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Direct evidence for activated CD8+ T cell transmigration across portal vein endothelial cells in liver graft rejection

View Article: PubMed Central - PubMed

ABSTRACT

Background: Lymphocyte recruitment into the portal tract is crucial not only for homeostatic immune surveillance but also for many liver diseases. However, the exact route of entry for lymphocytes into portal tract is still obscure. We investigated this question using a rat hepatic allograft rejection model.

Methods: A migration route was analyzed by immunohistological methods including a recently developed scanning electron microscopy method. Transmigration-associated molecules such as selectins, integrins, and chemokines and their receptors expressed by hepatic vessels and recruited T-cells were analyzed by immunohistochemistry and flow cytometry.

Results: The immunoelectron microscopic analysis clearly showed CD8β+ cells passing through the portal vein (PV) endothelia. Furthermore, the migrating pathway seemed to pass through the endothelial cell body. Local vascular cell adhesion molecule-1 (VCAM-1) expression was induced in PV endothelial cells from day 2 after liver transplantation. Although intercellular adhesion molecule-1 (ICAM-1) expression was also upregulated, it was restricted to sinusoidal endothelia. Recipient T-cells in the graft perfusate were CD25+CD44+ICAM-1+CXCR3+CCR5– and upregulated α4β1 or αLβ2 integrins. Immunohistochemistry showed the expression of CXCL10 in donor MHCIIhigh cells in the portal tract as well as endothelial walls of PV.

Conclusions: We show for the first time direct evidence of T-cell transmigration across PV endothelial cells during hepatic allograft rejection. Interactions between VCAM-1 on endothelia and α4β1 integrin on recipient effector T-cells putatively play critical roles in adhesion and transmigration through endothelia. A chemokine axis of CXCL10 and CXCR3 also may be involved.

Electronic supplementary material: The online version of this article (doi:10.1007/s00535-016-1169-1) contains supplementary material, which is available to authorized users.

No MeSH data available.


Kinetics of recipient cell infiltration into the graft after transplantation. Triple immunoenzyme staining by recipient MHCI (blue), type IV collagen (brown) and BrdU (red) in control liver (a), at 1d (b), 2d (c), 3d (d), 5d (e), 7d (f) and 10d (g) after LTx. Note recipient MHCI+ cell infiltration around the PV (PV) was dominant compared to the hepatic vein (HV) area over time. h, i Recruitment of TCRαβ+ (h, blue) or CD8β+ (i, blue) cells to portal tract on day 4, depicted by serial sections. Many TCRαβ+ or CD8β+ cells were seen in the portal tract and some of them were actively proliferating by incorporating BrdU. Some cells attached to the PV vessel walls (inset of h and i). Representative figures of three rats. Bd bile duct, PV portal vein, HV hepatic vein. Scale bars: a–g 200 μm; h, i 100 μm; inset of h and i 50 μm
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Fig1: Kinetics of recipient cell infiltration into the graft after transplantation. Triple immunoenzyme staining by recipient MHCI (blue), type IV collagen (brown) and BrdU (red) in control liver (a), at 1d (b), 2d (c), 3d (d), 5d (e), 7d (f) and 10d (g) after LTx. Note recipient MHCI+ cell infiltration around the PV (PV) was dominant compared to the hepatic vein (HV) area over time. h, i Recruitment of TCRαβ+ (h, blue) or CD8β+ (i, blue) cells to portal tract on day 4, depicted by serial sections. Many TCRαβ+ or CD8β+ cells were seen in the portal tract and some of them were actively proliferating by incorporating BrdU. Some cells attached to the PV vessel walls (inset of h and i). Representative figures of three rats. Bd bile duct, PV portal vein, HV hepatic vein. Scale bars: a–g 200 μm; h, i 100 μm; inset of h and i 50 μm

Mentions: In this transplantation setting, donor graft was acutely rejected by ~11 days [10]. In the graft, recipient MHCI-positive cells were almost absent on day 1 and appeared from day 2 and progressively increased. Notably prominent recipient cell infiltration was observed in the portal tract, which gradually expanded by the infiltrated cells (Fig. 1a–g). At the late stage, the sinusoidal area decreased to ~30 % of the total surface area (Fig. 1g) [6]. The majority of infiltrating cells on day 4 were positive for T-cell receptor (TCR)αβ, CD8β (Fig. 1h, i), or CD4 (not shown) with a high labeling index of BrdU, suggesting that infiltrated cells were mostly activated T-cells. These cellular kinetics results were consistent with a previously reported LTx model using a similar MHC-disparate combination, in which effector T-cell recruitment led to acute rejection [6].Fig. 1


