β-adrenergic receptors (βARs) are critical regulators of acute cardiovascular physiology. In response to elevated catecholamine stimulation during development of congestive heart failure (CHF), ...chronic activation of Gs-dependent β₁AR and Gi-dependent β₂AR pathways leads to enhanced cardiomyocyte death, reduced β₁AR expression, and decreased inotropic reserve. β-blockers act to block excessive catecholamine stimulation of βARs to decrease cellular apoptotic signaling and normalize β₁AR expression and inotropy. Whereas these actions reduce cardiac remodeling andmortality outcomes, the effects are not sustained. Converse to G-protein–dependent signaling, β-arrestin–dependent signaling promotes cardiomyocyte survival. Given that β₂AR expression is unaltered in CHF, a β-arrestin–biased agonist that operates through the β₂AR represents a potentially useful therapeutic approach. Carvedilol, a currently prescribed nonselective β-blocker, has been classified as a β-arrestin–biased agonist that can inhibit basal signaling from βARs and also stimulate cell survival signaling pathways. To understand the relative contribution of β-arrestin bias to the efficacy of select β-blockers, a specific β-arrestin–biased pepducin for the β₂AR, intracellular loop (ICL)1–9, was used to decouple β-arrestin–biased signaling from occupation of the orthosteric ligand-binding pocket. With similar efficacy to carvedilol, ICL1–9 was able to promote β₂AR phosphorylation, β-arrestin recruitment, β₂AR internalization, and β-arrestin–biased signaling. Interestingly, ICL1–9 was also able to induce β₂AR- and β-arrestin–dependent and Ca2+-independent contractility in primary adult murine cardiomyocytes, whereas carvedilol had no efficacy. Thus, ICL1–9 is an effective tool to access a pharmacological profile stimulating cardioprotective signaling and inotropic effects through the β₂AR and serves as a model for the next generation of cardiovascular drug development.
We validated a modified risk algorithm based on the Asia-Pacific Colorectal Screening (APCS) score that included body mass index (BMI) for prediction of advanced neoplasia.
Among 5744 Chinese ...asymptomatic screening participants undergoing a colonoscopy in Hong Kong from 2008 to 2012, a random sample of 3829 participants acted as the derivation cohort. The odds ratios for significant risk factors identified by binary logistic regression analysis were used to build a scoring system ranging from 0 to 6, divided into "average risk" (AR): 0; "moderate risk" (MR): 1-2; and "high risk" (HR): 3-6. The other 1915 subjects formed a validation cohort, and the performance of the score was assessed.
The prevalence of advanced neoplasia in the derivation and validation cohorts was 5.4% and 6.0%, respectively (P = 0.395). Old age, male gender, family history of colorectal cancer, smoking, and BMI were significant predictors in multivariate regression analysis. A BMI cut-off at > 23 kg/m
had better predictive capability and lower number needed to screen than that of > 25 kg/m
. Utilizing the score developed, 8.4%, 57.4%, and 34.2% in the validation cohort were categorized as AR, MR, and HR, respectively. The corresponding prevalence of advanced neoplasia was 3.8%, 4.3%, and 9.3%. Subjects in the HR group had 2.48-fold increased prevalence of advanced neoplasia than the AR group. The c-statistics of the modified score had better discriminatory capability than that using predictors of APCS alone (c-statistics = 0.65 vs 0.60).
Incorporating BMI into the predictors of APCS score was found to improve risk prediction of advanced neoplasia and reduce colonoscopy resources.
Transient receptor potential channel melastatin 2 (TRPM2) is highly expressed in cancer and has an essential function in preserving viability through maintenance of mitochondrial function and ...antioxidant response. Here, the role of TRPM2 in cell survival was examined in neuroblastoma cells with TRPM2 deletion with CRISPR technology. Viability was significantly decreased in TRPM2 knockout after doxorubicin treatment. RNA sequence analysis and RT-qPCR revealed reduced RNAs encoding master transcription regulators FOXM1 and E2F1/2 and downstream cell cycle targets including Cyclin B1, CDK1, PLK1, and CKS1. CHIP analysis demonstrated decreased FOXM1 binding to their promoters. Western blotting confirmed decreased expression, and increased expression of CDK inhibitor p21, a CKS1 target. In cells with TRPM2 deletion, cell cycle progression to S and G2/M phases was reduced after treatment with doxorubicin. RNA sequencing also identified decreased DNA repair proteins in cells with TRPM2 deletion after doxorubicin treatment, and DNA damage was increased. Wild type TRPM2, but not Ca
-impermeable mutant E960D, restored live cell number and reconstituted expression of E2F1, FOXM1, and cell cycle/DNA repair proteins. FOXM1 expression alone restored viability. TRPM2 is a potential therapeutic target to reduce tumor proliferation and increase doxorubicin sensitivity through modulation of FOXM1, E2F1, and cell cycle/DNA repair proteins.
