ER-anchored CRTH2 antagonizes collagen biosynthesis and organ fibrosis via binding LARP6.

Excessive deposition of extracellular matrix, mainly collagen protein, is the hallmark of organ fibrosis. The molecular mechanisms regulating fibrotic protein biosynthesis are unclear. Here, we find that chemoattractant receptor homologous molecule expressed on TH2 cells (CRTH2), a plasma membrane receptor for prostaglandin D2, is trafficked to the endoplasmic reticulum (ER) membrane in fibroblasts in a caveolin-1-dependent manner. ER-anchored CRTH2 binds the collagen mRNA recognition motif of La ribonucleoprotein domain family member 6 (LARP6) and promotes the degradation of collagen mRNA in these cells. In line, CRTH2 deficiency increases collagen biosynthesis in fibroblasts and exacerbates injury-induced organ fibrosis in mice, which can be rescued by LARP6 depletion. Administration of CRTH2 N-terminal peptide reduces collagen production by binding to LARP6. Similar to CRTH2, bumetanide binds the LARP6 mRNA recognition motif, suppresses collagen biosynthesis, and alleviates bleomycin-triggered pulmonary fibrosis in vivo. These findings reveal a novel anti-fibrotic function of CRTH2 in the ER membrane via the interaction with LARP6, which may represent a therapeutic target for fibrotic diseases.

PGD2/DP1 axis promotes liver regeneration by secreting Wnt2 in Kupffer cells in mice.

BACKGROUND AND AIMS: The liver possesses a remarkable regenerative capacity in response to injuries or viral infections. Various growth factors and cytokines are involved in regulating liver regeneration. Prostaglandin (PG) D2, a pro-resolution lipid mediator, is the most abundant hepatic prostanoid. However, the role of PGD2 in the injury-induced liver regeneration remains unclear. APPROACH AND RESULTS: Two-thirds partial hepatectomy (70% PH), massive hepatectomy (85% resection), and carbon tetrachloride-induced chronic injury were performed in mice to study the mechanisms of live regeneration. Hepatic PGD2 production was elevated in mice after PH. Global deletion of D prostanoid receptor (DP) 1, but not DP2, slowed PH-induced liver regeneration in mice, as evidenced by lower liver weight to body weight ratio, less Ki67+ hepatocyte proliferation, and G2/M phase hepatocytes. Additionally, DP1 deficiency specifically in resident Kupffer cells (KCs), and not in endothelial cells or hepatic stellate cells, retarded liver regeneration in mice post-PH. Conversely, the overexpression of exogenous DP1 in KCs accelerated liver regeneration in mice. Mechanistically, DP1 activation promoted Wnt2 transcription in a PKA/CREB-dependent manner in resident KCs and mediated hepatocyte proliferation through Frizzled8/ß-catenin signaling. Adeno-associated virus vector serotype 8 (AAV8)-mediated Frizzled8 knockdown in hepatocytes attenuated accelerated liver regeneration in KC-DP1 transgenic mice post-PH. Treatment with the DP1 receptor agonist BW245C promotes PH-induced liver regeneration in mice. CONCLUSIONS: DP1 activation mediates crosstalk between KCs and hepatocytes through Wnt2, and facilitates liver regeneration. Hence, DP1 may serve as a novel therapeutic target in acute and chronic liver diseases.

HMGB2 Release Promotes Pulmonary Hypertension and Predicts Severity and Mortality of Patients With Pulmonary Arterial Hypertension.

