Viral pneumonia is a global health burden with a high mortality rate, especially in the elderly and in patients with underlying diseases. Recent studies have found that myeloid-derived suppressor ...cells (MDSCs) are abundant in these patient groups; however, their roles in the progression of viral pneumonia remain unclear. In this study, we observed a substantial increase in MDSCs in a mouse model of renal ischemia/reperfusion (I/R) injury and in older mice. When intranasal polyinosinic-polycytidylic acid (poly(I:C)) administration was used to mimic viral pneumonia, mice with renal I/R injury exhibited more severe lung inflammation than sham mice challenged with poly(I:C). In addition, MDSC depletion attenuated lung inflammation in mice with I/R injury. Similar results were obtained in older mice compared with those in young mice. Furthermore, adoptive transfer of in
vitro
-differentiated MDSCs exacerbated poly(I:C)-induced lung inflammation. Taken together, these experimental results suggest that the increased proportion of MDSCs in mice with renal I/R injury and in older mice exacerbates poly(I:C)-induced lung inflammation. These findings have important implications for the treatment and prevention of severe lung inflammation caused by viral pneumonia.
Epstein–Barr virus (EBV) is a major cause of immunosuppression-related B-cell lymphomas and Hodgkin lymphoma (HL). In these malignancies, EBV latent membrane protein 1 (LMP1) and LMP2A provide ...infected B cells with surrogate CD40 and B-cell receptor growth and survival signals. To gain insights into their synergistic in vivo roles in germinal center (GC) B cells, from which most EBV-driven lymphomas arise, we generated a mouse model with conditional GC B-cell LMP1 and LMP2A coexpression. LMP1 and LMP2A had limited effects in immunocompetent mice. However, upon T- and NK-cell depletion, LMP1/2A caused massive plasmablast outgrowth, organ damage, and death. RNA-sequencing analyses identified EBV oncoprotein effects on GC B-cell target genes, including up-regulation of multiple proinflammatory chemokines and master regulators of plasma cell differentiation. LMP1/2A coexpression also up-regulated key HL markers, including CD30 and mixed hematopoietic lineage markers. Collectively, our results highlight synergistic EBV membrane oncoprotein effects on GC B cells and provide a model for studies of their roles in immunosuppression-related lymphoproliferative diseases.
Because mammalian cardiomyocytes largely cease to proliferate immediately after birth, the regenerative activity of the heart is limited. To date, much effort has been made to clarify the regulatory ...mechanism of cardiomyocyte proliferation because the amplification of cardiomyocytes could be a promising strategy for heart regenerative therapy. Recently, it was reported that the inhibition of glycogen synthase kinase (GSK)‐3 promotes the proliferation of neonatal rat cardiomyocytes (NRCMs) and human iPS cell‐derived cardiomyocytes (hiPSC‐CMs). Additionally, Yes‐associated protein (YAP) induces cardiomyocyte proliferation. The purpose of this study was to address the importance of YAP activity in cardiomyocyte proliferation induced by GSK‐3 inhibitors (GSK‐3Is) to develop a novel strategy for cardiomyocyte amplification. Immunofluorescent microscopic analysis using an anti‐Ki‐67 antibody demonstrated that the treatment of NRCMs with GSK‐3Is, such as BIO and CHIR99021, increased the ratio of proliferative cardiomyocytes. YAP was localized in the nuclei of more than 95% of cardiomyocytes, either in the presence or absence of GSK‐3Is, indicating that YAP was endogenously activated. GSK‐3Is increased the expression of β‐catenin and promoted its translocation into the nucleus without influencing YAP activity. The knockdown of YAP using siRNA or pharmacological inhibition of YAP using verteporfin or CIL56 dramatically reduced GSK‐3I‐induced cardiomyocyte proliferation without suppressing β‐catenin activation. Interestingly, the inhibition of GSK‐3 also induced the proliferation of hiPSC‐CMs under sparse culture conditions, where YAP was constitutively activated. In contrast, under dense culture conditions, in which YAP activity was suppressed, the proliferative effects of GSK‐3Is on hiPSC‐CMs were not detected. Importantly, the activation of YAP by the knockdown of α‐catenin restored the proproliferative activity of GSK‐3Is. Collectively, YAP activation potentiates the GSK‐3I‐induced proliferation of cardiomyocytes. The blockade of GSK‐3 in combination with YAP activation resulted in remarkable amplification of cardiomyocytes.
