Cryo-EM images of phase-separated lipid bilayer vesicles analyzed with a machine-learning approach.

Lateral lipid heterogeneity (i.e., raft formation) in biomembranes plays a functional role in living cells. Three-component mixtures of low- and high-melting lipids plus cholesterol offer a simplified experimental model for raft domains in which a liquid-disordered (Ld) phase coexists with a liquid-ordered (Lo) phase. Using such models, we recently showed that cryogenic electron microscopy (cryo-EM) can detect phase separation in lipid vesicles based on differences in bilayer thickness. However, the considerable noise within cryo-EM data poses a significant challenge for accurately determining the membrane phase state at high spatial resolution. To this end, we have developed an image-processing pipeline that utilizes machine learning (ML) to predict the bilayer phase in projection images of lipid vesicles. Importantly, the ML method exploits differences in both the thickness and molecular density of Lo compared to Ld, which leads to improved phase identification. To assess accuracy, we used artificial images of phase-separated lipid vesicles generated from all-atom molecular dynamics simulations of Lo and Ld phases. Synthetic ground-truth data sets mimicking a series of compositions along a tieline of Ld + Lo coexistence were created and then analyzed with various ML models. For all tieline compositions, we find that the ML approach can correctly identify the bilayer phase with >90% accuracy, thus providing a means to isolate the intensity profiles of coexisting Ld and Lo phases, as well as accurately determine domain-size distributions, number of domains, and phase-area fractions. The method described here provides a framework for characterizing nanoscopic lateral heterogeneities in membranes and paves the way for a more detailed understanding of raft properties in biological contexts.

A Novel PINK1 p.F385S Loss-of-Function Mutation in an Indian Family with Parkinson's Disease.

BACKGROUND: Most Parkinson's disease (PD) loci have shown low prevalence in the Indian population, highlighting the need for further research. OBJECTIVE: The aim of this study was to characterize a novel phosphatase tensin homolog-induced serine/threonine kinase 1 (PINK1) mutation causing PD in an Indian family. METHODS: Exome sequencing of a well-characterized Indian family with PD. A novel PINK1 mutation was studied by in silico modeling using AlphaFold2, expression of mutant PINK1 in human cells depleted of functional endogenous PINK1, followed by quantitative image analysis and biochemical assessment. RESULTS: We identified a homozygous chr1:20648535-20648535 T>C on GRCh38 (p.F385S) mutation in exon 6 of PINK1, which was absent in 1029 genomes from India and in other known databases. PINK1 F385S lies within the highly conserved Deutsche Forschungsgemeinschaft (DFG) motif, destabilizes its active state, and impairs phosphorylation of ubiquitin at serine 65 and proper engagement of parkin upon mitochondrial depolarization. CONCLUSIONS: We characterized a novel nonconservative mutation in the DFG motif of PINK1, which causes loss of its ubiquitin kinase activity and inhibition of mitophagy. © 2024 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Loss of the auxiliary α2δ1 voltage-sensitive calcium channel subunit impairs bone formation and anabolic responses to mechanical loading.

Voltage-sensitive calcium channels (VSCCs) influence bone structure and function, including anabolic responses to mechanical loading. While the pore-forming (α1) subunit of VSCCs allows Ca2+ influx, auxiliary subunits regulate the biophysical properties of the pore. The α2δ1 subunit influences gating kinetics of the α1 pore and enables mechanically induced signaling in osteocytes; however, the skeletal function of α2δ1 in vivo remains unknown. In this work, we examined the skeletal consequences of deleting Cacna2d1, the gene encoding α2δ1. Dual-energy X-ray absorptiometry and microcomputed tomography imaging demonstrated that deletion of α2δ1 diminished bone mineral content and density in both male and female C57BL/6 mice. Structural differences manifested in both trabecular and cortical bone for males, while the absence of α2δ1 affected only cortical bone in female mice. Deletion of α2δ1 impaired skeletal mechanical properties in both sexes, as measured by three-point bending to failure. While no changes in osteoblast number or activity were found for either sex, male mice displayed a significant increase in osteoclast number, accompanied by increased eroded bone surface and upregulation of genes that regulate osteoclast differentiation. Deletion of α2δ1 also rendered the skeleton insensitive to exogenous mechanical loading in males. While previous work demonstrates that VSCCs are essential for anabolic responses to mechanical loading, the mechanism by which these channels sense and respond to force remained unclear. Our data demonstrate that the α2δ1 auxiliary VSCC subunit functions to maintain baseline bone mass and strength through regulation of osteoclast activity and also provides skeletal mechanotransduction in male mice. These data reveal a molecular player in our understanding of the mechanisms by which VSCCs influence skeletal adaptation.