BACKGROUND: Pulmonary hypertension (PH) is a progressive and life-threatening disease characterized by pulmonary vascular remodeling, which involves aberrant proliferation and apoptosis resistance of the pulmonary arterial smooth muscle cells (PASMCs), resembling the hallmark characteristics of cancer. In cancer, the HMGB2 (high-mobility group box 2) protein promotes the pro-proliferative/antiapoptotic phenotype. However, the function of HMGB2 in PH remains uninvestigated. METHODS: Smooth muscle cell (SMC)-specific HMGB2 knockout or HMGB2-OE (HMGB2 overexpression) mice and HMGB2 silenced rats were used to establish hypoxia+Su5416 (HySu)-induced PH mouse and monocrotaline-induced PH rat models, respectively. The effects of HMGB2 and its underlying mechanisms were subsequently elucidated using RNA-sequencing and cellular and molecular biology analyses. Serum HMGB2 levels were measured in the controls and patients with pulmonary arterial (PA) hypertension. RESULTS: HMGB2 expression was markedly increased in the PAs of patients with PA hypertension and PH rodent models and was predominantly localized in PASMCs. SMC-specific HMGB2 deficiency or silencing attenuated PH development and pulmonary vascular remodeling in hypoxia+Su5416-induced mice and monocrotaline-treated rats. SMC-specific HMGB2 overexpression aggravated hypoxia+Su5416-induced PH. HMGB2 knockdown inhibited PASMC proliferation in vitro in response to PDGF-BB (platelet-derived growth factor-BB). In contrast, HMGB2 protein stimulation caused the hyperproliferation of PASMCs. In addition, HMGB2 promoted PASMC proliferation and the development of PH by RAGE (receptor for advanced glycation end products)/FAK (focal adhesion kinase)-mediated Hippo/YAP (yes-associated protein) signaling suppression. Serum HMGB2 levels were significantly increased in patients with PA hypertension, and they correlated with disease severity, predicting worse survival. CONCLUSIONS: Our findings indicate that targeting HMGB2 might be a novel therapeutic strategy for treating PH. Serum HMGB2 levels could serve as a novel biomarker for diagnosing PA hypertension and determining its prognosis.

Contribution of platelets to disruption of the blood-brain barrier during arterial baroreflex dysfunction.

BACKGROUND: Arterial baroreflex dysfunction, like many other central nervous system disorders, involves disruption of the blood-brain barrier, but what causes such disruption in ABR dysfunction is unclear. Here we explored the potential role of platelets in this disruption. METHODS: ABR dysfunction was induced in rats using sinoaortic denervation, and the effects on integrity of the blood-brain barrier were explored based on leakage of Evans blue or FITC-dextran, while the effects on expression of CD40L in platelets and of key proteins in microvascular endothelial cells were explored using immunohistochemistry, western blotting and enzyme-linked immunosorbent assay. Similar experiments were carried out in rat brain microvascular endothelial cell line, which we exposed to platelets taken from rats with ABR dysfunction. RESULTS: Sinoaortic denervation permeabilized the blood-brain barrier and downregulated zonula occludens-1 and occludin in rat brain, while upregulating expression of CD40L on the surface of platelets and stimulating platelet aggregation. Similar effects of permeabilization and downregulation were observed in healthy rats that received platelets from animals with ABR dysfunction, and in rat brain microvascular endothelial cells, but only in the presence of lipopolysaccharide. These effects were associated with activation of NF-κB signaling and upregulation of matrix metalloprotease-9. These effects of platelets from animals with ABR dysfunction were partially blocked by neutralizing antibody against CD40L or the platelet inhibitor clopidogrel. CONCLUSION: During ABR dysfunction, platelets may disrupt the blood-brain barrier when CD40L on their surface activates NF-kB signaling within cerebral microvascular endothelial cells, leading to upregulation of matrix metalloprotease-9. Our findings imply that targeting CD40L may be effective against cerebral diseases involving ABR dysfunction.

Arterial Baroreflex Dysfunction Promotes Neuroinflammation by Activating the Platelet CD40L/Nuclear Factor Kappa B Signaling Pathway in Microglia and Astrocytes.

Arterial baroreflex (ABR) dysfunction has previously been associated with neuroinflammation, the most common pathological feature of neurological disorders. However, the mechanisms mediating ABR dysfunction-induced neuroinflammation are not fully understood. In the present study, we investigated the role of platelet CD40 ligand (CD40L) in neuroinflammation in an in vivo model of ABR dysfunction, and microglia and astrocyte activation in vitro. ABR dysfunction was induced in Sprague‒Dawley rats by sinoaortic denervation (SAD). We used ELSA and immunofluorescence to assess the effect of platelet CD40L on glial cell polarization and the secretion of inflammatory factors. By flow cytometry, we found that rats subjected to SAD showed a high level of platelet microaggregation and upregulation of CD40L on the platelet surface. The promotion of platelet invasion and accumulation was also observed in the brain tissues of rats subjected to SAD. In the animal model and cultured N9 microglia/C6 astrocytoma cells, platelet CD40L overexpression promoted neuroinflammation and activated M1 microglia, A1 astrocytes, and the nuclear factor kappa B (NFκB) signaling pathway. These effects were partially blocked by inhibiting platelet activity with clopidogrel or inhibiting CD40L-mediated signaling. Our results suggest that during ABR dysfunction, CD40L signaling in platelets converts microglia to the M1 phenotype and astrocytes to the A1 phenotype, activating NFκB and resulting in neuroinflammation. Thus, our study provides a novel understanding of the pathogenesis of ABR dysfunction-induced neuroinflammation and indicates that targeting platelet CD40L is beneficial for treating central nervous system (CNS) disorders associated with ABR dysfunction.

