Certified Community Behavioral Health Clinic Demonstration Impact on Health Care Utilization Among Non-Medicaid Patients with Severe Mental Illnesses.

In 2017, the Certified Community Behavioral Health Clinic (CCBHC) demonstration was implemented in New York State to redesign care delivery and financing for behavioral health services. Although CCBHC primarily targeted Medicaid patients, it was hypothesized that the clinic-level benefits of CCBHC were expected to impact even non-Medicaid patients treated in CCBHCs. To test this hypothesis, this study conducted a health insurance claims data analysis of non-Medicaid (ie, commercial and Medicare) patients with severe mental illnesses, comparing a cohort of CCBHC-treated patients with a propensity score-matched comparison cohort of patients treated by non-CCBHC clinics on rates of mental health service utilization, hospitalization, and emergency department (ED) visits. The data suggested CCBHC was associated with more than 10% increase in outpatient mental health service utilization by the patients' second year of CCBHC exposure, accompanied by similarly significant reductions in the rates of all-cause ED visits and non-psychiatric hospitalization. These findings suggest that for behavioral health clinics that serve a sufficiently large population of Medicaid, the impact of innovative clinical redesign attributable to CCBHC is likely to extend to all patients treated by them.

Culture of the Intact Postnatal Naked Mole-Rat Ovary: From Meiotic Prophase to Single-Cell RNASeq.

Recently, we reported that, in the naked mole-rat (Heterocephalus glaber) ovary, there is mitotic expansion of the primordial germ cells (PGCs), and the initiation of the meiotic program occurs postnatally. This is opposite to almost all other mammals, including humans and mice, whose reproductive cycle begins very early in development. In both mouse and human, the ovaries become populated with PGCs in utero; these PGCs will later generate the oogonia. After mitotic proliferation, these cells will trigger the meiotic program and initiate meiotic prophase I. Given that all these processes happen in utero, their analysis has been very challenging; so the ability to study them postnatally and to manipulate them with inhibitors or other substances, in the naked mole-rat, opens new possibilities in the field. In this chapter, we present a comprehensive collection of protocols that permit the culture of whole naked mole-rat ovaries, followed by analysis of germ cells, from PGCs to oocytes, in meiotic prophase I, as well the obtention of single-cell suspension or single-nuclei suspension for RNASeq.

Association between psychosocial factors and left ventricular assist device implant outcomes: A systematic review.

BACKGROUND: Psychosocial assessment is a core component of the multidisciplinary evaluation for left ventricular assist device (LVAD) implantation. The degree to which psychosocial conditions are considered a contraindication to LVAD implantation continues to be debated. This systematic review examines modifiable psychosocial factors as predictors of outcomes in patients undergoing LVAD implantation. METHODS: We conducted a systematic review following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guideline. The search resulted in 2509 articles. After deduplication, abstract and full-text review, 20 relevant articles were identified. RESULTS: Included studies evaluated socioeconomic status (n = 6), caregiver characteristics (n = 6), non-adherence (n = 6), substance use (n = 13), and psychiatric disorder (n = 8). The most commonly measured outcomes were all-cause death, readmission rate, and adverse events. Studies varied widely in definition of each psychosocial factor and selected outcomes. No psychosocial factor was consistently associated with a specific outcome in all studies. Socioeconomic status was generally not associated with outcomes. Non-adherence, psychiatric disorder, and substance use were associated with higher risks of mortality, adverse events, and/or readmission. Findings on caregiver characteristics were mixed. CONCLUSION: Of the psychosocial factors studied, non-adherence, psychiatric disorder, and substance use were the most consistently associated with an increased risk of mortality, readmission, and/or adverse events. Heterogeneity in research methodology and study quality across studies precludes firm conclusions regarding the impact of psychosocial factors on long-term patient outcomes. The results of this review reveal a need for adequately powered studies that use uniform definitions of psychosocial factors to clarify relationships between these factors and outcomes after LVAD implantation.

Impact of the COVID-19 Pandemic on Low-Value Testing and Treatment of Bronchiolitis.

