HNEAP Regulates Necroptosis of Cardiomyocytes by Suppressing the m5 C Methylation of Atf7 mRNA.

PIWI-interacting RNAs (piRNAs) are highly expressed in various cardiovascular diseases. However, their role in cardiomyocyte death caused by ischemia/reperfusion (I/R) injury, especially necroptosis, remains elusive. In this study, a heart necroptosis-associated piRNA (HNEAP) is found that regulates cardiomyocyte necroptosis by targeting DNA methyltransferase 1 (DNMT1)-mediated 5-methylcytosine (m5 C) methylation of the activating transcription factor 7 (Atf7) mRNA transcript. HNEAP expression level is significantly elevated in hypoxia/reoxygenation (H/R)-exposed cardiomyocytes and I/R-injured mouse hearts. Loss of HNEAP inhibited cardiomyocyte necroptosis and ameliorated cardiac function in mice. Mechanistically, HNEAP directly interacts with DNMT1 and attenuates m5 C methylation of the Atf7 mRNA transcript, which increases Atf7 expression level. ATF7 can further downregulate the transcription of Chmp2a, an inhibitor of necroptosis, resulting in the reduction of Chmp2a level and the progression of cardiomyocyte necroptosis. The findings reveal that piRNA-mediated m5 C methylation is involved in the regulation of cardiomyocyte necroptosis. Thus, the HNEAP-DNMT1-ATF7-CHMP2A axis may be a potential target for attenuating cardiac injury caused by necroptosis in ischemic heart disease.

Circular RNA FEACR inhibits ferroptosis and alleviates myocardial ischemia/reperfusion injury by interacting with NAMPT.

BACKGROUND: Emerging research has reported that circular RNAs (circRNAs) play important roles in cardiac cell death after myocardial ischemia and reperfusion (I/R). Ferroptosis, a new form of cell death discovered in recent years, has been proven to participate in the regulation of myocardial I/R. This study used circRNA sequencing to explore the key circRNA in the regulation of cardiac ferroptosis after I/R and study the mechanisms of potential circRNA function. METHODS: We performed circRNA sequencing to explore circRNAs differentially expressed after myocardial I/R. We used quantitative polymerase chain reactions to determine the circRNA expression in different tissues and detect the circRNA subcellular localization in the cardiomyocyte. Gain- and loss-of-function experiments were aimed to examine the function of circRNAs in cardiomyocyte ferroptosis and cardiac tissue damage after myocardial I/R. RNA pull-down was applied to explore proteins interacting with circRNA. RESULTS: Here, we identified a ferroptosis-associated circRNA (FEACR) that has an underlying regulatory role in cardiomyocyte ferroptosis. FEACR overexpression suppressed I/R-induced myocardial infarction and ameliorated cardiac function. FEACR inhibition induces ferroptosis in cardiomyocytes and FEACR overexpression inhibits hypoxia and reoxygenation-induced ferroptosis. Mechanistically, FEACR directly bound to nicotinamide phosphoribosyltransferase (NAMPT) and enhanced the protein stability of NAMPT, which increased NAMPT-dependent Sirtuin1 (Sirt1) expression, which promoted the transcriptional activity of forkhead box protein O1 (FOXO1) by reducing FOXO1 acetylation levels. FOXO1 further upregulated the transcription of ferritin heavy chain 1 (Fth1), a ferroptosis suppressor, which resulted in the inhibition of cardiomyocyte ferroptosis. CONCLUSIONS: Our finding reveals that the circRNA FEACR-mediated NAMPT-Sirt1-FOXO1-FTH1 signaling axis participates in the regulation of cardiomyocyte ferroptosis and protects the heart function against I/R injury. Thus, FEACR and its downstream factors could be novel targets for alleviating ferroptosis-related myocardial injury in ischemic heart diseases.

Influence of rutin on the effects of neonatal cigarette smoke exposure-induced exacerbated MMP-9 expression, Th17 cytokines and NF-κB/iNOS-mediated inflammatory responses in asthmatic mice model.

