Bioengineered Hydrogels Recapitulate Fibroblast Heterogeneity in Cancer.

Recently mapped transcriptomic landscapes reveal the extent of heterogeneity in cancer-associated fibroblasts (CAFs) beyond previously established single-gene markers. Functional analyses of individual CAF subsets within the tumor microenvironment are critical to develop more accurate CAF-targeting therapeutic strategies. However, there is a lack of robust preclinical models that reflect this heterogeneity in vitro. In this study, single-cell RNA sequencing datasets acquired from head and neck squamous cell carcinoma tissues to predict microenvironmental and cellular features governing individual CAF subsets are leveraged. Some of these features are then incorporated into a tunable hyaluronan-based hydrogel system to culture patient-derived CAFs. Control over hydrogel degradability and integrin adhesiveness enabled derivation of the predominant myofibroblastic and inflammatory CAF subsets, as shown through changes in cell morphology and transcriptomic profiles. Last, using these hydrogel-cultured CAFs, microtubule dynamics are identified, but not actomyosin contractility, as a key mediator of CAF plasticity. The recapitulation of CAF heterogeneity in vitro using defined hydrogels presents unique opportunities for advancing the understanding of CAF biology and evaluation of CAF-targeting therapeutics.

Comparison of the µ-opioid receptor antagonists methocinnamox (MCAM) and naloxone to reverse and prevent the ventilatory depressant effects of fentanyl, carfentanil, 3-methylfentanyl, and heroin in male rats.

The number of opioid overdose deaths has increased significantly over the past decade. The life-threatening effect of opioids is hypoventilation that can be reversed by the µ-opioid receptor (MOR) antagonist naloxone; however, because of the very short duration of action of naloxone, re-emergence of MOR agonist-induced hypoventilation can occur, requiring additional doses of naloxone. The MOR antagonist methocinnamox (MCAM) antagonizes hypoventilation by the non-morphinan fentanyl and the morphinan heroin in laboratory animals with an unusually long duration of action. Whole-body plethysmography was used to compare the potency and effectiveness of MCAM and naloxone for preventing and reversing hypoventilation by fentanyl, heroin, and the ultra-potent and longer-acting fentanyl analogs carfentanil and 3-methylfentanyl in male rats breathing normal air. Sessions comprised a 45-minute habituation period followed by intravenous (i.v.) administration of saline or an acute dose of MOR agonist. The rank order of potency to decrease ventilation was 3-methylfentanyl > carfentanil > fentanyl > heroin. MCAM (0.0001-0.1 mg/kg) and naloxone (0.0001-0.01 mg/kg) dose-dependently reversed hypoventilation by 3-methylfentanyl (0.01 mg/kg), carfentanil (0.01 mg/kg), fentanyl (0.1 mg/kg), or heroin (3.2 mg/kg). For prevention studies, MCAM, naloxone, or vehicle was administered i.v. 22, 46, or 70 hours prior to a MOR agonist. When administered 22 hours earlier, MCAM (0.1-1.0 mg/kg) but not naloxone (1.0 mg/kg) prevented hypoventilation by each MOR agonist. This study demonstrates the effectiveness of MCAM to reverse and prevent hypoventilation by MOR agonists including ultra-potent fentanyl analogs that have a long duration of action. Significance Statement The number of opioid overdose deaths increased over the past decade despite the availability of antagonists that can prevent and reverse the effects of opioids. This study demonstrates the effectiveness and long duration of action of the µ-opioid receptor (MOR) antagonist methocinnamox (MCAM) for reversing and preventing hypoventilation by MOR agonists including ultra-potent fentanyl analogs. These results provide support for the notion that MCAM has the potential to positively impact the ongoing opioid crisis by reversing and preventing opioid overdose.

Cost-effectiveness of system-level mental health strategies for young people in the Australian Capital Territory: a dynamic simulation modelling study.

