1. Volume 123, Issue 4, Pages 1179-1190 (October 2002)
Intestinal infection with Giardia spp. reduces epithelial barrier function in a myosin light chain kinase–dependent fashion 
Kevin G.–E. Scott, Jonathon B. Meddings, David R. Kirk, Susan P. Lees–Miller, André G. Buret 
Gastroenterology 
Volume 123, Issue 4, Pages (October 2002)
DOI: /gast
Copyright © 2002 American Gastroenterological Association Terms and Conditions

2. Fig. 1
Jejunal colonization by G. muris trophozoites in immunocompetent euthymic (■) and nude athymic (▵) mice. No trophozoite colonization was observed in either control group at any time point during the study. Values are expressed as mean ± standard error under the mean log10 trophozoites per cm of jejunum. *P < 0.05 vs. euthymic-infected, **P < 0.01 vs. euthymic-infected; n = 14–28 per group at any time point.
Gastroenterology  , DOI: ( /gast )
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3. Fig. 2
Paracellular permeability in the small intestine (A; lactulose:mannitol FE), stomach (B; sucrose FE) and large intestine (C; sucralose FE) of euthymic (■) and athymic (▵) mice infected with G. muris. Values for infected animals are expressed as percent control which was calculated and as mean ± standard error under the mean. *P < 0.05 euthymic-infected vs. euthymic control, §P < 0.05 athymic infected versus athymic control, n = 6–16 per group at any time point.
Gastroenterology  , DOI: ( /gast )
Copyright © 2002 American Gastroenterological Association Terms and Conditions

4. Fig. 3
Paracellular permeability of nontransformed human intestinal SCBN monolayers, measured as transepithelial flux of FITC-dextran Values were obtained from control monolayers, monolayers incubated with G. lamblia S2 sonicates for 2 hours, monolayers exposed to S2 sonicates and coincubated with the MLCK inhibitor ML-9, and monolayers incubated with ML-9 alone. Values are expressed as mean ± standard error under the mean of the percent apical fluorescence that crossed 1 cm2 of the epithelial barrier into the basolateral chamber in 1 hour. *P < 0.05 vs. control, §P < 0.05 vs. S2; n = 12 per group.
Gastroenterology  , DOI: ( /gast )
Copyright © 2002 American Gastroenterological Association Terms and Conditions

5. Fig. 4
(A) Phosphorylation of MLC [(B) density of MLC phosphorylation normalized to the amount of protein evident by (C) immunoblot density] by MLCK is increased after a 2-hour exposure of nontransformed human intestinal monolayers to G. lamblia sonicates when compared with controls. This phosphorylation of MLC is prevented by addition of the MLCK inhibitor ML-9. ML-9 alone had no effect on the degree of phosphorylation compared with controls. A representative autoradiograph (B) and immunoblot (C) is also shown. Values are expressed as mean ± standard error under the mean of the ratio between autoradiograph density and immunoblot density. *P < 0.05 vs. control, §P < 0.05 vs. S2; n = 6 per group.
Gastroenterology  , DOI: ( /gast )
Copyright © 2002 American Gastroenterological Association Terms and Conditions

6. Fig. 5
Representative photomicrographs of nontransformed human intestinal SCBN monolayers stained for cytoskeletal F-actin. Preparations of monolayers included (A) unmanipulated control monolayers, (B) monolayers exposed to Giardia lamblia S2 sonicates for 2 hours, (C) monolayers coincubated with S2 sonicates and the MLCK inhibitor ML-9 for 2 hours, and (D) ML-9 alone. (A) In control monolayers, F-actin appears concentrated at the periphery of the enterocytes and spreads diffusely through the cytoplasm. (B) Exposure to G. lamblia S2 sonicates causes a characteristic flocculation of F-actin. (C) Coincubation of G. lamblia sonicates and ML-9 totally inhibited the reorganization of F-actin. (D) ML-9 alone has no effect on cytoskeletal F-actin. All micrographs were obtained at the same magnification (400×).
Gastroenterology  , DOI: ( /gast )
Copyright © 2002 American Gastroenterological Association Terms and Conditions

7. Fig. 6
(A) Quantification of F-actin flocculation and (B) ZO-1 relocation. Coincubation of SCBN monolayers with G. lamblia sonicates causes an increase in both F-actin flocculation and ZO-1 relocation. The MLCK inhibitor ML-9 prevents these cytoskeletal and tight junctional alterations when coincubated with Giardia and shows no effect alone. Values are expressed as mean ± standard error under the mean of the number of cells showing the abnormality out of 100 epithelial cells (%). *P < 0.05 vs. control; §P < 0.05 vs. S2; n = 6 per group, each from a separate monolayer.
Gastroenterology  , DOI: ( /gast )
Copyright © 2002 American Gastroenterological Association Terms and Conditions

8. Fig. 7
Representative confocal laser scanning micrographs of non-transformed human intestinal monolayers stained for tight junctional ZO-1. (A) Preparations included unmanipulated control monolayers, (B) monolayers exposed to G. lamblia S2 sonicates for 2 hours, (C) monolayers coincubated with G. lamblia sonicates and the MLCK inhibitor ML-9 for 2 hours, or (D) ML-9 alone. (A) Typical distribution of ZO-1 in control monolayers can be seen at the periphery of the enterocytes. (B) Exposure to G. lamblia sonicates for 2 hours causes a perinuclear relocation of ZO-1, an (C) effect prevented by coincubation of the MLCK inhibitor ML-9. (D) ML-9 alone has no effects on cellular arrangement of ZO-1. All micrographs were obtained at the same magnification (400×).
Gastroenterology  , DOI: ( /gast )
Copyright © 2002 American Gastroenterological Association Terms and Conditions

9. Fig. 8
Ratios of ZO-1 band intensities from insoluble (membrane-associated) versus soluble (cytosolic) in SCBN monolayers (A) exposed to vehicle medium, G. lamblia sonicates, Giardia sonicates plus the specific MLCK inhibitor ML-9, or ML-9 alone. Giardia decreases the ZO-1 membrane:cytosol ratio by reducing ZO-1 in the insoluble membrane fraction. This relocation of ZO-1 was inhibited by coincubation with ML-9. Representative immunoblots of ZO-1 from (B) insoluble and (C) soluble fractions are also shown.
Gastroenterology  , DOI: ( /gast )
Copyright © 2002 American Gastroenterological Association Terms and Conditions