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BACKGROUND: Healthcare providers (HPs) play a critical role in disseminating information about infant health and development. Two key developmental areas for infants are physical activity (PA) and communication. Few studies have examined HPs views of these topics. Thus, HPs may need support to promote these early development outcomes in infants. Thus, the study explored HPs' perceptions of infant PA and communication. METHODS: A total of 13 HPs with a wide variety of backgrounds were recruited. Zoom semi-structured interviews were based on the Health Belief Model. Deductive content analysis was utilized to analyze data. RESULTS: HPs knew little about PA guidelines. HPs were concerned about an infant's PA because of limited playtime and tummy time. Most HPs advised parents on PA and motor development milestones, with crawling, walking, and tummy time. HPs also wanted to learn about PA measurement tools. PA promotion was hindered by parents' busy schedules, lack of time, and language barriers. Most HPs advised parents on verbal and nonverbal communication. Many HPs were unfamiliar with communication assessment methods and wanted to learn more. CONCLUSIONS: HPs had limited knowledge about PA guidelines and communication measurement tools. Efforts are needed to identify easily accessible ways to educate HPs that could be disseminated to parents.
Subject(s)
Attitude of Health Personnel , Communication , Humans , Infant , Female , Male , Adult , Exercise/psychology , Child Development/physiology , Parents/psychology , Health Knowledge, Attitudes, Practice , Health Personnel/psychology , Health Promotion , Qualitative ResearchABSTRACT
Antibodies to high-frequency antigens are rarely implicated in cases of hemolytic disease of the fetus and newborn (HDFN), yet they pose a challenge to both clinical staff and transfusion medicine, especially with the identification of the implicating antibody and the arrangement of compatible blood for intrauterine transfusion. Here we report one such interesting case of HDFN caused by an alloantibody to a high-frequency antigen belonging to the Rhesus (Rh) blood group system. The patient presented at the 19th week with Rh-isoimmunized pregnancy. She received six intrauterine transfusions (IUTs) at different intervals during the antenatal period. Arranging the blood of this rare blood group required great efforts from hospital administration, clinicians, and social workers. At 31 weeks, the fetus developed a non-reassuring non-stress test (NST). Hence, the baby was delivered by cesarean section. The baby fared well in the neonatal period. With great efforts and support from social health workers, the Japanese Red Cross society, the administration, and non-government organizations, the impossible became possible.
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The present work reports the rapid sweat detection inside a PPE kit using a flexible humidity sensor based on hydrothermally synthesized ZnO (zinc oxide) nanoflowers (ZNFs). Physical characterization of ZNFs was done using scanning electron microscopy (SEM), X-ray diffraction (XRD), Fourier transmission infrared spectroscopy (FTIR), UV-visible, particle size analysis, Raman analysis, and X-ray photoelectron spectroscopy (XPS) analysis, and the hydrophilicity was investigated by using contact angle measurement. Fabrication of a flexible sensor was done by deposition on the paper substrate using the spin coating technique. It exhibited high sensitivity and low response and recovery times in the humidity range 10-95%RH. The sensor demonstrated the highest sensitivity of 296.70 nF/%RH within the humidity range 55-95%RH, and the rapid response and recovery times were also calculated and found as 5.10/1.70 s, respectively. The selectivity of the proposed sensor was also analyzed, and it is highly sensitive to humidity. The humidity sensing characteristics were theoretically witnessed in terms of the highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) and electronic properties of sensing materials in ambient and humid conditions. These theoretical results are evidence of the interaction of ZnO with humidity. Overall, the present study provides a scope of architecture-enabled paper-based humidity sensors for the detection of sweat levels inside PPE kits for health workers.