Direct evidence for activated CD8+ T cell transmigration across portal vein endothelial cells in liver graft rejection
Kinetics of recipient cell infiltration into the graft after transplantation. Triple immunoenzyme staining by recipient MHCI (blue), type IV collagen (brown) and BrdU (red) in control liver (a), at 1d (b), 2d (c), 3d (d), 5d (e), 7d (f) and 10d (g) after LTx. Note recipient MHCI+ cell infiltration around the PV (PV) was dominant compared to the hepatic vein (HV) area over time. h, i Recruitment of TCRαβ+ (h, blue) or CD8β+ (i, blue) cells to portal tract on day 4, depicted by serial sections. Many TCRαβ+ or CD8β+ cells were seen in the portal tract and some of them were actively proliferating by incorporating BrdU. Some cells attached to the PV vessel walls (inset of h and i). Representative figures of three rats. Bd bile duct, PV portal vein, HV hepatic vein. Scale bars: a–g 200 μm; h, i 100 μm; inset of h and i 50 μm
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Fig1: Kinetics of recipient cell infiltration into the graft after transplantation. Triple immunoenzyme staining by recipient MHCI (blue), type IV collagen (brown) and BrdU (red) in control liver (a), at 1d (b), 2d (c), 3d (d), 5d (e), 7d (f) and 10d (g) after LTx. Note recipient MHCI+ cell infiltration around the PV (PV) was dominant compared to the hepatic vein (HV) area over time. h, i Recruitment of TCRαβ+ (h, blue) or CD8β+ (i, blue) cells to portal tract on day 4, depicted by serial sections. Many TCRαβ+ or CD8β+ cells were seen in the portal tract and some of them were actively proliferating by incorporating BrdU. Some cells attached to the PV vessel walls (inset of h and i). Representative figures of three rats. Bd bile duct, PV portal vein, HV hepatic vein. Scale bars: a–g 200 μm; h, i 100 μm; inset of h and i 50 μm
Mentions: In this transplantation setting, donor graft was acutely rejected by ~11 days [10]. In the graft, recipient MHCI-positive cells were almost absent on day 1 and appeared from day 2 and progressively increased. Notably prominent recipient cell infiltration was observed in the portal tract, which gradually expanded by the infiltrated cells (Fig. 1a–g). At the late stage, the sinusoidal area decreased to ~30 % of the total surface area (Fig. 1g) [6]. The majority of infiltrating cells on day 4 were positive for T-cell receptor (TCR)αβ, CD8β (Fig. 1h, i), or CD4 (not shown) with a high labeling index of BrdU, suggesting that infiltrated cells were mostly activated T-cells. These cellular kinetics results were consistent with a previously reported LTx model using a similar MHC-disparate combination, in which effector T-cell recruitment led to acute rejection [6].Fig. 1

View Article: PubMed Central - PubMed

ABSTRACT

Background: Lymphocyte recruitment into the portal tract is crucial not only for homeostatic immune surveillance but also for many liver diseases. However, the exact route of entry for lymphocytes into portal tract is still obscure. We investigated this question using a rat hepatic allograft rejection model.

Methods: A migration route was analyzed by immunohistological methods including a recently developed scanning electron microscopy method. Transmigration-associated molecules such as selectins, integrins, and chemokines and their receptors expressed by hepatic vessels and recruited T-cells were analyzed by immunohistochemistry and flow cytometry.

Results: The immunoelectron microscopic analysis clearly showed CD8β+ cells passing through the portal vein (PV) endothelia. Furthermore, the migrating pathway seemed to pass through the endothelial cell body. Local vascular cell adhesion molecule-1 (VCAM-1) expression was induced in PV endothelial cells from day 2 after liver transplantation. Although intercellular adhesion molecule-1 (ICAM-1) expression was also upregulated, it was restricted to sinusoidal endothelia. Recipient T-cells in the graft perfusate were CD25+CD44+ICAM-1+CXCR3+CCR5– and upregulated α4β1 or αLβ2 integrins. Immunohistochemistry showed the expression of CXCL10 in donor MHCIIhigh cells in the portal tract as well as endothelial walls of PV.

Conclusions: We show for the first time direct evidence of T-cell transmigration across PV endothelial cells during hepatic allograft rejection. Interactions between VCAM-1 on endothelia and α4β1 integrin on recipient effector T-cells putatively play critical roles in adhesion and transmigration through endothelia. A chemokine axis of CXCL10 and CXCR3 also may be involved.

Electronic supplementary material: The online version of this article (doi:10.1007/s00535-016-1169-1) contains supplementary material, which is available to authorized users.

No MeSH data available.