Alan D. Enriquez

Unique distributions of the gap junction proteins connexin29, connexin32, and connexin47 in oligodendrocytes

Oligodendrocytes of adult rodents express three different connexins: connexin29 (Cx29), Cx32, and Cx47. In this study, we show that Cx29 is localized to the inner membrane of small myelin sheaths, whereas Cx32 is localized on the outer membrane of large myelin sheaths; Cx29 does not colocalize with Cx32 in gap junction plaques. All oligodendrocytes appear to express Cx47, which is largely restricted to their perikarya. Cx32 and Cx47 are colocalized in many gap junction plaques on oligodendrocyte somata, particularly in gray matter. Cx45 is detected in the cerebral vasculature, but not in oligodendrocytes or myelin sheaths. This diversity of connexins in oligodendrocytes (in different populations of cells and in different subcellular compartments) likely reflects functional differences between these connexins and perhaps the oligodendrocytes themselves.

Loss-of-function GJA12/Connexin47 mutations cause Pelizaeus–Merzbacher-like disease

Recessive mutations in GJA12/Cx47, the gene encoding the gap junction protein connexin47 (Cx47), cause Pelizaeus-Merzbacher-like disease (PMLD), which is characterized by severe CNS dysmyelination. Three missense PMLD mutations, P87S, Y269D and M283T, were expressed in communication-incompetent HeLa cells, and in each case the mutant proteins appeared to at least partially accumulate in the ER. Cells expressing each mutant did not pass Lucifer Yellow or neurobiotin in scrape loading assays, in contrast to robust transfer in cells expressing wild type Cx47. Dual whole-cell patch clamping of transfected Neuro2A cells demonstrated that none of the mutants formed functional channels, in contrast to wild type Cx47. Immunostaining sections of primate brains demonstrated that oligodendrocytes express Cx47, which is primarily localized to their cell bodies. Thus, the Cx47 mutants associated with PMLD likely disrupt the gap junction coupling between astrocytes and oligodendrocytes.

Cx29 and Cx32, two connexins expressed by myelinating glia, do not interact and are functionally distinct.

In rodents, oligodendrocytes and myelinating Schwann cells express connexin32 (Cx32) and Cx29, which have different localizations in the two cell types. We show here that, in contrast to Cx32, Cx29 does not form gap junction plaques or functional gap junctions in transfected cells. Furthermore, when expressed together, Cx29 and Cx32 are not colocalized and do not coimmunoprecipitate. To determine the structural basis of their divergent behavior, we generated a series of chimeric Cx32‐Cx29 proteins by exchanging their intracellular loops and/or their C‐terminal cytoplasmic tails. Although some chimerae reach the cell membrane, others appear to be largely localized intracellularly; none form gap junction plaques or functional gap junctions. Substituting the C‐terminus or the intracellular loop and the C‐terminus of Cx32 with those of Cx29 does not disrupt their colocalization or coimmunoprecipitation with Cx32. Substituting the C‐terminus of Cx29 with that of Cx32 does not disrupt the coimmunoprecipitation or the colocalization with Cx29, whereas substituting both the intracellular loop and the C‐terminus of Cx32 with those of Cx29 diminishes the coimmunoprecipitation with Cx29. Conversely, the Cx32 chimera that contains the intracellular loop of Cx29 coimmunoprecipitates with Cx29, indicating that the intracellular loop participates in Cx29‐Cx29 interactions. These data indicate that homomeric interactions of Cx29 and especially Cx32 largely require other domains: the N‐terminus, transmembrane domains, and extracellular loops. Substituting the intracellular loop and/or tail of Cx32 with those of Cx29 appears to prevent Cx32 from forming functional gap junctions.

