Figure 6.

TM9SF4-shRNA1 aggravated the starvation-induced cell death in HEK293 cells. HEK293 cells were stably transfected with lenti-scrambled-shRNA or lenti-TM9SF4-shRNA1. Amino acid starvation was carried out in EBSS for 2−6 h in (a), or 1−3 h in (b and c). (a) Cell viability assay by MTT. The data are expressed as the percent of metabolic activity relative to non-starved cells. (b and c) Cell death assay based on Apopxin green indicator and 7-amino-actinomycin (7-AAD) using Flow Cytometry. Apopxin green(−)/7-AAD (+) cells (Q1) were considered necrotic; Apopxin green(+)/7-AAD(+) double-positive cells (Q2) were considered late apoptotic; Apopxin green(−)/7-AAD(−) cells (Q3) were considered alive; and Apopxin green(+)/7-AAD(−) cells (Q4) were considered early apoptotic. Quantitative analysis of the apoptotic (Q2+Q4) cells by fluorescence-activated cell sorting (FACS) analysis is summarized in (c). Summary data are presented as mean±S.E.M. (n=7 in a; n=4 in c). *P<0.05