Background Valve repair for ischemic mitral regurgitation (IMR) with undersized annuloplasty rings is characterized by high IMR recurrence rates. Patient-specific preoperative imaging-based risk ...stratification for recurrent IMR would optimize results. We sought to determine if prerepair three-dimensional (3D) echocardiography combined with a novel valve-modeling algorithm would be predictive of IMR recurrence 6 months after repair. Methods Intraoperative transesophageal real-time 3D echocardiography was performed in 50 patients undergoing undersized ring annuloplasty for IMR and in 21 patients with normal mitral valves. A customized image analysis protocol was used to assess 3D annular geometry and regional leaflet tethering. IMR recurrence (≥ grade 2) was assessed with two-dimensional transthoracic echocardiography 6 months after repair. Results Preoperative annular geometry was similar in all IMR patients, and preoperative leaflet tethering was significantly higher in patients with recurrent IMR (n=13) than in patients in whom IMR did not recur (n=37) (tethering index: 3.91 ± 1.01 vs 2.90 ± 1.17, p = 0.008; tethering angles of A3: 23.5° ± 8.9° vs 14.4° ± 11.4°, p = 0.012; P2: 44.4° ± 8.8° vs 28.2° ± 17.0°, p = 0.002; and P3: 35.2° ± 6.0° vs. 18.6° ± 12.7°, p < 0.001). Multivariate logistic regression analysis revealed the preoperative P3 tethering angle as an independent predictor of IMR recurrence with an optimal cutoff value of 29.9° (area under the curve, 0.92; 95% confidence interval, 0.84 to 1.00; p < 0.001). Conclusions 3D echocardiography combined with valve modeling is predictive of recurrent IMR. Preoperative regional leaflet tethering of segment P3 is a strong independent predictor of IMR recurrence after undersized ring annuloplasty. In patients with a preoperative P3 tethering angle of 29.9° or larger, chordal-sparing valve replacement rather than valve repair should be strongly considered.
BAG3 is a cellular protein that is expressed predominantly in skeletal and cardiac muscle but can also be found in the brain and in the peripheral nervous system. BAG3 functions in the cell include: ...serving as a co-chaperone with members of the heat-shock protein family of proteins to facilitate the removal of misfolded and degraded proteins, inhibiting apoptosis by interacting with Bcl2 and maintaining the structural integrity of the Z-disk in muscle by binding with CapZ. The importance of BAG3 in the homeostasis of myocytes and its role in the development of heart failure was evidenced by the finding that single allelic mutations in BAG3 were associated with familial dilated cardiomyopathy. Furthermore, significant decreases in the level of BAG3 have been found in end-stage failing human heart and in animal models of heart failure including mice with heart failure secondary to trans-aortic banding and in pigs after myocardial infarction. Thus, it becomes relevant to understand the cellular biology and molecular regulation of BAG3 expression in order to design new therapies for the treatment of patients with both hereditary and non-hereditary forms of dilated cardiomyopathy.
G protein-coupled receptors (GPCRs) are central to many physiological processes. Regulation of this superfamily of receptors is controlled by GPCR kinases (GRKs), some of which have been implicated ...in heart failure. GSK180736A, developed as a Rho-associated coiled-coil kinase 1 (ROCK1) inhibitor, was identified as an inhibitor of GRK2 and co-crystallized in the active site. Guided by its binding pose overlaid with the binding pose of a known potent GRK2 inhibitor, Takeda103A, a library of hybrid inhibitors was developed. This campaign produced several compounds possessing high potency and selectivity for GRK2 over other GRK subfamilies, PKA, and ROCK1. The most selective compound, 12n (CCG-224406), had an IC50 for GRK2 of 130 nM, >700-fold selectivity over other GRK subfamilies, and no detectable inhibition of ROCK1. Four of the new inhibitors were crystallized with GRK2 to give molecular insights into the binding and kinase selectivity of this class of inhibitors.
Cardiovascular disease is a leading cause of co-morbidity in HIV-1 positive patients, even those in whom plasma virus levels are well-controlled. The pathogenic mechanism of HIV-1-associated ...cardiomyopathy is unknown, but has been presumed to be mediated indirectly, owing to the absence of productive HIV-1 replication in cardiomyocytes. We sought to investigate the effect of the HIV-1 auxiliary protein, Nef, which is suspected of extracellular release by infected CD4+ T cells on protein quality control and autophagy in cardiomyocytes. After detection of Nef in the serum of HIV-1 positive patients and the accumulation of this protein in human and primate heart tissue from HIV-1/SIV-infected cells we employed cell and molecular biology approaches to investigate the effect of Nef on cardiomyocyte-homeostasis by concentrating on protein quality control (PQC) pathway and autophagy. We found that HIV-1 Nef-mediated inhibition of autophagy flux leads to cytotoxicity and death of cardiomyocytes. Nef compromises autophagy at the maturation stage of autophagosomes by interacting with Beclin 1/Rab7 and dysregulating TFEB localization and cellular lysosome content. These effects were reversed by rapamycin treatment. Our results indicate that HIV-1 Nef-mediated inhibition of cellular PQC is one possible mechanism involved in the development of HIV-associated cardiomyopathy.
