Developing a process to measure actual harm from medication errors in paediatric inpatients: From design to implementation.

AIMS: The potential harm associated with medication errors is widely reported, but data on actual harm are limited. When actual harm has been measured, assessment processes are often poorly described, limiting their ability to be reproduced by other studies. Our aim was to design and implement a new process to assess actual harm resulting from medication errors in paediatric inpatient care. METHODS: Prescribing errors were identified through retrospective medical record reviews (n = 26 369 orders) and medication administration errors through direct observation (n = 5137 administrations) in a tertiary paediatric hospital. All errors were assigned potential harm severity ratings on a 5-point scale. Multidisciplinary panels reviewed case studies for patients assigned the highest three potential severity ratings and determined the following: actual harm occurrence and severity level, plausibility of a link between the error(s) and identified harm(s) and a confidence rating if no harm had occurred. RESULTS: Multidisciplinary harm panels (n = 28) reviewed 566 case studies (173 prescribing related and 393 administration related) and found evidence of actual harm in 89 (prescribing = 22, administration = 67). Eight cases of serious harm cases were found (prescribing = 1, administration = 7) and no cases of severe harm. The panels were very confident in 65% of cases (n = 302) where no harm was found. Potential and actual harm ratings varied. CONCLUSIONS: This harm assessment process provides a systematic method for determining actual harm from medication errors. The multidisciplinary nature of the panels was critical in evaluating specific clinical, therapeutic and contextual considerations including care delivery pathways, therapeutic dose ranges and drug-drug and drug-disease interactions.

Perceived Access to Health Care of Indigenous Peoples in Canada with Rheumatoid Arthritis and their First-Degree Relatives.

OBJECTIVE: There are complex and interrelated factors that lead to inequitable healthcare delivery in Canada. Many of the factors that underlie these inequities for Canada's geographically dispersed Indigenous Peoples remain underexamined. METHODS: A cohort of 831 First Nations (FN) individuals from urban and remote communities were recruited into a longitudinal study of rheumatoid arthritis (RA) risk from 2005-2017. Data from each participant's initial enrollment visit was assessed using a survey that captured concerns with health care access. RESULTS: We found that remote participants with RA reported poor access compared to remote First-Degree Relatives (FDR, p<0.001), this difference was not observed for urban RA participants. We observed substantial differences based on sex; Females perceived access to care to be more difficult than males in both urban and remote cohorts (p<0.001). We also observed that male participants with RA reported poor access to care compared to male FDR. Importantly, access to care in remote communities appeared to improve over the duration of the study (p=0.01). In a logistic regression analysis, female sex, remote location, and older age were independent predictors of poor access to care. Predictors of poor access in participants with RA were also female sex, remote location and older age. CONCLUSION: FN peoples living in remote communities, particularly those with an established RA diagnosis, report more problems accessing healthcare. Sex-based inequities exist, with FN females reporting greater difficulties in accessing appropriate healthcare, irrespective of RA diagnosis. Addressing these sex-based inequities should be a high priority for improving healthcare delivery.

Risk Factors Associated with Medication Administration Errors in Children: A Prospective Direct Observational Study of Paediatric Inpatients.

INTRODUCTION: Limited evidence exists regarding medication administration errors (MAEs) on general paediatric wards or associated risk factors exists. OBJECTIVE: The aim of this study was to identify nurse, medication, and work-environment factors associated with MAEs among paediatric inpatients. METHODS: This was a prospective, direct observational study of 298 nurses in a paediatric referral hospital in Sydney, Australia. Trained observers recorded details of 5137 doses prepared and administered to 1530 children between 07:00 h and 22:00 h on weekdays and weekends. Observation data were compared with medication charts to identify errors. Clinical errors, potential severity and actual harm were assessed. Nurse characteristics (e.g. age, sex, experience), medication type (route, high-risk medications, use of solvent/diluent), and work variables (e.g. time of administration, weekday/weekend, use of an electronic medication management system [eMM], presence of a parent/carer) were collected. Multivariable models assessed MAE risk factors for any error, errors by route, potentially serious errors, and errors involving high-risk medication or causing actual harm. RESULTS: Errors occurred in 37.0% (n = 1899; 95% confidence interval [CI] 35.7-38.3) of administrations, 25.8% (n = 489; 95% CI 23.8-27.9) of which were rated as potentially serious. Intravenous infusions and injections had high error rates (64.7% [n = 514], 95% CI 61.3-68.0; and 77.4% [n = 188], 95% CI 71.7-82.2, respectively). For intravenous injections, 59.7% (95% CI 53.4-65.6) had potentially serious errors. No nurse characteristics were associated with MAEs. Intravenous route, early morning and weekend administrations, patient age ≥ 11 years, oral medications requiring solvents/diluents and eMM use were all significant risk factors. MAEs causing actual harm were 45% lower using an eMM compared with paper charts. CONCLUSION: Medication error prevention strategies should target intravenous administrations and not neglect older children in hospital. Attention to nurses' work environments, including improved design and integration of medication technologies, is warranted.

