Gut-homing CD4+ T cell receptor αβ+ T cells in the pathogenesis of murine inflammatory bowel disease

We studied which T cell subsets from the gut-associated lymphoid tissue (GALT) can migrate out of the gut mucosa and repopulate GALT compartments of an immunodeficient (semi)syngeneic host. Many distinct lymphocyte subsets were found in GALT of immunocompetent H-2^d (BALB/c, BALB/c^dm2, C. B-17+/+) mice. No antigen receptor-expressing lymphoid cells were found in GALT of congenic C. B-17 scid/scid (scid) mice. The heterotopic transplantation of a full-thickness gut wall graft from the ileum or colon of immunocompetent (C. B-17+/+, BALB/c^dm2) donor mice onto immunodeficient scid mice selectively reconstituted a CD3⁺ T cell receptor αβ⁺ CD4⁺ T cell subset. CD4⁺ cells of this subset expressed the surface phenotype of mucosa-seeking, memory T cells. In the immunodeficient scid host, this gut-derived CD4⁺ T cell subset was found in spleen, peritoneal cavity, mesenteric lymph nodes (LN), epithelial layer and lamina propria of the small and large intestine, but not in peripheral LN. Scid mice heterotopically transplanted with gut from a congenic, immunocompetent donor developed clinical and histological signs of inflammatory bowel disease (IBD). Hence, the selective repopulation of GALT compartments with CD4⁺ T cells from normal GALT plays an essential role in the pathogenesis of IBD in an immunodeficient host.     

Co-expression of CD8α in CD4+ T cell receptor αβ+ T cells migrating into the murine small intestine epithelial layer

We investigated the surface phenotype of CD3⁺CD4⁺ T cell receptor (TCR) αβ⁺ T cells repopulating the intestinal lymphoid tissues of C.B-17 scidlscid (severe-combined immunodeficient; scid) (H-2^d, L^d+) mice. CD4⁺ CD8^? T cells were cell sorter-purified from various secondary and tertiary lymphoid organs of congenic C.B-17 +/+ (H-2^d, L^d+) or semi-syngeneic dm2 (H-2^d, L^d?) immunocompetent donor mice. After transfer of 10⁵ cells into young scid mice, a mucosa-homing, memory CD44^hi CD45RB^lo CD4⁺ T cell population was selectively engrafted. Large numbers of single-positive (SP) CD3⁺ CD2⁺ CD28⁺ CD4⁺ CD^8? T cells that expressed the α₄ integrin chain CD49d were found in the spleen, the mesenteric lymph nodes, the peritoneal cavity and the gut lamina propria of transplanted scid mice. Unexpectedly, large populations of donor-type doublepositive (DP) CD4⁺ CD8α⁺ CD8β^? T cells with high expression of the CD3/TCR complex appeared in the epithelial layer of the small intestine of transplanted scid mice. In contrast to SP CD4⁺ T cells, the intraepithelial DP T cells showed low expression of the CD2 and the CD28 co-stimulator molecules, and of the α₄ integrin chain CD49d, but expressed high levels of the α_IEL integrin chain CD103. The TCR-Vβ repertoire of DP but not SP intraepithelial CD4⁺ T cells was biased towards usage of the Vβ6 and Vβ8 viable domains. Highly purified populations of SP and DP CD4⁺ T cell populations from the small intestine epithelial layer of transplanted scid mice had different abilities to repopulate secondary scid recipient mice: SP CD4⁺ T cells repopulated various lymphoid tissues of the immunodeficient host, while intraepithelial DP CD4⁺ T cells did not. Hence, a subset of CD3⁺ CD4⁺ TCRαβ⁺ T cells apparently undergoes striking phenotypic changes when it enters the microenvironment of the small intestine epithelial layer.     

