Identification and characterization of two novel KCNH2 mutations contributing to long QT syndrome.

We identified two different inherited mutations in KCNH2 gene, or human ether-a-go-go related gene (hERG), which are linked to Long QT Syndrome. The first mutation was in a 1-day-old infant, whereas the second was in a 14-year-old girl. The two KCNH2 mutations were transiently transfected into either human embryonic kidney (HEK) cells or human induced pluripotent stem-cell derived cardiomyocytes. We performed associated multiscale computer simulations to elucidate the arrhythmogenic potentials of the KCNH2 mutations. Genetic screening of the first and second index patients revealed a heterozygous missense mutation in KCNH2, resulting in an amino acid change (P632L) in the outer loop of the channel and substitution at position 428 from serine to proline (S428P), respectively. Heterologous expression of P632L and S428P into HEK cells produced no hERG current compared to the wild type (WT). Moreover, the co-transfection of WT and P632L yielded no hERG current; however, the co-transfection of WT and S428P yielded partial hERG current. Action potentials were prolonged in a complete or partial blockade of hERG current from computer simulations which was more severe in Purkinje than ventricular myocytes. Three dimensional simulations revealed a higher susceptibility to reentry in the presence of hERG current blockade. Our experimental findings suggest that both P632L and S428P mutations may impair the KCNH2 gene. The Purkinje cells exhibit a more severe phenotype than ventricular myocytes, and the hERG current blockade renders the ventricles an arrhythmogenic substrate from computer modeling.

Improving efficiency and reducing costs in robotic surgery: a Lean Six Sigma approach to optimize turnover time.

Operating room (OR) turnover time (TOT) is the time it takes to prepare an OR for the next surgery after the previous one has been completed. Reducing OR TOT can improve the efficiency of the OR, reduce costs, and improve surgeons' and patients' satisfaction. The objective of this study is to evaluate the effectiveness of an operating room (OR) turnover time (TOT) reduction initiative using the Lean Six Sigma methodology (DMAIC) in the bariatric and thoracic service lines. Performance improvement strategies consist of simplifying steps (surgical tray optimization) and concurrent steps (parallel task execution). We compared 2-month pre-implementation vs. post-implementation. A paired t-test was used to assess whether the difference in the measurements was statistically significant. The study found that TOT was reduced by 15.6% from an average of 35.6 ± 8.1 to minutes 30.09 ± 9.7 min (p < 0.05). Specifically, in the bariatric service line, TOT was reduced by 17.15% and in the thoracic service line, TOT was reduced by 9.6%. No adverse events related to the initiative were reported. The results of this study indicate that the TOT reduction initiative was effective in reducing TOT. The efficient use of operating rooms is crucial in hospital management, as it not only impacts finances but also affects the satisfaction of surgical teams and patients. This study shows the effectiveness of Lean Six Sigma methodology in reducing TOT and improving the efficiency in the OR.

Introducing the E100: will the improvement make a significant difference? Financial analysis based on sleeve gastrectomy.

Robotic-assisted surgery has been shown to offer significant benefits for both patients and surgeons. However, the high cost of the equipment remains a major barrier to its widespread adoption in the medical field. To ensure cost-effectiveness of these procedures, it is important to implement strategies that can help reduce the costs involved. One potential strategy to reduce costs is to compare the performance of different generators used in these procedures. In this study, we compared the performance of two generators: the E100 (Intuitive Surgical, Inc.) and the ERBE VIO dV 2.0 (Elektromedizin GmbH). The analysis focused on several key metrics, including the number of times the generators were activated, the average time per seal, total sealing time, and console time. The financial impact of transitioning to the E100 was also evaluated based on annual volume. We analyzed a total of 1457 sleeve gastrectomies, with 746 performed using the ERBE generator and 711 performed using the E100. There were no significant differences in preoperative BMI or bleeding complication rates between the two groups. The average activation of the generator per case was similar in both groups. However, the sealing time was 42.3% lower, and the average console time was 8 min shorter in cases where the E100 was used. Based on our financial analysis, transitioning to the E100 generator would result in cost savings of approximately $33-$34 K per year. This suggests that introducing the new generator is a successful strategy for reducing costs associated with robotic-assisted procedures.

Utilizing Probiotics for the Prevention and Treatment of Gastrointestinal Diseases.

Probiotics are heavily advertised to promote a healthy gastrointestinal tract and boost the immune system. This review article summarizes the history and diversity of probiotics, outlines conventional in vitro assays and in vivo models, assesses the pharmacologic effects of probiotic and pharmaceutical co-administration, and the broad impact of clinical probiotic utilization for gastrointestinal disease indications.

Acetylation contributes to hypertrophy-caused maturational delay of cardiac energy metabolism.

