Abstract
ABSTRACTMaladaptive signaling by pro-inflammatory cytokines (PICs), such as TNFα, IL1β and IFNɣ, can activate downstream signaling cascades that are implicated in the development and progression of multiple inflammatory diseases. Despite playing critical roles in pathogenesis, the availability of in vivo models in which to model tissue-specific induction of PICs is limited. To bridge this gap, we have developed a novel multi-gene expression system dubbed Cre-enabled and tetracycline-inducible transgenic system for conditional, tissue-specific expression of pro-inflammatory cytokines (CETI-PIC3). This binary transgenic system permits the stoichiometric co-expression of proteins Tumor necrosis factor a (Tnfa), Interleukin-1 beta (Il1b) and Interferon gamma (Ifng1), and H2B-GFP fluorescent reporter in a dose-dependent manner. Furthermore, cytokine misexpression is enabled only in tissue domains that can be defined by Cre recombinase expression. We have validated this system in zebrafish using an insulin:cre line. In doubly transgenic fish, quantitative real-time polymerase chain reaction demonstrated increased expression levels of tnfa, il1b and ifng1 mRNA. Moreover, specific expression in pancreatic β cells was demonstrated by both Tnfa immunofluorescence and GFP fluorescence. Cytokine-overexpressing islets elicited specific responses: β cells exhibited increased expression of genes associated with reactive oxidative species-mediated stress and endoplasmic reticulum stress, surveilling and infiltrating macrophages were increased, and β cell death was promoted. This powerful and versatile model system can be used for modeling, analysis and therapy development of diseases with an underlying inflammatory etiology.This article has an associated First Person interview with the first author of the paper.
Highlights
Pro-inflammatory cytokines (PICs) are signaling molecules that are primarily produced by immune cells that coordinate and amplify certain immune responses
Lethality was observed at higher doses, but there was no difference in survival between the CETI-pro-inflammatory cytokines 3 (PIC3) embryos and WT embryos across all treatment regimens tested; this indicates that the lethality is due to the toxicity of high doxycycline dosing, not leaky Cre-independent expression of reverse tetracycline trans activator (rtTA) and/or non-specific PIC3 cassette expression
To confirm that the CETI-PIC3 system is competent to drive the induction of the PIC3 pro-inflammatory cytokine expression cassette in a titratable manner, the levels of tnfa, ifng1 and il1b, and H2B-GFP mRNA were measured using quantitative real-time polymerase chain reaction analysis of whole zebrafish larvae
Summary
Pro-inflammatory cytokines (PICs) are signaling molecules that are primarily produced by immune cells (including activated macrophages and T cells) that coordinate and amplify certain immune responses. Pro-inflammatory cytokines play a critical role in diseases with an underlying maladaptive inflammatory pathology (Dinarello, 2000) Cytokines such as interferon gamma (IFNɣ), interleukin 1 beta (IL1β) and tumor necrosis factor alpha (TNFα; known as TNF) can drive beta (β) cell dysfunction, damage and death in diabetes mellitus (Hatanaka et al, 2006; Dandona et al, 2004). Feedback amplification loops of cytokine signaling have been shown to further attract more innate immune cells and augment the innate immune response (Németh and Mócsai, 2016; Canna and Behrens, 2012) These pathways are important mediators of disease in islet injury observed in diabetes and in tissue injury caused by inflammation, rendering pro-inflammatory cytokines crucial mediators and aggravators of many disease states
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
