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. 2012 Jun;80(6):2165–2176. doi: 10.1128/IAI.06389-11

Fig 5.

Fig 5

EhMSP-1 silencing reduces E. histolytica cytopathic effect and enhances phagocytic ability. Cytotoxic ability is unaffected. (A) Effect of EhMSP-1 silencing on amebic lysis of lymphocytes, as reflected by release of the fluorescent dye BCECF (ameba/Jurkat cell ratio = 1:10, 30 min, 37°C) (mean and SE relative to G3 empty vector control, combined data from two experiments, n = 6). (B) Effect of EhMSP-1 silencing on amebic phagocytosis of live and apoptotic Jurkat lymphocytes. Phagocytosis of fluorescently labeled healthy or apoptotic Jurkat lymphocytes by EhMSP-1-silenced and control trophozoites was assayed by flow cytometry and expressed as the phagocytic index (ameba/Jurkat cell ratio = 1:2, 20 min, 37°C) (mean and SE relative to G3 empty vector controls, representative data from three experiments, n = 3). *, P = 0.01; **, P = 0.003. (C) Effect of EhMSP-1 silencing on amebic destruction of host cell monolayers. EhMSP-1-silenced (■) or G3 empty vector control (○) trophozoites (1 × 105/well) were added to confluent CHO cell monolayers and incubated at 37°C for the indicated times, after which the cell monolayers were washed, fixed, and stained with methylene blue. After washing, the remaining dye was extracted and quantified by measuring absorbance (600 nm) with a spectrophotometer. Data were expressed as the percentage of monolayer remaining relative to monolayers incubated with G3 empty vector control cells for 20 min (mean and SE, combined data from three independent experiments, n = 9). *, P = 0.005; **, P = 0.0003. (D) Effects of phenanthroline (100 μM) and E64 (100 μM) on tissue culture monolayer destruction by wild-type E. histolytica trophozoites. HM-1:IMSS strain E. histolytica trophozoites (1 × 105/well) were added to confluent CHO cell monolayers and incubated at 37°C for 40 min, after which cell monolayer destruction was assayed as for panel C. Data were expressed as the percentage of readings from monolayers to which no trophozoites were added (mean and SE, combined data from two independent experiments, n = 6). *, P = 0.05.