Little expression of cytokine mrna by fresh tumour-infiltrating mononuclear leukocytes from glioma and lung adenocarcinoma

We investigated whether cytokine genes were activated in human tumour-infiltrating mononuclear leukocytes (TIML) obtained from six lung adenocarcinomas and seven glioblastomas. TIML were extracted by mechanical disruption and isolated by double density gradient of Ficoll. We performed mRNA reverse transcription-polymerase chain reaction (RT-PCR) on these fresh (noncultured) TIML and autologous peripheral blood mononuclear leukocytes (PBML) using primers for the cytokines IL-1β, IL-6, IL-2, IL-4, GM-CSF, IFN-γ and TNF-β. In addition, we compared patients‘ TIML and PBML populations with healthy normal and α-CD3 activated PBML as an optimally activated reference population. Gel bands of RT-PCR products were quantitated in relative units (RU) as a function of their size and intensity by computerized image-analysis. Lung and brain patients‘ TIML showed IL-1β and IL-6 cytokine mRNA expressed in the average of 2-log RU but not significantly different from autologous and normal healthy PBML. IL-2, IFN-γ and TNF-β also did not appear expressed in the TIML at higher levels than in autologous or healthy normal PBML. However in two thirds of patients, lung TIML could be distinguished from autologous PBML by specific expression of GM-CSF and from healthy normal PBML by expression of IL-4. Similarly, most brain TIML expressed mRNA significantly above healthy normal PBML for GM-CSF and IL-4. In comparison with α-CD3 activated healthy PBML, our results suggest that lung and brain TIML had detectable cytokine mRNA, but they seemed poorly activated in total number of genes and amount of cytokine mRNA. Absence, insufficient amounts or deficient combinations of cytokine production by TIML may be factors that prevented an optimal activation of the immune response in these tumours.