Prostaglandin D2 signaling and cardiovascular homeostasis.

Cardiovascular diseases are the leading cause of death worldwide. A chronic inflammatory response is a common pathological alteration in diverse cardiovascular diseases. Prostaglandin (PG) D2, a key lipid mediator derived from arachidonic acid metabolism, promotes resolution of inflammation and regulated T cell function through its receptors. Accumulated evidence has shown that dysregulated PGD2 signaling is involved in the pathogenesis of cardiovascular diseases, including atherosclerosis, hypertension, pulmonary hypertension, abdominal aortic aneurysm, and myocardial ischemia. Here, we summarized the recent progresses on PGD2 in cardiovascular homeostasis and discussed potential therapeutic translation by targeting PGD2 signaling.

Mechanism and Origins of Enantioselectivity of Cobalt-Catalyzed Intermolecular Hydroarylation/Cyclization of 1,6-Enynes with N-Pyridylindoles.

Density functional theory calculations were performed to investigate the cobalt-catalyzed intermolecular hydroarylation/cyclization of 1,6-enynes with N-pyridylindoles. The computations reveal that the reaction begins with the oxidative cyclization of 1,6-enyne to afford the five-membered cobaltacycle, from which the metal-assisted σ-bond metathesis/C-C reductive elimination led to the final hydroarylation/cyclization product. The initial oxidative cyclization constitutes the rate-determining step of the overall reaction. The steric repulsion and π···π interaction were found to play a crucial role in dictating the experimentally observed enantioselectivity.

DP1 Activation Reverses Age-Related Hypertension Via NEDD4L-Mediated T-Bet Degradation in T Cells.

BACKGROUND: Blood pressure often rises with aging, but exact mechanisms are still not completely understood. With aging, the level of proinflammatory cytokines increases in T lymphocytes. Prostaglandin D2, a proresolution mediator, suppresses Type 1 T helper (Th1) cytokines through D-prostanoid receptor 1 (DP1). In this study, we aimed to investigate the role of the prostaglandin D2/DP1 axis in T cells on age-related hypertension. METHODS: To clarify the physiological and pathophysiological roles of DP1 in T cells with aging, peripheral blood samples were collected from young and older male participants, and CD4+ T cells were sorted for gene expression, prostaglandin production, and Western blot assays. Mice blood pressure was quantified by invasive telemetric monitor. RESULTS: The prostaglandin D2/DP1 axis was downregulated in CD4+ T cells from older humans and aged mice. DP1 deletion in CD4+ T cells augmented age-related hypertension in aged male mice by enhancing Th1 cytokine secretion, vascular remodeling, CD4+ T cells infiltration, and superoxide production in vasculature and kidneys. Conversely, forced expression of exogenous DP1 in T cells retarded age-associated hypertension in mice by reducing Th1 cytokine secretion. Tumor necrosis factor α neutralization or interferon γ deletion ameliorated the age-related hypertension in DP1 deletion in CD4+ T cells mice. Mechanistically, DP1 inhibited Th1 activity via the PKA (protein kinase A)/p-Sp1 (phosphorylated specificity protein 1)/neural precursor cell expressed developmentally downregulated 4-like (NEDD4L) pathway-mediated T-box-expressed-in-T-cells (T-bet) ubiquitination. T-bet deletion or forced NEDD4L expression in CD4+ T cells attenuated age-related hypertension in CD4+ T cell-specific DP1-deficient mice. DP1 receptor activation by BW245C prevented age-associated blood pressure elevation and reduced vascular/renal superoxide production in male mice. CONCLUSIONS: The prostaglandin D2/DP1 axis suppresses age-related Th1 activation and subsequent hypertensive response in male mice through increase of NEDD4L-mediated T-bet degradation by ubiquitination. Therefore, the T cell DP1 receptor may be an attractive therapeutic target for age-related hypertension.