Experiences and Outcomes of Indigenous Patients Undergoing Bariatric Surgery: a Mixed-Method Scoping Review.

Obesity and type 2 diabetes (T2D) are growing global health concerns. Evidence suggests that Indigenous peoples are at higher lifetime risk of obesity and its associated conditions. Obesity increases the risk of T2D, cardiovascular disease, and all-cause mortality. Bariatric surgery is the most sustained and effective intervention for treating obesity-associated medical problems. This review aims to explore the experiences and outcomes of Indigenous peoples undergoing bariatric surgery in Canada, the USA, Australia, and New Zealand (CANZUS). Analysis of quantitative data revealed that Indigenous patients had fewer bariatric procedures, poorer clinic attendance, similar weight loss outcomes and slightly higher post-operative complication rates. Qualitative data analysis revealed that Indigenous patients living with obesity have a desire to improve their health and quality of life.

Stepped-Wedge Cluster RCT to Assess the Effects of an Electronic Medication System on Medication Administration Errors.

Medication errors are a leading cause of preventable harm in hospitals. Electronic medication systems (EMS) have shown success in reducing the risk of prescribing errors, but considerable less evidence is available about whether these systems support a reduction in medication administration errors in paediatrics. Using a stepped wedge cluster randomized controlled trial we investigated changes in medication administration error rates following the introduction of an EMS in a paediatric referral hospital in Sydney, Australia. Direct observations of 284 nurses as they prepared and administered 4555 medication doses was undertaken and observational data compared against patient records to identify administration errors. We found no significant change in administration errors post EMS (adjusted Odds Ratio [aOR] 1.09; 95% CI 0.89-1.32) and no change in rates of potentially serious administration errors (aOR 1.18; 95%CI 0.9-1.56), or those resulting in actual harm (aOR 0.92; 95%CI 0.34-2.46). Errors in administration of medications by some routes increased post EMS. In the first 70 days of EMS use medication administration error rates were largely unchanged.

Comparison between [68Ga]Ga-PSMA-617 and [18F]FET PET as Imaging Biomarkers in Adult Recurrent Glioblastoma.

The aim of this prospective clinical study was to evaluate the potential of the prostate specific membrane antigen (PSMA) targeting ligand, [68Ga]-PSMA-Glu-NH-CO-NH-Lys-2-naphthyl-L-Ala-cyclohexane-DOTA ([68Ga]Ga-PSMA-617) as a positron emission tomography (PET) imaging biomarker in recurrent glioblastoma patients. Patients underwent [68Ga]Ga-PSMA-617 and O-(2-[18F]-fluoroethyl)-L-tyrosine ([18F]FET) PET scans on two separate days. [68Ga]Ga-PSMA-617 tumour selectivity was assessed by comparing tumour volume delineation and by assessing the intra-patient correlation between tumour uptake on [68Ga]Ga-PSMA-617 and [18F]FET PET images. [68Ga]Ga-PSMA-617 tumour specificity was evaluated by comparing its tumour-to-brain ratio (TBR) with [18F]FET TBR and its tumour volume with the magnetic resonance imaging (MRI) contrast-enhancing (CE) tumour volume. Ten patients were recruited in this study. [68Ga]Ga-PSMA-617-avid tumour volume was larger than the [18F]FET tumour volume (p = 0.063). There was a positive intra-patient correlation (median Pearson r = 0.51; p < 0.0001) between [68Ga]Ga-PSMA-617 and [18F]FET in the tumour volume. [68Ga]Ga-PSMA-617 had significantly higher TBR (p = 0.002) than [18F]FET. The [68Ga]Ga-PSMA-617-avid tumour volume was larger than the CE tumour volume (p = 0.0039). Overall, accumulation of [68Ga]-Ga-PSMA-617 beyond [18F]FET-avid tumour regions suggests the presence of neoangiogenesis in tumour regions that are not overly metabolically active yet. Higher tumour specificity suggests that [68Ga]-Ga-PSMA-617 could be a better imaging biomarker for recurrent tumour delineation and secondary treatment planning than [18F]FET and CE MRI.

