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Background: Adverse environmental exposure during the prenatal period can lead to diseases in the offspring, including hypertension. Whether or not the hypertensive phenotype can be transgenerationally transmitted is not known. Methods: Pregnant Sprague Dawley rats were intraperitoneally injected with lipopolysaccharide (LPS) on gestation days 6, 8, 10, and 12 to generate the prenatal LPS exposure model. Blood pressure was monitored by both telemetry and tail-cuff method. RNA sequencing was performed to analyze transcriptome alteration in the kidney of the third generation. Tempol and spironolactone were used to test the potential preventative and therapeutic effect of targeting reactive oxygen species and mineralocorticoid receptor signaling, respectively. Molecular biological experiments were performed to illustrate the mechanism of epigenetic and transcription regulation. Results: Prenatal LPS exposure can impair the ability to excrete a salt load and induce hypertension from the first to the third generations, with the fourth and fifth generations, inducing salt-sensitive hypertension. Compared with control pups, the transcriptome in the kidney of the hypertensive third-generation prenatal LPS–exposed offspring have upregulation of the Ras-related C3 botulinum toxin substrate 1 ( Rac1 ) gene and activation of mineralocorticoid receptor signaling. Furthermore, we found that LPS exposure during pregnancy triggered oxidative stress that upregulated KDM3B (histone lysine demethylase 3B) in the oocytes of first-generation female rats, leading to an inheritable low level of H3K9me2 (histone H3 lysine 9 dimethylation), resulting in the transgenerational upregulation of Rac1 . Based on these findings, we treated the LPS-exposed pregnant rats with the reactive oxygen species scavenger, tempol, which successfully prevented hypertension in the first-generation offspring and the transgenerational inheritance of hypertension. Conclusions: These findings show that adverse prenatal exposure induces transgenerational hypertension through an epigenetic-regulated mechanism and identify potentially preventive and therapeutic strategies for hypertension.

Activation of the angiotensin II type 2 receptor (AT2R) induces diuresis and natriuresis. Increased expression or/and activity of G-protein-coupled receptor kinase 4 (GRK4) or genetic variants (e.g., GRK4γ142V) cause sodium retention and hypertension. Whether GRK4 plays a role in the regulation of AT2R in the kidney remains unknown. In the present study, we found that spontaneously hypertensive rats (SHRs) had increased AT2R phosphorylation and impaired AT2R-mediated diuretic and natriuretic effects, as compared with normotensive Wistar-Kyoto (WKY) rats. The regulation by GRK4 of renal AT2R phosphorylation and function was studied in human (h) GRK4γ transgenic mice. hGRK4γ142V transgenic mice had increased renal AT2R phosphorylation and impaired AT2R-mediated natriuresis, relative to hGRK4γ wild-type (WT) littermates. These were confirmed in vitro; AT2R phosphorylation was increased and AT2R-mediated inhibition of Na+-K+-ATPase activity was decreased in hGRK4γ142V, relative to hGRK4γ WT-transfected renal proximal tubule (RPT) cells. There was a direct physical interaction between renal GRK4 and AT2R that was increased in SHRs, relative to WKY rats. Ultrasound-targeted microbubble destruction of renal GRK4 decreased the renal AT2R phosphorylation and restored the impaired AT2R-mediated diuresis and natriuresis in SHRs. In vitro studies showed that GRK4 siRNA reduced AT2R phosphorylation and reversed the impaired AT2R-mediated inhibition of Na+-K+-ATPase activity in SHR RPT cells. Our present study shows that GRK4, at least in part, impairs renal AT2R-mediated diuresis and natriuresis by increasing its phosphorylation; inhibition of GRK4 expression and/or activity may be a potential strategy to improve the renal function of AT2R.

The novel coronavirus disease 2019 continues to spread widely, not only causing physical disorders in patients but also impairing mental health, bringing a heavy burden on global public health. This study aimed to evaluate the anxiety and depression status of patients infected with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) omicron variant in Shanghai.This study aimed to retrospectively analyze 2000 patients infected with the SARS-CoV-2 Omicron variant. Data from patients assessed with demographic information, anxiety and depressive symptoms were collected using a questionnaire. Clinical and laboratory data were collected using electronic medical system. Anxiety and depression were assessed using the Self-Rating Anxiety Scale, the Generalized Anxiety Disorder Scale, and the Patient Health Questionnaire. Clinical information and laboratory indicators included age, sex, blood pressure, blood glucose, basic disease, time of diagnosis onset, duration of hospitalization, vaccination status of novel coronavirus disease 2019, and virus-negative conversion time.This study will provide evidence-based suggestions for early psychological intervention in patients infected with the SARS-CoV-2 Omicron Variant.

Aims Exposure to maternal diabetes is associated with increased prevalence of hypertension in the offspring. The mechanisms underlying the prenatal programming of hypertension remain unclear. Because endoplasmic reticulum (ER) stress plays a key role in vascular endothelial dysfunction in hypertension, we investigated whether aberrant ER stress causes endothelial dysfunction and high blood pressure in the offspring of dams with diabetes. Methods and results Pregnant Sprague-Dawley rats were intraperitoneally injected with streptozotocin (35 mg/kg) or citrate buffer at day 0 of gestation. Compared with control mother offspring (CMO), the diabetic mother offspring (DMO) had higher blood pressure and impaired endothelium-dependent relaxation (EDR) in mesenteric arteries, accompanied by decreased AMPK phosphorylation and PPARδ expression, increased ER stress markers and reactive oxygen species (ROS) levels. The inhibition of ER stress reversed these aberrant changes in DMO. Ex vivo treatment of mesenteric arteries with an AMPK agonist (A769662) or a PPARδ agonist (GW1516) improved the impaired EDR in DMO and reversed the tunicamycin-induced ER stress, ROS production, and EDR impairment in mesenteric arteries from CMO. The effects of A769662 were abolished by co-treatment with GSK0660 (PPARδ antagonist), whereas the effects of GW1516 were unaffected by Compound C (AMPK inhibitor). Conclusions These results suggest an abnormal fetal programming of vascular endothelial function in offspring of rats with maternal diabetes that is associated with increased ER stress, which can be ascribed to down-regulation of AMPK/PPARδ signaling cascade. Translational perspective Increasing evidence indicates that fetal programming is an important factor that contributes to the development of cardiovascular disease including hypertension and atherosclerosis later in life. This study explains the roles of AMPK/PPARδ/ER stress signaling cascade and endothelial dysfunction in maternal diabetes-programed adult hypertension in the offspring and provides a potential target for the prevention and therapy of this disease.