Loss of DP1 Aggravates Vascular Remodeling in Pulmonary Arterial Hypertension via mTORC1 Signaling.

Rationale: Vascular remodeling, including smooth muscle cell hypertrophy and proliferation, is the key pathological feature of pulmonary arterial hypertension (PAH). Prostaglandin I2 analogs (beraprost, iloprost, and treprostinil) are effective in the treatment of PAH. Of note, the clinically favorable effects of treprostinil in severe PAH may be attributable to concomitant activation of DP1 (D prostanoid receptor subtype 1).Objectives: To study the role of DP1 in the progression of PAH and its underlying mechanism.Methods: DP1 levels were examined in pulmonary arteries of patients and animals with PAH. Multiple genetic and pharmacologic approaches were used to investigate DP1-mediated signaling in PAH.Measurements and Main Results: DP1 expression was downregulated in hypoxia-treated pulmonary artery smooth muscle cells and in pulmonary arteries from rodent PAH models and patients with idiopathic PAH. DP1 deletion exacerbated pulmonary artery remodeling in hypoxia-induced PAH, whereas pharmacological activation or forced expression of the DP1 receptor had the opposite effect in different rodent models. DP1 deficiency promoted pulmonary artery smooth muscle cell hypertrophy and proliferation in response to hypoxia via induction of mTORC1 (mammalian target of rapamycin complex 1) activity. Rapamycin, an inhibitor of mTORC1, alleviated the hypoxia-induced exacerbation of PAH in DP1-knockout mice. DP1 activation facilitated raptor dissociation from mTORC1 and suppressed mTORC1 activity through PKA (protein kinase A)-dependent phosphorylation of raptor at Ser791. Moreover, treprostinil treatment blocked the progression of hypoxia-induced PAH in mice in part by targeting the DP1 receptor.Conclusions: DP1 activation attenuates hypoxia-induced pulmonary artery remodeling and PAH through PKA-mediated dissociation of raptor from mTORC1. These results suggest that the DP1 receptor may serve as a therapeutic target for the management of PAH.

T-Cell Mineralocorticoid Receptor Controls Blood Pressure by Regulating Interferon-Gamma.

RATIONALE: Hypertension remains to be a global public health burden and demands novel intervention strategies such as targeting T cells and T-cell-derived cytokines. Mineralocorticoid receptor (MR) antagonists have been clinically used to treat hypertension. However, the function of T-cell MR in blood pressure (BP) regulation has not been elucidated. OBJECTIVE: We aim to determine the role of T-cell MR in BP regulation and to explore the mechanism. METHODS AND RESULTS: Using T-cell MR knockout mouse in combination with angiotensin II-induced hypertensive mouse model, we demonstrated that MR deficiency in T cells strikingly decreased both systolic and diastolic BP and attenuated renal and vascular damage. Flow cytometric analysis showed that T-cell MR knockout mitigated angiotensin II-induced accumulation of interferon-gamma (IFN-γ)-producing T cells, particularly CD8+ population, in both kidneys and aortas. Similarly, eplerenone attenuated angiotensin II-induced elevation of BP and accumulation of IFN-γ-producing T cells in wild-type mice. In cultured CD8+ T cells, T-cell MR knockout suppressed IFN-γ expression whereas T-cell MR overexpression and aldosterone both enhanced IFN-γ expression. At the molecular level, MR interacted with NFAT1 (nuclear factor of activated T-cells 1) and activator protein-1 in T cells. Finally, T-cell MR overexpressing mice manifested more elevated BP compared with control mice after angiotensin II infusion and such difference was abolished by IFN-γ-neutralizing antibodies. CONCLUSIONS: MR may interact with NFAT1 and activator protein-1 to control IFN-γ in T cells and to regulate target organ damage and ultimately BP. Targeting MR in T cells specifically may be an effective novel approach for hypertension treatment.

Thromboxane Governs the Differentiation of Adipose-Derived Stromal Cells Toward Endothelial Cells In Vitro and In Vivo.