The Role of Implantable Cardioverter-Defibrillators in Patients With Continuous Flow Left Ventricular Assist Devices

Background—The prognosis for patients experiencing ventricular arrhythmias (VAs) while on continuous flow left ventricular assist device (LVAD) support has not been well elucidated. Accordingly, the role of implantable cardioverter-defibrillators (ICDs) in this patient population remains undefined. Methods and Results—Records of 106 consecutive patients undergoing implantation of the HeartMate II LVAD at a single center were reviewed. For patients surviving >30 days postimplant (98 patients), the impact of VAs and ICDs on survival was analyzed. Mean age was 56.6±11.4 years, 82.1% were male, 42.5% had an ischemic cardiomyopathy, 87.7% were bridge to transplantation, and median length of support was 217 days. Twenty-one (19.8%) patients died, 60 (56.6%) survived to transplantation, and 25 patients (23.6%) reached the end of study, had the LVAD explanted, or were lost to follow-up. Post-LVAD VAs occurred in 37 patients (34.9%) but were not associated with increased mortality (hazard ratio, 0.58 [0.18–1.90]). Sixty-two (63.3%) patients had an active ICD, and 36 (36.7%) patients had no ICD or an inactivated ICD post-LVAD. Patients with an ICD were more likely to be INTERMACS (Interagency Registry for Mechanically Assisted Circulatory Support) level 3 or 4 at the time of implant (54.8% versus 33.3%; P=0.04). An appropriate shock was delivered in 27.3% of patients, but the presence of an active ICD was not associated with improved survival (hazard ratio, 1.12 [0.37–3.35]). Conclusions—VAs are common in patients with continuous flow LVADs. Although some episodes may be clinically significant, VAs are not associated with a worse prognosis, and concomitant ICDs in these patients may not reduce mortality.

Human oligodendrocytes express Cx31.3: function and interactions with Cx32 mutants

Murine oligodendrocytes express the gap junction (GJ) proteins connexin32 (Cx32), Cx47, and Cx29. CNS phenotypes in patients with X-linked Charcot-Marie-Tooth disease may be caused by dominant effects of Cx32 mutations on other connexins. Here we examined the expression of Cx31.3 (the human ortholog of murine Cx29) in human brain and its relation to the other oligodendrocyte GJ proteins Cx32 and Cx47. Furthermore, we investigated in vitro whether Cx32 mutants with CNS manifestations affect the expression and function of Cx31.3. Cx31.3 was localized mostly in the gray matter along small myelinated fibers similar to Cx29 in rodent brain and was co-expressed with Cx32 in a subset of human oligodendrocytes. In HeLa cells Cx31.3 was localized at the cell membrane and appeared to form hemichannels but no GJs. Cx32 mutants with CNS manifestations were retained intracellularly, but did not alter the cellular localization or function of co-expressed Cx31.3. Thus, Cx31.3 shares many characteristics with its ortholog Cx29. Cx32 mutants with CNS phenotypes do not affect the trafficking or function of Cx31.3, and may have other toxic effects in oligodendrocytes.

Contemporary Management of Arrhythmias During Pregnancy

Cardiac arrhythmias are among the most common cardiac complications encountered during pregnancy.1 In some, pregnancy may trigger exacerbations of pre-existing arrhythmias, whereas in others arrhythmias may manifest for the first time.2 Fortunately, severe arrhythmias requiring aggressive or invasive therapies are rare. There are unfortunately few randomized studies, little data on the efficacy or safety of antiarrhythmic drugs (AADs), or even explicit guidelines to support decision making on pregnant women with arrhythmias. Thus, much of the clinical care is guided by knowledge of the physiology of pregnancy and educated risk/benefit decisions made in collaboration with high-risk obstetric colleagues in Maternal-Fetal Medicine, as well as with the patient. The precise mechanism of increased arrhythmia burden during pregnancy is unclear, but it is likely because of a combination of hemodynamic, hormonal, and autonomic changes. Increases in effective circulating blood volume of 30% to 50% are seen beginning at 8 weeks of gestation and peaking at ≈34 weeks. Cardiac output is increased as well, with an average of 6.7 L/min in the first trimester and ≤8.7 L/min in the third trimester. This is the result of a 35% increase in stroke volume and a 15% increase in heart rate. The increase in plasma volume causes stretching of atrial and ventricular myocytes, and this may result in early after depolarizations, shortened refractoriness, slowed conduction, and spatial dispersion through activation of stretch-activated ion channels.3,4 A larger heart can also potentially sustain re-entry more easily because of an increase in path length of potential reentrant circuits. The increase in heart rate during pregnancy, seen predominantly in the third trimester, may also predispose to arrhythmia, as a high resting heart rate has been associated with markers of arrhythmogenesis.5 Hormonal and autonomic changes may also contribute to arrhythmogenesis. Estradiol and progesterone have been …

Needle-in-needle epicardial access: Preliminary observations with a modified technique for facilitating epicardial interventional procedures.

to flex over the course of insertion, and tactile assessment of force and perception of cardiac motion was limited. In this report, we describe our initial experience with the “needle-inneedle” (NIN) technique of epicardial access in which a short 18G needle is inserted under the sternum and the long 21G needle is inserted through the 18G needle. It is hoped that this technique improves the stability of the small needle and potential tactile feedback, thus reducing the risk of significant pericardial bleeding. We report preliminary results of its procedural success and acute complications compared with the standard Sosa technique, 1 which uses a 17G or 18G Touhy needle.