Comparative assessment of facemask therapy with and without skeletal anchorage in growing Class III patients with unilateral cleft lip and palate (UCLP): A single-center, prospective randomized clinical trial.

OBJECTIVE: The purpose of this study was to evaluate the maxillary protraction effect of facemask therapy with and without skeletal anchorage in growing Class III patients with unilateral cleft lip and palate (UCLP). MATERIALS AND METHODS: Thirty patients (aged 9-13 years) with UCLP having a GOSLON score 3 were selected for this prospective clinical study. The patients were allocated into two groups using computer generated random number table. Group I (facemask therapy along with two I shaped miniplates, FM + MP) and Group II (facemask mask along with tooth-anchored appliance, FM). Skeletal and dental parameters were evaluated on pre- and post-treatment lateral cephalograms and pharyngeal airway on cone-beam computed tomography systems (CBCT) for assessment of the treatment changes. RESULTS: Both methods proved to be effective with statistically significant improvements in skeletal and dental parameters (p < .05). Skeletal parameters (e.g., SNA, convexity-point A, ANB) with the FM + MP group showed greater change compared to those with FM group (SNA, 2.56°; convexity-point A, 1.22°; ANB, 0.35°). Significant proclination of maxillary incisors was observed in the FM group as compared to FM + MP group (U1 to NA, 5.4°; 3.37 mm). A statistically significant increase in pharyngeal airway volume was noted in both groups (p < .05). CONCLUSION: While both therapies are effective in protracting the maxilla in growing patients with UCLP, the FM + MP allows for a greater skeletal correction, minimizing the dental side effects seen with FM therapy alone. Thus, FM + MP appears to be a promising adjunct in reducing the severity of Class III skeletal correction needed in patients with cleft lip and palate (CLP).

GST polymorphism as a predictive biomarker for modulating the susceptibility to chronic obstructive pulmonary disease: A North Indian study.

Chronic obstructive pulmonary disease (COPD) is commonly characterized by shortness of breath, coughing or expectoration. Smoking is the leading cause of COPD development, but only a small percentage of smokers develop symptoms, implying a genetic component. Glutathione S-transferase enzymes are responsible for detoxifying cigarette smoke components. The role of glutathione S-transferase T1 (GSTT1) and glutathione S-transferase M1 (GSTM1) gene polymorphism was assessed with COPD susceptibility and associated clinical parameters in the North Indian population. This was a cross-sectional study involving 200 COPD patients and 200 healthy individuals, with peripheral blood sampling and adequate questionnaires. Multiplex PCR was used for genotyping GSTT1 and GSTM1 gene polymorphism. Logistic regression was used to calculate the odds ratio and 95% confidence intervals to assess the COPD risk and GST polymorphisms. The GSTT1 gene deletion rate was higher in COPD cases (34.5%) than in healthy individuals (20.5%). A statistical relationship between the GSTT1(-) null genotype and COPD risk was observed (odds ratio = 2.04, 95% CI = 1.30-3.20, P = 0.0019). After adjusting for covariates like age, sex and smoking status, a significant association was found for GSTT1(-) null genotype and COPD risk (adjusted odds ratio = 2.90, 95% CI = 1.43-5.87, P = 0.003). The GSTT1(-) genotype was also significantly correlated with clinical parameters for COPD risk. Another primary observation was that females with the GSTT1(-) null genotype were more vulnerable to COPD than males with the same gene deletion. The GSTT1(-) null genotype strongly correlates with COPD development, while no association was observed in the GSTM1(-) null genotype in the North Indian population.

Impact of Vaping on Lungs: An Indian Prospect.

