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The Journey of Hope (JoH) is an evidence-based behavioral health prevention intervention model designed for disaster-affected children and youth. While the in-person JoH model has been extensively delivered and studied, the virtual implementation of this intervention, specifically tailored for the unique needs of children and youth during the COVID-19 pandemic, has not been previously explored. In this context, authors of this article adapted the JoH to a virtual delivery model, aiming to provide access to behavioral health preventive services for children and youth during the COVID-19 pandemic when in-person services were not feasible. This adaptation, utilizing the dynamic adaptation process (DAP) and community-based participatory research (CBPR) principles, was carefully designed to meet pandemic-related challenges while maintaining the core components of the original intervention. The primary objectives of this article are twofold: (1) to provide an overview of the theoretical and conceptual underpinnings of the JoH-C19, including the framework that guided the adaptation, and (2) to describe the virtually adapted curriculum and initial pilot of the JoH-C19. This adaptation represents a crucial step in ensuring the accessibility of virtual behavioral health interventions for young populations facing various collective traumas and challenges in a rapidly evolving world., (© 2024 National Association of Social Workers.)

The "orgasm gap" for women who have sex with men remains a pressing gender equity issue. Recent research found that women who pursued orgasm as a goal were more likely to have one. The current research replicated this relationship between orgasm goal pursuit and orgasm frequency for heterosexual women, and found that this relationship did not exist for heterosexual men (Study 1). Then, across two experimental studies, we examined how heterosexual women vary their orgasm goal pursuit across sexual encounters. In Study 2, women who read that a hypothetical sexual encounter would be "quick" reported less intent to pursue orgasm than women who were told they could "take their time" or received no time information. In Study 3, women who read that their hypothetical sexual partner seemed selfish reported less intent to pursue orgasm than women who were given a non-selfish partner or no partner information. Importantly, these effects were mediated by women's perceived orgasm likelihood in the scenario. These results suggest that women shift their pursuit of orgasm depending on cues which signal whether orgasm will be feasible. This research used self-regulation theory to understand women's motivations for pursuing orgasm during sexual encounters with men, with implications for reducing the orgasm gap.

Newly synthesised glycoproteins enter the rough endoplasmic reticulum through a translocation pore. The translocon associated protein (TRAP) complex is located close to the pore. In a patient with a homozygous start codon variant in TRAPγ (SSR3), absence of TRAPγ causes disruption of the TRAP complex, impairs protein translocation into the endoplasmic reticulum and affects transport, for example, into the brush-border membrane. Furthermore, we observed an unbalanced non-occupancy of N-glycosylation sites. The major clinical features are intrauterine growth retardation, facial dysmorphism, congenital diarrhoea, failure to thrive, pulmonary disease and severe psychomotor disability., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Kangaroo mother care (KMC) has been proved to be a safe and cost-effective standard of care for preterm babies. China hasn't adopted the KMC practice widely until recently. We aim to assess barriers and facilitators of KMC adoption in neonatal intensive care units (NICUs) and postnatal wards in China., Methods: We conducted clinical observations and semi-structured interviews with nurses, physicians, and parents who performed KMC in seven NICUs and postnatal wards housed in five hospitals in different provinces of China between August and September 2018. The interviews provided first-hand stakeholder perspectives on barriers and facilitators of KMC implementation and sustainability. We further explored health system's readiness and families' willingness to sustain KMC practice following its pilot introduction. We coded data for emerging themes related to financial barriers, parent- and hospital-level perceived barriers, and facilitators of KMC adoption, specifically those unique in the Chinese context., Results: Five hospitals with KMC pilot programs were selected for clinical observations and 38 semi-structured interviews were conducted. Common cultural barriers included concerns with the conflict with traditional postpartum confinement (Zuo-yue-zi) practice and grandparents' resistance, while a strong family support is a facilitator for KMC adoption. Some parents reported anxiety and guilt associated with having a preterm baby, which can be a parental-level barrier to KMC. Hospital-level factors such as fear of nosocomial infection and shortage of staff and spaces impeded the KMC implementation, and supportive community and peer group organized by the hospital contributed to KMC uptake. Financial barriers included lodging costs for caregivers and supply costs for hospitals., Conclusions: We provided a comprehensive in-depth report on the multi-level KMC barriers and facilitators in China. We recommend policy interventions specifically addressing these barriers and facilitators and increase family and peer support to improve KMC adoption in China. We also recommend that well-designed local cultural and economic feasibility and acceptability studies should be conducted before the KMC uptake.

The role of aquatic resources in ancient economies and paleodiet is important for understanding the evolution of prehistorical societies. Charred food remains from ancient pottery are valuable molecular evidence of dietary habits in antiquity. However, conventional archaeometric approaches applied in their analysis lack organismal specificity, are affected by abundant environmental contaminants, do not elucidate food processing recipes and are limited in the inland regions where diverse dietary resources are available. We performed proteomics analysis of charred organic deposits adhered on early ceramics from Mesolithic-Neolithic inland site Friesack 4 (Brandenburg, Germany). One of pots-a small coarse bowl radiocarbon dated to the end of the 5th millennium BC-was attributed to Endmesolithic pottery. Proteomics of foodcrust from this vessel identified fine carp roe meal and revealed details of a prehistorical culinary recipe. Ancient proteins were unequivocally distinguished from contemporary contaminants by computing deamidation ratios of glutamine residues. These data paint a broader picture of the site-specific exploitation of aquatic resources and contribute to better understanding of the dietary context of Neolithic transition in European inland., Competing Interests: The authors have declared that no competing interests exist.

It is still not entirely clear how α-galactosidase A (GAA) deficiency translates into clinical symptoms of Fabry disease (FD). The present communication investigates the effects of the mutation N215S in FD on the trafficking and processing of lysosomal GAA and their potential association with alterations in the membrane lipid composition. Abnormalities in lipid rafts (LRs) were observed in fibroblasts isolated from a male patient with FD bearing the mutation N215S. Interestingly, LR analysis revealed that the distribution of cholesterol and flotillin-2 are distinctly altered in the Fabry fibroblasts when compared with that of the wild-type cells. Furthermore, increased levels of glycolipid globotriaosylceramide 3 (Gb3) and sphingomyelin (SM) were observed in non-raft membrane fractions of Fabry cells. Substrate reduction with N -butyldeoxynojirimycin (NB-DNJ) in vitro was capable of reversing these abnormalities in this patient. These data led to the hypothesis that alterations of LRs may contribute to the pathophysiology of Morbus Fabry. Furthermore, it may be suggested that substrate reduction therapy with NB-DNJ might be a promising approach for the treatment of GAA deficiency at least for the selected patients., (© 2017 The Author(s).)

