Genome mining yields putative disease-associated ROMK variants with distinct defects.

Bartter syndrome is a group of rare genetic disorders that compromise kidney function by impairing electrolyte reabsorption. Left untreated, the resulting hyponatremia, hypokalemia, and dehydration can be fatal, and there is currently no cure. Bartter syndrome type II specifically arises from mutations in KCNJ1, which encodes the renal outer medullary potassium channel, ROMK. Over 40 Bartter syndrome-associated mutations in KCNJ1 have been identified, yet their molecular defects are mostly uncharacterized. Nevertheless, a subset of disease-linked mutations compromise ROMK folding in the endoplasmic reticulum (ER), which in turn results in premature degradation via the ER associated degradation (ERAD) pathway. To identify uncharacterized human variants that might similarly lead to premature degradation and thus disease, we mined three genomic databases. First, phenotypic data in the UK Biobank were analyzed using a recently developed computational platform to identify individuals carrying KCNJ1 variants with clinical features consistent with Bartter syndrome type II. In parallel, we examined genomic data in both the NIH TOPMed and ClinVar databases with the aid of Rhapsody, a verified computational algorithm that predicts mutation pathogenicity and disease severity. Subsequent phenotypic studies using a yeast screen to assess ROMK function-and analyses of ROMK biogenesis in yeast and human cells-identified four previously uncharacterized mutations. Among these, one mutation uncovered from the two parallel approaches (G228E) destabilized ROMK and targeted it for ERAD, resulting in reduced cell surface expression. Another mutation (T300R) was ERAD-resistant, but defects in channel activity were apparent based on two-electrode voltage clamp measurements in X. laevis oocytes. Together, our results outline a new computational and experimental pipeline that can be applied to identify disease-associated alleles linked to a range of other potassium channels, and further our understanding of the ROMK structure-function relationship that may aid future therapeutic strategies to advance precision medicine.

In vivo quasi-elastic light scattering detects molecular changes in the lenses of adolescents with Down syndrome.

Down syndrome (DS) is the most common chromosomal disorder in humans. DS is associated with increased prevalence of several ocular sequelae, including characteristic blue-dot cerulean cataract. DS is accompanied by age-dependent accumulation of Alzheimer's disease (AD) amyloid-ß (Aß) peptides and amyloid pathology in the brain and comorbid early-onset Aß amyloidopathy and colocalizing cataracts in the lens. Quasi-elastic light scattering (QLS) is an established optical technique that noninvasively measures changes in protein size distributions in the human lens in vivo. In this cross-sectional study, lenticular QLS correlation time was decreased in adolescent subjects with DS compared to age-matched control subjects. Clinical QLS was consistent with alterations in relative particle hydrodynamic radius in lenses of adolescents with DS. These correlative results suggest that noninvasive QLS can be used to evaluate molecular changes in the lenses of individuals with DS.

Rapid single cell evaluation of human disease and disorder targets using REVEAL: SingleCell™.

BACKGROUND: Single-cell (sc) sequencing performs unbiased profiling of individual cells and enables evaluation of less prevalent cellular populations, often missed using bulk sequencing. However, the scale and the complexity of the sc datasets poses a great challenge in its utility and this problem is further exacerbated when working with larger datasets typically generated by consortium efforts. As the scale of single cell datasets continues to increase exponentially, there is an unmet technological need to develop database platforms that can evaluate key biological hypotheses by querying extensive single-cell datasets. Large single-cell datasets like Human Cell Atlas and COVID-19 cell atlas (collection of annotated sc datasets from various human organs) are excellent resources for profiling target genes involved in human diseases and disorders ranging from oncology, auto-immunity, as well as infectious diseases like COVID-19 caused by SARS-CoV-2 virus. SARS-CoV-2 infections have led to a worldwide pandemic with massive loss of lives, infections exceeding 7 million cases. The virus uses ACE2 and TMPRSS2 as key viral entry associated proteins expressed in human cells for infections. Evaluating the expression profile of key genes in large single-cell datasets can facilitate testing for diagnostics, therapeutics, and vaccine targets, as the world struggles to cope with the on-going spread of COVID-19 infections. MAIN BODY: In this manuscript we describe REVEAL: SingleCell, which enables storage, retrieval, and rapid query of single-cell datasets inclusive of millions of cells. The array native database described here enables selecting and analyzing cells across multiple studies. Cells can be selected using individual metadata tags, more complex hierarchical ontology filtering, and gene expression threshold ranges, including co-expression of multiple genes. The tags on selected cells can be further evaluated for testing biological hypotheses. One such example includes identifying the most prevalent cell type annotation tag on returned cells. We used REVEAL: SingleCell to evaluate the expression of key SARS-CoV-2 entry associated genes, and queried the current database (2.2 Million cells, 32 projects) to obtain the results in < 60 s. We highlighted cells expressing COVID-19 associated genes are expressed on multiple tissue types, thus in part explains the multi-organ involvement in infected patients observed worldwide during the on-going COVID-19 pandemic. CONCLUSION: In this paper, we introduce the REVEAL: SingleCell database that addresses immediate needs for SARS-CoV-2 research and has the potential to be used more broadly for many precision medicine applications. We used the REVEAL: SingleCell database as a reference to ask questions relevant to drug development and precision medicine regarding cell type and co-expression for genes that encode proteins necessary for SARS-CoV-2 to enter and reproduce in cells.

