Language：English   Publishing type：Research paper (scientific journal)  

The biological significance of deadenylation in global gene expression is not fully understood. Here, we show that the CCR4-NOT deadenylase complex maintains expression of mRNAs, such as those encoding transcription factors, cell cycle regulators, DNA damage response-related proteins, and metabolic enzymes, at appropriate levels in the liver. Liver-specific disruption of Cnot1, encoding a scaffold subunit of the CCR4-NOT complex, leads to increased levels of mRNAs for transcription factors, cell cycle regulators, and DNA damage response-related proteins because of reduced deadenylation and stabilization of these mRNAs. CNOT1 suppression also results in an increase of immature, unspliced mRNAs (pre-mRNAs) for apoptosis-related and inflammation-related genes and promotes RNA polymerase II loading on their promoter regions. In contrast, mRNAs encoding metabolic enzymes become less abundant, concomitant with decreased levels of these pre-mRNAs. Lethal hepatitis develops concomitantly with abnormal mRNA expression. Mechanistically, the CCR4-NOT complex targets and destabilizes mRNAs mainly through its association with Argonaute 2 (AGO2) and butyrate response factor 1 (BRF1) in the liver. Therefore, the CCR4-NOT complex contributes to liver homeostasis by modulating the liver transcriptome through mRNA deadenylation.

DOI： 10.26508/lsa.201900494

Language：English  

Angiotensin-converting enzyme 2 (ACE2) is critically involved in cardiovascular physiology and pathology, and is currently clinically evaluated to treat acute lung failure. Here we show that the B38-CAP, a carboxypeptidase derived from Paenibacillus sp. B38, is an ACE2-like enzyme to decrease angiotensin II levels in mice. In protein 3D structure analysis, B38-CAP homolog shares structural similarity to mammalian ACE2 with low sequence identity. In vitro, recombinant B38-CAP protein catalyzed the conversion of angiotensin II to angiotensin 1-7, as well as other known ACE2 target peptides. Treatment with B38-CAP suppressed angiotensin II-induced hypertension, cardiac hypertrophy, and fibrosis in mice. Moreover, B38-CAP inhibited pressure overload-induced pathological hypertrophy, myocardial fibrosis, and cardiac dysfunction in mice. Our data identify the bacterial B38-CAP as an ACE2-like carboxypeptidase, indicating that evolution has shaped a bacterial carboxypeptidase to a human ACE2-like enzyme. Bacterial engineering could be utilized to design improved protein drugs for hypertension and heart failure.

DOI： 10.1038/s41467-020-14867-z

Language：English   Publishing type：Research paper (scientific journal)  

Genome-Scale CRISPR/Cas9 Screening Reveals Squalene Epoxidase as a Susceptibility Factor for Cytotoxicity of Malformin A1.
Malformin A1 (MA1) is a fungus-produced cyclic pentapeptide. MA1 exhibits teratogenicity to plants, fibrinolysis-enhancing activity, and cytotoxicity to mammalian cells. To clarify the cytotoxic mechanism of MA1, we screened for the genes involved in the cytotoxicity of MA1 in monocytoid U937 cells by using a CRISPR/Cas9-based genome-wide knockout library. Screening was performed by positive selection for cells that were resistant to MA1 treatment, and single guide RNAs (sgRNAs) integrated into MA1-resistant cells were analyzed by high-throughput sequencing. As a result of the evaluation of sgRNAs that were enriched in MA1-resistant cells, SQLE, which encodes squalene epoxidase, was identified as a candidate gene. SQLE-depleted U937 cells were viable in the presence of MA1, and squalene epoxidase inhibitor conferred MA1 resistance to wild-type cells. These results indicate that squalene epoxidase is implicated in the cytotoxicity of MA1. This finding represents a new insight into applications of MA1 for treating ischemic diseases.

DOI： 10.1002/cbic.201800769

Language：Others   Publishing type：Research paper (scientific journal)  

Pulmonary phagocyte-derived NPY controls the pathology of severe influenza virus infection.
© 2018, The Author(s), under exclusive licence to Springer Nature Limited. Crosstalk between the autonomic nervous system and the immune system by means of the sympathetic and parasympathetic pathways is a critical process in host defence. Activation of the sympathetic nervous system results in the release of catecholamines as well as neuropeptide Y (NPY). Here, we investigated whether phagocytes are capable of the de novo production of NPY, as has been described for catecholamines. We show that the synthesis of NPY and its Y1 receptor (Y1R) is increased in phagocytes in lungs following severe influenza virus infection. The genetic deletion of Npy or Y1r specifically in phagocytes greatly improves the pathology of severe influenza virus infection, which is characterized by excessive virus replication and pulmonary inflammation. Mechanistically, it is the induction of suppressor of cytokine signalling 3 (SOCS3) via NPY–Y1R activation that is responsible for impaired antiviral response and promoting pro-inflammatory cytokine production, thereby enhancing the pathology of influenza virus infection. Thus, direct regulation of the NPY–Y1R–SOCS3 pathway on phagocytes may act as a fine-tuner of an innate immune response to virus infection, which could be a therapeutic target for lethal influenza virus infection.

DOI： 10.1038/s41564-018-0289-1

Language：English   Publishing type：Research paper (scientific journal)  

Loss of Apelin Augments Angiotensin II-Induced Cardiac Dysfunction and Pathological Remodeling.
Apelin is an inotropic and cardioprotective peptide that exhibits beneficial effects through activation of the APJ receptor in the pathology of cardiovascular diseases. Apelin induces the expression of angiotensin-converting enzyme 2 (ACE2) in failing hearts, thereby improving heart function in an angiotensin 1⁻7-dependent manner. Whether apelin antagonizes the over-activation of the renin⁻angiotensin system in the heart remains elusive. In this study we show that the detrimental effects of angiotensin II (Ang II) were exacerbated in the hearts of aged apelin-gene-deficient mice. Ang II-mediated cardiac dysfunction and hypertrophy were augmented in apelin knockout mice. The loss of apelin increased the ratio of angiotensin-converting enzyme (ACE) to ACE2 expression in the Ang II-stressed hearts, and Ang II-induced cardiac fibrosis was markedly enhanced in apelin knockout mice. mRNA expression of pro-fibrotic genes, such as transforming growth-factor beta (TGF-β) signaling, were significantly upregulated in apelin knockout hearts. Consistently, treatment with the ACE-inhibitor Captopril decreased cardiac contractility in apelin knockout mice. In vitro, apelin ameliorated Ang II-induced TGF-β expression in primary cardiomyocytes, accompanied with reduced hypertrophy. These results provide direct evidence that endogenous apelin plays a crucial role in suppressing Ang II-induced cardiac dysfunction and pathological remodeling.

DOI： 10.3390/ijms20020239

Language：English   Publishing type：Research paper (scientific journal)  

Shortening and removal of the polyadenylate [poly(A)] tail of mRNA, a process called deadenylation, is a key step in mRNA decay that is mediated through the CCR4-NOT (carbon catabolite repression 4-negative on TATA-less) complex. In our investigation of the regulation of mRNA deadenylation in the heart, we found that this complex was required to prevent cell death. Conditional deletion of the CCR4-NOT complex components Cnot1 or Cnot3 resulted in the formation of autophagic vacuoles and cardiomyocyte death, leading to lethal heart failure accompanied by long QT intervals. Cnot3 bound to and shortened the poly(A) tail of the mRNA encoding the key autophagy regulator Atg7. In Cnot3-depleted hearts, Atg7 expression was posttranscriptionally increased. Genetic ablation of Atg7, but not Atg5, increased survival and partially restored cardiac function of Cnot1 or Cnot3 knockout mice. We further showed that in Cnot3-depleted hearts, Atg7 interacted with p53 and modulated p53 activity to induce the expression of genes encoding cell death-promoting factors in cardiomyocytes, indicating that defects in deadenylation in the heart aberrantly activated Atg7 and p53 to promote cell death. Thus, mRNA deadenylation mediated by the CCR4-NOT complex is crucial to prevent Atg7-induced cell death and heart failure, suggesting a role for mRNA deadenylation in targeting autophagy genes to maintain normal cardiac homeostasis.

DOI： 10.1126/scisignal.aan3638

Language：English   Publishing type：Research paper (scientific journal)  

Apela (also known as Elabela, Ende, and Toddler) is a small signaling peptide that activates the G-proteincoupled receptor Aplnr to stimulate cell migration during zebrafish gastrulation. Here, using CRISPR/Cas9 to generate a null, reporter-expressing allele, we study the role of Apela in the developing mouse embryo. We found that loss of Apela results in low-penetrance cardiovascular defects that manifest after the onset of circulation. Three-dimensional micro- computed tomography revealed a higher penetrance of vascular remodeling defects, from which some mutants recover, and identified extraembryonic anomalies as the earliest morphological distinction in Apela mutant embryos. Transcriptomics at late gastrulation identified aberrant upregulation of erythroid and myeloid markers in mutant embryos prior to the appearance of physical malformations. Double-mutant analyses showed that loss of Apela signaling impacts early Aplnr-expressing mesodermal populations independently of the alternative ligand Apelin, leading to lethal cardiac defects in some Apela null embryos.