Characterizing the first event of biological production of calcium carbonate requires a combination of microscopy approaches. First, intracellular pH distribution and calcium ions can be observed using live microscopy over time. This allows identification of the life stage and the tissue with the feature of interest for further electron microscopy studies. Life stage and tissues of interest are typically higher in pH and Ca signals. Here, using H. elegans, we present a protocol to characterize the presence of calcium carbonate structures in a biological specimen on the scanning electron microscope (SEM), using energy-dispersive X-ray spectroscopy (EDS) to visualize elemental composition, using electron backscatter diffraction (EBSD) to determine the presence of crystalline structures, and using transmission electron microscopy (TEM) to analyze the composition and structure of the material. In this protocol, a focused ion beam (FIB) is used to isolate samples with dimension suitable for TEM analysis. As FIB is a site specific technique, we demonstrate how information from the previous techniques can be used to identify the region of interest, where Ca signals are highest.

Tunneling nanotubes (TNTs) are small membranous tubes of 50-1000 nm diameter observed to connect cells in culture. Transfer of subcellular organelles through TNTs was observed in vitro and in vivo, but the formation and significance of these structures is not well understood. A polydimethylsiloxane biochip-based coculture model was devised to constrain TNT orientation and explore both TNT-formation and TNT-mediated mitochondrial transfer. Two parallel microfluidic channels connected by an array of smaller microchannels enabled localization of stem cell and cardiomyocyte populations while allowing connections to form between them. Stem cells and cardiomyocytes were deposited in their respective microfluidic channels, and stem cell-cardiomyocyte pairs were formed via the microchannels. Formation of TNTs and transfer of stained mitochondria through TNTs was observed by 24 h real-time video recording. The data show that stem cells are 7.7 times more likely to initiate contact by initial extension of filopodia. By 24 h, 67% of nanotube connections through the microchannels are composed of cardiomyocyte membrane. Filopodial extension and retraction by stem cells draws an extension of TNTs from cardiomyocytes. MitoTracker staining shows that unidirectional transfer of mitochondria between stem cell-cardiomyocyte pairs invariably originates from stem cells. Control experiments with cardiac fibroblasts and cardiomyocytes show little nanotube formation between homotypic or mixed cell pairs and no mitochondrial transfer. These data identify a novel biological process, unidirectional mitochondrial transfer, mediated by heterotypic TNT connections. This suggests that the enhancement of cardiomyocyte function seen after stem-cell injection may be due to a bioenergetic stimulus provided by mitochondrial transfer.

The molecular basis of gastrointestinal intolerances in a severe case of Niemann-Pick type C disease was analyzed in an intestinal biopsy specimen. The enzyme activities of intestinal sucrase-isomaltase and maltase-glucoamylase are reduced in the patient, while that of lactase is comparable to the control. The association of SI with lipid rafts is reduced in the patient's biopsy as a consequence of altered composition of membrane microdomains. As association with lipid rafts influences the intracellular transport and the enzyme activities of sucrase-isomaltase and maltase-glucoamylase, these data explain reduced carbohydrate digestion in the intestinal lumen and delineate the effect of deficient cholesterol and sphingolipid homeostasis in development of gastrointestinal symptoms in NPC patients.

Background: Hypoxia-inducible factor-1 α (HIF-1 α ) and NF- κ B play important roles in the inflammatory response after hemorrhagic shock and resuscitation (H/R). Here, the role of myeloid HIF-1 α in liver hypoxia, injury, and inflammation after H/R with special regard to NF- κ B activation was studied., Methods: Mice with a conditional HIF-1 α knockout (KO) in myeloid cell-line and wild-type (WT) controls were hemorrhaged for 90 min (30 ± 2 mm Hg) and resuscitated. Controls underwent only surgical procedures., Results: After six hours, H/R enhanced the expression of HIF-1 α -induced genes vascular endothelial growth factor (VEGF) and adrenomedullin (ADM). In KO mice, this was not observed. H/R-induced liver injury in HIF-1 α KO was comparable to WT. Elevated plasma interleukin-6 (IL-6) levels after H/R were not reduced by HIF-1 α KO. Local hepatic hypoxia was not significantly reduced in HIF-1 α KO compared to controls after H/R. H/R-induced NF- κB phosphorylation in liver did not significantly differ between WT and KO., Conclusions: Here, deleting HIF-1 α in myeloid cells and thereby in Kupffer cells was not protective after H/R. This data indicates that other factors, such as NF- κB, due to its upregulated phosphorylation in WT and KO mice, contrary to HIF-1 α, are rather key modulators of inflammation after H/R in our model.

Background: In hemorrhagic shock with subsequent resuscitation (H/R), increased pro-inflammatory changes contribute to tissue injury and mortality in rodent models. Ethanol (EtOH) is assumed to modulate the inflammatory response and the subsequent organ injury after H/R. Therefore, we determined the contribution of acute ethanol gavage on intestinal inflammation and injury as well as survival after H/R in rats., Methods: Fourteen hours before H/R, female LEWIS rats were gavaged with single dose of EtOH or saline (5 g/kg, 30% EtOH, H/R&#95;EtOH group or H/R&#95;ctrl group). Then, rats were hemorrhaged to a mean arterial blood pressure of 30 ± 2 mmHg for 60 min and resuscitated. Control groups underwent surgical procedures and gavage without H/R (sham&#95;ctrl group and sham&#95;EtOH group). Tissue was harvested 2 h after resuscitation. Mortality was assessed 72 h after H/R., Results: Ethanol gavage increased survival after H/R from 20% to 80%, but amplified plasma alanineaminotransferase (ALT) release compared to saline gavage (2847 ± 406 vs. 1159 ± 200 IU/L, p < 0.05). Intestinal mucosal damage index, intestinal permeability, ileal myeloperoxidase levels as indicators of polymorphonuclear leukocyte (PMNL) infiltration and systemic IL-6 levels as well as ileal IL-6 and TNF gene expressions after H/R were reduced and partly restored after ethanol gavage when compared to the saline gavaged group after H/R., Conclusions: Taken together, we propose that acute ethanol gavage prior to H/R 1) did not enhance intestinal mucosa injury after H/R and 2) suppressed the H/R-induced inflammatory response. Both findings seem to contribute to the ethanol-induced survival benefit after H/R in our model.

