Identification of binding partners for the cytoplasmic loop of connexin43: a novel interaction with ß-tubulin.

Connexin43 (Cx43), a component of gap junctions, has a relatively large carboxy-terminal region with multiple proteomic interactions. Proteomic interactions with its cytoplasmic loop, however, are poorly defined. The goal of this study is to examine proteomic interactions involving the cytoplasmic loop (CL) of Cx43. The authors utilized various techniques, including glutathione-S-transferase (GST) pull-down, immunoblot analysis, two-dimensional (2D) gel electrophoresis, and mass spectrometry, to elucidate binding partners for Cx43-CL. The authors identified novel interactions with Cx43-CL involving α- and ß-tubulin, myelin basic protein, and Purα. Because tubulin interacts with the C-terminus of Cx43 (Cx43-CT), the authors further investigated the nature of the interaction between ß-tubulin and Cx43-CL. ß-Tubulin binds with the full length of Cx43-CL with approximately one-fifth the affinity of the interaction between Cx43-CT and ß-tubulin. This study demonstrates novel proteomic interactions involving Cx43-CL that may lead to a more complete understanding of trafficking and gating of gap junction channels.

Decreased connexin43 expression in the mouse heart potentiates pacing-induced remodeling of repolarizing currents.

Gap junction redistribution and reduced expression, a phenomenon termed gap junction remodeling (GJR), is often seen in diseased hearts and may predispose toward arrhythmias. We have recently shown that short-term pacing in the mouse is associated with changes in connexin43 (Cx43) expression and localization but not with increased inducibility into sustained arrhythmias. We hypothesized that short-term pacing, if imposed on murine hearts with decreased Cx43 abundance, could serve as a model for evaluating the electrophysiological effects of GJR. We paced wild-type (normal Cx43 abundance) and heterozygous Cx43 knockout (Cx43+/-; 66% mean reduction in Cx43) mice for 6 h at 10-15% above their average sinus rate. We investigated the electrophysiological effects of pacing on the whole animal using programmed electrical stimulation and in isolated ventricular myocytes with patch-clamp studies. Cx43+/- myocytes had significantly shorter action potential durations (APD) and increased steady-state (Iss) and inward rectifier (I(K1)) potassium currents compared with those of wild-type littermate cells. In Cx43+/- hearts, pacing resulted in a significant prolongation of ventricular effective refractory period and APD and significant diminution of Iss compared with unpaced Cx43+/- hearts. However, these changes were not seen in paced wild-type mice. These data suggest that Cx43 abundance plays a critical role in regulating currents involved in myocardial repolarization and their response to pacing. Our study may aid in understanding how dyssynchronous activation of diseased, Cx43-deficient myocardial tissue can lead to electrophysiological changes, which may contribute to the worsened prognosis often associated with pacing in the failing heart.

Short-term pacing in the mouse alters cardiac expression of connexin43.

BACKGROUND: Cardiac insults such as ischemia, infarction, hypertrophy and dilatation are often accompanied by altered abundance and/or localization of the connexin43 gap junction protein, which may predispose towards arrhythmic complications. Models of chronic dyssynchronous cardiac activation have also been shown to result in redistribution of connexin43 in cardiomyocytes. We hypothesized that alterations in connexin43 expression and localization in the mouse heart might be induced by ventricular pacing over a short period of time. RESULTS: The subdiaphragmatic approach was used to pace a series of wild type mice for six hours before the hearts were removed for analysis. Mice were paced at 10-15% above their average anesthetized sinus rate and monitored to ensure 1:1 capture. Short-term pacing resulted in a significant reduction in connexin43 mRNA abundance, a partial redistribution of connexin43 from the sarcolemma to a non-sarcolemmal fraction, and accumulation of ubiquitinated connexin43 without a significant change in overall connexin43 protein levels. These early pacing-induced changes in connexin43 expression were not accompanied by decreased cardiac function, prolonged refractoriness or increased inducibility into sustained arrhythmias. CONCLUSION: Our data suggest that short-term pacing is associated with incipient changes in the expression of the connexin43 gap junction, possibly including decreased production and a slowed rate of degradation. This murine model may facilitate the study of early molecular changes induced by pacing and may ultimately assist in the development of strategies to prevent gap junction remodeling and the associated arrhythmic complications of cardiac disease.

Experience with endovascular abdominal aortic aneurysm repair in nonagenarians.

PURPOSE: To report a single-institution experience with endovascular abdominal aortic aneurysm (AAA) repair (EVAR) in nonagenarians. METHODS: A retrospective review was performed of all patients >90 years old undergoing EVAR over an 8-year period at a major academic medical center. The patient population was investigated for the presence of various comorbidities, initial aneurysm size, successful aneurysm exclusion, perioperative complications, disposition, endoleaks, secondary interventions, and overall survival. RESULTS: EVAR was performed in 18 male nonagenarians (mean age 92.4 years, range 90- 95). Mean aneurysm diameter was 7.3 cm (range 5.5-9.8). The cohort had an average of 3.2 comorbid conditions. Sixteen patients were treated electively, while 2 patients underwent emergent repair for contained rupture and bleeding aortoenteric fistula, respectively. Immediate technical success was 100%. Perioperative local/vascular complications occurred in 4 (22%) patients. Perioperative systemic complications occurred in 3 (17%) patients. There were 2 (11%) perioperative (<30 days) deaths. Three (17%) patients required secondary interventions. Mean survival in patients who expired during the follow-up period beyond the first 30 days was 34 months (range 8-78). Mean survival in 8 patients who are still alive is 17.4 months (range 9-39). CONCLUSION: Endovascular AAA repair in nonagenarians is associated with a high rate of technical success and relatively low morbidity rate. Survival times following successful hospital discharge are significant. Suitable patients over 90 years of age may benefit from an endovascular AAA repair.