Disparities in rheumatoid arthritis outcomes for North American Indigenous populations.

Advances in rheumatoid arthritis (RA) management have significantly improved clinical outcomes of this disease; however, some Indigenous North Americans (INA) with RA have not achieved the high rates of treatment success observed in other populations. We review factors contributing to poor long-term outcomes for INA with RA. We conducted a narrative review of studies evaluating RA in INA supplemented with regional administrative health and clinical cohort data on clinical outcomes and health care utilisation. We discuss factors related to conducting research in INA populations including studies of RA prevention. NA with RA have a high burden of genetic and environmental predisposing risk factors that may impact disease phenotype, delayed or limited access to rheumatology care and advanced therapy. These factors may contribute to the observed increased rates of persistent synovitis, premature end-stage joint damage and mortality. Novel models of care delivery that are culturally sensitive and address challenges associated with providing speciality care to patients residing in remote communities with limited accessibility are needed. Progress in establishing respectful research partnerships with INA communities has created a foundation for ongoing initiatives to address care gaps including those aimed at RA prevention. This review highlights some of the challenges of diagnosing, treating, and ultimately perhaps preventing, RA in INA populations.

Inclusivity in Leader Selection: An 8-Step Process to Promote Representation of Women and Racial/Ethnic Minorities in Leadership.

At a systemic level, organizations need to take intentional steps to build inclusion, equity, and diversity at all levels. In accordance with this need, organizations have been catalyzed by national conversations surrounding gender and racial/ethnic discrimination to generate sustainable change that addresses the disenfranchisement of women and racial/ethnic minorities. Although progress toward addressing the systemic issues that perpetuate these inequities has been made in recent years, research indicates that underrepresentation at the leadership level persists in academic medicine. Further, those in more senior roles are more likely to select, sponsor, and/or mentor individuals like themselves, thereby depriving minority populations of experiences directly correlated with career development and advancement. Hence, the authors posit a focus on the characteristics and competencies of a leader along with a structured selection process is an effective intervention to reduce bias and support inclusion by recalibrating the representation of leadership within academic medical centers. To this end, the authors developed a sequential 8-step leader selection process informed by their model of leadership characteristics and competencies. This process includes a policy update, selection of interview panels, training of panelists, screening the candidate pool, structured interview guides, final candidate slates, assessments of final candidates, and development of newly selected leaders. By following this process, the authors' organization has seen an increase in the representation of women and racial/ethnic minority leaders, an increase in employees' favorable perceptions specific to representation, and data indicative of developing and maintaining an internal diverse leadership candidate pipeline. Ultimately, inclusion makes stronger and more resilient organizations. By following a standardized process grounded in leadership characteristics and competencies, academic medical centers can see changes in their leadership that mirror the populations they lead and serve. Using such processes can lead to the kind of systemic change needed to create inclusive environments.

Short- and long-term effects of an electronic medication management system on paediatric prescribing errors.

