Trappin-2/Elafin Modulate Innate Immune Responses of Human Endometrial Epithelial Cells to PolyI∶C

Abstract

BackgroundUpon viral recognition, innate and adaptive antiviral immune responses are initiated by genital epithelial cells (ECs) to eradicate or contain viral infection. Such responses, however, are often accompanied by inflammation that contributes to acquisition and progression of sexually transmitted infections (STIs). Hence, interventions/factors enhancing antiviral protection while reducing inflammation may prove beneficial in controlling the spread of STIs. Serine antiprotease trappin-2 (Tr) and its cleaved form, elafin (E), are alarm antimicrobials secreted by multiple cells, including genital epithelia.Methodology and Principal FindingsWe investigated whether and how each Tr and E (Tr/E) contribute to antiviral defenses against a synthetic mimic of viral dsRNA, polyinosine-polycytidylic acid (polyI∶C) and vesicular stomatitis virus. We show that delivery of a replication-deficient adenovector expressing Tr gene (Ad/Tr) to human endometrial epithelial cells, HEC-1A, resulted in secretion of functional Tr, whereas both Tr/E were detected in response to polyI∶C. Moreover, Tr/E were found to significantly reduce viral replication by either acting directly on virus or through enhancing polyI∶C-driven antiviral protection. The latter was associated with reduced levels of pro-inflammatory factors IL-8, IL-6, TNFα, lowered expression of RIG-I, MDA5 and attenuated NF-κB activation. Interestingly, enhanced polyI∶C-driven antiviral protection of HEC-Ad/Tr cells was partially mediated through IRF3 activation, but not associated with higher induction of IFNβ, suggesting multiple antiviral mechanisms of Tr/E and the involvement of alternative factors or pathways.Conclusions and SignificanceThis is the first evidence of both Tr/E altering viral binding/entry, innate recognition and mounting of antiviral and inflammatory responses in genital ECs that could have significant implications for homeostasis of the female genital tract.

Antiviral Activity of Trappin-2 and Elafin In Vitro and In Vivo against Genital Herpes

“…Furthermore, we and others have shown that Tr/E exert immunomodulatory activity, where depending on the environment, they can either dampen inflammation or promote immunostimulatory events and prime the immune system (35)(36)(37). Tr/E are found at mucosal surfaces and in secretions, including cervicovaginal lavage fluid (CVL) (22,23,38); in tissues, like skin, placenta, and genital and gastrointestinal tracts (38)(39)(40)(41)(42)(43)(44); and in multiple cell types, such as neutrophils, macrophages, keratinocytes, and epithelial cells (12,22,45), including genital ECs (22,38).…”

“…Tr and E proteins were tested following protein reconstitution as per the manufacturer's instruction: (i) human recombinant 6ϫHis-Tr (with a C terminus His tag) (R&D Systems, Burlington, ON, Canada) (35,49); (ii) commercial human recombinant E (without a tag) HC4011 (Hycult Biotech, Uden, Netherlands) (49). Poly(I·C) (P1530; Sigma-Aldrich, Oakville, ON, Canada) was reconstituted in the phosphate-buffered saline (PBS).…”

“…The replication-deficient adenoviral constructs (Ad) used in this study have been described in detail elsewhere (35,(51)(52)(53). To express human Tr, the Ad/Tr vector, encoding a gene for 95-aa human Tr, was used (52,53).…”

“…Tr/E are regarded as alarm antiproteases, since they are produced mainly in response to proinflammatory stimuli, such as lipopolysaccharide (LPS) (46), TNF-␣ (47), and IL-1␤ (38,48). Additionally, Tr/E were recently shown to be induced and secreted by genital ECs in response to poly(I·C), a surrogate of a viral double-stranded RNA (dsRNA) (22,35,49), thus further supporting their importance in the inflammatory environment.…”

“…We and others recently demonstrated the importance of Tr/E in increased antiviral protection against HIV-1 as well as enhanced poly(I·C)-induced antiviral immune responses (22,23,30,35,49). Specifically, elevated Tr/E levels in CVLs of HIV-exposed seronegative commercial sex workers (CSWs) were found to be associated with mucosal HIV-1 resistance (23) and to significantly contribute to CVL's natural anti-HIV-1 activity in vitro (49).…”