Electronic cigarettes, or vape pens, have gained popularity among young people due to their attractive appearance, but they also have numerous side effects. These devices come in various shapes, sizes, and costs, with many brands and various flavors to choose from. As of now, there are around 2,807 people hospitalized in the United States due to vaping. Electronic cigarettes are illegal in many countries, with laws varying from country to country. The health service has been criticized for the implementation of the ban, with states implementing month-long drives and submitting reports. The Indian Police have also conducted raids and arrested five individuals under the 2019 ban on Electronic Cigarettes (Production, Manufacture, Trade, Transport, Sale, Distribution, Storage, and Advertisement) Act. The history of vapes can be traced back to the invention of the first electric vaporizer in 1927 by Joseph Robinson. Other pioneers like Herbert Gilbert and Jed Rose developed nicotine fixes using refined smoke. The vape was made in 2003 by Chinese smoker Hon Lik, who created the device as a better option to traditional smoking.

A Rare Case of Visual Hallucinations Associated With Hyponatremia.

Visual hallucinations are rare occurrences in patients presenting with hyponatremia. When they occur, the patient experiences these hallucinations with their eyes closed (vs. opened) and is insightful about the false perception. We present the case of a 64-year-old male diagnosed with hyponatremia caused by a gastrointestinal illness, which led to visual hallucinations. The patient was treated with electrolyte infusion, and the hallucinations were resolved. Detailed history-taking is significant when dealing with such cases as hallucinations to differentiate it from other causes. Hallucination caused by hyponatremia can resolve with prompt correction of sodium, and the patients can be reassured.

Determinants and Stratification of Microvascular Complications of Type 2 Diabetes Mellitus.

Background Diabetes mellitus (DM) is a prevalent metabolic disorder characterized by high blood sugar levels. It is classified into type 1 (T1DM) and type 2 (T2DM), which have different mechanisms and complications. The global prevalence of diabetes, particularly T2DM, has increased significantly in recent decades, leading to a need for standardized data collection of macrovascular and microvascular complications to track disease progression and guide treatment options. This study aims to assess and correlate the prevalence and severity of microvascular complications in patients with T2DM. Methodology This observational, cross-sectional study was conducted at Poonam Multispeciality Hospital in Ahmedabad, India. A total of 4,123 diabetic patients admitted to the hospital were included. Information on sociodemographics and medical history was collected using standardized forms. Fundus photography and fluorescein angiography were performed to assess diabetic retinopathy, and estimated glomerular filtration rate and albumin-to-creatinine ratio were measured to evaluate renal function. Neurological examinations were conducted to score diabetic neuropathy. Chi-square tests were used to determine associations between medical history with diabetic retinopathy and nephropathy, and t-tests were used to compare diabetic neuropathy scores. Kendall's Tau correlation was used to determine correlations between diabetic retinopathy and nephropathy. P-values <0.05 were considered statistically significant. Results The overall prevalence of diabetic retinopathy, nephropathy, and neuropathy was 37.5%. Of the patients included, 47.9% had diabetic nephropathy and 28.9% had diabetic neuropathy. A significant association was observed between the severity of diabetic retinopathy and age, body mass index, duration of diabetes, and hemoglobin A1c (HbA1C) levels. Similarly, significant associations were found between these factors and the severity of diabetic nephropathy. Unpaired t-tests revealed significant differences in diabetic neuropathy examination scores based on the duration of diabetes and Hba1C levels. Moreover, correlation analysis indicated a low, positive correlation between diabetic retinopathy and diabetic nephropathy. Conclusions This study provides insights into the prevalence, severity, and associations of microvascular complications in patients with T2DM, contributing to the understanding and management of these conditions. Additionally, the research revealed a direct association between diabetic retinopathy and different stages of chronic kidney disease determined by the Kidney Disease Improving Global Outcome guidelines.

Association of IL4RA and IL-8 polymorphism in predicting susceptibility towards tuberculosis.

