Identification of SMAD3 as a Novel Mediator of Inflammation in Human Myometrium In Vitro.

Martha Lappas

doi:10.1155/2018/3140420

Abstract

Preterm birth remains the primary cause of early neonatal death and is a major determinant for long-term health consequences. Aberrant intrauterine inflammation and infection are known to augment the synthesis of proinflammatory cytokines and induce uterine contractions, which can subsequently lead to preterm birth. The transforming growth factor-β (TGF-β) superfamily members regulate numerous cellular processes through the activation of intracellular mediators known as mothers against decapentaplegic homolog (SMADs). Studies in nongestational tissues have shown that SMAD3 plays a role in immune regulation and inflammation; however, its role in human labour remains unknown. Thus, the present study aimed at (i) characterising the expression of SMAD3 in the human myometrium; (ii) determining the effect of bacterial and viral products and proinflammatory cytokines on SMAD3 transcriptional activity in primary human myometrial cells; and (iii) investigating the effect of SMAD3 siRNA knockdown on the production of prolabour mediators in primary human myometrial cells. Phosphorylated (i.e., active) SMAD3 protein expression was lower in the myometrium after spontaneous term labour compared to the myometrium from nonlabouring women. Using a luciferase assay, the proinflammatory cytokines IL-1β and TNF, and viral analogue polyinosinic : polycytidylic acid (poly(I : C)) significantly reduced SMAD3 transcriptional activity in human primary myometrial cells. Loss-of-function studies found that SMAD3 knockdown in myometrial cells significantly increased IL-1β- and poly(I : C)-induced proinflammatory cytokines (IL-1A, IL-6), chemokines (IL-8, MCP-1), the adhesion molecule ICAM-1, COX-2 mRNA expression, and subsequent PGF2α release. In conclusion, SMAD3 deficiency is associated with increased production of proinflammatory and prolabour mediators in the human myometrium.

Highlights

The most common cause of early neonatal death is preterm birth
Human myometrium was obtained at term Caesarean section in the absence of labour and at term Caesarean section during spontaneous labour onset
The protein expression of total and phosphorylated Ser 423/425 SMAD3 (p-SMAD3) was assessed by Western blotting; data are expressed as polyclonal antiphosphorylated SMAD3 (pSMAD3) expression relative to total SMAD3 levels

Summary

Introduction

The most common cause of early neonatal death is preterm birth. It is responsible for approximately 1 million neonatal deaths globally each year [1]. Preterm infants have greatly increased rates of long-term disabilities including cerebral palsy, intellectual handicap, and chronic lung disease [2]. Such complications lead to long-term morbidity through childhood and extend into adult life, casting an enormous financial burden on the health system [3]. The presence of intrauterine infection and/or inflammation results in the activation of the maternal immune response and the influx of leukocytes into the myometrium and cervix and increased production of proinflammatory cytokines such as IL-1β and TNF within these tissues [8, 9]. In addition to further inducing cytokines, IL-1β and TNF can induce chemokine production (such as IL-8 and MCP-1) [10] or increase the production of phospholipid-derived mediators (such as prostaglandins) which are known to play an important role in myometrial contractions and cervical remodelling [5, 11,12,13,14]

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