HumanIslets: An integrated platform for human islet data access and analysis.

Comprehensive molecular and cellular phenotyping of human islets can enable deep mechanistic insights for diabetes research. We established the Human Islet Data Analysis and Sharing (HI-DAS) consortium to advance goals in accessibility, usability, and integration of data from human islets isolated from donors with and without diabetes at the Alberta Diabetes Institute (ADI) IsletCore. Here we introduce HumanIslets.com, an open resource for the research community. This platform, which presently includes data on 547 human islet donors, allows users to access linked datasets describing molecular profiles, islet function and donor phenotypes, and to perform various statistical and functional analyses at the donor, islet and single-cell levels. As an example of the analytic capacity of this resource we show a dissociation between cell culture effects on transcript and protein expression, and an approach to correct for exocrine contamination found in hand-picked islets. Finally, we provide an example workflow and visualization that highlights links between type 2 diabetes status, SERCA3b Ca2+-ATPase levels at the transcript and protein level, insulin secretion and islet cell phenotypes. HumanIslets.com provides a growing and adaptable set of resources and tools to support the metabolism and diabetes research community.

Scalable Bioreactor-based Suspension Approach to Generate Stem Cell-derived Islets From Healthy Donor-derived iPSCs.

BACKGROUND: Induced pluripotent stem cells (iPSCs) offer the potential to generate autologous iPSC-derived islets (iPSC islets), however, remain limited by scalability and product safety. METHODS: Herein, we report stagewise characterization of cells generated following a bioreactor-based differentiation protocol. Cell characteristics were assessed using flow cytometry, quantitative reverse transcription polymerase chain reaction, patch clamping, functional assessment, and in vivo functional and immunohistochemistry evaluation. Protocol yield and costs are assessed to determine scalability. RESULTS: Differentiation was capable of generating 90.4% PDX1+/NKX6.1+ pancreatic progenitors and 100% C-peptide+/NKX6.1+ iPSC islet cells. However, 82.1%, 49.6%, and 0.9% of the cells expressed SOX9 (duct), SLC18A1 (enterochromaffin cells), and CDX2 (gut cells), respectively. Explanted grafts contained mature monohormonal islet-like cells, however, CK19+ ductal tissues persist. Using this protocol, semi-planar differentiation using 150 mm plates achieved 5.72 × 104 cells/cm2 (total 8.3 × 106 cells), whereas complete suspension differentiation within 100 mL Vertical-Wheel bioreactors significantly increased cell yield to 1.1 × 106 cells/mL (total 105.0 × 106 cells), reducing costs by 88.8%. CONCLUSIONS: This study offers a scalable suspension-based approach for iPSC islet differentiation within Vertical-Wheel bioreactors with thorough characterization of the ensuing product to enable future protocol comparison and evaluation of approaches for off-target cell elimination. Results suggest that bioreactor-based suspension differentiation protocols may facilitate scalability and clinical implementation of iPSC islet therapies.

Development and First Clinical Use of an Extracorporeal Artificial Multiorgan System in Acute-on-Chronic Liver Failure Patients.

Multiple organ failure (MOF) is a common and deadly condition. Patients with liver cirrhosis with acute-on-chronic liver failure (AOCLF) are particularly susceptible. Excess fluid accumulation in tissues makes routine hemodialysis generally ineffective because of cardiovascular instability. Patients with three or more organ failures face a mortality rate of more than 90%. Many cannot survive liver transplantation. Extracorporeal support systems like MARS (Baxter, Deerfield, IL) and Prometheus (Bad Homburg, Germany) have shown promise but fall short in bridging patients to transplantation. A novel Artificial Multi-organ Replacement System (AMOR) was developed at the University of Washington Medical Center. AMOR removes protein-bound toxins through a combination of albumin dialysis, a charcoal sorbent column, and a novel rinsing method to prevent sorbent column saturation. It removes excess fluid through hemodialysis. Ten AOCLF patients with over three organ failures were treated by the AMOR system. All patients showed significant clinical improvement. Fifty percent of the cohort received liver transplants or recovered liver function. AMOR was successful in removing large amounts of excess body fluid, which regular hemodialysis could not. AMOR is cost-effective and user-friendly. It removes excess fluid, supporting the other vital organs such as liver, kidneys, lungs, and heart. This pilot study's results encourage further exploration of AMOR for treating MOF patients.

A role and mechanism for redox sensing by SENP1 in ß-cell responses to high fat feeding.