Polyclonal expansion of adoptively transferred CD4+ αβ T cells in the colonic lamina propria of scid mice with colitis

The adoptive transfer of low numbers of peripheral, non-fractionated CD4⁺ αβ T cells into histocompatible, severely immunodeficient (scid) hosts induces a colitis. This disease developed in C.B-17 scid/scid hosts after the injection of 10⁵CD4⁺ T cells purified from different peripheral lymphoid organs of immunocompetent C.B-17 +/+ or BALB/c^dm2 donor mice. Irrespective of their tissue origin, transferred CD4⁺ T cells selectively repopulated the scid host with gut-seeking CD4⁺ T cells. A chronic inflammatory bowel disease (IBD) developed as polyclonal populations of mucosa-seeking memory/effector CD4⁺ T cells accumulated in the gut lamina propria and epithelial layer of the adoptive host. The manifestation of colitis in the scid host correlated with the in situ polyclonal activation and expansion of adoptively transferred CD4⁺ T cells in the colonic lamina propria. Attempts were unsuccessful to select in vivo an oligoclonal CD4⁺ T cell population with an enhanced IBD-inducing potential by repeatedly reinjecting 10⁵ donor-type CD4⁺ T cells from the colonic lamina propria of transplanted scid mice with an early and severe IBD into new scid hosts. The data indicate that the preferential repopulation of gut-associated lymphoid tissues with immunocompetent CD4⁺ T cells, and their polyclonal activation and in situ expansion in the lamina propria of the histocompatible, immunodeficient host are critical events in the pathogenesis of an IBD in this model.     

CD3+ T cells in severe combined immunodeficiency (scid) mice. II. Transplantation of dm2 lymphoid cells into semi-allogeneic scid mice.

Intravenous injection of nonfractionated BALB/c-H-2dm2 (dm2) (Ld-) spleen cells into 4-week-old, semi-allogeneic (H-2d, Ld+) C.B-17 scid/scid severe combined immunodeficient (scid) mice (2 x 10(7) cells/mouse) reconstituted T lymphopoiesis in thymi and repopulated the lymphoid white pulp in spleens of these immunodeficient recipients. Transplantation of dm2 thymocytes into young scid mice (5 x 10(7) cells/mouse) established a donor-derived CD3+ T cell population in spleens of recipient scid mice, in which CD4+T cells predominated. This was demonstrated by marker analyses of thymocytes and splenocytes, and determinations of serum immunoglobulin levels in transplanted scid mice. Transfer of splenocytes from young primary scid recipients into young secondary or tertiary recipients (3 x 10(6) cells/mouse) engrafted preferentially dm2-derived CD3+CD4+CD8- T cells in spleens of scid mice despite the strong selective Ld-associated alloantigenic stimulus for CD8+ T cells. Intravenous injections of nonfractionated dm2 spleen cells (2 x 10(7) cells/mouse) or thymocytes 5 x 10(7) cells/mouse) into 10- to 12-month-old, "leaky" scid mice induced severe clinical signs of graft-vs.-host disease (GVHD) in all scid recipients. Lymphoid repopulation of spleen and thymus in old scid recipients was incomplete. This GVHD was not transferrable by injecting 3 x 10(6) spleen cells from old diseased primary scid recipients into secondary or tertiary young scid recipient mice. In these serial transfers, dm2-derived CD3+CD4+CD8- T cells were again preferentially engrafted in spleens of scid recipients. Transfer of purified CD4+ dm2 T cells into young scid mice (2 x 10(5) to 5 x 10(5) cells/mouse) engrafted this T cell subset into the spleen of semi-allogeneic scid recipients. This was revealed by histological examinations, surface marker analyses, in vitro isolation of donor-type CD3+CD4+ T cell lines from spleens of transplanted scid mice, and serial transfer experiments. These data indicated that the CD4+ T cell compartment of scid mice can be selectively repopulated by semi-allogeneic T cells. Injections of purified CD8+ dm2 T-cells into young scid mice (2 x 10(5) cells/mouse) did not establish a CD8+ T cell graft in spleens of recipients. It was necessary to inject transplanted scid mice biweekly with 10(4) units recombinant interleukin 2 to establish and/or maintain transferred dm2 CD8+ T cells in spleens of these recipients, dm2 CD8+ T cell-transplanted and interleukin 2-treated scid mice did not develop any evidence of GVHD over the 9-week observation period.     