Mycobacterium tuberculosis is an infectious bacterial disease frequently affecting the lungs. With two fatalities from tuberculosis (TB) occurring every three minutes, India has the highest disease burden. The aetiology of tuberculosis has been linked to IL-8 and IL-4RA. Thus, the impact of the IL4RAQ576R and IL8 gene polymorphism on TB susceptibility was assessed. 301 healthy and 301 TB patients participated in a cross-sectional study. PCR RFLP was performed to identify the genotype of the IL4RAQ576R and IL-8 +781C/T gene polymorphism. The odds ratio and 95% confidence intervals were calculated using logistic regression to evaluate the risk of TB with IL4RAQ576R and IL-8 +781C/T polymorphism. A significant association was found between IL-4RA (p=0.04) and IL-8 +781 C/T (p= 0.03) in tuberculosis. Further, when clinical symptoms were compared with both polymorphisms, two of them, i.e., cough in IL-4RA576R (p=0.04) and breathlessness (p=0.01) in IL-8 +781C/T, showed a significant association. Moreover, different combinations of the SNPS were made, and the 3 risk allele shows a significant protective role (p=0.02). There is considerable evidence which shows that M. tuberculosis causes TB, an infectious disease that is genetically predisposed. The results of our study also showed that IL-4 RA Q576R and IL-8 +781 C/T played a significant protective function against tuberculosis, confirming the claim mentioned earlier. However, only the cough in IL-4RA576R and the dyspnea in IL-8 +781C/T exhibited a significant co-relation in TB patients when symptoms were examined. Additionally, the combined effects of the two SNPs were investigated, and it was discovered that the 3-risk allele has a strong association with tuberculosis. Therefore, the polymorphisms mentioned earlier, which may also be influenced by ethnicity, may significantly impact the chance of developing tuberculosis.

Comparison of the Efficacy of Light's Criteria With Serum-Effusion Albumin Gradient and Pleural Effusion Glucose.

Introduction While Light's criteria exhibit high sensitivity (98%) in detecting exudative pleural effusions, the capacity to rule out transudates is relatively limited. A previous study showed that approximately one-fifth of patients with congestive cardiac failure on diuretics also met the criteria for exudate. This study compares the diagnostic value of Light's criteria, the serum-effusion albumin gradient (SEAG) method, and pleural effusion glucose levels for accurately categorizing pleural effusion as transudate or exudate. Methodology We conducted this cross-sectional observational study in a tertiary care hospital in Ahmedabad, India. Two hundred patients with pleural effusion undergoing thoracentesis were included. Laboratory parameters measured in pleural fluid analysis included pleural fluid protein, pleural fluid lactate dehydrogenase (LDH), pleural fluid albumin, and pleural fluid glucose. Serum protein, serum LDH, and serum albumin were also collected. Mean values and standard deviations (SDs) were calculated for analysis. Results A significant difference was observed in the mean value of exudative and transudative effusions for each parameter (pleural fluid protein/serum fluid protein ratio, pleural fluid LDH/serum fluid LDH ratio, pleural fluid LDH, SEAG, and pleural fluid glucose) (P < 0.001). Light's criteria demonstrated the highest efficacy in diagnosing exudates (accuracy = 97.50%), while SEAG demonstrated the highest efficacy in diagnosing transudates (accuracy = 97.50%). Conclusion SEAG is an effective alternative diagnostic tool for identifying transudates misclassified by Light's criteria. Its use can contribute to prompt diagnosis and timely treatment of patients with pleural effusion, improving patient outcomes.

Combined Radiological and Endoscopic Evaluation of Sino Nasal Anatomical Variations in Patients of Chronic Rhinosinusitis: A North Indian Study.

To determine the prevalence of anatomical variations of nasal cavity and paranasal sinuses of patients with chronic rhinosinusitis (CRS) on CT scan imaging. To correlate the Anatomical variations with the extent of CRS. 100 patients attending the ENT outpatient department with clinically diagnosed CRS were selected for study based on inclusion and exclusion criteria. Patients were subjected to CT scan and diagnostic nasal endoscopy. The correlation of anatomical variation with severity of CRS based on radiological score and endoscopic score was observed. Deviated nasal septum was the most common anatomical variation observed in 71% cases. Followed by Agger nasi (68%), concha bullosa (55%), Onodi cell (25%), Haller cell (14%), frontal sinus hypoplasia (2%) and uncinate bulla (1%) respectively. Statistically significant relationship of radiological score with left side Bullous Concha Bullosa and highly statistically significant relationship with Haller Cell was observed. While statistically significant relationship between Deviated Nasal Septum on left side with endoscopic score was also observed. Correlation of anatomical variation with CRS concludes on the note that some variations cause impaired sinus drainage and ventilation leading to recurrent sinusitis. Also, incidence of these variations was comparable to other studies done in asymptomatic population therefore, simply detection of a solitary anatomical variant itself does not determine predisposition to disease or the pathogenesis of the CRS and that we should have a critical look out for these anatomical variations from point of view of surgical management.