Cost-effectiveness of suppressing hepatitis B virus DNA in immune tolerant patients to prevent hepatocellular carcinoma and cirrhosis

Background  For patients with hepatitis B virus (HBV) infection in the immune tolerant phase, the current standard of care is to not offer treatment. However, the recent Risk Evaluation of the Viral Load Elevation and Associated Liver Disease/Cancer‐In study results show a striking relationship between high HBV DNA levels and risk for hepatocellular carcinoma and cirrhosis.

Impact of general anesthesia on initiation and stability of VT during catheter ablation

BACKGROUND Radiofrequency ablation of ventricular tachycardia (VT) may be performed with general anesthesia (GA) or conscious sedation; however, comparative data are limited. OBJECTIVE The purpose of the study was to assess the effects of GA on VT inducibility and stability. METHODS A retrospective comparison of 226 patients undergoing radiofrequency ablation for scar-related VT under GA or intravenous conscious sedation was performed. Data were then prospectively collected in 73 patients undergoing noninvasive programmed stimulation (NIPS) while awake, followed by GA and invasive programmed stimulation for VT induction. RESULTS In the retrospective study, groups did not differ in VT inducibility, complications, or abolition of clinical VT. Intravenous hemodynamic support was used more often in the GA group. In the prospective group, 12 patients (16%) were noninducible with NIPS. Of the 61 patients with inducible VT with NIPS, 5 (8%) were noninducible with GA, 25 (41%) were inducible with more aggressive simulation, and 31 (51%) were inducible with the same or less aggressive stimulation. Of the 56 patients who were inducible with NIPS and under GA, 28 (50%) had the same induced VTs and 28 (50%) had different induced VTs. In 23 of 56 patients, the clinical VT morphology was known. The clinical VT was reproduced with NIPS in 17 of 23 patients (74%) and under GA in 13 of 23 patients (59%). Under GA, nonclinical VTs were more often induced in patients with a lower ejection fraction and nonischemic cardiomyopathy. CONCLUSION GA does not prevent inducible VT in the majority of patients. GA is associated with an increased use of hemodynamic support, but this did not adversely affect VT stability or procedure outcomes.

Correlates and Prognosis of Early Recurrence After Catheter Ablation for Ventricular Tachycardia due to Structural Heart Disease

Background—Catheter ablation for ventricular tachycardia (VT) from structural heart disease has a significant risk of recurrence, but the optimal duration for in-hospital monitoring is not defined. This study assesses the timing, correlates, and prognostic significance of early VT recurrence after ablation. Methods and Results—Of 370 patients (313 men; aged 63.0±13.2 years) who underwent a first radiofrequency ablation for sustained monomorphic VT associated with structural heart disease from 2008 to 2012, sustained VT recurred in 81 patients (22%) within 7 days. In multivariable analysis, early recurrence was associated with New York Heart Association classification ≥III (odds ratio [OR] 1.90, 95% confidence interval [CI] 1.03–3.48; P=0.04), dilated cardiomyopathy (OR 1.93, 95% CI 1.03–3.57; P=0.04), prevalence of VT storm before the procedure (OR 2.62, 95% CI 1.48–4.65; P=0.001), a greater number of induced VTs (OR 1.24, 95% CI 1.07–1.45; P=0.006), and acute failure or no final induction test (OR 1.88, 95% CI 1.03–3.40; P=0.04). During a median of 2.5 (1.2, 4.0) years of follow-up, early VT recurrence was an independent correlates of mortality (hazard ratio 2.59, 95% CI 1.52–4.34; P=0.0005). Conclusions—Patients who have early recurrences of VT after ablation are a high risk group who may be identifiable from their clinical profile. Further study is warranted to define the optimal treatment strategies for this patient group.