Adoptive Transfer of Low Numbers of CD4+ T Cells into SCID Mice chronically treated with Soluble IL-4 Receptor does not prevent Engraftment of IL-4-Producing T Cells

After intravenous injection of 10⁵ purified, lymph node (LN)-derived dm2 (H-2^d/L^d) CD4⁺ T cells into young C.B-17 scidjscid (severe combined immunodeficiency, SCID) mice (H-2^d/L^d+), the transplanted L^d- T cells show a selective pattern of engraftment: they repopulate the spleen, the lamina propria of the small intestine and the mesenteric LN (but not other peripheral LN) of the immunodeficient host. CD4⁺ cells repopulating different lymphoid organs of the SCID recipient mice produce interleukin-2 (IL-2) and interleukin-4 (IL-4) in response to polyclonal stimulation in vitro. Some evidence has recently been provided that cytokines (e.g. IL-4) present at the site of antigen stimulation in vivo decisively influence the pattern of cytokines expressed by T cells activated at these sites. We therefore asked if neutralization of IL-4 by chronic treatment of SCID mice with high doses of recombinant soluble IL-4 receptor (sIL-4R) changes the IL-4 or IL-2 expression pattern of CD4⁺ T cells adoptively transferred into young SCID recipients. Transplanted SCID mice were chronically treated with two different, recombinant murine sIL-4R proteins. The experimental series further included groups of transplanted SCID mice treated with a recombinant human sIL-4R protein (which does not bind murine IL-4), treated with the anti-murine IL-4 monoclonal antibody (MoAb) 11B11, or non-treated. Transplanted SCID mice treated with the recombinant murine sIL-4R protein preparations displayed detectable sIL-4R serum levels, which demonstrates that the substitution therapy could maintain neutralizing serum levels of anti-IL-4 activity in SCID mice. By contrast, no serum sIL-4R levels were detectable in the sensitive ELISA readout in transplanted SCID mice which were non-treated, treated with the MoAb 11B11, or treated with the recombinant human sIL-4R protein. The efficiency and the pattern of CD4⁺ T-cell engraftment, and the lymphokine-producing phenotype of the engrafted dm2 CD4⁺ cells, was not affected by the continuous IL-4-neutralizing treatment of mice with either the MoAb 11B11 or the soluble IL-4R preparations. Hence, in contrast to the published evidence of the dramatic effect of IL-4 on the lymphokine-producing phenotype of CD4⁺ T cells stimulated in vitro or in vivo, the chronic suppression in vivo of IL-4 activity (by either different sIL4-R protein constructs, or by the anti-IL-4 MoAb 11B11) did not lead to preferential engraftment of Th1-type CD4⁺ T cells after adoptive transfer of CD4⁺ T-cell populations into an immunodeficient recipient.     

Cytotoxic reactivity of gut lamina propria CD4+ αβ T cells in SCID mice with colitis

Polyclonal, mucosa-seeking memory/effector CD4⁺ T cells containing a large fraction of blasts activated in situ accumulate in the gut lamina propria of severe-combined immunodeficient (SCID) mice developing colitis after CD4⁺ T cell transplantation. CD4⁺ T cells isolated from different repopulated lymphoid tissues of transplanted SCID mice proliferate in vitro in the presence of interleukin (IL)-2 + IL-7. CD3 ligation enhances this cytokine-supported proliferation in CD4⁺ T cells from the spleen and the mesenteric lymph node of transplanted SCID mice; CD3 ligation suppresses the cytokine-supported proliferation in CD4⁺ T cells from the gut lamina propria in a cell density- and dose-dependent manner. Almost all CD4⁺ T cells from repopulated lymphoid tissues of transplanted SCID mice express CD95 (Fas) on the cell surface, and a large fraction of CD4⁺ T cells from the gut lamina propria of transplanted SCID mice express the Fas ligand on the surface. Gut lamina propria CD4⁺ T cells show Fas-dependent cytotoxicity. A large fraction of gut lamina propria CD4⁺ T cells that infiltrate the inflamed colon in transplanted SCID mice are activated in situ and many CD4⁺ T cells are apoptotic. Hence, a large fraction of colitis-inducing CD4⁺ T cells undergo activation-induced cell death in situ and can damage other cells through Fas-dependent cytotoxicity.     

