Onset of Alzheimer disease in apolipoprotein ɛ4 carriers is earlier in butyrylcholinesterase K variant carriers.

BACKGROUND: The authors sought to examine the impact of the K-variant of butyrylcholinesterase (BCHE-K) carrier status on age-at-diagnosis of Alzheimer disease (AD) in APOE4 carriers. METHODS: Patients aged 50-74 years with cerebrospinal fluid (CSF) biomarker-confirmed AD, were recruited to clinical trial (NCT03186989 since June 14, 2017). Baseline demographics, disease characteristics, and biomarkers were evaluated in 45 patients according to BCHE-K and APOE4 allelic status in this post-hoc study. RESULTS: In APOE4 carriers (N = 33), the mean age-at-diagnosis of AD in BCHE-K carriers (n = 11) was 6.4 years earlier than in BCHE-K noncarriers (n = 22, P < .001, ANOVA). In APOE4 noncarriers (N = 12) there was no observed influence of BCHE-K. APOE4 carriers with BCHE-K also exhibited slightly higher amyloid and tau accumulations compared to BCHE-K noncarriers. A predominantly amyloid, limited tau, and limbic-amnestic phenotype was exemplified by APOE4 homozygotes with BCHE-K. In the overall population, multiple regression analyses demonstrated an association of amyloid accumulation with APOE4 carrier status (P < .029), larger total brain ventricle volume (P < .021), less synaptic injury (Ng, P < .001), and less tau pathophysiology (p-tau181, P < .005). In contrast, tau pathophysiology was associated with more neuroaxonal damage (NfL, P = .002), more synaptic injury (Ng, P < .001), and higher levels of glial activation (YKL-40, P = .01). CONCLUSION: These findings have implications for the genetic architecture of prognosis in early AD, not the genetics of susceptibility to AD. In patients with early AD aged less than 75 years, the mean age-at-diagnosis of AD in APOE4 carriers was reduced by over 6 years in BCHE-K carriers versus noncarriers. The functional status of glia may explain many of the effects of APOE4 and BCHE-K on the early AD phenotype. TRIAL REGISTRATION: NCT03186989 since June 14, 2017.

Exploratory Tau Biomarker Results From a Multiple Ascending-Dose Study of BIIB080 in Alzheimer Disease: A Randomized Clinical Trial.

Importance: Accumulation of hyperphosphorylated, tangled microtubule-associated protein tau (MAPT) is a pathological hallmark of Alzheimer disease (AD) associated with disease progression and cognitive decline. Objective: To evaluate the effect of tau synthesis reduction on tau biomarkers in patients with mild AD. Design, Setting, and Participants: This randomized clinical trial was a double-blind, placebo-controlled 36-week multiple-ascending dose (MAD) phase 1b trial (October 2017 to September 2020), followed by a 64- or 71-week open-label long-term extension (LTE) (October 2019 to May 2022). After being assessed for eligibility at 12 sites in Canada and Europe, participants with mild AD and confirmed amyloid pathology were randomized 3:1 (BIIB080:placebo) in 4 dose cohorts. Intervention: Intrathecal administration of BIIB080, a MAPT-targeting antisense oligonucleotide, or placebo. Active dose arms included 10 mg every 4 weeks, 30 mg every 4 weeks, 60 mg every 4 weeks, and 115 mg every 12 weeks during the MAD period and 60 mg every 12 weeks or 115 mg every 12 weeks during the LTE. Main Outcome and Measures: The original primary end point was safety. Additionally, BIIB080, total tau (t-tau), and phosphorylated tau 181 (p-tau181) cerebrospinal fluid (CSF) concentrations were evaluated. Tau positron emission tomography (PET) was collected in a substudy, and standard uptake value ratios (SUVRs) were calculated in a priori-defined composite regions of interest. Results: Of 102 participants assessed for eligibility, 46 participants with mild AD were enrolled; 23 (50%) were female, and mean (SD) age was 65.8 (5.70) years. BIIB080 was generally well tolerated and was associated with a dose-dependent reduction in CSF t-tau and p-tau181 in the MAD period (56% reduction; 95% CI, 50% to 62%; and 51% reduction; 95% CI, 38% to 63%, of CSF t-tau in the 2 higher-dose cohorts) that continued and/or was maintained through quarterly dosing in the LTE. Tau PET demonstrated reduced accumulation vs placebo at week 25 (n = 13). At week 100, tau PET showed a reduction from baseline across all regions assessed (n = 12), with the largest reductions from baseline observed in the temporal composite (-0.71 SUVR; 95% CI, -1.40 to -0.02). A moderate correlation was observed between model-predicted cumulative CSF drug exposure and tau PET change. Conclusions and Relevance: In this randomized clinical trial, BIIB080 reduced tau biomarkers, including CSF t-tau, CSF p-tau181, and tau PET, which is associated with cognitive decline, in participants with mild AD. Effects of BIIB080 on biomarkers and clinical outcomes are being further evaluated in a phase 2 trial. Trial Registration: ClinicalTrials.gov Identifier: NCT03186989.

