The emerging roles of Hedgehog signaling in tumor immune microenvironment.

The Hedgehog (Hh) signaling pathway is pervasively involved in human malignancies, making it an effective target for cancer treatment for decades. In addition to its direct role in regulating cancer cell attributes, recent work indicates that it has an immunoregulatory effect on tumor microenvironments. An integrated understanding of these actions of Hh signaling pathway in tumor cells and tumor microenvironments will pave the way for novel tumor treatments and further advances in anti-tumor immunotherapy. In this review, we discuss the most recent research about Hh signaling pathway transduction, with a particular emphasis on its role in modulating tumor immune/stroma cell phenotype and function, such as macrophage polarity, T cell response, and fibroblast activation, as well as their mutual interactions between tumor cells and nonneoplastic cells. We also summarize the recent advances in the development of Hh pathway inhibitors and nanoparticle formulation for Hh pathway modulation. We suggest that targeting Hh signaling effects on both tumor cells and tumor immune microenvironments could be more synergistic for cancer treatment.

Overexpression of KCNJ2 enhances maturation of human-induced pluripotent stem cell-derived cardiomyocytes.

BACKGROUND: Although human-induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) are a promising cell resource for cardiovascular research, these cells exhibit an immature phenotype that hampers their potential applications. The inwardly rectifying potassium channel Kir2.1, encoded by the KCNJ2 gene, has been thought as an important target for promoting electrical maturation of iPSC-CMs. However, a comprehensive characterization of morphological and functional changes in iPSC-CMs overexpressing KCNJ2 (KCNJ2 OE) is still lacking. METHODS: iPSC-CMs were generated using a 2D in vitro monolayer differentiation protocol. Human KCNJ2 construct with green fluorescent protein (GFP) tag was created and overexpressed in iPSC-CMs via lentiviral transduction. The mixture of iPSC-CMs and mesenchymal cells was cocultured with decellularized natural heart matrix for generation of 3D human engineered heart tissues (EHTs). RESULTS: We showed that mRNA expression level of KCNJ2 in iPSC-CMs was dramatically lower than that in human left ventricular tissues. KCNJ2 OE iPSC-CMs yielded significantly increased protein expression of Kir2.1 and current density of Kir2.1-encoded IK1. The larger IK1 linked to a quiescent phenotype that required pacing to elicit action potentials in KCNJ2 OE iPSC-CMs, which can be reversed by IK1 blocker BaCl2. KCNJ2 OE also led to significantly hyperpolarized maximal diastolic potential (MDP), shortened action potential duration (APD) and increased maximal upstroke velocity. The enhanced electrophysiological maturation in KCNJ2 OE iPSC-CMs was accompanied by improvements in Ca2+ signaling, mitochondrial energy metabolism and transcriptomic profile. Notably, KCNJ2 OE iPSC-CMs exhibited enlarged cell size and more elongated and stretched shape, indicating a morphological phenotype toward structural maturation. Drug testing using hERG blocker E-4031 revealed that a more stable MDP in KCNJ2 OE iPSC-CMs allowed for obtaining significant drug response of APD prolongation in a concentration-dependent manner. Moreover, KCNJ2 OE iPSC-CMs formed more mature human EHTs with better tissue structure and cell junction. CONCLUSIONS: Overexpression of KCNJ2 can robustly enhance maturation of iPSC-CMs in electrophysiology, Ca2+ signaling, metabolism, transcriptomic profile, cardiomyocyte structure and tissue engineering, thus providing more accurate cellular model for elucidating cellular and molecular mechanisms of cardiovascular diseases, screening drug-induced cardiotoxicity, and developing personalized and precision cardiovascular medicine.

Chick early amniotic fluid component improves heart function and protects against inflammation after myocardial infarction in mice.