A dramatic increase in cardiac fatty acid oxidation occurs following birth. However, cardiac hypertrophy secondary to congenital heart diseases (CHDs) delays this process, thereby decreasing cardiac energetic capacity and function. Cardiac lysine acetylation is involved in modulating fatty acid oxidation. We thus investigated what effect cardiac hypertrophy has on protein acetylation during maturation. Eighty-four right ventricular biopsies were collected from CHD patients and stratified according to age and the absence (n = 44) or presence of hypertrophy (n = 40). A maturational increase in protein acetylation was evident in nonhypertrophied hearts but not in hypertrophied hearts. The fatty acid ß-oxidation enzymes, long-chain acyl CoA dehydrogenase (LCAD) and ß-hydroxyacyl CoA dehydrogenase (ßHAD), were hyperacetylated and their activities positively correlated with their acetylation after birth in nonhypertrophied hearts but not hypertrophied hearts. In line with this, decreased cardiac fatty acid oxidation and reduced acetylation of LCAD and ßHAD occurred in newborn rabbits subjected to cardiac hypertrophy due to an aortocaval shunt. Silencing the mRNA of general control of amino acid synthesis 5-like protein 1 reduced acetylation of LCAD and ßHAD as well as fatty acid oxidation rates in cardiomyocytes. Thus, hypertrophy in CHDs prevents the postnatal increase in myocardial acetylation, resulting in a delayed maturation of cardiac fatty acid oxidation.

Screening and Analysis of Janelia FlyLight Project Enhancer-Gal4 Strains Identifies Multiple Gene Enhancers Active During Hematopoiesis in Normal and Wasp-Challenged Drosophila Larvae.

A GFP expression screen has been conducted on >1000 Janelia FlyLight Project enhancer-Gal4 lines to identify transcriptional enhancers active in the larval hematopoietic system. A total of 190 enhancers associated with 87 distinct genes showed activity in cells of the third instar larval lymph gland and hemolymph. That is, gene enhancers were active in cells of the lymph gland posterior signaling center (PSC), medullary zone (MZ), and/or cortical zone (CZ), while certain of the transcriptional control regions were active in circulating hemocytes. Phenotypic analyses were undertaken on 81 of these hematopoietic-expressed genes, with nine genes characterized in detail as to gain- and loss-of-function phenotypes in larval hematopoietic tissues and blood cells. These studies demonstrated the functional requirement of the cut gene for proper PSC niche formation, the hairy, Btk29A, and E2F1 genes for blood cell progenitor production in the MZ domain, and the longitudinals lacking, dFOXO, kayak, cap-n-collar, and delilah genes for lamellocyte induction and/or differentiation in response to parasitic wasp challenge and infestation of larvae. Together, these findings contribute substantial information to our knowledge of genes expressed during the larval stage of Drosophila hematopoiesis and newly identify multiple genes required for this developmental process.

Activating PPARα prevents post-ischemic contractile dysfunction in hypertrophied neonatal hearts.

RATIONALE: Post-ischemic contractile dysfunction is a contributor to morbidity and mortality after the surgical correction of congenital heart defects in neonatal patients. Pre-existing hypertrophy in the newborn heart can exacerbate these ischemic injuries, which may partly be due to a decreased energy supply to the heart resulting from low fatty acid ß-oxidation rates. OBJECTIVE: We determined whether stimulating fatty acid ß-oxidation with GW7647, a peroxisome proliferator-activated receptor-α (PPARα) activator, would improve cardiac energy production and post-ischemic functional recovery in neonatal rabbit hearts subjected to volume overload-induced cardiac hypertrophy. METHODS AND RESULTS: Volume-overload cardiac hypertrophy was produced in 7-day-old rabbits via an aorto-caval shunt, after which, the rabbits were treated with or without GW7647 (3 mg/kg per day) for 14 days. Biventricular working hearts were subjected to 35 minutes of aerobic perfusion, 25 minutes of global no-flow ischemia, and 30 minutes of aerobic reperfusion. GW7647 treatment did not prevent the development of cardiac hypertrophy, but did prevent the decline in left ventricular ejection fraction in vivo. GW7647 treatment increased cardiac fatty acid ß-oxidation rates before and after ischemia, which resulted in a significant increase in overall ATP production and an improved in vitro post-ischemic functional recovery. A decrease in post-ischemic proton production and endoplasmic reticulum stress, as well as an activation of sarcoplasmic reticulum calcium ATPase isoform 2 and citrate synthase, was evident in GW7647-treated hearts. CONCLUSIONS: Stimulating fatty acid ß-oxidation in neonatal hearts may present a novel cardioprotective intervention to limit post-ischemic contractile dysfunction.

bantam miRNA is important for Drosophila blood cell homeostasis and a regulator of proliferation in the hematopoietic progenitor niche.