DOI： 10.1016/j.celrep.2017.08.014

Language：English   Publishing type：Research paper (scientific journal)  

Aims Elabela/Toddler/Apela (ELA) has been identified as a novel endogenous peptide ligand for APJ/Apelin receptor/ Aplnr. ELA plays a crucial role in early cardiac development of zebrafish as well as in maintenance of self-renewal of human embryonic stem cells. Apelin was the first identified APJ ligand, and exerts positive inotropic heart effects and regulates the renin-angiotensin system. The aim of this study was to investigate the biological effects of ELA in the cardiovascular system.
Methods and results Continuous infusion of ELA peptide significantly suppressed pressure overload-induced cardiac hypertrophy, fibrosis and impaired contractility in mice. ELA treatment reduced mRNA expression levels of genes associated with heart failure and fibrosis. The cardioprotective effects of ELA were diminished in APJ knockout mice, indicating that APJ is the key receptor for ELA in the adult heart. Mechanistically, ELA downregulated angiotensin-converting enzyme (ACE) expression in the stressed hearts, whereas it showed little effects on angiotensin-converting enzyme 2 (ACE2) expression, which are distinct from the effects of Apelin. FoxM1 transcription factor, which induces ACE expression in the stressed hearts, was downregulated by ELA but not by Apelin. ELA antagonized angiotensin IIinduced hypertension, cardiac hypertrophy, and fibrosis in mice.
Conclusion The ELA-APJ axis protects from pressure overload-induced heart failure possibly via suppression of ACE expression and pathogenic angiotensin II signalling. The different effects of ELA and Apelin on the expression of ACE and ACE2 implicate fine-tuned mechanisms for a ligand-induced APJ activation and downstream signalling.

DOI： 10.1093/cvr/cvx061

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Transcription factor EBF1 (early B-cell factor 1) regulates early B-cell differentiation by poising or activating lineage-specific genes and repressing genes associated with alternative cell fates. To identify proteins that regulate the diverse functions of EBF1, we used SILAC (stable isotope labeling by amino acids in cell culture)-based mass spectrometry of proteins associated with endogenous EBF1 in pro-B cells. This analysis identified most components of the multifunctional CCR4-NOT complex, which regulates transcription and mRNA degradation. CNOT3 interacts with EBF1, and we identified histidine 240 in EBF1 as a critical residue for this interaction. Complementation of Ebf1(-/-)progenitors with EBF1H240A revealed a partial block of pro-B-cell differentiation and altered expression of specific EBF1 target genes that show either reduced transcription or increased mRNA stability. Most deregulated EBF1 target genes show normal occupancy by EBF1H240A, but we also detected genes with altered occupancy, suggesting that the CCR4-NOT complex affects multiple activities of EBF1. Mice with conditional Cnot3 inactivation recapitulate the block of early B-cell differentiation, which we found to be associated with an impaired autoregulation of Ebf1 and reduced expression of pre-B-cell receptor components. Thus, the interaction of the CCR4-NOT complex with EBF1 diversifies the function of EBF1 in a context-dependent manner and may coordinate transcriptional and post-transcriptional gene regulation.

DOI： 10.1101/gad.285452.116

Language：English   Publishing type：Research paper (scientific journal)  

Angiotensin converting enzyme 2 (ACE2) is a negative regulator of the renin-angiotensin system (RAS), catalyzing the conversion of Angiotensin II to Angiotensin 1-7. Apelin is a second catalytic substrate for ACE2 and functions as an inotropic and cardioprotective peptide. While an antagonistic relationship between the RAS and apelin has been proposed, such functional interplay remains elusive. Here we found that ACE2 was d.ownregulated in apelin-deficient mice. Pharmacological or genetic inhibition of angiotensin II type 1 receptor (AT1R) rescued the impaired contractility and hypertrophy of apelin mutant mice, which was accompanied by restored ACE2 levels. Importantly, treatment with angiotensin 1-7 rescued hypertrophy and heart dysfunctions of apelin-knockout mice. Moreover, apelin, via activation of its receptor, APJ, increased ACE2 promoter activity in vitro and upregulated ACE2 expression in failing hearts in vivo. Apelin treatment also increased cardiac contractility and ACE2 levels in AT1R-deficient mice. These data demonstrate that ACE2 couples the RAS to the apelin system, adding a conceptual framework for the apelin-ACE2-angiotensin 1-7 axis as a therapeutic target for cardiovascular diseases.

DOI： 10.1172/JCI69608

Language：English   Publishing type：Research paper (scientific journal)  

Influenza A viruses are a major cause of mortality. Given the potential for future lethal pandemics, effective drugs are needed for the treatment of severe influenza such as that caused by H5N1 viruses. Using mediator lipidomics and bioactive lipid screen, we report that the omega-3 polyunsaturated fatty acid (PUFA)-derived lipid mediator protectin D1 (PD1) markedly attenuated influenza virus replication via RNA export machinery. Production of PD1 was suppressed during severe influenza and PD1 levels inversely correlated with the pathogenicity of H5N1 viruses. Suppression of PD1 was genetically mapped to 12/15-lipoxygenase activity. Importantly, PD1 treatment improved the survival and pathology of severe influenza in mice, even under conditions where known antiviral drugs fail to protect from death. These results identify the endogenous lipid mediator PD1 as an innate suppressor of influenza virus replication that protects against lethal influenza virus infection. © 2013 Elsevier Inc.

DOI： 10.1016/j.cell.2013.02.027

Language：English   Publishing type：Research paper (scientific journal)  

Rationale: Patients who developed acute respiratory distress syndrome (ARDS) after infection with severe respiratory viruses (e.g., severe acute respiratory syndrome coronavirus, H5N1 avian influenza virus), exhibited unusually high levels of CXCL10, which belongs to the non-ELR (glutamic-leucine-arginine) CXC chemokine superfamily. CXCL10 may not be a bystander to the severe virus infection but may directly contribute to the pathogenesis of neutrophil-mediated, excessive pulmonary inflammation.
Objectives: We investigated the contribution of CXCL10 and its receptor CXCR3 axis to the pathogenesis of ARDS with nonviral and viral origins.
Methods: We induced nonviral ARDS by acid aspiration and viral ARDS by intratracheal influenza virus infection in wild-type mice and mice deficient in CXCL10, CXCR3, IFNAR1 (IFN-alpha/beta receptor 1), or TIR domain-containing adaptor inducing IFN-beta (TRIF).
Measurements and Main Results: We found that the mice lacking CXCL10 or CXCR3 demonstrated improved severity and survival of nonviral and viral ARDS, whereas mice that lack IFNAR1 did not control the severity of ARDS in vivo. The increased levels of CXCL10 in lungs with ARDS originate to a large extent from infiltrated pulmonary neutrophils, which express a unique CXCR3 receptor via TRIF. CXCL10-CXCR3 acts in an autocrine fashion on the oxidative burst and chemotaxis in the inflamed neutrophils,leading to fulminant pulmonary inflammation.
Conclusions: CXCL10-CXCR3 signaling appears to be a critical factor for the exacerbation of the pathology of ARDS. Thus, the CXCL10-CXCR3 axis could represent a prime therapeutic target in the treatment of the acute phase of ARDS of nonviral and viral origins.

DOI： 10.1164/rccm.201203-0508OC

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CXCL10-CXCR3 enhances the development of neutrophil-mediated fulminant lung injury of viral and nonviral origin.
Patients who developed acute respiratory distress syndrome (ARDS) after infection with severe respiratory viruses (e.g., severe acute respiratory syndrome-coronavirus, H5N1 avian influenza virus), exhibited unusually high levels of CXCL10, which belongs to the non-ELR (glutamic-leucine-arginine) CXC chemokine superfamily. CXCL10 may not be a bystander to the severe virus infection but may directly contribute to the pathogenesis of neutrophil-mediated, excessive pulmonary inflammation.<br />
We investigated the contribution of CXCL10 and its receptor CXCR3 axis to the pathogenesis of ARDS with nonviral and viral origins.<br />
We induced nonviral ARDS by acid aspiration and viral ARDS by intratracheal influenza virus infection in wild-type mice and mice deficient in CXCL10, CXCR3, IFNAR1 (IFN-α/β receptor 1), or TIR domain-containing adaptor inducing IFN-β (TRIF).<br />
We found that the mice lacking CXCL10 or CXCR3 demonstrated improved severity and survival of nonviral and viral ARDS, whereas mice that lack IFNAR1 did not control the severity of ARDS in vivo. The increased levels of CXCL10 in lungs with ARDS originate to a large extent from infiltrated pulmonary neutrophils, which express a uniq

DOI： 10.1164/rccm.201203-0508OC

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Heart diseases are the most common causes of morbidity and death in humans. Using cardiac-specific RNAi-silencing in Drosophila, we knocked down 7061 evolutionarily conserved genes under conditions of stress. We present a first global road-map of pathways potentially playing conserved roles in the cardiovascular system. One critical pathway identified was the CCR4-Not complex implicated in transcriptional and posttranscriptional regulatory mechanisms. Silencing of CCR4-Not components in adult Drosophila resulted in myofibrillar disarray and dilated cardiomyopathy. Heterozygous not3 knockout mice showed spontaneous impairment of cardiac contractility and increased susceptibility to heart failure. These heart defects were reversed via inhibition of HDACs, suggesting a mechanistic link to epigenetic chromatin remodeling. In humans, we show that a common NOT3 SNP correlates with altered cardiac QT intervals, a known cause of potentially lethal ventricular tachyarrhythmias. Thus, our functional genome-wide screen in Drosophila can identify candidates that directly translate into conserved mammalian genes involved in heart function.

DOI： 10.1016/j.cell.2010.02.023

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Multiple lung pathogens such as chemical agents, H5N1 avian flu, or SARS cause high lethality due to acute respiratory distress syndrome. Here we report that Toll-like receptor 4 (TLR4) mutant mice display natural resistance to acid-induced acute lung injury (ALI). We show that TLR4-TRIF-TRAF6 signaling is a key disease pathway that controls the severity of ALI. The oxidized phospholipid (OxPL) OxPAPC was identified to induce lung injury and cytokine production by lung macrophages via TLR4-TRIF. We observed OxPL production in the lungs of humans and animals infected with SARS, Anthrax, or H5N1. Pulmonary challenge with an inactivated H5N1 avian influenza virus rapidly induces ALI and OxPL formation in mice. Loss of TLR4 or TRIF expression protects mice from H5N1-induced ALI. Moreover, deletion of ncf1, which controls ROS production, improves the severity of H5N1-mediated ALI. Our data identify oxidative stress and innate immunity as key lung injury pathways that control the severity of ALI.