Electronic medication management (eMM) systems are designed to improve safety, but there is little evidence of their effectiveness in paediatrics. This study assesses the short-term (first 70 days of eMM use) and long-term (one-year) effectiveness of an eMM system to reduce prescribing errors, and their potential and actual harm. We use a stepped-wedge cluster randomised controlled trial (SWCRCT) at a paediatric referral hospital, with eight clusters randomised for eMM implementation. We assess long-term effects from an additional random sample of medication orders one-year post-eMM. In the SWCRCT, errors that are potential adverse drug events (ADEs) are assessed for actual harm. The study comprises 35,260 medication orders for 4821 patients. Results show no significant change in overall prescribing error rates in the first 70 days of eMM use (incident rate ratio [IRR] 1.05 [95%CI 0.92-1.21], but a 62% increase (IRR 1.62 [95%CI 1.28-2.04]) in potential ADEs suggesting immediate risks to safety. One-year post-eMM, errors decline by 36% (IRR 0.64 [95%CI 0.56-0.72]) and high-risk medication errors decrease by 33% (IRR 0.67 [95%CI 0.51-0.88]) compared to pre-eMM. In all periods, dose error rates are more than double that of other error types. Few errors are associated with actual harm, but 71% [95%CI 50-86%] of patients with harm experienced a dose error. In the short-term, eMM implementation shows no improvement in error rates, and an increase in some errors. A year after eMM error rates significantly decline suggesting long-term benefits. eMM optimisation should focus on reducing dose errors due to their high frequency and capacity to cause harm.

Can Studying Genetically Predisposed Individuals Inform Prevention Strategies for RA?

Rheumatoid arthritis (RA) is a prevalent autoimmune disorder in which complex genetic predisposition interacts with multiple environmental factors to precipitate chronic and progressive immune-mediated joint inflammation. Currently, in most affected individuals, ongoing suppression of the inflammation is required to prevent irreversible damage and functional loss. The delineation of a protracted preclinical period in which autoimmunity is initially established and then evolves to become pathogenic provides unprecedented opportunities for interventions that have the potential to prevent the onset of this lifelong disease. Clinical trials aimed at assessing the impact of specific prevention strategies require the identification of individuals who are at high risk of future RA development. Currently, these risk factors include a strong family history of RA, and the detection of circulating RA-associated autoantibodies, particularly anti-citrullinated protein antibodies (ACPA). Yet, even in such individuals, there remains considerable uncertainty about the likelihood and the timeframe for future disease development. Thus, individuals who are approached to participate in such clinical trials are left weighing the risks and benefits of the prevention measures, while having large gaps in our current understanding. To address this challenge, we have undertaken longitudinal studies of the family members of Indigenous North American RA patients, this population being known to have a high prevalence of RA, early age of onset, and familial clustering of cases. Our studies have indicated that the concepts of "risk" and "prevention" need to be communicated in a culturally relevant manner, and proposed prevention interventions need to have an appropriate balance of effectiveness, safety, convenience, and cultural acceptability. We have focused our proposed prevention studies on immunomodulatory/anti-inflammatory nutritional supplements that appear to strike such a complex balance.

Analysis of data to Advance Personalised Therapy with MR-Linac (ADAPT-MRL).

BACKGROUND: Analysis of Data to Advance Personalised Therapy with MR-Linac (ADAPT-MRL) is a multi-site, multinational, observational cohort registry designed to collect data on the use of the magnetic resonance linear accelerator (MR-Linac) for radiation therapy and patient outcomes. The registry will provide a linked repository of technical and clinical data that will form a platform for prospective studies and technology assessment. METHODS: Design: This registry aims to include an estimated 10,000 eligible participants across Australia and other countries over a 7- to 10-year period. Participants will undergo treatment and assessments in accordance with standard practice. Toxicity and survival outcomes will be assessed at baseline, during treatment, and with 3 monthly follow-up until 24 months, patient reported outcome measures will also be collected. Participants with a variety of cancers will be included. DISCUSSION: Data obtained from the ADAPT-MRL registry is expected to provide evidence on the safety and efficacy of the MR-Linac, a new technical innovation in radiation oncology. We expect this registry will generate data that will be used to optimise treatment techniques, MR-Linac software algorithms, evaluate participants' outcomes and toxicities and to create a repository of adapted plans, anatomical and functional MR sequences linked to participants' outcomes.

Early lung ultrasound affords little to the prediction of bronchopulmonary dysplasia.

