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Pivotal role of dendritic cell-derived CXCL10 in the retention of T helper cell 1 lymphocytes in secondary lymph nodes.

Yoneyama H, Narumi S, Zhang Y, Murai M, Baggiolini M, Lanzavecchia A, Ichida T, Asakura H, Matsushima K - J. Exp. Med. (2002)

Bottom Line: Blockade of CXCL10 dramatically altered the distribution of cluster-forming BrdU+CD4+ T cells.BrdU+CD4+ T cells in the hepatic LNs were selectively diminished while those in the circulation were significantly increased by treatment with anti-CXCL10 monoclonal antibody.This was accompanied by accelerated infiltration of memory T cells into the periphery of hepatic granuloma sites, most of them were in cell cycle and further produced higher amount of IFN-gamma leading to exacerbation of liver injury.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Preventive Medicine, School of Medicine and Core Research and Evolutional Science and Technology (CREST), The University of Tokyo, Tokyo 113-0033, Japan.

ABSTRACT
Various immune diseases are considered to be regulated by the balance of T helper (Th)1 and Th2 subsets. Although Th lymphocytes are believed to be generated in draining lymph nodes (LNs), in vivo Th cell behaviors during Th1/Th2 polarization are largely unexplored. Using a murine granulomatous liver disease model induced by Propionibacterium acnes, we show that retention of Th1 cells in the LNs is controlled by a chemokine, CXCL10/interferon (IFN) inducible protein 10 produced by mature dendritic cells (DCs). Hepatic LN DCs preferentially produced CXCL10 to attract 5'-bromo-2'-deoxyuridine (BrdU)+CD4+ T cells and form clusters with IFN-gamma-producing CD4+ T cells by day 7 after antigen challenge. Blockade of CXCL10 dramatically altered the distribution of cluster-forming BrdU+CD4+ T cells. BrdU+CD4+ T cells in the hepatic LNs were selectively diminished while those in the circulation were significantly increased by treatment with anti-CXCL10 monoclonal antibody. This was accompanied by accelerated infiltration of memory T cells into the periphery of hepatic granuloma sites, most of them were in cell cycle and further produced higher amount of IFN-gamma leading to exacerbation of liver injury. Thus, mature DC-derived CXCL10 is pivotal to retain Th1 lymphocytes within T cell areas of draining LNs and optimize the Th1-mediated immune responses.