GSK‐3 inhibitors (GSK‐3Is) induce cardiomyocyte proliferation via β‐catenin. The activation of YAP potentiates GSK‐3I‐induced proliferation without modulating β‐catenin signaling. The blockade of GSK‐3 in combination with YAP activation could be a promising strategy for the amplification of cardiomyocytes.
There is an urgent need to develop novel drugs to reduce the mortality from severe infectious diseases with the emergence of new pathogens, including Coronavirus disease 2019 (COVID-19). Although ...current drugs effectively suppress the proliferation of pathogens, immune cell activation, and inflammatory cytokine functions, they cannot completely reduce mortality from severe infections and sepsis. In this study, we focused on the endothelial cell-specific protein, Roundabout 4 (Robo4), which suppresses vascular permeability by stabilizing endothelial cells, and investigated whether enhanced Robo4 expression could be a novel therapeutic strategy against severe infectious diseases. Endothelial-specific overexpression of Robo4 suppresses vascular permeability and reduces mortality in lipopolysaccharide (LPS)-treated mice. Screening of small molecules that regulate Robo4 expression and subsequent analysis revealed that two competitive small mothers against decapentaplegic (SMAD) signaling pathways, activin receptor-like kinase 5 (ALK5)-SMAD2/3 and ALK1-SMAD1/5, positively and negatively regulate Robo4 expression, respectively. An ALK1 inhibitor was found to increase Robo4 expression in mouse lungs, suppress vascular permeability, prevent extravasation of melanoma cells, and decrease mortality in LPS-treated mice. The inhibitor suppressed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-induced endothelial barrier disruption and decreased mortality in mice infected with SARS-CoV-2. These results indicate that enhancing Robo4 expression is an efficient strategy to suppress vascular permeability and mortality in severe infectious diseases, including COVID-19, and that small molecules that upregulate Robo4 can be potential therapeutic agents against these diseases.
A systemic inflammatory response leads to widespread organ dysfunction, such as kidney dysfunction. Plasminogen activator inhibitor-1 (PAI-1) is involved in the pathogenesis of inflammatory kidney ...injury; however, the regulatory mechanism of PAI-1 in injured kidneys remains unclear. PAI-1 is induced by interleukin (IL)-6 in patients with sepsis. In addition, the stabilization of IL-6 is regulated by the adenine–thymine-rich interactive domain-containing protein 5a (Arid5a). Therefore, the aim of the present study was to examine the involvement of Arid5a/IL-6/PAI-1 signaling in lipopolysaccharide (LPS)-induced inflammatory kidney injury. LPS treatment to C57BL/6J mice upregulated Pai-1 mRNA in the kidneys. Enzyme-linked immunosorbent assay (ELISA) revealed that PAI-1 expression was induced in the culture supernatants of LPS-treated human umbilical vein endothelial cells, but not in those of LPS-treated human kidney 2 (HK-2) cells, a tubular cell line. Combined with single-cell analysis, endothelial cells were found to be responsible for PAI-1 elevation in LPS-treated kidneys. Administration of TM5441, a PAI-1 inhibitor, reduced the urinary albumin/creatinine ratio, concomitant with downregulation of Il-6 and Arid5a mRNA expressions. IL-6 treatment in LPS model mice further upregulated Pai-1 mRNA expression compared with LPS alone, accompanied by renal impairment. Furthermore, the expression of Il-6 and Pai-1 mRNA was lower in Arid5a knockout mice than in wild-type mice after LPS treatment. Taken together, the vicious cycle of Arid5a/IL-6/PAI-1 signaling is involved in LPS-induced kidney injury.