Background and Purpose: There is an escalating prevalence of obesity in youth that increases the risk for cardiovascular alterations such as left ventricular hypertrophy (LVH). The purpose of this study is to identify the most effective electrical voltage measurement for determining LVH in youth who are overweight and obese., Methods: A retrospective chart review was conducted to determine sensitivity, specificity, and the receiver operator characteristic (ROC) curve of 4 popular electrical voltage measures., Results: Our findings indicated the sensitivity and specificity for Cornell product (50.0%; 96.2%), Cornell voltage (52.9%; 98.0%), Romhilt Estes (50.0%; 100.0%), and Sokolow-Lyon index (60.0%; 86.4%) consecutively., Conclusion: The Romhilt-Estes and Cornell voltage measures displayed the highest specificity and could prove to be beneficial as a screening method to rule out LVH in overweight and obese youth.

Sucrase-isomaltase (SI) is a highly N- and O-glycosylated intestinal brush border membrane protein. SI is sorted with high fidelity to the apical membrane via O-linked glycans that mediate its association with lipid rafts or detergent-resistant membranes (DRMs). Here, we show that N- and O-glycosylation and DRMs are implicated in the regulation of the function of SI in intestinal Caco-2 cells. The activities of sucrase (SUC) and isomaltase (IM) increase substantially in DRMs when N- and O-glycosylation patterns are intact. Disruption of DRMs by solubilization with Triton X-100 at 37 degrees C substantially reduces the activities of SUC and IM. Furthermore, modulation of O-glycosylation by benzyl-2-acetamido-2-deoxy-alpha-d-galactopyranoside and N-glycosylation by deoxymannojirimycin is linked to a decreased capacity of SI to associate with DRMs, with a subsequent reduction of the enzymatic activities of SUC and IM. This is the first report that reveals a direct role of N- and O-glycans in association with DRMs in regulating the function of a membrane glycoprotein.

Background: Albuterol is a beta-2 agonist that has been demonstrated to increase muscle strength in studies in animals and humans. Based on a pilot study of extended-release albuterol Repetabs in children with dystrophinopathies, the authors conducted a randomized, double-blind, placebo-controlled study with a crossover design., Methods: Fourteen boys with Duchenne or Becker muscular dystrophy, 6 to 11 years old, completed two treatment periods (albuterol and placebo), 12 weeks each, separated by a 12-week washout period. As the albuterol Repetab formulation was no longer available, an alternate extended release albuterol was used (Volmax, 12 mg per day). Outcome measurements included 1) lean body mass, 2) fat mass, 3) isometric knee extensor and flexor moments, 4) manual muscle testing, and 5) timed functional tests., Results: Lean body mass was significantly higher for subjects following albuterol treatment compared to placebo treatment, while fat mass was significantly lower. No differences were found in isometric knee moments or manual muscle tests. Time to run/walk 30 feet was improved following albuterol., Conclusions: Short-term treatment with extended release albuterol may increase lean body mass, decrease fat mass, and improve functional measures in patients with dystrophinopathies. However, the significant change in strength of specific muscle groups found in the pilot study was not observed in the present study. These findings may be attributed to differences in the drug release and kinetics between Repetab and Volmax formulations as they affect the concentration of available beta-2 receptors on the muscle cell surface differently.

Human aminopeptidase N (APN) is used as a routine marker for myelomonocytic cells in hematopoietic malignant disorders. Its gene and surface expressions are increased in cases of malignant transformation, inflammation, or T cell activation, whereas normal B and resting T cells lack detectable APN protein expression. In this study we elucidated the intracellular distribution, expression pattern, and enzymatic activity of a naturally occurring mutation in the coding region of the APN gene. At physiological temperatures the mutant protein is enzymatically inactive, persists as a mannose-rich polypeptide in the endoplasmic reticulum, and is ultimately degraded by an endoplasmic reticulum-associated degradation pathway. It shows in part the distinct behavior of a temperature-sensitive mutant with a permissive temperature of 32 degrees C, leading to correct sorting of the Golgi compartment accompanied by the acquisition of proper glycosylation but without reaching the cell-surface membrane and without regaining its enzymatic activity. Because the patient bearing this mutation suffered from leukemia, possible links to the pathogenesis of leukemia are discussed.

One sorting mechanism of apical and basolateral proteins in epithelial cells is based on their solubility profiles with Triton X-100. Nevertheless, apical proteins themselves are also segregated beyond the trans-Golgi network by virtue of their association or nonassociation with cholesterol/sphingolipid-rich microdomains (Jacob, R., and Naim, H. Y. (2001) Curr. Biol. 11, 1444-1450). Therefore, extractability with Triton X-100 does not constitute an absolute criterion of protein sorting. Here, we investigate the solubility patterns of apical and basolateral proteins with other detergents and demonstrate that the mild detergent Tween 20 is adequate to discriminate between apical and basolateral proteins during early stages in their biosynthesis. Although the mannose-rich forms of the apical proteins sucrase-isomaltase, lactase-phlorizin hydrolase, aminopeptidase N, and dipeptidylpeptidase IV reveal similar solubility profiles comprising soluble and nonsoluble fractions, the basolateral proteins, vesicular stomatitis virus G protein, major histocompatibility complex class I, and CD46 are entirely soluble with this detergent. The insoluble Tween 20 membranes are enriched in phosphatidylinositol and phosphatidylglycerol compatible with their synthesis in the endoplasmic reticulum and the existence of a novel class of detergent-resistant membranes. The association of the mannose-rich biosynthetic forms of the apical proteins, sucraseisomaltase, lactase-phlorizin hydrolase, aminopeptidase N, and dipeptidylpeptidase IV with the Tween 20-resistant membranes suggests an early polarized sorting mechanism prior to maturation in the Golgi apparatus.