BACKGROUND: Viral testing and treatments such as systemic steroids and inhaled corticosteroids are low-value care for routine bronchiolitis. We sought to determine the impact of the COVID-19 pandemic on low-value care in young children with bronchiolitis. METHODS: This was a retrospective, cross-sectional study using the Pediatric Health Information Systems database. We included children <2 years seen in a pediatric emergency department for bronchiolitis. We selected a priori 3 study periods: September 2018 to February 2020 (prepandemic), March 2020 to August 2022 (early pandemic), and September 2022 to January 2023 (late pandemic). Low-value care included respiratory syncytial virus testing, chest radiography, albuterol, or corticosteroids and was compared across the 3 time periods. RESULTS: At least 1 element of low-value care was provided in 45%, 47%, and 44% of encounters in the prepandemic, early pandemic, and late pandemic periods, respectively. There was little variation in the use of albuterol and chest radiography across time periods and a slight increase in systemic corticosteroid use from prepandemic to early and late pandemic groups. Viral testing increased from 36% prepandemic to 65% early pandemic and 67% late pandemic, which appeared to be driven by SARS-CoV-2 testing and combination viral testing. CONCLUSIONS: There was no clinically significant change in low-value care for bronchiolitis during the pandemic. Because of SARS-CoV-2 testing, however, overall frequency of viral testing increased dramatically over time. This marked increase in overall viral testing should be taken into consideration for future quality improvement efforts.

Cannabis Legalization and Resource Use for Ingestions by Young Children.

OBJECTIVE: In conjunction with widening legalization, there has been a rapid rise in unintentional cannabis ingestions in young children. We sought to determine if the legal status of recreational cannabis was associated with resource use in young children with cannabis poisoning. METHODS: This retrospective cross-sectional study of the Pediatric Health Information System included emergency department encounters between January 2016 and April 2023 for children <6 years of age with a diagnosis indicating cannabis ingestion. The primary exposure was recreational cannabis legalization status in the state in which the encounter occurred. We used logistic regression models to determine the association of recreational cannabis legality with resource utilization outcomes, adjusting for demographic covariates. RESULTS: We included 3649 children from 47 hospitals; 29% of encounters occurred in places in which recreational cannabis was legal. Compared with environments in which recreational cannabis was illegal, cannabis-legal locations had lower uses of advanced neuroimaging (24% vs 35%; adjusted odds ratio [aOR], 0.65; 95% confidence interval [CI]: 0.55-0.77), lumbar puncture (1.6% vs 3.8%; aOR, 0.42; 95% CI: 0.24-0.70), ICU admission (7.9% vs 11%; aOR, 0.71; 95% CI: 0.54-0.93), and mechanical ventilation (0.8% vs 2.9%; aOR, 0.30; 95% CI: 0.14-0.58). Urine testing was more common in places in which recreational cannabis was legal (71% vs 58%; aOR, 1.87; 95% CI: 1.59-2.20). CONCLUSIONS: State-level legalization of recreational cannabis was associated with a significant decrease in the utilization of advanced medical resources in cases of cannabis intoxication in children. These findings suggest the need for a focus on policies and procedures to minimize invasive testing in cases of cannabis intoxication in children.

Cost-effectiveness of erosion mitigation to meet water clarity targets in the Manawatu-Whanganui Region of New Zealand.

Soil erosion is a significant environmental issue worldwide. It affects water quality, biodiversity, and land productivity. New Zealand government agencies and regional councils work to mitigate soil erosion through policies, management programmes, and funding for soil conservation projects. Information about cost-effectiveness is crucial for planning, targeting, and implementing erosion mitigation to achieve improvements in sediment-related water quality. While there is a good understanding of the costs of erosion mitigation measures, there is a dearth of literature on their cost-effectiveness in reducing sediment loads and improving water quality at the catchment level. In this study, we estimate the cost-effectiveness of erosion mitigation measures in meeting visual water clarity targets. The analysis utilizes the spatially explicit SedNetNZ erosion process and sediment budget modelling in the Manawatu-Whanganui Region and region-specific mitigation costs. The erosion mitigation measures considered in the analysis include afforestation, bush retirement, riparian retirement, space-planted trees, and gully tree planting. We modelled two scenarios with on-farm erosion mitigation implemented across the region from 2021 to 2100, resulting in a 48% and 60% reduction of total sediment load. We estimate the marginal costs to achieve the visual national bottom line for water clarity, as assessed by the length of waterways that meet the clarity targets. We also estimate the marginal costs of improving average water clarity, which can be linked with non-market valuation studies when conducting a cost-benefit analysis. We find that gully tree planting and space-planted trees are the most cost-effective mitigation measures and that riparian retirement is the least cost-effective. Moreover, cost-effectiveness is highly dependent on current land use and the biophysical features of the landscape. Our estimates can be used in cost-benefit analysis to plan and prioritize soil erosion mitigation at the catchment and regional levels.