DOI： 10.1016/j.cell.2008.02.043

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Apelin constitutes a novel endogenous peptide system suggested to be involved in a broad range of physiological functions, including cardiovascular function, heart development, control of fluid homeostasis, and obesity. Apelin is also a catalytic substrate for angiotensin- converting enzyme 2, the key severe acute respiratory syndrome receptor. The in vivo physiological role of Apelin is still elusive. Here we report the generation of Apelin gene -targeted mice. Apelin mutant mice are viable and fertile, appear healthy, and exhibit normal body weight, water and food intake, heart rates, and heart morphology. Intriguingly, aged Apelin knockout mice developed progressive impairment of cardiac contractility associated with systolic dysfunction in the absence of histological abnormalities. We also report that pressure overload induces upregulation of Apelin expression in the heart. Importantly, in pressure overload -induced heart failure, loss of Apelin did not significantly affect the hypertrophy response, but Apelin mutant mice developed progressive heart failure. Global gene expression arrays and hierarchical clustering of differentially expressed genes in hearts of banded Apelin(-/y) and Apelin(-/y) mice showed concerted upregulation of genes involved in extracellular matrix remodeling and muscle contraction. These genetic data show that the endogenous peptide Apelin is crucial to maintain cardiac contractility in pressure overload and aging.

DOI： 10.1161/CIRCRESAHA.107.158659

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During several months of 2003, a newly identified illness termed severe acute respiratory syndrome (SARS) spread rapidly through the world(1-3). A new coronavirus (SARS-CoV) was identified as the SARS pathogen(4-7) which triggered severe pneumonia and acute, often lethal, lung failure(8). Moreover, among infected individuals influenza such as the Spanish flu(9,10) and the emergence of new respiratory disease viruses(11,12) have caused high lethality resulting from acute lung failure(13). In cell lines, angiotensin-converting enzyme 2 (ACE2) has been identified as a potential SARS-CoV receptor(14). The high lethality of SARS-CoV infections, its enormous economic and social impact, fears of renewed outbreaks as well as the potential misuse of such viruses as biologic weapons make it paramount to understand the pathogenesis of SARS-CoV. Here we provide the first genetic proof that ACE2 is a crucial SARS-CoV receptor in vivo. SARS-CoV infections and the Spike protein of the SARS-CoV reduce ACE2 expression. Notably, injection of SARS-CoV Spike into mice worsens acute lung failure in vivo that can be attenuated by blocking the renin-angiotensin pathway. These results provide a molecular explanation why SARS-CoV infections cause severe and often lethal lung failure and suggest a rational therapy for SARS and possibly other respiratory disease viruses.

DOI： 10.1038/nm1267

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Acute respiratory distress syndrome (ARDS), the most severe form of acute lung injury, is a devastating clinical syndrome with a high mortality rate (30-60%) (refs 1-3). Predisposing factors for ARDS are diverse(1,3) and include sepsis, aspiration, pneumonias and infections with the severe acute respiratory syndrome (SARS) corona-virus(4,5). At present, there are no effective drugs for improving the clinical outcome of ARDS(1-3). Angiotensin-converting enzyme (ACE) and ACE2 are homologues with different key functions in the renin-angiotensin system(6-8). ACE cleaves angiotensin I to generate angiotensin II, whereas ACE2 inactivates angiotensin II and is a negative regulator of the system. ACE2 has also recently been identified as a potential SARS virus receptor and is expressed in lungs(9,10). Here we report that ACE2 and the angiotensin II type 2 receptor (AT(2)) protect mice from severe acute lung injury induced by acid aspiration or sepsis. However, other components of the renin-angiotensin system, including ACE, angiotensin II and the angiotensin II type 1a receptor (AT(1)a), promote disease pathogenesis, induce lung oedemas and impair lung function. We show that mice deficient for Ace show markedly improved disease, and also that recombinant ACE2 can protect mice from severe acute lung injury. Our data identify a critical function for ACE2 in acute lung injury, pointing to a possible therapy for a syndrome affecting millions of people worldwide every year.

DOI： 10.1038/nature03712

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Hepatocyte growth factor (HGF) is involved in malignant behavior of cancers as a mediator in tumor-stromal interactions through enhancing tumor invasion and metastasis. We found recently that NK4, a four-kringle fragment of HGF, functions as both an HGF-antagonist and an angiogenesis inhibitor. We have now determined whether blockade of the HGF-c-Met/HGF receptor pathway and tumor angiogenesis by administration of recombinant NK4 would inhibit growth, invasion, and metastasis of human pancreatic carcinoma implanted into the pancreas of nude mice. When treatment with NK4 or anti-HGF neutralizing antibody was initiated from the third day after orthotopic injection of SUIT-2 human pancreatic cancer cells, both NK4 and anti-HGF antibody suppressed the conversion of orthotopic pancreatic tumors from carcinoma in situ to aberrantly invading cancers during days 3-14. On the other hand, when the treatment was begun on day 10, a time when cancer cells were already invading surrounding tissues, NK4 but not anti-HGF antibody inhibited tumor growth, peritoneal dissemination, and ascites accumulation at 4 weeks after the inoculation. Antitumor effects of NK4 correlated with decreased microvessel density in pancreatic tumors thereby indicating that the antlangiogenic activity of NK4 may have mainly contributed to its antitumor effects. Moreover, although NK4-treatment was initiated from the end stage (day 24 after tumor inoculation), NK4 prolonged survival time of mice, and the suppression of peritoneal dissemination, ascites accumulation, and invasion of metastasized cancer cells into the peritoneal wall were remarkable. We propose that simultaneous targeting of both tumor angiogenesis and the HGF-mediated invasion-metastasis may prove to be a new approach to treating patients with pancreatic cancer.

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NK4 composed of the N-terminal hairpin and subsequent four-kringle domains of Hepatocyte growth factor (HGF) is bifunctional, acting as a competitive antagonist for HGF and an angiogenesis inhibitor. In this study, we determined whether or not four-kringle domains of HGF (K1-4) have anti-angiogenic activity. For this purpose, we prepared recombinant K1-4 and NK4, using the baculovirus expression system. Although NK4 antagonized HGF-induced DNA synthesis of rat hepatocytes, cell scattering of MDCK cells and the c-Met/HGF receptor tyrosine phosphorylation in endothelial cells, K1-4 failed to antagonize HGF-induced DNA synthesis, cell scattering and the c-Met/HGF receptor tyrosine phosphorylation in endothelial cells, thus, indicating that K1-4 lacks HGF-antagonist activity. However, endothelial proliferation and migration induced by HGF was inhibited by K1-4, similar to the case seen with NK4. Furthermore, K1-4 inhibited the proliferation and migration of human dermal microvascular endothelial cells induced by vascular endothelial growth factor or by basic fibroblast growth factor. We propose that kringle 1-4 of HGF inhibits angiogenic responses in endothelial cells, independently of HGF-c-Met signaling pathways. © 2000 Academic Press.

DOI： 10.1006/bbrc.2000.4034

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Language：English   Publishing type：Research paper (scientific journal)  

The effect of immune checkpoint inhibitors is extremely limited in patients with pancreatic ductal adenocarcinoma (PDAC) due to the suppressive tumor immune microenvironment (TIME). Autophagy, which has been shown to play a role in anti-tumor immunity, has been proposed as a therapeutic target for PDAC. Here, single-cell RNA-sequencing of autophagy-deficient murine PDAC tumors revealed that autophagy inhibition in cancer cells induced dendritic cell (DC) activation. Analysis of human PDAC tumors substantiated a negative correlation between autophagy and DC activation signatures. Mechanistically, autophagy inhibition increased intracellular accumulation of tumor antigens, which could activate DCs. Administration of chloroquine (CQ), an autophagy inhibitor, in combination with Flt3 ligand (Flt3L)-induced DC infiltration inhibited tumor growth and increased tumor-infiltrating T lymphocytes. However, autophagy inhibition in cancer cells also induced CD8+ T cell exhaustion with high expression of immune checkpoint LAG3. A triple therapy comprising CQ, Flt3L, and an anti-LAG3 antibody markedly reduced tumor growth in orthotopic syngeneic PDAC mouse models. Thus, targeting autophagy in cancer cells and activating DCs sensitizes PDAC tumors to immune checkpoint inhibitor therapy, warranting further development of this treatment approach to overcome immunosuppression in pancreatic cancer.

DOI： 10.1158/0008-5472.CAN-24-0830

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：Nature Structural and Molecular Biology  

Eukaryotic translation initiation factor (eIF)4A-a DEAD-box RNA-binding protein-plays an essential role in translation initiation. Recent reports have suggested helicase-dependent and helicase-independent functions for eIF4A, but the multifaceted roles of eIF4A have not been fully explored. Here we show that eIF4A1 enhances translational repression during the inhibition of mechanistic target of rapamycin complex 1 (mTORC1), an essential kinase complex controlling cell proliferation. RNA pulldown followed by sequencing revealed that eIF4A1 preferentially binds to mRNAs containing terminal oligopyrimidine (TOP) motifs, whose translation is rapidly repressed upon mTORC1 inhibition. This selective interaction depends on a La-related RNA-binding protein, LARP1. Ribosome profiling revealed that deletion of EIF4A1 attenuated the translational repression of TOP mRNAs upon mTORC1 inactivation. Moreover, eIF4A1 increases the interaction between TOP mRNAs and LARP1 and, thus, ensures stronger translational repression upon mTORC1 inhibition. Our data show the multimodality of eIF4A1 in modulating protein synthesis through an inhibitory binding partner and provide a unique example of the repressive role of a universal translational activator.