Allergic asthma is one of the most enduring diseases of the airway. The T-helper cells and regulatory T-cells are critically involved in inflammatory responses, mucus hypersecretion, airway remodelling and in airway hyper-responsiveness. Cigarette smoke (CS) has been found to aggravate inflammatory responses in asthma. Though currently employed drugs are effective, associated side effects demand identification and development of novel drugs with negligible or no adverse effects. Rutin, plant-derived flavonoid has been found to possess antioxidant and anti-inflammatory effects. We investigated the ability of rutin to modulate T-cells and inhibit inflammation in experimentally-induced asthma in cigarette smoke exposed mice. Separate groups of neonatal mice were exposed to CS for 10 days from post-natal days 2 to 11. After 2 weeks, the mice were sensitized and challenged with ovalbumin (OVA). Treatment group were given rutin (37.5 or 75 mg/kg body weight) during OVA sensitization and challenge. Rutin treatment was found to significantly inhibit cellular infiltration in the airways and Th2 and Th17 cytokine levels as well. Flow cytometry revealed effectively raised CD4+CD25+Fox3+ Treg cells and supressed Th17 cell population on rutin treatment. Airway hyper-responsiveness observed following CS and OVA challenge were inhibited by rutin. NF-κB and iNOS, chief regulators of inflammatory responses robustly activated by CS and OVA were down-regulated by rutin. Rutin also inhibited the expression of matrix metalloproteinase 9, thereby aiding in prevention of airway remodelling in asthma thereby revealing to be a potent candidate in asthma therapy.

Clinical observations on attention-deficit hyperactivity disorder (ADHD) in children with frontal lobe epilepsy.

The objective was to investigate the prevalence of attention-deficit hyperactivity disorder (ADHD) in children with frontal lobe epilepsy and related factors. The medical records of 190 children diagnosed with frontal lobe epilepsy at Qilu Hospital of Shandong University between 2006 and 2011 were retrospectively collected, and a follow-up analysis of the prevalence of ADHD in these children was conducted. Of the 161 children with an effective follow-up, 59.0% (95/161) with frontal lobe epilepsy suffered from ADHD as well. Analysis of epilepsy and ADHD-related factors indicated that the incidence of ADHD was 89.4% (76/85) in children with abnormal electroencephalogram (EEG) discharges on the most recent EEG, which was significantly higher than the ADHD incidence of 25% (19/76) in children with normal readings on the most recent EEG (P < .01). Children with frontal lobe epilepsy have a high incidence of ADHD. Sustained abnormal discharge on the electroencephalogram is associated with increased comorbidity of ADHD with frontal lobe epilepsy.

Phase I Study to Determine MTD of Docetaxel and Cisplatin with Concurrent Radiation Therapy for Stage III Non-Small Cell Lung Cancer.

OBJECTIVE: To evaluate the maximum tolerated dose (MTD) of docetaxel (DCT) and cisplatin (DDP) concurrently with three dimensional (3D) conformal radiotherapy or IMRT for patients with locally advanced non-small cell lung cancer (stage IIIa and IIIb) after 2-4 cycles of induction chemotherapy. METHODS: Fourteen patients with histological/cytological proven stage III non-small-cell lung cancer were eligible. 3D or IMRT radiotherapy (60-70Gy in 30-35 fractions, 6-7weeks, 2 Gy/fraction) was delivered concurrently with cisplatin and docetaxel, 2 cycles during concurrent chemoradiotherapy (CCRT). The level I dosage was composed of 56 mg/m(2) DCT, on day 1 and 28mg/m(2) DDP, on day 1 and day 2. The level II was composed of 60 mg/m(2) DCT, on day 1 and 30 mg/ m(2) DDP, on day 1 and day 2. The level III was composed of 64 mg/m(2) DCT, on day 1 and 32 mg/ m(2) DDP, on day 1 and day 2. RESULTS: Fourteen patients were allocated and finished concurrent chemoradiotherapy. The dose-limiting neutropenia was at the dose Level III (64 mg/m(2)) and occurred in 2 of 5 patients. No dose limiting non-hematologic or hematologic toxicity occurred in the other patients. CONCLUSIONS: Patients with locally advanced non-small cell lung cancer may tolerate 60mg/m(2) docetaxel and 60mg/m(2) cisplatin for 2 cycles during concurrent radiotherapy after 2-3 cycles of induction chemotherapy.

[Expression and activity of RhoA/Rho kinase in remodeling of high blood flow pulmonary vessels.].