RATIONALE: Autologous adipose-derived stromal cells (ASCs) offer great promise as angiogenic cell therapy for ischemic diseases. Because of their limited self-renewal capacity and pluripotentiality, the therapeutic efficacy of ASCs is still relatively low. Thromboxane has been shown to play an important role in the maintenance of vascular homeostasis. However, little is known about the effects of thromboxane on ASC-mediated angiogenesis. OBJECTIVE: To explore the role of the thromboxane-prostanoid receptor (TP) in mediating the angiogenic capacity of ASCs in vivo. METHODS AND RESULTS: ASCs were prepared from mouse epididymal fat pads and induced to differentiate into endothelial cells (ECs) by vascular endothelial growth factor. Cyclooxygenase-2 expression, thromboxane production, and TP expression were upregulated in ASCs on vascular endothelial growth factor treatment. Genetic deletion or pharmacological inhibition of TP in mouse or human ASCs accelerated EC differentiation and increased tube formation in vitro, enhanced angiogenesis in in vivo Matrigel plugs and ischemic mouse hindlimbs. TP deficiency resulted in a significant cellular accumulation of ß-catenin by suppression of calpain-mediated degradation in ASCs. Knockdown of ß-catenin completely abrogated the enhanced EC differentiation of TP-deficient ASCs, whereas inhibition of calpain reversed the suppressed angiogenic capacity of TP re-expressed ASCs. Moreover, TP was coupled with Gαq to induce calpain-mediated suppression of ß-catenin signaling through calcium influx in ASCs. CONCLUSION: Thromboxane restrained EC differentiation of ASCs through TP-mediated repression of the calpain-dependent ß-catenin signaling pathway. These results indicate that TP inhibition could be a promising strategy for therapy utilizing ASCs in the treatment of ischemic diseases.

2, 3, 7, 8-Tetrachlorodibenzo-p-dioxin promotes endothelial cell apoptosis through activation of EP3/p38MAPK/Bcl-2 pathway.

Endothelial injury or dysfunction is an early event in the pathogenesis of atherosclerosis. Epidemiological and animal studies have shown that 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD) exposure increases morbidity and mortality from chronic cardiovascular diseases, including atherosclerosis. However, whether or how TCDD exposure causes endothelial injury or dysfunction remains largely unknown. Cultured human umbilical vein endothelial cells (HUVECs) were exposed to different doses of TCDD, and cell apoptosis was examined. We found that TCDD treatment increased caspase 3 activity and apoptosis in HUVECs in a dose-dependent manner,at doses from 10 to 40 nM. TCDD increased cyclooxygenase enzymes (COX)-2 expression and its downstream prostaglandin (PG) production (mainly PGE2 and 6-keto-PGF1α ) in HUVECs. Interestingly, inhibition of COX-2, but not COX-1, markedly attenuated TCDD-triggered apoptosis in HUVECs. Pharmacological inhibition or gene silencing of the PGE2 receptor subtype 3 (EP3) suppressed the augmented apoptosis in TCDD-treated HUVECs. Activation of the EP3 receptor enhanced p38 MAPK phosphorylation and decreased Bcl-2 expression following TCDD treatment. Both p38 MAPK suppression and Bcl-2 overexpression attenuated the apoptosis in TCDD-treated HUVECs. TCDD increased EP3-dependent Rho activity and subsequently promoted p38MAPK/Bcl-2 pathway-mediated apoptosis in HUVECs. In addition, TCDD promoted apoptosis in vascular endothelium and delayed re-endothelialization after femoral artery injury in wild-type (WT) mice, but not in EP3-/- mice. In summary, TCDD promotes endothelial apoptosis through the COX-2/PGE2 /EP3/p38MAPK/Bcl-2 pathway. Given the cardiovascular hazard of a COX-2 inhibitor, our findings indicate that the EP3 receptor and its downstream pathways may be potential targets for prevention of TCDD-associated cardiovascular diseases.

Niacin Promotes Cardiac Healing after Myocardial Infarction through Activation of the Myeloid Prostaglandin D2 Receptor Subtype 1.

