphogenetic proteins (BMP) 4 and 7 produced by ectodermal and mesodermal cell layers likely initiate a differentiation program of EECs during the transition from the area vitellina to the area vasculosa. BMPs, by inducing SMAD signaling, promote the up‐regulation of endocytic receptor expression and thereby provide the EECs with the molecular machinery to produce triglyceride‐rich lipoprotein particles. Conclusion: This paracrine signaling cascade may constitute the basis for the EEC‐mediated mechanism underlying the efficient uptake, degradation, resynthesis, and transfer of yolk‐derived nutrients into the embryonic circulation, which assures proper energy supply and development of the growing fetus. Key Findings: Endodermal epithelial cells gain their nutrient transport competence during the transition from the area vitellina to the area vasculosa.BMPs signal from the ecto/mesodermal cell sheet to the underlying endoderm, thereby initiating endoderm specification.BMPs thereby provide the endoderm with nutrient transport competence which allows proper nutrition of the growing embryo. [ABSTRACT FROM AUTHOR]

Primary melanocytes isolated from skin and expanded in culture have been widely used for laboratory research and clinical applications. The conventional method to isolate primary melanocytes from skin usually requires about 3–4 weeks of culture for melanocytes to grow sufficiently to passage. Considering that melanocytes comprise only 3%–7% of epidermal cells in normal human skin, it would be extremely helpful to increase the isolation efficiency and shorten the initial culture time to quickly meet various application needs. Here, we report that adding Y‐27632, a Rho kinase inhibitor, into the initial culture medium for 2 days can dramatically increase the yield of melanocytes. We found that Y‐27632 can promote keratinocyte attachment and survival in the melanocyte culture system, resulting in not only better recovery, but also increased proliferation of melanocytes by a paracrine signaling pathway. More specifically, Y‐27632 significantly induced keratinocyte expression of stem cell factor, which played an important role in enhancing the growth of melanocytes. In summary, Y‐27632 could profoundly enhance the yield of primary melanocytes in the initial culture through paracrine effects on keratinocytes. [ABSTRACT FROM AUTHOR]

Accumulation of adipose tissue around and within muscles is highly correlated with reduced strength, functional limitations, and poor rehabilitative outcomes. Given the intimate physical contact between these tissues, paracrine cross-talk is a likely mediator of this association. The recent discovery that muscle-associated adipose tissue exhibits features of beige fat has suggested that this cross-talk may be modifiable, as beige fat can be stimulated to assume features of brown fat. In this work, we describe a novel intermuscular fat transplant model in the mouse rotator cuff to investigate cross-talk between muscle and adipose tissue. Specifically, we examine the role of transplanted fat phenotype on muscle regeneration by transplanting pieces of classical brown (interscapular), beige (inguinal), or white (epididymal) adipose tissue in conjunction with cardiotoxin injection to the adjacent supraspinatus muscle. Transplantation of brown fat, but not beige or white, significantly increased muscle mass, fiber cross-sectional area and contractile force production compared with sham injury. This effect was not seen when cardiotoxin was delivered to a distant muscle, or when adjacent muscles were injected with saline indicating that the effect is localized and specifically targeting the regenerative process. Thus, we conclude that local signaling between fat and muscle varies by phenotype and that brown fat supports regeneration. Clinical significance: Our findings suggest that the phenotype of muscle-associated fat could be a novel therapeutic target to modulate fat-muscle signaling. [ABSTRACT FROM AUTHOR]

Vascularization of the fracture site and cell‐mediated deposition of the mineralized matrix are crucial determinants for successful bone regeneration after injury. Ceramic biomaterials such as bioactive glasses (BAGs) that release bioactive ions have shown promising results in bone defect regeneration. However, it remains unclear how the dosage and composition of bioactive ions influence the angiogenic and osteogenic behavior of primary human mesenchymal stromal cells (MSCs). Here, we show that exposure to ionic dissolution products from 1393 and 45S5 BAGs can evoke distinct angiogenic and osteogenic responses from primary MSCs in a dose‐ and composition‐dependent manner. Significantly higher concentrations of the pro‐angiogenic factors VEGF, HGF, PIGF, angiopoietin, and angiogenin were detected in conditioned media (CM) from MSCs exposed to 45S5, but not 1393, BAGs. Application of this CM to human umbilical vein endothelial cells (HUVECs) resulted in robust 2D tube formation in vitro. Osteogenic differentiation of MSCs was assessed by gene expression analysis and mineralization assays. Low concentrations (0.1% w/v) of 1393 BAGs significantly enhanced the gene expression of RUNX2 and ALP and induced an earlier onset of matrix mineralization compared to all other groups. We further tested whether simultaneous exposure to both BAGs would improve both angiogenic secretion and osteogenic differentiation of MSCs, and did not find evidence to support this hypothesis. Our results provide evidence of BAG composition‐dependent enhancement of primary human MSCs' regenerative function, besides also underlining the importance of an in vitro evaluation of the dose–response relationship to translate BAG based approaches into safe and effective clinical therapies. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2827–2837, 2018., 2018 [ABSTRACT FROM AUTHOR]