Given the volume of pediatric orthotopic heart transplants (OHTs) at several centers, it is now possible to generate pediatric-specific, single-center OHT survival data. The transplant experience for 152 pediatric OHT patients at our institution was reviewed. The following were noted for each patient: graft survival; immunosuppressant therapy; initial diagnosis; cause of graft failure; clinical status at time of transplant; donor and recipient blood type, sex, weight, and age; ischemic time; previous cardiac surgery; race; and immune status. A series of Kaplan-Meier survival curves were constructed. Univariate comparisons of survival curves were performed with the Breslow test to determine equality of each pair of curves. Only immunosuppression with tacrolimus and an initial diagnosis of noncongenital heart disease positively influenced survival in pediatric OHT patients (p < or = 0.021 and p < or = 0.03, respectively). The more recently transplanted patients, managed with tacrolimus, had less mortality early after OHT (acute rejection) and less mortality during the period 2 or 3 years after OHT. No other factors, including prior cardiothoracic surgery, sex matching, and race matching, significantly influenced survival. Recently transplanted patients managed with tacrolimus-based immunosuppression and patients with noncongenital cardiomyopathy have significantly superior graft survival.

The authors conducted a randomized, crossover, double-blind, placebo-controlled pilot study of albuterol in nine boys with dystrophinopathies. Primary outcomes were 1) isometric knee extensor and flexor strength; and 2) manual muscle testing (MMT). Isometric knee extensor strength and MMT scores were significantly higher in patients taking albuterol vs placebo. Therefore, 12-week treatment with extended-release albuterol may increase strength in patients with dystrophinopathies. A larger, double-blind, randomized study is necessary to confirm these results.

Postoperative arrhythmia is a major cause of morbidity and mortality after cardiac surgery for congenital heart disease. Rhythm disturbances that may be well tolerated in a normal heart often cause hemodynamic instability when they occur in the immediate postoperative period. In the face of pre-existing myocardial dysfunction resulting from preoperative pressure or volume overload, patients with congenital heart conditions are especially vulnerable to rhythm disturbances after cardiac surgery. Cardiopulmonary bypass, intraoperative injury to the conduction system and myocardium, postoperative metabolic abnormalities, electrolyte disturbances, and increased adrenergic tone in response to the stress of the surgery or inotropic agents are also known factors associated with increased risk of arrhythmia in the immediate postoperative period. Surgically related arrhythmia can also present in the late postoperative period, particularly in association with surgical incision sites and surgically induced hemodynamic abnormalities. Early and late postoperative arrhythmias are important risk factors for morbidity and mortality after surgical treatment of many forms of congenital heart disease. This review describes the incidence of the most common forms of arrhythmia and recent advances in their diagnosis and treatment.

Background: Although amiodarone has been referred to as a class III antiarrhythmic agent, it also possesses electrophysiologic characteristics of the three other classes (classes I and IV and minor class II effects). Previous studies have demonstrated that amiodarone inhibits Ca2+ channel current in intact cardiac myocytes. However, it is not clear whether this response reflects a pure class IV effect (direct Ca2+ channel inhibition) or a class II effect (beta-adrenergic receptor blockade) of amiodarone., Methods: In the current study, the effects of amiodarone on Ca2+ current were studied in the absence of sympathetic regulation using a Xenopus oocyte expression system. The L-type Ca2+ channel alpha1C subunit was coexpressed with the alpha2delta and beta2a subunits in enzymatically digested Xenopus oocytes. Ca2+ currents were recorded using the cut-open oocyte preparation., Results: We found that perfusion of 10 microM isoproterenol produced no significant change in peak Ca2+ current (from 223+/-33 to 210+/-29 nA, mean+/-SEM, n=5, P=not significant), indicating the absence of a functional stimulatory sympathetic signal pathway in these oocytes. After 10 minutes of exposure to 10 microM amiodarone, Ca2+ current amplitude was significantly decreased from 174+/-33 to 100+/-26 nA (n=8, P<0.01; control group: 220+/-33 to 212+/-29 nA, n=5, P=not significant). These effects were similar to those of 10 microM nifedipine (201+/-48 to 108+/-48 nA, n=6, P<0.05), a typical Ca2+ channel blocker. On the other hand, neither amiodarone nor nifedipine significantly altered the Ca2+ current activation or inactivation kinetics., Conclusions: These results demonstrate that amiodarone inhibits Ca2+ current in the absence of a functional intrinsic beta-adrenergic stimulatory system and, therefore, represents a true class IV effect.

The purpose of the present study was to examine the role of G(i2)alpha in Ca(2+) channel regulation using G(i2)alpha gene knockout mouse ventricular myocytes. The whole cell voltage-clamp technique was used to study the effects of the muscarinic agonist carbachol (CCh) and the beta-adrenergic agonist isoproterenol (Iso) on cardiac L-type Ca(2+) currents in both 129Sv wild-type (WT) and G(i2)alpha gene knockout (G(i2)alpha-/-) mice. Perfusion with CCh significantly inhibited the Ca(2+) current in WT cells, and this effect was reversed by adding atropine to the CCh-containing solution. In contrast, CCh did not affect Ca(2+) currents in G(i2)alpha-/- ventricular myocytes. Addition of CCh to Iso-containing solutions attenuated the Iso-stimulated Ca(2+) current in WT cardiomyocytes but not in G(i2)alpha-/- cells. These findings demonstrate that, whereas the Iso-G(s)alpha signal pathway is intact in G(i2)alpha gene knockout mouse hearts, these cells lack the inhibitory regulation of Ca(2+) channels by CCh. Therefore, G(i2)alpha is necessary for the muscarinic regulation of Ca(2+) channels in the mouse heart. Further studies are needed to delineate the possible interaction of G(i) and other cell signaling proteins and to clarify the level of interaction of G protein-coupled regulation of L-type Ca(2+) current in the heart.

The purpose of the present study was to determine whether age-related changes in the expression and function of the cardiac isoform of the sarcoplasmic reticulum Ca(2+)-ATPase (SERCA2a) play a role in SR Ca(2+)release and cell contraction. SERCA2a protein levels and subcellular localization were compared between fetal, neonatal, juvenile and adult New Zealand White rabbits. Studies of SERCA function in isolated myocytes were performed in situ by examining the rate of reloading of the SR Ca(2+)stores following caffeine-induced depletion. We found that significant quantities of SERCA2a were present early in immature heart and that SERCA2a expression reached adult levels within 15-30 days after birth. Furthermore, SERCA2a protein is present as a series of transverse striations within the cell as early as 1 day of age. In contrast to previous studies of SERCA in vitro, the SERCA protein function in situ was found to be comparable between neonatal and adult myocytes in maintaining SR Ca(2+)stores. These results indicate that the paucity of SR Ca(2+)release in immature ventricular cardiac myocytes is not the result of immaturity in SERCA2a expression., (Copyright 2000 Academic Press.)