BACKGROUND: Regional mental health planning is a key challenge for decision makers because mental health care is a complex, dynamic system. Economic evaluation using a system dynamics modelling approach presents an opportunity for more sophisticated planning and important evidence on the value of alternative investments. We aimed to investigate the cost-effectiveness of eight systems-based interventions targeted at improving the mental health and wellbeing of children, adolescents, and young adults in the Australian Capital Territory (ACT). METHODS: We assessed eight interventions for children and young people (aged ≤25 years) with low, moderate, and high-to-very-high psychological distress: technology-enabled integrated care, emergency department-based suicide prevention, crisis response service, family education programme, online parenting programme, school-based suicide prevention programme, trauma service for youths, and multicultural-informed care. We developed a system dynamics model for the ACT through a participatory process and calibrated the model with historical data, including population demographics, the prevalence of psychological distress, and mental health services provision. We calculated incremental cost-effectiveness ratios compared with business as usual for cost (AU$) per: quality-adjusted life-year (QALY), suicide death avoided, self-harm related hospital admissions avoided, and mental health-related emergency department presentation, using a 10-year time horizon for health-care and societal perspectives. We investigated uncertainty through probabilistic sensitivity analysis and deterministic sensitivity analysis, including using a 30-year timeframe. FINDINGS: From a societal perspective, increased investment in technology-enabled integrated care, family education, an online parenting programme, and multicultural-informed care were expected to improve health outcomes (incremental QALYs 4517 [95% UI -3135 to 14 507] for technology-enabled integrated care; 339 [91 to 661] for family education; 724 [114 to 1149] for the online parenting programme; and 137 [88 to 194] for multicultural-informed care) and reduce costs ($-91·4 million [-382·7 to 100·7]; $-12·8 million [-21·0 to -6·6]; $-3·6 million  [-6·3 to 0·2]; and $-3·1 million [-4·5 to -1·8], respectively) compared with business as usual using a 10-year time horizon. The incremental net monetary benefit for the societal perspective for these four interventions was $452 million (-351 to 1555), $40 million (14 to 74), $61 million (9 to 98), and $14 million (9 to 20), respectively, compared with business as usual, when QALYs were monetised using a willingness to pay of $79 930 per QALY. Synergistic effects are anticipated if these interventions were to be implemented concurrently. The univariate and probabilistic sensitivity analyses indicated a high level of certainty in the results. Although emergency department-based suicide prevention and school-based suicide prevention were not cost effective in the base case (41 QALYs [0 to 48], incremental cost $4·1 million [1·2 to 8·2] for emergency department-based suicide prevention; -234 QALYs [-764 to 12], incremental cost $90·3 million [72·2 to 111·0] for school-based suicide prevention) compared with business as usual, there were scenarios for which these interventions could be considered cost effective. A dedicated trauma service for young people (9 QALYs gained [4 to 16], incremental cost $8·3 million [6·8 to 10·0]) and a crisis response service (-11 QALYs gained [-12 to -10], incremental cost $7·8 million [5·1 to 11·0]) were unlikely to be cost effective in terms of QALYs. INTERPRETATION: Synergistic effects were identified, supporting the combined implementation of technology-enabled integrated care, family education, an online parenting programme, and multicultural-informed care. Synergistic effects, emergent outcomes in the form of unintended consequences, the capability to account for service capacity constraints, and ease of use by stakeholders are unique attributes of a system dynamics modelling approach to economic evaluation. FUNDING: BHP Foundation.

Bioengineered hydrogels enhance ex vivo preservation of patient-derived tumor explants for drug evaluation.

Ex vivo patient-derived tumor slices (PDTS) are currently limited by short-term viability in culture. Here, we show how bioengineered hydrogels enable the identification of key matrix parameters that significantly enhance PDTS viability compared to conventional culture systems. As demonstrated using single-cell RNA sequencing and high-dimensional flow cytometry, hydrogel-embedded PDTS tightly preserved cancer, cancer-associated fibroblast, and various immune cell populations and subpopulations in the corresponding original tumor. Cell-cell communication networks within the tumor microenvironment, including immune checkpoint ligand-receptor interactions, were also maintained. Remarkably, our results from a co-clinical trial suggest hydrogel-embedded PDTS may predict sensitivity to immune checkpoint inhibitors (ICIs) in head and neck cancer patients. Further, we show how these longer term-cultured tumor explants uniquely enable the sampling and detection of temporal evolution in molecular readouts when treated with ICIs. By preserving the compositional heterogeneity and complexity of patient tumors, hydrogel-embedded PDTS provide a valuable tool to facilitate experiments targeting the tumor microenvironment.

Ventilatory Effects of Fentanyl, Heroin, and d-Methamphetamine, Alone and in Mixtures in Male Rats Breathing Normal Air .