OBJECTIVE: To test the hypothesis that lung ultrasound (LUS) performed in the first week of life would predict bronchopulmonary dysplasia (BPD). Secondary outcomes included the utility of LUS in predicting interim respiratory interventions. DESIGN: A prospective observational cohort study in preterm infants born <28 weeks' gestation in the single tertiary statewide neonatal intensive care unit in Western Australia. METHODS: A rigorous protocol for LUS acquisition on day 1, day 3-4, day 7, day 28 and 36 weeks' postmenstrual age (PMA) was implemented with blinded analysis using a modified, previously validated LUS score. BPD was defined by both recent National Institute of Child Health and Human Development categorical criteria and a continuous physiological variable using a modified Shift test. RESULTS: Of the 100 infants studies, primary outcome data were available for the 96 infants, surviving to 36 weeks' PMA. In a univariate logistic regression analysis, LUS on days 3-4 and day 7 accurately predicted BPD (day 3-4 OR (95% CI)=1.54 (1.03 to 2.42), p=0.044; day 7 OR (95% CI)=1.66 (1.07 to 2.70), p=0.031). The predictive value of LUS was insignificant in a multivariate model in which gestational age was the dominant predictor. LUS accurately predicted interim respiratory outcomes including surfactant administration, duration of intubation and extubation to non-invasive support at 48 hours. CONCLUSIONS: LUS in the first week of life predicted BPD. However, LUS offers little additive accuracy to current gestational age-based models. TRIAL REGISTRATION NUMBER: ACTRN12617000208303.

Associations between double-checking and medication administration errors: a direct observational study of paediatric inpatients.

BACKGROUND: Double-checking the administration of medications has been standard practice in paediatric hospitals around the world for decades. While the practice is widespread, evidence of its effectiveness in reducing errors or harm is scarce. OBJECTIVES: To measure the association between double-checking, and the occurrence and potential severity of medication administration errors (MAEs); check duration; and factors associated with double-checking adherence. METHODS: Direct observational study of 298 nurses, administering 5140 medication doses to 1523 patients, across nine wards, in a paediatric hospital. Independent observers recorded details of administrations and double-checking (independent; primed-one nurse shares information which may influence the checking nurse; incomplete; or none) in real time during weekdays and weekends between 07:00 and 22:00. Observational medication data were compared with patients' medical records by a reviewer (blinded to checking-status), to identify MAEs. MAEs were rated for potential severity. Observations included administrations where double-checking was mandated, or optional. Multivariable regression examined the association between double-checking, MAEs and potential severity; and factors associated with policy adherence. RESULTS: For 3563 administrations double-checking was mandated. Of these, 36 (1·0%) received independent double-checks, 3296 (92·5%) primed and 231 (6·5%) no/incomplete double-checks. For 1577 administrations double-checking was not mandatory, but in 26·3% (n=416) nurses chose to double-check. Where double-checking was mandated there was no significant association between double-checking and MAEs (OR 0·89 (0·65-1·21); p=0·44), or potential MAE severity (OR 0·86 (0·65-1·15); p=0·31). Where double-checking was not mandated, but performed, MAEs were less likely to occur (OR 0·71 (0·54-0·95); p=0·02) and had lower potential severity (OR 0·75 (0·57-0·99); p=0·04). Each double-check took an average of 6·4 min (107 hours/1000 administrations). CONCLUSIONS: Compliance with mandated double-checking was very high, but rarely independent. Primed double-checking was highly prevalent but compared with single-checking conferred no benefit in terms of reduced errors or severity. Our findings raise questions about if, when and how double-checking policies deliver safety benefits and warrant the considerable resource investments required in modern clinical settings.

Decolonising qualitative research to explore the experiences of Manitoba's urban Indigenous population living with type 2 diabetes mellitus, obesity and bariatric surgery.

