Effects of hypoxia on interleukin-2 mRNA expression by T lymphocytes

Objective: To investigate the effects of hypoxia on T-lymphocyte expression of IL-2 messenger RNA (mRNA), after cell activation with phorbol ester and ionomycin. Design: Prospective, controlled, cellular trial. Setting: University research laboratory. Subjects: EL4.6.1 cells, a murine T- cell lymphoma line. Interventions: Tissue culture media was deoxygenated and flushed continuously with 100% helium to maintain a PO₂ of 30 to 40 torr (<40 torr [<5.3 kPa]), or flushed with 10% oxygen/90% helium to maintain a PO₂ of 45 to 55 torr (>45 torr [>6.0 kPa]). The pH was maintained between 7.3 and 7.6. The media was inoculated with EL4 cells. Aliquots of cells were obtained at intervals and divided into two groups: an immediate group, stimulated immediately, and an overnight group that was returned to normal incubator conditions of 5% CO₂/humidified room air for 18 hrs before stimulation. Measurements and Main Results: Gas tension, pH, cell count, and viability were determined for each aliquot. Cells were stimulated with phorbol myristate acetate and ionomycin for 4 hrs, at which time levels of interleukin-2 (IL-2) messenger RNA (mRNA) and gamma actin mRNA were measured by solution hybridization and enzyme immunoassay. The results were expressed as IL-2 mRNA/gamma actin mRNA ratio, normalized to baseline room air values. Cell viability and housekeeping functions (gamma actin mRNA expression) were unaffected by hypoxia. Cells exposed to a PO₂ of <40 torr (<5.3 kPa) demonstrated a dramatic reduction in IL-2 mRNA expression with increasing duration of hypoxia. These effects persisted after an 18-hr recovery period. There was no effect on IL-2 mRNA expression when cells were exposed to a PO₂ of >45 torr (>6.0 kPa). Conclusions: The regulation of IL-2 transcription in the T lymphocyte appears to be exquisitely sensitive to changes in oxygen tension. Exposure to a PO₂ of <40 torr (<5.3 kPa) causes prolonged impairment of IL-2 mRNA expression. IL-2 is an important growth factor for T and NK cells, and plays a pivotal role in the regulation of the host's immune response. The long-lasting effects of brief hypoxic exposure may, in part, explain the critically ill patient's predisposition to infectious complications.