Prevention of kidney fibrosis is an essential requisite for effective therapy in preventing chronic kidney disease (CKD). Here, we identify Old astrocyte specifically induced substance (OASIS)/cAMP ...responsive element‐binding protein 3‐like 1 (CREB3l1), a CREB/ATF family transcription factor, as a candidate profibrotic gene that drives the final common pathological step along the fibrotic pathway in CKD. Although microarray data from diseased patient kidneys and fibrotic mouse model kidneys both exhibit OASIS/Creb3l1 upregulation, the pathophysiological roles of OASIS in CKD remains unknown. Immunohistochemistry revealed that OASIS protein was overexpressed in human fibrotic kidney compared with normal kidney. Moreover, OASIS was upregulated in murine fibrotic kidneys, following unilateral ureteral obstruction (UUO), resulting in an increase in the number of OASIS‐expressing pathological myofibroblasts. In vitro assays revealed exogenous TGF‐β1 increased OASIS expression coincident with fibroblast‐to‐myofibroblast transition and OASIS contributed to TGF‐β1–mediated myofibroblast migration and increased proliferation. Significantly, in vivo kidney fibrosis induced via UUO or ischemia/reperfusion injury was ameliorated by systemic genetic knockout of OASIS, accompanied by reduced myofibroblast proliferation. Microarrays revealed that the transmembrane glycoprotein Bone marrow stromal antigen 2 (Bst2) expression was reduced in OASIS knockout myofibroblasts. Interestingly, a systemic anti‐Bst2 blocking antibody approach attenuated kidney fibrosis in normal mice but not in OASIS knockout mice after UUO, signifying Bst2 functions downstream of OASIS. Finally, myofibroblast‐restricted OASIS conditional knockouts resulted in resistance to kidney fibrosis. Taken together, OASIS in myofibroblasts promotes kidney fibrosis, at least in part, via increased Bst2 expression. Thus, we have identified and demonstrated that OASIS signaling is a novel regulator of kidney fibrosis.
Glycoprotein 130 is the common receptor subunit for the interleukin (IL)-6 cytokine family. Previously, we reported that pretreatment of IL-11, an IL-6 family cytokine, activates the glycoprotein 130 ...signaling pathway in cardiomyocytes and prevents ischemia/reperfusion injury in vivo; however, its long-term effects on cardiac remodeling after myocardial infarction (MI) remain to be elucidated.
MI was generated by ligating the left coronary artery in C57BL/6 mice. Real-time reverse transcription polymerase chain reaction analyses showed that IL-11 mRNA was remarkably upregulated in the hearts exposed to MI. Intravenous injection of IL-11 activated signal transducer and activator of transcription 3 (STAT3), a downstream signaling molecule of glycoprotein 130, in cardiomyocytes in vivo, suggesting that cardiac myocytes are target cells of IL-11 in the hearts. Twenty-four hours after coronary ligation, IL-11 was administered intravenously, followed by consecutive administration every 24 hours for 4 days. IL-11 treatment reduced fibrosis area 14 days after MI, attenuating cardiac dysfunction. Consistent with a previous report that STAT3 exhibits antiapoptotic and angiogenic activity in the heart, IL-11 treatment prevented apoptotic cell death of the bordering myocardium adjacent to the infarct zone and increased capillary density at the border zone. Importantly, cardiac-specific ablation of STAT3 abrogated IL-11-mediated attenuation of fibrosis and was associated with left ventricular enlargement. Moreover, with the use of cardiac-specific transgenic mice expressing constitutively active STAT3, cardiac STAT3 activation was shown to be sufficient to prevent adverse cardiac remodeling.
IL-11 attenuated cardiac fibrosis after MI through STAT3. Activation of the IL-11/glycoprotein 130/STAT3 axis may be a novel therapeutic strategy against cardiovascular diseases.