Canine junctin is a 26-kDa transmembrane protein found in the sarcoplasmic reticulum (SR) membrane in cardiac and skeletal muscle. Junctin has recently been shown to bind directly to calsequestrin, the ryanodine receptor, and triadin. Junctin is thought to play a role in facilitating (and perhaps regulating) Ca(2+) release from the SR. Immature heart exhibits decreased utilization of SR Ca(2+) stores for cell contraction. We have cloned human and rabbit cardiac junctin and investigated the expression of junctin in developing rabbit heart. Human junctin was cloned from an adult cardiac cDNA library. Rabbit junctin was cloned by RT-PCR. Northern blot analysis demonstrates a single primary mRNA transcript of approximately 2.8 kb in hearts from both species. Sequence analysis demonstrates greater than 97% homology between the predicted amino acid sequences of human, rabbit, and canine junctin in the putative transmembrane domain and subsequent initial 61 amino acid portion of the putative luminal domain. These domains also exhibit sequence homology with triadin. The C-terminal region shows much lower (72 to 75%) sequence homology among the three species. In addition, Northern blot analysis demonstrates that the expression of junctin increases markedly in postnatal rabbit myocardium. These findings suggest that the putative transmembrane domain and subsequent initial portion of the putative luminal domain of junctin play an important role in the binding of junctin to calsequestrin, the ryanodine receptor, and triadin in the postnatal heart. Furthermore, the previously described increase in SR Ca(2+) release with development is associated with the increased expression of junctin., (Copyright 2000 Academic Press.)

Measurements from the winter of 1994-95 indicating removal of total reactive nitrogen from the Arctic stratosphere by particle sedimentation were used to constrain a microphysical model. The model suggests that denitrification is caused predominantly by nitric acid trihydrate particles in small number densities. The denitrification is shown to increase Arctic ozone loss substantially. Sensitivity studies indicate that the Arctic stratosphere is currently at a threshold of denitrification. This implies that future stratospheric cooling, induced by an increase in the anthropogenic carbon dioxide burden, is likely to enhance denitrification and to delay until late in the next century the return of Arctic stratospheric ozone to preindustrial values.

The purpose of this article is to alert nurse practitioners to elements of the pediatric history or physical examination that could herald urgent/emergent conditions. Five common symptoms along with suggested key questions and physical examination parameters for evaluation of these possible "red flags" are presented.

Radiofrequency (RF) ablation is a nonsurgical technique using catheter-directed RF energy for treating cardiac arrhythmias in children and adults. Previous reports have suggested that sequestration of calcium (Ca2+) by the sarcoplasmic reticulum may partially protect mature cardiac myocytes from the effects of RF energy. The purposes of this study were to determine whether differences exist between neonatal and adult myocyte responses to RF energy and if myocyte damage is a Ca2+-dependent process. Because immature myocardium is functionally deficient in sarcoplasmic reticulum, we hypothesized that immature myocytes would be more susceptible to damage induced by RF energy. Isolated ventricular myocytes were obtained from neonatal and adult New Zealand White rabbits by enzymatic dissociation, then placed in a perfusion chamber designed to deliver RF energy or a heated perfusate solution. Measurements of bath temperature, cell morphology, and contractile response to electrical stimuli were recorded. RF energy application associated with increased perfusate temperature resulted in cell death, but not when the temperature rise was inhibited. Thus, the acute damage to cells exposed to RF energy appears to be mediated by thermal energy. After exposure to thermal energy, neonatal cells underwent contracture at lower temperatures than did adult cells. Perfusion with solutions containing low Ca2+ concentrations, comparable to intracellular diastolic Ca2+ levels, had a protective effect for both neonatal and adult myocytes. These findings indicate that acute cell damage after exposure to RF energy is mediated by a Ca2+-dependent process. Furthermore, immature myocardium is particularly susceptible to RF-mediated cell damage, possibly secondary to reduced Ca2+ sequestration by the sarcoplasmic reticulum.

Mature myocardium utilizes calcium released by the sarcoplasmic reticulum (SR) for cell contraction. Transient exposure of mature myocytes to caffeine is known to directly trigger Ca2+ release from the SR. In contrast, neonatal rabbit heart cells rely on transsarcolemmal Ca2+ influx for tension generation. SR function is decreased in immature heart and appears to play a minimal role as a calcium source. Accordingly, we hypothesized that neonatal rabbit myocytes would not respond to a caffeine pulse. Isolated neonatal and adult myocytes were paced to load the SR with calcium and then exposed to a 1-s pulse of 10 mM caffeine. As previously described, adult myocytes exhibited a brisk contraction in response to caffeine. Unexpectedly, neonatal myocytes also exhibited a similar, brisk response. These caffeine-induced contractions were not dependent on extracellular Ca2+ but were dependent upon the loading of SR Ca2+ stores. When SR Ca2+ stores were depleted by exposure to caffeine, mature myocytes exhibited only small, slow contractions in response to electrical field stimulation. Replenishing the SR Ca2+ stores resulted in normal, brisk contractions. In contrast, electrically stimulated contractions in immature myocytes were largely unaffected by caffeine-induced SR depletion. Thus, although neonatal myocytes are capable of loading and releasing calcium from the SR, such SR calcium release is not normally required for contraction in the developing heart. The minor role of SR Ca2+ release in immature rabbit heart may not result from immaturity of the SR, but rather from an inadequate mechanism to trigger SR calcium release.

For human B lymphocytes, Epstein-Barr virus (EBV) is a polyclonal activator, inducing both proliferation and Ig secretion. It is also a transforming virus capable of generating immortalized B cell lines. These early and late functions of EBV are not apparently connected. The receptor for EBV, CD21, also serves as a receptor for some complement components and is called CR2. This molecule associates with CD19 and TAPA-1 on the surface of B cells. This complex is involved in signaling B cells and participates in many responses. We have observed that simultaneous ligation of CD40 and the CD21 complex, by exposure to anti-CD40 MAbs and EBV, enhances both the short-term proliferation as well as the long-term transformation rate of human B lymphocytes. B cell proliferation shows synergy between anti-CD40 MAb and EBV. CD19 also appears to be involved in the synergistic activation of B cells through CD40 and CD21, since ligation of CD19 with anti-CD19 MAbs, either prior to or concomitant with exposure to anti-CD40 and EBV, markedly inhibits both proliferation and subsequent B cell transformation. These observations do not elucidate the mechanisms of B cell transformation employed by EBV but the do suggest a relationship between early proliferation and later transformation induced by the virus. Anti-CD40 enhances both these effects and anti-CD19 is capable of inhibiting both.

