A Role for Calcium Influx in the Regulation of Mitochondrial Calcium in Endothelial Cells

By using an endothelial cell line (ECV304), derived from human umbilical vein and transfected with recombinant aequorin targeted to the mitochondrial matrix, we find that stimulation with ATP evokes long lasting increases in mitochondrial Ca2+ ([Ca2+]m) that largely depend on Ca2+ influx. In these cells, the release of stored Ca2+ is inefficient at elevating [Ca2+]m. Consequently it appears that in ECV304 cells, bulk cytosolic Ca2+ ([Ca2+]c) is the main determinant of [Ca2+]m changes. In ECV304 cells4% of mitochondria are within 700 nm of the endoplasmic reticulum as opposed to 65% in HeLa cells, whereas 14% are within 700 nm of the inner surface of the plasma membrane, compared with6% in HeLa cells. Following Ca2+ depletion, readdition of extracellular Ca2+ evokes an increase in [Ca2+]m but not in [Ca2+]c. Under these conditions, microdomains of high [Ca2+]c may occur beneath the plasma membrane of ECV304 cells resulting in the preferential elevation of Ca2+ in mitochondria located in this region. A model is discussed in which the localization of mitochondria with respect to Ca2+ sources is the main determinant of their in situ Ca2+ uptake kinetics. Thus, in any given cell type mitochondria may be localized to suit the energy and metabolic demands of their physiological actions

Automated Classification of Depression from Structural Brain Measures across Two Independent Community-based Cohorts

ACKNOWLEDGEMENTS: This study was supported and funded by the Wellcome Trust Strategic Award ‘Stratifying Resilience and Depression Longitudinally’ (STRADL) (Reference 104036/Z/14/Z), and the Medical Research Council Mental Health Pathfinder Award ‘Leveraging routinely collected and linked research data to study the causes and consequences of common mental disorders’ (Reference MRC-MC_PC_17209). MAH is supported by research funding from the Dr Mortimer and Theresa Sackler Foundation. The research was conducted using the UK Biobank resource, with application number 4844. Structural brain imaging data from the UK Biobank was processed at the University of Edinburgh Centre for Cognitive Ageing and Cognitive Epidemiology (CCACE) http://www.ccace.ed.ac.uk/), which is a part of the crosscouncil Lifelong Health and Wellbeing Initiative (MR/K026992/1). CCACE received funding from Biotechnology and Biological Sciences Research Council (BBSRC), Medical Research Council (MRC), and was also supported by Age UK as part of The Disconnected Mind project. This work has made use of the resources provided by the Edinburgh Compute and Data Facility (ECDF) (http://www.ecdf.ed.ac.uk/)Peer reviewedPublisher PD

Disrupted limbic-prefrontal effective connectivity in response to fearful faces in lifetime depression

Background: Multiple brain imaging studies of negative emotional bias in major depressive disorder (MDD) have used images of fearful facial expressions and focused on the amygdala and the prefrontal cortex. The results have, however, been inconsistent, potentially due to small sample sizes (typically N < 50 ). It remains unclear if any alterations are a characteristic of current depression or of past experience of depression, and whether there are MDD-related changes in effective connectivity between the two brain regions.Methods: Activations and effective connectivity between the amygdala and dorsolateral prefrontal cortex (DLPFC) in response to fearful face stimuli were studied in a large population-based sample from Generation Scotland. Participants either had no history of MDD ( N = 664 in activation analyses, N = 474 in connectivity analyses) or had a diagnosis of MDD during their lifetime (LMDD, N = 290 in activation analyses, N = 214 in connectivity analyses). The within-scanner task involved implicit facial emotion processing of neutral and fearful faces.Results: Compared to controls, LMDD was associated with increased activations in left amygdala ( PFWE = 0.031 , k E = 4 ) and left DLPFC ( PFWE = 0.002 , k E = 33 ), increased mean bilateral amygdala activation ( β = 0.0715, P = 0.0314 ), and increased inhibition from left amygdala to left DLPFC, all in response to fearful faces contrasted to baseline. Results did not appear to be attributable to depressive illness severity or antidepressant medication status at scan time.Limitations: Most studied participants had past rather than current depression, average severity of ongoing depression symptoms was low, and a substantial proportion of participants were receiving medication. The study was not longitudinal and the participants were only assessed a single time.Conclusions: LMDD is associated with hyperactivity of the amygdala and DLPFC, and with stronger amygdala to DLPFC inhibitory connectivity, all in response to fearful faces, unrelated to depression severity at scan time. These results help reduce inconsistency in past literature and suggest disruption of ‘bottom-up’ limbic-prefrontal effective connectivity in depression

Safety and immunogenicity of an FP9-vectored candidate tuberculosis vaccine (FP85A), alone and with candidate vaccine MVA85A in BCG-vaccinated healthy adults: a phase I clinical trial.