Pancreatic ß-cells respond to metabolic stress by upregulating insulin secretion, however the underlying mechanisms remain unclear. Here we show, in ß-cells from overweight humans without diabetes and mice fed a high-fat diet for 2 days, insulin exocytosis and secretion are enhanced without increased Ca2+ influx. RNA-seq of sorted ß-cells suggests altered metabolic pathways early following high fat diet, where we find increased basal oxygen consumption and proton leak, but a more reduced cytosolic redox state. Increased ß-cell exocytosis after 2-day high fat diet is dependent on this reduced intracellular redox state and requires the sentrin-specific SUMO-protease-1. Mice with either pancreas- or ß-cell-specific deletion of this fail to up-regulate exocytosis and become rapidly glucose intolerant after 2-day high fat diet. Mechanistically, redox-sensing by the SUMO-protease requires a thiol group at C535 which together with Zn+-binding suppresses basal protease activity and unrestrained ß-cell exocytosis, and increases enzyme sensitivity to regulation by redox signals.

Multisectoral Collaborations to Increase Recruitment and Retention of Diverse Older Adults in Biomedical Research.

BACKGROUND: Older adults, especially minoritized racial-ethnic groups, are historically underrepresented in biomedical research. This study summarizes the development and assesses the impact of a review board involving a multisectoral group of stakeholders with the goal of increasing the diversity of older adults in biomedical research. METHODS: A 25-member board of community members, caregivers, researchers, and clinicians from Upstate New York reviewed 3 projects presented by researchers, clinician-scientists, and a pharmaceutical company between January and December 2022. For each biomedical research project, the reviews provided guidance to increase the recruitment and retention of diverse older adults engaged in the study. Review board members and presenters completed surveys to provide feedback on their experience in this collaboration. RESULTS: There was consistent positive feedback from all members and presenters. From member surveys, feedback trended positive in meetings throughout the year. Community members and caregivers initially indicated discomfort in expressing their views; however, these concerns subsided over time. Presenters had a very positive experience in the review board's impact on their recruitment strategy and study design, and therefore very likely to use this service again. Recommendations were made to adjust membership criteria, presentation format, and funding to sustain this effort. CONCLUSIONS: Lack of diversity for older adults represented in biomedical research contributes to ethical and generalizability ramifications. The positive feedback from all stakeholders in our multisectoral board of community members, caregivers, researchers, and clinicians offers a promising structure for developing similar strategies to increase diversity within and beyond biomedical aging research in other communities.

IL-6 trans-signaling in a humanized mouse model of scleroderma.

Fibrosis is regulated by interactions between immune and mesenchymal cells. However, the capacity of cell types to modulate human fibrosis pathology is poorly understood due to lack of a fully humanized model system. MISTRG6 mice were engineered by homologous mouse/human gene replacement to develop an immune system like humans when engrafted with human hematopoietic stem cells (HSCs). We utilized MISTRG6 mice to model scleroderma by transplantation of healthy or scleroderma skin from a patient with pansclerotic morphea to humanized mice engrafted with unmatched allogeneic HSC. We identified that scleroderma skin grafts contained both skin and bone marrow-derived human CD4 and CD8 T cells along with human endothelial cells and pericytes. Unlike healthy skin, fibroblasts in scleroderma skin were depleted and replaced by mouse fibroblasts. Furthermore, HSC engraftment alleviated multiple signatures of fibrosis, including expression of collagen and interferon genes, and proliferation and activation of human T cells. Fibrosis improvement correlated with reduced markers of T cell activation and expression of human IL-6 by mesenchymal cells. Mechanistic studies supported a model whereby IL-6 trans-signaling driven by CD4 T cell-derived soluble IL-6 receptor complexed with fibroblast-derived IL-6 promoted excess extracellular matrix gene expression. Thus, MISTRG6 mice transplanted with scleroderma skin demonstrated multiple fibrotic responses centered around human IL-6 signaling, which was improved by the presence of healthy bone marrow-derived immune cells. Our results highlight the importance of IL-6 trans-signaling in pathogenesis of scleroderma and the ability of healthy bone marrow-derived immune cells to mitigate disease.

Early Identification of Cognitive Impairment: Utility of the Mini-Cog in Non-Clinical Settings.

While we currently cannot cure Alzheimer's disease or change the course of the disease, there are advantages to early detection. Routine, evidence based, brief cognitive screens offer destigmatized opportunities for diagnosis and improve the possibility of early identification of cognitive impairment. This community-based participatory research project evaluated the use of the Mini-Cog™ instrument to detect cognitive impairment in vulnerable community-dwelling older adults when administered by trained social services providers. Over 9 months, a case manager screened 69 clients ages 65 to 94 (mean 74.67) who met inclusion criteria for the pilot; 84.1% were female, 53.6% were Black, 26% were living with undetected cognitive impairment. Although participants agreed to Mini-Cog™ screening, two-thirds with Mini-Cog™ scores indicating cognitive impairment refused referrals for further evaluation. Future interventions should reduce stigma by educating the public about dementia and engaging members of racial and cultural communities in outreach.