A wave of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant rapidly resulted in a steep increase in the infected population and an overloaded healthcare system. Effective medications for Omicron are currently limited. The previous observational study supports the efficacy and safety of Reyanning (RYN) mixture in the treatment of coronavirus disease 2019 (COVID-19).To evaluate the efficacy of RYN in asymptomatic and mildly infected patients with SARS-CoV-2 infection.This study was a prospective, open-label, randomized controlled trial. We consecutively recruited 2830 patients from Shanghai New International Expo Center mobile cabin hospital and randomized them in a 1:1 ratio to receive RYN plus standard care or receive standard care alone. The primary outcomes were the negative conversion of nucleic acid. Secondary outcomes included the hospital duration, new-onset symptoms, proportion of disease progression, and the viral load measured by the cycle threshold (Ct) value.A total of 1393 patients in the intervention group and 1407 patients in the control group completed the study. The negative conversion time of nucleic acid was significantly shortened in the intervention group (median: 6 d vs. 7 d, Hazard ratio: 0.768, 95CI %: 0.713-0.828, p0.0001). The negative conversion rate of nucleic acid was significantly higher in the intervention group (Day 3: 32.4% vs. 18.3%; Day7: 65.3% vs. 55.2%, p0.001). The hospitalization duration was significantly shortened in the intervention group (median: 8 d vs. 9 d, Hazard ratio: 0.759, 95% CI: 0.704-0.818, p0.0001). The proportion of new-onset fever (2.4% vs. 4.1%, p = 0.012), coughing (12.2% vs. 14.8%, p = 0.046), and expectoration (6.0% vs. 8.0%, p = 0.032) in the intervention group was significantly lower. RYN treatment increased Ct values and reduced the viral load. No disease progression and serious adverse events were reported during the study.RYN is a safe and effective treatment that can accelerate virus clearance and promote disease recovery in asymptomatic and mild Omicron infections.

To analyze the clinical characteristics and prognosis of patients infected with novel coronavirus Omicron variant in Shanghai, as to provide a reference for epidemic prevention, clinical diagnosis, and treatment.Altogether 4 264 novel coronavirus Omicron variant-infected patients with positive results of nucleic acid admitted to Shanghai New International Expo Center N3 Mobile Cabin Hospital from April 2 to May 7, 2022, were included. The demographic and baseline clinical characteristics, treatment strategy, prognosis, and different factors affecting the length of hospital stay were analyzed.A total of 4 264 novel coronavirus variant Omicron-infected cases were collected, including 3 111 cases (73.0%) asymptomatic infections and 1 153 cases (27.0%) mild infections. The overall median age was 45 (33, 55) years old with a range from 2 years old to 81 years old. The male to female ratio was 1.37:1. Altogether 3 305 cases (77.5%) had been vaccinated, of which 3 166 cases completed more than 2 doses. The upper respiratory tract symptoms such as cough and expectoration were the most common clinical manifestations of these infected patients. During the course of the disease, patients with asymptomatic infection were mainly treated with traditional Chinese medicine (TCM, 55.1%) and clinical observation (36.8%), and those with mild infection were mainly treated with TCM (42.2%) or integrated Chinese and Western medicine (30.4%). All patients were cured and discharged. The overall median length of hospital stay and the negative conversion time of nucleic acid were 9 (6, 10) days and 8 (5, 9) days, respectively. Compared with the asymptomatic infected patients, the hospitalization duration and the nucleic acid negative conversion time of the mildly infected patients were slightly longer [days: 10 (8, 11) vs. 9 (5, 10); 8 (6, 10) vs. 7 (4, 9), both P0.001]. Multiple linear regression analysis showed that the increasing age and mild infection were associated with longer hospitalization duration, and the treatment of TCM or integrated Chinese and Western medicine was associated with shortened length of hospital stay (all P0.05).The current novel coronavirus Omicron variant epidemic in Shanghai mainly caused asymptomatic and mild infections. The young and middle-aged population had a relatively high infection rate. The upper respiratory tract symptoms such as cough and expectoration were the most common clinical symptoms. Elderly and confirmed patients had prolonged hospitalization duration, while for patients receiving TCM treatment, the hospitalization duration was shortened.

Angiotensin II type 1 receptor (AT1R) is a vital therapeutic target for hypertension. Sorting nexin 1 (SNX1) participates in the sorting and trafficking of the renal dopamine D5 receptor, while angiotensin and dopamine are counterregulatory factors in the regulation of blood pressure. The effect of SNX1 on AT1R is not known. We hypothesized that SNX1, through arterial AT1R sorting and trafficking, is involved in blood pressure regulation. CRISPR/Cas9 system-generated SNX1-/- mice showed dramatic elevations in blood pressure compared to their wild-type littermates. The angiotensin II-mediated contractile reactivity of the mesenteric arteries and AT1R expression in the aortas were also increased. Moreover, immunofluorescence and immunoprecipitation analyses revealed that SNX1 and AT1R were colocalized and interacted in the aortas of wild-type mice. In vitro studies revealed that AT1R protein levels and downstream calcium signaling were upregulated in A10 cells treated with SNX1 siRNA. This may have resulted from decreased AT1R protein degradation since the AT1R mRNA levels showed no changes. AT1R protein was less degraded when SNX1 was downregulated, as reflected by a cycloheximide chase assay. Furthermore, proteasomal rather than lysosomal inhibition increased AT1R protein content, and this effect was accompanied by decayed binding of ubiquitin and AT1R after SNX1 knockdown. Confocal microscopy revealed that AT1R colocalized with PSMD6, a proteasomal marker, and the colocalization was reduced after SNX1 knockdown. These findings suggest that SNX1 sorts AT1R for proteasomal degradation and that SNX1 impairment increases arterial AT1R expression, leading to increased vasoconstriction and blood pressure.

