Cancer-associated Fibroblasts Promote Cancer Stemness In Lung Cancer By Activation Of H3K18ac

Background:Lung cancer is one of the most common malignant tumors. It is also one of the tumors with the highest death rate in the world. With the development of cancer research, researchers found that Cancer microenvironment (CME) plays a very important role in the tumor occurrence and development. Cancer associated fibroblasts (CAFs),as important stromal element in the tumor microenvironment, provide effective support for the occurrence, development and metastasis of the tumors. Researches targeting on CAFs can provide a new strategy for tumor therapy.The occurrence and development of tumors is not only dependent on genetic factors, but also affected by epigenetic modification. Epigenetic modification includes DNA methylation, histone modification, chromatin remodeling and so on. Acetylation is one of the important modifications of histone proteins, which can lead to changes in the structure of chromosomes and the level of gene transcription, driving tumorigenesis.So far, researches on CAFs in lung cancer are mainly focused on the genetic level, while the epigenetic changes are poorly studied. In this study, we explore the effect of lung-cancer-associated fibroblasts on non-small cell lung cancer and its related mechanisms,including detecting the change on histone acetylation.Methods:1. Separate and purify CAFs and NFsHuman peritumoral and NSCLC tissues were minced with scalpels and enzymatically dissociated by collagenase IV. The suspension was then centrifugated to separate the epithelial and fibroblast cells. The primarily isolated fibroblasts were purified by using Anti-Epcam beads.2. Establish a co-culture system of CAFs and lung cancer cellsWe use CAFs to co-culture with lung adenocarcinoma cell lines in transwell for 3 days. Real time PCR, Western blot was used to detect the ips factors. Flow cytometry was used to detect the stem cell markers. The global changes of several histone acetylation marks in CAF co-cultured lung cancer cells were examined.3. We use western blot and immunohistochemistry to detect the expression of H3K18ac in specimens.4.We establish the H3K18A mutanted lung cancer cells,which the histone 18 lysine residues were mutated into alanine (H3K18A), and detect the change of sternness.Results:1.CAFs have a high expression of a-SMA and FSP-1 compared to NFs.2.CAFs promote the stemness and tumorigenicity of lung cancer cell lines.3.The expression H3K18ac was enhanced in lung cancer cell lines after co-culture with CAFs.H3K18ac is high expressed in lung adenocarcinoma.The H3 mutant K18A lung cancer cells were low in expression of stemness markers, epithelial markers compared to H3 wild type lines when co-cultured with CAF.6. TGF-β can induce the high expression of H3K18ac in lung cancer cells.Conclusions:CAF enhanced H3K18ac expression in lung cancer through secreting TGF-β, leading to the enhancement of sternness, eventually promoting tumor progression. H3K18ac was a potential essential epigenetic regulatory mechanism for CAF induced lung cancer progression.