A series of diabetic ketoacidosis associated with the use of sodium-glucose co-transporter-2 inhibitors in secondary care.

BACKGROUND AND AIMS: Sodium-glucose co-transporter-2 inhibitors (SGLT-2i) are associated with diabetic ketoacidosis (DKA), however limited case series are published. METHODS: We evaluated the characteristics of patients admitted with SGLT-2i associated DKA. RESULTS: Over 4 months, 22 patients were identified; 45.5% of DKA was not associated with concurrent illness. CONCLUSION: DKA is not uncommonly associated with SGLT2i with no clear patient factors associated with severity.

A series of patients with hospital-acquired diabetic ketoacidosis (HADKA): a descriptive analysis.

BACKGROUND: Hospital-acquired diabetic ketoacidosis (HADKA) can complicate hospital admission in people with type 1 diabetes (T1D) and type 2 diabetes (T2D). We aimed to determine the characteristics of such patients and the reasons for HADKA. METHODS: A retrospective analysis of patients referred to diabetes services with HADKA at Morriston Hospital between January 2016 and January 2022 was undertaken. Patients that were included were admitted without diabetic ketoacidosis (DKA), were aged 18 years and over, and who subsequently developed DKA in hospital. RESULTS: Twenty-five patients were included with a mean age of 65.2 years; nine (32.0%) were men, 13 (52.0%) had T1D and 12 (48.0%) had T2D. Patients had a mean pre-admission glycated haemoglobin of 84.7 mmol/mol, and 17 (68.0%) were insulin-treated. Most were admitted under medicine (n=14; 56.0%) and the remainder under surgery (n=11; 44.0%). More common reasons for HADKA were erroneous insulin administration (n=9; 36.0%), infection (n=7; 28.0%) and surgery (n=5; 20.0%).Five (20.0%) patients required intensive care admission, and the mean length of hospital stay was 42.6 days (range 2-173). Three (12.0%) patients died during the hospital admission. CONCLUSION: HADKA was identified in a significant number of patients at our hospital and was associated with significant mortality. Earlier recognition of ketonaemia and associated medication use may prevent HADKA and improve outcomes.

Lockdown babies: Birth and new parenting experiences during the 2020 Covid-19 lockdown in South Africa, a cross-sectional study.

BACKGROUND: Perceived birth experiences of parents can have a lasting impact on children. We explored the birth and new parenting experiences of South African parents in 2020 during the Covid-19 lockdown. METHODS: We conducted a cross-sectional online survey with consenting parents of babies born in South Africa during 2020. Factors associated with negative birth emotions and probable depression were estimated using logistic regression. RESULTS: Most of the 520 respondents were females (n = 496, 95%) who gave birth at private hospitals (n = 426, 86%). Mothers reported having overall positive birth emotions (n = 399, 80%). Multivariable analysis showed that having a preterm baby (aOR 2.89; CI 1.51-5.53) and the mother self-reporting that Covid-19 affected her birth experience (aOR 4.25; CI 2.08-8.68) increased the odds of mothers reporting predominantly negative emotions about their birth. The mother having her preferred delivery method reduced the odds of having negative birth emotions (aOR 0.41; CI 0.25-0.66). Multivariable analysis showed that having predominantly negative emotions about the birth increased the odds of probable minor depression (aOR 3.60; CI 1.93-6.70). Being older reduced the odds of having probable minor depression (25-34 years aOR 0.36; CI 0.10-1.32; 35 years or older aOR 0.25; CI 0.06-0.91). CONCLUSIONS: Lockdown exacerbated many birth and parenting challenges including mental health and health care access. However, overall experiences were positive and there was a strong sense of resilience amongst parents.