Expression of selectin-binding epitopes and cytokines by CD4+ T cells repopulating scid mice with colitis

Recruitment into the gut of CD4⁺ T cells and their activation in the colonic lamina propria (LP) are key events in the development of colitis in scid mice reconstituted with CD4⁺ T cells from immunocompetent, congenic donor mice. This study investigated the expression of cytokines and selectin-binding epitopes by CD4⁺ T cells repopulating different tissues of the adoptive scid host. Cells from the inflamed colonic LP of transplanted scid mice produced high amounts of IL-12, IFN-γ and TNF-α but only low amounts IL-4 and IL-10. Intracellular cytokine staining confirmed the presence of large numbers of IFN-γ- and TNF-α-producing effector CD4⁺ T cells in the colonic LP of scid mice with colitis but also in non-inflamed tissues [spleen (S), peritoneal cavity (PC) and mesenteric lymph nodes (mLN)] of the adoptive host. Cells from these tissues furthermore produced large amounts of IL-12. Ligands for endothelial selectins are involved in recruiting T cells into inflamed tissues. We have analyzed the expression of selectin-binding epitopes on CD4⁺ T cells repopulating different tissues of the adoptive scid host. We found that a large fraction of CD4⁺ T cells from inflamed colonic LP and from non-inflamed PC, mLN and S expressed high levels of P- and E-selectin-binding epitopes (P-L^hi) in transplanted scid mice, but not in congenic, immunocompetent control mice. Although P-L^hi CD4⁺ T cells were enriched in IFN-γ-producing subsets from most (but not all) tissues, we also found large numbers of in vivo generated P-L^lo CD4⁺ T cells producing pro-inflammatory cytokines. This was in contrast to in vitro generated Th1 CD4⁺ T blasts that were almost exclusively P-L^hi. In this mouse model, production of Th1-type pro-inflammatory cytokines and expression of surface epitopes binding endothelial selectins are hence strikingly up-regulated in CD4⁺ T cells residing in inflamed and non-inflamed tissues during the development of colitis.     

Proliferation and apoptosis of lamina propria CD4+ T cells from scid mice with inflammatory bowel disease

Scid mice transplanted with low numbers of syngeneic CD4⁺ T cells, develop a chronic and lethal inflammatory bowel disease (IBD) within 4 – 6 months. We have used in vivo 5-bromo-2-deoxy-uridine (BrdU) labeling to assess the proliferation of lamina propria-derived CD4⁺ T cells in diseased scid mice. The hourly rate of renewal of colonic lamina propria CD4⁺ T cells in diseased mice was 7 % compared with 1.5 % in normal BALB/c control mice. Transplantation of scid mice with in vitro activated CD4⁺ T cells accelerated the disease onset and development in a cell dose-dependent fashion when compared with non-activated CD4⁺ T cells. In pulse-chase experiments it was shown that BrdU-labeled cells disappeared rapidly from the lamina propria of diseased mice. DNA analysis revealed that this was due to the presence of nearly four times as many apoptotic CD4⁺ T cells in diseased than in control mice. Further analyses showed that the apoptotic lamina propria CD4⁺ T cells were derived from cells having entered the cell cycle within the previous 8 h. These data clearly demonstrate that vigorous CD4⁺ T cell proliferation and death are involved throughout the course of IBD.     

