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Blood, 15 May 2008, Vol. 111, No. 10, pp. 5242-5251. Prepublished online as a Blood First Edition Paper on February 19, 2008; DOI 10.1182/blood-2007-09-107953.
TRANSPLANTATION Effects of donor T-cell trafficking and priming site on graft-versus-host disease induction by naive and memory phenotype CD4 T cells1 Department of Laboratory Medicine, Yale University School of Medicine, New Haven, CT; 2 Cancer Center and Department of Pediatrics, Division of Bone Marrow Transplantation, University of Minnesota, Minneapolis; Departments of 3 Pathology & Dermatology and 4 Pathology, Yale University School of Medicine, New Haven, CT; 5 Department of Pathology, University of Pittsburgh School of Medicine, PA; 6 Department of Medical Biochemistry and Immunology, School of Medicine, Cardiff University, Cardiff, United Kingdom; 7 Section of Medical Oncology and Department of Immunobiology, Yale University School of Medicine, New Haven, CT; and 8 Departments of Laboratory Medicine and Immunobiology, Yale University School of Medicine, New Haven, CT
Graft-versus-host disease (GVHD) remains a major cause of morbidity and mortality in allogeneic stem cell transplantation. Effector memory T cells (TEM) do not cause GVHD but engraft and mount immune responses, including graft-versus-tumor effects. One potential explanation for the inability of TEM to cause GVHD is that TEM lack CD62L and CCR7, which are instrumental in directing naive T cells (TN) to lymph nodes (LN) and Peyer patches (PP), putative sites of GVHD initiation. Thus TEM should be relatively excluded from LN and PP, possibly explaining their inability to cause GVHD. We tested this hypothesis using T cells deficient in CD62L or CCR7, transplant recipients lacking PNAd ligands for CD62L, and recipients without LN and PP or LN, PP, and spleen. Surprisingly, CD62L and CCR7 were not required for TN-mediated GVHD. Moreover, in multiple strain pairings, GVHD developed in recipients that lacked LN and PP. Mild GVHD could even be induced in mice lacking all major secondary lymphoid tissues (SLT). Conversely, enforced constitutive expression of CD62L on TEM did not endow them with the ability to cause GVHD. Taken together, these data argue against the hypothesis that TEM fail to induce GVHD because of inefficient trafficking to LN and PP.
The application of allogeneic stem cell transplantation is limited by graft-versus-host disease (GVHD). GVHD can be prevented by purging T cells from the graft; however, donor T cells are important for facilitating stem cell engraftment, providing immediate posttransplantation immunity, and eliminating residual leukemic cells.1 A key challenge is to balance the positive and negative effects of donor T cells to optimize outcome. We showed that purified CD4+ TN (CD62L+CD44–CD25–) initiated severe GVHD, whereas effector memory T cells (TEM) (CD62L–CD44+CD25–) did not but mounted a strong recall responses to a vaccination antigen.2 Chen and colleagues3 demonstrated that total TEM (CD4 and CD8 cells) did not induce GVHD but if primed in vivo against leukemia, these cells induced graft-versus-leukemia responses. These studies and others2–5 suggested that transplanting memory T cells may provide some of the benefits of donor T cells without the risk of GVHD. However, the reason for reduced GVHD induction by TEM is unclear. One fundamental difference between TN and TEM is their trafficking patterns. Naive T cells (TN) primarily circulate through the blood, spleen, and lymph nodes (LN) and are excluded from nonlymphoid tissues. In contrast, TEM generally bypass LN but can enter parenchymal tissues as well as spleen.6 These circulation patterns are largely determined by the expression of CD62L, CCR7, and CD44. TN express CD62L and CCR7, whereas TEM express neither but up-regulate CD44, which promotes entry to inflamed nonlymphoid tissues.7,8 CD62L binds to peripheral lymph node addressin (PNAd) ligands on high endothelial venules, is essential for T cell entry into peripheral lymph nodes (pLN), and promotes but is not required for T cell migration to mesenteric LN (mLN) and Peyer patches (PP).9,10 CCR7 is essential for optimal homing of TN to all LN and to a lesser extent governs entry to PP.11,12 In GVHD, donor TN and TEM encounter host antigens regardless of where they traffic. Secondary lymphoid organs, including LN and PP, contain high concentrations of "professional" antigen-presenting cells that display host antigens and are required to activate donor T cells.13–15 Therefore, it is reasonable to hypothesize that donor T cells will be effectively primed to recipient antigens in LN and PP. Thus, the inability of TEM to induce GVHD could be because they lack CD62L and CCR7 and, relative to TN, they are restricted from LN and PP. A corollary of this idea is that TN must be primed in LN and PP to induce GVHD.