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Visualization of DC–T cell clusters in the hepatic LNs. (a–c) Triple immunostaining in the hepatic LNs on days 0 (a) and 7 (b and c). (a) DEC-205+ (brown) cells and a little number BrdU+ (red) CD4+ (blue) cells at day 0. Note that these cells did not form clusters. (b) Numerous clusters between DEC-205+ (brown) cells and BrdU+ (red) CD4+ (blue) cells (arrow) were seen in the paracortex at day 7. *HEV. Scale bar, 100 μm. (c) Contact between BrdU+ (red) CD4+ (blue) cells and DEC-205+ (brown) cells (arrow) at the periphery of a small granulomatous cluster. Scale bar, 20 μm. (d) The numbers of BrdU+CD4+ cells in the paracortex of hepatic LNs determined by 15-mm2 stained cryosections. T cell area of hepatic LNs was defined by B220 or CD3ɛ staining (reference 25). (e) Antigen-specific proliferation of hepatic LN T cells sorted from normal and P. acnes–primed mice at day 2. White bar, medium alone (unstimulated); black bar, P. acnes–stimulated; striped bar, OVA-stimulated. (f) IFN-γ and IL-4 productions by sorted LN CD4+ T cells at day 7. C, medium alone as negative control. (d–f) Representative data from five independent experiments. Mean ± SD. n = 6. Student's t test, *P < 0.05. (g) Double immunostaining for B220 or CD4 (red) and IFN-γ (green) in the hepatic LNs on day 7. IFN-γ+ cells were selectively located in the B220− paracortex (left). IFN-γ+CD4+ cells (yellow) were selectively located at the paracortex. The outlines show the B cell follicle border. F, follicle; PC, paracortex. Scale bar, 40 μm. IFN-γ+CD4+ cells (merge; yellow; white arrowheads) were located at the periphery of a small granulomatous clusters. Right bottom panel depicts double staining for IFN-γ (green) and DEC-205 (red) at day 7. IFN-γ+ cells (white arrowheads) were located at the periphery of DEC-205+ DC accumulation. Scale bar, 20 μm.
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fig2: Visualization of DC–T cell clusters in the hepatic LNs. (a–c) Triple immunostaining in the hepatic LNs on days 0 (a) and 7 (b and c). (a) DEC-205+ (brown) cells and a little number BrdU+ (red) CD4+ (blue) cells at day 0. Note that these cells did not form clusters. (b) Numerous clusters between DEC-205+ (brown) cells and BrdU+ (red) CD4+ (blue) cells (arrow) were seen in the paracortex at day 7. *HEV. Scale bar, 100 μm. (c) Contact between BrdU+ (red) CD4+ (blue) cells and DEC-205+ (brown) cells (arrow) at the periphery of a small granulomatous cluster. Scale bar, 20 μm. (d) The numbers of BrdU+CD4+ cells in the paracortex of hepatic LNs determined by 15-mm2 stained cryosections. T cell area of hepatic LNs was defined by B220 or CD3ɛ staining (reference 25). (e) Antigen-specific proliferation of hepatic LN T cells sorted from normal and P. acnes–primed mice at day 2. White bar, medium alone (unstimulated); black bar, P. acnes–stimulated; striped bar, OVA-stimulated. (f) IFN-γ and IL-4 productions by sorted LN CD4+ T cells at day 7. C, medium alone as negative control. (d–f) Representative data from five independent experiments. Mean ± SD. n = 6. Student's t test, *P < 0.05. (g) Double immunostaining for B220 or CD4 (red) and IFN-γ (green) in the hepatic LNs on day 7. IFN-γ+ cells were selectively located in the B220− paracortex (left). IFN-γ+CD4+ cells (yellow) were selectively located at the paracortex. The outlines show the B cell follicle border. F, follicle; PC, paracortex. Scale bar, 40 μm. IFN-γ+CD4+ cells (merge; yellow; white arrowheads) were located at the periphery of a small granulomatous clusters. Right bottom panel depicts double staining for IFN-γ (green) and DEC-205 (red) at day 7. IFN-γ+ cells (white arrowheads) were located at the periphery of DEC-205+ DC accumulation. Scale bar, 20 μm.

Mentions: After administration of P. acnes, DEC-205+ mature DCs accumulated in the paracortex (Fig. 2 , a and b) clustered with CD4+ T cells (Fig. 2 b). Part of the CD4+ T cells at the periphery of accumulated DCs were labeled with BrdU (Fig. 2, b and c), suggesting that CD4+ T cells proliferated at these sites. To characterize the T cell subsets responding to DC-derived CXCL10, we next investigated the kinetics of proliferation, cytokine production, and migration potential of the hepatic LN T cells. The frequency of BrdU+CD4+ T cells (per mm2) in the paracortex of hepatic LNs rapidly increased to a maximum level by day 2 and remained stable thereafter (Fig. 2 d). P. acnes-primed LN T cells exhibited antigen-specific proliferation by day 2 (Fig. 2 e). Sorted LN CD4+ T cells at day 2 produced considerable levels of IFN-γ and IL-4 (Fig. 2 f). In contrast, the IL-4 level was decreased nearly to background while the IFN-γ level was dramatically increased by LN CD4+ T cells at day 7 (Fig. 2 f). These results suggested that BrdU+CD4+ T cells on day 2 reflected proliferating, nonpolarized Th cells while BrdU+CD4+ T cells by day 7 corresponded to further polarized Th1 cells. IFN-γ–producing CD4+ T cells were selectively detected in the paracortex (Fig. 2 g) and were located at the periphery of small granulomatous clusters (Fig. 2 g). DEC-205+ DCs were actually clustered with IFN-γ–producing cells (Fig. 2 g).