Background: Fibronectin extracellular matrix is essential for embryogenesis. Its assembly is a cell-mediated process where secreted fibronectin dimers bind to integrin receptors on receiving cells, which actively assemble fibronectin into a fibrillar matrix. During development, paracrine communication between tissues is crucial for coordinating morphogenesis, typically being mediated by growth factors and their receptors. Recent reports of situations where fibronectin is produced by one tissue and assembled by another, with implications on tissue morphogenesis, suggest that fibronectin assembly may also be a paracrine communication event in certain contexts. Results: Here we addressed which tissues express fibronectin ( Fn1) while also localizing assembled fibronectin matrix and determining the mRNA expression and/or protein distribution pattern of integrins α5 and αV, α chains of the major fibronectin assembly receptors, during early chick and mouse development. We found evidence supporting a paracrine system in fibronectin matrix assembly in several tissues, including immature mesenchymal tissues, components of central and peripheral nervous system and developing muscle. Conclusions: Thus, similarly to growth factor signaling, fibronectin matrix assembly during early development can be both autocrine and paracrine. We therefore propose that it be considered a cell-cell communication event at the same level and significance as growth factor signaling during embryogenesis. Developmental Dynamics 245:520-535, 2016. © 2015 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Cardiac fibrosis is an essential pathological process in pressure overload (PO)-induced heart failure. Recently, myocyte-fibroblast communication is proven to be critical in heart failure, in which, pathological growth of cardiomyocytes (CMs) may promote fibrosis via miRNAs-containing exosomes (Exos). Peli1 regulates the activation of NF-κB and AP-1, which has been demonstrated to engage in miRNA transcription in cardiomyocytes. Therefore, we hypothesized that Peli1 in CMs regulates the activation of cardiac fibroblasts (CFs) through an exosomal miRNA-mediated paracrine mechanism, thereby promoting cardiac fibrosis. We found that CM-conditional deletion of Peli1 improved PO-induced cardiac fibrosis. Moreover, Exos from mechanical stretch (MS)-induced WT CMs (WT MS-Exos) promote activation of CFs, Peli1-/- MS-Exos reversed it. Furthermore, miRNA microarray and qPCR analysis showed that miR-494-3p was increased in WT MS-Exos while being down regulated in Peli1-/- MS-Exos. Mechanistically, Peli1 promoted miR-494-3p expression via NF-κB/AP-1 in CMs, and then miR-494-3p induced CFs activation by inhibiting PTEN and amplifying the phosphorylation of AKT, SMAD2/3, and ERK. Collectively, our study suggests that CMs Peli1 contributes to myocardial fibrosis via CMs-derived miR-494-3p-enriched exosomes under PO, and provides a potential exosomal miRNA-based therapy for cardiac fibrosis. [ABSTRACT FROM AUTHOR]

Recent findings in several organ systems show that cytoneme-mediated signaling transports signaling proteins along cellular extensions and targets cell-to-cell exchanges to synaptic contacts. This mechanism of paracrine signaling may be a general one that is used by many (or all) cell types in many (or all) organs. We briefly review these findings in this perspective. We also describe the properties of several signaling systems that have previously been interpreted to support a passive diffusion mechanism of signaling protein dispersion, but can now be understood in the context of the cytoneme mechanism. Also watch the . [ABSTRACT FROM AUTHOR]

Fibroblast growth factor (FGF2) regulates endothelial and melanoma cell migration. The binding of FGF2 to its receptor requires N-sulfated heparan sulfate (HS) glycosamine. We have previously reported that Epac1, an exchange protein activated by cAMP, increases N-sulfation of HS in melanoma. Therefore, we examined whether Epac1 regulates FGF2-mediated cell-cell communication. Conditioned medium (CM) of melanoma cells with abundant expression of Epac1 increased migration of human umbilical endothelial cells (HUVEC) and melanoma cells with poor expression of Epac1. CM-induced increase in migration was inhibited by antagonizing FGF2, by the removal of HS and by the knockdown of Epac1. In addition, knockdown of Epac1 suppressed the binding of FGF2 to FGF receptor in HUVEC, and in vivo angiogenesis in melanoma. Furthermore, knockdown of Epac1 reduced N-sulfation of HS chains attached to perlecan, a major secreted type of HS proteoglycan that mediates the binding of FGF2 to FGF receptor. These data suggested that Epac1 in melanoma cells regulates melanoma progression via the HS-FGF2-mediated cell-cell communication. [ABSTRACT FROM AUTHOR]