JinHong Formula (JHF) was derived from the famous Rhubarb and Moutan Decoction which was prescribed for appendicitis. It was originally recorded in the classic of "Jingui Yaolve" written by Zhang Zhongjing. It is a kind of traditional Chinese medicine, widely used in the treatment of inflammation. However, the clinical effect of JHF for sepsis and its comprehensive mechanism in sepsis remained largely unknown.The aim of our study was to evaluate the clinical effect of JHF in the treatment of sepsis, and to explore its mechanism from the perspective of network pharmacology.The single-center randomized clinical trial was conducted to assess the effect of JHF in the treatment of sepsis. Additionally, we used the Chinese herbal medicine pharmacology database and analysis platform to identify the active components and therapeutic target of JHF. Numerous well-known disease target databases have been used to screen therapeutic target proteins for sepsis. Furthermore, we have established a Protein-Protein Interaction (PPI) network and carried out Gene Onotology/Kyoto Encyclopedia of Genes and Genomes (GO/KEGG) enrichment analysis. In order to conclude which active compounds from JHF may be responsible for signaling pathway, we performed network analysis.The study included 114 patients. By comparing participants with and without JHF, the results suggested that JHF significantly reduced all-cause mortality on 28 and 60 days after intervention, and improved Sequential Organ Failure Assessment (SOFA) on 7th day after intervention as well as. JHF had an effect of anti-inflammatories and antioxidants (SOD). By using network pharmacological analysis, we identified 72 active components and 426 target genes of JHF, and successfully constructed a "JHF-compound target-sepsis" network. 116 mentioned targets revealed by GO/KEGG enrichment analysis played a significant role in the inflammatory reaction and immunoregulation via interleukin-17 (IL-17) and tumor necrosis factor (TNF) signaling pathway. Moreover, the analysis of "pathway target-active component" revealed that Sennidin A, Rheidin A, Rheidin B, Rheidin C, (E)-4-Phenyl-3-Buten-2-One, Osmanthuside H, Esculetin, and Caffeicacid were responsible for IL-17, TNF signaling pathways.JHF contains potential active substance of anti-inflammatory and antioxidant. These active compounds may come into play through IL-17 and TNF signaling pathways. For sepsis, JHF may be a promising and effective treatment strategy.

Aims: Cardiac hypertrophy, in the long-term, is a maladaptive response to the change in hemodynamics needed to maintain cardiac output that leads to heart failure and sudden death. However, the underlying regulatory mechanisms causing cardiac hypertrophy remain to be elucidated. Recent studies have highlighted the importance of long non-coding RNAs (lncRNAs) in many biological processes and diseases. However, knowledge of the role of lncRNAs in cardiac diseases is still limited. Methods and Results：We identified the role of NPPA-AS1 in cardiac hypertrophy and remodeling. NPPA-AS1 is a heart-enriched and -conserved lncRNA, located in the antisense strand of the atrial natriuretic peptide (NPPA) gene. The cardiac expression of NPPA-AS1 was markedly upregulated early and sustained, in response to stress caused by transverse aortic constriction (TAC) in mice. NPPA-AS1 levels were also increased in failing human hearts. To determine the functional role of NPPA-AS1 in the heart in vivo, we inactivated NPPA-AS1 in mice using CRISPR-Cas9. Under basal conditions, there was no body or cardiac morphological or functional phenotype in NPPA-AS1-inactivated mice. However, in vivo germline inactivation of NPPA-AS1 minimized the TAC-induced cardiac hypertrophy and fibrosis and normalized the cardiac size, weight, and function. GapmeR-mediated or AAV9-shRNA-mediated silencing of NPPA-AS1 could also block and attenuate TAC-induced pathological cardiac remodeling, which reveals its clinical translation potential. The beneficial cardiac effects of inhibition of NPPA-AS1 were related to the recruitment of transcriptional coactivator EP300 to bind with transcription factor GATA4 to promote GATA4 acetylation and inhibition of hypertrophic gene expression. Conclusions: Our studies show that NPPA-AS1 recruits the transcriptional factor GATA4, increasing EP300/GATA4 binding and subsequent GATA4 acetylation to promote pathological cardiac hypertrophy. Inhibition of NPPA-AS1 could serve as a potential therapeutic strategy in the treatment of maladaptive cardiac hypertrophy and remodeling.

Cytidine base editors are powerful genetic tools that catalyse cytidine to thymidine conversion at specific genomic loci, and further improvement of the editing range and efficiency is critical for their broader applications. Through insertion of a non-sequence-specific single-stranded DNA-binding domain from Rad51 protein between Cas9 nickase and the deaminases, serial hyper cytidine base editors were generated with substantially increased activity and an expanded editing window towards the protospacer adjacent motif in both cell lines and mouse embryos. Additionally, hyeA3A-BE4max selectively catalysed cytidine conversion in TC motifs with a broader editing range and much higher activity (up to 257-fold) compared with eA3A-BE4max. Moreover, hyeA3A-BE4max specifically generated a C-to-T conversion without inducing bystander mutations in the haemoglobin gamma gene promoter to mimic a naturally occurring genetic variant for amelioration of β-haemoglobinopathy, suggesting the therapeutic potential of the improved base editors.

Acute myocardial infarction (MI) is one of the leading causes of death in humans. Our previous studies showed that gastrin alleviated acute myocardial ischaemia–reperfusion injury. We hypothesize that gastrin might protect against heart injury after MI by promoting angiogenesis. An MI model was simulated by ligating the anterior descending coronary artery in adult male C57BL/6J mice. Gastrin was administered twice daily by intraperitoneal injection for 2 weeks after MI. We found that gastrin reduced mortality, improved myocardial function with reduced infarct size and promoted angiogenesis. Gastrin increased HIF-1α and VEGF expression. Downregulation of HIF-1α expression by siRNA reduced the proliferation, migration and tube formation of human umbilical vein endothelial cells. These results indicate that gastrin restores cardiac function after MI by promoting angiogenesis via the HIF-1α/VEGF pathway.