Automated Audiometry in Quiet and Simulated Exam Room Noise for Listeners with Normal Hearing and Impaired Hearing.

BACKGROUND: Up to 80% of audiograms could be automated which would allow more time for provision of specialty services. Ideally, automated audiometers would provide accurate results for listeners with impaired hearing as well as normal hearing. Additionally, accurate results should be provided both in controlled environments like a sound-attenuating room but also in test environments that may support greater application when sound-attenuating rooms are unavailable. Otokiosk is an iOS-based system that has been available for clinical use, but there are not yet any published validation studies using this product. PURPOSE: The purpose of this project was to complete a validation study on the OtoKiosk automated audiometry system in quiet and in low-level noise, for listeners with normal hearing and for listeners with impaired hearing. RESEARCH DESIGN: Pure tone air conduction thresholds were obtained for each participant for three randomized conditions: standard audiometry, automated testing in quiet, and automated testing in noise. Noise, when present, was 35 dBA overall and was designed to emulate an empty medical exam room. STUDY SAMPLE: Participants consisted of 11 adults with hearing loss and 15 adults with normal hearing recruited from the local area. DATA COLLECTION AND ANALYSIS: Thresholds were measured at 500, 1,000, 2,000, and 4,000 Hz using the Otokiosk system that incorporates a modified Hughson-Westlake method. Results were analyzed using descriptive statistics and also by a linear mixed-effects model to compare thresholds obtained in each condition. RESULTS: Across condition and participant group 73.6% of thresholds measured with OtoKiosk were within ± 5 dB of the conventionally measured thresholds; 92.8% were within ± 10 dB. On average, differences between tests were small. Pairwise comparisons revealed thresholds were ∼3.5-4 dB better with conventional audiometry than with the mobile application in quiet and in noise. Noise did not affect thresholds measured with OtoKiosk. CONCLUSIONS: The OtoKiosk automated hearing test measured pure tone air conduction thresholds from 500 to 4,000 Hz at slightly higher thresholds than conventional audiometry, but less than the smallest typical 5 dB clinical step-size. Our results suggest OtoKiosk is a reasonable solution for sound booths and exam rooms with low-level background noise.

Dialectical behaviour therapy 'Skills for Living' for young people leaving care: A qualitative exploration of participants' experiences.

It is acknowledged that care leavers experience an accelerated transition into adulthood, despite often having complex psychosocial needs with limited support networks. The 'Skills for Living' programme was designed to improve the psychological wellbeing of care leavers and offers an adapted Dialectical Behaviour Therapy skills group as its primary intervention. This paper provides a qualitative evaluation of the programme. Semi-structured interviews were undertaken with 10 participants, and the data were analysed using thematic analysis (Braun & Clarke, 2006). Four key themes emerged: 'Initial Apprehension and Reluctance to Participate', 'Connection, Understanding and Validation', 'Confidence with Social Skills', and 'Emotional Acceptance and Self-Soothing'. Clinical implications and recommendations are discussed.

Rehabilitation of Concussion and Persistent Postconcussive Symptoms.

Concussion and persistent postconcussive symptoms (PPCS) are encountered by clinicians in sports medicine, pediatrics, neurology, physiatry, emergency medicine, and primary care. Clinical management may require a multidisciplinary approach. This article presents a structured method for the diagnosis of concussion and PPCS in the outpatient setting, which includes a history, physical examination, and additional tests as clinically indicated to help identify underlying symptom generators. Treatment for concussion and PPCS should be individualized, based on predominant signs and symptoms, and can include subsymptom threshold aerobic exercise, cervical physical therapy, vestibulo-ocular rehabilitation, behavioral and cognitive psychotherapy, and some symptom-specific pharmacological therapies.

Pressure Injury Prevention in Patients with Prolonged ED Stays Prior to Admission.