Abstract
Runt-related transcription factor 2 (Runx2), a regulator of osteoblast differentiation, is pathologically involved in vascular calcification; however, the significance of Runx2 in cardiac ...homeostasis remains unclear. Here, we investigated the roles of Runx2 in cardiac remodeling after myocardial infarction (MI). The expression of Runx2 mRNA and protein was upregulated in murine hearts after MI. Runx2 was expressed in heart-infiltrating myeloid cells, especially in macrophages, at the border zone of post-infarct myocardium. To analyze the biological functions of Runx2 in cardiac remodeling, myeloid cell-specific
Runx2
deficient (CKO) mice were exposed to MI. After MI, ventricular weight/tibia length ratio was increased in CKO mice, concomitant with severe cardiac dysfunction. Cardiac fibrosis was exacerbated in CKO mice, consistent with the upregulation of collagen 1a1 expression. Mechanistically, immunohistochemical analysis using anti-CD31 antibody showed that capillary density was decreased in CKO mice. Additionally, conditioned culture media of myeloid cells from Runx2 deficient mice exposed to MI induced the tube formation of vascular endothelial cells to a lesser extent than those from control mice. RNA-sequence showed that the expression of pro-angiogenic or anti-angiogenic factors was altered in macrophages from Runx2-deficient mice. Collectively,
Runx2
+
myeloid cells infiltrate into post-infarct myocardium and prevent adverse cardiac remodeling, at least partially, by regulating endothelial cell function.
Excessive levels of reactive oxygen species (ROS) and impaired Ca2+ homeostasis play central roles in the development of multiple cardiac pathologies, including cell death during ischemia-reperfusion ...(I/R) injury. In several organs, treatment with 2-aminoethoxydiphenyl borate (2-APB) was shown to have protective effects, generally believed to be due to Ca2+ channel inhibition. However, the mechanism of 2-APB-induced cardioprotection has not been fully investigated. Herein we investigated the protective effects of 2-APB treatment against cardiac pathogenesis and deciphered the underlying mechanisms. In neonatal rat cardiomyocytes, treatment with 2-APB was shown to prevent hydrogen peroxide (H2O2) -induced cell death by inhibiting the increase in intracellular Ca2+ levels. However, no 2-APB-sensitive channel blocker inhibited H2O2-induced cell death and a direct reaction between 2-APB and H2O2 was detected by 1H-NMR, suggesting that 2-APB chemically scavenges extracellular ROS and provides cytoprotection. In a mouse I/R model, treatment with 2-APB led to a considerable reduction in the infarct size after I/R, which was accompanied by the reduction in ROS levels and neutrophil infiltration, indicating that the anti-oxidative properties of 2-APB plays an important role in the prevention of I/R injury in vivo as well. Taken together, present results indicate that 2-APB treatment induces cardioprotection and prevents ROS-induced cardiomyocyte death, at least partially, by the direct scavenging of extracellular ROS. Therefore, administration of 2-APB may represent a promising therapeutic strategy for the treatment of ROS-related cardiac pathology including I/R injury.
Mammalian cardiomyocytes substantially lose proliferative capacity immediately after birth, limiting adult heart regeneration after injury. However, clinical myocarditis appears to be self-limiting ...with tissue-reparative properties. Here, we investigated the molecular mechanisms underlying the recovery from myocarditis with regard to cardiomyocyte proliferation using an experimental autoimmune myocarditis (EAM) model. Three weeks after EAM induction (EAM3w), cardiac tissue displayed infiltration of inflammatory cells with cardiomyocyte apoptosis. However, by EAM5w, the myocardial damage was remarkably attenuated, associated with an increase in cardiomyocytes that were positively stained with cell cycle markers at EAM3w. Cardiomyocyte fate mapping study revealed that the proliferating cardiomyocytes primarily derived from pre-existing cardiomyocytes. Signal transducer and activator of transcription 3 (STAT3) was robustly activated in cardiomyocytes during inflammation, accompanied by induction of interleukin-6 family cytokines. Cardiomyocyte-specific ablation of STAT3 gene suppressed the frequency of cycling cardiomyocytes in the recovery period without influencing inflammatory status, resulting in impaired tissue repair and cardiac dysfunction. Finally, microarray analysis revealed that the expression of regeneration-related genes, metallothioneins and clusterin, in cardiomyocytes was decreased by STAT3 gene deletion. These data show that adult mammalian cardiomyocytes restore regenerative capacity with cell cycle reentry through STAT3 as the heart recovers from myocarditis-induced cardiac damage.
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