DOI： 10.1038/s41594-024-01321-7

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：Journal of Experimental and Clinical Cancer Research  

BACKGROUND: Although immune checkpoint blockade (ICB) therapy has proven to be extremely effective at managing certain cancers, its efficacy in treating pancreatic ductal adenocarcinoma (PDAC) has been limited. Therefore, enhancing the effect of ICB could improve the prognosis of PDAC. In this study, we focused on the histamine receptor H1 (HRH1) and investigated its impact on ICB therapy for PDAC. METHODS: We assessed HRH1 expression in pancreatic cancer cell (PCC) specimens from PDAC patients through public data analysis and immunohistochemical (IHC) staining. The impact of HRH1 in PCCs was evaluated using HRH1 antagonists and small hairpin RNA (shRNA). Techniques including Western blot, flow cytometry, quantitative reverse transcription polymerase chain reaction (RT-PCR), and microarray analyses were performed to identify the relationships between HRH1 and major histocompatibility complex class I (MHC-I) expression in cancer cells. We combined HRH1 antagonism or knockdown with anti-programmed death receptor 1 (αPD-1) therapy in orthotopic models, employing IHC, immunofluorescence, and hematoxylin and eosin staining for assessment. RESULTS: HRH1 expression in cancer cells was negatively correlated with HLA-ABC expression, CD8+ T cells, and cytotoxic CD8+ T cells. Our findings indicate that HRH1 blockade upregulates MHC-I expression in PCCs via cholesterol biosynthesis signaling. In the orthotopic model, the combined inhibition of HRH1 and αPD-1 blockade enhanced cytotoxic CD8+ T cell penetration and efficacy, overcoming resistance to ICB therapy. CONCLUSIONS: HRH1 plays an immunosuppressive role in cancer cells. Consequently, HRH1 intervention may be a promising method to amplify the responsiveness of PDAC to immunotherapy.

DOI： 10.1186/s13046-024-03060-5

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：Journal of Inflammation Research  

BACKGROUND: Acute lung injury (ALI) is caused by a variety of illnesses, including aspiration pneumonia and sepsis. The CCR4-NOT complex is a large multimeric protein complex that degrades mRNA through poly(A) tail shortening, whereas it also contributes to regulation of transcription and translation. Cnot3 is a scaffold component of the CCR4-NOT complex and is essential for the integrity of the complex; loss of Cnot3 leads to depletion of whole complex. While the significance of cytokine mRNA degradation in limiting inflammation has been established, the roles of CCR4-NOT complex-mediated in ALI remain elusive. METHODS: The effects of Cnot3 haploinsufficiency in the pathology and cytokine expression were analyzed in the mouse lungs of acid aspiration-induced acute lung injury. The decay rate and transcription activity of cytokine mRNAs under Cnot3 heterozygous deletion were analyzed in lipopolysaccharide (LPS) -stimulated mouse embryonic fibroblasts (MEFs). RESULTS: Tamoxifen-induced heterozygous deletion of Cnot3 in adult mice (Cnot3 Hetz) did not show body weight loss or any apparent abnormality. Under acid aspiration-induced acute lung injury, Cnot3 Hetz mice exhibited increased pulmonary edema, worse lung pathologies and more severe inflammation compared with wild type mice. mRNA expression of pro-inflammatory genes Il1b and Nos2 were significantly upregulated in the lungs of Cnot3 Hetz mice. Consistently, mRNA expression of Il1b and Nos2 was upregulated in LPS-stimulated Cnot3 Hetz MEFs. Mechanistically, while heterozygous depletion of Cnot3 stabilized both Il1b and Nos2 mRNAs, the nascent pre-mRNA level of Il1b was upregulated in Cnot3 Hetz MEFs, implicating Cnot3-mediated transcriptional repression of Il1b expression in addition to destabilization of Il1b and Nos2 mRNAs. PU.1 (Spi1) was identified as a causative transcription factor to promote Il1b expression under Cnot3 haploinsufficient conditions. CONCLUSION: CNOT3 plays a protective role in ALI by suppressing expression of pro-inflammatory genes Il1b and Nos2 through both post-transcriptional and transcriptional mechanisms, including mRNA stability control of Spi1.

DOI： 10.2147/JIR.S468612

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While group-2 innate lymphoid cells (ILC2s) are highly proliferative in allergic inflammation, the removal of overactivated ILC2s in allergic diseases has not been investigated. We previously showed that chronic airway allergy induces "exhausted-like" dysfunctional ILC2s expressing T cell immunoreceptor with Ig and ITIM domains (TIGIT). However, the physiological relevance of these cells in chronic allergy remains elusive. To precisely identify and monitor TIGIT+ ILC2s, we generated TIGIT lineage tracer mice. Chronic allergy stably induced TIGIT+ ILC2s, which were highly activated, apoptotic, and were quickly removed from sites of chronic allergy. Transcripts from coding genes were globally suppressed in the cells, possibly due to reduced chromatin accessibility. Cell death in TIGIT+ ILC2s was enhanced by interactions with CD155 expressed on macrophages, whereas genetic ablation of Tigit or blockade by anti-TIGIT antagonistic antibodies promoted ILC2 survival, thereby deteriorating chronic allergic inflammation. Our work demonstrates that TIGIT shifts the fate of ILC2s toward activation-induced cell death, which could present a new therapeutic target for chronic allergies.

DOI： 10.1084/jem.20222005

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：Biochemical and Biophysical Research Communications  

Apelin (APL), an endogenous ligand for APJ, has been reported to be upregulated in a murine model of acute colitis induced by sodium dextran sulfate, as well as inflammatory bowel diseases (IBD) in humans. However, the mechanisms and functions of APL/APJ axis in the pathogenesis of IBD are unclear. We herein analyzed CD4+ T cells to determine the functions of APL in a murine model of chronic colitis induced in Rag deficient mice (Rag-/-). In colonic tissues of wild-type mice (WT), we found that APL was expressed especially in the lamina propria lymphocytes, where CD4+ T cells are dominant, rather than the epithelial cells. Unexpectedly, the APL expression was rather downregulated in the colonic tissue of the chronic colitis group compared to the control groups (Rag-/- before colitis induction and WT). The APL expression was downregulated when naïve T cells were differentiated into effecter T cells. A lack of APL resulted in decreased naïve T cells and increased effecter T cells in secondary lymphoid organs. A synthetic APL peptide, [Pyr1]-APL-13, increased IL-10 and decreased IFN-γ productions by effecter T cells. Administration of [Pyr1]-APL-13 improved survival rate in association with lessened colitis severity and decreased pro-inflammatory cytokine production. This is the first report showing immunological function of APL specifically on T cells, and these results indicate that APL/APJ axis may be a novel therapeutic target for IBD.

DOI： 10.1016/j.bbrc.2023.01.068

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BACKGROUND: The mechanism underlying cancer cell metastasis from the tumor to regional lymph nodes is not yet fully understood. We hypothesized that peritumoral neutrophil accumulation promotes regional lymph node metastasis in thoracic esophageal squamous cell cancer. METHODS: Between 2010 and 2019, 126 thoracic esophageal squamous cell cancer patients received curative (R0) esophagectomy without preoperative treatment in our hospital. Using paraffin-embedded resected tumors, we performed immunohistochemical analysis of CD16b-positive neutrophil accumulation in the peritumoral area, which was defined as a 1-mm region centered on the border separating the malignant cell nests from the host tissue. The relationship between the density of peritumoral CD16b staining and pathological lymph node metastasis or 5-year overall survival was evaluated. RESULTS: Although the clinicopathological characteristics of CD16b-high and CD16b-low patients did not differ, greater pathological lymph node metastasis (P < .001) and lymphatic invasion by the tumor (P = .024) and a poorer 5-year survival (P = .010) were seen in CD16b-high patients. Moreover, CD16b-positive neutrophil density was generally higher in the peritumoral area than within the tumor itself. Univariate and multivariate analyses showed that CD16b-positive neutrophil accumulation was an independent factor for lymph node metastasis with an odds ratio >25 (P < .001). On the other hand, blood neutrophil counts did not correlate with lymph node metastasis. CONCLUSION: Peritumoral accumulation of CD16b-positive neutrophils is an independent factor strongly correlated with lymph node metastasis in thoracic esophageal squamous cell cancer.

DOI： 10.1016/j.surg.2021.11.022

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Angiotensin-converting enzyme 2 (ACE2) is the carboxypeptidase to degrade angiotensin II (Ang II) to angiotensin 1-7 (Ang 1-7) and improves the pathologies of cardiovascular disease and acute respiratory distress syndrome (ARDS)/acute lung injury. B38-CAP is a bacteria-derived ACE2-like carboxypeptidase as potent as human ACE2 and ameliorates hypertension, heart failure and SARS-CoV-2-induced lung injury in mice. Recombinant B38-CAP is prepared with E. coli protein expression system more efficiently than recombinant soluble human ACE2. Here we show therapeutic effects of B38-CAP on abdominal sepsis- or acid aspiration-induced acute lung injury. ACE2 expression was downregulated in the lungs of mice with cecal ligation puncture (CLP)-induced sepsis or acid-induced lung injury thereby leading to upregulation of Ang II levels. Intraperitoneal injection of B38-CAP significantly decreased Ang II levels while upregulated angiotensin 1-7 levels. B38-CAP improved survival rate of the mice under sepsis. B38-CAP suppressed the pathologies of lung inflammation, improved lung dysfunction and downregulated elevated cytokine mRNA levels in the mice with acute lung injury. Thus, systemic treatment with an ACE2-like enzyme might be a potential therapeutic strategy for the patients with severe sepsis or ARDS.