Sex and race differences of cerebrospinal fluid metabolites in healthy individuals.

INTRODUCTION: Analyses of cerebrospinal fluid (CSF) metabolites in large, healthy samples have been limited and potential demographic moderators of brain metabolism are largely unknown. OBJECTIVE: Our objective in this study was to examine sex and race differences in 33 CSF metabolites within a sample of 129 healthy individuals (37 African American women, 29 white women, 38 African American men, and 25 white men). METHODS: CSF metabolites were measured with a targeted electrochemistry-based metabolomics platform. Sex and race differences were quantified with both univariate and multivariate analyses. Type I error was controlled for by using a Bonferroni adjustment (0.05/33 = .0015). RESULTS: Multivariate Canonical Variate Analysis (CVA) of the 33 metabolites showed correct classification of sex at an average rate of 80.6% and correct classification of race at an average rate of 88.4%. Univariate analyses revealed that men had significantly higher concentrations of cysteine (p < 0.0001), uric acid (p < 0.0001), and N-acetylserotonin (p = 0.049), while women had significantly higher concentrations of 5-hydroxyindoleacetic acid (5-HIAA) (p = 0.001). African American participants had significantly higher concentrations of 3-hydroxykynurenine (p = 0.018), while white participants had significantly higher concentrations of kynurenine (p < 0.0001), indoleacetic acid (p < 0.0001), xanthine (p = 0.001), alpha-tocopherol (p = 0.007), cysteine (p = 0.029), melatonin (p = 0.036), and 7-methylxanthine (p = 0.037). After the Bonferroni adjustment, the effects for cysteine, uric acid, and 5-HIAA were still significant from the analysis of sex differences and kynurenine and indoleacetic acid were still significant from the analysis of race differences. CONCLUSION: Several of the metabolites assayed in this study have been associated with mental health disorders and neurological diseases. Our data provide some novel information regarding normal variations by sex and race in CSF metabolite levels within the tryptophan, tyrosine and purine pathways, which may help to enhance our understanding of mechanisms underlying sex and race differences and potentially prove useful in the future treatment of disease.

Diagnostic uncertainties, ethical tensions, and accounts of role responsibilities in genetic counseling communication.

Diagnostic uncertainties are intricately associated with genomic testing-especially concerning new technologies such as exome sequencing-with test results being either inconclusive or generating secondary findings or showing variants of uncertain significance. In the process of genetic counseling, diagnostic uncertainties have to be managed even when test results for an individual client are either positive or negative because of differential implications for family members. Previous studies have investigated diagnostic uncertainties in relation to clients wanting to know or not know the test results; here, we extend this line of research by addressing how genetic counselors and clients account for the management of diagnostic uncertainties vis-à-vis the attendant ethical tensions in the complex communicative environment in the clinic setting. Our dataset from the Norwegian context is longitudinal, consisting of ten audio-recorded pre-test genetic counseling sessions. It involves one extended family with a high burden of colorectal cancer. Through theme-oriented discourse analysis, we demonstrate how diagnostic uncertainties give rise to tensions concerning risks and benefits of knowing in both professional and familial spheres, which then map onto accounts of various role responsibilities. For instance, in looking for certainty via advanced genomic testing to reduce diagnostic uncertainty for clients, genetic counselors are confronted with tensions regarding what can be communicated and made known because of their role responsibilities toward what may be regarded as scientific others and clinical others. Likewise, clients are faced with tensions concerning wanting to know/not know, which invokes various familial others and may align or not align with genetic counselors' preferences, especially relating to management of diagnostic uncertainties and secondary findings.