Summary Development of spermatogonia and spermatocytes are the critical steps of spermatogenesis, impacting on male fertility. Investigation of the related regulators benefits the understanding of male reproduction. The proteasome system has been reported to regulate spermatogenesis, but the mechanisms and key contributing factors in vivo are poorly explored. Here we found that ablation of REGγ, a proteasome activator, resulted in male subfertility. Analysis of the mouse testes after birth showed there was a decreased number of PLZF+ spermatogonia and spermatocytes. Molecular analysis found that REGγ loss significantly increased the abundance of p53 protein in the testis, and directly repressed PLZF transcription in cell lines. Of note, allelic p53 haplodeficiency partially rescued the defects in spermatogenesis observed in REGγ-deficient mice. In summary, our results identify REGγ-p53-PLZF to be a critical pathway that regulates spermatogenesis and establishes a new molecular link between the proteasome system and male reproduction., Graphical Abstract, Highlights • REGγ loss results in male subfertility • REGγ loss results in a decrease of spermatocytes and PLZF+ spermatogonial cells • p53 protein, increased in REGγ−/− mouse testes, represses PLZF expression • Allelic p53 haplodeficiency partially rescues defects in REGγ−/− mouse spermatogenesis, In this article, Lei Li and colleagues show that ablation of REGγ, a proteasome activator, resulted in male subfertility, partially because of a decreased number of PLZF+ spermatogonia and spermatocytes. Mechanistically, REGγ loss significantly increased the abundance of p53 protein, and transcriptionally represses PLZF expression. Allelic p53 haplodeficiency partially rescued the defects in spermatogenesis observed in REGγ-deficient mice.

BACKGROUND: Genetic variants of coding genes related to blood pressure regulation participate in the pathogenesis of hypertension and determines the response to specific antihypertensive drugs. G protein-coupled receptor kinase 4 (GRK4) and its variants are of great importance in pathogenesis of hypertension. However, little is known about role of GRK4 variants in determine circadian rhythm of blood pressure and response to candesartan in hypertension. The aim of this study was to analyze the correlation of GRK4 variants and circadian rhythm of blood pressure, and to explore their effect on antihypertensive efficiency of candestartan. METHODS: In this study, a total of 1239 cases were eligible, completed ambulatory blood pressure monitoring (ABPm) observation and exon sequencing of G protein-coupled receptor kinase 4 (GRK4). ABPm was obtained before and after 4-week treatment of candesartan. Diurnal variation of systolic blood pressure and antihypertensive effect of candesartan were then assessed. RESULTS: Compared to GRK4 wild type (GRK4-WT), patients with GRK4 variants were more likely to be non-dippers (odds ratio (OR) 6.672, 95% confidence interval (CI) 5.124–8.688, P < .001), with GRK4 A142V (OR 5.888, 95% CI 4.332–8.003, P < .001), A486V (OR 7.102, 95% CI 5.334–9.455, P < .001) and GRK4 R65L (OR 3.273, 95% CI 2.271–4.718, P < .001), respectively. Correlation analysis revealed that non-dippers rhythm of blood pressure were associated with GRK4 variants (r = .420, P < .001), with GRK4 A142V (r = .416, P < .001), A486V (r = .465, P < .001) and GRK4 R65L (r = .266, P < .001), respectively. When given 4-week candesartan, patients with GRK4 variants showed better antihypertensive effect as to drop in blood pressure (24 h mSBP, 21.21 ± 4.99 vs 12.34 ± 4.78 mmHg, P < .001) and morning peak (MP-SBP, 16.54 ± 4.37 vs 11.52 ± 4.14 mmHg, P < .001), as well as greater increase in trough to peak ratio (SBP-T/P, .71 ± .07 vs .58 ± .07, P < .001) and smoothness index (SBP-SI, 1.44 ± .16 vs 1.17 ± .11, P < .001) than those with GRK4 WT. CONCLUSION: This study indicates that hypertensive patients with GRK4 variants are more likely to be non-dippers. What’s more, patients with GRK4 variants possess a significantly better antihypertensive response to candesartan than those with GRK4 WT.

The endothelin receptor type B (ETBR) regulates water and electrolyte balance and blood pressure, in part, by inhibiting renal sodium transport. Our preliminary study found that the ETBR-mediated diuresis and natriuresis are impaired in hypertension with unknown mechanism. Persistently increased activity of G protein-coupled receptor kinase 4 (GRK4), caused by increased expression or genetic variants (eg, GRKγ142V), impairs the ability of the kidney to excrete a sodium load, in part, by impairing renal dopamine D(1) receptor function through persistent phosphorylation. Our present study found that although renal ETBR expression was not different between Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHRs), renal ETBR phosphorylation was higher in SHRs. The role of hyper-phosphorylation in impaired ETBR-function was supported by results in human (h) GRK4γ transgenic mice. Stimulation of ETBR by BQ3020-induced natriuresis in human (h) GRK4γ wild-type (WT) mice. However, in hGRK4γ 142V transgenic mice, the renal ETBR was hyperphosphorylated and ETBR-mediated natriuresis and diuresis were not evident. There were co-localization and co-immunoprecipitation of ETBR and GRK4 in renal proximal tubule (RPT) cells from both WKY and SHRs but was greater in the latter than the former group. SiRNA-mediated downregulation of GRK4 expression, recovered the impaired inhibitory effect of ETBR on Na(+)-K(+)-ATPase activity in RPT cells from SHR. In vivo downregulation of renal GRK4 expression, via ultrasound-targeted microbubble destruction, decreased ETBR phosphorylation and restored ETBR-mediated natriuresis and diuresis in SHRs. This study provides a mechanism by which GRK4, via regulation of renal ETBR function, participates in the pathogenesis of hypertension.

