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Genetic dissection of the cellular pathways and signaling mechanisms in modeled tumor necrosis factor-induced Crohn's-like inflammatory bowel disease.

Kontoyiannis D, Boulougouris G, Manoloukos M, Armaka M, Apostolaki M, Pizarro T, Kotlyarov A, Forster I, Flavell R, Gaestel M, Tsichlis P, Cominelli F, Kollias G - J. Exp. Med. (2002)

Bottom Line: Tissue-specific activation of the mutant TNF allele by Cre/loxP-mediated recombination indicated that either myeloid- or T cell-derived TNF can exhibit full pathogenic capacity.Interestingly, TNF-mediated intestinal pathology was exacerbated in the absence of MAPKAP kinase 2, yet strongly attenuated in a Cot/Tpl2 or JNK2 kinase-deficient genetic background.Our data establish the existence of redundant cellular pathways operating downstream of TNF in inflammatory bowel disease, and demonstrate the therapeutic potential of selective kinase blockade in TNF-mediated intestinal pathology.

View Article: PubMed Central - PubMed

Affiliation: Institute for Immunology, Biomedical Sciences Research Center "Al. Fleming," Vari 166-72, Greece. Division of Gastroenterology and Hepatology, University of Virginia Health Sciences Center, Charlottesville, VA 22906, USA.

ABSTRACT
Recent clinical evidence demonstrated the importance of tumor necrosis factor (TNF) in the development of Crohn's disease. A mouse model for this pathology has previously been established by engineering defects in the translational control of TNF mRNA (Tnf(Delta)(ARE) mouse). Here, we show that development of intestinal pathology in this model depends on Th1-like cytokines such as interleukin 12 and interferon gamma and requires the function of CD8(+) T lymphocytes. Tissue-specific activation of the mutant TNF allele by Cre/loxP-mediated recombination indicated that either myeloid- or T cell-derived TNF can exhibit full pathogenic capacity. Moreover, reciprocal bone marrow transplantation experiments using TNF receptor-deficient mice revealed that TNF signals are equally pathogenic when directed independently to either bone marrow-derived or tissue stroma cell targets. Interestingly, TNF-mediated intestinal pathology was exacerbated in the absence of MAPKAP kinase 2, yet strongly attenuated in a Cot/Tpl2 or JNK2 kinase-deficient genetic background. Our data establish the existence of redundant cellular pathways operating downstream of TNF in inflammatory bowel disease, and demonstrate the therapeutic potential of selective kinase blockade in TNF-mediated intestinal pathology.

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MK2 and JNK2 kinase signals associate with the modulation of intestinal mononuclear apoptosis in the context of TNF-mediated IBD. (A) Quantitation of the infiltrating and apoptotic cells and the percentage of apoptotic cells in the infiltrate (apoptotic index) in the lamina propria of signaling-deficient TnfΔARE/+ mice. *, values of statistical significance relative to the TnfΔARE/+ group. Representative photomicrographs of paraffin-embedded ileal sections from (B) TnfΔARE/+ mice bred into (C) MK2- or (D) Jnk2-deficient backgrounds followed by the in situ detection of apoptotic cells. Alkaline phosphatase–labeled apoptotic cells were visualized using Fast Blue substrate and counterstaining with Nuclear Fast Red. B–D, ×300.
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fig6: MK2 and JNK2 kinase signals associate with the modulation of intestinal mononuclear apoptosis in the context of TNF-mediated IBD. (A) Quantitation of the infiltrating and apoptotic cells and the percentage of apoptotic cells in the infiltrate (apoptotic index) in the lamina propria of signaling-deficient TnfΔARE/+ mice. *, values of statistical significance relative to the TnfΔARE/+ group. Representative photomicrographs of paraffin-embedded ileal sections from (B) TnfΔARE/+ mice bred into (C) MK2- or (D) Jnk2-deficient backgrounds followed by the in situ detection of apoptotic cells. Alkaline phosphatase–labeled apoptotic cells were visualized using Fast Blue substrate and counterstaining with Nuclear Fast Red. B–D, ×300.