Genetic counseling/consultation in South-East Asia: a report from the workshop at the 10th Asia pacific conference on human genetics.

This paper reports on the workshop 'Genetic Counseling/Consultations in South-East Asia' at the 10(th) Asia Pacific Conference on Human Genetics in Kuala Lumpur, Malaysia, in December 2012. The workshop brought together professionals and language/communication scholars from South-East Asia, and the UK. The workshop aimed at addressing culture- and context-specific genetic counseling/consultation practices in South-East Asia. As a way of contextualizing genetic counseling/consultation in South-East Asia, we first offer an overview of communication-oriented research generally, drawing attention to consultation and counseling as part of a communicative continuum with distinctive interactional features. We then provide examples of genetic counseling/consultation research in Hong Kong. As other countries in South-East Asia have not yet embarked on communication-oriented empirical research, we report on the current practices of genetic counseling/consultation in these countries in order to identify similarities and differences as well as key obstacles that could be addressed through future research. Three issues emerged as 'problematic': language, religion and culture. We suggest that communication-oriented research can provide a starting point for evidence-based reflections on how to incorporate a counseling mentality in genetic consultation. To conclude, we discuss the need for creating a platform for targeted training of genetic counselors based on communication-oriented research findings.

Genome mining yields new disease-associated ROMK variants with distinct defects.

Bartter syndrome is a group of rare genetic disorders that compromise kidney function by impairing electrolyte reabsorption. Left untreated, the resulting hyponatremia, hypokalemia, and dehydration can be fatal. Although there is no cure for this disease, specific genes that lead to different Bartter syndrome subtypes have been identified. Bartter syndrome type II specifically arises from mutations in the KCNJ1 gene, which encodes the renal outer medullary potassium channel, ROMK. To date, over 40 Bartter syndrome-associated mutations in KCNJ1 have been identified. Yet, their molecular defects are mostly uncharacterized. Nevertheless, a subset of disease-linked mutations compromise ROMK folding in the endoplasmic reticulum (ER), which in turn results in premature degradation via the ER associated degradation (ERAD) pathway. To identify uncharacterized human variants that might similarly lead to premature degradation and thus disease, we mined three genomic databases. First, phenotypic data in the UK Biobank were analyzed using a recently developed computational platform to identify individuals carrying KCNJ1 variants with clinical features consistent with Bartter syndrome type II. In parallel, we examined ROMK genomic data in both the NIH TOPMed and ClinVar databases with the aid of a computational algorithm that predicts protein misfolding and disease severity. Subsequent phenotypic studies using a high throughput yeast screen to assess ROMK function-and analyses of ROMK biogenesis in yeast and human cells-identified four previously uncharacterized mutations. Among these, one mutation uncovered from the two parallel approaches (G228E) destabilized ROMK and targeted it for ERAD, resulting in reduced protein expression at the cell surface. Another ERAD-targeted ROMK mutant (L320P) was found in only one of the screens. In contrast, another mutation (T300R) was ERAD-resistant, but defects in ROMK activity were apparent after expression and two-electrode voltage clamp measurements in Xenopus oocytes. Together, our results outline a new computational and experimental pipeline that can be applied to identify disease-associated alleles linked to a range of other potassium channels, and further our understanding of the ROMK structure-function relationship that may aid future therapeutic strategies. Author Summary: Bartter syndrome is a rare genetic disorder characterized by defective renal electrolyte handing, leading to debilitating symptoms and, in some patients, death in infancy. Currently, there is no cure for this disease. Bartter syndrome is divided into five types based on the causative gene. Bartter syndrome type II results from genetic variants in the gene encoding the ROMK protein, which is expressed in the kidney and assists in regulating sodium, potassium, and water homeostasis. Prior work established that some disease-associated ROMK mutants misfold and are destroyed soon after their synthesis in the endoplasmic reticulum (ER). Because a growing number of drugs have been identified that correct defective protein folding, we wished to identify an expanded cohort of similarly misshapen and unstable disease-associated ROMK variants. To this end, we developed a pipeline that employs computational analyses of human genome databases with genetic and biochemical assays. Next, we both confirmed the identity of known variants and uncovered previously uncharacterized ROMK variants associated with Bartter syndrome type II. Further analyses indicated that select mutants are targeted for ER-associated degradation, while another mutant compromises ROMK function. This work sets-the-stage for continued mining for ROMK loss of function alleles as well as other potassium channels, and positions select Bartter syndrome mutations for correction using emerging pharmaceuticals.

