Defects at the Posttranscriptional Level Account for the Low TCRζ Chain Expression Detected in Gastric Cancer Independently of Caspase-3 Activity

Abstract

Background. Reduced TCRζ chain surface has been reported in T cells from patients with different inflammatory conditions and cancer. However, the causes of this diminished expression in cancer remain elusive. Methods. T cell-enriched populations of blood or tissue (tumoral and nontumoral) origin from 44 patients with gastric adenocarcinoma and 33 healthy subjects were obtained. Samples were subjected to cytofluorimetry, Western blot analysis, TCRζ cDNA sequencing experiments, measurement of TCRζ mRNA levels, and caspase-3 activity assays. Results. Cytofluorimetry revealed a decreased TCRζ expression in T cells of patients, assessed either as percentage of cells expressing this chain (blood: control subjects $99.8\pm 0.1\%$ , patients $98.8\pm 1.1\%$ $P<0.001$ ; tissue: control subjects $96.7\pm 0.9\%$ , patients tumoral tissue $67.9\pm 27.0\%$ , patients nontumoral tissue $82.8\pm 12.6\%$ , $P=0.019$ ) or mean fluorescence intensity (MFI) value (blood: control subjects $102.2\pm 26.0$ ; patients $58.0\pm 12.3$ , $P=0.001$ ; tissue: control subjects $99.4\pm 21.4$ ; patients tumoral tissue $41.6\pm 21.4$ ; patients nontumoral tissue $62.3\pm 16.6$ , $P=0.001$ ). Other chains pertaining to the TCR-CD3 complex (CD3ε) showed no significant differences (MFI values). Subsequent TCRζ cDNA sequencing experiments or measurements of TCRζ mRNA levels disclosed no differences between patients and control subjects. Evaluation of caspase-3 activity showed higher levels in T cell extracts of patients, and this activity could be decreased by 70% with the use of the inhibitor Ac-DEVD-FMK, although CD3ζ expression levels did not recover. Conclusions. These results further place the defect responsible for the low TCRζ expression in cancer at the posttranscriptional level and suggests contrary to what has been proposed in other pathologies that elevated caspase-3 activity is not the causative agent.