DOI： 10.1371/journal.pone.0270920

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Publishing type：Research paper (scientific journal)   Publisher：Japanese Pharmacological Society  

DOI： 10.1254/jpssuppl.95.0_2-o-041

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Language：Others   Publishing type：Research paper (scientific journal)  

The duration of viral shedding is determined by a balance between de novo infection and removal of infected cells. That is, if infection is completely blocked with antiviral drugs (100% inhibition), the duration of viral shedding is minimal and is determined by the length of virus production. However, some mathematical models predict that if infected individuals are treated with antiviral drugs with efficacy below 100%, viral shedding may last longer than without treatment because further de novo infections are driven by entry of the virus into partially protected, uninfected cells at a slower rate. Using a simple mathematical model, we quantified SARS-CoV-2 infection dynamics in non-human primates and characterized the kinetics of viral shedding. We counterintuitively found that treatments initiated early, such as 0.5 d after virus inoculation, with intermediate to relatively high efficacy (30–70% inhibition of virus replication) yield a prolonged duration of viral shedding (by about 6.0 d) compared with no treatment.

DOI： 10.26508/lsa.202101049

Language：English   Publishing type：Research paper (scientific journal)  

The spatial organization of chromatin is known to be highly dynamic in response to environmental stress. However, it remains unknown how chromatin dynamics contributes to or modulates disease pathogenesis. Here, we show that upon influenza virus infection, the H4K20me3 methyltransferase Suv4-20h2 binds the viral protein NP, which results in the inactivation of Suv4-20h2 and the dissociation of cohesin from Suv4-20h2. Inactivation of Suv4-20h2 by viral infection or genetic deletion allows the formation of an active chromatin loop at the HoxC8-HoxC6 loci coincident with cohesin loading. HoxC8 and HoxC6 proteins in turn enhance viral replication by inhibiting the Wnt-β-catenin mediated interferon response. Importantly, loss of Suv4-20h2 augments the pathology of influenza infection in vivo. Thus, Suv4-20h2 acts as a safeguard against influenza virus infection by suppressing cohesin-mediated loop formation.

DOI： 10.1016/j.isci.2021.102660

Language：English   Publishing type：Research paper (scientific journal)  

Idiopathic pure red cell aplasia (PRCA) and secondary PRCA associated with thymoma and large granular lymphocyte leukemia are generally considered to be immune-mediated. The PRCA2004/2006 study showed that poor responses to immunosuppression and anemia relapse were associated with death. PRCA may represent the prodrome to MDS. Thus, clonal hematopoiesis may be responsible for treatment failure. We investigated gene mutations in myeloid neoplasm-associated genes in acquired PRCA. We identified 21 mutations affecting amino acid sequences in 11 of the 38 adult PRCA patients (28.9%) using stringent filtering of the error-prone sequences and SNPs. Four PRCA patients showed 7 driver mutations in TET2, DNMT3A and KDM6A, and 2 PRCA patients carried multiple mutations in TET2. Five PRCA patients had mutations with high VAFs exceeding 0.3. These results suggest that clonal hematopoiesis by stem/progenitor cells might be related to the pathophysiology of chronic PRCA in certain adult patients.

DOI： 10.1038/s41598-021-81890-5

Language：English   Publishing type：Research paper (scientific journal)  

Language：Others   Publishing type：Research paper (scientific journal)  

DOI： 10.1096/fj.202000483r

Language：Others   Publishing type：Research paper (scientific journal)  

© 2020 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd Esophageal squamous cell carcinoma (ESCC) is one of the most fatal types of malignant tumors worldwide. Epitranscriptome, such as N6-methyladenosine (m6A) of mRNA, is an abundant post-transcriptional mRNA modification and has been recently implicated to play roles in several cancers, whereas the significance of m6A modifications is virtually unknown in ESCC. Analysis of tissue microarray of the tumors in 177 ESCC patients showed that higher expression of m6A demethylase ALKBH5 correlated with poor prognosis and that ALKBH5 was an independent prognostic factor of the survival of patients. There was no correlation between the other demethylase FTO and prognosis. siRNA knockdown of ALKBH5 but not FTO significantly suppressed proliferation and migration of human ESCC cells. ALKBH5 knockdown delayed progression of cell cycle and accumulated the cells to G0/G1 phase. Mechanistically, expression of CDKN1A (p21) was significantly up-regulated in ALKBH5-depleted cells, and m6A modification and stability of CDKN1A mRNA were increased by ALKBH5 knockdown. Furthermore, depletion of ALKBH5 substantially suppressed tumor growth of ESCC cells subcutaneously transplanted in BALB/c nude mice. Collectively, we identify ALKBH5 as the first m6A demethylase that accelerates cell cycle progression and promotes cell proliferation of ESCC cells, which is associated with poor prognosis of ESCC patients.

DOI： 10.1111/gtc.12792

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1080/15476286.2019.1709747.

Language：English   Publishing type：Research paper (scientific journal)  

The CCR4-NOT Deadenylase Complex Maintains Adipocyte Identity.
Shortening of poly(A) tails triggers mRNA degradation; hence, mRNA deadenylation regulates many biological events. In the present study, we generated mice lacking the Cnot1 gene, which encodes an essential scaffold subunit of the CCR4-NOT deadenylase complex in adipose tissues (Cnot1-AKO mice) and we examined the role of CCR4-NOT in adipocyte function. Cnot1-AKO mice showed reduced masses of white adipose tissue (WAT) and brown adipose tissue (BAT), indicating abnormal organization and function of those tissues. Indeed, Cnot1-AKO mice showed hyperinsulinemia, hyperglycemia, insulin resistance, and glucose intolerance and they could not maintain a normal body temperature during cold exposure. Muscle-like fibrous material appeared in both WAT and BAT of Cnot1-AKO mice, suggesting the acquisition of non-adipose tissue characteristics. Gene expression analysis using RNA-sequencing (RNA-seq) showed that the levels of adipose tissue-related mRNAs, including those of metabolic genes, decreased, whereas the levels of inflammatory response-related mRNAs increased. These data suggest that the CCR4-NOT complex ensures proper adipose tissue function by maintaining adipocyte-specific mRNAs at appropriate levels and by simultaneously suppressing mRNAs that would impair adipocyte function if overexpressed.

DOI： 10.3390/ijms20215274

Language：Others  

Apelin inhibition prevents resistance and metastasis associated with anti-angiogenic therapy.

DOI： 10.15252/emmm.201809266

Language：English   Publishing type：Research paper (scientific journal)  

Pharmacological interventions to enhance fibrinolysis are effective for treating thrombotic disorders. Utilizing the in vitro U937 cell line-based fibrin degradation assay, we had previously found a cyclic pentapeptide malformin A1 (MA1) as a novel activating compound for cellular fibrinolytic activity. The mechanism by which MA1 enhances cellular fibrinolytic activity remains unknown. In the present study, we show that RSK1 is a crucial mediator of MA1-induced cellular fibrinolysis. Treatment with rhodamine-conjugated MA1 showed that MA1 localizes mainly in the cytoplasm of U937 cells. Screening with an antibody macroarray revealed that MA1 induces the phosphorylation of RSK1 at Ser380 in U937 cells. SL0101, an inhibitor of RSK, inhibited MA1-induced fibrinolytic activity, and CRISPR/Cas9-mediated knockout of RSK1 but not RSK2 suppressed MA1-enhanced fibrinolysis in U937 cells. Synthetic active MA1 derivatives also induced the phosphorylation of RSK1. Furthermore, MA1 treatment stimulated phosphorylation of ERK1/2 and MEK1/2. PD98059, an inhibitor of MEK1/2, inhibited MA1-induced phosphorylation of RSK1 and ERK1/2, indicating that MA1 induces the activation of the MEK-ERK-RSK pathway. Moreover, MA1 upregulated the expression of urokinase-type plasminogen activator (uPA) and increased uPA secretion. These inductions were abrogated in RSK1 knockout cells. These results indicate that RSK1 is a key regulator of MA1-induced extracellular fibrinolytic activity.

DOI： 10.1038/s41598-018-23745-0

Language：English   Publishing type：Research paper (scientific journal)  

The multi-subunit CCR4 (carbon catabolite repressor 4)-NOT (Negative on TATA) complex serves as a central coordinator of all different steps of eukaryotic gene expression. Here we performed a systematic and comparative analysis of cells where the CCR4-NOT subunits CNOT1, CNOT2 or CNOT3 were individually downregulated using doxycycline-inducible shRNAs. Microarray experiments showed that downregulation of either CNOT subunit resulted in elevated expression of major histocompatibility complex class II (MHC II) genes which are found in a gene cluster on chromosome 6. Increased expression of MHC II genes after knock-down or knock-out of either CNOT subunit was seen in a variety of cell systems and also in naive macrophages from CNOT3 conditional knock-out mice. CNOT2-mediated repression of MHC II genes occurred also in the absence of the master regulator class II transactivator (CIITA) and did not cause detectable changes of the chromatin structure at the chromosomal MHC II locus. CNOT2 downregulation resulted in an increased de novo transcription of mRNAs whereas tethering of CNOT2 to a regulatory region governing MHC II expression resulted in diminished transcription. These results expand the known repertoire of CCR4-NOT members for immune regulation and identify CNOT proteins as a novel group of corepressors restricting class II expression.

