Background. Glutamine is crucial in the growth and protection of intestinal mucosa; the molecular mechanisms for these effects are not known. Heat shock proteins (hsp) are stress-responsive genes that protect cells from various injuries. The purpose of this study was to determine whether glutamine alters hsp expression in intestinal cells after injury. Methods. IEC-6 (rat intestinal crypt) cells were grown to 80% confluence, rinsed free of medium and serum, and then grown in medium containing various concentrations of L-glutamine (0.5 to 8.0 mmol/L) for 24 hours. Cells were then exposed to heat shock (43°C for 1 hour) and RNA and protein were extracted at 0, 2, 8, and 24 hours after heat shock. Northern hybridizations were performed using an hsp 70 complementary DNA. Western blots were performed by using antibodies specific to the inducible and constitutive forms of hsp 70. Results. Northern blot analysis demonstrated a fourfold stimulation of hsp 70 messenger RNA 2 hours after heat shock in IEC-6 cells supplemented with 8.0 mmol/L glutamine compared with 0.5 and 1.0 mmol/L glutamine. In addition, inducible Hsp 70 protein levels were increased threefold in IEC-6 cells given glutamine (8.0 mmol/L) at 24 hours after heat shock. Conclusions. Glutamine augments hsp 70 messenger RNA and protein expression in gut-derived IEC-6 cells after injury. Our findings demonstrate a novel mechanism for the enhanced cytoprotection of gut mucosa conferred by administration of glutamine and suggest that supplemental glutamine may be required after injury to enhance expression of stress-responsive genes in the gut.