Objectives The mechanisms responsible for the postnatal loss of mammalian cardiac regenerative capacity are not fully elucidated. The aim of the present study is to investigate the role of progesterone in cardiac regeneration and explore underlying mechanism. Materials and Methods Effect of progesterone on cardiomyocyte proliferation was analysed by immunofluorescent staining. RNA sequencing was performed to screen key target genes of progesterone, and yes‐associated protein (YAP) was knocked down to demonstrate its role in pro‐proliferative effect of progesterone. Effect of progesterone on activity of YAP promoter was measured by luciferase assay and interaction between progesterone receptor and YAP promoter by electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP). Adult mice were subjected to myocardial infarction, and then, effects of progesterone on adult cardiac regeneration were analysed. Results Progesterone supplementation enhanced cardiomyocyte proliferation in a progesterone receptor‐dependent manner. Progesterone up‐regulated YAP expression and knockdown of YAP by small interfering RNA reduced progesterone‐mediated cardiomyocyte proliferative effect. Progesterone receptor interacted with the YAP promoter, determined by ChIP and EMSA; progesterone increased luciferase activity of YAP promoter and up‐regulated YAP target genes. Progesterone administration also promoted adult cardiomyocyte proliferation and improved cardiac function in myocardial infarction. Conclusion Our data uncover a role of circulating progesterone withdrawal as a novel mechanism for the postnatal loss of mammalian cardiac regenerative potential. Progesterone promotes both neonatal and adult cardiomyocyte proliferation by up‐regulating YAP expression., Progesterone promotes cardiomyocyte proliferation by transcriptional activation of YAP.

Background: Long noncoding RNA (lncRNA) can regulate various physiological and pathological processes through multiple molecular mechanisms in cis and in trans. However, the role of lncRNAs in cardiac hypertrophy is yet to be fully elucidated. Methods: A mouse lncRNA microarray was used to identify differentially expressed lncRNAs in the mouse hearts following transverse aortic constriction–induced pressure overload comparing to the sham-operated samples. The direct impact of one lncRNA, Ahit, on cardiomyocyte hypertrophy was characterized in neonatal rat cardiomyocytes in response to phenylephrine by targeted knockdown and overexpression. The in vivo function of Ahit was analyzed in mouse hearts by using cardiac-specific adeno-associated virus, serotype 9–short hairpin RNA to knockdown Ahit in combination with transverse aortic constriction. Using catRAPID program, an interaction between Ahit and SUZ12 (suppressor of zeste 12 protein homolog) was predicted and validated by RNA immunoprecipitation and immunoblotting following RNA pull-down. Chromatin immunoprecipitation was performed to determine SUZ12 or H3K27me3 occupancy on the MEF2A (myocyte enhancer factor 2A) promoter. Finally, the expression of human Ahit (leukemia-associated noncoding IGF1R activator RNA 1 [LUNAR1]) in the serum samples from patients of hypertrophic cardiomyopathy was tested by quantitative real-time polymerase chain reaction. Results: A previously unannotated lncRNA, antihypertrophic interrelated transcript ( Ahit ), was identified to be upregulated in the mouse hearts after transverse aortic constriction. Inhibition of Ahit induced cardiac hypertrophy, both in vitro and in vivo, associated with increased expression of MEF2A, a critical transcriptional factor involved in cardiac hypertrophy. In contrast, overexpression of Ahit significantly attenuated stress-induced cardiac hypertrophy in vitro. Furthermore, Ahit was significantly upregulated in serum samples of patients diagnosed with hypertensive heart disease versus nonhypertrophic hearts (1.46±0.17 fold, P =0.0325). Mechanistically, Ahit directly bound and recruited SUZ12, a core PRC2 (polycomb repressive complex 2) protein, to the promoter of MEF2A, triggering its trimethylation on H3 lysine 27 (H3K27me3) residues and mediating the downregulation of MEF2A, thereby preventing cardiac hypertrophy. Conclusions: Ahit is a lncRNA with a significant role in cardiac hypertrophy regulation through epigenomic modulation. Ahit is a potential therapeutic target of cardiac hypertrophy.

Both renin–angiotensin systems and insulin participate in kidney-involved blood pressure regulation. Activation of angiotensin II type 2 receptor (AT 2 R) decreases sodium reabsorption in renal proximal tubule (RPT) cells, whereas insulin produces the opposite effect. We presume that AT 2 R has an inhibitory effect on insulin receptor expression in RPT cells, which may affect renal sodium transport and therefore be of physiological or pathological significance. Our present study found that activation of AT 2 R inhibited insulin receptor expression in a concentration and time-dependent manner in RPT cells from Wistar-Kyoto (WKY) rats. In the presence of a protein kinase C (PKC) inhibitor (PKC inhibitor peptide 19–31, 10 −6 mol/L) or a phosphatidylinositol 3 kinase inhibitor (wortmannin, 10 −6 mol/L), the inhibitory effect of AT 2 R on insulin receptor was blocked, indicating that both PKC and phosphatidylinositol 3 kinase were involved in the signaling pathway. There was a linkage between AT 2 R and insulin receptor which was determined by both laser confocal microscopy and coimmunoprecipitation. However, the effect of AT 2 R activation on insulin receptor expression was different in RPT cells from spontaneously hypertensive rats (SHRs). Being contrary to the effect in WKY RPT cells, AT 2 R stimulation increased insulin receptor in SHR RPT cells. Insulin (10 −7 mol/L, 15 minutes) enhanced Na + -K + -ATPase activity in both WKY and SHR RPT cells. Pretreatment with CGP42112 decreased the stimulatory effect of insulin on Na + -K + -ATPase activity in WKY RPT cells, whereas pretreatment with CGP42112 increased it in SHR RPT cells. It is suggested that activation of AT 2 R inhibits insulin receptor expression and function in RPT cells. The lost inhibitory effect of AT 2 R on insulin receptor expression may contribute to the pathophysiology of hypertension.

