Probiotic Lactobacillus rhamnosus GR-1 is a unique prophylactic agent that suppresses infection-induced myometrial cell responses

Bona Kim,Oksana Shynlova,Stephen Lye

doi:10.1038/s41598-019-41133-0

Abstract

Preterm birth (PTB) is a multifactorial syndrome affecting millions of neonates worldwide. Intrauterine infection can induce PTB through the secretion of pro-inflammatory cytokines and untimely activation of uterine contractions. In pregnant mice, prophylactic administration of probiotic Lactobacillus rhamnosus GR-1 supernatant (GR1SN) prevented lipopolysaccharide (LPS)-induced PTB and reduced cytokine expression in the uterine muscle (myometrium). In this study we sought to delineate the mechanisms by which GR1SN suppressed cytokine secretion in the myometrium. We observed that L. rhamnosus GR-1 uniquely secretes heat-resistant but trypsin-sensitive factors, which significantly suppressed LPS-induced secretion of pro-inflammatory cytokines IL-6, IL-8, and MCP-1 in the human myometrial cell line, hTERT-HM. This effect was unique to GR1SN and could not be replicated using supernatant derived from non-GR-1 commensal lactobacilli species: L. rhamnosus GG, L. lactis, L. casei, or L. reuteri RC-14. Furthermore, pre-incubation of hTERT-HM cells with low-dose Pam3CSK (a TLR1/2 synthetic agonist which mimics LPS action) prior to LPS administration also significantly decreased LPS-induced cytokine secretion. This study highlights the distinct capacity of protein-like moieties secreted by L. rhamnosus GR-1 to inhibit pro-inflammatory cytokine production by human myometrial cells, potentially through a TLR1/2-mediated mechanism.

Highlights

Preterm birth (PTB; birth of an infant prior to 37 weeks of gestation) is a global medical concern that affects 13 million mothers and infants, annually
Because of high patient-to-patient sample variation in cytokine expression, we chose to use a human myometrial cell line for the current study. hTERT-HM cells stimulated with LPS (100 ng/mL) secreted levels of IL-6, IL-8, and MCP-1 proteins (Fig. 1, n = 7) that were comparable to levels secreted by primary myometrial cells
Pre-treatment of hTERT-HM cells with low dose GR-1 supernatant (GR1SN)-MRS (1% v/v) for 16 hours prior to LPS significantly suppressed secretions of IL-6 (31%, P < 0.01), IL-8 (54%, P < 0.001), and MCP-1 (65.5%, P < 0.001) compared to cells exposed to LPS only (Fig. 2, n = 9)

Summary

Introduction

Preterm birth (PTB; birth of an infant prior to 37 weeks of gestation) is a global medical concern that affects 13 million mothers and infants, annually. Prophylactic administration of GR-1 supernatant (GR1SN) to pregnant mice resulted in significant delay in lipopolysaccharide (LPS)-induced PTB which was associated with reduced expression of multiple pro-inflammatory cytokines IL-6, TNF-α, CSF-2, IL-3, IL-9, IL-12, IL-13, and IL-17 by pregnant uterine tissues, including the myometrial smooth muscle layer (Yang et al, 2014). In vitro GR1SN exposure to human macrophages resulted in increased expression of granulocyte-colony stimulating factor (G-CSF/CSF3) and reduced secretion of the pro-inflammatory cytokine, tumor-necrosis factor (TNF-α)[12]. Premature induction of pro-inflammatory cytokines, including TNF-α and IL-6, and chemokines such as IL-8 and MCP-1 by local or systemic infection can initiate pro-labour pathways, through increased expression of uterotonins Using a human myometrial cell line (hTERT-HM), we examined in vitro whether GR1SN can abrogate the LPS-induced inflammatory response mediated through TLR signalling

Methods

Results

Conclusion