PURPOSE: The purpose of this project was to examine whether initiating a standardized pressure injury (PI) assessment and prevention protocol early in adult patients' ED stay reduces hospital-acquired PIs (HAPIs) in those patients admitted from the ED to acute care inpatient medical units. METHODS: A nurse-led evidence-based practice team studied the problem of increasing HAPIs on four acute care inpatient units and found that, among patients who had been admitted to inpatient care from the ED, longer ED boarding times correlated with a higher rate of HAPIs. ED staff and acute care unit nurses collaborated to develop new protocols to prevent HAPIs in the ED, including staff education and standardized assessments and prevention care for at-risk patients. Data collection was performed at three time periods over approximately two and a half years: baseline, intervention, and postintervention. RESULTS: The incidence rate for HAPIs decreased from 3.56 per 1,000 patient-days at baseline to 1.31 per 1,000 patient-days during the intervention period. This reduction was sustained over the next five months, during which the HAPI incidence rate was 1.53 per 1,000 patient-days. IMPLICATIONS: At a time when ED length of stay is difficult to manage and continues to increase, the use of evidence-based interventions and protocols can reduce the rate of PIs in high-risk patients waiting for hospital admission, leading to a reduction in PI rates and overall hospital costs.

Quantitative Measurements of LRRK2 in Human Cerebrospinal Fluid Demonstrates Increased Levels in G2019S Patients.

Leucine-rich repeat kinase 2 (LRRK2) mutations are among the most significant genetic risk factors for developing late onset Parkinson's disease (PD). To understand whether a therapeutic can modulate LRRK2 levels as a potential disease modifying strategy, it is important to have methods in place to measure the protein with high sensitivity and specificity. To date, LRRK2 measurements in cerebrospinal fluid (CSF) have used extracellular vesicle enrichment via differential ultracentrifugation and western blot detection. Our goal was to develop a methodology which could be deployed in a clinical trial, therefore throughput, robustness and sensitivity were critical. To this end, we developed a Stable Isotope Standard Capture by Anti-peptide Antibody (SISCAPA) assay which is capable of detecting LRRK2 from 1 ml of human CSF. The assay uses a commercially available LRRK2 monoclonal antibody (N241A/34) and does not require extracellular vesicle enrichment steps. The assay includes stable isotope peptide addition which allows for absolute quantitation of LRRK2 protein. We determined that the assay performed adequately for CSF measurements and that blood contamination from traumatic lumbar puncture does not pose a serious analytical challenge. We then applied this technique to 106 CSF samples from the MJFF LRRK2 Cohort Consortium which includes healthy controls, sporadic PD patients and LRRK2 mutation carriers with and without PD. Of the 105 samples that had detectable LRRK2 signal, we found that the PD group with the G2019S LRRK2 mutation had significantly higher CSF LRRK2 levels compared to all other groups. We also found that CSF LRRK2 increased with the age of the participant. Taken together, this work represents a step forward in our ability to measure LRRK2 in a challenging matrix like CSF which has implications for current and future LRRK2 therapeutic clinical trials.

Native Chromatin Proteomics Reveals a Role for Specific Nucleoporins in Heterochromatin Organization and Maintenance.

Spatially and functionally distinct domains of heterochromatin and euchromatin play important roles in the maintenance of chromosome stability and regulation of gene expression, but a comprehensive knowledge of their composition is lacking. Here, we develop a strategy for the isolation of native Schizosaccharomyces pombe heterochromatin and euchromatin fragments and analyze their composition by using quantitative mass spectrometry. The shared and euchromatin-specific proteomes contain proteins involved in DNA and chromatin metabolism and in transcription, respectively. The heterochromatin-specific proteome includes all proteins with known roles in heterochromatin formation and, in addition, is enriched for subsets of nucleoporins and inner nuclear membrane (INM) proteins, which associate with different chromatin domains. While the INM proteins are required for the integrity of the nucleolus, containing ribosomal DNA repeats, the nucleoporins are required for aggregation of heterochromatic foci and epigenetic inheritance. The results provide a comprehensive picture of heterochromatin-associated proteins and suggest a role for specific nucleoporins in heterochromatin function.

Multiplexed quantitative phosphoproteomics of cell line and tissue samples.

