The effect of a population bottleneck on the evolution of genetic variance/covariance structure.

It is well known that standard population genetic theory predicts decreased additive genetic variance (V(a) ) following a population bottleneck and that theoretical models including interallelic and intergenic interactions indicate such loss may be avoided. However, few empirical data from multicellular model systems are available, especially regarding variance/covariance (V/CV) relationships. Here, we compare the V/CV structure of seventeen traits related to body size and composition between control (60 mating pairs/generation) and bottlenecked (2 mating pairs/generation; average F = 0.39) strains of mice. Although results for individual traits vary considerably, multivariate analysis indicates that V(a) in the bottlenecked populations is greater than expected. Traits with patterns and amounts of epistasis predictive of enhanced V(a) also show the largest deviations from additive expectations. Finally, the correlation structure of weekly weights is not significantly different between control and experimental lines but correlations between necropsy traits do differ, especially those involving the heart, kidney and tail length.

Implantable cardioverter defibrillators in heart failure.

Implantable cardioverter defibrillators provide effective and reliable treatment of spontaneous VT and VF. These devices can be expected to decrease the risk for arrhythmic death in patients with heart failure but do not improve overall survival when death from severe pump dysfunction is imminent. Appropriate patient selection is a major aspect of arrhythmia management. Future devices will incorporate features that have the potential to reduce atrial arrhythmias, improve ventricular function, monitor hemodynamics, and prevent sudden arrhythmic death.

Recalls and safety alerts involving pacemakers and implantable cardioverter-defibrillator generators.

CONTEXT: Unanticipated pacemaker and implantable cardioverter-defibrillator (ICD) generator malfunctions sometimes warrant recall by the US Food and Drug Administration (FDA). Despite increasingly frequent device implantation, pacemaker and ICD recalls and safety alerts (advisories) remain poorly characterized. OBJECTIVES: To determine pacemaker and ICD generator advisory rates in the United States, to identify trends in these rates, and to examine their clinical and financial implications. DESIGN AND SETTING: Analysis of weekly FDA Enforcement Reports issued between January 1990 and December 2000 to identify all advisories involving pacemaker or ICD generators in the United States. Recalls and safety alerts involving lead malfunctions were not included. MAIN OUTCOME MEASURES: Number of pacemakers and ICD generators in the United States subject to FDA recall or safety alert in 1990-2000; annual pacemaker and ICD advisory rates in the United States in 1990-2000; and estimated cost of device advisories. RESULTS: During the study period, 52 advisories (median [25th and 75th percentiles], 4 [4 and 7] per year) involving 408 500 pacemakers and 114 645 ICDs (523 145 total devices) were issued. Hardware malfunctions (35 advisories affecting 280 641 devices) and computer errors (10 advisories affecting 216 533 devices) accounted for 95% of device recalls. Implantable cardioverter-defibrillators were recalled more frequently than pacemakers (mean [SD], 16.4 [1.6] vs 6.7 [0.8] advisories per 100 person-years; P<.001). Between 1995 and 2000, the annual advisory rate increased for both pacemakers (P for trend <.001) and ICDs (P for trend =.02). An estimated 1.3 million device checks and analyses and 36 187 device replacements resulted from the advisories and cost approximately $870 million. CONCLUSIONS: Pacemaker and ICD recalls and safety alerts occur frequently, affect many patients, and appear to be increasing in number and rate. With the growing number of device implants and expanding indications for device therapy, the number of patients affected by device advisories will likely continue to increase.

Provoked and spontaneous high-frequency, low-amplitude, respirophasic noise transients in patients with implantable cardioverter defibrillators.