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BACKGROUND: Essential to a patient-centered approach to imaging individuals with chest pain is knowledge of differences in radiation effective dose across imaging modalities. Body mass index (BMI) is an important and underappreciated predictor of effective dose. This study evaluated the impact of BMI on estimated radiation exposure across imaging modalities. METHODS AND RESULTS: This was a retrospective analysis of patients with concern for cardiac ischemia undergoing positron emission tomography (PET)/computed tomography (CT), cadmium zinc telluride single-photon emission CT (SPECT) myocardial perfusion imaging, or coronary CT angiography (CCTA) using state-of-the-art imaging modalities and optimal radiation-sparing protocols. Radiation exposure was calculated across BMI categories based on established cardiac imaging-specific conversion factors. Among 9046 patients (mean±SD age, 64.3±13.1 years; 55% men; mean±SD BMI, 30.6±6.9 kg/m2), 4787 were imaged with PET/CT, 3092 were imaged with SPECT/CT, and 1167 were imaged with CCTA. Median (interquartile range) radiation effective doses were 4.4 (3.9-4.9) mSv for PET/CT, 4.9 (4.0-6.3) mSv for SPECT/CT, and 6.9 (4.0-11.2) mSv for CCTA. Patients at a BMI <20 kg/m2 had similar radiation effective dose with all 3 imaging modalities, whereas those with BMI ≥20 kg/m2 had the lowest effective dose with PET/CT. Radiation effective dose and variability increased dramatically with CCTA as BMI increased, and was 10 times higher in patients with BMI >45 kg/m2 compared with <20 kg/m2 (median, 26.9 versus 2.6 mSv). After multivariable adjustment, PET/CT offered the lowest effective dose, followed by SPECT/CT, and then CCTA (P<0.001). CONCLUSIONS: Although median radiation exposure is modest across state-of-the-art PET/CT, SPECT/CT, and CCTA systems using optimal radiation-sparing protocols, there are significant variations across modalities based on BMI. These data are important for making patient-centered decisions for ischemic testing.
Subject(s)
Coronary Artery Disease , Radiation Exposure , Male , Humans , Middle Aged , Aged , Female , Body Mass Index , Positron Emission Tomography Computed Tomography , Retrospective Studies , Radiation Dosage , Radiation Exposure/adverse effects , Chest Pain , Coronary Angiography/methodsABSTRACT
OBJECTIVE: This study aims to investigate the relationship between thyroid computed tomography (CT) density and serum thyroid stimulating hormone (TSH) levels in patients with hypothyroidism. METHODS: The research involved 60 hypothyroid patients aged 18 years and older, with TSH levels exceeding 5 mIU/mL or those already receiving thyroid supplementation therapy. These individuals had undergone non-contrast CT chest scans due to concurrent respiratory concerns. The thyroid CT densities, as evaluated through non-contrast chest CT, were compared to those of euthyroid patients with normal TSH levels. Correlation analysis was conducted to assess the relationship between serum TSH levels and thyroid CT densities. RESULTS: Hypothyroid patients exhibited significantly reduced thyroid CT densities (P<0.0001). The correlation analysis revealed an inverse correlation between TSH levels and thyroid CT densities in hypothyroid patients (r=-0.365, p=0.020) and a positive correlation (r=0.270, p=0.036) in euthyroid patients. CONCLUSION: This study demonstrates a substantial correlation between thyroid CT density and serum TSH levels. As TSH levels increase in hypothyroid patients, thyroid CT density tends to decrease.
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Objective: To study the perinatal outcome in fetuses diagnosed with congenital diaphragmatic hernia (CDH). Methods: Thirty-two pregnant women with antenatal diagnosis of CDH in fetus, who delivered between 2018 and 2021, were included in the study. Postnatally eventration of diaphragm was diagnosed in 3 neonates and were excluded. Results: The median gestational age at diagnosis was 23 weeks (IQR: 216-261 weeks). The mean O/E LHR was 34.88 ± 9.03%, and the O/E LHR was significantly lower in fetuses who did not survive (40.81 ± 4.25 vs 31.26 ± 9.33; p = 0.0037). On ROC analysis, at a cutoff of ≤ 32.93, O/E LHR had a specificity of 100% with a sensitivity of 72.22% in predicting mortality. Cases with liver herniation were not significantly different between survivors versus non-survivors. The overall survival rate was 37.93%, and the leading cause of death was severe persistent pulmonary hypertension. Conclusion: O/E LHR can predict mortality in neonates with antenatal diagnosis of CDH. The presence of pulmonary hypertension was the leading cause of death in these neonates.