DOI： 10.1038/s41598-017-03708-7

Language：English   Publishing type：Research paper (scientific journal)  

For acute respiratory distress syndrome (ARDS), mechanical ventilation (MV) is a life-saving intervention without alternative; however, MV can cause ventilator-induced lung injury. Reactive oxygen species (ROS) play important roles in the pathogenesis of both ARDS and ventilator-induced lung injury. Lecithinized superoxide dismutase (PC-SOD) overcomes the limitations of superoxide dismutase such as low tissue affinity and low stability in plasma. In this study, we examined the effect of PC-SOD on tissue injury, edema, and inflammation in the lung and other organs of mice subjected to cecal ligation and puncture (CLP), LPS administration, or MV. The severity of the lung injury was assessed on the basis of vascular permeability, histopathologic evaluation, and lung mechanics. Intravenous PC-SOD administration (the first administered just before CLP) increased the survival rate and decreased vascular permeability in mice subjected to CLP. PC-SOD, but not dexamethasone or sivelestat sodium hydrate (sivelestat), suppressed CLP-induced kidney injury and systemic inflammation. PC-SOD also suppressed vascular permeability, tissue injury, and inflammation in the lung induced by LPS administration. Moreover, PC-SOD, but not dexamethasone or sivelestat, suppressed vascular permeability, edema, tissue injury, and mechanical alterations in the lung induced by MV. In vivo imaging analysis of ROS revealed that CLP, LPS administration, and MV increased the level of ROS and that this increase was suppressed by PC-SOD. The results of this study thus suggest that, on the basis of its ROS-reducing properties, intravenous administration of PC-SOD may be beneficial for patients at high risk of developing ARDS.

DOI： 10.1165/rcmb.2016-0158OC

Language：Others   Publishing type：Research paper (scientific journal)  

Vps34 regulates myofibril proteostasis to prevent hypertrophic cardiomyopathy.

DOI： 10.1172/jci.insight.89462

Language：English   Publishing type：Research paper (scientific journal)  

The formation of blood clots in blood vessels causes severe ischemic diseases such as cerebral infarction and myocardial infarction. While searching for microbial products that increase fibrinolytic activity using an in vitro fibrin degradation assay, we found malformin Al, a disulfide form of cyclo(-D-Cys-n-Cys-L-Valn-Leu-L-Ile-), as an active compound. In this study, we synthesized malformin derivatives using a solid phase peptide synthesis method and evaluated their fibrinolytic activity and cytotoxicity. Reduction of the disulfide bond and linearization of the cyclic peptide frame decreased the pro-fibrinolytic activity. Substitution of a branched-chain amino acid with lysine resulted in loss of activity. However, protection of the amino group in the lysine derivatives by the tert-butoxycarbonyl (Boc) group rescued the inactivity. Furthermore, the phenylalanine derivatives also exhibited a similar pro-fibrinolytic effect compared to malformin Al. These results suggest that the disulfide bond, the cyclic peptide frame, and the bulky hydrophobic side chains play a crucial role in the pro-fibrinolytic activity of malformin. The effective dose of the active derivatives for the in vitro fibrin degradation showed similar ranges (1-5 mu M), while the order of cytotoxic potency for the active derivatives was as follows: Phe-derivatives > BocLys-derivatives > malformin Al > reduced form. These results showed no correlation between pro-fibrinolytic activity and cytotoxicity, suggesting the possibility of the synthesis for non-toxic malformin derivatives possessing the activity. (C) 2016 The Author(s). Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license.

DOI： 10.1016/j.bmcl.2016.09.045

Language：English   Publishing type：Research paper (scientific journal)  

Background: Nanoparticles have become a key technology in multiple industries. However, there are growing reports of the toxicity of nanomaterials to humans. In particular, nanomaterials have been linked to lung diseases. The molecular mechanisms of nanoparticle toxicity are largely unexplored.
Methods: Acute lung injury was induced in wild-type mice and angiotensin-coverting enzyme 2 (ACE2) knockout mice by the intratracheal instillation of cationic polyamidoamine dendrimer (PAMAM) nanoparticles. For rescue experiments, losartan (15 mg/kg in PBS) was injected intraperitoneally 30 min before nanoparticle administration.
Results: Some PAMAM nanoparticles, but not anionic PAMAM nanoparticles or carbon nanotubes, triggered acute lung failure in mice. Mechanistically, cationic nanoparticles can directly bind ACE2, decrease its activity and down-regulate its expression level in lung tissue, resulting in deregulation of the renin-angiotensin system. Gene inactivation of Ace2 can exacerbate lung injury. Importantly, the administration of losartan, which is an angiotensin II type I receptor antagonist, can ameliorate PAMAM nanoparticle-induced lung injury.
Conclusions: Our data provide molecular insight into PAMAM nanoparticle-induced lung injury and suggest potential therapeutic and screening strategies to address the safety of nanomaterials.

DOI： 10.1186/s12989-015-0080-x

Language：English   Publishing type：Research paper (scientific journal)  

Cholesterol metabolism is closely interrelated with cardiovascular disease in humans. Dietary supplementation with omega-6 polyunsaturated fatty acids including arachidonic acid (AA) was shown to favorably affect plasma LDL-C and HDL-C. However, the underlying mechanisms are poorly understood. By combining data from a GWAS screening in >100,000 individuals of European ancestry, mediator lipidomics, and functional validation studies in mice, we identify the AA metabolome as an important regulator of cholesterol homeostasis. Pharmacological modulation of AA metabolism by aspirin induced hepatic generation of leukotrienes (LTs) and lipoxins (LXs), thereby increasing hepatic expression of the bile salt export pump Abcb11. Induction of Abcb11 translated in enhanced reverse cholesterol transport, one key function of HDL. Further characterization of the bioactive AA-derivatives identified LX mimetics to lower plasma LDL-C. Our results define the AA metabolome as conserved regulator of cholesterol metabolism, and identify AA derivatives as promising therapeutics to treat cardiovascular disease in humans.

DOI： 10.1016/j.cmet.2014.09.004

Language：English   Publishing type：Research paper (scientific journal)  

Dilated cardiomyopathy (DCM) is characterized by ventricular dilation associated with systolic dysfunction, which could be caused by mutations in lamina/C gene (LMNA). LMNA-linked DCM is severe in males in both human patients and a knock-in mouse model carrying a homozygous p.H222P mutation (Lmna(H222P/H222P)). The aim of this study was to investigate the molecular mechanisms underlying the gender difference of LMNA-linked DCM.
A whole-exome analysis of a multiplex family with DCM exhibiting the gender difference revealed a DCM-linked LMNA mutation, p.R225X. Immunohistochemical analyses of neonatal rat cardiomyocytes expressing mutant LMNA constructs and heart samples from the LMNA-linked DCM patients and Lmna(H222P/H222P) mice demonstrated a nuclear accumulation of androgen receptor (AR) and its co-activators, serum response factor, and four-and-a-half LIM protein-2. Role of sex hormones in the gender difference was investigated in vivo using the Lmna(H222P/H222P) mice, where male and female mice were castrated and ovariectomized, respectively, or treated with testosterone or an antagonist of AR. Examination of the mice by echocardiography, followed by the analyses of histological changes and gene/protein expression profiles in the hearts, confirmed the involvement of testicular hormone in the disease progression and enhanced cardiac remodelling in the Lmna(H222P/H222P) mice.
These observations indicated that nuclear accumulation of AR was associated with the gender difference in LMNA-linked DCM.

DOI： 10.1093/cvr/cvt106

Language：English  

Angiotensin-converting enzyme 2 (ACE2) is a negative regulator of the renin-angiotensin system, and functions as the key SARS coronavirus receptor and stabilizer of neutral amino acid transporters. ACE2 catalyzes the conversion of angiotensin II to angiotensin 1-7, thereby counterbalancing ACE activity. Accumulating evidence indicates that the enzymatic activity of ACE2 has a protective role in cardiovascular diseases. Loss of ACE2 can be detrimental, as it leads to functional deterioration of the heart and progression of cardiac, renal, and vascular pathologies. Recombinant soluble human ACE2 protein has been demonstrated to exhibit beneficial effects in various animal models, including cardiovascular diseases. ACE2 is a multifunctional enzyme and thus potentially acts on other vasoactive peptides, such as Apelin, a vital regulator of blood pressure and myocardium contractility. In addition, ACE2 is structurally a chimeric protein that has emerged from the duplication of 2 genes: homology with ACE at the carboxypeptidase domain and homology with Collectrin in the transmembrane C-terminal domain. ACE2 has been implicated in the pathology of Hartnup's disease, a disorder of amino acid homeostasis, and, via its function in amino acid transport, it has been recently revealed that ACE2 controls intestinal inflammation and diarrhea, thus regulating the gut microbiome. This review summarizes and discusses the structure and multiple functions of ACE2 and the relevance of this key enzyme in disease pathogenesis. (Circ J 2013; 77: 301-308)

DOI： 10.1253/circj.CJ-12-1544

Language：English   Publishing type：Research paper (scientific journal)  

Malnutrition affects up to one billion people in the world and is a major cause of mortality(1,2). In many cases, malnutrition is associated with diarrhoea and intestinal inflammation, further contributing to morbidity and death(2). The mechanisms by which unbalanced dietary nutrients affect intestinal homeostasis are largely unknown. Here we report that deficiency in murine angiotensin I converting enzyme (peptidyl-dipeptidase A) 2 (Ace2), which encodes a key regulatory enzyme of the renin-angiotensin system (RAS), results in highly increased susceptibility to intestinal inflammation induced by epithelial damage. The RAS is known to be involved in acute lung failure(3), cardiovascular functions(4) and SARS infections(5). Mechanistically, ACE2 has a RAS-independent function, regulating intestinal amino acid homeostasis, expression of antimicrobial peptides, and the ecology of the gut microbiome. Transplantation of the altered microbiota from Ace2 mutant mice into germ-free wild-type hosts was able to transmit the increased propensity to develop severe colitis. ACE2-dependent changes in epithelial immunity and the gut microbiota can be directly regulated by the dietary amino acid tryptophan. Our results identify ACE2 as a key regulator of dietary amino acid homeostasis, innate immunity, gut microbial ecology, and transmissible susceptibility to colitis. These results provide a molecular explanation for how amino acid malnutrition can cause intestinal inflammation and diarrhoea.