Changes in HIV knowledge and interest among nursing and public health students at a large Midwest University: Outcomes of implementing the National HIV Curriculum.

BACKGROUND: Ending the HIV epidemic requires additional healthcare and public health workers who are competent in HIV prevention and treatment. The National HIV Curriculum was developed to increase competency in HIV among healthcare workers in the US. OBJECTIVES: The purpose of the current study was to examine the impact of implementing the National HIV Curriculum (NHC) for nursing and public health students. DESIGN: This study employed a single-arm, cohort intervention design. SETTING: This study was conducted at large, public university in the Midwestern United States of America in a state noted for high HIV transmission. PARTICIPANTS: Undergraduate nursing, graduate nursing, and undergraduate public health students participated in this study. METHODS: An online survey of nursing and public health students was conducted following implementation of the NHC at a large, public university in the Midwest. Students were assessed on knowledge and interest of HIV using a bootstrapped paired-samples t-test approach. RESULTS: Participants (N = 175) were enrolled in the undergraduate nursing program (n = 72, 41.14 %), graduate nursing (n = 37, 21.14 %) public health (n = 37, 21.14 %), medicine (n = 10, 5.71 %), and biological, biomedical, and health sciences discipline (n = 19, 10.86 %). Overall, results suggest a consistent gain in knowledge of working with individuals living with HIV of 1.42 points (on a 4-point scale). About half (47.43 %) of all students increased interest to work with individuals living with HIV in the future. CONCLUSION: The NHC increased knowledge and interest in students across a broad range of nursing, public health, medicine, and other disciplines. This study suggests that universities can integrate the curriculum across undergraduate and graduate programs. Students at varying degree levels may benefit from the NHC. Future longitudinal studies should be conducted on the career choices of those students exposed to the NHC.

A genome-wide CRISPR screen identifies CALCOCO2 as a regulator of beta cell function influencing type 2 diabetes risk.

Identification of the genes and processes mediating genetic association signals for complex diseases represents a major challenge. As many of the genetic signals for type 2 diabetes (T2D) exert their effects through pancreatic islet-cell dysfunction, we performed a genome-wide pooled CRISPR loss-of-function screen in a human pancreatic beta cell line. We assessed the regulation of insulin content as a disease-relevant readout of beta cell function and identified 580 genes influencing this phenotype. Integration with genetic and genomic data provided experimental support for 20 candidate T2D effector transcripts including the autophagy receptor CALCOCO2. Loss of CALCOCO2 was associated with distorted mitochondria, less proinsulin-containing immature granules and accumulation of autophagosomes upon inhibition of late-stage autophagy. Carriers of T2D-associated variants at the CALCOCO2 locus further displayed altered insulin secretion. Our study highlights how cellular screens can augment existing multi-omic efforts to support mechanistic understanding and provide evidence for causal effects at genome-wide association studies loci.

Applying the 2022 Cardiovascular and Pulmonary Entry-Level Physical Therapist Competencies to Physical Therapist Education and Practice.

BACKGROUND AND PURPOSE: Competency-based education (CBE) is an emerging topic within physical therapy (PT). It has emerged to assure all stakeholders that physical therapist education program graduates are proficient in the requisite knowledge, skills, and behaviors (KSBs) essential for entry-level practice. Competencies have existed within cardiovascular and pulmonary (CVP) PT since 1980, updated in 2008, and most recently updated in 2022. This article discusses how individuals should apply the 2022 CVP competencies to clinical practice and education. POSITION AND RATIONALE: The 2022 CVP competencies were developed using a modified mixed-method Delphi approach. These competencies set a level of proficiency for KSBs used within entry-level CVP PT practice following the patient-client management model. The position put forward in this article describes how and why multiple stakeholder groups should apply these entry-level competencies specifically for graduates of physical therapist education programs who are entering practice (entry level). The competencies provide a more detailed description of expected proficiency for entry-level CVP PT practice than currently available documents. These competencies may form the basis for developing entrustable professional activities (EPAs). DISCUSSION AND CONCLUSION: The establishment of entry-level competencies is essential for use by multiple stakeholders to inform physical therapist curriculum, provide clinical instructors with a reference for expected levels of proficiency during final student clinical experiences, guide content on the Federation of State Boards of PT national licensure examination, and prepare employers to provide needed continued professional development, based on the clinical environment. These competencies lend themselves to the future development of EPAs in the PT profession for CVP PT.