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We have previously identified alveolar type II cell as the cell-of-origin of KrasG12D-induced lung adenocarcinoma using cell lineage-specific inducible Cre mouse models. Using gain-of-function and loss-of-function genetic models, we discovered that active Notch signaling and low Sox2 levels dictate the ability of type II cells to proliferate and progress into lung adenocarcinoma upon KrasG12D activation. Here, we examine the phenotype of type II cells after Kras activation and find evidence for proliferation of cells that coexpress type I and type II markers. Three-dimensional organoid culture and transplantation studies determine that these dual-positive cells are highly plastic and tumor initiating in vivo. RNA sequencing analysis reveals that these dual-positive cells are enriched in Ras/MAPK, EGFR, and Notch pathways. Furthermore, the proliferation of these cells requires active Notch signaling and is inhibited by genetic/chemical Sox2 upregulation. Our findings could provide new therapeutic strategies to target KRAS-activated lung adenocarcinomas. SIGNIFICANCE: Identification of progenitor like tumor-initiating cells in KRAS-mutant lung adenocarcinoma may allow development of novel targeted therapeutics.
Subject(s)
Adenocarcinoma of Lung , Adenocarcinoma , Lung Neoplasms , Mice , Animals , Proto-Oncogene Proteins p21(ras)/genetics , Adenocarcinoma/genetics , Lung Neoplasms/genetics , Cell Plasticity , Cell Proliferation/genetics , Adenocarcinoma of Lung/geneticsABSTRACT
Hepatitis C Virus (HCV) co-infection and its genotypic distribution in people living with Human Immunodeficiency Virus (HIV) show global inconsistency. Therefore, the present study aimed to investigate the prevalence and genotypic distribution patterns of HCV, along with viral load, in people living with HIV. This cross-sectional study was conducted at SRL Diagnostics Nepal, Pvt. Ltd. in 203 HIV-seropositive patients attending the Tribhuvan University Teaching Hospital (TUTH), Maharajgunj, Kathmandu, Nepal from October 2021 to May 2022. The viral load and HCV genotypes were estimated from RNA extracted from the blood sample (plasma) of PLHIV by using a standard Q-PCR protocol. HCV infection was considered as a core variable, whereas covariates used for this study were duration of HIV infection, age, sex, and ART regimen. Out of total 203 PLHIV, the estimated prevalence of HCV co-infection was 115 (56.6%). Male gender was a unique characteristic associated with a high prevalence of HCV co-infection compared to females. The HCV viral load among PLHIV ranged from 34 to 3,000,000 IU/mL. Among HCV co-infected PLHIV, 56 (48.69%) had a low level of HCV viral load. Interestingly, only 3 (2.6%) patients had an HCV viral load higher than 3,000,000 IU/mL. Diverse HCV genotypes were found in the population, including genotypes 1, 1a, 3a, 5a, and 6. However, genotype 3 was the most prevalent HCV variant among HCV-co-infected PLHIV, with a distribution of 36 (61.1%) and viral load ranging from 34 to 3000 IU/mL. HCV co-infection is frequent in the Nepalese population of people living with HIV, particularly due to HCV genotypic variant 3. The findings of this study could be useful for the management and clearance of the HCV co-infection in PLHIV, aiming to provide a good quality of life.
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Preschool teachers are uniquely positioned to impact children's physical activity levels, yet the relationship between teachers' and children's physical activity levels has not been widely investigated. The purpose of this study was to explore preschool teachers' physical activity levels, practices, and perceptions and how this relates to children's physical activity at preschool centers. This convergent mixed methods study included eight teachers and 20 children from four preschool classrooms. Accelerometers were used to measure their physical activity. Pearson correlations were used to explore the relationship between teachers' and children's physical activity levels. Direct observation was used to contextualize children's physical activity while at preschool. Teachers completed a semi-structured interview to explore their physical activity perceptions and practices. On average, teachers and children were physically active for 50.2 ± 9.3% and 29.5 ± 7.0% of their time at preschool, respectively. A significant positive correlation (p = 0.02; r = 0.98) was found between teachers' and children's percent of time at preschool per day in moderate-to-vigorous physical activity. Children mostly engaged in low-intensity activities (stationary play; light walking) during free play both indoors and outdoors and were mostly sedentary during teacher-initiated group times. All teachers stated they had a positive influence on children's physical activity. Teachers commonly reported pain or health conditions as a barrier to their physical activity. There was a positive relationship between teachers' and children's physical activity. More research is needed to confirm this relationship and to explore the impacts of high amounts of occupational physical activity on teachers' health. Supplementary Information: The online version contains supplementary material available at 10.1007/s10643-023-01486-8.
