Research Article: Effect of Silibinin in Reducing Inflammatory Pathways in In Vitro and In Vivo Models of Infection-Induced Preterm Birth

Date Published: March 19, 2014

Publisher: Public Library of Science

Author(s): Ratana Lim, Carrington J. Morwood, Gillian Barker, Martha Lappas, Paul Proost.


Infection-induced preterm birth is the largest cause of infant death and of neurological disabilities in survivors. Silibinin, from milk thistle, exerts potent anti-inflammatory activities in non-gestational tissues. The aims of this study were to determine the effect of silibinin on pro-inflammatory mediators in (i) human fetal membranes and myometrium treated with bacterial endotoxin lipopolysaccharide (LPS) or the pro-inflammatory cytokine IL-1β, and (ii) in preterm fetal membranes with active infection. The effect of silibinin on infection induced inflammation and brain injury in pregnant mice was also assessed. Fetal membranes and myometrium (tissue explants and primary cells) were treated with 200 μM silibinin in the presence or absence of 10 μg/ml LPS or 1 ng/ml IL-1β. C57BL/6 mice were injected with 70 mg/kg silibinin with or without 50 μg LPS on embryonic day 16. Fetal brains were collected after 6 h. In human fetal membranes, silibinin significantly decreased LPS-stimulated expression of IL-6 and IL-8, COX-2, and prostaglandins PGE2 and PGF2α. In primary amnion and myometrial cells, silibinin also decreased IL-1β-induced MMP-9 expression. Preterm fetal membranes with active infection treated with silibinin showed a decrease in IL-6, IL-8 and MMP-9 expression. Fetal brains from mice treated with silibinin showed a significant decrease in LPS-induced IL-8 and ninjurin, a marker of brain injury. Our study demonstrates that silibinin can reduce infection and inflammation-induced pro-labour mediators in human fetal membranes and myometrium. Excitingly, the in vivo results indicate a protective effect of silibinin on infection-induced brain injury in a mouse model of preterm birth.

Partial Text

Preterm birth is the single leading cause of neonatal death worldwide, after exclusion of congenital defects, and can lead to numerous long-term health consequences for those surviving babies [1]. Spontaneous preterm birth, which accounts for almost 70% of all preterm births, may result from preterm labour with intact membranes or preterm pre-labour rupture of membranes (PPROM) [2]. Intrauterine infection, commonly presented as chorioamnionitis, is an acute inflammation of the membranes and chorion of the placenta, typically due to ascending bacterial infection. Chorioamnionitis complicates a third of patients with preterm labour [3] and is the most common complication associated with PPROM [4]. Chorioamnionitis predisposes the preterm infant to numerous organ disease, affecting cardiopulmonary, cerebral, and renal systems [5]; developmental delay and lifelong neurological impairments, such as mental retardation, cerebral palsy and learning and behavioural deficits, are caused by perinatal brain damage [6]–[8]. Besides the emotional burden on families, direct and indirect costs of preterm birth accounts for billions of health care dollars each year.

A diet consisting of many serves of fruits and vegetables has been attributed to many health benefits, such as decreased risk of coronary heart disease [63] and decreased incidence of preeclampsia [64], [65]. It is unclear as to why they are beneficial, and it may be due to the presence of dietary phytophenols. In preeclamptic women, the polyphenolic plant flavanoid silibinin has been shown to reduce oxidative metabolism and cytokine production in peripheral blood mononuclear cells [34], [35]. However, the effect of silibinin on mediators of labour in human gestational tissues has not previously been explored. The data presented in this study demonstrate that in human gestational tissues, silibinin decreases LPS and IL-1β-induced mRNA expression and secretion of pro-inflammatory cytokines, COX-2 mRNA expression and resultant prostaglandin release (fetal membranes, but not myometrium), and MMP-9 mRNA expression and activity. In fetal membranes from spontaneous preterm labour and with active infection (confirmed by pathology), silibinin treatment decreased pro-inflammatory cytokine expression and release, COX-2 gene expression and prostaglandin release, and MMP-9 gene expression. We also investigated the ability of silibinin to decrease inflammation and brain injury using a mouse model of infection-induced preterm birth. Fetal brains from C57BL/6 time mated mice treated with silibinin showed a decrease in LPS-induced IL-8 and ninjurin expression. However, silibinin did not change LPS-induced inflammation in placenta or myometrium.