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Lack of CC chemokine ligand 2 differentially affects inflammation and fibrosis according to the genetic background in a murine model of steatohepatitis.

Galastri S, Zamara E, Milani S, Novo E, Provenzano A, Delogu W, Vizzutti F, Sutti S, Locatelli I, Navari N, Vivoli E, Caligiuri A, Pinzani M, Albano E, Parola M, Marra F - Clin. Sci. (2012)

Bottom Line: Sirius Red staining demonstrated pericellular fibrosis in zone 3, and image analysis showed a significantly lower matrix accumulation in CCL2-KO mice.This was associated with reduced hepatic expression of TGF-β (transforming growth factor-β), type I procollagen, TIMP-1 (tissue inhibitor of metalloproteinases-1) and α-smooth muscle actin.We conclude that, in experimental murine steatohepatitis, the effects of CCL2 deficiency are markedly dependent on the genetic background.

View Article: PubMed Central - PubMed

Affiliation: Dipartimento di Medicina Interna, University of Florence, Florence, Italy.

ABSTRACT
Expression of CCL2 (CC chemokine ligand 2) (or monocyte chemoattractant protein-1) regulates inflammatory cell infiltration in the liver and adipose tissue, favouring steatosis. However, its role in the pathogenesis of steatohepatitis is still uncertain. In the present study, we investigated the development of non-alcoholic steatohepatitis induced by an MCD diet (methionine/choline-deficient diet) in mice lacking the CCL2 gene on two different genetic backgrounds, namely Balb/C and C57/Bl6J. WT (wild-type) and CCL2-KO (knockout) mice were fed on a lipid-enriched MCD diet or a control diet for 8 weeks. In Balb/C mice fed on the MCD diet, a lack of CCL2 was associated with lower ALT (alanine transaminase) levels and reduced infiltration of inflammatory cells, together with a lower generation of oxidative-stress-related products. Sirius Red staining demonstrated pericellular fibrosis in zone 3, and image analysis showed a significantly lower matrix accumulation in CCL2-KO mice. This was associated with reduced hepatic expression of TGF-β (transforming growth factor-β), type I procollagen, TIMP-1 (tissue inhibitor of metalloproteinases-1) and α-smooth muscle actin. In contrast, in mice on a C57Bl/6 background, neither ALT levels nor inflammation or fibrosis were significantly different comparing WT and CCL2-KO animals fed on an MCD diet. In agreement, genes related to fibrogenesis were expressed to comparable levels in the two groups of animals. Comparison of the expression of several genes involved in inflammation and repair demonstrated that IL (interleukin)-4 and the M2 marker MGL-1 (macrophage galactose-type C-type lectin 1) were differentially expressed in Balb/C and C57Bl/6 mice. No significant differences in the degree of steatosis were observed in all groups of mice fed on the MCD diet. We conclude that, in experimental murine steatohepatitis, the effects of CCL2 deficiency are markedly dependent on the genetic background.

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CCL2 deficiency does not affect the development of fibrosis or the expression of pro-fibrogenic genes following administration of an MCD diet in C57Bl/6 miceWT or CCL2-KO mice were fed for 8 weeks on the control diet or the MCD diet. Groups are as in the legend to Figure 1. (A) Sirius-Red-stained slides were subjected to image analysis, and the area occupied by fibrotic tissue was measured in WT and CCL2-KO mice fed on a control diet or an MCD diet. (B–E) RNA was isolated from liver tissue and the expression of TGF-β (B), type I procollagen (C), TIMP-1 (D) and α-smooth muscle actin (E) were measured by RT-PCR. *P<0.01 compared with WT animals fed on the control diet; **P<0.01 compared with WT animals fed on the MCD diet.
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Figure 6: CCL2 deficiency does not affect the development of fibrosis or the expression of pro-fibrogenic genes following administration of an MCD diet in C57Bl/6 miceWT or CCL2-KO mice were fed for 8 weeks on the control diet or the MCD diet. Groups are as in the legend to Figure 1. (A) Sirius-Red-stained slides were subjected to image analysis, and the area occupied by fibrotic tissue was measured in WT and CCL2-KO mice fed on a control diet or an MCD diet. (B–E) RNA was isolated from liver tissue and the expression of TGF-β (B), type I procollagen (C), TIMP-1 (D) and α-smooth muscle actin (E) were measured by RT-PCR. *P<0.01 compared with WT animals fed on the control diet; **P<0.01 compared with WT animals fed on the MCD diet.

Mentions: We next evaluated whether a similar inflammatory response to the MCD diet was associated with a similar fibrogenic response in the absence of CCL2 in C57Bl/6 mice. Image analysis of the area stained with Sirius Red demonstrated a marked and significant increase in mice fed on the MCD diet compared with control animals (Figure 6A). However, no differences comparing WT mice and mice lacking CCL2 were observed. In agreement with this observation, intrahepatic expression of genes involved in fibrogenesis was not reduced in the absence of CCL2 in C57Bl/6 mice fed on the MCD diet, and the expression levels of α1-procollagen were even higher (Figures 6C–6E). Accordingly, α-smooth muscle actin expression, a marker of activated myofibroblasts, was similar irrespective of the presence of CCL2 (Figure 6E). These results demonstrate that the effects of CCL2 on hepatic necro-inflammation and fibrosis induced by the MCD diet are dependent on the mouse genetic background.