In support of this hypothesis, in GVHD models, donor T cells homed to secondary lymphoid organs within 24 hours after infusion but did not infiltrate target organs until days later.16,17 Moreover, when TN and TEM were transplanted separately, TN accumulated in LN and PP, whereas TEM did not.16 Also consistent with an important role for LN priming is that interrupting the engagement of LN homing receptors CD62L and Here we use several approaches to test the hypothesis that TEM do not induce GVHD because they are restricted from LN and PP and the related hypothesis that TN must access these SLT to cause GVHD. Contrary to our expectations, blocking homing of TN to LN—thereby rendering them more like TEM—had little impact on GVHD, whereas TEM genetically modified to improve access to LN still failed to induce GVHD. Overall, these results suggest that differential ability to traffic to LN/PP plays a limited role in explaining why TN cause GVHD and TEM do not.
Mice
BALB/c, C57BL/6 (B6), 129/Sv, and CD45.1 (B6) mice were from the National Cancer Institute (Frederick, MD). B6.C mice (B6-H2d congenics) were from The Jackson Laboratory (Bar Harbor, ME). B6bm12 and IAb–/– (B6 major histocompatibility complex [MHC] II-deficient) were from Taconic Farms (Hudson, NY). BALB.B mice were from Harlan (Indianapolis, IN). CD62L–/– (B6)9 and GST2/3–/– (B6)21 mice were gifts from Tom Tedder (Duke University Medical Center, Durham, NC) and Steve Rosen (University of California-San Francisco), respectively. CCR7–/– (BALB/c)11 were a gift from Martin Lipp and Uta Hopken (Max-Delbrück-Center for Molecular Medicine, Berlin, Germany). Alymphoplasia (B6) (aly/aly)22 and heterozygous littermates (aly/+) were obtained from Fadi Lakkis (University of Pittsburgh School of Medicine, PA). Mice transgenic for CD62L with a mutated protease site (L Splenectomy Mice were anesthetized with ketamine (100 mg/kg)/xylazine (10 mg/kg) solution via intraperitoneal injection and received 0.3 mg/kg meloxicam intraperitoneally before surgery for pain relief. Splenectomies were performed as described previously.25 Postoperative treatment included ibuprofen in the drinking water (30 mg/kg/day) for 3 days after surgery and antibiotic (trimethoprim/sulfamethoxazole)-supplemented water for 7 days after surgery. Mice were allowed to recover for 3 to 5 weeks before undergoing bone marrow transplantation (BMT). Bone marrow transplantation
Transplantation was performed according to protocols approved the by Yale University or University of Minnesota IACUC. Recipients received total body irradiation as indicated from a 137Cs source (Yale) or x-irradiator (Minnesota). Three to 5 hours later, all recipients received T cell–depleted bone marrow (BM) with or without donor T cells via tail vein injection. Unfractionated spleen cells and spleen cells enriched for CD4+ or CD4+CD25– cells were transplanted to induce GVHD depending on the GVHD model. CD25+ cells were depleted in some experiments to reduce the GVHD-modulating effects of regulatory T cells.26–28 Transplanted T cell type and doses are indicated in the figure legends. For studies performed at Yale University, animals were fed moistened chow and water supplemented with trimethoprim/sulfamethoxazole for 2 weeks after BMT. For wild-type (WT) or MHC II–/– Cell separations BM cells were isolated and prepared as described previously.2 Remaining Thy1.2-positive cells were routinely less than 0.5% of BM cells as determined by flow cytometry. CD4+CD25– cells were enriched from RBC-depleted splenocytes by negative selection using streptavidin-coated magnetic activated cell sorting (MACS) beads and an AutoMACS (Miltenyi Biotec, Auburn, CA) after incubating with blocking antibody (anti-FcR, clone 2.4G2), biotinylated anti-CD8 (TIB105) and biotinylated anti-CD25 (PC61). Resulting cells had less than 1% CD4+CD25+ or CD8+ cells as determined by flow cytometry. CD4+ cells were positively selected using streptavidin-coated MACS beads and biotinylated anti-CD4 (GK1.5) on an AutoMACS. Resulting cells were more than 95% CD4+ as determined by flow cytometry. The type of CD4 purification used and whether CD25+ cells were depleted is stated in the text legend of each figure; for simplicity groups receiving any type of CD4+ cell are designated "CD4" on the graphs themselves. TN and TEM CD4+CD25– subsets were isolated by staining positively selected CD4+ cells with