The number of drug overdoses and deaths has increased significantly over the past decade and co-use of opioids and stimulants is associated with greater likelihood of overdose and decreased likelihood of accessing treatment, compared with use of opioids alone. Potential adverse effects of opioid/stimulant mixtures, particularly methamphetamine, are not well characterized. Two structurally different drugs with agonist properties at µ-opioid receptors (MOR), fentanyl and heroin, and d-methamphetamine, alone and in mixtures, were assessed for their effects on ventilation in rats breathing normal air. Whole-body phethysmography chambers were equipped with a tower and swivel allowing infusions to indwelling intravenous catheters. After a 45-minute habituation period, saline, fentanyl, heroin, or d-methamphetamine, alone and in mixtures, was administered. Five minutes later, the opioid receptor antagonist naloxone or vehicle was injected. Fentanyl (0.0032-0.1 mg/kg) and heroin (0.32-3.2 mg/kg) decreased ventilation [frequency (f) and tidal volume (VT)] in a dose-related manner whereas d-methamphetamine (0.1-3.2 mg/kg) increased f to >400% of control and decreased VT to <60% of control, overall increasing minute volume (product of f and VT) to >240% of control. When combined, d-methamphetamine (0.1-3.2 mg/kg) attenuated the ventilatory depressant effects of fentanyl (0.1 mg/kg) and heroin (3.2 mg/kg). d-Methamphetamine did not alter the potency of naloxone to reverse the ventilatory depressant effects of fentanyl or heroin. These studies demonstrate that d-methamphetamine can attenuate the ventilatory depressant effects of moderate doses of opioid receptor agonists while not altering the potency of naloxone to reverse opioid hypoventilation. SIGNIFICANCE STATEMENT: Co-use of opioids and stimulants is associated with greater likelihood of overdose and decreased likelihood of accessing treatment, compared with use of opioids alone. Potential adverse effects of opioid/stimulant mixtures are not well characterized. This study reports that 1) d-methamphetamine attenuates the ventilatory depressant effects of moderate doses of two structurally different opioid receptor agonists, fentanyl and heroin, and 2) d-methamphetamine does not alter potency or effectiveness of naloxone to reverse the ventilatory depressant effects of these opioid receptor agonists.

Pharmacokinetic and Pharmacodynamic Consequences of Cytochrome P450 3A Inhibition on Mitragynine Metabolism in Rats.

Mitragynine, an opioidergic alkaloid present in Mitragyna speciosa (kratom), is metabolized by cytochrome P450 3A (CYP3A) to 7-hydroxymitragynine, a more potent opioid receptor agonist. The extent to which conversion to 7-hydroxymitragynine mediates the in vivo effects of mitragynine is unclear. The current study examined how CYP3A inhibition (ketoconazole) modifies the pharmacokinetics of mitragynine in rat liver microsomes in vitro. The study further examined how ketoconazole modifies the discriminative stimulus and antinociceptive effects of mitragynine in rats. Ketoconazole [30 mg/kg, oral gavage (o.g.)] increased systemic exposure to mitragynine (13.3 mg/kg, o.g.) by 120% and 7-hydroxymitragynine exposure by 130%. The unexpected increase in exposure to 7-hydroxymitragynine suggested that ketoconazole inhibits metabolism of both mitragynine and 7-hydroxymitragynine, a finding confirmed in rat liver microsomes. In rats discriminating 3.2 mg/kg morphine from vehicle under a fixed-ratio schedule of food delivery, ketoconazole pretreatment increased the potency of both mitragynine (4.7-fold) and 7-hydroxymitragynine (9.7-fold). Ketoconazole did not affect morphine's potency. Ketoconazole increased the antinociceptive potency of 7-hydroxymitragynine by 4.1-fold. Mitragynine (up to 56 mg/kg, i.p.) lacked antinociceptive effects both in the presence and absence of ketoconazole. These results suggest that both mitragynine and 7-hydroxymitragynine are cleared via CYP3A and that 7-hydroxymitragynine is formed as a metabolite of mitragynine by other routes. These results have implications for kratom use in combination with numerous medications and citrus juices that inhibit CYP3A. SIGNIFICANCE STATEMENT: Mitragynine is an abundant kratom alkaloid that exhibits low efficacy at the µ-opioid receptor (MOR). Its metabolite, 7-hydroxymitragynine, is also an MOR agonist but with higher affinity and efficacy than mitragynine. Our results in rats demonstrate that cytochrome P450 3A (CYP3A) inhibition can increase the systematic exposure of both mitragynine and 7-hydroxymitragynine and their potency to produce MOR-mediated behavioral effects. These data highlight potential interactions between kratom and CYP3A inhibitors, which include numerous medications and citrus juices.

Improving access to mental health care: a system dynamics model of direct access to specialist care and accelerated specialist service capacity growth.

