High throughput screening of cytokines, chemokines and matrix metalloproteinases in wound fluid induced by mammary surgery

Abstract

Objective: To clarify the composition of wound fluid (WF) and investigate the impact of WF on breast cancer cell lines.

Methods: The proliferation and migration of WF-treated breast cancer cells MDA-MB-231 and MCF-7 were assessed with colony formation test, MTT cell proliferation test and scratch wound test. The quantitative profiles of WF were analyzed using Bio-Plex Pro kits.

Results: The proliferation and migration of WF-treated breast cancer cells were significantly higher than that of untreated cells. Fifteen cytokines, 29 chemokines and 9 matrix metalloproteinases (MMPs) were assessed in WF. The concentrations of these factors were influenced by post-surgery days, neoadjuvant chemotherapy (NAC), TNM stage, pathological type and molecular subtype. The WF harvested from patients underwent NAC showed significant higher profiles of interleukin-1β (IL-1β), IL-4, IL-6, IL-17F, IL-21, IL-23, IL-25, IL-31, Interferon γ (IFNγ), CD40 ligand (CD40L), tumor necrosis factor α (TNFα), CXCL1, CXCL2, CXCL5, CCL3, CCL7 and CCL20.

Conclusions: Surgery-induced WF promotes the proliferation and migration of breast cancer cells. The composition of WF is influenced by various clinical features and provides potential therapeutic targets to control local recurrence and tumor progression.

Keywords: breast cancer; proliferation; wound fluid.

Figure 1. Colony formation assays: WF promotes MCF-7 and MDA-MB-231 cell growth

Less colonies form (both MCF-7 and MDA-MB-231 cells) in the absence of WF compared to WF-treated groups. A. 0.1%, 0.5% and 1%WF promotes colony formation of MCF-7 cells. C. The numbers of colonies form by MDA-MB-231 cells in 0.5% and 1.0% WF-treated groups are significant higher than that in 0.1%WF-treated group and the control group. B. and D. No significant difference of colony numbers are observed between benign disease and breast cancer groups (both MCF-7 and MDA-MB-231 cells).

Figure 2. MTT proliferation assays: WF increases MCF-7 and MDA-MB-231 cell proliferation

A. and C. The proliferation rates of MCF-7 and MDA-MB-231 cells are significantly increased after treated with WF for 48 hours when compared to the control group. B. and D. No significant difference of proliferation rate is observed between benign disease and breast cancer groups (both MCF-7 and MDA-MB-231 cells). E. MDA-MB-231 cells treated with WF from breast cancer patients underwent neoadjuvant chemotherapy (NAC) show higher rate of proliferation.

Figure 3. Scratch wounds assays: WF promotes MDA-MB-231cells migration

Scratch wounds for MDA-MB-231 cells at 0 h and 24 h after introducing the wound and treated with WF are shown. Scratch closure change of control and WF groups for MDA-MB-231 cells was evaluated at 24 h. Scratch fold changes in 0.5% and 1% WF treated groups are significantly higher than that of the control group (P = 0.041 and P = 0.006). The scratch closure change of the cells treated with 0.5%WF was similar to that of 1% WF. Cells treated with 1% WF from breast cancer tend to migrate more rapidly than those treated with WF from benign disease (P = 0.017).