Inactivation of CFTR by CRISPR/Cas9 alters transcriptional regulation of inflammatory pathways and other networks.

BACKGROUND: Individuals with cystic fibrosis (CF) experience elevated inflammation in multiple organs, but whether this reflects an inherent feature of CF cells or is a consequence of a pro-inflammatory environment is not clear. METHOD: Using CRISPR/Cas9-mediated mutagenesis of CFTR, 17 subclonal cell lines were generated from Caco-2 cells. Clonal lines with functional CFTR (CFTR+) were compared to those without (CFTR-) to directly address the role of CFTR in inflammatory gene regulation. RESULTS: All lines maintained CFTR mRNA production and formation of tight junctions. CFTR+ lines displayed short circuit currents in response to forskolin, while the CFTR- lines did not. Baseline expression of cytokines IL6 and CXCL8 (IL8) was not different between the lines regardless of CFTR genotype. All lines responded to TNFα and IL1ß by increasing IL6 and CXCL8 mRNA levels, but the CFTR- lines produced more CXCL8 mRNA than the CFTR+ lines. Transcriptomes of 6 CFTR- and 6 CFTR+ lines, before and after stimulation by TNFα, were compared for differential expression as a function of CFTR genotype. While some genes appeared to be differentially expressed simply because of CFTR's absence, others required stimulation for differences to be apparent. CONCLUSION: Together, these data suggest cells respond to CFTR's absence by modulating transcriptional networks, some of which are only apparent when cells are exposed to different environmental contexts, such as inflammation. With regards to inflammation, these data suggest a model in which CFTR's absence leads to a poised, pro-inflammatory state of cells that is only revealed by stimulation.

Psychometric properties of the Generalized Anxiety Disorder Inventory in a Canadian sample.

The Generalized Anxiety Disorder Inventory is a recently developed self-report measure that assesses symptoms of generalized anxiety disorder. Its psychometric properties have not been investigated further since its original development. The current study investigated its psychometric properties in a Canadian student/community sample. Exploratory principal component analysis replicated the original three-component structure. The total scale and subscales demonstrated excellent internal consistency reliability (α = 0.84-0.94) and correlated strongly with the Penn State Worry Questionnaire (r = 0.41-0.74, all ps <0.001) and Generalized Anxiety Disorder-7 (r = 0.55-0.84, all ps <0.001). However, only the total scale and cognitive subscale (r = 0.48-0.49, all ps <0.05) significantly predicted generalized anxiety disorder diagnosis established by diagnostic interview. The somatic subscale in particular may require revision to improve predictive validity. Revision may also be necessary given changes in required somatic symptoms for generalized anxiety disorder diagnostic criteria in more recent versions of the Diagnostic and Statistical Manual of Mental Disorders (i.e. although major changes occurred from Diagnostic and Statistical Manual of Mental Disorders-III-R to Diagnostic and Statistical Manual of Mental Disorders-IV, changes in Diagnostic and Statistical Manual of Mental Disorders-5 were minimal) and the possibility of changes in the upcoming 11th revision of the International Classification of Diseases.