OBJECTIVE: To simulate the impact on population mental health indicators of allowing people to book some Medicare-subsidised sessions with psychologists and other mental health care professionals without a referral (direct access), and of increasing the annual growth rate in specialist mental health care capacity (consultations). DESIGN: System dynamics model, calibrated using historical time series data from the Australian Bureau of Statistics, HealthStats NSW, the Australian Institute of Health and Welfare, and the Australian Early Development Census. Parameter values that could not be derived from these sources were estimated by constrained optimisation. SETTING: New South Wales, 1 September 2021 - 1 September 2028. MAIN OUTCOME MEASURES: Projected mental health-related emergency department presentations, hospitalisations following self-harm, and deaths by suicide, both overall and for people aged 15-24 years. RESULTS: Direct access (for 10-50% of people requiring specialist mental health care) would lead to increases in the numbers of mental health-related emergency department presentations (0.33-1.68% of baseline), hospitalisations with self-harm (0.16-0.77%), and deaths by suicide (0.19-0.90%), as waiting times for consultations would increase, leading to disengagement and consequently to increases in adverse outcomes. Increasing the annual rate of growth of mental health service capacity (two- to fivefold) would reduce the frequency of all three outcomes; combining direct access to a proportion of services with increased growth in capacity achieved substantially greater gains than an increase in service capacity alone. A fivefold increase in the annual service growth rate would increase capacity by 71.6% by the end of 2028, compared with current projections; combined with direct access to 50% of mental health consultations, 26 616 emergency department presentations (3.6%), 1199 hospitalisations following self-harm (1.9%), and 158 deaths by suicide (2.1%) could be averted. CONCLUSION: The optimal combination of increased service capacity growth (fivefold) and direct access (50% of consultations) would have double the impact over seven years of accelerated capacity growth alone. Our model highlights the risks of implementing individual reforms without knowledge of their overall system effect.

Predictive modelling of deliberate self-harm and suicide attempts in young people accessing primary care: a machine learning analysis of a longitudinal study.

PURPOSE: Machine learning (ML) has shown promise in modelling future self-harm but is yet to be applied to key questions facing clinical services. In a cohort of young people accessing primary mental health care, this study aimed to establish (1) the performance of models predicting deliberate self-harm (DSH) compared to suicide attempt (SA), (2) the performance of models predicting new-onset or repeat behaviour, and (3) the relative importance of factors predicting these outcomes. METHODS: 802 young people aged 12-25 years attending primary mental health services had detailed social and clinical assessments at baseline and 509 completed 12-month follow-up. Four ML algorithms, as well as logistic regression, were applied to build four distinct models. RESULTS: The mean performance of models predicting SA (AUC: 0.82) performed better than the models predicting DSH (AUC: 0.72), with mean positive predictive values (PPV) approximately twice that of the prevalence (SA prevalence 14%, PPV: 0.32, DSH prevalence 22%, PPV: 0.40). All ML models outperformed standard logistic regression. The most frequently selected variable in both models was a history of DSH via cutting. CONCLUSION: History of DSH and clinical symptoms of common mental disorders, rather than social and demographic factors, were the most important variables in modelling future behaviour. The performance of models predicting outcomes in key sub-cohorts, those with new-onset or repetition of DSH or SA during follow-up, was poor. These findings may indicate that the performance of models of future DSH or SA may depend on knowledge of the individual's recent history of either behaviour.

Interactive Effects of µ-Opioid and Adrenergic-α 2 Receptor Agonists in Rats: Pharmacological Investigation of the Primary Kratom Alkaloid Mitragynine and Its Metabolite 7-Hydroxymitragynine.

The primary kratom alkaloid mitragynine is proposed to act through multiple mechanisms, including actions at µ-opioid receptors (MORs) and adrenergic-α 2 receptors (Aα 2Rs), as well as conversion in vivo to a MOR agonist metabolite (i.e., 7-hydroxymitragynine). Aα 2R and MOR agonists can produce antinociceptive synergism. Here, contributions of both receptors to produce mitragynine-related effects were assessed by measuring receptor binding in cell membranes and, in rats, pharmacological behavioral effect antagonism studies. Mitragynine displayed binding affinity at both receptors, whereas 7-hydroxymitragynine only displayed MOR binding affinity. Compounds were tested for their capacity to decrease food-maintained responding and rectal temperature and to produce antinociception in a hotplate test. Prototypical MOR agonists and 7-hydroxymitragynine, but not mitragynine, produced antinociception. MOR agonist and 7-hydroxymitragynine rate-deceasing and antinociceptive effects were antagonized by the opioid antagonist naltrexone but not by the Aα 2R antagonist yohimbine. Hypothermia only resulted from reference Aα 2R agonists. The rate-deceasing and hypothermic effects of reference Aα 2R agonists were antagonized by yohimbine but not naltrexone. Neither naltrexone nor yohimbine antagonized the rate-decreasing effects of mitragynine. Mitragynine and 7-hydroxymitragynine increased the potency of the antinociceptive effects of Aα 2R but not MOR reference agonists. Only mitragynine produced hypothermic effects. Isobolographic analyses for the rate-decreasing effects of the reference Aα 2R and MOR agonists were also conducted. These results suggest mitragynine and 7-hydroxymitragynine may produce antinociceptive synergism with Aα 2R and MOR agonists. When combined with Aα 2R agonists, mitragynine could also produce hypothermic synergism. SIGNIFICANCE STATEMENT: Mitragynine is proposed to target the µ-opioid receptor (MOR) and adrenergic-α2 receptor (Aα2R) and to produce behavioral effects through conversion to its MOR agonist metabolite 7-hydroxymitragynine. Isobolographic analyses indicated supra-additivity in some dose ratio combinations. This study suggests mitragynine and 7-hydroxymitragynine may produce antinociceptive synergism with Aα2R and MOR agonists. When combined with Aα2R agonists, mitragynine could also produce hypothermic synergism.