Pivotal role of dendritic cell-derived CXCL10 in the retention of T helper cell 1 lymphocytes in secondary lymph nodes.

Yoneyama H, Narumi S, Zhang Y, Murai M, Baggiolini M, Lanzavecchia A, Ichida T, Asakura H, Matsushima K - J. Exp. Med. (2002)

Visualization of DC–T cell clusters in the hepatic LNs. (a–c) Triple immunostaining in the hepatic LNs on days 0 (a) and 7 (b and c). (a) DEC-205+ (brown) cells and a little number BrdU+ (red) CD4+ (blue) cells at day 0. Note that these cells did not form clusters. (b) Numerous clusters between DEC-205+ (brown) cells and BrdU+ (red) CD4+ (blue) cells (arrow) were seen in the paracortex at day 7. *HEV. Scale bar, 100 μm. (c) Contact between BrdU+ (red) CD4+ (blue) cells and DEC-205+ (brown) cells (arrow) at the periphery of a small granulomatous cluster. Scale bar, 20 μm. (d) The numbers of BrdU+CD4+ cells in the paracortex of hepatic LNs determined by 15-mm2 stained cryosections. T cell area of hepatic LNs was defined by B220 or CD3ɛ staining (reference 25). (e) Antigen-specific proliferation of hepatic LN T cells sorted from normal and P. acnes–primed mice at day 2. White bar, medium alone (unstimulated); black bar, P. acnes–stimulated; striped bar, OVA-stimulated. (f) IFN-γ and IL-4 productions by sorted LN CD4+ T cells at day 7. C, medium alone as negative control. (d–f) Representative data from five independent experiments. Mean ± SD. n = 6. Student's t test, *P < 0.05. (g) Double immunostaining for B220 or CD4 (red) and IFN-γ (green) in the hepatic LNs on day 7. IFN-γ+ cells were selectively located in the B220− paracortex (left). IFN-γ+CD4+ cells (yellow) were selectively located at the paracortex. The outlines show the B cell follicle border. F, follicle; PC, paracortex. Scale bar, 40 μm. IFN-γ+CD4+ cells (merge; yellow; white arrowheads) were located at the periphery of a small granulomatous clusters. Right bottom panel depicts double staining for IFN-γ (green) and DEC-205 (red) at day 7. IFN-γ+ cells (white arrowheads) were located at the periphery of DEC-205+ DC accumulation. Scale bar, 20 μm.
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fig2: Visualization of DC–T cell clusters in the hepatic LNs. (a–c) Triple immunostaining in the hepatic LNs on days 0 (a) and 7 (b and c). (a) DEC-205+ (brown) cells and a little number BrdU+ (red) CD4+ (blue) cells at day 0. Note that these cells did not form clusters. (b) Numerous clusters between DEC-205+ (brown) cells and BrdU+ (red) CD4+ (blue) cells (arrow) were seen in the paracortex at day 7. *HEV. Scale bar, 100 μm. (c) Contact between BrdU+ (red) CD4+ (blue) cells and DEC-205+ (brown) cells (arrow) at the periphery of a small granulomatous cluster. Scale bar, 20 μm. (d) The numbers of BrdU+CD4+ cells in the paracortex of hepatic LNs determined by 15-mm2 stained cryosections. T cell area of hepatic LNs was defined by B220 or CD3ɛ staining (reference 25). (e) Antigen-specific proliferation of hepatic LN T cells sorted from normal and P. acnes–primed mice at day 2. White bar, medium alone (unstimulated); black bar, P. acnes–stimulated; striped bar, OVA-stimulated. (f) IFN-γ and IL-4 productions by sorted LN CD4+ T cells at day 7. C, medium alone as negative control. (d–f) Representative data from five independent experiments. Mean ± SD. n = 6. Student's t test, *P < 0.05. (g) Double immunostaining for B220 or CD4 (red) and IFN-γ (green) in the hepatic LNs on day 7. IFN-γ+ cells were selectively located in the B220− paracortex (left). IFN-γ+CD4+ cells (yellow) were selectively located at the paracortex. The outlines show the B cell follicle border. F, follicle; PC, paracortex. Scale bar, 40 μm. IFN-γ+CD4+ cells (merge; yellow; white arrowheads) were located at the periphery of a small granulomatous clusters. Right bottom panel depicts double staining for IFN-γ (green) and DEC-205 (red) at day 7. IFN-γ+ cells (white arrowheads) were located at the periphery of DEC-205+ DC accumulation. Scale bar, 20 μm.
Mentions: After administration of P. acnes, DEC-205+ mature DCs accumulated in the paracortex (Fig. 2 , a and b) clustered with CD4+ T cells (Fig. 2 b). Part of the CD4+ T cells at the periphery of accumulated DCs were labeled with BrdU (Fig. 2, b and c), suggesting that CD4+ T cells proliferated at these sites. To characterize the T cell subsets responding to DC-derived CXCL10, we next investigated the kinetics of proliferation, cytokine production, and migration potential of the hepatic LN T cells. The frequency of BrdU+CD4+ T cells (per mm2) in the paracortex of hepatic LNs rapidly increased to a maximum level by day 2 and remained stable thereafter (Fig. 2 d). P. acnes-primed LN T cells exhibited antigen-specific proliferation by day 2 (Fig. 2 e). Sorted LN CD4+ T cells at day 2 produced considerable levels of IFN-γ and IL-4 (Fig. 2 f). In contrast, the IL-4 level was decreased nearly to background while the IFN-γ level was dramatically increased by LN CD4+ T cells at day 7 (Fig. 2 f). These results suggested that BrdU+CD4+ T cells on day 2 reflected proliferating, nonpolarized Th cells while BrdU+CD4+ T cells by day 7 corresponded to further polarized Th1 cells. IFN-γ–producing CD4+ T cells were selectively detected in the paracortex (Fig. 2 g) and were located at the periphery of small granulomatous clusters (Fig. 2 g). DEC-205+ DCs were actually clustered with IFN-γ–producing cells (Fig. 2 g).