Base-Promoted Synthesis of Isoquinolines through a Tandem Reaction of 2-Methyl-arylaldehydes and Nitriles.

A convenient method for preparing 3-aryl isoquinolines via a base-promoted tandem reaction is presented. Simply combining commercially available 2-methyl-arylaldehydes, benzonitriles, NaN(SiMe3)2, and Cs2CO3 enabled the synthesis of a variety of isoquinolines (23 examples, ≤90% yield). Among the syntheses of isoquinolines, the transition metal-free method described here is straightforward, practical, and operationally simple.

Sex Differences in Delirium after Coronary Artery Bypass Graft Surgery and Perioperative Neuropsychiatric Conditions: A Secondary Analysis of a Cohort Study.

BACKGROUND: Biological sex influences the risk of depression and cognitive impairment, but its role in relation to postoperative delirium is unclear. This analysis investigates sex differences in delirium risk after coronary artery bypass graft (CABG) surgery and sex-related differences in relation to affective and cognitive symptoms. METHODS: This is a secondary analysis of the Neuropsychiatric Outcomes After Heart Surgery (NOAHS) study, a single-site, observational study of a CABG surgery cohort (n = 149). Preoperative characteristics are stratified by sex, and baseline variables that differ by sex are evaluated to understand whether sex modifies their relationships with delirium. We also evaluate sex differences in one-month depression and cognition. RESULTS: Female sex is associated with several delirium risk factors, including higher risk of preoperative depression and middle cerebral artery (MCA) stenosis. MCA stenosis was statistically associated with delirium only among women (OR 15.6, 95% CI 1.5, 164.4); mild cognitive impairment (MCI) was associated with delirium only in men (OR 4.6, 95% CI 1.2, 17.9). Other sex-based differences failed to reach statistical significance. Depression remained commoner among women 1 month post-CABG. CONCLUSIONS: Women in this CABG cohort were more likely to have depression at baseline and 1 month postoperatively, as well as MCA stenosis and postoperative delirium. Sex might modify the relationship between post-CABG delirium and its risk factors including MCA stenosis and MCI. Cerebrovascular disease deserves study as a potential explanation linking female sex and a range of poor outcomes among women with coronary heart disease.

How to differentiate induced pluripotent stem cells into sensory neurons for disease modelling: a functional assessment.

BACKGROUND: Human induced pluripotent stem cell (iPSC)-derived peripheral sensory neurons present a valuable tool to model human diseases and are a source for applications in drug discovery and regenerative medicine. Clinically, peripheral sensory neuropathies can result in maladies ranging from a complete loss of pain to severe painful neuropathic disorders. Sensory neurons are located in the dorsal root ganglion and are comprised of functionally diverse neuronal types. Low efficiency, reproducibility concerns, variations arising due to genetic factors and time needed to generate functionally mature neuronal populations from iPSCs remain key challenges to study human nociception in vitro. Here, we report a detailed functional characterization of iPSC-derived sensory neurons with an accelerated differentiation protocol ("Anatomic" protocol) compared to the most commonly used small molecule approach ("Chambers" protocol). Anatomic's commercially available RealDRG™ were further characterized for both functional and expression phenotyping of key nociceptor markers. METHODS: Multiple iPSC clones derived from different reprogramming methods, genetics, age, and somatic cell sources were used to generate sensory neurons. Manual patch clamp was used to functionally characterize both control and patient-derived neurons. High throughput techniques were further used to demonstrate that RealDRGs™ derived from the Anatomic protocol are amenable to high throughput technologies for disease modelling. RESULTS: The Anatomic protocol rendered a purer culture without the use of mitomycin C to suppress non-neuronal outgrowth, while Chambers differentiations yielded a mix of cell types. Chambers protocol results in predominantly tonic firing when compared to Anatomic protocol. Patient-derived nociceptors displayed higher frequency firing compared to control subject with both, Chambers and Anatomic differentiation approaches, underlining their potential use for clinical phenotyping as a disease-in-a-dish model. RealDRG™ sensory neurons show heterogeneity of nociceptive markers indicating that the cells may be useful as a humanized model system for translational studies. CONCLUSIONS: We validated the efficiency of two differentiation protocols and their potential application for functional assessment and thus understanding the disease mechanisms from patients suffering from pain disorders. We propose that both differentiation methods can be further exploited for understanding mechanisms and development of novel treatments in pain disorders.