Mesenchymal stem cells (MSCs) exert therapeutic effect on treating acute myocardial infarction. Recent evidence showed that paracrine function rather than direct differentiation predominately contributes to the beneficial effects of MSCs, but how the paracrine factors function are not fully elucidated. In the present study, we tested if extracellular vesicles (EVs) secreted by MSC promotes angiogenesis in infracted heart via microRNAs. Immunostaining of CD31 and matrigel plug assay were performed to detect angiogenesis in a mouse myocardial infarction (MI) model. The cardiac function and structure was examined with echocardiographic analysis. Capillary-like tube formation, migration and proliferation of human umbilical vein endothelial cells (HUVECs) were determined. As a result, MSC-EVs significantly improved angiogenesis and cardiac function in post-MI heart. MSC-EVs increased the proliferation, migration and tube formation capacity of HUVECs. MicroRNA (miR)-210 was found to be enriched in MSC-EVs. The EVs collected from MSCs with miR-210 silence largely lost the pro-angiogenic effect both in-vitro and in-vivo. The miR-210 target gene Efna3, which plays a role in angiogenesis, was down-regulated by MSC-EVs treatment in HUVECs. In conclusion, MSC-EVs are sufficient to improve angiogenesis and exert therapeutic effect on MI, its pro- angiogenesis effect might be associated with a miR-210-Efna3 dependent mechanism. This article is part of a Special Issue entitled: Genetic and epigenetic control of heart failure - edited by Jun Ren & Megan Yingmei Zhang.

BACKGROUND Essential hypertension is associated with increased plasma concentrations of extracellular vesicles (EVs). We aimed to determine the role of monocyte miR-27a in EVs on arterial Mas receptor expression, and its involvement in the pathogenesis of hypertension. METHODS THP-1 cells were transfected with miR-27a mimic and miR-27a inhibitor, and EVs were collected. Mas receptor expression and endothelial nitric oxide synthase (eNOS) phosphorylation were determined by immunoblotting. Sprague–Dawley (SD) rats received EVs via tail-vein injection. Blood pressure (BP) was measured with the tail-cuff method. The vasodilatory response of mesenteric arteries was measured using a small vessel myograph. RESULTS EVs from THP-1 cells increased rat BP by impairing Ang-(1–7)-mediated vasodilation in mesenteric arteries, which was further exaggerated by EVs from lipopolysaccharides-treated THP-1 cells. As the receptor and key signaling of Ang-(1–7), next experiments found that Mas receptor expression and eNOS phosphorylation were decreased in mesenteric arteries from EVs-treated SD rats. Screening studies found miR-27a in EVs may be involved in this process. Through transfection with miR-27a inhibitor or miR-27a mimic, we found that miR-27a downregulates Mas receptor expression in endothelial cells. Injection of EVs from miR-27a-transfected HEK-293 cells decreased Mas receptor and eNOS phosphorylation in mesenteric arteries, impaired Ang-(1–7)-mediated vasodilation and increased BP. Earlier effects were reversed using cells with downregulation of miR-27 in EVs. CONCLUSIONS Monocyte miR-27a in EVs decreases Mas receptor expression and eNOS phosphorylation in endothelium, impairs Ang-(1–7)-mediated vasodilation, and causes hypertension. Understanding the contributions of EVs in the pathogenesis of hypertension may facilitate their use as a diagnostic biomarker.

Cardiac troponin I (cTnI), a biomarker for myocardial damage and risk stratification, may be involved in the pathogenesis of cardiovascular diseases, which was ascribed to the effect of cTnI auto-antibodies. Whether or not cTnI itself has a direct impact on acute myocardial injury is unknown. To exclude the influence of cTnI antibody on the cardiac infarct size, we studied the effect of cTnI shortly after myocardial ischaemia–reperfusion (I/R) injury when cTnI antibodies were not elevated. Pretreatment with cTnI augmented the myocardial infarct size caused by I/R, accompanied by an increase in inflammatory markers in the blood and myocardium. Additional experiments using human umbilical vein endothelial cells (HUVECs) showed that the detrimental effect of cTnI was related to cTnI-induced increase in vascular cell adhesion molecule-1 (VCAM-1) expression and VCAM-1 mediated adhesion of human monocytes (THP-1) to HUVECs, which could be neutralized by VCAM-1 antibody. Both toll-like receptor 4 (TLR4) and nuclear factor-κB (NF-κB) were involved in the signalling pathway, because blockade of either TLR4 or NF-κB inhibited the cTnI's effect on VCAM-1 expression and adhesion of monocytes to endothelial cells. Moreover, TLR4 inhibition reduced cTnI-augmented cardiac injury in rats with I/R injury. We conclude that cTnI exacerbates myocardial I/R injury by inducing the adhesion of monocytes to vascular endothelial cells via activation of the TLR4/NF-κB pathway. Inhibition of TLR4 may be an alternative strategy to reduce cTnI-induced myocardial I/R injury.

Background Dopamine receptors induce natriuresis in kidney. Previous studies have shown interactions between different subtypes of dopamine receptors in renal proximal tubule (RPT) cells. We hypothesize that D3 receptors have an interaction with D4 receptors in RPT cells from normotensive rats (Wistar-Kyoto, WKY) and spontaneously hypertensive rats (SHRs). Methods Immunoblotting and reverse transcriptase-polymerase chain reaction (RT-PCR) were used to examine the expression of D3 and D4 receptors. Na-K-ATPase activity was used to measure the function of receptors. The distribution and colocalization of D3 and D4 receptors were detected by confocal microscopy and co-immunoprecipitation. Results D3 receptor agonist PD128907 increased the mRNA and protein expression of D4 receptors in RPT cells from WKY rats, but decreased that from SHRs. In the presence of PLC blocker (U73122, 10-mol/l) or PKC inhibitor 19 -31 (10-mol/l), the up-regulation of D3 receptor on D4 receptor was lost in WKY cells. Moreover, stimulation with PD128907 for 30 minutes decreased D4 receptor degradation in WKY cells, not in SHR cells. D3 and D4 receptors colocalized and co-immunoprecipitated in RPT cells. PD128907 increased co-immunoprecipitation of D3 and D4 receptors in WKY RPT cells, but not in SHR RPT cells. Pre-treatment with D3 receptor agonist also increases D4 receptor mediated inhibitory effect on Na-K-ATPase activity in WKY cells, but not in SHR cells. Conclusion Renal D3 receptor regulates the expression and function of D4 receptor in RPT cells via PLC /PKC signaling pathway, the loss of this interaction might be involved in the pathogenesis of hypertension.