An assessment of masticatory efficiency and occlusal load distribution in adolescent patients undergoing orthodontic treatment with functional jaw orthopedics: A prospective cohort study.

INTRODUCTION: Functional jaw orthopaedics, produces a radical change in the occlusal scheme and the masticatory apparatus, particularly in patients with Class II malocclusion. It remains to be seen how the changes brought about by a functional appliance alter the masticatory ability of a growing child, who needs the necessary nutrition to properly grow the craniofacial region. MATERIALS AND METHODS: Pretreatment and Post-treatment values of masticatory efficiency and the distribution of the occlusal load at centric occlusion were evaluated and compared for 20 patients with Class II division 1 malocclusion undergoing functional jaw orthopaedics. RESULTS: Significant increase in the masticatory efficiency was seen during and after treatment (p < 0.5) There was an increase in the anterior distribution of occlusal load associated with a concomitant decrease in the posterior region at centric occlusion. CONCLUSION: Improvement in the masticatory efficiency was observed after treatment of a retrognathic mandible with functional jaw orthopaedics in the adolescent participants with Class II malocclusion. This highlights the importance of treatment with functional jaw orthopaedics, which apart from providing esthetic and functional improvement also improves the ability of a growing child to extract proper nutrition from his/her diet.

Neurofibrolipoma of Hypopharynx: A Rare Entity.

A Large pedunculated neurofibrolipoma involving the hypopharynx is a very rare entity. Patient with hypopharyngeal polyp usually presents with progressive dyaphagia, sensation of persistent lump in throat and difficulty in breathing. Regurgitation of the mass into the airway can led to fatal asphyxiation on several occasions and may rarely cause death also if not evaluated and treated timely. Malignant degeneration of these large polyps occurs infrequently. We encountered a case with similar complaints in our outpatient department. After complete clinical examination, radiological investigations and anaesthetic evaluation; patient was taken for elective tracheostomy first and subsequently followed by successful transoral endoscopic excision of hypopharyngeal polypoidal mass. The histopathological examination of mass revealed it to be a neurofibrolipoma.

A Three-Dimensional Scale for the Qualitative and Quantitative Assessments of Secondary Alveolar Bone Grafting (SABG) in Unilateral Cleft Lip and Palate Patients Using Cone-Beam Computed Tomography (CBCT).

Background The objective of our study was to derive an objective assessment scale for three-dimensional (3D) qualitative and quantitative evaluation of secondary alveolar bone grafting (SABG) using cone-bone computed tomography (CBCT) in patients with unilateral cleft lip and palate (UCLP). Methods CBCT scans for pre- and 3-month post-SABG were reviewed for bone volume, height, width, and density of the bony bridge formed in the cleft defect in 20 patients with UCLP. Basic descriptive and principal component analysis was used to extract the various sub-components of the scale. Spearman's correlation was used to check the validity of the scale, and intra-class coefficient (ICC) and Cronbach's α were calculated to establish the reliability and retest applicability of the scale. Results Each CBCT scan was assessed in five areas: cementoenamel junction (CEJ), root apex, root midpoint, 3 and 6 mm below CEJ, and tabulated in percentiles of 20, 25, 40, 50, 60, and 75 for all the parameters (bone volume, density, and width). These scores were validated when correlated to the scale given by Kamperos et al. Cronbach's α for the domains demonstrated acceptable to excellent internal consistency. The ICC showed good test-retest reliability having a range of scores from 0.89 to 0.94. Conclusion The proposed scale for the 3D assessment of SABG in patients with UCLP provides gradation for the objective assessment of the bony bridge. This gradation enables the qualitative and quantitative assessments of the bony bridge, thus allowing each clinician to judge SABG more conclusively.