Background: Amiodarone is an effective antiarrhythmic drug used to treat a wide variety of ventricular and supraventricular tachyarrhythmias. Recent voltage clamp studies indicate that amiodarone may possess a variety of antiarrhythmic effects., Methods: The tight-seal, whole-cell voltage clamp technique was used to investigate the acute effects of amiodarone on L-type Ca2+ channel kinetics in isolated neonatal ventricular myocytes., Results: We found that acute perfusion with 1 mumol/L amiodarone inhibited peak inward Ca2+ current by 39.1% (4.85 +/- 0.42 to 2.95 +/- 0.6 pA/pF, n = 10, p < 0.001) without changing the shape of the current-voltage relation. In addition, amiodarone shifted Ca2+ channel steady-state inactivation to more negative membrane potentials. In the absence of amiodarone, half inactivation of the Ca2+ current occurred at a membrane potential of -23.8 +/- 0.2 mV compared to -34.2 +/- 0.6 mV after addition of amiodarone (n = 11, p < 0.01). Furthermore, amiodarone significantly delayed Ca2+ current recovery from previous inactivation., Conclusions: These results provide evidence that amiodarone blocks voltage-dependent Ca2+ current in isolated neonatal rabbit ventricular myocytes by a variety of different mechanisms. The inhibitory effect of amiodarone on L-type Ca2+ current may represent an important facet of amiodarone's acute antiarrhythmic activity in the immature heart.

This study was designed to assess the effects of isoflurane (ISO) on the electrophysiological properties of the accessory pathway, atrium, ventricle, and AV node in children with the Wolff-Parkinson-White (WPW) syndrome. The results of programmed electrical stimulation were analyzed in 51 patients (4 months to 17 years of age) with WPW. The study population was divided into two groups. Twenty-seven patients received local anesthesia and intramuscular injection of meperidine, promethazine, and chlorpromazine (MPC group). Twenty-four patients received general anesthesia with ISO inhalation (ISO group). We compared the antegrade effective refractory period of the accessory pathway (antegrade APERP), ventricular effective refractory period (VERP), atrial effective refractory period (AERP), AH interval, and cycle length of circus movement tachycardia (CMT-CL) in 12 pairs of age and sex matched patients selected from the MPC and ISO groups. Of the 12 pairs of age and sex matched patients, antegrade APERP in patients who received ISO (299 +/- 17 ms, mean +/- SEM) was significantly longer as compared with matched patients in the MPC group (262 +/- 5 ms, P < 0.025). The VERP and AERP in patients from the ISO group were significantly prolonged compared with the MPC patients (239 +/- 7 vs 210 +/- 8 ms, P < 0.025, and 228 +/- 11 vs 180 +/- 6 ms, P < 0.01, respectively). There was no significant difference in the AH interval or CMT-CL between the two subgroups. Thus, ISO prolongs the antegrade APERPs as well as the effective refractory periods of atrial and ventricular muscle in children with WPW, while the AH interval and CMT-CL appear to be unaffected. Care must be taken in interpreting measurements of the antegrade APERP made in patients under general anesthesia for RF ablation of accessory pathways.

Protons inhibit Ca2+ current and contraction in heart muscle. The present study compares the effects of lowering the pH of the bath solution on single-cell contraction, action potential configuration and Ca2+ currents between neonatal and adult rabbit hearts. We found that a reduction of extracellular pH from 7.3 to 6.3 decreased cell contraction amplitude to 84.3% of control in isolated neonatal myocytes. A comparable change in extracellular pH resulted in a decrease in cell contraction to 56.2% of control in adult cells. Similarly, tension generation in intact neonatal papillary muscles was less sensitive to a decrease in external pH as compared to papillary muscles from adult animals. In addition, acidosis caused a less pronounced inhibition of Ca2+ current in neonatal cells than in adult cells (85 +/- 4% nu 62 +/- 4% of control, pH = 6.3, P < 0.001; 63 +/- 5% nu 32 +/- 5% of control, pH = 5.8, P < 0.001). Thus, the effect of external acidosis on myocardial contractility is commensurate with the effect on trans-sarcolemmal Ca2+ current. The membrane potential at which peak Ca2+ current occurred was not altered by low pH in neonatal cells but was shifted toward positive potentials by 17.7 mV in adult myocytes. Further, low external pH solution reduced Ca2+ current conductance more in adult cells than in neonatal cells. Moreover, action potential configuration in neonatal cells was altered less by acidosis as compared with adult cells. These findings may help explain the greater resistance of neonatal hearts to extracellular acidosis.

This overview of cardiac ion channel development does not suggest any particular theme underlying the expression or regulation of all channel subtypes. Calcium and potassium channels generally exhibit increased expression in more mature hearts. However, this increase in channel number or activity as determined under voltage clamp conditions may not be translated into increased activity in vivo. Concomitant changes in other physiological factors such as local intracellular Ca2+ accumulation, increased resting membrane potential and decreased heart rate in mature heart may inhibit or augment channel activity. Na(+)-Ca2+ exchange activity appears to decrease with development, possibly reflecting its decreasing role in both systolic and diastolic Ca2+ regulation. Na+ channel activity follows a middle course, exhibiting little change in channel conductance. The reported shift in the voltage dependence of channel inactivation toward more negative membrane potentials may reflect a concomitant shift in the resting membrane potential in mature heart. However, this change is in the direction opposite to that reported for L-type Ca2+ channel inactivation, suggesting that the regulation of these channels is not modulated by a common factor such as membrane surface charge. A detailed characterization of multiple channel subtypes in mature myocardium has resulted in significant advances in models of the cardiac action potential and excitation-contraction coupling. Recently, developmental changes in ion channel physiology have been described, setting the stage for a comparable elucidation of the ontogeny of the cardiac action potential. Ca2+ and K+ channel currents generally become more prominent with development. In contrast, developmental changes in Na+ currents are less dramatic and Na(+)-Ca2+ exchange currents appear to decrease with age. These changes may, in part, be reflected by the increasingly important role of transsarcolemmal Ca2+ influx in triggering Ca2+ release from the SR in mature heart as compared to its direct role of providing Ca2+ for cell contraction in immature heart. These developmental changes in ion channel expression and function are likely to have a significant effect on the generation of the action potential in individual myocytes. Developmental changes in the characteristics of the action potential may then have a major influence on the initiation, propagation and termination of autonomic, triggered, and re-entrant arrhythmias. Progress in this area provides an essential foundation for the evolution of a systematic approach to pediatric arrhythmias comparable to that under development for mature heart [3].