Optimizing Strategies for Improving Mental Health in Victoria, Australia during the COVID-19 Era: A System Dynamics Modelling Study.

The ongoing COVID-19 pandemic has impacted the mental health of populations and highlighted the limitations of mental health care systems. As the trajectory of the pandemic and the economic recovery are still uncertain, decision tools are needed to help evaluate the best interventions to improve mental health outcomes. We developed a system dynamics model that captures causal relationships among population, demographics, post-secondary education, health services, COVID-19 impact, and mental health outcomes. The study was conducted in the Australian state of Victoria. The model was calibrated using historical data and was stratified by age group and by geographic remoteness. Findings demonstrate that the most effective intervention combination includes economic, social, and health sector initiatives. Assertive post-suicide attempt care is the most impactful health sector intervention, but delaying implementation reduces the potency of its impact. Some evidence-based interventions, such as population-wide community awareness campaigns, are projected to worsen mental health outcomes when implemented on their own. Systems modelling offers a powerful decision-support tool to test alternative strategies for improving mental health outcomes in the Victorian context.

Dynamic networks of psychological symptoms, impairment, substance use, and social support: The evolution of psychopathology among emerging adults.

BACKGROUND: Subthreshold/attenuated syndromes are established precursors of full-threshold mood and psychotic disorders. Less is known about the individual symptoms that may precede the development of subthreshold syndromes and associated social/functional outcomes among emerging adults. METHODS: We modeled two dynamic Bayesian networks (DBN) to investigate associations among self-rated phenomenology and personal/lifestyle factors (role impairment, low social support, and alcohol and substance use) across the 19Up and 25Up waves of the Brisbane Longitudinal Twin Study. We examined whether symptoms and personal/lifestyle factors at 19Up were associated with (a) themselves or different items at 25Up, and (b) onset of a depression-like, hypo-manic-like, or psychotic-like subthreshold syndrome (STS) at 25Up. RESULTS: The first DBN identified 11 items that when endorsed at 19Up were more likely to be reendorsed at 25Up (e.g., hypersomnia, impaired concentration, impaired sleep quality) and seven items that when endorsed at 19Up were associated with different items being endorsed at 25Up (e.g., earlier fatigue and later role impairment; earlier anergia and later somatic pain). In the second DBN, no arcs met our a priori threshold for inclusion. In an exploratory model with no threshold, >20 items at 19Up were associated with progression to an STS at 25Up (with lower statistical confidence); the top five arcs were: feeling threatened by others and a later psychotic-like STS; increased activity and a later hypo-manic-like STS; and anergia, impaired sleep quality, and/or hypersomnia and a later depression-like STS. CONCLUSIONS: These probabilistic models identify symptoms and personal/lifestyle factors that might prove useful targets for indicated preventative strategies.

Hepatocellular carcinoma organoid co-cultures mimic angiocrine crosstalk to generate inflammatory tumor microenvironment.

