Generation of Cell Models with Stable Expression of a Caspase-3 Activity Fluorescent Sensor.

Abstract

A lentiviral vector was created for inserting the commercial fluorogenic protease biosensor for caspase-3, ZipGFP-Casp3, into cell cultures. Three cultures of mesenchymal stem cells (MSCs) from the liver and placenta, dermal fibroblasts, and colorectal carcinoma clones of HT-29 and Caco2 cell lines with knockout of the main cancer stem cell marker CD133, along with control clones of these lines, were selected as the target cultures for studying apoptosis. All cultures were transduced with the ZipGFP-Casp3 sensor. The transduced cells were sorted, and in some derived cultures, cells undergoing spontaneous apoptosis and showing stable GFP fluorescence were noted. Additionally, experiments were conducted to study the sensitivity of transduced placental mesenchymal stem cells, as well as clones of the HT-29 line with CD133 knockout and its control clone, to classical apoptosis inducers staurosporine and TRAIL. The obtained results confirm that the derived cell cultures are good models for studying apoptosis in MSC and in the cancer stem cell population.