Niacin is a well established drug used to lower cholesterol and prevent cardiovascular disease events. However, niacin also causes cutaneous flushing side effects due to release of the proresolution mediator prostaglandin D2 (PGD2). Recent randomized clinical trials have demonstrated that addition of niacin with laropiprant [a PGD2 receptor subtype 1 (DP1) blocker] to statin-based therapies does not significantly decrease the risk of cardiovascular disease events, but increases the risk of serious adverse events. Here, we tested whether, and how, niacin beneficial effects on myocardial ischemia require the activation of the PGD2/DP1 axis. Myocardial infarction (MI) was reproduced by ligation of the left anterior descending branch of the coronary artery in mice. We found that niacin increased PGD2 release in macrophages and shifted macrophages to M2 polarization both in vitro and in vivo by activation of DP1 and accelerated inflammation resolution in zymosan-induced peritonitis in mice. Moreover, niacin treatment facilitated wound healing and improved cardiac function after MI through DP1-mediated M2 bias and timely resolution of inflammation in infarcted hearts. In addition, we found that niacin intake also stimulated M2 polarization of peripheral monocytes in humans. Collectively, niacin promoted cardiac functional recovery after ischemic myocardial infarction through DP1-mediated M2 polarization and timely resolution of inflammation in hearts. These results indicated that DP1 inhibition may attenuate the cardiovascular benefits of niacin.

Rare SNP rs12731181 in the miR-590-3p Target Site of the Prostaglandin F2α Receptor Gene Confers Risk for Essential Hypertension in the Han Chinese Population.

OBJECTIVE: To investigate whether rs12731181 (A→G) interrupted miR-590-3p-mediated suppression of the prostaglandin F2α receptor (FP) and whether it is associated with essential hypertension in the Chinese population. APPROACH AND RESULTS: We found that miR-590-3p regulates human FP gene expression by binding to its 3'-untranslated region. rs12731181 (A→G) altered the binding affinity between miR-590-3p and its FP 3'-untranslated region target, thus reducing the suppression of FP expression, which, in turn, enhanced FP receptor-mediated contractility of vascular smooth muscle cells. Overexpression of FP augmented vascular tone and elevated blood pressure in mice. An association study was performed to analyze the relationship between the FP gene and essential hypertension in the Han Chinese population. The results indicated that the rs12731181 G allele was associated with susceptibility to essential hypertension. Carriers of the AG genotype exhibited significantly higher blood pressure than those of the AA genotype. FP gene expression was significantly higher in human peripheral leukocytes from individuals with the AG genotype than that in leukocytes from individuals with the AA genotype. CONCLUSIONS: rs12731181 in the seed region of the miR-590-3p target site is associated with increased risk of essential hypertension and represents a new paradigm for FP involvement in blood pressure regulation.

Myeloid-derived suppressor cell function is diminished in aspirin-triggered allergic airway hyperresponsiveness in mice.

BACKGROUND: Myeloid-derived suppressor cells (MDSCs) have recently been implicated in the pathogenesis of asthma, but their regulation in patients with aspirin-intolerant asthma (AIA) remains unclear. OBJECTIVE: We sought to characterize MDSC accumulation and pathogenic functions in allergic airway inflammation mediated by COX-1 deficiency or aspirin treatment in mice. METHODS: Allergic airway inflammation was induced in mice by means of ovalbumin challenge. The distribution and function of MDSCs in mice were analyzed by using flow cytometry and pharmacologic/gene manipulation approaches. RESULTS: CD11b(+)Gr1(high)Ly6G(+)Ly6C(int) MDSCs (polymorphonuclear MDSCs [PMN-MDSCs]) recruited to the lungs are negatively correlated with airway inflammation in allergen-challenged mice. Aspirin-treated and COX-1 knockout (KO) mice showed significantly lower accumulation of PMN-MDSCs in the inflamed lung and immune organs accompanied by increased TH2 airway responses. The TH2-suppressive function of PMN-MDSCs was notably impaired by COX-1 deletion or inhibition, predominantly through downregulation of arginase-1. COX-1-derived prostaglandin E2 promoted PMN-MDSC generation in bone marrow through E prostanoid 2 and 4 receptors (EP2 and EP4), whereas the impaired arginase-1 expression in PMN-MDSCs in COX-1 KO mice was mediated by dysregulation of the prostaglandin E2/EP4/cyclic AMP/protein kinase A pathway. EP4 agonist administration alleviated allergy-induced airway hyperresponsiveness in COX-1 KO mice. Moreover, the immunosuppressive function of PMN-MDSCs from patients with AIA was dramatically decreased compared with that from patients with aspirin-tolerant asthma. CONCLUSION: The immunosuppressive activity of PMN-MDSCs was diminished in both allergen-challenged COX-1 KO mice and patients with AIA, probably through an EP4-mediated signaling pathway, indicating that activation of PMN-MDSCs might be a promising therapeutic strategy for asthma, particularly AIA.