Perforated Marginal Ulcer.

Marginal ulcers are a late complication of gastric bypass surgery. A marginal ulcer is a term for ulcers that develop at the margins of a gastrojejunostomy, primarily on the jejunal side. A perforated ulcer involves the entire thickness of an organ, creating an opening on both surfaces. We will present an intriguing case of a 59-year-old Caucasian female who arrived at the emergency department with diffused chest and abdominal pain that began in her left shoulder and went down to the right lower quadrant area. The patient was in visible pain with restlessness, and her abdomen was moderately distended. The computed tomography (CT) showed possible perforation in the gastric bypass surgery area, but the results were inconclusive. The patient had laparoscopic cholecystectomy ten days prior, and the pain began right after surgery. The patient underwent an open abdominal exploratory surgery, with the closure of the perforated marginal ulcer. The fact that the patient had undergone another surgery and had pain immediately afterward also obscured the diagnosis. This case shows the rare presentation of the patientäs diverse signs and symptoms and inconclusive reports that led to the open abdominal exploratory surgery that finally confirmed the diagnosis. This case highlights the importance of a thorough past medical history, including surgical history. The past surgical history led the team to zone in on the gastric bypass area, leading to an accurate differential diagnosis.

The Association between Nursing Skill Mix and Patient Outcomes in a Mental Health Setting: An Observational Feasibility Study.

Higher levels of educational preparation for nurses are associated with lower mortality rates in both medical and surgical wards. In mental health inpatient wards, few studies have examined whether specialist mental health nurse training has any impact on patient outcomes. The aim of this retrospective observational study was to establish the feasibility of extracting and linking nurse education and inpatient outcome data from hospital administrative sources to inform the design of future mental health nursing skill mix studies. Study participants were people experiencing mental ill-health and admitted to psychiatric inpatient care for at least 24 h. The exposure was the ratio of mental health nurses to comprehensive nurses for each patient for each day of their admission. The outcome was readmission for psychiatric inpatient care within 12 months of discharge from the index admission. Confounders were patient demographic (age, gender) and clinical characteristics (diagnosis, legal status, community follow-up). Forty-four patients included in the study were inpatients for a total of 595 days. The median hospital stay was 12 days (IQR = 7-17). In total, 11 (25%) patients were readmitted. In the readmitted and not readmitted groups, the median skill mix ratio was 5 (IQR = 5-7) and 5 (1-6), respectively. It was feasible to extract and code patient and nurse data from hospital databases and link them together. However, a substantial amount of manual post hoc recoding was required to enable us to calculate the exposure (mental health to comprehensive nurse ratio) in a precise way. It may be realistic to automate our methodology in an appropriately powered mental health nursing skill mix study. Australian and New Zealand clinical trial registry: ACTRN12619001337167p.

Factors contributing to nurse resignation during COVID-19: A qualitative descriptive study.

AIMS: To explore and describe registered nurses' perceptions and experiences of work well-being extending from what inspired them to join the healthcare organization, what created a great day at work for them, through to what may have supported them to stay. DESIGN: Qualitative descriptive study. METHODS: Thirty-nine Australian nurses who resigned in 2021 from two metropolitan healthcare organizations in Victoria were interviewed in 2022, each for 30-60 min. The semi-structured interview transcripts were transcribed verbatim and analysed inductively and thematically. RESULTS: Four themes were constructed for each of the key research questions. Inspiration to join the organizations transpired through organizational reputation, recruitment experiences, right position and right time, fit and feel. A great day at work was created through relationships with colleagues, experiences with managers, adequate resourcing and delivering quality care. Factors contributing to nurses resigning included COVID-19, uncertainty of role, workload and rostering, and finally, not feeling supported, respected and valued. Factors that may have supported the nurses to stay included flexible work patterns and opportunities, improved workplace relationships, workload management and support, and supportive systems and environments. Cutting across these themes were five threads: (1) relationships, (2) communication, (3) a desire to learn and develop, (4) work-life balance and (5) providing quality patient care. CONCLUSIONS: Novel ways of working and supporting individuals, teams and organizations are needed to maintain and sustain nurses. The nurses' inspiration, what created a great day at work, and support needed to stay highlighted the importance of workplace initiatives to build nursing career pathways, provide equitable opportunities for professional development, workload and roster flexibility and implement professional relationship-enhancing actions to foster authentic civility. IMPACT: This study contributes an in-depth exploration of the perceptions and experiences of nurses who resigned from two healthcare organizations and provides a description of (1) what inspired these nurses to initially join the organization, (2) what they perceived created a great day at work for them, (3) the factors contributing to their resignation and (4) what may have supported them to stay. The reasons nurses resign from an organization were identified as complex and multi-factorial, with opportunities for promoting nursing career pathways, addressing equity in opportunities and implementing professional relationship-enhancing actions. These contributions add both context and opportunity to strengthen organizational initiatives to attract, sustain and retain nurses.