Sulfonamides as N-Centered Radical Precursors for C-N Coupling Reactions To Generate Amidines.

Nitrogen-centered radicals (NCRs) are valuable intermediates for the construction of C-N bonds. Traditional methods for the generation of NCRs employ toxic radical initiators, transition metal catalysts, photocatalysts, or organometallic reagents. Herein, we report a novel strategy for the generation of NCRs toward the construction of C-N bonds under transition-metal-free conditions. Thus, super-electron-donor (SED) 2-azaallyl anions undergo single-electron transfer (SET) with sulfonamides, forming aminyl radicals (R2N•, R = alkyl) and culminating in the generation of amidines bearing various functional groups (33 examples, up to 96% yield). Broad substrate scope and gram-scale telescoped preparation demonstrate the practicality of this method. Radical clock and electron paramagnetic resonance (EPR) experiments support the proposed radical coupling pathway between the generated N-centered radical and the C-centered 2-azaallyl radical.

Serum IL-36ß levels are associated with Insulin sensitivity in paediatric patients with obesity.

Although the orchestrating role of Interleukin-36 cytokines in regulating inflammation at barrier tissue sites, is well established, whether they play a significant role in the settings of metabolic health and disease, has yet to be fully established. Several recent studies have demonstrated that IL-36 cytokine expression is elevated among adult patients with obesity, and can play roles in regulating both insulin sensitivity and driving inflammation. In this report, we have extended these analyses to paediatric patients and identified an association between elevated serum levels of expression of the specific Interleukin-36 subfamily member, IL-36ß, among children with obesity displaying insulin sensitivity, compared to children with obesity who are insulin resistant. While these data further indicate a possible protective role for IL-36 in metabolic health, they also differ with previous findings from an adult patient cohort, where elevated levels of the related cytokine, IL-36γ, were found to occur in association with improved metabolic health. While highlighting important differences between paediatric and adult patient cohorts in the context of metabolic disease associated with obesity, these data underscore the need for a deeper mechanistic analysis of the role of IL-36 cytokines in disease.

Automated Screening to Enhance Proactive Consultation-Liaison Psychiatry Services in Acute Medicine Units: Evaluation of Service Outcomes.

Objective: Proactive consultation-liaison (C-L) psychiatry aims to meet the mental health needs of medical-surgical populations-many of which go unmet by the conventional C-L model-through systematic screening and integrated care. We implemented an automated screening list to enhance case identification of an existing proactive C-L service and evaluated service metrics along with clinician- and patient-reported outcomes.Methods: Service outcomes were evaluated using historical and contemporary comparison data. Adjusted difference-in-difference analyses were used to determine change in consult characteristics, mean length of stay (LOS), and scores on Hospital Consumer Assessment of Healthcare Providers and Systems (HCAHPS). Practitioners and nurses were surveyed regarding service satisfaction, perceived safety, and burnout.Results: During the intervention, the consult rate was 3-fold higher than at baseline. Change in time to consultation was equivocal. Overall mean LOS was not reduced, but observed LOS was 1.2 days shorter than expected among non-COVID patients receiving psychiatric consultation (P = not significant). Mean patient-rated hospital satisfaction on HCAHPS was 1 point higher on intervention units during the intervention. Surveys revealed broad satisfaction with this model among practitioners and improved perception of safety among nurses.Conclusions: Proactive C-L psychiatry enhanced by automated screening was associated with improved service utilization and evidence suggestive of LOS reduction among those most likely to receive direct benefit from this model of care. Further, both patient and clinician ratings were improved during the intervention. Proactive C-L psychiatry provides benefits to patients, clinicians, and health systems and may be poised to achieve the Triple Aim in health care.Prim Care Companion CNS Disord 2024;26(2):23m03647. Author affiliations are listed at the end of this article.