Lack of CC chemokine ligand 2 differentially affects inflammation and fibrosis according to the genetic background in a murine model of steatohepatitis.

Galastri S, Zamara E, Milani S, Novo E, Provenzano A, Delogu W, Vizzutti F, Sutti S, Locatelli I, Navari N, Vivoli E, Caligiuri A, Pinzani M, Albano E, Parola M, Marra F - Clin. Sci. (2012)

CCL2 deficiency does not affect the development of fibrosis or the expression of pro-fibrogenic genes following administration of an MCD diet in C57Bl/6 miceWT or CCL2-KO mice were fed for 8 weeks on the control diet or the MCD diet. Groups are as in the legend to Figure 1. (A) Sirius-Red-stained slides were subjected to image analysis, and the area occupied by fibrotic tissue was measured in WT and CCL2-KO mice fed on a control diet or an MCD diet. (B–E) RNA was isolated from liver tissue and the expression of TGF-β (B), type I procollagen (C), TIMP-1 (D) and α-smooth muscle actin (E) were measured by RT-PCR. *P<0.01 compared with WT animals fed on the control diet; **P<0.01 compared with WT animals fed on the MCD diet.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3369401&req=5

Figure 6: CCL2 deficiency does not affect the development of fibrosis or the expression of pro-fibrogenic genes following administration of an MCD diet in C57Bl/6 miceWT or CCL2-KO mice were fed for 8 weeks on the control diet or the MCD diet. Groups are as in the legend to Figure 1. (A) Sirius-Red-stained slides were subjected to image analysis, and the area occupied by fibrotic tissue was measured in WT and CCL2-KO mice fed on a control diet or an MCD diet. (B–E) RNA was isolated from liver tissue and the expression of TGF-β (B), type I procollagen (C), TIMP-1 (D) and α-smooth muscle actin (E) were measured by RT-PCR. *P<0.01 compared with WT animals fed on the control diet; **P<0.01 compared with WT animals fed on the MCD diet.
Mentions: We next evaluated whether a similar inflammatory response to the MCD diet was associated with a similar fibrogenic response in the absence of CCL2 in C57Bl/6 mice. Image analysis of the area stained with Sirius Red demonstrated a marked and significant increase in mice fed on the MCD diet compared with control animals (Figure 6A). However, no differences comparing WT mice and mice lacking CCL2 were observed. In agreement with this observation, intrahepatic expression of genes involved in fibrogenesis was not reduced in the absence of CCL2 in C57Bl/6 mice fed on the MCD diet, and the expression levels of α1-procollagen were even higher (Figures 6C–6E). Accordingly, α-smooth muscle actin expression, a marker of activated myofibroblasts, was similar irrespective of the presence of CCL2 (Figure 6E). These results demonstrate that the effects of CCL2 on hepatic necro-inflammation and fibrosis induced by the MCD diet are dependent on the mouse genetic background.

Bottom Line: Sirius Red staining demonstrated pericellular fibrosis in zone 3, and image analysis showed a significantly lower matrix accumulation in CCL2-KO mice.This was associated with reduced hepatic expression of TGF-β (transforming growth factor-β), type I procollagen, TIMP-1 (tissue inhibitor of metalloproteinases-1) and α-smooth muscle actin.We conclude that, in experimental murine steatohepatitis, the effects of CCL2 deficiency are markedly dependent on the genetic background.

View Article: PubMed Central - PubMed

Affiliation: Dipartimento di Medicina Interna, University of Florence, Florence, Italy.

ABSTRACT
Expression of CCL2 (CC chemokine ligand 2) (or monocyte chemoattractant protein-1) regulates inflammatory cell infiltration in the liver and adipose tissue, favouring steatosis. However, its role in the pathogenesis of steatohepatitis is still uncertain. In the present study, we investigated the development of non-alcoholic steatohepatitis induced by an MCD diet (methionine/choline-deficient diet) in mice lacking the CCL2 gene on two different genetic backgrounds, namely Balb/C and C57/Bl6J. WT (wild-type) and CCL2-KO (knockout) mice were fed on a lipid-enriched MCD diet or a control diet for 8 weeks. In Balb/C mice fed on the MCD diet, a lack of CCL2 was associated with lower ALT (alanine transaminase) levels and reduced infiltration of inflammatory cells, together with a lower generation of oxidative-stress-related products. Sirius Red staining demonstrated pericellular fibrosis in zone 3, and image analysis showed a significantly lower matrix accumulation in CCL2-KO mice. This was associated with reduced hepatic expression of TGF-β (transforming growth factor-β), type I procollagen, TIMP-1 (tissue inhibitor of metalloproteinases-1) and α-smooth muscle actin. In contrast, in mice on a C57Bl/6 background, neither ALT levels nor inflammation or fibrosis were significantly different comparing WT and CCL2-KO animals fed on an MCD diet. In agreement, genes related to fibrogenesis were expressed to comparable levels in the two groups of animals. Comparison of the expression of several genes involved in inflammation and repair demonstrated that IL (interleukin)-4 and the M2 marker MGL-1 (macrophage galactose-type C-type lectin 1) were differentially expressed in Balb/C and C57Bl/6 mice. No significant differences in the degree of steatosis were observed in all groups of mice fed on the MCD diet. We conclude that, in experimental murine steatohepatitis, the effects of CCL2 deficiency are markedly dependent on the genetic background.

Show MeSH
Related in: MedlinePlus