Isolation and characterization of the primary epithelial breast cancer cells and the adjacent normal epithelial cells from Iranian women’s breast cancer tumors

Abstract

As an experimental model, most studies rely on established human cancer cell lines; however, some genetical or phenotypical differences exist between these cells and their original tumor. Therefore, primary cells isolated directly from tissue are believed to be more biologically relevant tools for studying human and animal biology. Here, we aimed to isolate primary epithelial cancer and normal cells from breast tumors of Iranian women, for the first time. Thus, we isolated the epithelial and fibroblast cells from biopsy samples of patients with breast cancer based on differential centrifugation followed by culture in selective media. Normal epithelial cells obtained from the tissue biopsy away from the core of the tumor, based on the pathological diagnosis. Flow cytometry analysis indicated the positive immunoreactivity of the isolated epithelial cells against CD24 and Epithelial Specific Antigen (ESA/EpCAM), while they displayed a concomitant low expression of CD44 and CD49f. In contrat to fibroblasts, the qPCR data indicated the expression of luminal intracellular cytokeratin (Ck18) in both normal and cancer epithelial cells, but there was no expression of myoepithelial/basal markers, CK5 and vimentin. The epithelial cancer cells were reactive to cytokeratin 19 (CK19) antibody, whereas the normal epithelial cells were not. The expression of calmodulin-like protein (CLP) was also lower in the cancer epithelial cells than in the normal ones. In conclusion, primary epithelial normal and cancer cells, in addition to the fibroblasts were isolated and characterized from breast tumor of Iranian patients; and CLP expression is suggested as a susceptibility marker for breast cancer screening.

Diagnostic and prognostic impact of cytokeratin 18 expression in human tumors: a tissue microarray study on 11,952 tumors

“…CK18 is an intermediate filament protein that forms heteropolymers with its co-expressed complementary type II keratin partner CK8, which assembles into keratin filaments—the major structural component in the cytoplasm of epithelial cells (Moll et al 1982 ; Fuchs and Weber 1994 ). CK18 is primarily expressed in single-layered or “simple” epithelial tissues of, for example, the liver, kidney, breast, prostate, gastrointestinal tract as well as in cancers arising from CK18 positive epithelial cells (Oshima et al 1996 ; Cajaiba et al 2006 ; Skinnider et al 2005 ; Faridi et al 2018 ). Beside the important structural function, CK18 was also shown to play a role in apoptosis (Caulin et al ( 2000 ); Gilbert et al 2001 ), cell cycle progression (Galarneau et al 2007 ), and cancer-related signaling pathways.…”

From Donor to the Lab: A Fascinating Journey of Primary Cell Lines

“…Mechanical disaggregation of tumour tissues comprises mincing with a scalpel, filtration through 50-100 µm mesh, homogenisation, vortexing, pipetting, or application of extreme osmolarity stress (Mitra et al, 2013). The most frequently used method for mechanical dissociation is cutting tissue samples with scissors or scalpel to the size of 1-2 mm 3 (Phuc et al, 2010;Ali et al, 2017;Faridi et al, 2018;Ghaderi et al, 2019). Specimens prepared this way can be subsequently placed in a Petri dish and cultured with an explant method, or mechanical disaggregation may be combined with an enzymatic process for better results (Nushtaeva et al, 2019).…”

Synthesis and cytotoxic activity of N-(4-bromo)-benzoyl-N’phenylthiourea and 4-(tert-butyl)-N-benzoylurea on primary cells of HER2-Positive breast cancer

“…Breast tissue obtained from the biopsy was isolated to obtain primary cells 25,26 . Then the isolated cells were identified using CD133 biomarkers by immunofluorescence to find out that the isolated cells were cancer cells.…”