Visualizing lipid membrane structure with cryo-EM: past, present, and future.

The development of electron cryomicroscopy (cryo-EM) has evolved immensely in the last several decades and is now well-established in the analysis of protein structure both in isolation and in their cellular context. This review focuses on the history and application of cryo-EM to the analysis of membrane architecture. Parallels between the levels of organization of protein structure are useful in organizing the discussion of the unique parameters that influence membrane structure and function. Importantly, the timescales of lipid motion in bilayers with respect to the timescales of sample vitrification is discussed and reveals what types of membrane structure can be reliably extracted in cryo-EM images of vitrified samples. Appreciating these limitations, a review of the application of cryo-EM to examine the lateral organization of ordered and disordered domains in reconstituted and biologically derived membranes is provided. Finally, a brief outlook for further development and application of cryo-EM to the analysis of membrane architecture is provided.

Miro1 R272Q disrupts mitochondrial calcium handling and neurotransmitter uptake in dopaminergic neurons.

The Rho GTPase Miro1, located at the mitochondrial outer membrane is known to properly distribute mitochondria to synapses, aid calcium buffering and initiate PINK1-Parkin mediated mitophagy. Several heterozygous RHOT1/Miro1 variants were identified in sporadic Parkinson's disease patients. Miro1 R272Q is located within a calcium binding domain, but the functional outcome of this point mutation and its contribution to the development of disease are unclear. To address this, we introduced a heterozygous RHOT1/Miro1 R272Q point mutation in healthy induced pluripotent stem cells. In dopaminergic neurons, Miro1 R272Q does not affect Miro1 protein levels, CCCP-induced mitophagy, nor mitochondrial movement yet causes the fragmentation of mitochondria with reduction of cristae and ATP5A. Inhibition of the mitochondrial calcium uniporter phenocopied Miro1 R272Q cytosolic calcium response to Thapsigargin in active neurons, a similar effect was observed during the calcium buffering phase in Miro1 knockdown neuroblastoma cells. Altered mitochondrial calcium regulation is associated with reduced mitochondrial respiration and reduced catecholamine neurotransmitter uptake. Synaptic changes are not coupled to dopamine distribution or dopamine transporters but are linked to Miro1 R272Q-related calcium handling via the mitochondria concomitant with defective dopamine regulation at the mitochondrial surface by monoamine oxidase. We conclude that the Miro1 R272Q heterozygous point mutation dampens mitochondrial-calcium regulation and mitochondrial capacity via events at the outer membrane that are sufficient to disrupt dopaminergic function.

END TB initiative - involving youth - challenges and solutions.

WHO has given a special call to involve youth in END TB initiative. Involvement of youth, especially in the medical fraternity, in the TB control activities can ensure we reach our goal of ending TB in India by 2025. This study was planned to recruit and monitor young volunteers from medical colleges of Andhra Pradesh. 183 volunteers were recruited from 15 medical colleges and were tasked to spread awareness among general public. Work of these volunteers was monitored via Whatsapp group chats and videoconferencing. 4 follow-up meetings were held via videoconferencing and various hurdles and challenges were discussed. Major challenges faced included lack of physical meetings due to COVID-19 pandemic, which adversely affected their motivation and monitoring activities. Increased involvement from medical colleges, regular telecommunication and meetings can improve the work and motivation of these volunteers.