Myocardial infarction (MI) is the major cause of mortality around the world. We recently demonstrated that chick early amniotic fluid (ceAF) can effectively rescue ischemic heart injury, indicating that it has a therapeutic function in MI. However, its functional components and the underlying mechanisms remain to be clarified. Here, we demonstrated that a fraction of ceAF, peak 8 (P8), had a protective effect on acute MI. P8 significantly decreased cardiomyocyte cross-sectional areas and cardiomyocyte apoptosis in MI mice. Using a human embryonic stem cell-derived cardiomyocyte model, which was subjected to hypoxia and reoxygenation, mimicking MI state, we found that P8 treatment reduced apoptosis and reversed myocardial contractility. Mechanistically, P8 improved cardiac function by inhibiting NF-κB signaling and downregulating inflammatory cytokine expression. Using mass spectrometry, we identified that guanosine and deoxynucleoside were the main functional components of P8 that suppressed the inflammatory response in human embryonic stem cell-derived cardiomyocytes. Collectively, our data suggest that specific components from ceAF are promising therapeutic agents for ischemic heart injury and could be a potential supplement to current medications for MI.

ß-hydroxybutyrate inhibits ferroptosis-mediated pancreatic damage in acute liver failure through the increase of H3K9bhb.

Acute pancreatitis and hyperamylasemia are often seen in patients with acute liver failure (ALF). However, the underlying mechanisms remain elusive. This study describes pancreatic tissue damage and exocrine dysfunction in a mouse model of major-liver-resection-induced ALF. The analysis of 1,264 clinical cases of liver failure (LF) showed that the incidence of hyperamylasemia and hyperlipasemia in patients with LF is 5.5% and 20%, respectively. Metabolomic studies indicate that glutathione (GSH)-deficiency-caused ferroptosis contributes to pancreatic damage in mouse ALF. ß-hydroxybutyrate (ß-HB) is the only metabolite downregulated in the liver, serum, and pancreas. Our data suggest that ß-HB protects pancreatic cells and tissues from GSH-deficiency-caused ferroptosis. ß-HB administration in ALF mice restores the expression of ferroptosis-suppressor genes through histone H3 lysine 9 ß-hydroxybutyrylation (H3K9bhb)-mediated chromatin opening. Our findings highlight ß-HB as an endogenous metabolite regulating ferroptosis in the pancreas and extend our understanding of the pathophysiology of ALF-induced pancreatitis.

The regulation of skin homeostasis, repair and the pathogenesis of skin diseases by spatiotemporal activation of epidermal mTOR signaling.

The epidermis, the outmost layer of the skin, is a stratified squamous epithelium that protects the body from the external world. The epidermis and its appendages need constantly renew themselves and replace the damaged tissues caused by environmental assaults. The mechanistic target of rapamycin (mTOR) signaling is a central controller of cell growth and metabolism that plays a critical role in development, homeostasis and diseases. Recent findings suggest that mTOR signaling is activated in a spatiotemporal and context-dependent manner in the epidermis, coordinating diverse skin homeostatic processes. Dysregulation of mTOR signaling underlies the pathogenesis of skin diseases, including psoriasis and skin cancer. In this review, we discuss the role of epidermal mTOR signaling activity and function in skin, with a focus on skin barrier formation, hair regeneration, wound repair, as well as skin pathological disorders. We propose that fine-tuned control of mTOR signaling is essential for epidermal structural and functional integrity.

Therapeutic application of chick early amniotic fluid: effective rescue of acute myocardial ischemic injury by intravenous administration.

Myocardial regeneration has been considered a promising option for the treatment of adult myocardial injuries. Previously, a chick early amniotic fluid (ceAF) preparation was shown to contain growth-related factors that promoted embryonic growth and cellular proliferation, though the nature of the components within ceAF were not fully defined. Here we tested whether this ceAF preparation is similarly effective in the promotion of myocardial regeneration, which could provide an alternative therapeutic for intervening myocardial injury. In this study, a myocardial ischemic injury model was established in adult mice and pigs by multiple research entities, and we were able to show that ceAF can efficiently rescue damaged cardiac tissues and markedly improve cardiac function in both experimental models through intravenous administration. ceAF administration increased cell proliferation and improved angiogenesis, likely via down-regulation of Hippo-YAP signaling. Our data suggest that ceAF administration can effectively rescue ischemic heart injury, providing the key functional information for the further development of ceAF for use in attenuating myocardial injury.

Transcriptomics- and metabolomics-based integration analyses revealed the potential pharmacological effects and functional pattern of in vivo Radix Paeoniae Alba administration.