Bottom Line: Blockade of CXCL10 dramatically altered the distribution of cluster-forming BrdU+CD4+ T cells.BrdU+CD4+ T cells in the hepatic LNs were selectively diminished while those in the circulation were significantly increased by treatment with anti-CXCL10 monoclonal antibody.This was accompanied by accelerated infiltration of memory T cells into the periphery of hepatic granuloma sites, most of them were in cell cycle and further produced higher amount of IFN-gamma leading to exacerbation of liver injury.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Preventive Medicine, School of Medicine and Core Research and Evolutional Science and Technology (CREST), The University of Tokyo, Tokyo 113-0033, Japan.

ABSTRACT
Various immune diseases are considered to be regulated by the balance of T helper (Th)1 and Th2 subsets. Although Th lymphocytes are believed to be generated in draining lymph nodes (LNs), in vivo Th cell behaviors during Th1/Th2 polarization are largely unexplored. Using a murine granulomatous liver disease model induced by Propionibacterium acnes, we show that retention of Th1 cells in the LNs is controlled by a chemokine, CXCL10/interferon (IFN) inducible protein 10 produced by mature dendritic cells (DCs). Hepatic LN DCs preferentially produced CXCL10 to attract 5'-bromo-2'-deoxyuridine (BrdU)+CD4+ T cells and form clusters with IFN-gamma-producing CD4+ T cells by day 7 after antigen challenge. Blockade of CXCL10 dramatically altered the distribution of cluster-forming BrdU+CD4+ T cells. BrdU+CD4+ T cells in the hepatic LNs were selectively diminished while those in the circulation were significantly increased by treatment with anti-CXCL10 monoclonal antibody. This was accompanied by accelerated infiltration of memory T cells into the periphery of hepatic granuloma sites, most of them were in cell cycle and further produced higher amount of IFN-gamma leading to exacerbation of liver injury. Thus, mature DC-derived CXCL10 is pivotal to retain Th1 lymphocytes within T cell areas of draining LNs and optimize the Th1-mediated immune responses.

Show MeSH
Related in: MedlinePlus