Post-translational modifications (PTMs) of proteins increase a biological system's repertoire of regulatory tools to control cellular mechanisms. Protein phosphorylation is the most studied PTM and known to be dysregulated in many diseases, including cancer, and protein kinases are among the most important drug targets. Many proteins across the eukaryotic proteome are phosphorylated, and more than 50,000 unique protein phosphorylation sites have been identified in a single human cell line. Understanding the vast biological networks directed by protein phosphorylation requires deep quantitative mapping of the phosphoproteome across many samples. Multiplexed proteomics using isobaric labeling reagents to barcode proteome samples for simultaneous quantification has greatly increased the throughput of mass spectrometry-based proteomics and enabled the number of analyses required to understand complex biological systems. We are presenting a detailed protocol to use multiplexed proteomics for mapping phosphoproteomes in samples from cell culture experiments and in tissue samples. The protocol includes phosphopeptide enrichment with TiO2 and phosphotyrosine antibody technology. We are using tandem mass tag (TMT) reagents for barcoding the samples allowing parallel quantification of up to 11 samples. The mass spectrometry method is based on the MultiNotch MS3 method to generate quantitative data of high accuracy and reproducibility. Tandem mass spectrometry (MS2) based on regular collision-induced dissociation (CID) and higher-energy collisional dissociation (HCD) is used to maximize the number of quantified phosphopeptides. The protocol typically enables the quantification of more than 20,000 unique phosphoforms (unique patterns of peptide phosphorylations) from proteome samples of human origin requiring less than 8h of mass spectrometry time per sample.

Substrate-based kinase activity inference identifies MK2 as driver of colitis.

Inflammatory bowel disease (IBD) is a chronic and debilitating disorder that has few treatment options due to a lack of comprehensive understanding of its molecular pathogenesis. We used multiplexed mass spectrometry to collect high-content information on protein phosphorylation in two different mouse models of IBD. Because the biological function of the vast majority of phosphorylation sites remains unknown, we developed Substrate-based Kinase Activity Inference (SKAI), a methodology to infer kinase activity from phosphoproteomic data. This approach draws upon prior knowledge of kinase-substrate interactions to construct custom lists of kinases and their respective substrate sites, termed kinase-substrate sets that employ prior knowledge across organisms. This expansion as much as triples the amount of prior knowledge available. We then used these sets within the Gene Set Enrichment Analysis framework to infer kinase activity based on increased or decreased phosphorylation of its substrates in a dataset. When applied to the phosphoproteomic datasets from the two mouse models, SKAI predicted largely non-overlapping kinase activation profiles. These results suggest that chronic inflammation may arise through activation of largely divergent signaling networks. However, the one kinase inferred to be activated in both mouse models was mitogen-activated protein kinase-activated protein kinase 2 (MAPKAPK2 or MK2), a serine/threonine kinase that functions downstream of p38 stress-activated mitogen-activated protein kinase. Treatment of mice with active colitis with ATI450, an orally bioavailable small molecule inhibitor of the MK2 pathway, reduced inflammatory signaling in the colon and alleviated the clinical and histological features of inflammation. These studies establish MK2 as a therapeutic target in IBD and identify ATI450 as a potential therapy for the disease.

Evidence map of knowledge translation strategies, outcomes, facilitators and barriers in African health systems.

BACKGROUND: The need for research-based knowledge to inform health policy formulation and implementation is a chronic global concern impacting health systems functioning and impeding the provision of quality healthcare for all. This paper provides a systematic overview of the literature on knowledge translation (KT) strategies employed by health system researchers and policy-makers in African countries. METHODS: Evidence mapping methodology was adapted from the social and health sciences literature and used to generate a schema of KT strategies, outcomes, facilitators and barriers. Four reference databases were searched using defined criteria. Studies were screened and a searchable database containing 62 eligible studies was compiled using Microsoft Access. Frequency and thematic analysis were used to report study characteristics and to establish the final evidence map. Focus was placed on KT in policy formulation processes in order to better manage the diversity of available literature. RESULTS: The KT literature in African countries is widely distributed, problematically diverse and growing. Significant disparities exist between reports on KT in different countries, and there are many settings without published evidence of local KT characteristics. Commonly reported KT strategies include policy briefs, capacity-building workshops and policy dialogues. Barriers affecting researchers and policy-makers include insufficient skills and capacity to conduct KT activities, time constraints and a lack of resources. Availability of quality locally relevant research was the most reported facilitator. Limited KT outcomes reflect persisting difficulties in outcome identification and reporting. CONCLUSION: This study has identified substantial geographical gaps in knowledge and evidenced the need to boost local research capacities on KT practices in low- and middle-income countries. Evidence mapping is also shown to be a useful approach that can assist local decision-making to enhance KT in policy and practice.

Exploring the feasibility of using mobile phones to improve the management of clients with cervical cancer precursor lesions.