Hepatocellular carcinoma (HCC) is the sixth most common cancer and the second leading cause of cancer worldwide. Despite approvals of several therapeutics to treat advanced HCC in the past few years, the impact of anti-angiogenic treatment on HCC patient overall survival remains limited. This suggests there may be alternative, perfusion-independent roles of endothelial cells that support tumor progression. Thus, we leveraged a well-defined hydrogel system to establish co-culture models to mimic and characterize the angiocrine crosstalk between HCC and endothelial cells in vitro. Co-cultures of HCC cell lines or patient-derived xenograft organoids with endothelial cells exhibited the upregulation of MCP-1, IL-8 and CXCL16, suggesting that the HCC-endothelial interactions established in our models recapitulate known angiocrine signaling. Additionally, by subjecting co-cultures and mono-cultures to RNA sequencing, transcriptomic analysis revealed an upregulation in the expression of genes associated with tumor necrosis factor (TNF) signaling, such as that of chemokines, suggesting that endothelial cells induce HCC cells to generate an inflammatory microenvironment by recruiting immune cells. Finally, HCC-endothelial angiocrine crosstalk in the co-culture models polarized macrophages towards a pro-inflammatory and pro-angiogenic phenotype, paralleling a tumor-associated macrophage subset previously reported in HCC. Together, these findings suggest that these HCC-endothelial co-culture models may serve as important models to understand and target the interplay between angiogenesis and the immune milieu.

Premature mortality in early-intervention mental health services: a data linkage study protocol to examine mortality and morbidity outcomes in a cohort of help-seeking young people.

INTRODUCTION: Understanding the risk of premature death from suicide, accident and injury and other physical health conditions in people seeking healthcare for mental disorders is essential for delivering targeted clinical interventions and secondary prevention strategies. It is not clear whether morbidity and mortality outcomes in hospital-based adult cohorts are applicable to young people presenting to early-intervention services. METHODS AND ANALYSIS: The current data linkage project will establish the Brain and Mind Patient Research Register-Mortality and Morbidity (BPRR-M&M) database. The existing Brain and Mind Research Institute Patient Research Register (BPRR) is a cohort of 6743 young people who have accessed primary care-based early-intervention services; subsets of the BPRR contain rich longitudinal clinical, neurobiological, social and functional data. The BPRR will be linked with the routinely collected health data from emergency department (ED), hospital admission and mortality databases in New South Wales from January 2010 to November 2020. Mortality will be the primary outcome of interest, while hospital presentations will be a secondary outcome. The established BPRR-M&M database will be used to establish mortality rates and rates of ED presentations and hospital admissions. Survival analysis will determine how time to death or hospital presentation varies by identified social, demographic and clinical variables. Bayesian modelling will be used to identify predictors of these morbidity and mortality outcomes. ETHICS AND DISSEMINATION: The study has been reviewed and approved by the human research ethics committee of the Sydney Local Health District (2019/ETH00469). All data will be non-identifiable, and research findings will be disseminated through peer-reviewed journals and scientific conference presentations.

Predicting the emergence of full-threshold bipolar I, bipolar II and psychotic disorders in young people presenting to early intervention mental health services.

BACKGROUND: Predictors of new-onset bipolar disorder (BD) or psychotic disorder (PD) have been proposed on the basis of retrospective or prospective studies of 'at-risk' cohorts. Few studies have compared concurrently or longitudinally factors associated with the onset of BD or PDs in youth presenting to early intervention services. We aimed to identify clinical predictors of the onset of full-threshold (FT) BD or PD in this population. METHOD: Multi-state Markov modelling was used to assess the relationships between baseline characteristics and the likelihood of the onset of FT BD or PD in youth (aged 12-30) presenting to mental health services. RESULTS: Of 2330 individuals assessed longitudinally, 4.3% (n = 100) met criteria for new-onset FT BD and 2.2% (n = 51) met criteria for a new-onset FT PD. The emergence of FT BD was associated with older age, lower social and occupational functioning, mania-like experiences (MLE), suicide attempts, reduced incidence of physical illness, childhood-onset depression, and childhood-onset anxiety. The emergence of a PD was associated with older age, male sex, psychosis-like experiences (PLE), suicide attempts, stimulant use, and childhood-onset depression. CONCLUSIONS: Identifying risk factors for the onset of either BD or PDs in young people presenting to early intervention services is assisted not only by the increased focus on MLE and PLE, but also by recognising the predictive significance of poorer social function, childhood-onset anxiety and mood disorders, and suicide attempts prior to the time of entry to services. Secondary prevention may be enhanced by greater attention to those risk factors that are modifiable or shared by both illness trajectories.

The Lack of Contribution of 7-Hydroxymitragynine to the Antinociceptive Effects of Mitragynine in Mice: A Pharmacokinetic and Pharmacodynamic Study.