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Primary dysmenorrhea is a common menstrual disorder that significantly impacts women's quality of life, productivity, and healthcare utilization. In this randomized, double-blinded, placebo-controlled trial, sixty women with primary dysmenorrhea were randomly divided into two groups with thirty participants each, and were allocated either turmeric-boswellia-sesame formulation (treatment) or placebo. The participants were advised to take two softgels of 500 mg as a single dose of allocated study intervention (total dose 1000 mg) when their menstrual pain reached 5 or more on a numerical rating scale (NRS). Menstrual cramp pain intensity and relief were evaluated every 30 min post-dose until 6 h. Results indicated a promising role of turmeric-boswellia-sesame formulation for menstrual pain relief compared to the placebo. The mean total pain relief (TOTPAR) of the treatment group (18.9 ± 0.56) was found to be 12.6 times better than the placebo group (1.5 ± 0.39). The NRS analysis showed that there was a statistically significant difference in pain intensity between the treatment and placebo groups (p < 0.001) at every timepoint. Additionally, the sum of pain intensity difference at 6 h (SPID6) of the treatment group (34.32 ± 1.41) showed a significant difference (p < 0.0001) and was 20.19 times better when compared to placebo (1.7 ± 0.56). Based on the study results, the turmeric-boswellia-sesame formulation exhibited remarkable menstrual pain relief as compared to the placebo.
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Energy shortage has become a global issue in the twenty-firt century, as energy consumption grows at an alarming rate as the fossil fuel supply exhausts. Perovskite solar cells (PSCs) are a promising photovoltaic technology that has grown quickly in recent years. Its power conversion efficiency (PCE) is comparable to that of traditional silicon-based solar cells, and scale-up costs can be substantially reduced due to its utilization of solution-processable fabrication. Nevertheless, most PSCs research uses hazardous solvents, such as dimethylformamide (DMF) and chlorobenzene (CB), which are not suitable for large-scale ambient operations and industrial production. In this study, we have successfully deposited all of the layers of PSCs, except the top metal electrode, under ambient conditions using a slot-die coating process and nontoxic solvents. The fully slot-die coated PSCs exhibited PCEs of 13.86% and 13.54% in a single device (0.09 cm2) and mini-module (0.75 cm2), respectively.
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Biocomposites have gained tremendous advantages over synthetic composites due to their biocompatibility, sustainable degradation, and ability to easily combine with other substances. In the present study, we have prepared silk fibroin (SF) hydrogel, mulberry leaf extract (MLE), tasar pupal oil (TPO), and their composites, such as TPO-loaded SF hydrogel and MLE-loaded SF hydrogel, and characterized them by using a phase contrast microscope (PCM), scanning electron microscope (SEM) SEM- EDX, and Fourier transform infrared spectroscopy (FTIR). In addition, 1H-NMR was used for profiling of mulberry leaf extract and GC-MS was used to find tasar pupal oil composition. Further, the disc diffusion method evaluated their antimicrobial activities against S. aureus, E. coli, A. flavus, and A. brassicae. PCM, SEM, and FTIR results validated the conjugation of MLE and SF hydrogel composite; 1H-NMR confirmed the 41 metabolites in MLE, and GC-MS established the composition of tasar pupal oil. Since both composites, such as TPO-loaded SF hydrogel and MLE-loaded SF hydrogel, reduced the S. aureus and E. coli activities at all tested concentrations, the antibacterial results were unambiguous in their conclusion. S. aureus could only be inhibited by SF hydrogel at a high concentration (300 g/ml), despite suppressing E. coli growth at all tested concentrations. At 300 g/ml, MLE demonstrated antibacterial action against S. aureus. Furthermore, at a dosage of 300 g/ml, TPO inhibited both S. aureus and E. coli. Both mulberry leaf extract (at 200 and 300 g/ml) and the MLE-loaded SF hydrogel composite displayed antifungal activity against A. flavus at all tested concentrations (100, 200, and 300 g/ml).
