Modeling and Analysis of the Molecular Basis of Pain in Sensory Neurons

Intracellular calcium dynamics are critical to cellular functions like pain transmission. Extracellular ATP plays an important role in modulating intracellular calcium levels by interacting with the P2 family of surface receptors. In this study, we developed a mechanistic mathematical model of ATP-induced P2 mediated calcium signaling in archetype sensory neurons. The model architecture, which described 90 species connected by 162 interactions, was formulated by aggregating disparate molecular modules from literature. Unlike previous models, only mass action kinetics were used to describe the rate of molecular interactions. Thus, the majority of the 252 unknown model parameters were either association, dissociation or catalytic rate constants. Model parameters were estimated from nine independent data sets taken from multiple laboratories. The training data consisted of both dynamic and steady-state measurements. However, because of the complexity of the calcium network, we were unable to estimate unique model parameters. Instead, we estimated a family or ensemble of probable parameter sets using a multi-objective thermal ensemble method. Each member of the ensemble met an error criterion and was located along or near the optimal trade-off surface between the individual training data sets. The model quantitatively reproduced experimental measurements from dorsal root ganglion neurons as a function of extracellular ATP forcing. Hypothesized architecture linking phosphoinositide regulation with P2X receptor activity explained the inhibition of P2X-mediated current flow by activated metabotropic P2Y receptors. Sensitivity analysis using individual and the whole system outputs suggested which molecular subsystems were most important following P2 activation. Taken together, modeling and analysis of ATP-induced P2 mediated calcium signaling generated qualitative insight into the critical interactions controlling ATP induced calcium dynamics. Understanding these critical interactions may prove useful for the design of the next generation of molecular pain management strategies

A healthy school start - Parental support to promote healthy dietary habits and physical activity in children: Design and evaluation of a cluster-randomised intervention

<p>Abstract</p> <p>Background</p> <p>Childhood obesity is multi-factorial and determined to a large extent by dietary habits, physical activity and sedentary behaviours. Previous research has shown that school-based programmes are effective but that their effectiveness can be improved by including a parental component. At present, there is a lack of effective parental support programmes for improvement of diet and physical activity and prevention of obesity in children.</p> <p>Methods/Design</p> <p>This paper describes the rationale and design of a parental support programme to promote healthy dietary habits and physical activity in six-year-old children starting school. The study is performed in close collaboration with the school health care and is designed as a cluster-randomised controlled trial with a mixed methods approach. In total, 14 pre-school classes are included from a municipality in Stockholm county where there is large variation in socio-economic status between the families. The school classes are randomised to intervention (n = 7) and control (n = 7) groups including a total of 242 children. The intervention is based on social cognitive theory and consists of three main components: 1) a health information brochure; 2) two motivational interviewing sessions with the parents; and 3) teacher-led classroom activities with the children. The primary outcomes are physical activity in the children measured objectively by accelerometry, children's dietary and physical activity habits measured with a parent-proxy questionnaire and parents' self-efficacy measured by a questionnaire. Secondary outcomes are height, weight and waist circumference in the children. The duration of the intervention is six months and includes baseline, post intervention and six months follow-up measurements. Linear and logistic regression models will be used to analyse differences between intervention and control groups in the outcome variables. Mediator and moderator analysis will be performed. Participants will be interviewed.</p> <p>Discussion</p> <p>The results from this study will show if it is possible to promote a healthy lifestyle and a normal weight development among children from low-income districts with relatively limited efforts involving parents. Hopefully the study will provide new insights to the further development of effective programmes to prevent overweight and obesity in children.</p> <p>Trial registration</p> <p>ISRCTN: <a href="http://www.controlled-trials.com/ISRCTN32750699">ISRCTN32750699</a></p

A Liposome-Based Mycobacterial Vaccine Induces Potent Adult and Neonatal Multifunctional T Cells through the Exquisite Targeting of Dendritic Cells