Kratom (Mitragyna speciosa), a Southeast Asian tree, has been used for centuries in pain relief and mitigation of opium withdrawal symptoms. Mitragynine (MTG), the major kratom alkaloid, is being investigated for its potential to provide analgesia without the deleterious effects associated with typical opioids. Concerns have been raised regarding the active metabolite of MTG, 7-hydroxymitragynine (7HMG), which has higher affinity and efficacy at µ-opioid receptors than MTG. Here we investigated the hotplate antinociception, pharmacokinetics, and tissue distribution of MTG and 7HMG at equianalgesic oral doses in male and female C57BL/6 mice to determine the extent to which 7HMG metabolized from MTG accounts for the antinociceptive effects of MTG and investigate any sex differences. The mechanism of action was examined by performing studies with the opioid receptor antagonist naltrexone. A population pharmacokinetic/pharmacodynamic model was developed to predict the behavioral effects after administration of various doses of MTG and 7HMG. When administered alone, 7HMG was 2.8-fold more potent than MTG to produce antinociception. At equivalent effective doses of MTG and 7HMG, there was a marked difference in the maximum brain concentration of 7HMG achieved, i.e., 11-fold lower as a metabolite of MTG. The brain concentration of 7HMG observed 4 hours post administration, producing an analgesic effect <10%, was still 1.5-fold higher than the maximum concentration of 7HMG as a metabolite of MTG. These results provide strong evidence that 7HMG has a negligible role in the antinociceptive effects of MTG in mice. SIGNIFICANCE STATEMENT: Mitragynine (MTG) is being investigated for its potential to aid in pain relief, opioid withdrawal syndrome, and opioid use disorder. The active metabolite of MTG, 7-hydroxymitragynine (7HMG), has been shown to have abuse potential and has been implicated in the opioid-like analgesic effect after MTG administration. The results of this study suggest a lack of involvement of 7HMG in the antinociceptive effects of MTG in mice.

Identifying pathways to early-onset metabolic dysfunction, insulin resistance and inflammation in young adult inpatients with emerging affective and major mood disorders.

AIM: Young people with common mood disorders face the prospect of shortened life expectancy largely due to premature cardiovascular disease. Metabolic dysfunction is a risk factor for premature cardiovascular disease. There is an ongoing debate whether metabolic dysfunction can be simply explained by weight gain secondary to psychotropic medications or whether shared genetic vulnerability, intrinsic immune-metabolic disturbances or other system perturbations (e.g. dysregulated sympathetic nervous system, circadian dysfunction) are more relevant determinants of premature cardiovascular disease. Thus, we aimed to investigate underlying drivers of metabolic dysfunction and premature cardiovascular disease in young people in the early phases of common mood disorders. METHODS: We evaluated the relationships between insulin resistance (assessed by HOMA2-IR) and body mass index (BMI), sex, diagnosis, medication, inflammatory markers and hormonal factors in 327 inpatients with emerging affective and major mood disorders admitted to the Young Adult Mental Health Unit, St Vincent's Private Hospital, Sydney. RESULTS: While HOMA2-IR scores were positively associated with BMI (rs  = 0.465, p < .001), they were also higher in those prescribed mood stabilizers (p = .044) but were not associated with specific diagnoses, other medication types or the number of prescribed medications. Further, high-sensitivity C-reactive protein levels (but not thyroid-stimulating hormone and ferritin levels) were positively associated with HOMA2-IR (rs  = 0. 272, p < .001) and BMI (rs  = . 409, p < .001). CONCLUSIONS: In addition to BMI, other non-specific markers of inflammation are associated with early metabolic dysfunction in young people with emerging affective and major mood disorders.

Which Social, Economic, and Health Sector Strategies Will Deliver the Greatest Impacts for Youth Mental Health and Suicide Prevention? Protocol for an Advanced, Systems Modelling Approach.

Background: Current global challenges are generating extensive social disruption and uncertainty that have the potential to undermine the mental health, wellbeing, and futures of young people. The scale and complexity of challenges call for engagement with systems science-based decision analytic tools that can capture the dynamics and interrelationships between physical, social, economic, and health systems, and support effective national and regional responses. At the outset of the pandemic mental health-related systems models were developed for the Australian context, however, the extent to which findings are generalisable across diverse regions remains unknown. This study aims to explore the context dependency of systems modelling insights. Methods: This study will employ a comparative case study design, applying participatory system dynamics modelling across eight diverse regions of Australia to answer three primary research questions: (i) Will current regional differences in key youth mental health outcomes be exacerbated in forward projections due to the social and economic impacts of COVID-19?; (ii) What combination of social policies and health system strengthening initiatives will deliver the greatest impacts within each region?; (iii) To what extent are optimal strategic responses consistent across the diverse regions? We provide a detailed technical blueprint as a potential springboard for more timely construction and deployment of systems models in international contexts to facilitate a broader examination of the question of generalisability and inform investments in the mental health and wellbeing of young people in the post COVID-19 recovery. Discussion: Computer simulation is known as the third pillar of science (after theory and experiment). Simulation allows researchers and decision makers to move beyond what can be manipulated within the scale, time, and ethical limits of the experimental approach. Such learning when achieved collectively, has the potential to enhance regional self-determination, help move beyond incremental adjustments to the status quo, and catalyze transformational change. This research seeks to advance efforts to establish regional decision support infrastructure and empower communities to effectively respond. In addition, this research seeks to move towards an understanding of the extent to which systems modelling insights may be relevant to the global mental health response by encouraging researchers to use, challenge, and advance the existing work for scientific and societal progress.