The Drosophila hematopoietic system is utilized in this study to gain novel insights into the process of growth control of the hematopoietic progenitor niche in blood development. The niche microenvironment is an essential component controlling the balance between progenitor populations and differentiated, mature blood cells and has been shown to lead to hematopoietic malignancies in humans when misregulated. MicroRNAs are one class of regulators associated with blood malignancies; however, there remains a relative paucity of information about the role of miRNAs in the niche. Here we demonstrate that bantam miRNA is endogenously active in the Drosophila hematopoietic progenitor niche, the posterior signaling center (PSC), and functions in the primary hematopoietic organ, the lymph gland, as a positive regulator of growth. Loss of bantam leads to a significant reduction in the PSC and overall lymph gland size, as well as a loss of the progenitor population and correlative premature differentiation of mature hemocytes. Interestingly, in addition to being essential for proper lymph gland development, we have determined bantam to be a novel upstream component of the insulin signaling cascade in the PSC and have unveiled dMyc as one factor central to bantam activity. These important findings identify bantam as a new hematopoietic regulator, place it in an evolutionarily conserved signaling pathway, present one way in which it is regulated, and provide a mechanism through which it facilitates cellular proliferation in the hematopoietic niche.

Inhibition of carnitine palmitoyltransferase-1 activity alleviates insulin resistance in diet-induced obese mice.

Impaired skeletal muscle fatty acid oxidation has been suggested to contribute to insulin resistance and glucose intolerance. However, increasing muscle fatty acid oxidation may cause a reciprocal decrease in glucose oxidation, which might impair insulin sensitivity and glucose tolerance. We therefore investigated what effect inhibition of mitochondrial fatty acid uptake has on whole-body glucose tolerance and insulin sensitivity in obese insulin-resistant mice. C57BL/6 mice were fed a high-fat diet (60% calories from fat) for 12 weeks to develop insulin resistance. Subsequent treatment of mice for 4 weeks with the carnitine palmitoyltransferase-1 inhibitor, oxfenicine (150 mg/kg i.p. daily), resulted in improved whole-body glucose tolerance and insulin sensitivity. Exercise capacity was increased in oxfenicine-treated mice, which was accompanied by an increased respiratory exchange ratio. In the gastrocnemius muscle, oxfenicine increased pyruvate dehydrogenase activity, membrane GLUT4 content, and insulin-stimulated Akt phosphorylation. Intramyocellular levels of lipid intermediates, including ceramide, long-chain acyl CoA, and diacylglycerol, were also decreased. Our results demonstrate that inhibition of mitochondrial fatty acid uptake improves insulin sensitivity in diet-induced obese mice. This is associated with increased carbohydrate utilization and improved insulin signaling in the skeletal muscle, suggestive of an operating Randle Cycle in muscle.

Inhibition of serine palmitoyl transferase I reduces cardiac ceramide levels and increases glycolysis rates following diet-induced insulin resistance.

OBJECTIVE: Diet-induced obesity (DIO) leads to an accumulation of intra-myocardial lipid metabolites implicated in causing cardiac insulin resistance and contractile dysfunction. One such metabolite is ceramide, and our aim was to determine the effects of inhibiting de novo ceramide synthesis on cardiac function and insulin stimulated glucose utilization in mice subjected to DIO. MATERIALS AND METHODS: C57BL/6 mice were fed a low fat diet or subjected to DIO for 12 weeks, and then treated for 4 weeks with either vehicle control or the serine palmitoyl transferase I (SPT I) inhibitor, myriocin. In vivo cardiac function was assessed via ultrasound echocardiography, while glucose metabolism was assessed in isolated working hearts. RESULTS: DIO was not associated with an accumulation of intra-myocardial ceramide, but rather, an accumulation of intra-myocardial DAG (2.63±0.41 vs. 4.80±0.97 nmol/g dry weight). Nonetheless, treatment of DIO mice with myriocin decreased intra-myocardial ceramide levels (50.3±7.7 vs. 26.9±2.7 nmol/g dry weight) and prevented the DIO-associated increase in intra-myocardial DAG levels. Interestingly, although DIO impaired myocardial glycolysis rates (7789±1267 vs. 2671±326 nmol/min/g dry weight), hearts from myriocin treated DIO mice exhibited an increase in glycolysis rates. CONCLUSIONS: Our data reveal that although intra-myocardial ceramide does not accumulate following DIO, inhibition of de novo ceramide synthesis nonetheless reduces intra-myocardial ceramide levels and prevents the accumulation of intra-myocardial DAG. These effects improved the DIO-associated impairment of cardiac glycolysis rates, suggesting that SPT I inhibition increases cardiac glucose utilization.