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INTRODUCTION: Thyroid dysfunction is one of the most common endocrine disorders in people living with HIV (PLHIV). The abnormality in thyroid function has been linked with the adverse effects of prolonged antiretroviral therapy (ART) in PLHIV, but its prevalence remains obscure. The present study aimed to determine the prevalence of impaired thyroid function and its relationship to ART duration in Nepalese people living with HIV. METHODS: This cross-sectional clinical laboratory based study was conducted at SRL Diagnostics Nepal, Pvt. Ltd. from October 2021 to May 2022. Two hundred and three HIV-seropositive patients enrolled at Tribhuvan University Teaching Hospital (TUTH) in Kathmandu, Nepal were examined for their thyroid function test (TFT) by analyzing the serum T3, T4, and TSH concentrations using a fully automated COBAS e411 analyzer (Roche Diagnostics, USA) based on the electrochemiluminescence assay (ECLIA). RESULTS: Out of 203 PLHIV, 22 (10.83%) had a thyroid disorder, with subclinical hypothyroidism (n = 16, 72.73%) being the most common, followed by subclinical hyperthyroidism (n = 3, 13.63%). Thyroid dysfunction had no significant correlation with HIV/ART duration (p = 0.304) and sex (p = 0.419), whereas, the risk of thyroid dysfunction was induced with the rise in the age of the PLHIV (p = 0.002, Ï = 0.274). There were no significant differences in the mean serum T3, T4 and TSH values among different sexes and the HIV/ART duration, however a significant difference in the mean values of TSH (F (3, 199) = 3.231, p = 0.023) and T3 (F (3, 199) = 4.587, p = 0.004) among the different age-groups were shown. The mean T3 values also indicated a gradual decrease with increasing age. CONCLUSION: The study revealed subclinical hypothyroidism as the prevailing thyroid disorder associated with PLHIV. The risk of thyroid dysfunction in PLHIV was neither gender specific nor being attributed by the ART duration in Nepalese population; however, elderly PLHIV were highly susceptible to the risk of thyroid disorder.
Subject(s)
HIV Infections , Hypothyroidism , Humans , Aged , Cross-Sectional Studies , Nepal/epidemiology , Hypothyroidism/complications , Hypothyroidism/epidemiology , HIV Infections/drug therapy , Thyrotropin/therapeutic useABSTRACT
Diabetes is a major health challenge, and it is linked to a number of serious health issues, including cardiovascular disease (heart attack and stroke), diabetic nephropathy (kidney damage or failure), and birth defects. The detection of glucose has a direct and significant clinical importance in the management of diabetes. Herein, we demonstrate the application of in-situ synthesized Ti2C-TiO2 MXene nanocomposite for high throughput non-enzymatic electrochemical sensing of glucose. The nanocomposite was synthesized by controlled oxidation of Ti2C-MXene nanosheets using H2O2 at room temperature. The oxidation results in the opening up of Ti2C-MXene nanosheets and the formation of TiO2 nanocrystals on their surfaces as revealed in microscopic and spectroscopic analysis. Nanocomposite exhibited considerably high electrochemical response than parent Ti2C MXene, and hence utilized as a novel electrode material for enzyme-free sensitive and specific detection of glucose. Developed nanocomposite-based non-enzymatic glucose sensor (NEGS) displays a wide linearity range (0.1 µM-200 µM, R2 = 0.992), high sensitivity of 75.32 µA mM-1 cm-2, a low limit of detection (0.12 µM) and a rapid response time (~3s). NEGS has further shown a high level of repeatability and selectivity for glucose in serum spiked samples. The unveiled excellent sensing performance of NEGS is credited to synergistically improved electrochemical response of Ti2C MXene and TiO2 nanoparticles. All of these attributes highlight the potential of MXene nanocomposite as a next-generation NEGS for on the spot mass screening of diabetic patients.
