A20, an essential component of the ubiquitin-editing protein complex, is a negative regulator of inflammation in human myometrium and foetal membranes.

STUDY QUESTION: Does A20 regulate mediators involved in the terminal processes of human labour in primary myometrial and amnion cells?

SUMMARY ANSWER: A20 is a nuclear factor-kappa B (NF-κB) responsive gene that acts as a negative regulator of NF-κB-induced expression of pro-labour mediators.

WHAT IS KNOWN ALREADY: Inflammation is commonly implicated in spontaneous preterm birth and the processes involved in rupture of foetal membranes and uterine contractions. In myometrium and foetal membranes, the pro-inflammatory transcription factor NF-κB regulates the transcription of pro-labour mediators in response to inflammatory stimuli. In non-gestational tissues, A20 is widely recognised as an anti-inflammatory protein that inhibits inflammation-induced NF-κB signalling.

STUDY DESIGN, SIZE, DURATION: Primary human amnion and myometrial cells were used to determine the effect of pro-inflammatory mediators on A20 expression and the effect of A20 siRNA on the expression and secretion of pro-labour mediators. The expression of A20 was assessed in myometrium and foetal membranes from non-labouring and labouring women at preterm and or term (n = 8 or nine samples per group).

PARTICIPANTS/MATERIALS, SETTING, METHODS: The effects of pro-inflammatory mediators and of A20 siRNA in cell cultures were determined by quantitative RT-PCR (qRT-PCR), western blots, immunoassays, gelatin zymography and luciferase assays. A20 expression in tissue samples was assessed by qRT-PCR. Statistical significance was ascribed to a P value < 0.05.

MAIN RESULTS AND THE ROLE OF CHANCE: In primary cells isolated from myometrium and or amnion, the pro-inflammatory cytokines IL1B and TNF, the bacterial products flagellin and fsl-1, and the viral double stranded RNA analogue poly(I:C) significantly increased A20 mRNA expression via NF-κB. A20 siRNA studies in primary myometrial and amnion cells demonstrated an augmentation of inflammation-induced expression and or secretion of pro-inflammatory cytokines (IL1A, IL6), chemokines (CXCL1, CXCL8, CCL2), adhesion molecules (ICAM1, VCAM1), contraction-associated proteins (PTGS2, PTGFR, PGF2α) and the extracellular matrix degrading enzyme MMP9, as well as NF-κB activation. Inhibition of NF-κB activity significant attenuated inflammation-induced expression of pro-labour mediators in A20 siRNA transfected cells. Finally, A20 mRNA expression was decreased in myometrium and foetal membranes with labour, and in foetal membranes with chorioamnionitis.

LARGE SCALE DATA: Not applicable.

LIMITATIONS, REASONS FOR CAUTION: The conclusions of this study are solely reliant on the data from in vitro experiments using cells isolated from myometrium and amnion.

WIDER IMPLICATIONS OF THE FINDINGS: The results of this study raise the possibility that targeting A20 may be a therapeutic approach to reduce inflammation associated with spontaneous preterm birth.

STUDY FUNDING AND COMPETING INTEREST(S): Associate Professor Martha Lappas is supported by a Career Development Fellowship from the National Health and Medical Research Council (NHMRC; grant no. 1047025). Funding for this study was provided by the NHMRC (grant no. 1058786), Norman Beischer Medical Research Foundation and the Mercy Research Foundation. There are no competing interests.