# Increased Toll-Like Receptor-4 Signalling in Breast Tissue of High Fibroglandular Density

## TL;DR

This study explores how increased TLR4 signaling in high-density breast tissue may contribute to inflammation and breast cancer risk.

## Contribution

The study identifies elevated TLR4 signaling and HMGB1 as potential drivers of inflammation in high fibroglandular density breast tissue.

## Key findings

- TLR4, MYD88, NFKB, and HMGB1 are more highly expressed in epithelial cells of high fibroglandular density tissue.
- LPS treatment increases expression of TNFA and CCL2 genes in mammary epithelial cell organoids.
- TLR4 signaling may mediate local inflammation in high-density breast tissue, with HMGB1 as a possible trigger.

## Abstract

High mammographic breast density relates to the abundance of fibroglandular tissue in comparison to fatty tissue in the breast and is associated with increased breast cancer risk. Chronic low-level inflammation has been implicated as a driver of high density and cancer risk, however little is understood of the underlying cause of inflammation. This research aimed to investigate the role of the innate immune recognition receptor toll-like receptor-4 (TLR4) in inflammation associated with high fibroglandular density. Immunohistochemical analysis was performed on paired breast tissue samples of high and low fibroglandular density tissue (n = 22 pairs) to investigate the expression of TLR4, TLR4 agonists lipopolysaccharide (LPS) and damage response protein high-mobility group protein 1 (HMGB1), as well as activation of downstream mediators myeloid differentiation primary response 88 (MYD88) and nuclear factor kappa B (NFKB). Mammary epithelial cell organoids (n = 5) were cultured in vitro with LPS to investigate the expression of genes associated with inflammation. TLR4 was primarily expressed in basal epithelial cells, stromal macrophages, and some expression was detected in luminal epithelial cells. Increased expression of TLR4, MYD88, NFKB, and HMGB1 was observed in epithelial cells in high fibroglandular density tissue. There was increased expression of the genes encoding inflammatory cytokines tumour necrosis factor alpha (TNFA) and C-C motif ligand 2 (CCL2) in mammary epithelial cell organoids treated with LPS. The TLR4 signalling pathway may be a mediator of local breast inflammation associated with regions of high breast density and the damage response protein HMGB1 may be a trigger for TLR4 activation.