Construct Validity of an Instrument for Assessment of Reflective Writing-Based Portfolios of Medical Students.

PURPOSE: Assessment of reflective writing for medical students is challenging, and there is lack of an available instrument with good psychometric properties. The authors developed a new instrument for assessment of reflective writing-based portfolios and examined the construct validity of this instrument. METHODS: After an extensive literature review and pilot testing of the instrument, two raters assessed the reflective writing-based portfolios from years 2 and 3 medical students (n=135) on three occasions. The instrument consists of three criteria: organization, description of an experience and reflection on the experience. We calculated the reliability of scores using generalizability theory with a fully crossed design and two facets (raters and occasions). In addition, we measured criterion validity by testing correlations with students' scores using other assessment methods. RESULTS: The dependability (Φ) coefficient of the portfolio scores was 0.75 using two raters on three occasions. Students' portfolio scores represented 46.6% of the total variance across all score comparisons. The variance due to occasions was negligible, while the student-occasion interaction was small. The variance due to student-rater interaction represented 17.7%, and the remaining 27.7% of the variance was due to unexplained sources of error. The decision (D) study suggested that an acceptable dependability (Φ = 0.70 and 0.72) can be achieved by using two raters for one and two occasions, respectively. Finally, we found moderate to large effect-size correlations between students' scores in reflective writing-based portfolios and communication skills (r = 0.47) and PBL tutorials (r = 0.50). CONCLUSION: We demonstrated the presence of different sources of evidence that support construct validity of the study instrument. Further studies are warranted before utilizing this instrument for summative assessment of students' reflective writing-based portfolios in other medical schools.

In Vivo Quasi-Elastic Light Scattering Eye Scanner Detects Molecular Aging in Humans.

The absence of clinical tools to evaluate individual variation in the pace of aging represents a major impediment to understanding aging and maximizing health throughout life. The human lens is an ideal tissue for quantitative assessment of molecular aging in vivo. Long-lived proteins in lens fiber cells are expressed during fetal life, do not undergo turnover, accumulate molecular alterations throughout life, and are optically accessible in vivo. We used quasi-elastic light scattering (QLS) to measure age-dependent signals in lenses of healthy human subjects. Age-dependent QLS signal changes detected in vivo recapitulated time-dependent changes in hydrodynamic radius, protein polydispersity, and supramolecular order of human lens proteins during long-term incubation (~1 year) and in response to sustained oxidation (~2.5 months) in vitro. Our findings demonstrate that QLS analysis of human lens proteins provides a practical technique for noninvasive assessment of molecular aging in vivo.

Professional ambivalence: accounts of ethical practice in childhood genetic testing.

Childhood genetic testing raises complex ethical and moral dilemmas for both families and professionals. In the family sphere, the role of communication is a key aspect in the transmission of 'genetic responsibility' between adults and children. In the professional sphere, genetic responsibility is an interactional accomplishment emerging from the sometimes competing views over what constitutes the 'best interests' of the child in relation to parental preferences on the one hand, and professional judgements on the other. In the present paper we extend our previous research into parental accounts of childhood genetic testing and explore the ethical accounts of professionals in research interviews. Interviews (n = 20) were conducted with professional practitioners involved in the genetic diagnosis and management of children and their families. We first identify four inter-related themes-juxtaposition of parental rights vis-à-vis child's autonomy, elicitation of the child's autonomy, avoidance of parental responsibility and recognition of professional uncertainty. Then, using Rhetorical Discourse Analysis, we examine the range of discourse devices through which ethical accounts are situationally illustrated: contrast, reported speech, constructed dialogue, character and event work. An overarching device in these ethical accounts is the use of extreme case scenarios, which reconstruct dilemmas as justifications of professional conduct. While acknowledging ambivalence, our analysis suggests that professional judgement is not a simple matter of implementing ethical principles but rather of managing the practical conditions and consequences of interactions with parents and children. We conclude that more attention is needed to understand the way professional practitioners formulate judgements about ethical practice.