The safety and immunogenicity of a new candidate tuberculosis (TB) vaccine, FP85A was evaluated alone and in heterologous prime-boost regimes with another candidate TB vaccine, MVA85A. This was an open label, non-controlled, non-randomized Phase I clinical trial. Healthy previously BCG-vaccinated adult subjects were enrolled sequentially into three groups and vaccinated with FP85A alone, or both FP85A and MVA85A, with a four week interval between vaccinations. Passive and active data on adverse events were collected. Immunogenicity was evaluated by Enzyme Linked Immunospot (ELISpot), flow cytometry and Enzyme Linked Immunosorbent assay (ELISA). Most adverse events were mild and there were no vaccine-related serious adverse events. FP85A vaccination did not enhance antigen 85A-specific cellular immunity. When MVA85A vaccination was preceded by FP85A vaccination, cellular immune responses were lower compared with when MVA85A vaccination was the first immunisation. MVA85A vaccination, but not FP85A vaccination, induced anti-MVA IgG antibodies. Both MVA85A and FP85A vaccinations induced anti-FP9 IgG antibodies. In conclusion, FP85A vaccination was well tolerated but did not induce antigen-specific cellular immune responses. We hypothesize that FP85A induced anti-FP9 IgG antibodies with cross-reactivity for MVA85A, which may have mediated inhibition of the immune response to subsequent MVA85A. ClinicalTrials.gov identification number: NCT00653770

Blunted Medial Prefrontal Cortico-Limbic Reward-Related Effective Connectivity and Depression

Stratifying Resilience and Depression Longitudinally (STRADL) was supported by the Wellcome Trust through a Strategic Award (Grant No. 104036/Z/14/Z). Parts of the work were supported by a China Scholarship Council (Grant No. 201506040037 to SX), National Institutes of Health (Grant No. DA027764 to MRD), Lister Institute Prize Fellowship 2016–2021 (to DJS), Dr Mortimer and Theresa Sackler Foundation (AMM, HCW, and SML), Centre for Cognitive Ageing and Cognitive Epidemiology (IJD and AMM), Medical Research Council and Biotechnology and Biological Sciences Research Council (Grant No. MR/K026992/1), Royal College of Physicians of Edinburgh John, Margaret, Alfred and Stewart Sim fellowship (to HCW), and University of Edinburgh, Edinburgh Scientific Academic TmPCk College Fellowship (to HCW). The Chief Scientist Office of the Scottish Government Health Department (Grant No. CZD/16/6) and Scottish Funding Council (Grant No. HR03006) provided core support for Generation Scotland. Data acquisition was additionally supported by the Scottish Mental Health Research Network and Scottish Government’s Support for Science initiative. LR, HCW, and AMM, received financial support from Pfizer (formerly Wyeth) in relation to imaging studies of people with schizophrenia and bipolar disorder. AMM has previously received grant support from Lilly and Janssen. SML has received honoraria for lectures, chairing meetings, and consultancy work from Janssen in connection with brain imaging and therapeutic initiatives for psychosis. JDS has received funding via an honorarium associated with a lecture or Wyeth and funding from Indivior for a study on opioid dependency. No other disclosures were reported. The authors declare no conflict of interest.Peer reviewedPublisher PD

A single dose of ChAdOx1 Chik vaccine induces neutralising antibodies against four chikungunya virus lineages in a phase 1 clinical trial

Chikungunya virus (CHIKV) is a reemerging mosquito-borne virus that causes swift outbreaks. Major concerns are the persistent and disabling polyarthralgia in infected individuals. Here we present the results from a first-in-human trial of the candidate simian adenovirus vectored vaccine ChAdOx1 Chik, expressing the CHIKV full-length structural polyprotein (Capsid, E3, E2, 6k and E1). 24 adult healthy volunteers aged 18–50 years, were recruited in a dose escalation, open-label, nonrandomized and uncontrolled phase 1 trial (registry NCT03590392). Participants received a single intramuscular injection of ChAdOx1 Chik at one of the three preestablished dosages and were followed-up for 6 months. The primary objective was to assess safety and tolerability of ChAdOx1 Chik. The secondary objective was to assess the humoral and cellular immunogenicity. ChAdOx1 Chik was safe at all doses tested with no serious adverse reactions reported. The vast majority of solicited adverse events were mild or moderate, and self-limiting in nature. A single dose induced IgG and Tcell responses against the CHIKV structural antigens. Broadly neutralizing antibodies against the four CHIKV lineages were found in all participants and as early as 2 weeks after vaccination. In summary, ChAdOx1 Chik showed excellent safety, tolerability and 100% PRNT50 seroconversion after a single dose