INTRODUCTION: Obesity and type 2 diabetes mellitus (T2DM) are growing global health concerns associated with significant morbidity, mortality and healthcare expenditures. Due to histories of colonisation and contemporary marginalisation, Canada's Indigenous populations are disproportionately burdened by obesity, T2DM and many other chronic illnesses. Culturally appropriate research on experiences and outcomes of Indigenous patients undergoing bariatric surgery in Canada is scarce. This qualitative study protocol will use a decolonising approach guided by an Indigenous Elder to explore the perspectives and experiences of urban Indigenous Manitobans with respect to T2DM, obesity and bariatric surgery. This knowledge will guide the development and implementation of culturally sensitive bariatric care. METHODS AND ANALYSIS: Sequential sharing circles (SSCs) and semistructured conversational interviews that have been purposefully designed to be culturally relevant with the guidance of an Indigenous Elder and advisory group (IAG) will be carried out in Winnipeg, Manitoba, Canada. Indigenous adults who are obese (body mass index >35 kg/m2), have T2DM and live in an urban centre will be recruited. Three groups will be investigated: (1) those who have had bariatric surgery; (2) those on the wait list for bariatric surgery and (3) those not associated with a bariatric surgery programme. Each group of 10-12 participants will be guided through a semistructured script led by an Indigenous Elder. Elder-facilitated conversational interviews will also be completed following the SSCs. All content will be audio recorded and transcribed. Thematic analysis will be used to identify emerging patterns using a constructive grounded theory approach. ETHICS AND DISSEMINATION: This study has received ethical approval from the University of Manitoba Health Research Ethics Board. Findings will inform the development and implementation of culturally sensitive programmes at Manitoba's Centre for Metabolic and Bariatric Surgery. Results will be disseminated in peer-reviewed scientific journals, at obesity and Indigenous health conferences, and knowledge sharing ceremonies.

Changes in medication administration error rates associated with the introduction of electronic medication systems in hospitals: a multisite controlled before and after study.

BACKGROUND: Electronic medication systems (EMS) have been highly effective in reducing prescribing errors, but little research has investigated their effects on medication administration errors (MAEs). OBJECTIVE: To assess changes in MAE rates and types associated with EMS implementation. METHODS: This was a controlled before and after study (three intervention and three control wards) at two adult teaching hospitals. Intervention wards used an EMS with no bar-coding. Independent, trained observers shadowed nurses and recorded medications administered and compliance with 10 safety procedures. Observational data were compared against medication charts to identify errors (eg, wrong dose). Potential error severity was classified on a 5-point scale, with those scoring ≥3 identified as serious. Changes in MAE rates preintervention and postintervention by study group, accounting for differences at baseline, were calculated. RESULTS: 7451 administrations were observed (4176 pre-EMS and 3275 post-EMS). At baseline, 30.2% of administrations contained ≥1 MAE, with wrong intravenous rate, timing, volume and dose the most frequent. Post-EMS, MAEs decreased on intervention wards relative to control wards by 4.2 errors per 100 administrations (95% CI 0.2 to 8.3; p=0.04). Wrong timing errors alone decreased by 3.4 per 100 administrations (95% CI 0.01 to 6.7; p<0.05). EMS use was associated with an absolute decline in potentially serious MAEs by 2.4% (95% CI 0.8 to 3.9; p=0.003), a 56% reduction in the proportion of potentially serious MAEs. At baseline, 74.1% of administrations were non-compliant with ≥1 of 10 procedures and this rate did not significantly improve post-EMS. CONCLUSIONS: Implementation of EMS was associated with a modest, but significant, reduction in overall MAE rate, but halved the proportion of MAEs rated as potentially serious.

The Role of Work Environment in Training Sustainment: A Meta-Analysis.

OBJECTIVE: The purpose of this meta-analytic review is to examine the role of three work environment support variables (i.e., peer, supervisor, and organizational support) in training transfer and sustainment or long-term use of learned knowledge, skills, and attitudes (KSAs). BACKGROUND: Estimates demonstrate that little training is transferred to the job, wasting billions in organizational spending each year and resulting in significant loss to safety and individual and team performance. Prior research shows the importance of a supportive work environment to facilitating transfer; however, we know little of the relative importance of specific support variables. This study seeks to examine the unique roles of distinct support variables in training transfer. METHOD: A meta-analysis was conducted with multiple regressions to answer three primary research questions. RESULTS: All work environment support variables demonstrate moderate and positive correlations with transfer of training. Furthermore, multiple regressions demonstrate that each factor of the work environment explains unique variance as a predictor, with the model accounting for 32% of transfer and peer support accounting for most of R2. Motivation to transfer mediates the relationship between all three work environment support variables and transfer. Furthermore, three support variables are positively related to sustainment, with peer and supervisor support showing the strongest relationships. CONCLUSION: Findings illuminate the relative contribution of peer, supervisor, and organizational support to transfer and sustainment of training. As transfer continues to be an important yet understudied measure of the effectiveness of workplace training, these findings hold implications for both research and practice.