DOI： 10.1038/nature11228

Language：English   Publishing type：Research paper (scientific journal)  

Both acute and chronic apelin treatment have been shown to improve insulin sensitivity in mice. However, the effects of apelin on fatty acid oxidation (FAO) during obesity-related insulin resistance have not yet been addressed. Thus, the aim of the current study was to determine the impact of chronic treatment on lipid use, especially in skeletal muscles. High-fat diet (HFD)induced obese and insulin-resistant mice treated by an apelin injection (0.1 mu mol/kg/day i.p.) during 4 weeks had decreased fat mass, glycemia, and plasma levels of triglycerides and were protected from hyperinsulinemia compared with HFD PBS-treated mice. Indirect calorimetry experiments showed that apelin-treated mice had a better use of lipids. The complete FAO, the oxidative capacity, and mitochondrial biogenesis were increased in soleus of apelin-treated mice. The action of apelin was AMP-activated protein kinase (AMPK) dependent since all the effects studied were abrogated in HFD apelin-treated mice with muscle-specific inactive AMPK. Finally, the apelin-stimulated improvement of oxidative capacity led to decreased levels of acylcarnitines and enhanced insulin-stimulated glucose uptake in soleus. Thus, by promoting complete lipid use in muscle of insulin-resistant mice through mitochondria' biogenesis and fighter matching between FAO and the tricarboxylic acid cycle, apelin treatment could contribute to insulin sensitivity improvement. Diabetes 61:310-320, 2012

DOI： 10.2337/db11-0100

Language：English  

Angiotensin-converting enzyme (ACE) 2 is a homolog to the carboxypeptidase ACE, which generates angiotensin II, the main active peptide of renin-angiotensin system (RAS). After the cloning of ACE2 in 2000, three major ACE2 functions have been described so far. First ACE2 has emerged as a potent negative regulator of the RAS counterbalancing the multiple functions of ACE. By targeting angiotensin II ACE2 exhibits a protective role in the cardiovascular system and many other organs. Second ACE2 was identified as an essential receptor for the SARS coronavirus that causes severe acute lung failure. Downregulation of ACE2 strongly contributes to the pathogenesis of severe lung failure. Third, both ACE2 and its homologue Collectrin can associate with amino acid transporters and play essential role in the absorption of amino acids in the kidney and gut. In this review, we will discuss the multiple biological functions of ACE2. (C) 2010 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.pharmthera.2010.06.003

Language：English   Publishing type：Research paper (scientific journal)  

Macrophages have critical roles in the pathogenesis of atherosclerosis by activating the innate immune system and producing inflammatory cytokines. Accumulating evidence indicates that angiotensin type 1 receptor (AT1R) blockers exert anti-inflammatory effects in inflammatory diseases including atherosclerosis. In this study, we investigated the effect of losartan, an AT1R blocker, on the proinflammatory gene expression induced by bacterial lipopolysaccharide (LPS) in a well-defined in vitro human THP-1 macrophage system. We found that losartan significantly attenuated the LPS-induced expression of proinflammatory genes TNF-alpha, IL-8 and COX-2. However, exogenous angiotensin II (AngII) had no effect on LPS-induced inflammatory signaling despite the expression of AT1R. In addition, losartan did not block LPS-induced I kappa B phosphorylation, which is downstream of Toll-like receptor activation. Peroxisome proliferator-activated receptor-gamma (PPAR gamma) antagonists, GW9662 and T0070907, reversed the inhibitory effects of losartan on LPS-induced TNF-alpha and IL-8 expression in THP-1 macrophages. These observations suggest that losartan inhibits LPS-induced proinflammatory gene expression in macrophages by activating the PPAR gamma pathway rather than by the competitive inhibition of AT1R binding to AngII. Hypertension Research (2010) 33, 831-835; doi: 10.1038/hr.2010.79; published online 27 May 2010

DOI： 10.1038/hr.2010.79

Language：English  

Angiotensin-converting enzyme 2 (ACE2), a first homolog of ACE, regulates the renin-angiotensin system by counterbalancing ACE activity. Accumulating evidence in recent years has demonstrated a physiological and pathological role of ACE2 in the cardiovascular, renal and respiratory systems. For instance, in the acute respiratory distress syndrome (ARDS), ACE, AngII, and AT1R promote the disease pathogenesis, whereas ACE2 and the AT2R protect from ARDS. Importantly, ACE2 has been identified as a key SARS-coronavirus receptor and plays a protective role in SARS pathogenesis. Furthermore, the recent explosion of research into the ACE2 homolog, collectrin, has revealed a new physiological function of ACE2 as an amino acid transporter, which explains the pathogenic role of gene mutations in Hartnup disorder. This review summarizes and discusses the recently unveiled roles for ACE2 in disease pathogenesis. (Circ J 2010; 74: 405-410)

DOI： 10.1253/circj.CJ-10-0045

Language：English   Publishing type：Research paper (scientific journal)  

Background: PI3K gamma functions in the immune compartment to promote inflammation in response to G-protein-coupled receptor (GPCR) agonists and PI3K gamma also acts within the heart itself both as a negative regulator of cardiac contractility and as a pro-survival factor. Thus, PI3K gamma has the potential to both promote and limit M I/R injury.
Methodology/Principal Findings: Complete PI3K gamma(-/-) mutant mice, catalytically inactive PI3K gamma(KD/KD) (KD) knock-in mice, and control wild type (WT) mice were subjected to in vivo myocardial ischemia and reperfusion (M I/R) injury. Additionally, bone-marrow chimeric mice were constructed to elucidate the contribution of the inflammatory response to cardiac damage. PI3K gamma(-/-) mice exhibited a significantly increased infarction size following reperfusion. Mechanistically, PI3K gamma is required for activation of the Reperfusion Injury Salvage Kinase (RISK) pathway (AKT/ERK1/2) and regulates phospholamban phosphorylation in the acute injury response. Using bone marrow chimeras, the cardioprotective role of PI3K gamma was mapped to non-haematopoietic cells. Importantly, this massive increase in M I/R injury in PI3K gamma(-/-) mice was rescued in PI3K gamma kinase-dead (PI3K gamma(KD/KD)) knock-in mice. However, PI3K gamma(KD/KD) mice exhibited a cardiac injury similar to wild type animals, suggesting that specific blockade of PI3K gamma catalytic activity has no beneficial effects.
Conclusions/Significance: Our data show that PI3K gamma is cardioprotective during M I/R injury independent of its catalytic kinase activity and that loss of PI3K gamma function in the hematopoietic compartment does not affect disease outcome. Thus, clinical development of specific PI3K gamma blockers should proceed with caution.

DOI： 10.1371/journal.pone.0009350

Language：English   Publishing type：Research paper (scientific journal)  

Background: Although therapeutic angiogenesis is a most promising strategy for the treatment of myocardial infarction (MI), it remains unknown if and how endogenous angiogenesis inhibitors, such as endostatin, regulate angiogenesis in MI. In the present study the role of endostatin in left ventricular (LV) remodeling and heart failure was tested in a rat MI model.
Methods and Results: When exposed to hypoxia, rat cardiomyocytes showed increased expression of endostatin. After MI induction in the rat MI model, endostatin expression was upregulated in cardiomyocytes, and serum endostatin levels were significantly elevated. Anti-endostatin antibody treatment resulted in significantly higher mortality of MI rats than controls. The MI rats with endostatin neutralization displayed adverse LV remodeling and severe heart failure compared with control MI rats. Although angiogenesis was increased, tissue remodeling and interstitial fibrosis were further exaggerated in post-MI hearts by endostatin neutralization. Furthermore, the expression and protease activity of matrix metalloproteinases -2 and -9, and of angiotensin-converting enzyme were markedly elevated by endostatin neutralization.
Conclusions: Neutralization of endostatin worsens the symptoms and outcomes of MI in a rat model. The results imply that endogenous endostatin/collagen XVIII may suppress aberrant LV remodeling and heart failure after MI. (Circ J 2010; 74: 109-119)

DOI： 10.1253/circj.CJ-09-0486

Language：English   Publishing type：Research paper (scientific journal)  

The orphan transporter Slc6a18 (XT2) is highly expressed at the luminal membrane of kidney proximal tubules and displays similar to 50% identity with Slc6a19 (B(0)AT1), which is the main neutral amino acid transporter in both kidney and small intestine. As yet, the amino acid transport function of XT2 has only been experimentally supported by the urinary glycine loss observed in xt2 null mice. We report here that in Xenopus laevis oocytes, co-expressed ACE2 (angiotensin-converting enzyme 2) associates with XT2 and reveals its function as a Na+- and Cl--dependent neutral amino acid transporter. In contrast to its association with ACE2 observed in Xenopus laevis oocytes, our experiments with ace2 and collectrin null mice demonstrate that in vivo it is Collectrin, a smaller homologue of ACE2, that is required for functional expression of XT2 in kidney. To assess the function of XT2 in vivo, we reanalyzed its knock-out mouse model after more than 10 generations of backcrossing into C57BL/6 background. In addition to the previously published glycinuria, we observed a urinary loss of several other amino acids, in particular beta-branched and small neutral ones. Using telemetry, we confirmed the previously described link of XT2 absence with hypertension but only in physically restrained animals. Taken together, our data indicate that the formerly orphan transporter XT2 functions as a sodium and chloride-dependent neutral amino acid transporter that we propose to rename B(0)AT3.