An Innovative Approach to Integrating Nephrology Nursing into Acute and Critical Care.

There is a growing number of individuals living with chronic kidney disease in the United States and worldwide. There is also a nursing shortage and increased need for nurses, particularly in specialties like nephrology. Meeting these growing demands and improving conditions for nurses will take multiple approaches and broadening of current systems. An area of focus is the training and expansion of the nephrology nursing workforce. This article discusses a dialysis training program for acute and critical care nurses who are able to provide both bedside care and kidney replacement therapy in the hospital setting.

Zmiz1 is required for mature ß-cell function and mass expansion upon high fat feeding.

OBJECTIVE: Identifying the transcripts which mediate genetic association signals for type 2 diabetes (T2D) is critical to understand disease mechanisms. Studies in pancreatic islets support the transcription factor ZMIZ1 as a transcript underlying a T2D GWAS signal, but how it influences T2D risk is unknown. METHODS: ß-Cell-specific Zmiz1 knockout (Zmiz1ßKO) mice were generated and phenotypically characterised. Glucose homeostasis was assessed in Zmiz1ßKO mice and their control littermates on chow diet (CD) and high fat diet (HFD). Islet morphology and function were examined by immunohistochemistry and in vitro islet function was assessed by dynamic insulin secretion assay. Transcript and protein expression were assessed by RNA sequencing and Western blotting. In islets isolated from genotyped human donors, we assessed glucose-dependent insulin secretion and islet insulin content by static incubation assay. RESULTS: Male and female Zmiz1ßKO mice were glucose intolerant with impaired insulin secretion, compared with control littermates. Transcriptomic profiling of Zmiz1ßKO islets identified over 500 differentially expressed genes including those involved in ß-cell function and maturity, which we confirmed at the protein level. Upon HFD, Zmiz1ßKO mice fail to expand ß-cell mass and become severely diabetic. Human islets from carriers of the ZMIZ1-linked T2D-risk alleles have reduced islet insulin content and glucose-stimulated insulin secretion. CONCLUSIONS: ß-Cell Zmiz1 is required for normal glucose homeostasis. Genetic variation at the ZMIZ1 locus may influence T2D-risk by reducing islet mass expansion upon metabolic stress and the ability to maintain a mature ß-cell state.

The Feasibility of Virtual Fall Risk Screens and Evidence-Based Program Referrals Using an Algorithmic Approach.

AIMS: 1) Can virtual fall risk screens be performed safely? 2) Are older adults able to manage technology to participate in telehealth? 3) Does an algorithm aid in referral appropriate evidence-based (EBP) fall prevention programs? METHODS: An algorithm was piloted using the Zoom platform to screen for falls, to assign to intervention groups, and to guide referral to EBP. Statistical analysis of data included descriptive, parametric, and non-parametric tests. RESULTS: Forty-four participants, aged 55-94 years, were screened. A significant relationship between 30-second chair stand and referral between two programs was found (p<0.05). Spearman correlations revealed statistically significant negative correlation between 30-second chair stand and timed up-and-go (TUG) (r= -0.584; p=0.003). No safety incidents occurred. Ninety-five percent of screened participants managed technology requirements successfully. CONCLUSION: Virtual fall risk screens are feasible and offer clinicians an alternative means to screen and refer older adults for EBP.

Longitudinal Evaluation of a Deprescribing Protocol in Skilled Nursing Facilities.

Objective: To determine whether a deprescribing effort reduced several key classes of medications, and the overall number of medication classes per patient, among long-term residents of skilled nursing facilities (SNFs). Design: Retrospective, longitudinal pre/post evaluation. Data from before and during the implementation of the deprescribing effort (2017 through 2019) were compared with data from the post-intervention year (2020). Setting and Patients: Long-term resident data reported through annual comprehensive reviews conducted at two SNFs located in central New York State between 2017 and 2020 (N = 12,144). Interventions: Multifaceted, interdisciplinary deprescribing effort to reduce medications in SNF residence including clinician education, guideline development, and individual chart reviews began in 2019. Results: The mean number of medications prescribed per resident was lower at both facilities after the intervention (mean = 1.74 at both facilities) versus preintervention (1.90 at Facility 1, 1.86 at Facility 2). Significant decreases were observed in the usage rates for diuretics (-4.2%; P = 0.001), opioids (-3.8%; P = 0.001), and antipsychotics (-2.4%; P = 0.010). The raw antidepressant usage rate increased by 1.5% after the intervention but the change was not significant. Effects were robust to covariate adjustment. Conclusion: A combined, comprehensive approach to deprescribing was associated with a reduction in the overall number of medication classes per resident and in several key classes of medications. Additional research with more data and covariate control is in progress for verification of these findings.