INTRODUCTION: Ventricular oversensing (OS) of respirophasic noise transients may cause spurious detections and therapies and pacing inhibition among patients with implantable cardioverter defibrillators (ICDs). The incidence of OS and its relationship to clinical variables and ICD system design are unknown. METHODS AND RESULTS: Three hundred twenty-nine patients performed provocative respiratory maneuvers at rest during intrinsic rhythm and continuous ventricular pacing. OS resulting in spurious ventricular detections was provoked in 3 (0.9%) of 329 patients during intrinsic rhythm and 34 (10.3%) of 329 during pacing. Noise transients not recognized and marked as sensed events, but visually evident on the local endocardial ventricular electrogram, were provoked in an additional 23 (7.0%) of 329 patients. Multivariate logistic regression identified history of spontaneous OS (P < 0.0005, odds ratio 9.7, 95% confidence interval [CI] 1.9 to 50.0), automatic gain control device (P < 0.0005, odds ratio 5.3, 95% CI 2.6 to 10.8) or integrated bipolar lead (P = 0.05, odds ratio 2.6, 95% CI 1.0 to 7.25), and male gender (P = 0.008, odds ratio 3.7, 95% CI 1.2 to 11.1) as predictive of provocable OS. Spontaneous OS resulting in spurious ventricular detections and therapies occurred in 12 (3.6%) patients during follow-up. Eleven of 12 spontaneous episodes occurred in male patients during ventricular pacing; 11 of 12 patients had automatic gain control devices and integrated bipolar leads. CONCLUSION: OS is commonly provoked in ICD patients during ventricular pacing and may occur spontaneously, causing spurious tachyarrhythmia therapies and pacing inhibition. Differences in the incidence of spontaneous and provoked OS between ICD systems can be explained on the basis of unique features of automatic sensing systems and sensing lead design.

The N + 1 difference: a new measure for entrainment mapping.

OBJECTIVES: The purpose of this study was to develop and test a new entrainment mapping measurement, the N + 1 difference. BACKGROUND: Entrainment mapping is useful for identifying re-entry circuit sites but is often limited by difficulty in assessing: 1) changes in QRS complexes or P-waves that indicate fusion, and 2) the postpacing interval (PPI) recorded directly from the stimulation site. METHODS: In computer simulations of re-entry circuits, the interval from a stimulus that reset tachycardia to a timing reference during the second beat after the stimulus was compared with the timing of local activation at the site during tachycardia to define an interval designated the N + 1 difference. The N + 1 difference was compared with the PPI-tachycardia cycle length (TCL) difference in simulations and at 65 sites in 10 consecutive patients with ventricular tachycardia (VT) after myocardial infarction and at 45 sites in 10 consecutive patients with atrial flutter. RESULTS: In simulations, the N + 1 difference was equal to the PPI-TCL difference. During mapping of VT and atrial flutter, the N + 1 difference correlated well with the PPI-TCL difference (r > or = 0.91, p < 0.0001), identifying re-entry circuit sites with sensitivity of > or = 86% and specificity of > or = 90%. Accuracy was similar using either the surface electrocardiogram or an intracardiac electrogram (Eg) as the timing reference. CONCLUSIONS: The N + 1 difference allows entrainment mapping to be used to identify re-entry circuit sites when it is difficult to evaluate Egs at the mapping site or fusion in the surface electrocardiogram.

Genetic architecture of adiposity in the cross of LG/J and SM/J inbred mice.

The genetic basis of variation in obesity in human populations is thought to be owing to many genes of relatively small effect and their interactions. The LG/J by SM/J intercross of mouse inbred strains provides an excellent model system in which to investigate multigenic obesity. We previously mapped a large number of quantitative trait loci (QTLs) affecting adult body weight in this cross. We map body composition traits, adiposity, and skeletal size, in a replicate F2 intercross of the same two strains containing 510 individuals. Using interval-mapping methods, we located eight QTLs affecting adiposity (Adip1-8). Two of these adiposity loci also affected tail length (Adip4 and Adip6) along with seven additional tail length QTLs (Skl1-7). A further four QTLs (Wt1-4) affect adult weight but not body composition. These QTLs have relatively small effects, typically about 0.2-0.4 standard deviation units, and account for between 3% and 10% of the variance in individual characters. All QTLs participated in epistatic interactions with other QTLs. Most of these interactions were due to additive-by-additive epistasis, which can nullify the apparent effects of single loci in our population. Adip8 interacts with all the other adiposity QTLs and seems to play a central role in the genetic system affecting obesity in this cross. Only two adiposity QTLs, Adip4 and Adip6, also affect tail length, indicating largely separate genetic control of variation in adiposity and skeletal size. Body size and obesity QTLs in the same locations as those discovered here are commonly found in mapping experiments with other mouse strains.