Subject(s)
Diabetes Mellitus , Nanocomposites , Diabetes Mellitus/diagnosis , Glucose/analysis , Humans , Hydrogen Peroxide/analysis , Nanocomposites/chemistry , Titanium/chemistryABSTRACT
Background: Barriers to clinical trial enrollment have been the subject of extensive research; however, the rate of clinical trial participation has not improved significantly over time. Studies often emphasize patient-related barriers, but institutional and organizational barriers in the health care system may have a more substantial impact on clinical trial participation. Objective: To qualitatively identify perceived barriers to clinical trial participation based on perspectives from healthcare providers. Design: Qualitative research design with a phenomenological approach was used. A purposive sample of 18 healthcare providers participated in an in-depth focus group session. Participants were involved in cancer care and clinical research from a large hospital in the United States Midwest region. Data were transcribed, coded, and systematically analyzed through thematic content analysis. Results: The data revealed four levels of barriers to clinical trial enrollment, with emergent themes within each level: patient (beliefs or trust, distance to trial sites, health insurance coverage, language, and immigration status), provider (limited awareness of trial, time constraint, and non-cooperation from colleagues), clinical (eligibility criteria and clinical design), and institutional (policy and limited logistic support). Conclusion: Healthcare providers face complex, multifaceted, and interrelated barriers to clinical trial enrollment. To overcome these barriers, health care organizations need to commit more human and financial resources, break down boundaries for more efficient inter-departmental cooperation, develop more coordinated efforts in promoting trial awareness and participation, and remove unnecessary regulatory barriers.
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The purpose of this research was to investigate the adsorption of arsenic (As) from aqueous solutions using MPAC-500 and MPAC-600 (magnetic-activated carbons synthesized from the peel of Pisum sativum (pea) pyrolyzed at 500 °C and 600 °C temperatures, respectively). The potential of both biosorbents for As adsorption was determined in batch and column mode. The characterization of both biosorbents was performed by energy dispersive spectroscopy, scanning electron microscope, pHZPC, particle size distribution, X-ray diffraction, zeta potential and Fourier-transform infrared spectroscopy. It was found that the efficiency of MPAC-600 was better than MPAC-500 for the adsorption of As(III) and As(V) ions. The adsorption capacities of MPAC-500 and MPAC-600 in removing As(III) were 0.7297 mg/g and 1.3335 mg/g, respectively, while the values of Qmax for As(V) on MPAC-500 and MPAC-600 were 0.4930 mg/g and 0.9451 mg/g, respectively. The Langmuir isotherm model was found to be the best fit for adsorption of As(III) by MPAC-500 and MPAC-600, as well as adsorption of As(V) by MPAC-500. The Freundlich isotherm model, on the other hand, was optimal for As(V) removal with MPAC-600. With R2 values close to unity, the pseudo-second-order kinetics were best fitted to the adsorption process of both As species. The Thomas model was used to estimate the breakthrough curves. The effects of coexisting oxyanions and regeneration studies were also carried out to examine the influence of oxyanions on As adsorption and reusability of biosorbents.
Subject(s)
Arsenic , Water Pollutants, Chemical , Adsorption , Arsenic/chemistry , Biomass , Hydrogen-Ion Concentration , Kinetics , Magnetic Phenomena , Pisum sativum , Spectroscopy, Fourier Transform Infrared , Thermodynamics , Water , Water Pollutants, Chemical/chemistryABSTRACT
INTRODUCTION: Fostering physical activity, muscle strengthening and communication skills in diverse environments are vital to ensuring healthy infant development; however, promotion of these skills may be impacted by the COVID-19 pandemic. Therefore, the purpose of this study was to explore healthcare workers, parents and childcare providers' perceptions of the pandemic's influence on how they engage with infants to promote physical activity, muscle strength and communication. METHODS: 37 subjects (12 = parents; 12 = childcare providers, 13 = healthcare workers) participated in a semi-structured interview. Data were analyzed via an inductive content analysis. RESULTS: The majority of caregivers identified concerns related to: limitations in social interactions (especially masks impacting communication), lack of access to peer modeling, fewer opportunities for physical exploration, and a need for creative activities in diverse environments (e.g., home/childcare) for infant development during and after the pandemic. CONCLUSIONS: Caregivers are concerned about the role COVID-19 is having on infant development. Additional resources on how to promote infant physical activity, muscle strength and communication despite challenges associated with the COVID-19 pandemic are needed.