BACKGROUND: In the search for more potent and safer tuberculosis vaccines, CAF01 was identified as a remarkable formulation. Based on cationic liposomes and including a synthetic mycobacterial glycolipid as TLR-independent immunomodulator, it induces strong and protective T helper-1 and T helper-17 adult murine responses to Ag85B-ESAT-6, a major mycobacterial fusion protein. Here, we assessed whether these properties extend to early life and how CAF01 mediates its adjuvant properties in vivo. METHODS/FINDINGS: Following adult or neonatal murine immunization, Ag85B-ESAT-6/CAF01 similarly reduced the post-challenge bacterial growth of M. bovis BCG, whereas no protection was observed using Alum as control. This protection was mediated by the induction of similarly strong Th1 and Th17 responses in both age groups. Multifunctional Th1 cells were already elicited after a single vaccine dose and persisted at high levels for at least 6 months even after neonatal priming. Unexpectedly, this potent adjuvanticity was not mediated by a massive targeting/activation of dendritic cells: in contrast, very few DCs in the draining lymph nodes were bearing the labeled antigen/adjuvant. The increased expression of the CD40 and CD86 activation markers was restricted to the minute portion of adjuvant-bearing DCs. However, vaccine-associated activated DCs were recovered several days after immunization. CONCLUSION: The potent adult and neonatal adjuvanticity of CAF01 is associated in vivo with an exquisite but prolonged DC uptake and activation, fulfilling the preclinical requirements for novel tuberculosis vaccines to be used in early life

Reduced Secretion of YopJ by Yersinia Limits In Vivo Cell Death but Enhances Bacterial Virulence

Numerous microbial pathogens modulate or interfere with cell death pathways in cultured cells. However, the precise role of host cell death during in vivo infection remains poorly understood. Macrophages infected by pathogenic species of Yersinia typically undergo an apoptotic cell death. This is due to the activity of a Type III secreted effector protein, designated YopJ in Y. pseudotuberculosis and Y. pestis, and YopP in the closely related Y. enterocolitica. It has recently been reported that Y. enterocolitica YopP shows intrinsically greater capacity for being secreted than Y. pestis YopJ, and that this correlates with enhanced cytotoxicity observed for high virulence serotypes of Y. enterocolitica. The enzymatic activity and secretory capacity of YopP from different Y. enterocolitica serotypes have been shown to be variable. However, the underlying basis for differential secretion of YopJ/YopP, and whether reduced secretion of YopJ by Y. pestis plays a role in pathogenesis during in vivo infection, is not currently known. It has also been reported that similar to macrophages, Y. enterocolitica infection of dendritic cells leads to YopP-dependent cell death. We demonstrate here that in contrast to Y. enterocolitica, Y. pseudotuberculosis infection of bone marrow–derived dendritic cells does not lead to increased cell death. However, death of Y. pseudotuberculosis–infected dendritic cells is enhanced by ectopic expression of YopP in place of YopJ. We further show that polymorphisms at the N-terminus of the YopP/YopJ proteins are responsible for their differential secretion, translocation, and consequent cytotoxicity. Mutation of two amino acids in YopJ markedly enhanced both translocation and cytotoxicity. Surprisingly, expression of YopP or a hypersecreted mutant of YopJ in Y. pseudotuberculosis resulted in its attenuation in oral mouse infection. Complete absence of YopJ also resulted in attenuation of virulence, in accordance with previous observations. These findings suggest that control of cytotoxicity is an important virulence property for Y. pseudotuberculosis, and that intermediate levels of YopJ-mediated cytotoxicity are necessary for maximal systemic virulence of this bacterial pathogen

The TGF-β/Smad pathway induces breast cancer cell invasion through the up-regulation of matrix metalloproteinase 2 and 9 in a spheroid invasion model system

Transforming growth factor-beta (TGF-beta) has opposing roles in breast cancer progression by acting as a tumor suppressor in the initial phase, but stimulating invasion and metastasis at later stages. In contrast to the mechanisms by which TGF-beta induces growth arrest, the pathways that mediate tumor invasion are not well understood. Here, we describe a TGF-beta-dependent invasion assay system consisting of spheroids of MCF10A1 normal breast epithelial cells (M1) and RAS-transformed (pre-)malignant derivatives (M2 and M4) embedded in collagen gels. Both basal and TGF-beta-induced invasion of these cell lines was found to correlate with their tumorigenic potential; M4 showing the most aggressive behavior and M1 showing the least. Basal invasion was strongly inhibited by the TGF-beta receptor kinase inhibitor SB-431542, indicating the involvement of autocrine TGF-beta or TGF-beta-like activity. TGF-beta-induced invasion in premalignant M2 and highly malignant M4 cells was also inhibited upon specific knockdown of Smad3 or Smad4. Interestingly, both a broad spectrum matrix metalloproteinase (MMP) inhibitor and a selective MMP2 and MMP9 inhibitor mitigated TGF-beta-induced invasion of M4 cells, while leaving basal invasion intact. In line with this, TGF-beta was found to strongly induce MMP2 and MMP9 expression in a Smad3- and Smad4-dependent manner. This collagen-embedded spheroid system therefore offers a valuable screening model for TGF-beta/Smad- and MMP2- and MMP9-dependent breast cancer invasion.Urolog

