Post-traumatic stress disorder (PTSD) is psychiatric disease, which can occur following exposure to traumatic events. PTSD may be acute or chronic, and can have a waxing and waning course of symptoms. It has been hypothesized that proinflammatory cytokines and chemokines in the cerebrospinal fluid (CSF) or plasma might be mediators of the psychophysiological mechanisms relating a history of trauma exposure to changes in behavior and mental health disorders, and medical morbidity. Here we test the cytokine/chemokine hypothesis for PTSD by examining levels of 17 classical cytokines and chemokines in CSF, sampled at 0900 hours, and in plasma sampled hourly for 24 h. The PTSD and healthy control patients are from the NIMH Chronic PTSD and healthy control cohort, initially described by Bonne et al. (2011), in which the PTSD patients have relatively low comorbidity for major depressive disorder (MDD), drug or alcohol use. We find that in plasma, but not CSF, the bivariate MCP4 (CCL13)/ MCP1(CCL2) ratio is ca. twofold elevated in PTSD patients compared with healthy controls. The MCP-4/MCP-1 ratio is invariant over circadian time, and is independent of gender, body mass index or the age at which the trauma was suffered. By contrast, MIP-1β is a candidate biomarker for PTSD only in females, whereas TARC is a candidate biomarker for PTSD only in males. It remains to be discovered whether these disease-specific differences in circadian expression for these specific immune signaling molecules are biomarkers, surrogates, or drivers for PTSD, or whether any of these analytes could contribute to therapy.

Devising molecular strategies that overcome ΔF508 folding and trafficking defects comprise a central objective of CF therapeutic development. An understanding of disease mechanisms can be improved by new method(s) and/or compounds that redirect ΔF508-CFTR to the plasma membrane. RNA interference (siRNA) mediated knock-down of gene expression has proven to be a powerful tool for investigating protein function(s) and advancing drug discovery. BioFocus (a Galapagos company) has developed adenoviral vectors expressing small hairpin RNAs (shRNAs) for genomewide functional screening that allow robust transduction and durable gene repression. In this project, sixty-eight adenoviral-shRNA constructs (targeting 28 high priority genes) were provided by BioFocus to five collaborating laboratories in the United States and Canada. Putative gene targets were identified by a consortium-based review of the existing CF literature. Each research group established independent protocols to investigate effect(s) of gene knock-down (via Ad-shRNA) on ΔF508-CFTR maturation. Protocols included 1) A study of CF bronchial epithelial cells expressing the halide sensitive variant of eYFP and measurement of ΔF508-CFTR activity at the cell surface. 2) Short circuit current in primary human bronchial epithelial cells (ΔF508/ΔF508), 3) Appearance of rescued ΔF508 CFTR at the plasma membrane in CF bronchial epithelial cells monitored biochemically, 4) Effects on CFTR-dependent release of inflammatory markers (chemokines and cytokines) from IB3 cells (ΔF508/W1282X), and 5) Short circuit current and Western blotting in CFBE cells transduced with lentivirus encoding ΔF508-CFTR. Preliminary results indicate significant activity of BioFocus shRNAs in several of the independent protocols and laboratories, particularly against gene products such as AHSA1 and 2 (activators of HSP 90) and HDAC7A (a member of the histone deacetylase family). A profile of shRNAs found to improve ΔF508 processing, including spectrum of activity data and gene-network annotation of the relevant pathways, will be presented. Identifying the most robust molecular targets for ΔF508 CFTR correction (from among hundreds of candidates in the CFTR “interactome”) has been limited by the complexity of the relevant cellular pathways. The studies described here provide a means by which chaperones and other contributors to CFTR misprocessing can be evaluated, prioritized, and better understood in the future for development of new therapeutic approaches and delineating genetic modifiers that contribute to variation in severity of CF onset and disease progression. The project represents a collaboration among members of the CFTR Folding Consortium. Supported by the CFF and NIH.