Optimising methods for human thyroid primary cell culture isolation

Abstract

According to the GLOBOCAN 2020 database on cancer incidence and mortality, thyroid cancer (TC) has the ninth-highest cancer in- cidence worldwide. Basic research contributes to developing diag- nostic and treatment alternatives. Cell models play a crucial role in vitro study of the thyroid. Permanent thyroid cell lines widely used in laboratory research typically originate from tumors. However, there is a need to compare these tumor cells with cells originating from normal thyroid tissue. Primary cell lines are invaluable for exploring cancer biology and investigating novel treatments. This study aims is optimize an isolation protocol of human primary thyroid cells from histologically non-tumor, tumor, and metastasis tissues. Primary cul- tures are laborious to obtain and then difficult to maintain in culture. We tried five different methods for isolating primary cells that com- bine mechanical disaggregation and enzymatic digestion. These methods use distinct enzymatic compositions, incubation times, and mechanical approaches, including centrifugation. We have ar- rived at a simple, rapid, and effective method to culture cells from thyroid biopsies for subsequent studies. Our previous results have revealed that GEF-H1 plays a pro-tumor role in different tumoral cell lines. We investigate the expression of GEF-H1, which belongs to the RhoA-GTPase activator family, in primary cells. Using immu- nofluorescence staining, we observed significantly higher GEF-H1 protein expression in cytokeratin-19 positive thyroid carcinoma cells compared to normal thyroid cells (p<0.0001). Our results propose a cheap and easy isolation thyroid primary cells protocol and suggest that GEF-H1 could be used as a potential tumor biomarker in TC.