Efficient construction of functionalized pyrroloindolines through cascade radical cyclization/intermolecular coupling.

Pyrroloindolines are important structural units in nature and the pharmaceutical industry, however, most approaches to such structures involve transition-metal or photoredox catalysts. Herein, we describe the first tandem SET/radical cyclization/intermolecular coupling between 2-azaallyl anions and indole acetamides. This method enables the transition-metal-free synthesis of C3a-substituted pyrroloindolines under mild and convenient conditions. The synthetic utility of this transformation is demonstrated by the construction of an array of C3a-methylamine pyrroloindolines with good functional group tolerance and yields. Gram-scale sequential one-pot synthesis and hydrolysis reactions demonstrate the potential synthetic utility and scalability of this approach.

Construction of C-S and C-Se Bonds from Unstrained Ketone Precursors under Photoredox Catalysis.

A mild photoredox catalyzed construction of sulfides, disulfides, selenides, sulfoxides and sulfones from unstrained ketone precursors is introduced. Combination of this deacylative process with SN 2 or coupling reactions provides novel and convenient modular strategies toward unsymmetrical or symmetric disulfides. Reactivity studies favor a bromine radical that initiates a HAT (Hydrogen Atom Transfer) from the aminal intermediate resulting in expulsion of a C-centered radical that is intercepted to make C-S and C-Se bonds. Gram scale reactions, broad substrate scope and tolerance towards various functional groups render this method appealing for future applications in the synthesis of organosulfur and selenium complexes.

Mission Alliance Community Engagement Project: Exploring the Impact of COVID-19 on Social Isolation, Loneliness, Mental Health and Wellbeing in Veterans.

During the Coronavirus disease pandemic, many U.S. veterans with posttraumatic stress disorder (PTSD) experienced increased symptomology and worsened mental health and well-being due in part to social isolation and loneliness. The Mission Alliance project explored these ramifications and prioritized critical issues expressed by U.S. veterans and stakeholders (N = 182) during virtual regional meetings (N = 32). Field notes created specifically for this project were recorded and thematically analyzed. Emerging themes included: (1) social isolation: missed opportunities, collapsed social circles, work-life balance, fostering relationships, and evolving health care delivery; (2) loneliness: deteriorated mental health, suffered with PTSD together but alone, looked out for each other, ambivalence toward technology, and strained and broken systems; (3) mental health: sense of chaos, increased demand and decreased access, aggravation, implementation of tools, innovative solutions, fear and loss, and availability of resources; (4) wellbeing: sense of purpose, holistic perspective on well-being, recognition of balance, persisting stigma, redefined pressures, freedom to direct treatment, and reconnection and disconnection. A PTSD-related patient centered outcomes research (PCOR)/comparative effectiveness research (CER) agenda was developed from these themes. Establishment of a veteran and stakeholder network is suggested to support, facilitate, and promote the PTSD-related PCOR/CER agenda. Furthermore, enhancement of opportunities for veterans with PTSD and stakeholders to partner in PCOR/CER is required to develop and conduct projects that lead to PTSD-related comprehensive care of veterans affected by traumatic events with the potential to translate findings to other populations.

Glycolytic reprogramming fuels myeloid cell-driven hypercoagulability.