Induction of organ-selective CD4+ regulatory T cell homing

Compelling evidence suggests that Foxp3⁺CD25⁺CD4⁺ Treg play a fundamental role in immunoregulation. We have previously demonstrated that Treg have to enter peripheral tissues to suppress ongoing inflammation. However, relatively little is known about how Treg acquire the expression of homing receptors required for tissue‐ or inflammation‐specific migration. Migratory properties of conventional naïve T cells are shaped by the tissue microenvironment and organ‐specific dendritic cells during priming. Here, we show that this basic concept also holds true for CD25⁺CD4⁺ Treg: Priming of Treg within peripheral LN led to the expression of selectin ligands, which facilitate migration into inflamed skin, whereas activation within mesenteric LN led to induction of the integrin α₄β₇, which is required for migration into mucosal tissues. Furthermore, we could establish in vitro culture systems containing either dendritic cells from mesenteric and peripheral LN, or retinoic acid and IL‐12 as polarizing compounds to induce mucosa‐ and skin‐seeking Treg, respectively. Together, our results demonstrate that Treg, similarly to conventional T cells, can be configured with organ‐selective homing properties allowing efficient targeting into distinct tissues.     

Renal‐infiltrating CD11c+ cells are pathogenic in murine lupus nephritis through promoting CD4+ T cell responses

Lupus nephritis (LN) is a major manifestation of systemic lupus erythematosus (SLE), causing morbidity and mortality in 40–60% of SLE patients. The pathogenic mechanisms of LN are not completely understood. Recent studies have demonstrated the presence of various immune cell populations in lupus nephritic kidneys of both SLE patients and lupus‐prone mice. These cells may play important pathogenic or regulatory roles in situ to promote or sustain LN. Here, using lupus‐prone mouse models, we showed the pathogenic role of a kidney‐infiltrating CD11c⁺ myeloid cell population in LN. These CD11c⁺ cells accumulated in the kidneys of lupus‐prone mice as LN progressed. Surface markers of this population suggest their dendritic cell identity and differentiation from lymphocyte antigen 6 complex (Ly6C)^low mature monocytes. The cytokine/chemokine profile of these renal‐infiltrating CD11c⁺ cells suggests their roles in promoting LN, which was confirmed further in a loss‐of‐function in‐vivo study by using an antibody‐drug conjugate (ADC) strategy targeting CX₃CR1, a chemokine receptor expressed highly on these CD11c⁺ cells. However, CX₃CR1 was dispensable for the homing of CD11c⁺ cells into lupus nephritic kidneys. Finally, we found that these CD11c⁺ cells co‐localized with infiltrating T cells in the kidney. Using an ex‐ vivo co‐culture system, we showed that renal‐infiltrating CD11c⁺ cells promoted the survival, proliferation and interferon‐γ production of renal‐infiltrating CD4⁺ T cells, suggesting a T cell‐dependent mechanism by which these CD11c⁺ cells promote LN. Together, our results identify a pathogenic kidney‐infiltrating CD11c⁺ cell population promoting LN progression, which could be a new therapeutic target for the treatment of LN.     

Lymph node and circulating T cell characteristics are strongly correlated in end‐stage renal disease patients,but highly differentiated T cells reside within the circulation

Ageing is associated with changes in the peripheral T cell immune system, which can be influenced significantly by latent cytomegalovirus (CMV) infection. To what extent changes in circulating T cell populations correlate with T cell composition of the lymph node (LN) is unclear, but is crucial for a comprehensive understanding of the T cell system. T cells from peripheral blood (PB) and LN of end‐stage renal disease patients were analysed for frequency of recent thymic emigrants using CD31 expression and T cell receptor excision circle content, relative telomere length and expression of differentiation markers. Compared with PB, LN contained relatively more CD4⁺ than CD8⁺ T cells (P < 0·001). The percentage of naive and central memory CD4⁺ and CD8⁺ T cells and thymic output parameters showed a strong linear correlation between PB and LN. Highly differentiated CD28^null T cells, being CD27^–, CD57⁺ or programmed death 1 (PD‐1⁺), were found almost exclusively in the circulation but not in LN. An age‐related decline in naive CD4⁺ and CD8⁺ T cell frequency was observed (P = 0·035 and P = 0·002, respectively) within LN, concomitant with an increase in central memory CD8⁺ T cells (P = 0·033). Latent CMV infection increased dramatically the frequency of circulating terminally differentiated T cells, but did not alter T cell composition and ageing parameters of LN significantly. Overall T cell composition and measures of thymic function in PB and LN are correlated strongly. However, highly differentiated CD28^null T cells, which may comprise a large part of circulating T cells in CMV‐seropositive individuals, are found almost exclusively within the circulation.     

