Anti-Ischemic Effects of PIK3IP1 Are Mediated through Its Interactions with the ETA-PI3Kγ-AKT Axis.

Oxidative stress, caused by the accumulation of reactive oxygen species (ROS) during acute myocardial infarction (AMI), is one of the main factors leading to myocardial cell damage and programmed cell death. Phosphatidylinositol-3-kinase-AKT (PI3K-AKT) signaling is essential for regulating cell proliferation, differentiation, and apoptosis. Phosphoinositide-3-kinase (PI3K)-interacting protein 1 (PIK3IP1) is an intrinsic inhibitor of PI3K in various tissues, but its functional role during AMI remains unknown. In this study, the anti-ischemic role of PIK3IP1 in an in vitro AMI setting was evaluated using H9c2 cells. The MTT assay demonstrated that cell viability decreased significantly via treatment with H2O2 (200-500 µM). The TUNEL assay results revealed substantial cellular apoptosis following treatment with 200 µM H2O2. Under the same conditions, the expression levels of hypoxia-inducible factor (HIF-1α), endothelin-1 (ET-1), bcl-2-like protein 4 (BAX), and cleaved caspase-3 were elevated, whereas those of PIK3IP1, LC3II, p53, and Bcl-2 decreased significantly. PIK3IP1 overexpression inhibited H2O2-induced and PI3K-mediated apoptosis; however, PIK3IP1 knockdown reversed this effect, suggesting that PIK3IP1 functions as an anti-apoptotic molecule. To identify both the upstream and downstream molecules associated with PIK3IP1, ET-1 receptor type-specific antagonists (BQ-123 and BQ-788) and PI3K subtype-specific antagonists (LY294002 and IPI-549) were used to determine the participating isoforms. Co-immunoprecipitation was performed to identify the binding partners of PIK3IP1. Our results demonstrated that ROS-induced cardiac cell death may occur through the ETA-PI3Kγ-AKT axis, and that PIK3IP1 inhibits binding with both ETA and PI3Kγ. Taken together, these findings reveal that PIK3IP1 plays an anti-ischemic role by reducing the likelihood of programmed cell death via interaction with the ETA-PI3Kr-AKT axis.

Quantitative proteomic analyses reveal that GPX4 downregulation during myocardial infarction contributes to ferroptosis in cardiomyocytes.

Ischaemic heart disease (IHD) is the leading cause of death worldwide. Although myocardial cell death plays a significant role in myocardial infarction (MI), its underlying mechanism remains to be elucidated. To understand the progression of MI and identify potential therapeutic targets, we performed tandem mass tag (TMT)-based quantitative proteomic analysis using an MI mouse model. Gene ontology (GO) analysis and gene set enrichment analysis (GSEA) revealed that the glutathione metabolic pathway and reactive oxygen species (ROS) pathway were significantly downregulated during MI. In particular, glutathione peroxidase 4 (GPX4), which protects cells from ferroptosis (an iron-dependent programme of regulated necrosis), was downregulated in the early and middle stages of MI. RNA-seq and qRT-PCR analyses suggested that GPX4 downregulation occurred at the transcriptional level. Depletion or inhibition of GPX4 using specific siRNA or the chemical inhibitor RSL3, respectively, resulted in the accumulation of lipid peroxide, leading to cell death by ferroptosis in H9c2 cardiomyoblasts. Although neonatal rat ventricular myocytes (NRVMs) were less sensitive to GPX4 inhibition than H9c2 cells, NRVMs rapidly underwent ferroptosis in response to GPX4 inhibition under cysteine deprivation. Our study suggests that downregulation of GPX4 during MI contributes to ferroptotic cell death in cardiomyocytes upon metabolic stress such as cysteine deprivation.

A novel system-level approach using RNA-sequencing data identifies miR-30-5p and miR-142a-5p as key regulators of apoptosis in myocardial infarction.

This study identified microRNAs involved in myocardial infarction (MI) through a novel system-level approach using RNA sequencing data in an MI mouse model. This approach involved the extraction of DEGs and DEmiRs from RNA-seq data in sham and MI samples and the subsequent selection of two miRNAs: miR-30-5p (family) and miR-142a-5p, which were downregulated and upregulated in MI, respectively. Gene Set Enrichment Analysis (GSEA) using the predicted targets of the two miRNAs suggested that apoptosis is an essential gene ontology (GO)-associated term. In vitro functional assays using neonatal rat ventricular myocytes (NRVMs) demonstrated that miR-30-5p is anti-apoptotic and miR-142a-5p is pro-apoptotic. Luciferase assays showed that the apoptotic genes, Picalm and Skil, and the anti-apoptotic genes, Ghr and Kitl, are direct targets of miR-30-5p and miR-142a-5p, respectively. siRNA studies verified the results of the luciferase assays for target validation. The results of the system-level high throughput approach identified a pair of functionally antagonistic miRNAs and their targets in MI. This study provides an in-depth analysis of the role of miRNAs in the pathogenesis of MI which could lead to the development of therapeutic tools. The system-level approach could be used to identify miRNAs involved in variety of other diseases.

miR-374 promotes myocardial hypertrophy by negatively regulating vascular endothelial growth factor receptor-1 signaling.

Vascular endothelial growth factor (VEGF) is an essential cytokine that has functions in the formation of new blood vessels and regression of cardiac hypertrophy. VEGF/VEGF-receptor-1 (VEGFR1) signaling plays a key role in the regression of cardiac hypertrophy, whereas VEGF/VEGFR2 signaling leads to cardiac hypertrophy. In this study, we identified the prohypertrophic role of miR-374 using neonatal rat ventricular myocytes (NRVMs). Our results showed that overexpression of miR-374 activated G protein-coupled receptor-mediated prohypertrophic pathways by the inhibition of VEGFR1-dependent regression pathways. Luciferase assays revealed that miR-374 could directly target the 3'-untranslated regions of VEGFR1 and cGMP-dependent protein kinase-1. Collectively, these findings demonstrated that miR-374 was a novel pro-hypertrophic microRNA functioning to suppress the VEGFR1-mediated regression pathway. [BMB Reports 2017; 50(4): 208-213].