Processing of alpha-globin and ICP0 mRNA in cells infected with herpes simplex virus type 1 ICP27 mutants.

Herpes simplex virus (HSV) ICP27 is an essential and multifunctional regulator of viral gene expression that modulates RNA splicing, polyadenylation, and nuclear export. We have previously reported that ICP27 causes the cytoplasmic accumulation of unspliced alpha-globin pre-mRNA. Here we examined the effects of a series of ICP27 mutations that alter important functional regions of the protein on the processing and nuclear transport of alpha-globin and HSV ICP0 RNA. The results demonstrate that ICP27 mutants that are impaired for growth in noncomplementing cells, including mutants in the N- and C-terminal regions, are defective in the accumulation of alpha-globin pre-mRNA. Unexpectedly, several mutants that are competent to repress the expression of reporter genes in transient transfection assays failed to accumulate unspliced RNA, implying that different mechanisms are responsible for transrepression and pre-mRNA accumulation. Several mutants caused a marked increase in the length and heterogeneity of the alpha-globin mRNA poly(A) tail, suggesting that ICP27 may directly or indirectly affect the regulation of poly(A) polymerase. ICP27 was also required for the accumulation of multiple ICP0 intron-bearing transcripts, but this effect displayed a mutational sensitivity profile different from that of accumulation of unspliced alpha-globin RNA. Moreover, unlike spliced and unspliced alpha-globin RNAs, which were efficiently exported to the cytoplasm, spliced and intron-containing ICP0 transcripts were predominantly nuclear in localization, and ICP27 was not required for nuclear retention of the spliced message. We propose that these transcript- and ICP27 allele-specific differences may be explained by the presence of a strong cis-acting ICP27 response element in the alpha-globin transcript.

Mapping and radiofrequency catheter ablation of the three types of sustained monomorphic ventricular tachycardia in nonischemic heart disease.

INTRODUCTION: Sustained monomorphic ventricular tachycardia (VT) associated with nonischemic cardiomyopathy (CMP) is uncommon. Optimal approaches to catheter mapping and ablation are not well characterized, but they are likely to depend on the VT mechanism. The purpose of this study was to evaluate the mechanisms of sustained monomorphic VT encountered in nonischemic CMP and to assess the feasibility, safety, and efficacy of catheter radiofrequency ablation for treatment. METHODS AND RESULTS: Twenty-six consecutive patients with nonischemic CMP referred for management of recurrent VT were studied. In 16 (62%) patients, VT was related to a region of abnormal electrograms consistent with scar and the response to pacing suggested a reentrant mechanism. In 5 (19%) patients, VT was due to bundle branch or interfascicular reentry. In 7 (27%) patients, the VT mechanism was focal automaticity, 4 of whom had evidence of tachycardia-induced CMP. After catheter ablation targeting parts of reentrant circuits, VT was not inducible in 8 (53%) of 15 patients with scar-related reentry, was modified in 5 (33%) patients, and still was inducible in 2 (13%) patients. Ablation was successful in 5 of 5 patients with bundle branch reentry and in 6 of 7 patients with a focal automaticity mechanism. Overall, catheter ablation abolished clinical recurrence of VT in 20 (77%) of 26 patients during a follow-up of 15 +/- 12 months. CONCLUSION: Three different mechanisms of VT are encountered in patients with nonischemic CMP. The mapping and ablation approach varies with the type of VT. In this selected population, the overall efficacy was 77%.

Catheter ablation for hemodynamically unstable monomorphic ventricular tachycardia.