T cell specialization at environmental interfaces: T cells from the lung and the female genital tract of lpr and gld mice differ from their splenic and lymph node counterparts

Mice homozygous for lpr and gld accumulate CD4^?CD8^? (double-negative, DN) B220⁺CD5¹⁰Thy-1¹⁰ αβ T cells in the spleen and lymph nodes (LN), while mucosal gut T cells are normal. To study other mucosa-associated T cell populations, we examined T cell subsets separated according to expression of αβ T cell receptor, CD4, CD5, CD8, Thy-1 and B220 in the lung and the female genital tract (FGT) of adult MRL lpr, C3H lpr and C3H gld mice. αβ T cell accumulation was detected in both the FGT and the lungs of lpr and gld mice but, in contrast to the spleen and LN, equal proportions of DN B220⁺ and CD4⁺ of CD8⁺ (single-positive, SP) B220^? T cells were observed in these sites, and the T cells had an increased expression of Thy-1 and CD5. Staining for CD44, L-selectin, and CD45RB revealed a higher percentage of effector/memory T cells in lpr and gld lungs and FGT compared to spleens and LN. CD69 expression suggested chronic activation of DN and SP T cells in lpr and gld lungs and FGT. Thus, we show that FGT and lung resident T cells are affected by lpr and gld mutations, but that their phenotypes are distinct from those of systemic T cells. These data suggest that T cells associated with FGT and lung mucosal tissues represent a separate lineage from systemic T cells, and/or that the abnormal T cells in lpr and gld mice are selected against in mucosal surfaces exposed to environmental antigen.     

Nonspecific augmentation of lymph node T cells and I-E-independent selective deletion of VβbTl4+ T cells by Mtv-2 in the DDD mouse

DDD/1 (DDD) mice were characterized by marked paucity of T cells in lymph nodes (LN). In DDD-Mtv-2/Mtv-2 (DDD-Mtv-2) congenics, T cells were 4- to 18-fold increased depending on ages but B cells doubled at the most. Thymus weight also increased. In DDDfDDD-Mtv-2, DDD neonatally infected with Mtv-2-derived exogenous MMTV (MMTV-2), neither LN cells nor thymus weight increased. The V_β 5⁺ and V_β 8⁺ Tcell contents in LN were practically the same among three strains. The Mtv-2-induced expansion of LN Tcells was polyclonal and appeared indigenous to DDD mice. Both Mtv-2 and MMTV-2 induced progressive age-dependent deletion of V_β 14⁺CD4⁺ LN cells. Mtv-2 but not MMTV-2 caused deletion of V_β 14⁺CD8+ LN cells and mature V_β 14⁺CD4⁺ thymocytes. Thus, Mtv-2- and MMTV-2-induced V_β 14⁺ T cell deletion may reflect intrathymic and peripheral elimination, respectively. The absence of I-E gene expression in DDD indicates that V_β 14⁺ T cell deletion advances independently of I-E molecules in this experimental system.     

Development of T cells in SCID mice grafted with fetal thymus from AKR mice or F344 rats

