Kupffer Cells Abrogate Cholestatic Liver Injury in Mice Stephan Gehring, Elizabeth M. Dickson, Maryann E. San Martin, Nico van Rooijen, Elaine F. Papa, Mark W. Harty, Thomas F. Tracy, Stephen H. Gregory  Gastroenterology  Volume 130, Issue 3, Pages 810-822 (March 2006) DOI: 10.1053/j.gastro.2005.11.015 Copyright © 2006 American Gastroenterological Association Institute Terms and Conditions

Figure 1 (A) BDLs or sham operations were performed on mice that were not treated or were Kupffer cell–depleted by inoculation intravenously with Cl2MDP-L 3 days previously. On day 7 postsurgery, representative liver sections were prepared, stained with H&E, and examined microscopically (magnification 40×). (B) Stained liver section derived from a Kupffer cell–depleted, bile duct–ligated mouse (upper left) was magnified to show a necrotic field (200×, upper right panel), proliferating bile ducts (200×, lower left), and infiltrating leukocytes including neutrophils (white arrowheads) (400×, lower right). Gastroenterology 2006 130, 810-822DOI: (10.1053/j.gastro.2005.11.015) Copyright © 2006 American Gastroenterological Association Institute Terms and Conditions

Figure 2 Kupffer cell depletion exacerbates liver injury. Trichrome-stained liver sections derived from groups composed of 5 untreated or Kupffer cell–depleted mice on day 7 after BDL were subjected to photoimage analysis. Percent damaged area/total area ± SD was calculated from 4 mice treated comparably. A second experiment yielded similar results. Values are significantly different (P = .019; nonpaired Student t test). Gastroenterology 2006 130, 810-822DOI: (10.1053/j.gastro.2005.11.015) Copyright © 2006 American Gastroenterological Association Institute Terms and Conditions

Figure 3 LCM of Kupffer cells. Kupffer cells, identified by the presence of carbon particles in the livers of mice injected intravenously with India ink diluted 1:100 in saline 24 hours previously (top half), were obtained by LCM (bottom half). Gastroenterology 2006 130, 810-822DOI: (10.1053/j.gastro.2005.11.015) Copyright © 2006 American Gastroenterological Association Institute Terms and Conditions

Figure 4 Carbon-labeled liver cells obtained by LCM express Kupffer cell–specific mRNA transcripts. The Kupffer cell receptor and CSF-1 receptor transcripts extracted from the carbon-labeled cells and from total liver section scrapes derived from 3 separate mice were quantified by real-time reverse-transcription PCR. *Significantly more than extracted from total tissue scrapes (P < .001; nonpaired Student t test). Gastroenterology 2006 130, 810-822DOI: (10.1053/j.gastro.2005.11.015) Copyright © 2006 American Gastroenterological Association Institute Terms and Conditions

Figure 5 IL-6 message expression by Kupffer cells is increased in cholestatic livers. Kupffer cells were obtained by LCM from the livers of 3 BDL or 3 sham-operated mice at the times indicated. IL-6 transcripts extracted from Kupffer cells and from total liver section scrapes were quantified by real-time reverse-transcription PCR. *Significantly greater than all other groups (P < .05; 1-way analysis of variance). Gastroenterology 2006 130, 810-822DOI: (10.1053/j.gastro.2005.11.015) Copyright © 2006 American Gastroenterological Association Institute Terms and Conditions

Figure 6 Liver injury is increased in bile duct-ligated, IL-6–deficient (IL-6−/−) mice. Photoimage analysis of trichrome-stained liver sections prepared on day 3 postsurgery. Values (percent damaged area/total area ± SD) were calculated from 6 mice treated comparably; a second experiment yielded similar results. *Significantly more tissue injury than sham-operated, IL-6–deficient animals (P = .001; nonpaired Student t test). Gastroenterology 2006 130, 810-822DOI: (10.1053/j.gastro.2005.11.015) Copyright © 2006 American Gastroenterological Association Institute Terms and Conditions

Figure 7 Recombinant IL-6 protects Kupffer cell–depleted mice from cholestatic liver injury. Groups of mice not treated or rendered Kupffer cell–depleted 3 days previously were subjected to BDL or sham operations. Animals in 1 group (BDL + recombinant murine IL-6) received 2 μg of recombinant murine IL-6 (Peprotech Inc., Rocky Hill, NJ) inoculated subcutaneously at the base of the tail 1 hour before surgery. (A) Sera were collected on day 3 postsurgery and ALT levels were quantified. (B) Photoimage analysis of trichrome-stained liver dissected on day 3 postsurgery. Values are the means ± SD derived from 6 mice treated comparably; similar results were obtained in a second experiment. BDL, Kupffer cell–depleted group administered recombinant murine IL-6 is significantly less than comparable group not administered IL-6: *P = .002 (nonpaired Student t test); **P = .029 (Mann–Whitney rank-sum test). Gastroenterology 2006 130, 810-822DOI: (10.1053/j.gastro.2005.11.015) Copyright © 2006 American Gastroenterological Association Institute Terms and Conditions