Accessible detection of SARS-CoV-2 through molecular nanostructures and automated microfluidics.

Accurate and accessible nucleic acid diagnostics is critical to reducing the spread of COVID-19 and resuming socioeconomic activities. Here, we present an integrated platform for the direct detection of SARS-CoV-2 RNA targets near patients. Termed electrochemical system integrating reconfigurable enzyme-DNA nanostructures (eSIREN), the technology leverages responsive molecular nanostructures and automated microfluidics to seamlessly transduce target-induced molecular activation into an enhanced electrochemical signal. Through responsive enzyme-DNA nanostructures, the technology establishes a molecular circuitry that directly recognizes specific RNA targets and catalytically enhances signaling; only upon target hybridization, the molecular nanostructures activate to liberate strong enzymatic activity and initiate cascading reactions. Through automated microfluidics, the system coordinates and interfaces the molecular circuitry with embedded electronics; its pressure actuation and liquid-guiding structures improve not only analytical performance but also automated implementation. The developed platform establishes a detection limit of 7 copies of RNA target per µl, operates against the complex biological background of native patient samples, and is completed in <20 min at room temperature. When clinically evaluated, the technology demonstrates accurate detection in extracted RNA samples and direct swab lysates to diagnose COVID-19 patients.

Collaborative Equilibrium Coupling of Catalytic DNA Nanostructures Enables Programmable Detection of SARS-CoV-2.

Accessible and adaptable nucleic acid diagnostics remains a critical challenge in managing the evolving COVID-19 pandemic. Here, an integrated molecular nanotechnology that enables direct and programmable detection of SARS-CoV-2 RNA targets in native patient specimens is reported. Termed synergistic coupling of responsive equilibrium in enzymatic network (SCREEN), the technology leverages tunable, catalytic molecular nanostructures to establish an interconnected, collaborative architecture. SCREEN mimics the extraordinary organization and functionality of cellular signaling cascades. Through programmable enzyme-DNA nanostructures, SCREEN activates upon interaction with different RNA targets to initiate multi-enzyme catalysis; through system-wide favorable equilibrium shifting, SCREEN directly transduces a single target binding into an amplified electrical signal. To establish collaborative equilibrium coupling in the architecture, a computational model that simulates all reactions to predict overall performance and optimize assay configuration is developed. The developed platform achieves direct and sensitive RNA detection (approaching single-copy detection), fast response (assay reaction is completed within 30 min at room temperature), and robust programmability (across different genetic loci of SARS-CoV-2). When clinically evaluated, the technology demonstrates robust and direct detection in clinical swab lysates to accurately diagnose COVID-19 patients.

The use of hypercapnic conditions to assess opioid-induced respiratory depression in rats.

INTRODUCTION: Whole-body plethysmography (WBP) in unrestrained, non-anesthetized rodents is a preclinical method to assess the respiratory depressant effects of opioids, the leading cause of opioid overdose death in humans. However, low baseline respiration rates under normocapnic conditions (i.e., "floor" effect) can render the measurement of respiratory decreases challenging. We assessed hypercapnia-induced increases in respiration as a strategy to assess opioid-induced decreases in respiration in rats. METHODS: WBP was used to assess respiration frequency, tidal volume and minute volume in the presence of normocapnic and hypercapnic (8% CO2) conditions in rats during the rat diurnal period of the light cycle. The mu-opioid receptor agonist fentanyl was administered intravenously, and the hot plate test was used to assess acute antinociception. RESULTS AND DISCUSSION: Hypercapnia-induced increases in respiratory parameters (frequency, minute volume, and tidal volume) were decreased by fentanyl at doses that did not decrease the same parameters under the normocapnic conditions. These findings show that hypercapnia increases sensitivity to respiratory depressant effects of fentanyl, as compared with assessments during the rat diurnal period when activity and breathing rate are generally low, i.e., there is a floor effect. The current approach is highly sensitive to opioid-induced respiratory depression, and therefore provides a useful method for assessment in a pre-clinical setting.