A screening tool to prioritize public health risk associated with accidental or deliberate release of chemicals into the atmosphere

The Chemical Events Working Group of the Global Health Security Initiative has developed a flexible screening tool for chemicals that present a risk when accidentally or deliberately released into the atmosphere. The tool is generic, semi-quantitative, independent of site, situation and scenario, encompasses all chemical hazards (toxicity, flammability and reactivity), and can be easily and quickly implemented by non-subject matter experts using freely available, authoritative information. Public health practitioners and planners can use the screening tool to assist them in directing their activities in each of the five stages of the disaster management cycle

Genetic regulation of Nrnx1 expression: an integrative cross-species analysis of schizophrenia candidate genes

Neurexin 1 (NRXN1) is a large presynaptic transmembrane protein that has complex and variable patterns of expression in the brain. Sequence variants in NRXN1 are associated with differences in cognition, and with schizophrenia and autism. The murine Nrxn1 gene is also highly polymorphic and is associated with significant variation in expression that is under strong genetic control. Here, we use co-expression analysis, high coverage genomic sequence, and expression quantitative trait locus (eQTL) mapping to study the regulation of this gene in the brain. We profiled a family of 72 isogenic progeny strains of a cross between C57BL/6J and DBA/2J (the BXD family) using exon arrays and massively parallel RNA sequencing. Expression of most Nrxn1 exons have high genetic correlation (r>0.6) because of the segregation of a common trans eQTL on chromosome (Chr) 8 and a common cis eQTL on Chr 17. These two loci are also linked to murine phenotypes relevant to schizophrenia and to a novel human schizophrenia candidate gene with high neuronal expression (Pleckstrin and Sec7 domain containing 3). In both human and mice, NRXN1 is co-expressed with numerous synaptic and cell signaling genes, and known schizophrenia candidates. Cross-species co-expression and protein interaction network analyses identified glycogen synthase kinase 3 beta (GSK3B) as one of the most consistent and conserved covariates of NRXN1. By using the Molecular Genetics of Schizophrenia data set, we were able to test and confirm that markers in NRXN1 and GSK3B have epistatic interactions in human populations that can jointly modulate risk of schizophrenia

Evidence of causal effect of major depression on alcohol dependence: findings from the psychiatric genomics consortium

BACKGROUND Despite established clinical associations among major depression (MD), alcohol dependence (AD), and alcohol consumption (AC), the nature of the causal relationship between them is not completely understood. We leveraged genome-wide data from the Psychiatric Genomics Consortium (PGC) and UK Biobank to test for the presence of shared genetic mechanisms and causal relationships among MD, AD, and AC. METHODS Linkage disequilibrium score regression and Mendelian randomization (MR) were performed using genome-wide data from the PGC (MD: 135 458 cases and 344 901 controls; AD: 10 206 cases and 28 480 controls) and UK Biobank (AC-frequency: 438 308 individuals; AC-quantity: 307 098 individuals). RESULTS Positive genetic correlation was observed between MD and AD (rgMD−AD = + 0.47, P = 6.6 × 10−10). AC-quantity showed positive genetic correlation with both AD (rgAD−AC quantity = + 0.75, P = 1.8 × 10−14) and MD (rgMD−AC quantity = + 0.14, P = 2.9 × 10−7), while there was negative correlation of AC-frequency with MD (rgMD−AC frequency = −0.17, P = 1.5 × 10−10) and a non-significant result with AD. MR analyses confirmed the presence of pleiotropy among these four traits. However, the MD-AD results reflect a mediated-pleiotropy mechanism (i.e. causal relationship) with an effect of MD on AD (beta = 0.28, P = 1.29 × 10−6). There was no evidence for reverse causation. CONCLUSION This study supports a causal role for genetic liability of MD on AD based on genetic datasets including thousands of individuals. Understanding mechanisms underlying MD-AD comorbidity addresses important public health concerns and has the potential to facilitate prevention and intervention efforts