The aim of this study was to explore the effect of bone morphogenetic protein-2 (BMP-2) and fibroblast growth factor-2 (FGF-2)— paracrine factors implicated in both cardiac embryogenesis and cardiac repair following myocardial infarction (MI)—on murine bone marrow stem cell (mBMSC) differentiation in an ex vivo cardiac microenvironment. For this purpose, green fluorescent protein (GFP) expressing hematopoietic lineage negative (lin-) c-kit ligand (c-kit) and stem cell antigen-1 (Sca-1) positive (GFP-lin-/c-kit+/sca+) mBMSC were co-cultured with neonatal rat ventricular cardiomyocytes (NVCMs). GFP+ mBMSC significantly induced the expression of BMP-2 and FGF-2 in NVCMs, and approximately 4% GFP+ mBMSCs could be recovered from the co-culture at day 10. The addition of BMP-2 in concert with FGF-2 significantly enhanced the amount of integrated GFP+ mBMSCs by 5-fold (∼20%), whereas the addition of anti-BMP-2 and/or anti-FGF-2 antibodies completely abolished this effect. An analysis of calcium cycling revealed robust calcium transients in GFP+ mBMSCs treated with BMP-2/FGF-2 compared to untreated co-cultures. BMP-2 and FGF-2 addition led to a significant induction of early (NK2 transcription factor related, locus 5; Nkx2.5, GATA binding protein 4; GATA-4) and late (myosin light chain kinase [MLC-2v], connexin 43 [Cx43]) cardiac marker mRNA expression in mBMSCs following co-culture. In addition, re-cultured fluorescence-activated cell sorting (FACS)-purified BMP-2/FGF-2-treated mBMSCs revealed robust calcium transients in response to electrical field stimulation which were inhibited by the L-type calcium channel (LTCC) inhibitor, nifedipine, and displayed caffeine-sensitive intracellular calcium stores. In summary, our results show that mBMSCs can adopt a functional cardiac phenotype through treatment with factors essential to embryonic cardiogenesis that are induced after cardiac ischemia. This study provides the first evidence that mBMSCs with long-term self-renewal potential possess the capability to serve as a functional cardiomyocyte precursor through the appropriate paracrine input and cross-talk within an appropriate cardiac microenvironment. [ABSTRACT FROM AUTHOR]

In many sensory systems, stimulus sensitivity is dynamically modulated through mechanisms of peripheral adaptation, efferent input, or hormonal action. In this way, responses to sensory stimuli can be optimized in the context of both the environment and the physiological state of the animal. Although the gustatory system critically influences food preference, food intake and metabolic homeostasis, the mechanisms for modulating taste sensitivity are poorly understood. In this study, we report that glucagon-like peptide-1 (GLP-1) signaling in taste buds modulates taste sensitivity in behaving mice. We find that GLP-1 is produced in two distinct subsets of mammalian taste cells, while the GLP-1 receptor is expressed on adjacent intragemmal afferent nerve fibers. GLP-1 receptor knockout mice show dramatically reduced taste responses to sweeteners in behavioral assays, indicating that GLP-1 signaling normally acts to maintain or enhance sweet taste sensitivity. A modest increase in citric acid taste sensitivity in these knockout mice suggests GLP-1 signaling may modulate sour taste, as well. Together, these findings suggest a novel paracrine mechanism for the regulation of taste function. [ABSTRACT FROM AUTHOR]

Background: Cardiac mesenchymal cell (CMC) administration improves cardiac function in animal models of heart failure. Although the precise mechanisms remain unclear, transdifferentiation and paracrine signaling are suggested to underlie their cardiac reparative effects. We have shown that histone deacetylase 1 (HDAC1) inhibition enhances CMC cardiomyogenic lineage commitment. Here, we investigated the impact of HDAC1 on CMC cytokine secretion and associated paracrine-mediated activities on endothelial cell function., Methods and Results: CMCs were transduced with shRNA constructs targeting HDAC1 (shHDAC1) or nontarget (shNT) control. Cytokine arrays were used to assess the expression of secreted proteins in conditioned medium (CM) from shHDAC1 or shNT-transduced CMCs. In vitro functional assays for cell proliferation, protection from oxidative stress, cell migration, and tube formation were performed on human endothelial cells incubated with CM from the various treatment conditions. CM from shHDAC1-transduced CMCs contained more cytokines involved in cell growth/differentiation and more efficiently promoted endothelial cell proliferation and tube formation compared with CM from shNT. After evaluating key cytokines previously implicated in cell-therapy-mediated cardiac repair, we found that basic fibroblast growth factor was significantly upregulated in shHDAC1-transduced CMCs. Furthermore, shRNA-mediated knockdown of basic fibroblast growth factor in HDAC1-depleted CMCs inhibited the effects of shHDAC1 CM in promoting endothelial proliferation and tube formation-indicating that HDAC1 depletion activates CMC proangiogenic paracrine signaling in a basic fibroblast growth factor-dependent manner., Conclusions: These results reveal a hitherto unknown role for HDAC1 in the modulation of CMC cytokine secretion and implicate the targeted inhibition of HDAC1 in CMCs as a means to enhance paracrine-mediated neovascularization in cardiac cell therapy applications., (© 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.)