Dissociable consequences of moderate and high volume stress are mediated by the differential energetic demands of stress.

Exposure to traumatic stress leads to persistent, deleterious behavioral and biological changes in both human and non-human species. The effects of stress are not always consistent, however, as exposure to different stressors often leads to heterogeneous effects. The intensity of the stressor may be a key factor in determining the consequences of stress. While it is difficult to quantify intensity for many stress types, electric shock exposure provides us with a stressor that has quantifiable parameters (presentation length x intensity x number = shock volume). Therefore, to test the procedural differences in shock volume that may account for some reported heterogeneity, we used two common shock procedures. Learned helplessness is a commonly reported behavioral outcome, highlighted by a deficit in subsequent shuttle-box escape, which requires a relatively high-volume stress (HVS) of about 100 uncontrollable shocks. Conversely, stress-enhanced fear learning (SEFL) is another common behavioral outcome that requires a relatively moderate-volume stress (MVS) of only 15 shocks. We exposed rats to HVS, MVS, or no stress (NS) and examined the effects on subsequent fear learning and normal weight gain. We found doubly dissociable effects of the two levels of stress. MVS enhanced contextual fear learning but did not impact weight, while HVS produced the opposite pattern. In other words, more stress does not simply lead to greater impairment. We then tested the hypothesis that the different stress-induced sequalae arouse from an energetic challenge imposed on the hippocampus by HVS but not MVS. HVS rats that consumed a glucose solution did exhibit SEFL. Furthermore, rats exposed to MVS and glucoprivated during single-trial context conditioning did not exhibit SEFL. Consistent with the hypothesis that the inability of HVS to enhance fear learning is because of an impact on the hippocampus, HVS did enhance hippocampus-independent auditory fear learning. Finally, we provide evidence that stressors of different volumes produce dissociable changes in glutamate receptor proteins in the basolateral amygdala (BLA) and dorsal hippocampus (DH). The data indicate that while the intensity of stress is a critical determinant of stress-induced phenotypes that effect is nonlinear.

Immune cells and their inflammatory mediators modify ß cells and cause checkpoint inhibitor-induced diabetes.

Checkpoint inhibitors (CPIs) targeting programmed death 1 (PD-1)/programmed death ligand 1 (PD-L1) and cytotoxic T lymphocyte antigen 4 (CTLA-4) have revolutionized cancer treatment but can trigger autoimmune complications, including CPI-induced diabetes mellitus (CPI-DM), which occurs preferentially with PD-1 blockade. We found evidence of pancreatic inflammation in patients with CPI-DM with shrinkage of pancreases, increased pancreatic enzymes, and in a case from a patient who died with CPI-DM, peri-islet lymphocytic infiltration. In the NOD mouse model, anti-PD-L1 but not anti-CTLA-4 induced diabetes rapidly. RNA sequencing revealed that cytolytic IFN-γ+CD8+ T cells infiltrated islets with anti-PD-L1. Changes in ß cells were predominantly driven by IFN-γ and TNF-α and included induction of a potentially novel ß cell population with transcriptional changes suggesting dedifferentiation. IFN-γ increased checkpoint ligand expression and activated apoptosis pathways in human ß cells in vitro. Treatment with anti-IFN-γ and anti-TNF-α prevented CPI-DM in anti-PD-L1-treated NOD mice. CPIs targeting the PD-1/PD-L1 pathway resulted in transcriptional changes in ß cells and immune infiltrates that may lead to the development of diabetes. Inhibition of inflammatory cytokines can prevent CPI-DM, suggesting a strategy for clinical application to prevent this complication.

Adaptation of Threat Responses Within the Negative Valence Framework.

External threats are a major source of our experience of negatively valanced emotion. As a threat becomes closer and more real, our specific behavior patterns and our experiences of negative affect change in response to the perceived imminence of threat. Recognizing this, the National Institute of Mental Health's Research Domain Criteria (RDoC) Negative Valence system is largely based around different levels of threat imminence. This perspective describes the correspondence between the RDoC Negative Valence System and a particular neurobiological/neuroecological model of reactions to threat, the Predatory Imminence Continuum (PIC) Theory. Using the COVID-19 pandemic as an illustration, we describe both adaptive and maladaptive behavior patterns from this perspective to illustrate how behavior in response to a crisis may get shaped. We end with suggestions on how further consideration of the PIC suggests potential modifications of the negative valence systems RDoC.