Tumor Acidic Microenvironment-Responsive Promodulator Iron Oxide Nanoparticles for Photothermal-Enhanced Chemodynamic Immunotherapy of Cancer.

Cancer nanomedicine combined with immunotherapy has emerged as a promising strategy for the treatment of cancer. However, precise regulation of the activation of antitumor immunity in targeting tissues for safe and effective cancer immunotherapy remains challenging. Herein, we report a tumor acidic microenvironment-responsive promodulator iron oxide nanoparticle (termed as FGR) with pH-activated action for photothermal-enhanced chemodynamic immunotherapy of cancer. FGR is formed via surface-modifying iron oxide nanoparticles with a dextran-conjugated Toll-like receptor agonist (R848) containing an acid-labile bond. In an acidic tumor microenvironment, the acid-responsive bonds are hydrolyzed to trigger the specific release of R848 to promote the maturation of dendritic cells. In addition, iron oxide nanoparticles within FGR exert photothermal and chemodynamic effects under near-infrared laser irradiation to directly kill tumor cells and induce immunogenic cell death. The synergistic effect of the released immunogenic factors and the acid-activated TLR7/8 pathway stimulates the formation of strong antitumor immunity, resulting in increased infiltration of cytotoxic CD8+ T cells into tumor tissues. As a result, FGR achieves acid-responsive on-demand release and activation of modulators in tumor sites and mediates photothermal-enhanced chemodynamic immunotherapy to inhibit the growth and metastasis of melanoma. Therefore, this work proposes a general strategy for designing prodrug nanomedicines to accurately regulate cancer immunotherapy.

Nanopore long-read RNA sequencing reveals functional alternative splicing variants in human vascular smooth muscle cells.

Vascular smooth muscle cells (VSMCs) are the major contributor to vascular repair and remodeling, which showed high level of phenotypic plasticity. Abnormalities in VSMC plasticity can lead to multiple cardiovascular diseases, wherein alternative splicing plays important roles. However, alternative splicing variants in VSMC plasticity are not fully understood. Here we systematically characterized the long-read transcriptome and their dysregulation in  human aortic smooth muscle cells (HASMCs) by employing the Oxford Nanopore Technologies long-read RNA sequencing in HASMCs that are separately treated with platelet-derived growth factor, transforming growth factor, and hsa-miR-221-3P transfection. Our analysis reveals frequent alternative splicing events and thousands of unannotated transcripts generated from alternative splicing. HASMCs treated with different factors exhibit distinct transcriptional reprogramming modulated by alternative splicing. We also found that unannotated transcripts produce different open reading frames compared to the annotated transcripts. Finally, we experimentally validated the unannotated transcript derived from gene CISD1, namely CISD1-u, which plays a role in the phenotypic switch of HASMCs. Our study characterizes the phenotypic modulation of HASMCs from an insight of long-read transcriptome, which would promote the understanding and the manipulation of HASMC plasticity in cardiovascular diseases.

A prodrug hydrogel with tumor microenvironment and near-infrared light dual-responsive action for synergistic cancer immunotherapy.