Humanizing HIV/AIDS and its (re)stigmatizing effects: HIV public 'positive' speaking in India.

Social stigma has been inextricably linked with HIV and AIDS since the epidemic erupted in the early 1980s. The stigma that has built up around HIV and AIDS is generally regarded as having a negative impact on the quality of life of HIV-positive people and on general prevention efforts. Current attempts to combat HIV-related stigma focus on increasing the acceptance of HIV among the stigmatizing public and stigmatized individuals alike. In this, the global HIV-positive community is being increasingly called upon to ;humanize' the virus, not least through public displays of HIV 'positive' health and public ;positive' speaking. This article critically explores the constitutive effects and inherent power relations of HIV Positive Speakers' Bureaus (PSBs) as a platform for such a display. Adopting a post-structuralist discourse analytic approach, we explore accounts of positive-speaking and HIV health from HIV-related non-government organizations in India and in PSB training manuals. In particular, we highlight ways in which positive-speaking in India can be seen to have significant (re)stigmatizing effects by way of ambivalent and hyper-real configurations of HIV 'positive' identity and life.

Quality of life as a mode of governance: NGO talk of HIV 'positive' health in India.

Quality of life (QOL) is being increasingly emphasized in healthcare research and practice around the world as a desirable and measurable outcome of health policy, health-seeking behaviour and overall life satisfaction. Drawing on F notions of governmentality and biopower, we take the position that the promotion of a universal and apolitical concept of QOL, as it pertains to individual health and well-being, can be seen as being tied to a neoliberal rationality of global socio-economic and health governance. In understanding non-government organizations (NGOs) as a prime catalyst for socio-economic change under neoliberalism, we highlight ways in which HIV-related NGOs in India can be seen to deploy aspects of the prescriptive and regulatory QOL discourse in their promotions of an empowered HIV 'positive' health and subjectivity. Implications are discussed in terms of the inevitable tensions involved for many under-resourced HIV-positive people in India who, in the name of QOL, are called upon to identify and act as entrepreneurial and (self)empowered individuals in particular normalizing terms.

Managing self-responsibility through other-oriented blame: family accounts of genetic testing.

'Genetic responsibility' has emerged as a key notion for understanding how genetic risk reshapes patterns of choice, identification and obligation within families. Where previous research has examined the difficulties of managing responsibility for genetic testing and disclosure of the testing process and results, there is little work that examines these themes when accounts of genetic responsibility involve blame. In this paper, we explore how forms of responsible selfhood are managed through accounts of other-oriented blame in the family sphere. Interviews (n=20) were conducted in the United Kingdom with parents whose genetic condition may have consequences for other family members. Using rhetorical discourse analysis we show that a key discursive resource for managing blame and responsibility is the use of contrast -- via constructed dialogue, character/event work and extreme case formulation -- to either endorse or contest versions of responsible/moral selfhood. We conclude that claims of personal responsibility manifest in open disclosure of genetic information often entail the blaming of others within the family. By extension, blaming others has moral and relational significance when competing views of genetic responsibility are at stake and when genetic understandings are incongruent.

The micropolitics of responsibility vis-à-vis autonomy: parental accounts of childhood genetic testing and (non)disclosure.

Genetic testing and (non)disclosure of genetic information present ethical and moral dilemmas for the management of parental responsibility vis-à-vis the child's autonomy. Ethical guidelines aimed at professionals currently seek to defer childhood testing where there is no clear medical or psychosocial benefit. This version of autonomy is derived from a bioethical paradigm which brackets the individual rights and capacities of the child. In this paper we focus on situated parental accounts of responsibility/autonomy to understand the complex forms of relational work -i.e. the micropolitics of balancing rights and responsibilities - involving a range of inherited genetic disorders. Interviews (n= 20) were conducted with parents whose genetic condition may have had consequences for their children. Using rhetorical discourse analysis, we show how parents draw upon a number of rhetorical/discoursal devices to produce accounts where genetic responsibility is actually or potentially transmitted to the child. We identify three kinds of accounting practice: (1) aligned responsibility; (2) deferred responsibility; and (3) misaligned responsibility. Each of these practices demonstrates how parents position themselves responsibly by foregrounding figures and events onto which the child's autonomy is selectively mapped. Rather than simple representations, we regard these accounts as complex moral performances that seek alignment with broader bioethical discourses.