BACKGROUND: Cancer screening programs hold much potential for reducing the cervical cancer disease burden in developing countries. The aim of this study was to determine the feasibility of mobile health (mHealth) phone technology to improve management and follow-up of clients with cervical cancer precursor lesions. METHODS: A sequential mixed methods design was employed for this study. Quantitative data was collected using a cross-sectional survey of 364 women eligible for a Pap smear at public sector health services in Cape Town, South Africa. Information was collected on socio-demographic status; cell phone ownership and patterns of use; knowledge of cervical cancer prevention; and interest in Pap smear results and appointment reminders via SMS-text messages. Descriptive statistics, crude bivariate comparisons and logistic regression models were employed to analyze survey results. Qualitative data was collected through 10 in-depth interviews with primary health care providers and managers involved in cervical cancer screening. Four focus group discussions with 27 women attending a tertiary level colposcopy clinic were also conducted. Themes related to loss of mobile phones, privacy and confidentiality, interest in receiving SMS-text messages, text language and clinic-based management of a SMS system are discussed. Thematic analyses of qualitative data complemented quantitative findings. RESULTS: Phone ownership amongst surveyed women was 98% with phones mostly used for calls and short message service (SMS) functions. Over half (58%) of women reported loss/theft of mobile phones. Overall, there was interest in SMS interventions for receiving Pap smear results and appointment reminders. Reasons for interest, articulated by both providers and clients, included convenience, cost and time-saving benefits and benefits of not taking time off work. However, concerns were expressed around confidentiality of SMS messages, loss/theft of mobile phones, receiving negative results via SMS and accessibility/clarity of language used to convey messages. Responsibility for the management of a clinic-based SMS system was also raised. CONCLUSIONS: Results indicated interest and potential for mHealth interventions in improving follow-up and management of clients with abnormal Pap smears. Health system and privacy issues will need to be addressed for mHealth to achieve this potential. Next steps include piloting of specific SMS messages to test feasibility and acceptability in this setting.

Integrated in vivo multiomics analysis identifies p21-activated kinase signaling as a driver of colitis.

Inflammatory bowel disease (IBD) is a chronic disorder of the gastrointestinal tract that has limited treatment options. To gain insight into the pathogenesis of chronic colonic inflammation (colitis), we performed a multiomics analysis that integrated RNA microarray, total protein mass spectrometry (MS), and phosphoprotein MS measurements from a mouse model of the disease. Because we collected all three types of data from individual samples, we tracked information flow from RNA to protein to phosphoprotein and identified signaling molecules that were coordinately or discordantly regulated and pathways that had complex regulation in vivo. For example, the genes encoding acute-phase proteins were expressed in the liver, but the proteins were detected by MS in the colon during inflammation. We also ascertained the types of data that best described particular facets of chronic inflammation. Using gene set enrichment analysis and trans-omics coexpression network analysis, we found that each data set provided a distinct viewpoint on the molecular pathogenesis of colitis. Combining human transcriptomic data with the mouse multiomics data implicated increased p21-activated kinase (Pak) signaling as a driver of colitis. Chemical inhibition of Pak1 and Pak2 with FRAX597 suppressed active colitis in mice. These studies provide translational insights into the mechanisms contributing to colitis and identify Pak as a potential therapeutic target in IBD.

Comparing brief interventions for suicidal individuals not engaged in treatment: A randomized clinical trial.

BACKGROUND: Non-treatment-engaged individuals experiencing suicidal thoughts have been largely overlooked in the intervention literature, despite reviews suggesting most individuals who die by suicide were not in treatment immediately prior to their death. Most intervention studies recruit individuals from treatment providers, potentially neglecting those individuals who are not already engaged in services. These individuals clearly represent a group in need of additional empirical attention. METHODS: A randomized clinical trial was conducted to compare a single-session dialectical behavior therapy skills-based intervention to a relaxation training control condition. Ninety-three non-treatment-engaged subjects participated in a single in-person assessment, received one of the intervention protocols, and completed follow-up phone interviews for three months including measures of suicidal ideation, emotion dysregulation, and coping skills, as well as other relevant assessments. RESULTS: Both conditions reported significantly reduced levels of suicidal ideation, depression, and anxiety; however, analyses revealed no significant differences between conditions on the main outcome measures of suicidal ideation, emotion dysregulation, skills use, depression, or anxiety. LIMITATIONS: The two interventions may have been too similar to permit detection of differential effects with this sample size. Specifically, the control condition may have been too active and there may have been stylistic overlap by providers who delivered both interventions. CONCLUSIONS: Encouragingly, half of subjects contacted other mental health services during the follow-up period. Although the two interventions under investigation did not yield differential results, the significant changes in important domains across interventions suggest that brief interventions may hold promise for this difficult-to-reach population.