Human Interleukin-34 facilitates microglia-like cell differentiation and persistent HIV-1 infection in humanized mice.

BACKGROUND: Microglia are the principal innate immune defense cells of the centeral nervous system (CNS) and the target of the human immunodeficiency virus type one (HIV-1). A complete understanding of human microglial biology and function requires the cell's presence in a brain microenvironment. Lack of relevant animal models thus far has also precluded studies of HIV-1 infection. Productive viral infection in brain occurs only in human myeloid linage microglia and perivascular macrophages and requires cells present throughout the brain. Once infected, however, microglia become immune competent serving as sources of cellular neurotoxic factors leading to disrupted brain homeostasis and neurodegeneration. METHODS: Herein, we created a humanized bone-marrow chimera producing human "microglia like" cells in NOD.Cg-PrkdcscidIl2rgtm1SugTg(CMV-IL34)1/Jic mice. Newborn mice were engrafted intrahepatically with umbilical cord blood derived CD34+ hematopoietic stem progenitor cells (HSPC). After 3 months of stable engraftment, animals were infected with HIV-1ADA, a myeloid-specific tropic viral isolate. Virologic, immune and brain immunohistology were performed on blood, peripheral lymphoid tissues, and brain. RESULTS: Human interleukin-34 under the control of the cytomegalovirus promoter inserted in NSG mouse strain drove brain reconstitution of HSPC derived peripheral macrophages into microglial-like cells. These human cells expressed canonical human microglial cell markers that included CD14, CD68, CD163, CD11b, ITGB2, CX3CR1, CSFR1, TREM2 and P2RY12. Prior restriction to HIV-1 infection in the rodent brain rested on an inability to reconstitute human microglia. Thus, the natural emergence of these cells from ingressed peripheral macrophages to the brain could allow, for the first time, the study of a CNS viral reservoir. To this end we monitored HIV-1 infection in a rodent brain. Viral RNA and HIV-1p24 antigens were readily observed in infected brain tissues. Deep RNA sequencing of these infected mice and differential expression analysis revealed human-specific molecular signatures representative of antiviral and neuroinflammatory responses. CONCLUSIONS: This humanized microglia mouse reflected human HIV-1 infection in its known principal reservoir and showed the development of disease-specific innate immune inflammatory and neurotoxic responses mirroring what can occur in an infected human brain.

Human-like NSG mouse glycoproteins sialylation pattern changes the phenotype of human lymphocytes and sensitivity to HIV-1 infection.

BACKGROUND: The use of immunodeficient mice transplanted with human hematopoietic stem cells is an accepted approach to study human-specific infectious diseases such as HIV-1 and to investigate multiple aspects of human immune system development. However, mouse and human are different in sialylation patterns of proteins due to evolutionary mutations of the CMP-N-acetylneuraminic acid hydroxylase (CMAH) gene that prevent formation of N-glycolylneuraminic acid from N-acetylneuraminic acid. How changes in the mouse glycoproteins' chemistry affect phenotype and function of transplanted human hematopoietic stem cells and mature human immune cells in the course of HIV-1 infection are not known. RESULTS: We mutated mouse CMAH in the NOD/scid-IL2Rγc-/- (NSG) mouse strain, which is widely used for the transplantation of human cells, using the CRISPR/Cas9 system. The new strain provides a better environment for human immune cells. Transplantation of human hematopoietic stem cells leads to broad B cells repertoire, higher sensitivity to HIV-1 infection, and enhanced proliferation of transplanted peripheral blood lymphocytes. The mice showed no effect on the clearance of human immunoglobulins and enhanced transduction efficiency of recombinant adeno-associated viral vector rAAV2/DJ8. CONCLUSION: NSG-cmah-/- mice expand the mouse models suitable for human cells transplantation, and this new model has advantages in generating a human B cell repertoire. This strain is suitable to study different aspects of the human immune system development, provide advantages in patient-derived tissue and cell transplantation, and could allow studies of viral vectors and infectious agents that are sensitive to human-like sialylation of mouse glycoproteins.