DOI： 10.1074/jbc.M109.011171

Language：English   Publishing type：Research paper (scientific journal)  

Hepatocyte growth factor (HGF) promotes malignant development of cancer cells by enhancing invasion and metastasis. NK4, a competitive antagonist for HGF, is a bifunctional molecule that acts as a HGF antagonist and angiogenesis inhibitor. Although successful tumor inhibition by NK4 gene expression in tumor models has been demonstrated, the effects of systemic NK4 gene introduction are yet to be addressed. Here we show that systemic administration of a replication-defective adenovirus expressing NK4 (Ad.NK4) inhibits tumor growth and lung metastasis of B16F10 melanoma and Lewis lung carcinoma in syngeneic mice. Single tail-vein injection of Ad.NK4 achieved therapeutic levels of NK4 in the circulation and in multiple organs. Despite NK4 expression that was highest in the liver, toxicity in the liver was minimal. Ad.NK4-mediated growth inhibition was associated with decreased blood vessel density and increased apoptosis in tumor tissues, which suggests that NK4 suppressed tumor growth as an angiogenesis inhibitor. Metastasis of B16F10 melanoma and Lewis lung carcinoma cells to the lung was potently inhibited by systemic Ad.NK4-administration. Our results demonstrated that the adenovirus-mediated induction of high levels of circulating NK4 significantly inhibited in vivo tumor growth and distant metastasis without obvious side effects. NK4 gene therapy is thus a safe and promising strategy for the treatment of cancer patients, and further validation in clinical trials is needed. (Cancer Sci 2009; 100: 1351-1358)

DOI： 10.1111/j.1349-7006.2009.01184.x

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1093/ndt/gfp065

Language：English   Publishing type：Research paper (scientific journal)  

Background & Aims: Hartnup amino acid transporter B(0)AT1 (SLC6A19) is the major luminal sodium-dependent neutral amino acid transporter of small intestine and kidney proximal tubule. The expression of B(0)AT1 in kidney was recently shown to depend on its association with collectrin (Tmem27), a protein homologous to the membrane-anchoring domain of angiotensin-converting enzyme (ACE) 2. Methods: Because collectrin is almost absent from small intestine, we tested the hypothesis that it is ACE2 that interacts with B(0)AT1 in enterocytes. Furthermore, because B(0)AT1 expression depends on an associated protein, we tested the hypothesis that Hartnup-causing B(0)AT1 mutations differentially impact on B(0)AT1 interaction with intestinal and kidney accessory proteins. Results: Immunofluorescence, coimmunoprecipitation, and functional experiments using wild-type and ace2-null mice showed that expression of B(0)AT1 in small intestine critically depends on ACE2. Coexpressing new and previously identified Hartnup, disorder-causing missense mutations of B(0)AT1 with either collectrin or ACE2 in Xenopus laevis oocytes showed that the high-frequency D173N and the newly identified P26SL mutant B(0)AT1 transporters can still be activated by ACE2 but not collectrin coexpression. In contrast, the human A69T and R240Q B(0)AT1 mutants cannot be activated by either of the associated proteins, although they function as wild-type B(0)AT1 when expressed alone. Conclusions: We thus show that ACE2 is necessary for the expression of the Hartnup transporter in intestine and suggest that the differential functional. association of mutant B(0)AT1 transporters with ACE2 and collectrin in intestine and kidney, respectively, participates in the phenotypic heterogeneity of human Hartnup disorder.

DOI： 10.1053/j.gastro.2008.10.055

Language：English  

During several months of 2002, severe acute respiratory syndrome (SARS) caused by SARS-coronavirus (SARS-CoV) spread rapidly from China throughout the world, causing more than 800 deaths due to the development of acute respiratory distress syndrome (ARDS), which is the severe form of acute lung injury (ALI). Interestingly, a novel homologue of angiotensin-converting enzyme, termed angiotensin-converting enzyme 2 (ACE2), has been identified as a receptor for SARS-CoV. Angiotensin-converting enzyme and ACE2 share homology in their catalytic domain and provide different key functions in the renin-angiotensin system (RAS). Angiotensin-converting enzyme cleaves angiotensin I to generate angiotensin II, which is a key effector peptide of the system and exerts multiple biological functions, whereas ACE2 reduces angiotensin II levels. Importantly, our recent studies using ACE2 knockout mice have demonstrated that ACE2 protects murine lungs from ARDS. Furthermore, SARS-CoV infections and the Spike protein of the SARS-CoV reduce ACE2 expression. Notably, injection of SARS-CoV Spike into mice worsens acute lung failure in vivo, which can be attenuated by blocking the renin-angiotensin pathway, suggesting that the activation of the pulmonary RAS influences the pathogenesis of ALI/ARDS and SARS.

DOI： 10.1113/expphysiol.2007.040048

Language：Japanese  

[Lessons from SARS: a new potential therapy for acute respiratory distress syndrome (ARDS) with angiotensin converting enzyme 2 (ACE2)].

Language：Others  

Endogenous apelin maintains heart contractility in aging and pressure overload

Language：English  

Angiotensin-converting enzyme 2 (ACE2), a second angiotensin-converting enzyme (ACE), regulates the renin-angiotensin system by counterbalancing ACE activity. Accumulating evidence in recent years has demonstrated a physiological and pathological role of ACE2 in the cardiovascular systems. Recently, it has been shown that severe acute respiratory syndrome (SARS) coronavirus, the cause of SARS, utilizes ACE2 as an essential receptor for cell fusion and in vivo infections in mice. Intriguingly, ACE2 acts as a protective factor in various experimental models of acute lung failure and, therefore, acts not only as a key determinant for SARS virus entry into cells but also contributes to SARS pathogenesis. Here we review the role of ACE2 in disease pathogenesis, including lung diseases and cardiovascular diseases.

DOI： 10.1007/s00109-006-0094-9

Language：English  

The renin-angiotensin system (RAS) plays a key role in maintaining blood pressure homeostasis, as well as fluid and salt balance. Angiotensin II, a key effector peptide of the system, causes vasoconstriction and exerts multiple biological functions. Angiotensin-converting enzyme (ACE) plays a central role in generating angiotensin II from angiotensin I, and capillary blood vessels in the lung are one of the major sites of ACE expression and angiotensin II production in the human body. The RAS has been implicated in the pathogenesis of pulmonary hypertension and pulmonary fibrosis, both commonly seen in chronic lung diseases such as chronic obstructive lung disease. Recent studies indicate that the RAS also plays a critical role in acute lung diseases, especially acute respiratory distress syndrome (ARDS). ACE2, a close homologue of ACE, functions as a negative regulator of the angiotensin system and was identified as a key receptor for, SARS (severe acute respiratory syndrome) coronavirus infections. In the lung, ACE2 protects against acute lung injury in several animal models of ARDS. Thus, the RAS appears to play a critical role in the pathogenesis of acute lung injury. Indeed, increasing ACE2 activity might be a novel approach for the treatment of acute lung failure in several diseases.

DOI： 10.1016/j.coph.2006.03.001

Language：Others  

HGF/NK4 inhibited VEGF-induced angiogenesis in in vitro cultured endothelial cells and in vivo rabbit model.
Nakabayashi M, Morishita R, Nakagami H, Kuba K, Matsumoto K, Nakamura T, Tano Y, Kaneda Y, Diabetologia, 2003, vol. 46, no. 1, pp. 115-123, 2003

Language：English   Publishing type：Research paper (scientific journal)  

Invasion of various carcinoma cells follows their interaction with stromal cells. Hepatocyte growth factor (HGF), four-kringle-containing growth factor, is a mesenchymal or stromal-derived mediator which affects the growth and the invasiveness of carcinoma cells. We now have evidence that a four-kringle-containing antagonist for HGF, HGF/NK4 inhibits invasion of tumors in vivo, as well as in vitro. HGF/NK4 competitively inhibited the binding of HGF to Met/HGF receptors on GB-d1 human gallbladder carcinoma cells. HGF induced invasion of the cells through Matrigel basement membrane components and into collagen gels, but HGF-induced invasion was inhibited by HGF/NK4. Invasion of GB-d1 cells was induced by co-cultivation with stromal fibroblasts, which mimics tumor-stromal interaction, but it was almost completely suppressed by HGF/NK4. Likewise, invasive growth induced by HGF in collagen gels in GB-d1 cells, HuCC-T1 human cholangiocarcinoma cells, and ME-180 human uterus cervical carcinoma cells was also strongly inhibited by HGF/NK4. When GB-d1 cells were implanted subcutaneously into nude mouse, tumor cells invaded muscular tissue, but the infusion of HGF/NK4 inhibited this invasion. Furthermore, HGF/NK4 increased apoptotic cell death of GB-d1 cells and inhibited tumor growth in vivo. These results indicate that HGF/NK4 may inhibit growth and invasion of carcinoma cells, as mediated by HGF during tumor-stromal interactions. We propose that there is a unique therapeutic potential for HGF/NK4 to prevent tumor invasion and perhaps even metastasis.

DOI： 10.1038/sj.onc.1202231

Language：English  

Language：Japanese  

Language：Japanese  

Language：English  

Language：Japanese  

脂質シグナリングとその破綻がもたらす病態の理解 心肥大におけるホスホイノシタイド代謝酵素Vps34のタンパク質分解機構の役割

Language：English  

Language：English  

Language：English  

Language：English  

Language：English  

Angiotensin-converting enzyme (ACE) and ACE2 are highly homologous metalloproteases that provide essential catalytic functions in the renin-angiotensin system ( RAS). Angiotensin II is one key effector peptide of the RAS, inducing vasoconstriction and exerting multiple biological functions. ACE cleaves angiotensin I to generate angiotensin II, whereas ACE2 reduces angiotensin II levels. Accumulating evidence has demonstrated a physiological and pathological role of ACE2 in the cardiovascular systems. Intriguingly, the SARS coronavirus, the cause of severe acute respiratory syndrome ( SARS), utilizes ACE2 as an essential receptor for cell fusion and in vivo infections. Moreover, recent studies have demonstrated that ACE2 protects murine lungs from acute lung injury as well as SARS-Spike protein-mediated lung injury, suggesting a dual role of ACE2 in SARS infections and protection from ARDS.

DOI： 10.1007/s00018-007-6228-6

Type：Competition, symposium, etc. 

Type：Academic society, research group, etc. 

Type：Academic society, research group, etc. 

Type：Peer review 

Number of peer-reviewed articles in foreign language journals：7