Cardiac responsiveness to beta-adrenergic stimulation changes with age. Developmental changes in expression of guanine nucleotide-binding coupling protein (G protein) subunits may account for these physiologic changes. We measured steady-state levels of mRNA encoding the alpha-subunit of the specific G protein that stimulates adenylyl cyclase (Gs alpha) and three isoforms of beta-subunit of G proteins (G beta) in developing myocardium. Total RNA prepared from the right and left ventricles of fetal, neonatal, juvenile, and adult rabbits was size-fractionated, blotted, and probed with 32P-labeled cDNAs encoding rat Gs alpha, bovine G beta-1, human G beta-2, and human G beta-3. For standardization, these blots were subsequently hybridized with a 32P-labeled cDNA encoding glyceraldehyde 3-phosphate dehydrogenase (GAPD). Two-dimensional densitometric analysis of autoradiographs was used to quantify relative hybridization intensities. An age-dependent decrease in mRNAs encoding Gs alpha, G beta-1, and G beta-2 relative to mRNA encoding GAPD was observed in both ventricles, while G beta-3 mRNA was not detected. At all ages studied, levels of Gs alpha and G beta-1 mRNA were similar in the two ventricles. However, G beta-2 mRNA declined more in the left ventricle than in the right ventricle during maturation. Our results demonstrate developmental control in heart for mRNAs encoding several G protein subunits. In addition, differential declines in G beta-1 and G beta-2 mRNA in the right ventricle suggest that these G beta isoforms are regulated uniquely and may reflect functional roles for these G beta isoforms in different signaling cascades.

The anterograde effective refractory period of the accessory pathway is age-dependent in pediatric patients with the WPW syndrome. Thus, age should be considered when developing electrophysiologic criteria for the risk of hypotensive arrhythmias in these patients. In addition, general anesthesia must also be considered in interpreting age-related changes in the anterograde APERP, especially in children.

We investigated the effects of added beta gamma subunits of G proteins (G beta gamma) on beta-adrenergic responsiveness of transmembrane Ca2+ currents (ICa) in ventricular myocytes from neonatal rabbits. G beta 1 gamma 1 purified from retinal rods was dialyzed into cells via the voltage clamp micro-electrode. Stimulation of ICa by isoproterenol was not affected by added intracellular G beta 1 gamma 1 or by carbachol alone but was completely blocked by combined G beta 1 gamma 1 and carbachol. Pretreatment of cells with pertussis toxin or temporal separation of carbachol and isoproterenol allowed stimulation of ICa by isoproterenol in cells dialyzed with G beta 1 gamma 1. Carbachol and G beta 1 gamma 1 together also did not prevent stimulation of ICa by dibutyryl-cyclic AMP. Thus, rather than simply inactivating Gs alpha by mass action, G beta 1 gamma 1 acts in concert with carbachol to inhibit isoproterenol stimulation of ICa.

Recent studies have demonstrated a relative deficiency in voltage-gated Ca2+ currents (ICa) in immature myocardium. We hypothesized that contraction in developing heart results in part from Ca2+ influx via "reverse" Na+/Ca2+ exchange current (INa/Ca). Accordingly, INa/Ca and cell contraction amplitude were measured in single neonatal and adult rabbit ventricular myocytes. INa/Ca was dependent on Ca2+ concentration, Na+ concentration, and membrane potential and was blocked by 5 mM Ni2+ but not by the Ca(2+)-channel blocker nifedipine. In neonatal cells, contraction amplitude reached a plateau for depolarizations positive to 0 mV. In adult myocytes, contraction amplitude was maximal at 0 mV and decreased at positive membrane potentials. Inhibition of ICa with nifedipine did not affect maximal contraction amplitude in neonatal myocytes but almost completely suppressed contraction of adult cells. These data suggest that Ca2+ influx via ICa is not required for contraction of neonatal rabbit cardiac myocytes. Moreover, Ca2+ influx via reversal of the Na+/Ca2+ exchange mechanism may provide a significant portion of the Ca2+ regulating cell contraction, especially during depolarization to positive membrane potentials.

High-resolution IR remote-sensing measurements from space by means of a nadir-viewing geometry are particularly suited to the detection of trace gases and yield high temporal and horizontal resolutions on a global scale. To identify the potential of such a technique, an extensive feasibility study has been performed. The column amount of some trace gases, namely H(2)O, CH(4), N(2)O, CO, and O(3), may be determined with accuracies of approximately 10%. In addition, some information on the vertical distribution of these species is also possible. Concerning CFC-12, an accuracy of 10%-20% may be expected. Furthermore, it is believed that column amounts can be derived with an accuracy of 20% for HNO(3), and 50% for species like NO(2), OCS, and CFC-11.

To examine the ability of normal and autoimmune individuals to produce circulating anti-TNF alpha antibodies, plasma samples from 10 RA patients, 10 SLE patients and 5 normal subjects were assessed for anti-TNF alpha antibody. While every individual tested demonstrated circulating IgM anti-TNF alpha antibody, IgG anti-TNF alpha autoantibody was seen predominantly in autoimmune patients. Only 1 of 5 normal individuals, but 15 of 20 autoimmune individuals had plasma IgG anti-TNF alpha antibodies. To examine the ability of normal and autoimmune individuals to produce anti-TNF alpha autoantibody from their circulating lymphocytes, EBV transformation was performed. Oligoclonal immortal cell lines were successfully established from 13 patients and each one secreted detectable IgM anti-TNF alpha autoantibody. Transformed cells from only 1 of 5 normal individuals secreted IgM anti-TNF alpha autoantibody. These results indicate a higher prevalence of anti-TNF alpha autoantibody production among autoimmune individuals although normal individuals are also capable of producing these autoantibodies.