Transient receptor potential melastatin channel subfamily member 2 (TRPM2) has an essential role in protecting cell viability through modulation of oxidative stress. TRPM2 is highly expressed in ...cancer. When TRPM2 is inhibited, mitochondria are dysfunctional, ROS levels are increased, and cell viability is reduced. Here, the importance of NF-E2-related factor (Nrf2) in TRPM2-mediated suppression of oxidant stress was explored. In TRPM2 depleted cells, antioxidant cofactors glutathione, NADPH, and NADH were significantly reduced. Cytoplasmic and nuclear expression of Nrf2 and of IQGAP1, a modulator of Nrf2 stability regulated by intracellular calcium, were decreased. Antioxidant enzymes transcriptionally regulated by Nrf2 and involved in GSH, NADPH, and NADH generation were significantly lower including PRX1 and PRX3, GPX4, GSTP1, GCLC, and MTHFD2. The glutamine pathway leading to GSH production was suppressed, and ATP and GTP levels were impaired. Reconstitution with wild type TRPM2 or Nrf2, but not TRPM2 pore mutant E960D, rescued expression of enzymes downstream of Nrf2 and restored GSH and GTP. Cell viability, ROS, NADPH, NADH, and ATP levels were fully rescued by TRPM2 and partially by Nrf2. These data show that TRPM2 maintains cell survival following oxidative stress through modulation of antioxidant pathways and cofactors regulated by Nrf2.
Patients with heart failure and a non–left bundle branch block (non-LBBB) QRS pattern have a limited response to biventricular pacing (BVP).
A personalized cardiac resynchronization therapy (CRT) ...implantation approach guided by real-time electrocardiographic imaging (ECGi) was studied.
Twenty patients with left ventricular ejection fraction (LVEF) ≤ 35%, QRS duration ≥ 120 ms, and non-LBBB 13 (65%) with right bundle branch block and 7 (35%) with intraventricular conduction delay were recruited. During CRT implantation, right atrial, right ventricular, coronary sinus, His-bundle, and/or left bundle leads were inserted. The total activation time (TAT) with different pacing combinations were measured in real time during implantation by ECGi. The configuration producing the shortest TAT was chosen. Clinical response was defined as ≥1 New York Heart Association class improvement. Echocardiographic response was defined as left ventricular end-systolic volume reduction ≥ 15% and/or LVEF improvement ≥ 10% at 6 months.
After ECGi-guided CRT implantation, LVEF improved from 26% ± 6% to 34% ± 11% (P < .01) and New York Heart Association class improved from 3.0 ± 0.5 to 2.0 ± 0.6 (P < .01). Both clinical and echocardiographic response rates were 70%. The ECGi approach resulted in better acute electrical resynchronization over BVP as measured by TAT reduction (40% vs 14%; P < .01). The percentage of TAT reduction was found to be a strong predictor for echocardiographic response (area under the curve for the receiver operating characteristic curve 0.91; 95% confidence interval 0.78–1.00). A strong positive correlation between percentage TAT reduction and percentage LVEF improvement (Pearson R = 0.70; P = .001) was found.
ECGi-guided CRT implantation in patients with non-LBBB generates superior acute electrical resynchronization compared with BVP and is associated with favorable clinical and echocardiographic outcomes.
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The ubiquitin carboxyl‐terminal hydrolase L1 (UCHL1) is a carboxyl‐terminal ubiquitin hydrolase regulating cellular ubiquitin levels, recently suggested as a tumor suppressor. However, the role of ...UCHL1 in hepatocellular carcinoma (HCC) is not clear. We investigated the expression and DNA methylation of the UCHL1 in primary HCC, liver metastases from digestive carcinomas, and primary digestive cancers. UCHL1 is expressed in all normal tissues and immortalized normal epithelial cell lines, but was low or silenced in 77% (10/13) of HCC cell lines, which is well correlated with its promoter methylation status. Methylation was further detected in 44% (12/27) of HCCs, but less in metastatic tumors generated from colorectal and stomach in the liver (19%, 3/16; P < 0.05). Methylation was also detected in primary digestive tumors, including 71% (22/31) of colon, 77% (53/69) of gastric, and 40% (18/45) of esophageal carcinomas, but none or occasionally in paired adjacent nontumor tissues. Detailed methylation analysis of 49 CpG sites at a 540‐bp promoter region by bisulfite genomic sequencing confirmed the methylation. UCHL1 silencing could be reversed by chemical or genetic demethylation of the promoter, indicating direct epigenetic silencing. Restoring UCHL1 expression in silenced cell lines significantly inhibited their growth and colony formation ability by inhibiting cell proliferation, causing cell cycle arrest in G2/M phase and inducing apoptosis through the intrinsic caspase‐dependent pathway. Moreover, UCHL1 directly interacts with p53 and stabilizes p53 through the ubiquitination pathway. Conclusion: Epigenetic inactivation of UCHL1 is common in primary HCCs and other digestive tumors. UCHL1 appears to be a functional tumor suppressor involved in the tumorigenesis of HCCs and other digestive cancers. (HEPATOLOGY 2008;48:508–518.)
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