MicroRNA-101b attenuates cardiomyocyte hypertrophy by inhibiting protein kinase C epsilon signaling.

Previously, a surgical regression model identified microRNA-101b (miR-101b) as a potential inhibitor of cardiac hypertrophy. Here, we investigated the antihypertrophic mechanism of miR-101b using neonatal rat ventricular myocytes. miR-101b markedly suppressed agonist-induced cardiac hypertrophy as shown by cell size and fetal gene expression. By systems biology approaches, we identified protein kinase C epsilon (PKCε) as the major target of miR-101b. Our results from qRT-PCR, western blot, and luciferase reporter assays confirm that PKCε is a direct target of miR-101b. In addition, we found that effectors downstream of PKCε (p-AKT, p-ERK1/2, p-NFAT, and p-GSK3ß) are also affected by miR-101b. Our study reveals a novel inhibitory mechanism for miR-101b as a negative regulator of cardiac hypertrophy.

Incomplete colonoscopy: maximizing completion rates of gastroenterologists.

BACKGROUND: Cecal intubation is one of the goals of a quality colonoscopy; however, many factors increasing the risk of incomplete colonoscopy have been implicated. The implications of missed pathology and the demand on health care resources for return colonoscopies pose a conundrum to many physicians. The optimal course of action after incomplete colonoscopy is unclear. OBJECTIVES: To assess endoscopic completion rates of previously incomplete colonoscopies, the methods used to complete them and the factors that led to the previous incomplete procedure. METHODS: All patients who previously underwent incomplete colonoscopy (2005 to 2010) and were referred to St Paul's Hospital (Vancouver, British Columbia) were evaluated. Colonoscopies were re-attempted by a single endoscopist. Patient charts were reviewed retrospectively. RESULTS: A total of 90 patients (29 males) with a mean (± SD) age of 58 ± 13.2 years were included in the analysis. Thirty patients (33%) had their initial colonoscopy performed by a gastroenterologist. Indications for initial colonoscopy included surveillance or screening (23%), abdominal pain (15%), gastrointestinal bleeding (29%), change in bowel habits or constitutional symptoms (18%), anemia (7%) and chronic diarrhea (8%). Reasons for incomplete colonoscopy included poor preparation (11%), pain or inadequate sedation (16%), tortuous colon (30%), diverticular disease (6%), obstructing mass (6%) and stricturing disease (10%). Reasons for incomplete procedures in the remaining 21% of patients were not reported by the referring physician. Eighty-seven (97%) colonoscopies were subsequently completed in a single attempt at the institution. Seventy-six (84%) colonoscopies were performed using routine manoeuvres, patient positioning and a variable-stiffness colonoscope (either standard or pediatric). A standard 160 or 180 series Olympus gastroscope (Olympus, Japan) was used in five patients (6%) to navigate through sigmoid diverticular disease; a pediatric colonoscope was used in six patients (7%) for similar reasons. Repeat colonoscopy on the remaining three patients (3%) failed: all three required surgery for strictures (two had obstructing malignant masses and one had a severe benign obstructing sigmoid diverticular stricture). CONCLUSION: Most patients with previous incomplete colonoscopy can undergo a successful repeat colonoscopy at a tertiary care centre with instruments that are readily available to most gastroenterologists. Other modalities for evaluation of the colon should be deferred until a second attempt is made at an expert centre.

Endoscopic frontal sinusotomy-preventing recurrence or a route to revision?

OBJECTIVES/HYPOTHESIS: The Messerklinger technique is an endoscopic approach to sinus surgery designed to be minimally invasive and preserve mucosa and therefore physiological function. More recently there have been advocates for more radical endoscopic approaches to the frontal sinus such as the modified Lothrop procedure. This study aims to determine the effectiveness of endoscopic frontal sinusotomy in preventing recurrent frontal sinus disease and the need for any revision frontal sinus surgery. STUDY DESIGN: Retrospective data review. METHODS: A retrospective review of the chronic rhinosinusitis database at St. Paul's Sinus Centre was performed, randomly selecting 200 patients who had undergone primary bilateral functional endoscopic sinus surgery between 2000 and 2009. Any endoscopic or radiological recurrences listed on the database were counted along with the number of cases returned to theater for revision surgery. The preoperative Lund-Mackay score was also extracted from the database. RESULTS: In the 200 patients who had undergone their primary surgery at St. Paul's Hospital, the recurrence rate of frontal sinus disease was 19%, with less than one half (8%) requiring revision surgery. The Lund-Mackay scores showed no correlation between disease severity and the incidence of recurrence (P = .35), and there was no difference between polyp and nonpolyp forms of chronic rhinosinusitis (P = .14). A comparison with 100 patients in the database who had received their primary surgery at another center showed that the revision patients had a recurrence rate of 34% and a revision rate of 21%. The patients who did not receive surgical revision were treated satisfactorily with topical medications in the outpatient clinic. CONCLUSIONS: Meticulously performed endoscopic frontal sinusotomy with computer guidance appears to be an effective minimally invasive procedure to treat chronic frontal sinusitis secondary to outflow tract obstruction. Properly performed, it is effective in dealing with the most diseased frontal sinus. It offers clear advantages in reducing complications and recurrence rates in frontal sinus disease even in revision cases.