Epidemiological and experimental studies suggest that maternal diabetes mellitus programs hypertension that is associated with impaired sodium excretion in the adult offspring. However, the underlying mechanisms are not clear. Because dopamine receptor function is involved in the pathogenesis of hypertension, we hypothesized that impaired renal dopamine D 1 receptor function is also involved in the hypertension in offspring of maternal diabetes mellitus. Maternal diabetes mellitus was induced by a single intraperitoneal injection of streptozotocin (35 mg/kg) to pregnant Sprague-Dawley rats at day 0 of gestation. Compared with the offspring of mothers injected with citrate buffer (control mother offspring), the diabetic mother offspring (DMO) had increased systolic blood pressure and impaired D 1 receptor-mediated diuresis and natriuresis, accompanied by increased renal PKC (protein kinase C) expression and activity, GRK-2 (G protein–coupled receptor kinase-2) expression, D 1 receptor phosphorylation, D 1 receptor/Gαs uncoupling, and loss of D 1 receptor-mediated inhibition of Na + -K + -ATPase activity in renal proximal tubule cells from DMO. Inhibition of PKC reduced the increased GRK-2 expression and normalized D 1 receptor function in primary cultures of renal proximal tubule cells from DMO. In addition, DMO, relative to control mother offspring, in vivo, had increased oxidative stress, indicated by decreased renal glutathione and increased renal malondialdehyde and urine 8-isoprostane. Normalization of oxidative stress with tempol also normalized the renal D 1 receptor phosphorylation, D 1 receptor-mediated diuresis and natriuresis, and blood pressure in DMO. Our present study indicates that maternal diabetes mellitus–programed hypertension in the offspring is caused by impaired renal D 1 receptor function because of oxidative stress that is mediated by increased PKC-GRK-2 activity.

Genome-wide association studies (GWAS) have identified many genetic variants in genes related to lipid metabolism. However, how these variations affect lipid levels remains elusive. Long non-coding RNAs (lncRNAs) have been implicated in a variety of biological processes. We hypothesize lncRNAs are likely to be located within disease or trait-associated DNA regions to regulate lipid metabolism. The aim of this study was to investigate whether and how lncRNAs in lipid- associated DNA regions regulate cholesterol homeostasis in hepatocytes. In this study, we identified a novel long non-coding RNA in Lipid Associated Single nucleotide polymorphism gEne Region (LASER) by bioinformatic analysis. We report that LASER is highly expressed in both hepatocytes and peripheral mononuclear cells (PBMCs). Clinical studies showed that LASER expression is positively related with that of cholesterol containing apolipoprotein levels. In particular, we found that LASER is positively correlated with plasma PCSK9 levels in statin free patients. siRNAs mediated knock down of LASER dramatically reduces intracellular cholesterol levels and affects the expression of genes involved in cholesterol metabolism. Transcriptome analyses show that knockdown of LASER affects the expression of genes involved in metabolism pathways. We found that HNF-1α and PCSK9 were reduced after LASER knock-down. Interestingly, the reduction of PCSK9 can be blocked by the treatment of berberine, a natural cholesterol-lowering compound which functions as a HNF-1α antagonist. Mechanistically, we found that LASER binds to LSD1 (lysine-specific demethylase 1), a member of CoREST/REST complex, in nucleus. LASER knock-down enhance LSD1 targeting to genomic loci, resulting in decreased histone H3 lysine 4 mono-methylation at the promoter regions of HNF-1α gene. Conversely, LSD1 knock-down abolished the effect of LASER on HNF-1α and PCSK9 expressions. Finally, we found that statin treatment increased LASER expression, accompanied with increased PCSK9 expression, suggesting a feedback regulation of cholesterol on LASER expression. This observation may partly explain the statin escape during anti-cholesterol treatment. These findings identified a novel lncRNA in cholesterol homeostasis. Therapeutic targeting LASER might be an effective approach to augment the effect of statins on cholesterol levels in clinics.

Background Ischemia/reperfusion injury (IRI) is one of the most predominant complications of ischemic heart disease. Gastrin has emerged as a regulator of cardiovascular function, playing a key protective role in hypoxia. Serum gastrin levels are increased in patients with myocardial infarction, but the pathophysiogical significance of this finding is unknown. The purpose of this study was to determine whether and how gastrin protects cardiac myocytes from IRI. Methods and Results Adult male Sprague‐Dawley rats were used in the experiments. The hearts in living rats or isolated Langendorff‐perfused rat hearts were subjected to ischemia followed by reperfusion to induce myocardial IRI. Gastrin, alone or with an antagonist, was administered before the induction of myocardial IRI. We found that gastrin improved myocardial function and reduced the expression of myocardial injury markers, infarct size, and cardiomyocyte apoptosis induced by IRI. Gastrin increased the phosphorylation levels of ERK 1/2 (extracellular signal‐regulated kinase 1/2), AKT (protein kinase B), and STAT 3 (signal transducer and activator of transcription 3), indicating its ability to activate the RISK (reperfusion injury salvage kinase) and SAFE (survivor activating factor enhancement) pathways. The presence of inhibitors of ERK 1/2, AKT , or STAT 3 abrogated the gastrin‐mediated protection. The protective effect of gastrin was via CCK2R (cholecystokinin 2 receptor) because the CCK 2R blocker CI 988 prevented the gastrin‐mediated protection of the heart with IRI. Moreover, we found a negative correlation between serum levels of cardiac troponin I and gastrin in patients with unstable angina pectoris undergoing percutaneous coronary intervention, suggesting a protective effect of gastrin in human cardiomyocytes. Conclusions These results indicate that gastrin can reduce myocardial IRI by activation of the RISK and SAFE pathways.