OBJECTIVE: High pulmonary blood flow induced pulmonary hypertension (PH) is often associated with increased vasoconstriction and deteriorating pulmonary artery remodeling, of which the exact mechanism has not been completely elucidated. The involvement of RhoA/Rho-kinase pathway has been demonstrated in the pathogenesis of hypoxia and monocrotaline induced PH. Thus the purpose of this study was to test whether RhoA/Rho-kinase pathway is involved in the process of high pulmonary flow induced pulmonary artery remodeling in rats. METHODS: Wistar rats aged 4 weeks in the shunt group underwent left common carotid artery-external jugular vein shunt operation, those in control group received sham-operation. At weeks 1, 2, 4 and 8 of the study, rats underwent right ventricular systolic pressure (RVSP) measurement; blood gases were analyzed to calculate Qp/Qs. The morphologic alterations of the pulmonary arteries were observed under optical microscope. The mean percentage of media wall thickness (%MT) was also measured to assess the extent of medial wall thickness of moderate size pulmonary arteries. Proliferating smooth muscle cells (SMCs) were evaluated by proliferating cell nuclear antigen (PCNA) immunohistochemical staining. Apoptotic SMCs were detected by TUNEL method. RhoA activity in pulmonary arteries was detected using pull down assay. Rho kinase activity was quantified by the extent of MYPT1 phosphorylation with Western blotting. The expression of RhoA and Rho kinase (ROCK2) was also detected with Western blotting. RESULTS: Carotid artery-jugular vein shunt resulted in high pulmonary blood flow, of all rats in shunted groups, the mean Qp/Qs was 2.26 +/- 0.35, which were all considered large shunts. Compared with the control group, RVSP in shunt group increased significantly at both week 1 and week 8 (t = 8.799, t = 5.332, respectively, P < 0.01). Compared with the control group, moderate pulmonary artery medial wall thickening characterized by SMCs hyper-proliferation and hypertrophy in shunted group was firstly appeared at week 4 and became more significant at week 8, as indicated by MT% (t = 9.192, t = 11.185, respectively, P < 0.01). Compared with the control group, the percentage of PCNA-positive SMCs in shunted group increased significantly at week 1 (t = 2.438, P < 0.05), and reached the maximal level at week 2 (t = 7.213, P < 0.01), then, it decreased to a level significantly lower than that of the control group at week 4 (t = 4.183, P < 0.01), and continued to decrease to so low a level that proliferative SMCs was scarcely observed at week 8 (t = 6.152, P < 0.01). The percentage of TUNEL-positive SMCs decreased significantly compared with the control group at week 2 (t = 2.418, P < 0.05), and continued to decrease to a level that apoptotic SMCs was scarcely observed at week 8 (t = 4.582, P < 0.01). Compared with the control group, the expression of RhoA and ROCK2 increased significantly at week 1 (t = 6.056, t = 8.411, respectively, P < 0.01), and reached the maximal level at week 2 (t = 9.342, t = 10.437, respectively, P < 0.01), then began to decrease at week 4, however, both of them were still significantly higher than those of the control group at week 8 (t = 4.743, t = 4.455, respectively, P < 0.01). In line with the expression of RhoA and ROCK2, both RhoA and Rho kinase activity of shunted group increased significantly compared with the control group at week 1 (t = 10.246, t = 19.110, respectively, P < 0.01), and reached the maximal level at week 4 (t = 24.984, t = 16.124, respectively, P < 0.01), then decreased, however, both of them were still higher than those of the control group at week 8 (t = 4.934, t = 10.426, respectively, P < 0.01). CONCLUSION: Activated RhoA/Rho-kinase pathway is associated with both high pulmonary blood flow induced acute pulmonary vasoconstriction and chronic pulmonary artery remodeling in rats.

Inhibition of rho kinase attenuates high flow induced pulmonary hypertension in rats.