To examine the development of T cells within an allogeneic or xenogeneic environment, we engrafted the fetal thymus from AKR mice or F344 rats under the kidney capsule of SCID mice (mTG and rTG mice). T lymphopoiesis developed in SCID mice 2 months after transplantation, although the ratio of CD4/CD8 in both experimental groups was different from that of normal control. T cells in mTG mice did not show in vitro proliferation or cytotoxicity against either host-type C.B-17 (H-2^d) or donor-type AKR (H-2^k) cells, while they exerted potent activities against third-party BIO (H-2^b) cells. In contrast, T cells in rTG mice exhibited proliferation against both host-type C.B-17 and donor-type F344 rat cells. Consistently, graft-vs.-host disease symptoms developed in these mice and histological examination showed impressive infiltration of lymphocytes into the skin or into the mucosal layers of the stomach. Activated state of T cells in rTG mice was also evidenced by the positive expression of interleukin-2 receptor. Taken together, fetal thymus appears to contain progenitor cells which are sufficient for in vivo reconstitution of T lymphopoiesis, but species-specific environment is important for the induction of tolerance. In mTG mice, Vβ6⁺ T cells reactive to donor Mls^a determinants and Vβ3⁺ T cells reactive to host Mls^c determinants were deleted, suggesting that tolerance was regulated mainly by clonal deletion. By contrast, Vβ11⁺ T cells reactive to Mls^f determinants were not deleted possibly due to the lack of their ligands.     

Homeostatic proliferation of naïve CD8+ T cells depends on CD62L/L‐selectin‐mediated homing to peripheral LN

Adoptive transfer of naïve CD8⁺ T cells into lymphopenic recipients results both in spontaneous proliferation and in partial activation of T cells, a phenomenon termed homeostatic proliferation (HP). HP of CD8⁺ T cells is dependent on host IL‐7, IL‐15, and MHC‐class I and has been shown to prevent T‐cell tolerance, reverse T‐cell anergy and support T‐cell‐mediated tumor control in vivo. However, the initial anatomic site of HP is still under debate. Since we observed that the earliest detectable HP occurs within LN and that T cells undergoing HP retain a CD62L^bright phenotype, we investigated the functional role of CD62L for this process. We found that CD62L‐expression on T cells is required for optimal HP and HP was impaired in lymphotoxin‐αβ^?/? mice, indicating the necessity for intact host secondary lymphoid organ structures. Use of the LN egression inhibitor FTY720 indicated that LN structures were pivotal to yield homeostatically proliferated T cells detected in other compartments. Consistent with these results, HP‐supported control of MC57‐SIY tumors depended on CD62L. Our data indicate a critical role for CD62L and LN homing for the process of HP, which has implications for adoptive immunotherapy approaches of cancer.     

Human recombinant interferon-β influences T helper subset differentiation by regulating cytokine secretion pattern and expression of homing receptors

Type I interferons (IFN) are important regulators of both innate and acquired immunity. We have used an in vitro system of human CD4⁺ T cell differentiation to determine how IFN-β influences development of T helper (Th) subsets and homing receptor expression. IFN-β promoted differentiation of CD4⁺ T cells that produce low levels of both IFN-γ and lymphotoxin compared to interleukin (IL)-12-derived Th1 CD4⁺ T cells. IFN-β inhibited production of Th2 cytokines (IL-5 and IL-13) and augmented IL-12-mediated IL-10 secretion. In addition, IFN-β significantly enhanced L-selectin expression on CD4⁺ T cells and synergized with IL-12 to induce expression of cutaneous lymphocyte-associated antigen (CLA). This Th1 L-selectin⁺, CLA⁺ phenotype is characteristic of T cells found in normal human skin and suggests a role for type I IFN in the regulation of Th subset differentiation and tissue-specific homing receptors.     

Dual effects of the alloresponse by Th1 and Th2 cells on acute and chronic rejection of allotransplants

The contribution of direct and indirect alloresponses by CD4⁺ Th1 and Th2 cells in acute and chronic rejection of allogeneic transplants remains unclear. In the present study, we addressed this question using a transplant model in a single MHC class I‐disparate donor–recipient mouse combination. BALB/c‐dm2 (dm2) mutant mice do not express MHC class I L^d molecules and reject acutely L^d+ skin grafts from BALB/c mice. In contrast, BALB/c hearts placed in dm2 mice are permanently accepted in the absence of chronic allograft vasculopathy. In this model, CD4⁺ T cells are activated following recognition of a donor MHC class I determinant, L^d 61–80, presented by MHC Class II A^d molecules on donor and recipient APC. Pre‐transplantation of recipients with L^d 61–80 peptide emulsified in complete Freund's adjuvant induced a Th1 response, which accelerated the rejection of skin allografts, but it had no effect on cardiac transplants. In contrast, induction of a Th2 response to the same peptide abrogated the CD8⁺ cytotoxic T cells response and markedly delayed the rejection of skin allografts while it induced de novo chronic rejection of heart transplants. This shows that Th2 cells activated via indirect allorecognition can exert dual effects on acute and chronic rejection of allogeneic transplants.     