Mentions: The absence of any significant alterations amongst the TnfΔARE/+ peripheral lymphocytes in the presence or absence of MK2 and JNK2 kinases suggested that these signals may affect events in the locality of the intestine. Recent evidence indicated a strong correlation between the death of lymphocytic (or other) infiltrates in the intestine and the modulation of intestinal inflammation (14). Thus, we counted the number of apoptotic cells in inflamed ileal sections from TnfΔARE/+ mice in the absence of MK2 and Jnk2, using a TUNEL Assay. To circumvent the problem of the difference in inflammation between these groups we selected ileal samples with similar inflammatory indices. More specifically, we compared the number of apoptotic cells in inflamed ilea from 3-mo-old TnfΔARE/+ mice, 2-mo-old TnfΔARE/+ mk2−/− mice, and 7-mo-old TnfΔARE/+ JNK2−/− mice bearing comparable numbers of LPMCs (Fig. 6 A). A high incidence of apoptotic cells was observed in the compartment of the LPMCs in TnfΔARE/+ ilea at the age of 3 mo. (Fig. 6, A and B). Strikingly, the number of positive apoptotic cells was significantly lower in TnfΔARE/+ mk2−/−–deficient ilea suggesting a significant inhibition of cell death in the absence of MK2 (Fig.6, A and C). In contrast, the number of apoptotic cells in TnfΔARE/+ JNK2−/− ilea was dramatically increased relative to the lower cell numbers in the infiltrate indicating an increased rate of apoptosis (Fig. 6, A and D). Comparison of the apoptotic indices between 3-mo-old TnfΔARE/+ ilea and 6-mo-old TnfΔARE/+ Tpl2−/− did not reveal any significant differences (not depicted). Overall, our data suggest that Tpl2 and JNK2 kinases favor, whereas the MK2 kinase opposes, the development of IBD.


Genetic dissection of the cellular pathways and signaling mechanisms in modeled tumor necrosis factor-induced Crohn's-like inflammatory bowel disease.

Kontoyiannis D, Boulougouris G, Manoloukos M, Armaka M, Apostolaki M, Pizarro T, Kotlyarov A, Forster I, Flavell R, Gaestel M, Tsichlis P, Cominelli F, Kollias G - J. Exp. Med. (2002)

MK2 and JNK2 kinase signals associate with the modulation of intestinal mononuclear apoptosis in the context of TNF-mediated IBD. (A) Quantitation of the infiltrating and apoptotic cells and the percentage of apoptotic cells in the infiltrate (apoptotic index) in the lamina propria of signaling-deficient TnfΔARE/+ mice. *, values of statistical significance relative to the TnfΔARE/+ group. Representative photomicrographs of paraffin-embedded ileal sections from (B) TnfΔARE/+ mice bred into (C) MK2- or (D) Jnk2-deficient backgrounds followed by the in situ detection of apoptotic cells. Alkaline phosphatase–labeled apoptotic cells were visualized using Fast Blue substrate and counterstaining with Nuclear Fast Red. B–D, ×300.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2196068&req=5