Cardiac hypertrophy in the newborn delays the maturation of fatty acid ß-oxidation and compromises postischemic functional recovery.

During the neonatal period, cardiac energy metabolism progresses from a fetal glycolytic profile towards one more dependent on mitochondrial oxidative metabolism. In this study, we identified the effects of cardiac hypertrophy on neonatal cardiac metabolic maturation and its impact on neonatal postischemic functional recovery. Seven-day-old rabbits were subjected to either a sham or a surgical procedure to induce a left-to-right shunt via an aortocaval fistula to cause RV volume-overload. At 3 wk of age, hearts were isolated from both groups and perfused as isolated, biventricular preparations to assess cardiac energy metabolism. Volume-overload resulted in cardiac hypertrophy (16% increase in cardiac mass, P < 0.05) without evidence of cardiac dysfunction in vivo or in vitro. Fatty acid oxidation rates were 60% lower (P < 0.05) in hypertrophied hearts than controls, whereas glycolysis increased 246% (P < 0.05). In contrast, glucose and lactate oxidation rates were unchanged. Overall ATP production rates were significantly lower in hypertrophied hearts, resulting in increased AMP-to-ATP ratios in both aerobic hearts and ischemia-reperfused hearts. The lowered energy generation of hypertrophied hearts depressed functional recovery from ischemia. Decreased fatty acid oxidation rates were accompanied by increased malonyl-CoA levels due to decreased malonyl-CoA decarboxylase activity/expression. Increased glycolysis in hypertrophied hearts was accompanied by a significant increase in hypoxia-inducible factor-1α expression, a key transcriptional regulator of glycolysis. Cardiac hypertrophy in the neonatal heart results in a reemergence of the fetal metabolic profile, which compromises ATP production in the rapidly maturing heart and impairs recovery of function following ischemia.

Specific cell ablation in Drosophila using the toxic viral protein M2(H37A).

The expression of toxic viral proteins for the purpose of eliminating distinct populations of cells, while leaving the rest of an organism unaffected, is a valuable method for analyzing development. Using the Gal4-UAS system, we employed the M2(H37A) toxic ion channel of the influenza-A virus to selectively ablate the Drosophila eye-antennal imaginal discs, hemocytes, dorsal vessel and nervous tissue, and comparatively monitored the effects of expressing the apoptosis-promoting protein Reaper in identical cell populations. In this report, we demonstrate the effectiveness of M2(H37A)-mediated ablation as a new means to selectively eliminate cells of interest during Drosophila development.

High levels of fatty acids increase contractile function of neonatal rabbit hearts during reperfusion following ischemia.

In the neonatal heart the transition from using carbohydrates to using fatty acids has not fully matured and oxidative metabolism/ATP generation may be limiting contractile function after ischemia. This study tested the hypothesis that increasing fatty acid availability increases recovery of left ventricular (LV) work by increasing palmitate oxidation, tricarboxylic acid (TCA) cycle activity, and ATP generation. Isolated working hearts from 7-day-old rabbits were perfused with Krebs solution containing low (0.4 mM) or high (2.4 mM) palmitate and 5.5 mM glucose. Hearts were subjected to 35-min global ischemia before 40-min reperfusion, and rates of glycolysis, glucose oxidation, and palmitate oxidation were assessed. LV work was similar before ischemia but was greater during reperfusion in hearts perfused with 2.4 mM palmitate compared with hearts perfused with 0.4 mM palmitate [6.98 +/- 0.14 (n = 15) vs. 3.01 +/- 0.23 (n = 16) mJ.beat(-1).g dry wt(-1); P < 0.05]. This was accompanied by increased LV energy expenditure during reperfusion [35.98 +/- 0.16 (n = 8) vs. 19.92 +/- 0.18 (n = 6) mJ.beat(-1).g dry wt(-1); P < 0.05]. During reperfusion the rates of palmitate oxidation [237.5 +/- 28.10 (n = 7) vs. 86.0 +/- 9.7 (n = 6) nmol.g dry wt(-1).min(-1); P < 0.05], total TCA cycle activity [2.65 +/- 0.39 (n = 7) vs. 1.36 +/- 0.14 (n = 6) micromol acetyl-CoA.g dry wt(-1).min(-1); P < 0.05], and ATP generation attributable to palmitate oxidation [26.6 +/- 3.1 (n = 7) vs. 12.6 +/- 1.7 (n = 6) micromol.g dry wt(-1).min(-1); P < 0.05] were greater in hearts perfused with 2.4 mM palmitate. These data indicate that the neonatal heart has decreased energy reserve, and, in contrast to the mature heart, increasing availability of fatty acid substrate increases energy production and improves recovery of function after ischemia.