Splenic T helper cell type 1 cytokine profile and extramedullary haematopoiesis in severe combined immunodeficient (scid) mice with inflammatory bowel disease (IBD)

Scid mice develop a severe, chronic, and lethal IBD 3–6 months after engraftment of gut wall from immunocompetent congenic donors, induced by donor-derived CD4⁺ T cells migrating from the graft [ 7]. We have investigated intracellular T-helper type 1 (Th1) cytokines in the spleens of gut wall-transplanted scid mice with IBD. Increased fractions of interferon-gamma (IFN-γ), tumour necrosis factor-alpha (TNF-α) and IL-2-positive CD4⁺ T cells were found in the spleens of diseased mice compared with control mice. Moreover, a small but significant population of CD4⁺ T cells which stained positive for granulocyte-macrophage colony-stimulating factor (GM-CSF) was found in scid mice with IBD but was virtually absent in congenic non-scid control mice. Cloning of granulocyte/macrophage colony-forming cells (G/M-CFC) revealed that both non-transplanted scid mice and scid mice with IBD had an 8–14-fold increase in splenic G/M-CFC compared with control mice. No significant difference in the number of G/M-CFC per total spleen was found between non-transplanted and disease scid mice, although both groups of mice showed a nearly two-fold increase compared with control mice. G/M-CFC were never found in the thymus, liver or lymph nodes of diseased mice. Immunohistochemistry revealed that the multinucleated giant cells observed in the gut wall of diseased mice did not represent haematopoietic foci, but were derived from macrophages. These observations point towards a dominant role for Th1-type CD4⁺ T cells in the immunopathogenesis of IBD, whereas haematopoiesis does not seem to be affected by the development of the disease.     

Improved Engraftment of Human Spleen Cells in NOD/LtSz-scid/scid Mice as Compared with C.B-17-scid/scid Mice

T and B lymphocyte-deficient mice homozygous for the severe combined immunodeficiency (SCID) mutation can be immunologically engrafted with human lymphocytes. However, low levels of human peripheral blood mononuclear cell engraftment are commonly observed, impeding full use of this model We now demonstrate that strain background in mice homozygous for the scid mutation is a strong determinant of levels of human lymphocyte engraftment. NOD/LtSz-scid/scid mice support higher levels of engraftment of both human spleen and peripheral blood mononuclear cells than do C.B-17-scid/scid mice. We observed, using human spleen cell injected scid mice, 1), high levels of engraftment of the host peripheral lymphoid tissues with human CD45⁺ (leukocytes), CD3⁺ (T cells), CD4⁺ (helper/inducer), and CD8⁺ (suppressor/cytotoxic) lymphoid cells for up to 24 weeks in NOD/LtSz-scid/scid mice; 2), migration of high numbers of human lymphocytes to peripheral lymphoid and nonlymphoid organs in NOD/LtSz-scid/scid, but not in C.B-17-scid/scid mice; 3), higher levels of serum immunoglobulin of human origin in NOD/LtSz-scid/scid mice than in C.B-17-scid/scid mice; 4), histological lesions character-istic of human anti-mouse xenoreactivity in NOD/LtSz-scid/scid mice; and 5), human origin antibodies against filarial antigens after engraftment with naive human spleen cells. The use of NOD/LtSz-scid/scid mice as recipients to achieve significantly enhanced human lymphopoietic cell engraftment will now enable human immunity to be more easily studied in animal models.