fig6: MK2 and JNK2 kinase signals associate with the modulation of intestinal mononuclear apoptosis in the context of TNF-mediated IBD. (A) Quantitation of the infiltrating and apoptotic cells and the percentage of apoptotic cells in the infiltrate (apoptotic index) in the lamina propria of signaling-deficient TnfΔARE/+ mice. *, values of statistical significance relative to the TnfΔARE/+ group. Representative photomicrographs of paraffin-embedded ileal sections from (B) TnfΔARE/+ mice bred into (C) MK2- or (D) Jnk2-deficient backgrounds followed by the in situ detection of apoptotic cells. Alkaline phosphatase–labeled apoptotic cells were visualized using Fast Blue substrate and counterstaining with Nuclear Fast Red. B–D, ×300.
Mentions: The absence of any significant alterations amongst the TnfΔARE/+ peripheral lymphocytes in the presence or absence of MK2 and JNK2 kinases suggested that these signals may affect events in the locality of the intestine. Recent evidence indicated a strong correlation between the death of lymphocytic (or other) infiltrates in the intestine and the modulation of intestinal inflammation (14). Thus, we counted the number of apoptotic cells in inflamed ileal sections from TnfΔARE/+ mice in the absence of MK2 and Jnk2, using a TUNEL Assay. To circumvent the problem of the difference in inflammation between these groups we selected ileal samples with similar inflammatory indices. More specifically, we compared the number of apoptotic cells in inflamed ilea from 3-mo-old TnfΔARE/+ mice, 2-mo-old TnfΔARE/+ mk2−/− mice, and 7-mo-old TnfΔARE/+ JNK2−/− mice bearing comparable numbers of LPMCs (Fig. 6 A). A high incidence of apoptotic cells was observed in the compartment of the LPMCs in TnfΔARE/+ ilea at the age of 3 mo. (Fig. 6, A and B). Strikingly, the number of positive apoptotic cells was significantly lower in TnfΔARE/+ mk2−/−–deficient ilea suggesting a significant inhibition of cell death in the absence of MK2 (Fig.6, A and C). In contrast, the number of apoptotic cells in TnfΔARE/+ JNK2−/− ilea was dramatically increased relative to the lower cell numbers in the infiltrate indicating an increased rate of apoptosis (Fig. 6, A and D). Comparison of the apoptotic indices between 3-mo-old TnfΔARE/+ ilea and 6-mo-old TnfΔARE/+ Tpl2−/− did not reveal any significant differences (not depicted). Overall, our data suggest that Tpl2 and JNK2 kinases favor, whereas the MK2 kinase opposes, the development of IBD.

Bottom Line: Tissue-specific activation of the mutant TNF allele by Cre/loxP-mediated recombination indicated that either myeloid- or T cell-derived TNF can exhibit full pathogenic capacity.Interestingly, TNF-mediated intestinal pathology was exacerbated in the absence of MAPKAP kinase 2, yet strongly attenuated in a Cot/Tpl2 or JNK2 kinase-deficient genetic background.Our data establish the existence of redundant cellular pathways operating downstream of TNF in inflammatory bowel disease, and demonstrate the therapeutic potential of selective kinase blockade in TNF-mediated intestinal pathology.

View Article: PubMed Central - PubMed

Affiliation: Institute for Immunology, Biomedical Sciences Research Center "Al. Fleming," Vari 166-72, Greece. Division of Gastroenterology and Hepatology, University of Virginia Health Sciences Center, Charlottesville, VA 22906, USA.

ABSTRACT
Recent clinical evidence demonstrated the importance of tumor necrosis factor (TNF) in the development of Crohn's disease. A mouse model for this pathology has previously been established by engineering defects in the translational control of TNF mRNA (Tnf(Delta)(ARE) mouse). Here, we show that development of intestinal pathology in this model depends on Th1-like cytokines such as interleukin 12 and interferon gamma and requires the function of CD8(+) T lymphocytes. Tissue-specific activation of the mutant TNF allele by Cre/loxP-mediated recombination indicated that either myeloid- or T cell-derived TNF can exhibit full pathogenic capacity. Moreover, reciprocal bone marrow transplantation experiments using TNF receptor-deficient mice revealed that TNF signals are equally pathogenic when directed independently to either bone marrow-derived or tissue stroma cell targets. Interestingly, TNF-mediated intestinal pathology was exacerbated in the absence of MAPKAP kinase 2, yet strongly attenuated in a Cot/Tpl2 or JNK2 kinase-deficient genetic background. Our data establish the existence of redundant cellular pathways operating downstream of TNF in inflammatory bowel disease, and demonstrate the therapeutic potential of selective kinase blockade in TNF-mediated intestinal pathology.

Show MeSH
Related in: MedlinePlus