Real nursing? The development of telenursing.

AIM: This paper is a report of a study to understand the impact of telenursing from the perspective of nurses involved in its provision, and in more traditional roles. BACKGROUND: Nurse-led telephone helplines have recently been introduced across the United Kingdom, a major step in the development of nursing practice. METHOD: A structured questionnaire was sent to all nurses working in the NHS Direct (National Health Service Direct) Wales telephone service (n = 111). Ninety-two completed questionnaires were returned (response rate 83 per cent). Two focus groups were conducted: one with telephone service nurses (n = 8) and one with other nurses (n = 5). The data were collected in 2002. FINDINGS: Respondents represented a highly educated workforce from a range of healthcare specialties. They reported that they joined the telephone service for improved salary and flexible working. Two-thirds reported improved job satisfaction. All focus group participants reported that the development of nursing skills was affected by the use of decision support software and the remote nature of the consultation. Participants reported opportunities for skill development, although the role could be stressful. All agreed that the service was popular with callers, but the nurses from outside raised concerns about whether telenursing was 'real' nursing and about the evidence base for the service and access by disadvantaged groups. CONCLUSION: Differences between the groups reflect policy tensions between the need to develop new nursing skills, including the use of technology, to improve efficiency and recognition of the worth of hands-on nursing. These tensions must be addressed for the telephone service to function as part of an integrated healthcare system.

Editorial: Team work and team talk as distributed and coordinated action in healthcare delivery.

This special issue of Communication & Medicine is dedicated to the theme of teamwork and team talk in healthcare delivery.

The management of diagnostic uncertainty and decision-making in genetics case conferences.

In this paper we examine one type of intraprofessional collaborative activity, namely case conferences in a specialist genetics clinic. Our specific focus is on how clinical geneticists manage decision-making through team talk in the event of diagnostic uncertainty which is mainly attributable to limitations in the current state of genomic knowledge, 'uncertain significances' associated with genetic test results, and a lack of information/ evidence pertaining to cases under discussion. The case conference then becomes a means to minimise the uncertainty and arrive at decisions that optimise the significance of the results in terms of clients' life trajectories. Adopting theme-oriented activity analysis, we examine video-recorded data from five case conferences in Hong Kong. Beginning with a prototypical structural mapping of the case conference activity type, our analysis focuses on what we call 'uncertain cases'. Our findings highlight three discourse types constitutive of team talk: pedagogic talk, diagnostic talk and decisional talk. In paying particular attention to how uncertainty is formulated and negotiated, we suggest that access to and assessment of different kinds of evidence as well as the activity-specific expert role-positions of the participants are crucial with regard to establishing a correct diagnosis and/or striving towards a minimisation of current uncertainties.

Discourse types and (re)distribution of responsibility in simulated emergency team encounters.

Successful teamwork, constitutive of team talk, depends largely on shared responsibility in the coordination of tasks in a goal-oriented way. This paper examines how specific modes of talk or 'discourse types' are utilised by a healthcare team in simulated emergency care. The data corpus comprises six video-recorded simulation training sessions in an emergency department at a large Norwegian hospital. Our analysis focuses on the critical moment when the original healthcare team is joined by other specialists in an ad hoc manner, which necessitates the (re)distribution of expert responsibility in the management of the patient's condition. We examine the interactional trajectories and, in particular, the discourse types surrounding the critical moment which marks the incorporation of the new team members. The analysis centres on three discourse types (online commentary, offline commentary and metacommentary) that are utilised in accomplishing the multiple tasks in a collaborative and coordinated fashion. We suggest that team talk overlays and overlaps with distributed medical work in highly charged decision-making contexts such as emergency care. The findings have relevance for how healthcare professionals and students are trained in multidisciplinary team talk and teamwork.