BACKGROUND: Radix Paeoniae Alba (RPA) and other natural medicines have remarkable curative effects and are widely used in traditional Chinese Medicine (TCM). However, due to their multi-component and multi-target characteristics, it is difficult to study the detailed pharmacological mechanisms for those natural medicines in vivo. Therefore, their real effects on organisms is still uncertain. METHODS: RPA was selected as research object, the present study was designed to study the complex mechanisms of RPA in vivo by integrating and interpreting the transcriptomic based RNA-seq and metabolomic based NMR spectrum after RPA administration in mice. A variety of dimension-reduction algorithms and classifier models were applied to the processing of high-throughput data. RESULTS: Among serum metabolites, the contents of PC and glucose were significantly increased, while the contents of various amino acids, lipids and their metabolites were significantly decreased in mice after RPA administration. Based on the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases, differential analysis showed that the liver was the site where RPA exerted a significant effect, which confirmed the rationality of "meridian tropism" in the theory in TCM. In addition, RPA played a role in lipid metabolism by regulating genes encoding enzymes of the glycerolipid metabolism pathway, such as 1-acyl-sn-glycerol-3-phosphate acyltransferase (Agpat), phosphatidate phosphatase (Lpin), phospholipid phosphatase (Plpp) and endothelial lipase (Lipg). We also found that RPA regulates several substance addiction pathways in the brain, such as the cocaine addiction pathway, and the related targets were predicted based on the sequencing data from pathological model in the GEO database. The overall effective pattern of RPA was intuitively presented with a multidimensional radar map through a self-designed model which found that liver and brain were mainly regulated by RPA compared with the traditional meridian tropism theory. CONCLUSIONS: Overall this study expanded the potential application of RPA and provided possible targets and directions for further mechanism study, meanwhile, it also established a multi-dimensional evaluation model to represent the overall effective pattern of TCM for the first time. In the future, such study based on the high-throughput data sets can be used to interpret the theory of TCM and to provide a valuable research model and clinical medication reference for the TCM researchers and doctors.

Potential Crosstalk between Liver and Extra-liver Organs in Mouse Models of Acute Liver Injury.

Carbon tetrachloride (CCl4), Concanavalin A (ConA), bile duct ligation (BDL), and liver resection (LR) are four types of commonly used mouse models of acute liver injury. However, these four models belong to different types of liver cell damage while their application situations are often confounded. In addition, the systematic changes of multiple extra-liver organs after acute liver injury and the crosstalk between liver and extra-liver organs remain unclear. Here, we aim to map the morphological, metabolomic and transcriptomic changes systematically after acute liver injury and search for the potential crosstalk between the liver and the extra-liver organs. Significant changes of transcriptome were observed in multiple extra-liver organs after different types of acute liver injury despite dramatic morphological damage only occurred in lung tissues of the ConA/BDL models and spleen tissues in the ConA model. Liver transcriptomic changes initiated the serum metabolomic alterations which correlated to transcriptomic variation in lung, kidney, and brain tissues of BDL and LR models. The potential crosstalk might lead to pulmonary damage and development of hepatorenal syndrome (HRS) and hepatic encephalopathy (HE) during liver injury. Serum derived from acute liver injury mice damaged alveolar epithelial cells and human podocytes in vitro. Our data indicated that different types of acute liver injury led to different transcriptomic changes within extra-liver organs. Integration of serum metabolomics and transcriptomics from multiple tissues can improve our understanding of acute liver injury and its effect on the other organs.

Repair of Adult Mammalian Heart After Damages by Oral Intake of Gu Ben Pei Yuan San.

Adult mammalian heart repair after myocardial damage is highly inefficient due to the post-mitotic nature of cardiomyocytes. Interestingly, in traditional Chinese medicine (TCM), there are reported effective treatments of myocardial infarction (MI) and heart failure in adult humans by oral intake of a TCM concoction named Gu Ben Pei Yuan San (GBPYS), which is composed of Panax ginseng, velvet antler, Gekko gecko Linnaeus tail, human placenta, Trogopterus dung, Panax notoginseng, and amber. We fed mice with GBPYS after myocardial damages through everyday self-feeding. We then examined the effect of everyday oral intake of GBPYS on improving cardiac function and myocardial repair in adult mice after apical resection or MI. We found that long-term oral intake of GBPYS significantly improved cardiac function after myocardial damages in adult mice. BrdU, phospho-histone 3, and AuroraB staining indicated increased cell proliferation at the border zone of MI after TCM feeding. GBPYS feeding reduced organ inflammation, induced angiogenesis, and is non-toxic to mice after long-term oral intake. Further, serum derived from TCM-fed MI rats promoted division of both neonatal rat cardiomyocytes and human induced pluripotent stem cell (iPSC)-derived cardiomyocytes in vitro. Oral intake of GBPYS improved heart repair after myocardial damages in adult mice. Our results suggest that there are substances present in GBPYS that help improve adult mammalian heart repair after MI. Also, it could be a good choice of non-invasive alternative therapy for myocardial damages and heart failure after rigorous clinical study in the future.