INTRODUCTION: Hemodynamic collapse precludes extensive catheter mapping to identify focal target regions in many patients with ventricular tachycardia (VT) associated with heart disease. This study tested the feasibility of catheter ablation of poorly tolerated VTs by targeting a region identified during sinus rhythm. METHODS AND RESULTS: Ablation was attempted in five patients, ages 44 to 59 years, with left ventricular ejection fractions of 0.15 to 0.20 and poorly tolerated VT causing multiple implantable defibrillator therapies (6 to 30 episodes/month). VT was due to prior infarction in three patients and nonischemic cardiomyopathy in two. Target regions were sought that met the following criteria: (1) evidence of slow conduction from fractionated sinus rhythm electrograms and stimulus-QRS delays during pace mapping, and (2) evidence that the region contains the reentrant circuit exit from pace mapping. In 4 of 5 patients, a target region was identified and radiofrequency lesions applied. Ablation abolished all recurrences of VT in 3 of 4 patients during follow-up of 14 to 22 months. There were no complications. CONCLUSION: Ablation of poorly tolerated VT is feasible in some patients by mapping during sinus rhythm and performing ablation over a region of identifiable scar that contains abnormal conduction and a presumptive VT exit.

Herpes simplex virus ICP27 induces cytoplasmic accumulation of unspliced polyadenylated alpha-globin pre-mRNA in infected HeLa cells.

Transcripts of most intron-bearing cellular genes must be processed by the splicing machinery in order to efficiently accumulate and gain access to the cytoplasm. However, we found that herpes simplex virus induces cytoplasmic accumulation of both spliced and unspliced polyadenylated alpha-globin RNAs in infected HeLa cells. Accumulation of the unspliced RNA required the immediate-early protein ICP27, and ICP27 was sufficient (in combination with ICP4) to produce this effect in a transient-transfection assay. However, expression of ICP27 did not markedly alter the levels of fully spliced alpha-globin transcripts in infected cells. These data demonstrate that the previously documented effects of ICP27 on the cellular splicing apparatus do not greatly inhibit splicing of alpha-globin RNA and argue that ICP27 induces a splicing-independent pathway for alpha-globin RNA accumulation and nuclear export.

Mapping quantitative trait loci for murine growth: a closer look at genetic architecture.

Over 20 years ago, D. S. Falconer and others launched an important avenue of research into the quantitative of body size growth in mice. This study continues in that tradition by locating quantitative trait loci (QTLs) responsible for murine growth, such as age-specific weights and growth periods, and examining the genetic architecture for body weight. We identified a large number of potential QTLs in an earlier F2 intercross (Intercross I) of the SM/J and LG/J inbred mouse strains. Many of these QTLs are replicated in a second F2 intercross (Intercross II) between the same two strains. These replicated regions provide candidate regions for future fine-mapping studies. We also examined body size and growth QTLs using the combined data set from these two intercrosses, resulting in 96 microsatellite markers being scored for 1045 individuals. An examination of the genetic architecture for age-specific weight and growth periods resulted in locating 20 separate QTLs, which were mainly additive in nature, although dominance was found to affect early growth and body size. QTLs affecting early and late growth were generally distinct, mapping to separate chromosome locations. This QTL pattern indicates largely separate genetic and physiological systems for early and later murine growth, as Falconer suggested. We also found sex-specific QTLs for body size with implications for the evolution of sexual dimorphism.

Entrainment mapping and radiofrequency catheter ablation of ventricular tachycardia in right ventricular dysplasia.

