ANALYSIS OF EPITHELIAL-CELL STRESS-RESPONSE DURING INFECTION BY SHIGELLA-FLEXNERI

Shigella flexneri-infected macrophage cells undergo an apoptotic-like death as early as one hour after infection (A, Zychlinsky, M. C. Prevo st, and P. J. Sansonetti, Nature [London] 358:167-168, 1992). To deter mine the fate of infected epithelial cells, we characterized the viabi lity, morphology and several metabolic activities of HeLa cells after treatment with M90T, an invasive isolate of S. flexneri serotype 5, or BS176, a noninvasive isolate cured of the 220-kb virulence plasmid. U sing standard assays,we found that for at least 4 h after infection wi th M90T, HeLa cells remained viable and did not detach or lyse. The ul trastructural morphology of HeLa cells heavily infected with M90T was free of hallmarks associated with cells undergoing apoptosis. Consiste nt with the idea that intracellular bacterial growth is metabolically stressful to the host cell, we observed that, compared with BS176 trea ted-Hela cells, M90T-treated HeLa tells showed (i) a significant decre ase in the total pool size of nucleoside triphosphates, (ii) a reduced ability to incorporate extracellular radiolabeled methionine into the soluble and insoluble cell fractions, and (iii) a stimulation of gluc ose uptake. However, there was no detectable increase in expression of the stress-inducible hsp70 gene in M90T-infected HeLa cells or activa tion of the anaerobic metabolic pathway as determined by measuring tot al lactate levels. These results demonstrate clearly that the fate of S. flexneri-infected cells can vary dramatically between cell types an d agree with the hypothesis that the destruction of epithelial cells o bserved in experimental models of shigellosis is due to the host infla mmatory response and probably not bacterial intracellular multiplicati on per se.