Subject(s)
COVID-19 , Pandemics , Child , Communication , Humans , Infant , Qualitative Research , SARS-CoV-2ABSTRACT
We describe the case of an antenatally diagnosed massive cardiac tumor in a fetus requiring cardiorespiratory support immediately following birth. We further discuss the successful management of this case and highlight the importance of a multidisciplinary team in managing such complicated cases. (Level of Difficulty: Advanced.).
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UDP-N-Acetyl glucosamine-2 epimerase/N-acetyl mannosamine kinase (GNE) catalyzes key enzymatic reactions in the biosynthesis of sialic acid. Mutation in GNE gene causes GNE myopathy (GNEM) characterized by adult-onset muscle weakness and degeneration. However, recent studies propose alternate roles of GNE in other cellular processes beside sialic acid biosynthesis, particularly interaction of GNE with α-actinin 1 and 2. Lack of appropriate model system limits drug and treatment options for GNEM as GNE knockout was found to be embryonically lethal. In the present study, we have generated L6 rat skeletal muscle myoblast cell-based model system carrying one single Gne allele where GNE gene is knocked out at exon-3 using AAV mediated SEPT homology recombination (SKM-GNEHz). The cell line was heterozygous for GNE gene with one wild type and one truncated allele as confirmed by sequencing. The phenotype showed reduced GNE epimerase activity with little reduction in sialic acid content. In addition, the heterozygous GNE knockout cells revealed altered cytoskeletal organization with disrupted actin filament. Further, we observed increased levels of RhoA leading to reduced cofilin activity and causing reduced F-actin polymerization. The disturbed signaling cascade resulted in reduced migration of SKM-GNEHz cells. Our study indicates possible role of GNE in regulating actin dynamics and cell migration of skeletal muscle cell. The skeletal muscle cell-based system offers great potential in understanding pathomechanism and target identification for GNEM.
Subject(s)
Actins/metabolism , Alleles , Muscle, Skeletal/metabolism , Myoblasts/metabolism , Animals , Mutation , Phenotype , RatsABSTRACT
Accumulation of misfolded proteins in endoplasmic reticulum (ER) generates a stress condition in the cell. The cell combats ER stress by activating unfolded protein response (UPR) and ERAD (ER stress-associated degradation) pathway. Failure to restore favorable folding environment results in cell dysfunction and apoptosis. Various neurodegenerative disorders are characterized by the accumulation of misfolded protein, protein aggregates, and ER stress. GNE myopathy (GNEM) is a neuromuscular disorder pathologically characterized by rimmed vacuole formation due to the accumulation of protein aggregates. More than 200 mutations in key sialic acid biosynthetic enzyme UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) have been identified worldwide in the muscle biopsies of GNE myopathy patients. However, the cellular and molecular pathomechanism leading to the disease ar poorly understood. In the present study, the phenomenon of ER stress has been elucidated in GNE mutant cells overexpressing GNE mutations of Indian origin. The effect of GNE mutations on activation of UPR signaling via inositol-requiring transmembrane kinase/endoribonuclease 1 (IRE-1), protein kinase RNA-like endoplasmic reticulum kinase (PERK), and activating transcription factor-6 (ATF6) were deciphered to understand the effect of GNE mutations on these proteins. GRP78 was upregulated with increased X-box-binding protein-1 (XBP-1) splicing and CCAAT/enhancer-binding protein (C/EBP) homologous protein (CHOP) upregulation leading to increased apoptosis of GNE mutant cells. Insulin-like growth factor 1 (IGF-1) ligand rescued the cells from apoptotic phenotype by supporting cell survival mechanism. Our study indicates a balance of cell death and survival that decides cell fate and offers potential therapeutic targets to combat ER stress in diseases associated with dysfunctional UPR pathway.