Activated neutrophils induce an hMSH2-dependent G2/M checkpoint arrest and replication errors at a (CA)13-repeat in colon epithelial cells

Abstract

Objective:Chronic inflammation in ulcerative colitis is associated with increased risk for colorectal cancer. Its molecular pathway of cancer development is poorly understood. We investigated the role of neutrophil-derived cellular stress in an in vitro model of neutrophils as effectors and colon epithelial cells as targets, and tested for changes in cell cycle distribution and the appearance of replication errors.Design:Colon epithelial cells with different mismatch repair phenotypes were co-cultured with activated neutrophils. Target cells were analysed for cell cycle distribution and replication errors by flow cytometry. Changes in nuclear and DNA-bound levels of mismatch repair- and checkpoint-related proteins were analysed by western blotting.Results:Activated neutrophils cause an accumulation of target cells in G2/M, consistent with the installation of a DNA-damage checkpoint. Cells that do not express hMSH2, p53 or p21waf1/cip1 failed to undergo the G2/M arrest. Phosphorylation of p53 at site Ser15 and Chk1 at Ser317, as well as accumulation of p21waf1/cip1, was observed within 8–24 h. Superoxide dismutase and catalase were unable to overcome this G2/M arrest, possibly indicating that neutrophil products other than superoxide or H2O2 are involved in this cellular response. Finally, exposure to activated neutrophils increased the number of replication errors.Conclusions:By using an in vitro co-culture model that mimics intestinal inflammation in ulcerative colitis, we provide molecular evidence for an hMSH2-dependent G2/M checkpoint arrest and for the presence of replication errors.

Correlation between cell cycle proteins and hMSH2 in actinic cheilitis and lip cancer

“…Activation of p53 induces the expression of several genes, including cyclindependent kinase inhibitor p21 WAF1/CIP1 gene (p21) [18,19]. The DNA alkylating agents induce the phosphorylation and activation of p53, leading to an increase in p21 expression [6]. Thus, p21 may suppress tumors by its function in mediated p53-dependent G1 growth arrest [20].…”

Inflammation and skeletal metastasis

“…For example, reactive oxygen species (ROS), proteases, inflammatory cytokines and other detrimental factors released by macrophages and neutrophils at sites of inflammation have the potential to damage surrounding tissue by introducing genetic and epigenetic alterations in critical tumor suppressors or proto-oncogenes such as tumor protein p53 (TP53), phosphatase and tensin homolog (PTEN) and MYC. 5,[9][10][11][12][13] On the other hand, the activation of oncogenes and other cancerpromoting mutations or genetic rearrangements results in the induction of transcriptional programs that promote an inflammatory microenvironment. [14][15][16] When the tumor is incipient, both antitumorigenic and protumorigenic inflammatory signaling concur, but tumor progression results in the predominance of protumorigenic molecules that shape the microenvironment and significantly contribute to the cancer hallmark potentials.…”

MSH2 is required for cell proliferation, cell cycle control and cell invasiveness in colorectal cancer cells

“…ATM (ataxia-telangiectasia-mutated kinase), CHK1 and CHK2 have all been implicated in triggering the G2/M checkpoint response, and MMR genes are presumed to play a role in the G2/M cell cycle checkpoint [18]. Campregher et al [19] used an in vitro co-culture model that mimicked intestinal inflammation in ulcerative colitis, in which the activated neutrophils caused an accumulation of target cells in G2/M, which was consistent with the installation of a DNA-damage checkpoint. Cells that did not express MSH2, p53 or p21 waf1/cip1 failed to undergo the G2/M arrest.…”