OBJECTIVES: The purpose of this study was to determine if entrainment mapping techniques and predictors of successful ablation sites previously tested in coronary artery disease can be applied to ventricular tachycardia (VT) in arrhythmogenic right ventricular dysplasia (ARVD). BACKGROUND: VT in ARVD has not been well characterized. Reentry circuits in areas of abnormal myocardium are the likely cause, but these circuits have not been well defined. METHODS: Mapping of 19 VTs in 5 patients with ARVD was performed. At 58 sites pacing entrained VT and radiofrequency current (RF) was applied to assess acute termination of VT. RESULTS: Sites classified as exits, central/proximal, inner loop, outer loop, remote bystander and adjacent bystander were identified by entrainment criteria. The reentrant circuit sites were clustered predominantly around the tricuspid annulus and in the right ventricular outflow tract (RVOT). RF ablation acutely terminated VT at 13 sites or 22% of the applications. Of the 19 VTs, eight were rendered noninducible and three were modified to a longer cycle length. In 2 patients ablation at a single site abolished two VTs. CONCLUSION: VT in ARVD shows many of the characteristics of VT due to myocardial infarction. Entrainment mapping techniques can be used to characterize reentry circuits in ARVD. The use of entrainment mapping to guide ablation is feasible.

Identification and ablation of macroreentrant ventricular tachycardia with the CARTO electroanatomical mapping system.

Monomorphic ventricular tachycardias associated with regions of scar are most commonly due to reentry. Catheter based techniques have recently been described for mapping of reentry circuits. Fluoroscopic methods have obvious limitations when attempting to map large ventricular reentry circuit and localize target-sites of radiofrequency ablation. Three-dimensional right ventricular endocardial mapping was performed in a 38-year-old patient with ventricular tachycardia 28 years after surgical correction of tetralogy of Fallot by using the CARTO electroanatomical system. The map of electrogram voltage showed low amplitude electrograms over the anterior wall of the right ventricle extending into the right ventricular outflow tract, consistent with the location of the ventriculotomy scar. Recording of local activation time was combined with entrainment mapping to define the macroreentrant circuit during ventricular tachycardia. Since the activation propagated through a broad path around the right ventriculotomy scar, ablation was performed by creating a line of block, which was facilitated by tagging of the lesion sites on the endocardial activation map. Large ventricular reentry circuits can be identified and interrupted by creation of a line of block to interrupt a broad path. A practical approach to mapping combining analysis of electrogram voltage, activation sequence, and entrainment is presented.

Ablation of ventricular tachycardia due to a postinfarct ventricular septal defect: identification and transection of a broad reentry loop.

INTRODUCTION: Ventricular tachycardia (VT) after postinfarct ventricular septal defect (VSD) repair has not been well characterized. METHODS AND RESULTS: A 55-year-old man developed refractory VT after inferior wall infarction and VSD repair. Entrainment demonstrated a broad reentry circuit path (outer loop) between the tricuspid annulus and VSD patch. A series of radiofrequency (RF) lesions transected this path, abolishing VT and producing conduction block between the inferior and superior aspects of the basal right ventricular septum. CONCLUSION: Some VTs have broad reentry loops requiring ablation by a series of RF lesions across the path to create a line of block. This approach is analogous to that for atrial flutter.

Ablation therapy for cardiac arrhythmias.

Ablation has become an important and, in some cases, the first-line therapy for a number of tachyarrhythmias. The feasibility of treating arrhythmias with ablation was initially demonstrated with surgical ablation techniques. Recently, catheter ablation techniques have replaced the surgical approach in nearly all cases. Catheter ablation is highly effective for the Wolff-Parkinson-White syndrome, atrioventricular nodal reentry, and atrial ectopic tachycardia. It is effective for atrial flutter, although approximately one quarter of patients treated with catheter ablation continue to require therapy for concomitant atrial fibrillation. The surgical maze procedure has proved to be feasible for preventing atrial fibrillation. The risks and long-term efficacy of catheter ablation maze procedures for atrial fibrillation need to be defined. The efficacy of ablation for ventricular tachycardia varies with the type of tachycardia. Catheter ablation is very effective for the rare idiopathic ventricular tachycardias that occur in structurally normal hearts and for bundle-branch reentry ventricular tachycardia, which occurs most frequently in patients with dilated cardiomyopathy. When performed at an experienced center, surgical ablation is an excellent option for selected patients with ventricular tachycardia due to prior myocardial infarction who have a discrete aneurysm but otherwise well-preserved ventricular function. Catheter ablation shows promise for this arrhythmia, but it can be offered only to those patients who have relatively slow tachycardias that allow catheter mapping. Substantial advances in mapping and ablation technology will continue to occur, allowing nonpharmacologic control of cardiac arrhythmias to be achieved in an ever greater number of patients.