Heart Regeneration in Adult Mammals after Myocardial Damage.

Heart regeneration remains a critical question in current basic research and clinical practice. The adult mammalian heart exhibits a very limited regeneration capacity. In contrast, adult zebrafish and neonatal mice retain a remarkable ability of heart regeneration after damage. Understanding the mechanisms of heart regeneration would be very valuable to help design efficient treatment strategies against myocardial damage and heart failure. While inherent regeneration of the heart occurs after damage with varying efficiency among species, regeneration may also be induced exogenously. In this study, we briefly review the different approaches and current progress in improving heart regeneration.

Diarylethene based fluorescent switchable probes for the detection of amyloid-ß pathology in Alzheimer's disease.

Two fluorescent switchable diarylethene derivatives which exhibit high affinity for amyloid-ß aggregates with the increase of fluorescence intensity were reported. Moreover, the probes show excellent photochromic and anti-photobleaching properties both in vitro and in vivo.

Electroacupuncture improves cardiac function and remodeling by inhibition of sympathoexcitation in chronic heart failure rats.

Chronic heart failure (CHF) is responsible for significant morbidity and mortality worldwide, mainly as a result of neurohumoral activation. Acupuncture has been used to treat a wide range of diseases and conditions. In this study, we investigated the effects of electroacupuncture (EA) on the sympathetic nerve activity, heart function, and remodeling in CHF rats after ligation of the left anterior descending coronary artery. CHF rats were randomly selected to EA and control groups for acute and chronic experiments. In the acute experiment, both the renal sympathetic nerve activity and cardiac sympathetic afferent reflex elicited by epicardial application of capsaicin were recorded. In the chronic experiment, we performed EA for 30 min once a day for 1 wk to test the long-term EA effects on heart function, remodeling, as well as infarct size in CHF rats. The results show EA significantly decreased the renal sympathetic nerve activity effectively, inhibited cardiac sympathetic afferent reflex, and lowered the blood pressure of CHF rats. Treating CHF rats with EA for 1 wk dramatically increased left ventricular ejection fraction and left ventricular fraction shortening, reversed the enlargement of left ventricular end-systolic dimension and left ventricular end-diastolic dimension, and shrunk the infarct size. In this experiment, we demonstrated EA attenuates sympathetic overactivity. Additionally, long-term EA improves cardiac function and remodeling and reduces infarct size in CHF rats. EA is a novel and potentially useful therapy for treating CHF.

Angiotensin-(1-7) in the rostral ventrolateral medulla modulates enhanced cardiac sympathetic afferent reflex and sympathetic activation in renovascular hypertensive rats.