BACKGROUND: The RhoA/Rho kinase pathway may participate in the pathogenesis of hypoxia and monocrotaline induced pulmonary hypertension. This study tested whether RhoA/Rho kinase pathway is involved in the pathogenesis of high flow induced pulmonary hypertension in rats. METHODS: Male Wistar rats (4 weeks) were randomly divided into 4 shunt groups, 4 treated groups and 4 control groups. Shunt and treated groups underwent left common carotid artery/external jugular vein shunt operation. Control groups underwent sham operation. Treated groups received fasudil treatment and the others received same dose of saline. At weeks 1, 2, 4 and 8 of the study, right ventricular systolic pressure was measured and blood gases were analysed to calculate Qp/Qs. The weight ratio of right ventricle to left ventricle plus septum and the mean percentage of medial wall thickness in moderate sized pulmonary arteries were obtained. RhoA activity in pulmonary arteries was detected using Rho activity assay reagent. Rho kinase activity was quantified by the extent of MYPT1 phosphorylation with Western blot. Proliferating cells were evaluated using proliferating cell nuclear antigen immunohistological staining. RESULTS: Carotid artery/jugular vein shunt resulted in high pulmonary blood flow, both an acute and a chronic elevation of right ventricular systolic pressure, significant medial wall thickening characterized by smooth muscle cells proliferation, right ventricular hypertrophy and increased activation of RhoA and Rho kinase. Fasudil treatment lowered pulmonary artery systolic pressure, suppressed pulmonary artery smooth muscle cells proliferation, attenuated pulmonary artery medial wall thickening and inhibited right ventricular hypertrophy together with significant suppression of Rho kinase activity but not Rho activity. CONCLUSIONS: Activated RhoA/Rho kinase pathway is associated with both the acute pulmonary vasoconstriction and the chronic pulmonary artery remodelling of high flow induced pulmonary hypertension. Fasudil treatment could improve pulmonary hypertension by inhibiting Rho kinase activity.

A new modification of transanal Soave pull-through procedure for Hirschsprung's disease.

BACKGROUND: One stage transanal Soave pull-through procedure (TSPP) is a recent popular operation in the treatment of Hirschsprung's disease (HD). With no visible scar and a short hospital stay, it is well accepted by surgeons and mothers. In the conventional Soave procedure, a long rectal muscular cuff left for anocolic anastomosis might increase the incidence of postoperative enterocolitis and constipation. This study presents a modified transanal Soave pull-through procedure (MTSPP) which includes an oblique mucosectomy and an oblique anastomosis with a short split muscular cuff. METHODS: A review of two groups of HD patients was made: 112 underwent conventional transanal Soave procedure from 1999 to 2001 (group 1) and 140 underwent modified transanal Soave procedure from 2002 to 2004 (group 2). A comparison was made between the two groups on operative data and postoperative complications. The data included: age at the operation, operating time, blood loss, time to feeds and hospital stay, occurrence of postoperative enterocolitis or constipation, need for anal dilatation, postoperative bowel function and perianal skin problems. RESULTS: There was no significant difference between two groups with respect to age, gender, length of colon resected, operating time, blood loss and hospital stay. However occurrence of postoperative enterocolitis, constipation, anastomotic stricture and time needed for anal dilatation were evidently less in group 2 (MTSPP). The mean operating time in group 1 was (106 +/- 39) minutes with a range of 60 to 170 minutes; in group 2 was (101 +/- 36) minutes with a range of 66 to 190 minutes. The average length of the bowel resected in group 1 was (24 +/- 7) cm, range 15 to 58 cm; in group 2 was (26 +/- 8) cm, range 15 to 70 cm. Two patients, one in each group, required laparoscopic assistance because of long aganglionic colon. Another patient in group 2 required laparotomy because of total colonic aganglionosis. Postoperative complications in group 1 included: temporary perianal excoriation in 34 patients (26 were < 3 months of age), enterocolitis in 21, anastomotic stricture in 11, recurrent constipation in 12, cuff abscess in 1, anastomosis leak in 1, soiling in 3 and rectal prolapse in 1. In group 2 post operative complications included: transient perianal excoriation in 37 patients (30 were < 3 months of age), enterocolitis in 13, anastomotic stricture in 5, recurrent constipation in 6, anastomotic leak in 1, adhesive bowel obstruction in 1 and soiling in 4. Complete bowel continence was found in 97 children (86.6%) in group 1 and in 129 children (92.1%) in group 2 at one year followup after operation. CONCLUSIONS: Modified transanal Soave pull-through procedure for HD with oblique mucosectomy and anastomosis and a short split muscular cuff is a safe and feasible operation with low incidence of postoperative complication. It is an encouraging improvement of the conventional transanal Soave pull-through procedure. MTSPP is a preferable choice in the surgery of HD.