Cardiovascular diseases, such as hypertension, could be programmed in fetal life. Prenatal lipopolysaccharide (LPS) exposure in utero results in increased blood pressure in offspring, but the vascular mechanisms involved are unclear. Pregnant Sprague–Dawley rats were intraperitoneally injected with LPS (0.79 mg/kg) or saline (0.5 ml) on gestation days 8, 10, and 12. The offspring of LPS-treated dams had higher blood pressure and decreased acetylcholine (ACh)-induced relaxation and increased phenylephrine (PE)-induced contraction in endothelium-intact mesenteric arteries. Endothelium removal significantly enhanced the PE-induced contraction in offspring of control but not LPS-treated dams. The arteries pretreated with l -NAME to inhibit nitric oxide synthase (eNOS) in the endothelium or ODQ to inhibit cGMP production in the vascular smooth muscle had attenuated ACh-induced relaxation but augmented PE-induced contraction to a larger extent in arteries from offspring of control than those from LPS-treated dams. In addition, the endothelium-independent relaxation caused by sodium nitroprusside was also decreased in arteries from offspring of LPS-treated dams. The functional results were accompanied by a reduction in the expressions of eNOS and soluble guanylate cyclase (sGC) and production of NO and cGMP in arteries from offspring of LPS-treated dams. Furthermore, LPS-treated dam’s offspring arteries had increased oxidative stress and decreased antioxidant capacity. Three-week treatment with TEMPOL, a reactive oxygen species (ROS) scavenger, normalized the alterations in the levels of ROS, eNOS, and sGC, as well as in the production of NO and cGMP and vascular function in the arteries of the offspring of LPS-treated dams. In conclusion, prenatal LPS exposure programs vascular dysfunction of mesenteric arteries through increased oxidative stress and impaired NO–cGMP signaling pathway.

Angiotensin type 1a receptor (AT1aR) is thought to play an important role in the development of hypertension. However, it is unknown how the AT1aR expression in vascular tissue is changed during the development of hypertension or if the degree of methylation in the AT1aR promoter correlates with the expression of AT1aR. To address these questions, we measured AT1aR mRNA, protein expression, and methylation status of the AT1aR promoter in the aorta and mesenteric artery of male spontaneously hypertensive rats (SHRs) and age-matched Wistar-Kyoto (WKY) rats acting as controls at pre-hypertensive (4 weeks), evolving (10 weeks), and established (20 weeks) stages of hypertension. The expression of the AT1aR mRNA and protein was not different between the SHRs and WKY rats at 4 weeks. However, they were significantly greater in SHRs than in WKY rats at 20 weeks. Bisulfite sequencing revealed that the AT1aR promoter from the aorta and mesenteric artery of the SHRs was progressively hypo-methylated with age as compared with their WKY rat counterparts. These results suggest that the heightened AT1aR expression in SHRs is related to the AT1aR promoter hypo-methylation, which might be a consequence of the increased blood pressure and may be important in the maintenance of high blood pressure.

The abnormal migration of vascular smooth muscle cells (VSMCs) has been implicated to contribute to lesion formation in the adult vasculature. The renin-angiotensin-aldosterone system (RAAS) is intensively involved in the pathogenesis of a variety of cardiovascular diseases. There are increasing pieces of evidence for interactions between RAAS and dopamine receptors. We hypothesize that the D3 receptor has an inhibitory effect on angiotensin II (Ang II)/aldosterone-induced VSMC migration.In this study, embryonic thoracic aortic smooth muscle cells were cultured. VSMC migration was determined by the Boyden chamber and wound healing assays.VSMC migration was increased by Ang II (10(-10)-10(-7) mol/L) in a concentration-dependent manner, but not by aldosterone (10(-10)-10(-7) mol/L), and a synergistic effect of Ang II (10(-10) mol/L)/aldosterone (10(-10)mol/L) was also observed in VSMC migration. The migratory effects of Ang II alone/with aldosterone were attenuated by the activation of D3 receptors (10(-10)-10(-7) mol/L), although a D3 receptor agonist, PD128907, by itself, had no effect on VSMC migration. The inhibitory effect of the D3 receptor on Ang II/ aldosterone-mediated VSMC migration was blocked by the blocker of PKA (14-22 amide, 10(-7) mol/L), indicating that PKA was involved in the signaling pathway.These results indicate that activation of vascular D3 receptors inhibits Ang II/aldosterone-induced VSMC migration through the PKA signal pathway, which may be important in the regulation of vascular remodeling.

Limb remote ischemic preconditioning (RIPC) is an effective means of protection against ischemia/reperfusion (IR)-induced injury to multiple organs. Many studies are focused on identifying endocrine mechanisms that underlie the cross-talk between muscle and RIPC-mediated organ protection. We report that RIPC releases irisin, a myokine derived from the extracellular portion of fibronectin domain-containing 5 protein (FNDC5) in skeletal muscle, to protect against injury to the lung. Human patients with neonatal respiratory distress syndrome show reduced concentrations of irisin in the serum and increased irisin concentrations in the bronchoalveolar lavage fluid, suggesting transfer of irisin from circulation to the lung under physiologic stress. In mice, application of brief periods of ischemia preconditioning stimulates release of irisin into circulation and transfer of irisin to the lung subjected to IR injury. Irisin, via lipid raft-mediated endocytosis, enters alveolar cells and targets mitochondria. Interaction between irisin and mitochondrial uncoupling protein 2 (UCP2) allows for prevention of IR-induced oxidative stress and preservation of mitochondrial function. Animal model studies show that intravenous administration of exogenous irisin protects against IR-induced injury to the lung via improvement of mitochondrial function, whereas in UCP2-deficient mice or in the presence of a UCP2 inhibitor, the protective effect of irisin is compromised. These results demonstrate that irisin is a myokine that facilitates RIPC-mediated lung protection. Targeting the action of irisin in mitochondria presents a potential therapeutic intervention for pulmonary IR injury.