Neonatal myocardium is distinctly more sensitive to extracellular calcium levels than is mature myocardium. This has been ascribed to the poorly developed sarcoplasmic reticulum of neonatal myocardium. Recent evidence has suggested that there is an increased dependence of neonatal myocardium on the sodium-calcium exchange current, and that sodium-calcium exchange may be a major source of calcium influx in neonatal myocardial cells. We determined the effect of halothane on the sodium-calcium exchange current on single neonatal (2- to 5-day-old) rabbit ventricular myocytes by means of the whole cell voltage clamp. Lower (1.5%) halothane decreased sodium-calcium exchange current by 49%, from 29 +/- 3 to 15 +/- 6 pA. Higher (3%) halothane decreased this current by 66%, from 50 +/- 9 to 17 +/- 9 pA. Thus halothane has a reversible inhibition of sodium-calcium exchange current in neonatal myocardium. Inhibition of sodium-calcium exchange current would be expected to have a magnified effect on contractility in neonatal as opposed to adult myocardium, and could theoretically ameliorate reperfusion injury due to influx of calcium via the sodium-calcium exchanger.

We evaluated the effects of clinically relevant concentrations of halothane (1%) and ketamine (10(-4) M) on activation, inactivation, and recovery from inactivation of voltage-gated sarcolemmal calcium current (ICa) in single guinea pig ventricular myocytes, using the whole cell voltage clamp. Both anesthetics had qualitatively similar effects. The potential at half-activation was shifted from -18 to -23 mV for halothane (p < 0.03) and from -17 to -21 mV for ketamine (p = 0.005). There was no change in the slope of the activation curve for either anesthetic. The potential at half-inactivation was shifted from -29 to -40 mV with exposure to halothane (p < 0.001) and from -27 to -33 mV (p < 0.001) with exposure to ketamine. There was no change in the slope of the inactivation curve with either agent. The changes in time constant of recovery from steady-state inactivation with halothane did not reach statistical significance (178 vs. 207 ms, p = 0.20) and was significantly prolonged with exposure to ketamine (106 vs. 157 ms, p = 0.005). The two anesthetics show parallel shifts in activation, inactivation, and recovery from inactivation of ICa in ventricular myocardial cells. These findings in normal ventricular myocytes may help interpret the interactions of these anesthetics with other types of heart muscle, such as ischemic and immature myocardium.

Several monoclonal human IgM antibodies to recombinant human tumor necrosis factor-alpha (rhTNF alpha) have been generated and partially characterized. The F78-1A10-B5 monoclonal antibody (mAb) (B5) binds to rhTNF alpha with a titer comparable to three high-affinity neutralizing mouse mAbs, when tested by ELISA. However, the B5 mAb binds relatively weakly to soluble rhTNF alpha. It appears to bind to epitopes on rhTNF alpha distinct from those bound by the mouse mAbs for three reasons. First, preincubation of plate-bound rhTNF alpha with mouse mAbs does not decrease or compete subsequent B5 mAb binding. Second, rhTNF alpha complexed to the mouse mAbs can still be bound by B5 mAb. Third, the mouse mAbs neutralize TNF alpha cytotoxicity whereas the B5 mAb does not. Binding analyses indicate that this human IgM autoantibody binds to both human and mouse recombinant TNF alpha, but not to other antigens commonly recognized by polyreactive natural IgM autoantibodies. The high level of amino acid identity between the human and mouse TNF alpha molecules suggest that the B5 mAb is monospecific for a given epitope shared by these two forms of TNF alpha. This spectrum of characteristics makes B5 a novel mAb.

A human IgM monoclonal antibody (B5) recognizing human TNF alpha was established from peripheral blood lymphocytes by transformation with Epstein-Barr virus and subsequent cell fusion. The B5 monoclonal antibody (mAb) binds to cell surface TNF alpha (csTNF alpha) on human T cells, B cells, and monocytes. In addition, this autoantibody binds to csTNF alpha on a variety of lymphoid and monocyte lineage cell lines of human origin, as well as astrocytomas, a breast carcinoma, and a melanoma. Interestingly, the B5 mAb also binds to chimpanzee lymphocytes and to mouse T lymphoma cell line csTNF alpha. Many neutralizing mouse anti-TNF alpha mAbs do not exhibit comparable binding to csTNF alpha. This is consistent with the previous demonstration that B5 recognizes an epitope on TNF alpha distinct from those recognized by three neutralizing mouse anti-TNF alpha mAbs. B5 binding to csTNF alpha is specific since it can be inhibited by TNF alpha. No inhibition of B5 binding was seen by a neutralizing mouse anti-TNF alpha mAb. The B5 autoantibody appears to recognize the transmembrane form of TNF alpha and most likely also recognizes TNF alpha associated with its receptor. The unique specificity of this B5 autoantibody provides some additional insight into the complex physiology of cell surface-associated TNF alpha.

Measurement of transsarcolemmal voltage-gated Ca2+ current (ICa) in myocytes isolated from immature rabbit heart has demonstrated an unexpectedly low level of Ca2+ channel activity. We have characterized the kinetic properties of ICa in acutely isolated 21-day fetal, 1-5-day-old neonatal, and adult cardiac myocytes by the whole-cell voltage-clamp technique. The membrane potential for half-maximal steady-state inactivation became less negative with maturation (-24 +/- 3 [mean +/- SEM] mV, n = 5; -19 +/- 2 mV, n = 5; and -11 +/- 2 mV, n = 6 for fetal, neonatal, and adult myocytes, respectively; p < 0.005). In contrast, the membrane potential for half-maximal steady-state activation was not statistically different among the age groups studied. These parameters accurately predicted the voltage dependence of the sustained ICa present at the end of a 400-msec depolarization. This "window" current was significantly smaller in immature cells than in adult cells and occurred at a more negative membrane potential in the younger age groups. The time course of inactivation of ICa was not significantly different between age groups. However, ICa was inhibited by increasing the frequency of stimulation. This effect was most prominent in immature cells, particularly at more positive holding potentials. This developmental alteration in the frequency dependence of ICa was due in part to a prolonged time constant of recovery from inactivation in the younger age groups. In summary, the kinetic properties of ICa in immature cardiac cells place them at a relative disadvantage in terms of the total Ca2+ influx during a depolarization. Thus, the role of ICa in the control of cell contraction may change with development.