Repeat controlled human malaria infection of healthy UK adults with blood-stage plasmodium falciparum:Safety and parasite growth dynamics

In endemic settings it is known that natural malaria immunity is gradually acquired following repeated exposures. Here we sought to assess whether similar acquisition of blood-stage malaria immunity would occur following repeated parasite exposure by controlled human malaria infection (CHMI). We report the findings of repeat homologous blood-stage Plasmodium falciparum (3D7 clone) CHMI studies VAC063C (ClinicalTrials.gov NCT03906474) and VAC063 (ClinicalTrials.gov NCT02927145). In total, 24 healthy, unvaccinated, malaria-naïve UK adult participants underwent primary CHMI followed by drug treatment. Ten of these then underwent secondary CHMI in the same manner, and then six of these underwent a final tertiary CHMI. As with primary CHMI, malaria symptoms were common following secondary and tertiary infection, however, most resolved within a few days of treatment and there were no long term sequelae or serious adverse events related to CHMI. Despite detectable induction and boosting of anti-merozoite serum IgG antibody responses following each round of CHMI, there was no clear evidence of anti-parasite immunity (manifest as reduced parasite growth in vivo) conferred by repeated challenge with the homologous parasite in the majority of volunteers. However, three volunteers showed some variation in parasite growth dynamics in vivo following repeat CHMI that were either modest or short-lived. We also observed no major differences in clinical symptoms or laboratory markers of infection across the primary, secondary and tertiary challenges. However, there was a trend to more severe pyrexia after primary CHMI and the absence of a detectable transaminitis post-treatment following secondary and tertiary infection. We hypothesize that this could represent the initial induction of clinical immunity. Repeat homologous blood-stage CHMI is thus safe and provides a model with the potential to further the understanding of naturally acquired immunity to blood-stage infection in a highly controlled setting. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov, identifier NCT03906474, NCT02927145

Safety, Immunogenicity and Efficacy of Prime-Boost Vaccination with ChAd63 and MVA Encoding ME-TRAP against Plasmodium falciparum Infection in Adults in Senegal.

Malaria transmission is in decline in some parts of Africa, partly due to the scaling up of control measures. If the goal of elimination is to be achieved, additional control measures including an effective and durable vaccine will be required. Studies utilising the prime-boost approach to deliver viral vectors encoding the pre-erythrocytic antigen ME-TRAP (multiple epitope thrombospondin-related adhesion protein) have shown promising safety, immunogenicity and efficacy in sporozoite challenge studies. More recently, a study in Kenyan adults, similar to that reported here, showed substantial efficacy against P. falciparum infection. One hundred and twenty healthy male volunteers, living in a malaria endemic area of Senegal were randomised to receive either the Chimpanzee adenovirus (ChAd63) ME-TRAP as prime vaccination, followed eight weeks later by modified vaccinia Ankara (MVA) also encoding ME-TRAP as booster, or two doses of anti-rabies vaccine as a comparator. Prior to follow-up, antimalarials were administered to clear parasitaemia and then participants were monitored by PCR for malaria infection for eight weeks. The primary endpoint was time-to-infection with P. falciparum malaria, determined by two consecutive positive PCR results. Secondary endpoints included adverse event reporting, measures of cellular and humoral immunogenicity and a meta-analysis of combined vaccine efficacy with the parallel study in Kenyan adults.We show that this pre-erythrocytic malaria vaccine is safe and induces significant immunogenicity, with a peak T-cell response at seven days after boosting of 932 Spot Forming Cells (SFC)/106 Peripheral Blood Mononuclear Cells(PBMC) compared to 57 SFC/ 106 PBMCs in the control group. However, a vaccine efficacy was not observed: 12 of 57 ME-TRAP vaccinees became PCR positive during the intensive monitoring period as compared to 13 of the 58 controls (P = 0.80). This trial confirms that vaccine efficacy against malaria infection in adults may be rapidly assessed using this efficient and cost-effective clinical trial design. Further efficacy evaluation of this vectored candidate vaccine approach in other malaria transmission settings and age-de-escalation into the main target age groups for a malaria vaccine is in progress