Downregulation of Stemness Genes and Induction of Necrosis in Rat LA۷ Cancer Stem Cells Induced Tumors Treated with Starved Fibroblasts Culture Supernatant

Background: Stem cell differentiation therapy is a promising strategy in cancer treatment. we show that protein cocktail prepared from serum starved fibroblasts has therapeutic potential based on this strategy. Methods: The condition medium was prepared from foreskin isolated fibroblasts and analyzed by Liquid chromatography electrospray ionization mass spectrometry-mass spectrometry (LC-ESI-MS/MS). LA۷ mammary gland cancer stem cells originated tumors were induced in Sprague Dawley rats. The rats treated subcutaneously with DMEM (group A), condition medium (group B), or normal saline (group C) once daily for ۷ days. Then the tumors were removed and divided into the two parts, one part was used to quantify gene expression by stem-loop RT-qPCR assay and the other part was used for Hematoxylin & Eosin (H & E), Giemsa, and immunohistochemistry (IHC) staining. Results: All induced tumors appeared as sarcomatoid carcinoma (SC). Immunohistochemistry staining confirmed this conclusion by recognizing the tumor as Ki۶۷+, cytokeratin+, vimentine+, and estrogen receptor negative SC. RT-qPCR analysis revealed that Oct۴-, Sox-۲, Nanog- gene expression was much reduced in the condition medium treated tumors versus proper controls (p< ۰.۰۵). Tissue necrosis was more prevalent in this group while tumors volume was diminished almost by ۴۰%. The LC-ESI-MS/MS analysis unrevealed the stemness reducing and the cell death inducing proteins such as, pigment epithelium-derived factor (PEDF), insulin like growth factor binding protein-۵ (IGFBP-۵) and -۷ (IGFBP-۷) in the condition medium. Conclusions: This study showed that the substances released from starved human fibroblasts were able to down-regulate the stemness-related genes and induce necrosis in LA۷ derived tumors.

Keywords:

Breast cancer , Cancer Stem cells , Cell differentiation , Fibroblasts , Gene expression.