BACKGROUND: Myeloid cell metabolic reprogramming is a hallmark of inflammatory disease; however, its role in inflammation-induced hypercoagulability is poorly understood. OBJECTIVES: We aimed to evaluate the role of inflammation-associated metabolic reprogramming in regulating blood coagulation. METHODS: We used novel myeloid cell-based global hemostasis assays and murine models of immunometabolic disease. RESULTS: Glycolysis was essential for enhanced activated myeloid cell tissue factor expression and decryption, driving increased cell-dependent thrombin generation in response to inflammatory challenge. Similarly, inhibition of glycolysis enhanced activated macrophage fibrinolytic activity through reduced plasminogen activator inhibitor 1 activity. Macrophage polarization or activation markedly increased endothelial protein C receptor (EPCR) expression on monocytes and macrophages, leading to increased myeloid cell-dependent protein C activation. Importantly, inflammation-dependent EPCR expression on tissue-resident macrophages was also observed in vivo. Adipose tissue macrophages from obese mice fed a high-fat diet exhibited significantly enhanced EPCR expression and activated protein C generation compared with macrophages isolated from the adipose tissue of healthy mice. Similarly, the induction of colitis in mice prompted infiltration of EPCR+ innate myeloid cells within inflamed colonic tissue that were absent from the intestinal tissue of healthy mice. CONCLUSION: Collectively, this study identifies immunometabolic regulation of myeloid cell hypercoagulability, opening new therapeutic possibilities for targeted mitigation of thromboinflammatory disease.

Asthmalitis? Diagnostic Variability of Asthma and Bronchiolitis in Children <24 Months.

BACKGROUND AND OBJECTIVES: Bronchiolitis and asthma have similar acute clinical presentations in young children yet have opposing treatment recommendations. We aimed to assess the role of age and other factors in the diagnosis of bronchiolitis and asthma in children <24 months of age. METHODS: We conducted a retrospective cross-sectional analysis of the Pediatric Health Information System database. We included children aged <2 years diagnosed with bronchiolitis, asthma, wheeze, or bronchospasm in emergency department or hospital encounters from 2017 to 2021. We described variation by age and between institutions. We used mixed-effects models to assess factors associated with a non-bronchiolitis diagnosis in children 12 to 23 months of age. RESULTS: We included 554 158 encounters from 42 hospitals. Bronchiolitis made up 98% of encounters for children <3 months of age, whereas asthma diagnoses increased with age and were included in 44% of encounters at 23 months of age. Diagnosis patterns varied widely between hospitals. In children 12 to 23 months of age, the odds of a non-bronchiolitis diagnosis increased with month of age (odds ratio [OR] 1.13, 95% confidence interval [CI] 1.12-1.13), male sex (OR 1.37, 95% CI 1.35-1.40), non-Hispanic Black race (OR 1.54, 95% CI 1.50-1.58), number of previous encounters (OR 2.73, 95% CI 2.61-2.86, for 3 or more encounters), and previous albuterol use (OR 2.24, 95% CI 2.16-2.32). CONCLUSIONS: Non-bronchiolitis diagnoses and the use of inhaled bronchodilators and systemic steroids for acute wheezing respiratory illness increase with month of age in children aged 0 to 23 months. Better definitions of clinical phenotypes of bronchiolitis and asthma would allow for more appropriate treatment in acute care settings, particularly in children 12 to 23 months of age.

Do young children with known cannabis intoxication benefit from further neurological-based testing or imaging?

BACKGROUND: Recent work has demonstrated that children with unintentional cannabis ingestions often undergo extensive ancillary testing such as head imaging or lumbar puncture. To better understand the yield of these tests, our objective was to describe the frequency of additional significant diagnoses in children with cannabis ingestion. METHODS: We performed a retrospective cross-sectional study of the Pediatric Health Information System (PHIS) database, including ED encounters from January 2016 to April 2023 with a diagnosis indicating cannabis exposure in children <6 years of age. We assessed the frequency of co-diagnoses that would be found on head imaging, lumbar puncture, or toxicology testing. RESULTS: We included 4132 ED encounters for cannabis ingestion from 47 hospitals. Of these, 1243 (30%) received head imaging and 130 (3.1%) underwent lumbar puncture. There were 23 children (0.6%) with diagnosis of skull fracture or intracranial hemorrhage, 4 (<0.1%) with intracranial neoplasm, and 0 (0%) with a diagnosis for meningitis or intracranial abscess. Presence of discharge diagnosis for other drugs was also uncommon. The most frequent drug ingestion co-diagnoses were cocaine in 43 (1.0%) and opioids in 22 (0.5%) encounters. CONCLUSION: In children with cannabis intoxication, high rates of head imaging and lumbar puncture are likely driven by the signs of altered mental status at presentation. These data suggest that if cannabis ingestion is considered early and identified quickly with testing, neuroimaging, particularly that with ionizing radiation, may be low yield.