Mutations in active-site residues of the uracil-DNA glycosylase encoded by vaccinia virus are incompatible with virus viability.

The D4R gene of vaccinia virus encodes a functional uracil-DNA glycosylase that is essential for viral viability (D. T. Stuart, C. Upton, M. A. Higman, E. G. Niles, and G. McFadden, J. Virol. 67:2503-2513, 1993), and a D4R mutant, ts4149, confers a conditional lethal defect in viral DNA replication (A. K. Millns, M. S. Carpenter, and A. M. DeLange, Virology 198:504-513, 1994). The mutant ts4149 protein was expressed in vitro and assayed for uracil-DNA glycosylase activity. Less than 6% of wild-type activity was observed at permissive temperatures, but the ts4149 protein was completely inactive at the nonpermissive temperature. Mutagenesis of the ts4149 gene back to wild type (Arg-179-->Gly) restored full activity. The ts4149 protein was considerably reduced in lysates of cells infected at the permissive temperature, and its activity was undetectable, even in the presence of the uracil glycosylase inhibitor protein, which inhibits the host uracil-DNA glycosylases but not that of vaccinia virus. Thus the ts4149 protein is thermolabile, correlating uracil removal with vaccinia virus DNA replication. Three active-site amino acids of the vaccinia virus uracil-DNA glycosylase were mutated (Asp-68-->Asn, Asn-120-->Val, and His-181-->Leu), producing proteins that were completely defective in uracil excision but still retained the ability to bind DNA. Each mutated D4R gene was transfected into vaccinia virus ts4149-infected cells in order to assess the recombination events that allowed virus survival at 40 degrees C. Genetic analysis and sequencing studies revealed that the only viruses to survive were those in which recombination eliminated the mutant locus. We conclude that the uracil cleavage activity of the D4R protein is essential for its function in vaccinia virus DNA replication, suggesting that the removal of uracil residues plays an obligatory role.

Fusigenic liposome-mediated DNA transfer into cardiac myocytes.

Current methods of gene transfer into cultured cardiac myocytes have serious limitations, including low efficiency, toxicity or constraints on DNA insert size. The present study examined the effectiveness of hemagglutinating virus of Japan (HVJ) in promoting liposome-mediated DNA transfer into cultured neonatal rat cardiac myocytes. Fluorescein isothiocyanate-labeled oligonucleotides (F-ODN) or plasmid expression vectors encoding SV40 large T antigen (pActSVT) and beta-galactosidase (pAct beta-gal) were complexed with liposomes and the viral protein coat of HVJ. Plasmid vectors were complexed with the nuclear localizing protein HMG-1 prior to HVJ-liposome encapsulation. Neonatal myocytes were transfected by incubation with HVJ-liposome/DNA complexes on culture day 3 or 7. Using F-ODN, we were able to demonstrate significant uptake of DNA (transfection efficiencies of 80-90%) 1 h after transfection that persisted for 1 week in culture. Interestingly, F-ODN were concentrated in the myocyte nuclei for the first 4 days after transfection. Immunohistochemistry showed that 25-30% of myocytes transfected with either pActSVT or pAct beta-Gal expressed plasmid-encoded protein at 72 h whether they were transfected at day 3 or day 7 of culture, while cells transfected with blank vectors did not. Quantitative beta-galactosidase assays confirmed that the use of HVJ significantly enhanced liposome-mediated transfection. Cell toxicity was not apparent. Gene transfer via intracoronary injection also demonstrated the capacity of HVJ to mediate transfection of rabbit cardiac myocytes in vivo, with F-ODN-dependent fluorescence persisting for up to 1 week. We conclude that HVJ/liposome-mediated transfer is efficient for the transfection of both oligonucleotides and plasmids into cardiac myocytes both in vitro and in vivo, and may provide a new tool for the investigation of cardiac myocyte biology and disease.