LAMC1, Upregulated by TGFβ in Tumor Cells, Contributed to The Formation of Inflammatory Cancer-Associated Fibroblasts Via NF-kB/CXCL1/STAT3 in Esophageal Squamous Cell Carcinoma.

Abstract Background: The tumor microenvironment (TME) consists of a variety of cells that interact with each other through cytokines. As an important member of the TME, cancer-associated fibroblasts (CAFs) play an important role in the development of tumor cells, which is influenced by the heterogeneity of CAFs. Transforming growth factor β (TGFβ) not only plays a dual role in the progression of tumor cells directly but also influences tumor cells by regulating the heterogeneity of CAFs. Methods: we explored oncogenes regulated by TGFβ, which were involved in signaling molecules and interactions in the TME. We analyzed sequencing data of TCGA and GSE53625, as well as ESCC cell lines with or without TGFβ1 stimulation, and then we focused on laminin subunit gamma 1 (LAMC1). The upregulation of LAMC1 after TGF-β1 stimulation was examined by western blot (WB), quantitative real time PCR (qRT-PCR) and Chromatin immunoprecipitation (ChIP). We performed gain-of-function and loss-of-function assays to examine the effect of LAMC1 on proliferation and migration of ESCC cells. CAFs were isolated and cocultured with ESCC cells. And conditional medium of shLAMC1 ESCC cells and CAFs with different treatments were collected. RNA-seq of those cells were also performed. Luminex liquid suspension chip detection, ELISA, WB, qRT-PCR and rescue experiments were carried out to reveal the interaction of between ESCC cells and CAFs. Results: LAMC1 was highly expressed in ESCC, affecting the prognosis of patients. Moreover, LAMC1 could be upregulated by TGFβ1 through SMAD4 and SP1 synergistic activation. Further experiments showed that LAMC1 would promote the proliferation and migration of tumor cells mainly via Akt/NF-κB/MMP9 and MMP14. Additionally, LAMC1 would promote CXCL1 secretion mainly by activating NF-κB. Tumor-secreted CXCL1 remodeled the formation of inflammatory CAFs (iCAFs) through CXCR2/pSTAT3. The conditioned medium of iCAFs promoted the proliferation and migration of tumor cells. Conclusions: Our study identified the mechanism by which upregulation of LAMC1 by TGFβ in tumor cells not only promoted ESCC progression but also indirectly induced carcinogenesis by stimulating CXCL1 secretion and promoting the formation of iCAFs. This suggests that LAMC1 could be a potential therapeutic target and prognostic marker for ESCC.