How to differentiate induced pluripotent stem cells into sensory neurons for disease modelling: a comparison of two protocols.

Background: Human induced pluripotent stem cell (iPSC)-derived peripheral sensory neurons present a valuable tool to model human diseases and are a source for applications in drug discovery and regenerative medicine. Clinically, peripheral sensory neuropathies can result in maladies ranging from a complete loss of pain to severe painful neuropathic symptoms. Sensory neurons are located in the dorsal root ganglion and are comprised of functionally diverse neuronal types. Low efficiency, reproducibility concerns, variations arising due to genetic factors and time needed to generate functionally mature neuronal populations from iPSCs for disease modelling remain key challenges to study human nociception in vitro. Here, we report a detailed characterization of iPSC-derived sensory neurons with an accelerated differentiation protocol ("Anatomic" protocol) compared to the most commonly used small molecule approach ("Chambers" protocol). Methods: Multiple iPSC clones derived from different reprogramming methods, genetics, age, and somatic cell sources were used to generate sensory neurons. Expression profiling of sensory neurons was performed with Immunocytochemistry and in situ hybridization techniques. Manual patch clamp and high throughput cellular screening systems (Fluorescence imaging plate reader, automated patch clamp and multi-well microelectrode arrays recordings) were applied to functionally characterize the generated sensory neurons. Results: The Anatomic protocol rendered a purer culture without the use of mitomycin C to suppress non-neuronal outgrowth, while Chambers differentiations yielded a mix of cell types. High throughput systems confirmed functional expression of Na+ and K+ ion channels. Multi-well microelectrode recordings display spontaneously active neurons with sensitivity to increased temperature indicating expression of heat sensitive ion channels. Patient-derived nociceptors displayed higher frequency firing compared to control subject with both, Chambers and Anatomic differentiation approaches, underlining their potential use for clinical phenotyping as a disease-in-a-dish model. Conclusions: We validated the efficiency of two differentiation protocols and their potential application for understanding the disease mechanisms from patients suffering from pain disorders. We propose that both differentiation methods can be further exploited for understanding mechanisms and development of novel treatments in pain disorders.

Accelerated differentiation of human induced pluripotent stem cells into regionally specific dorsal and ventral spinal neural progenitor cells for application in spinal cord therapeutics.

Spinal cord injury can attenuate both motor and sensory function with minimal potential for full recovery. Research utilizing human induced pluripotent stem cell (hiPSC) -derived spinal cell types for in vivo remodeling and neuromodulation after spinal cord injury has grown substantially in recent years. However, the majority of protocols for the differentiation of spinal neurons are lengthy, lack the appropriate dorsoventral or rostrocaudal specification, and are not typically replicated in more than one cell line. Furthermore, most researchers currently utilize hiPSC-derived motor neurons for cell transplantation after injury, with very little exploration of spinal sensory neuron transplantation. The lack of studies that utilize sensory populations may be due in part to the relative scarcity of dorsal horn differentiation protocols. Building upon our previously published work that demonstrated the rapid establishment of a primitive ectoderm population from hiPSCs, we describe here the production of a diverse population of both ventral spinal and dorsal horn progenitor cells. Our work creates a novel system allowing dorsal and ventral spinal neurons to be differentiated from the same intermediate ectoderm population, making it possible to construct the dorsal and ventral domains of the spinal cord while decreasing variability. This technology can be used in tandem with biomaterials and pharmacology to improve cell transplantation for spinal cord injury, increasing the potential for neuroregeneration.