Enhancement of the cardiac sympathetic afferent reflex (CSAR) contributes to sympathetic excitation in hypertension. The aim of the present study was to determine whether angiotensin (Ang)-(1-7) in the rostral ventrolateral medulla (RVLM) modulated the enhanced CSAR and sympathetic activation, and the signaling pathways that mediated these effects in the 2-kidney, 1-clip renovascular hypertension model. Cardiac sympathetic afferent reflex was evaluated using renal sympathetic nerve activity and mean arterial pressure responses to epicardial capsaicin application in anesthetized sinoaortic-denervated and cervical-vagotomized rats. RVLM microinjection of Ang-(1-7) induced greater increases in renal sympathetic nerve activity and mean arterial pressure, and greater enhancement in CSAR in 2-kidney, 1-clip rats than in sham-operated rats, which was blocked by Mas receptor antagonist A-779, adenylyl cyclase inhibitors SQ22536 and MDL-12,330A, and protein kinase A inhibitors rp-adenosine-3',5'-cyclic monophosphorothionate and H-89. Mas receptor expression in RVLM was increased in 2-kidney, 1-clip rats. Treatment with A-779, SQ22536, MDL-12,330A, rp-adenosine-3',5'-cyclic monophosphorothionate, or H-89 in RVLM inhibited CSAR and decreased renal sympathetic nerve activity and mean arterial pressure in 2-kidney, 1-clip rats, whereas cAMP analogue dibutyryl-cAMP had the opposite effects. Ang-(1-7) in RVLM increased, whereas A-779 decreased the cAMP level and the epicardial capsaicin application-induced increases in the cAMP level in RVLM. These results indicate that Ang-(1-7) in the RVLM enhances the CSAR and increases the sympathetic outflow and blood pressure via Mas receptor activation. The increased endogenous Ang-(1-7) and Mas receptor activity in RVLM contributes to the enhanced CSAR and sympathetic activation in renovascular hypertension, and the cAMP-protein kinase A pathway is involved in these Ang-(1-7)-mediated effects in the RVLM.

Melanocortin 3/4 receptors in paraventricular nucleus modulate sympathetic outflow and blood pressure.

Central melanocortin 3/4 receptors (MC3/4Rs) are known to regulate energy balance. Activation of MC3/4Rs causes a greater increase in the firing activity of the PVN neurons in obese Zucker rats than in lean Zucker rats. The present study was undertaken to determine the roles of MC3/4Rs in the hypothalamic paraventricular nucleus (PVN) in modulating the sympathetic activity and blood pressure and its downstream pathway. Renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were recorded in anaesthetized rats. Microinjection of the MC3/4R agonist melanotan II (MTII) into the PVN increased the RSNA and MAP. The MC3/4R antagonist agouti-related peptide (AgRP) or SHU9119 decreased the RSNA and MAP, but the MC4R antagonist HS024 had no significant effect on the RSNA and MAP. The effects of MTII were abolished by pretreatment of the PVN with AgRP, SHU9119, the adenylate cyclase inhibitor SQ22536 or the protein kinase A inhibitor Rp-cAMP, and substantially attenuated by HS024. Microinjection of SQ22536 alone into the PVN had no significant effect on the RSNA and MAP, but Rp-cAMP caused significant decreases in the RSNA and MAP. Furthermore, MTII increased the cAMP level in the PVN. These results indicate that activation of MC3/4Rs in the PVN increases the sympathetic outflow and blood pressure via the cAMP-protein kinase A pathway. Melanocortin 3 receptors in the PVN may exert a tonic excitatory effect on sympathetic activity.

Inhibition of cardiac sympathetic afferent reflex and sympathetic activity by baroreceptor and vagal afferent inputs in chronic heart failure.

BACKGROUND: Cardiac sympathetic afferent reflex (CSAR) contributes to sympathetic activation and angiotensin II (Ang II) in paraventricular nucleus (PVN) augments the CSAR in vagotomized (VT) and baroreceptor denervated (BD) rats with chronic heart failure (CHF). This study was designed to determine whether it is true in intact (INT) rats with CHF and to determine the effects of cardiac and baroreceptor afferents on the CSAR and sympathetic activity in CHF. METHODOLOGY/PRINCIPAL FINDINGS: Sham-operated (Sham) or coronary ligation-induced CHF rats were respectively subjected to BD+VT, VT, cardiac sympathetic denervation (CSD) or INT. Under anesthesia, renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were recorded, and the CSAR was evaluated by the RSNA and MAP responses to epicardial application of capsaicin. Either CSAR or the responses of RSNA, MAP and CSAR to Ang II in PVN were enhanced in CHF rats treated with BD+VT, VT or INT. Treatment with VT or BD+VT potentiated the CSAR and the CSAR responses to Ang II in both Sham and CHF rats. Treatment with CSD reversed the capsaicin-induced RSNA and MAP changes and the CSAR responses to Ang II in both Sham and CHF rats, and reduced the RSNA and MAP responses to Ang II only in CHF rats. CONCLUSIONS: The CSAR and the CSAR responses to Ang II in PVN are enhanced in intact CHF rats. Baroreceptor and vagal afferent activities inhibit CSAR and the CSAR responses to Ang II in intact Sham and CHF rats.