Matrix metalloproteinase activity stimulates N-cadherin shedding and the soluble N-cadherin ectodomain promotes classical microglial activation.

BACKGROUND: Matrix metalloproteinases (MMPs) are a family of enzymes that are typically released from intracellular stores to act on specific extracellular substrates. MMP expression and activity can be increased in a neuronal activity-dependent manner, and further increased in response to tissue injury. MMP substrates include cell adhesion molecules (CAMs) that are abundantly expressed in the brain and well positioned for membrane proximal cleavage. Importantly, CAM integrity is important to synaptic structure and axon-myelin interactions, and shed ectodomains may themselves influence cellular function. METHODS: In the present study, we have examined proteolysis of N-cadherin (N-cdh) by MMP-7, a family member that has been implicated in disorders including HIV dementia, multiple sclerosis, and major depression. With in vitro digest assays, we tested N-cdh cleavage by increasing concentrations of recombinant enzyme. We also tested MMP-7 for its potential to stimulate N-cdh shedding from cultured neural cells. Since select CAM ectodomains may interact with cell surface receptors that are expressed on microglial cells, we subsequently tested the N-cdh ectodomain for its ability to stimulate activation of this cell type as determined by nuclear translocation of NF-κB, Iba-1 expression, and TNF-α release. RESULTS: We observed that soluble N-cdh increased Iba-1 levels in microglial lysates, and also increased microglial release of the cytokine TNF-α. Effects were associated with increased NF-κB immunoreactivity in microglial nuclei and diminished by an inhibitor of the toll-like receptor adaptor protein, MyD88. CONCLUSIONS: Together, these in vitro results suggest that soluble N-cdh may represent a novel effector of microglial activation, and that disorders with increased MMP levels may stimulate a cycle in which the products of excess proteolysis further exacerbate microglial-mediated tissue injury. Additional in vivo studies are warranted to address this issue.

Challenges in Targeting a Basic Helix-Loop-Helix Transcription Factor with Hydrocarbon-Stapled Peptides.

Basic helix-loop-helix (bHLH) transcription factors play critical roles in organism development and disease by regulating cell proliferation and differentiation. Transcriptional activity, whether by bHLH homo- or heterodimerization, is dependent on protein-protein and protein-DNA interactions mediated by α-helices. Thus, α-helical decoys have been proposed as potential targeted therapies for pathologic bHLH transcription. Here, we developed a library of stabilized α-helices of OLIG2 (SAH-OLIG2) to test the capacity of hydrocarbon-stapled peptides to disrupt OLIG2 homodimerization, which drives the development and chemoresistance of glioblastoma multiforme, one of the deadliest forms of human brain cancer. Although stapling successfully reinforced the α-helical structure of bHLH constructs of varying length, sequence-specific dissociation of OLIG2 dimers from DNA was not achieved. Re-evaluation of the binding determinants for OLIG2 self-association and stability revealed an unanticipated role of the C-terminal domain. These data highlight potential pitfalls in peptide-based targeting of bHLH transcription factors given the liabilities of their positively charged amino acid sequences and multifactorial binding determinants.

Multiplexed Quantitative Proteomics for High-Throughput Comprehensive Proteome Comparisons of Human Cell Lines.

The proteome is the functional entity of the cell, and perturbations of a cellular system almost always cause changes in the proteome. These changes are a molecular fingerprint, allowing characterization and a greater understanding of the effect of the perturbation on the cell as a whole. Monitoring these changes has therefore given great insight into cellular responses to stress and disease states, and analytical platforms to comprehensively analyze the proteome are thus extremely important tools in biological research. Mass spectrometry has evolved as the most relevant technology to characterize proteomes in a comprehensive way. However, due to a lack of throughput capacity of mass spectrometry-based proteomics, researchers frequently use measurement of mRNA levels to approximate proteome changes. Growing evidence of substantial differences between mRNA and protein levels as well as recent improvements in mass spectrometry-based proteomics are heralding an increased use of mass spectrometry for comprehensive proteome mapping. Here we describe the use of multiplexed quantitative proteomics using isobaric labeling with tandem mass tags (TMT) for the simultaneous quantitative analysis of five cancer cell proteomes in biological duplicates in one mass spectrometry experiment.