Immunotherapy has been used for cancer treatment, while it faces the common dilemmas of low therapeutic efficacy and serious immunotoxicity. In this study, we report the construction of a tumor microenvironment and near-infrared (NIR) light dual-responsive prodrug hydrogel for cancer synergistic immunotherapy in a more effective and safe manner. Such prodrug hydrogels were in-situ formed via calcium-induced gelation of alginate solution containing protoporphyrin IX (PpIX)-modified iron oxide (Fe3O4) nanoparticles and programmed death ligand 1 antibody (aPD-L1) prodrug nanoparticles crosslinked by reactive oxygen species (ROS)-responsive linkers. PpIX served as a photosensitizer to produce singlet oxygen (1O2) under NIR laser irradiation for photodynamic therapy (PDT), and Fe3O4 nanoparticles mediated chemodynamic therapy (CDT) to generate hydroxyl radical (·OH) via Fenton reaction in the tumor microenvironment. In view of the cumulative actions of PDT and CDT, amplified ROS was generated to not only induce immunogenic cell death (ICD), but also destroy ROS-responsive linkers to achieve on-demand release of aPD-L1 from prodrug nanoparticles. Boosted antitumor immunity was elicited in tumor-bearing mice due to the aPD-L1-mediated immune checkpoint blocking. As a result, the prodrug hydrogel-based synergistic immunotherapy could almost treat bilateral tumors and prevent lung and liver metastasis using 4T1 tumor mouse models. This study thus offers a dual-responsive prodrug hydrogel platform for precision cancer immunotherapy. STATEMENT OF SIGNIFICANCE: Via calcium-induced gelation of alginate, we constructed a prodrug hydrogel with tumor microenvironment and near-infrared light dual-responsive action for synergistic cancer immunotherapy. Such hydrogels can achieve on-demand release of aPD-L1 upon photoactivation in the tumor microenvironment. Through mediating photodynamic and chemodynamic therapy, the prodrug hydrogels can induce enhanced immunogenic cell death and synergistically improve the efficacy of aPD-L1-mediated immune checkpoint blocking. The prodrug hydrogel-based synergistic therapy almost deracinates the primary and distant tumors, and prevents lung and liver metastasis in tumor mouse models.

Tumor immunosuppressive microenvironment modulating hydrogels for second near-infrared photothermal-immunotherapy of cancer.

Immunotherapy has recently been seen as a hopeful therapeutic device to inhibit tumor growth and metastasis, while the curative efficacy is limited by intrinsic immunosuppressive tumor microenvironment. Herein, we reported a tumor immunosuppressive microenvironment modulating hydrogel (TIMmH) platform to achieve second near-infrared (NIR-II) photothermal therapy (PTT) combined immunotherapy for durable inhibition of breast cancer. This TIMmH platform was synthesized through co-loading of NIR-II photothermal nanoagent and an immunoadjuvant cytosine-phosphateguanosine oligodeoxynucleotides (CpG ODNs) into the alginate hydrogel (ALG). Upon the administration of ALG into the tumor, the TIMmH was in situ formed via the coordination effect with Ca2+, locally encapsulating the semiconducting polymer nanoparticles (SPIIN) and CpG in the colloid, achieving to prolong the accumulation time and prevent the premature damage and release of immunotherapeutic agents. Upon 1064-nm photoirradiation, the TIMmHSD was able to elevate the intratumoral temperature for the ablation of tumors, which could induce the apoptosis of tumor cells and achieve thermal immune activation by regulating of an immunosuppressive microenvironment. The TIMmH-mediated combined treatment effectively suppressed the growths of breast cancers, and even acquired a sustained inhibition of the lung metastasis. This study provides a novel tumor immunosuppressive microenvironment modulating hydrogel platform with NIR-II photoexcited capacity for the safe, effective and durable lung metastasis-inhibiting breast cancer treatment.

Systematic characterization of cancer transcriptome at transcript resolution.

Transcribed RNAs undergo various regulation and modification to become functional transcripts. Notably, cancer transcriptome has not been fully characterized at transcript resolution. Herein, we carry out a reference-based transcript assembly across >1000 cancer cell lines. We identify 498,255 transcripts, approximately half of which are unannotated. Unannotated transcripts are closely associated with cancer-related hallmarks and show clinical significance. We build a high-confidence RNA binding protein (RBP)-transcript regulatory network, wherein most RBPs tend to regulate transcripts involved in cell proliferation. We identify numerous transcripts that are highly associated with anti-cancer drug sensitivity. Furthermore, we establish RBP-transcript-drug axes, wherein PTBP1 is experimentally validated to affect the sensitivity to decitabine by regulating KIAA1522-a6 transcript. Finally, we establish a user-friendly data portal to serve as a valuable resource for understanding cancer transcriptome diversity and its potential clinical utility at transcript level. Our study substantially extends cancer RNA repository and will facilitate anti-cancer drug discovery.