streptavidin-Alexa 488 (Invitrogen, Carlsbad, CA), anti-CD45RB-phycoerythrin (PE) (C363.16A; BD Biosciences, San Jose, CA), CD25-PE/Cy7 (PC61; BD Biosciences) and CD44-APC (IM7; BD Biosciences). Cells were sorted into TN (CD45RBhighCD44–) and TEM (CD45RBlowCD44+) subsets using a FACSAria (BD Biosciences). We validated this approach by analysis of B6 splenocytes stained with anti-CD4, anti-CD25, anti-CD45RB, anti-CD44, and anti-CD62L. Back-gating showed the CD4+CD25–CD45RBhighCD44– TN population to be approximately 80% CD62L+ and the CD45RB–CD44+ TEM population to be approximately 90% CD62L–. Antibodies and fusion proteins Mel-14 protein was purified from serum-free tissue culture medium as described previously.29 LTβRIg and TNFRIg fusion proteins were kind gifts of Drs Jeff Browning and Evangelia Notidis (Biogen Idec, Cambridge, MA) and were used exactly as described previously.30 Rat IgG control protein was obtained from Sigma-Aldrich (St Louis, MO). GVHD analysis Recipients were weighed on day +1 or day –1 after BMT and every 3 to 4 days thereafter. Beginning on day +16, mice were monitored for clinical signs of GVHD: diarrhea, skin disease, and hunched posture. Mice that died of radiation toxicity early after transplantation (days 0-10) before the onset of GVHD were censored with the exception of experiments shown in Figure 3. Mice with evidence of severe GVHD (very hunched and/or lethargic) or having lost more than 20% of their original weight (after recovery from irradiation) or considered clinically moribund (University of Minnesota) were killed and scored as dead. The last weight recorded for killed animals remained in the dataset. Throughout this work, unless otherwise indicated, BM-only control mice among various types of recipients in the same experiment were indistinguishable and were thus pooled for the purposes of graphing and statistical analysis. Pathologic analysis Tissues were fixed in 10% phosphate-buffered formalin, embedded in paraffin, sectioned, and stained with hematoxylin and eosin. Slides were read by pathologists who were expert in liver (A.D.), gastrointestinal (A.D, A.P.-M., and D.J.), and skin disease (J.M.) without knowledge of experimental groups. Liver scoring criteria: portal inflammation (0-3), bile duct injury (0-3), central perivenulitis (0-3), and lobular necroinflammatory activity (0-4). Colon slides in Figure 3B were scored by D.J. Scoring criteria: inflammation (0-3), apoptosis (0-3), and crypt loss (0-3). Colon slides in Figure 3D were scored by A.P.M. using published criteria.31 Colon slides in Figures 4B and 5D were scored by A.D. Scoring criteria: inflammation (0-3), apoptosis (0-4), neutrophilic abscesses (0-3), crypt loss (0-3), and ulceration (0-3).2 Statistical methods
Significance for differences in weight loss was calculated by an unpaired t test. P values for survival curves were calculated by log rank test. P values for histology comparisons were calculated by Mann-Whitney or Fisher exact test (Figure 3F, BM controls vs CD4
Functional CD62L on donor T cells was not required for GVHD To test whether CD62L-mediated homing to recipient peripheral LN was required for GVHD, we transplanted into BALB/c recipient mice CD4+CD25– T cells from B6 wild-type (WT) or CD62L–/– donors9 (Figure 1A). Recipients of WT T cells were also treated on days –1, +2, and +5 with control rat IgG or an effective dose32 of blocking antibody to CD62L (Mel-1433), which markedly reduces T-cell migration to peripheral LNs in vivo.33 Mel-14 treatment resulted in a significant but slight delay in the kinetics of GVHD death compared with control treatment, although the final mortalities were identical (Figure 1A).
CD62L–/– CD4+CD25– (Figure 1A) or CD62L–/–CD4+ T cells (Figure 1B) caused GVHD-mediated mortality similar to that induced by WT T cells (Figure 1A,B, P > .3), confirming recently published results.20 GVHD is maximal with both CD4 and CD8 cells in the MHC-matched B6  BALB.B model,34 and total CD62L–/– T cells caused GVHD-related mortality indistinguishable from that of WT T cells (Figure 1C). Taken together, these experiments indicate that TN need not express CD62L to cause GVHD. Although Mel-14 treatment slightly reduced GVHD, given the results with CD62L-deficient T cells, this may be attributable to depletion of antibody-coated T cells and/or a partial blockade of neutrophil entry to inflamed sites.35,36 To test whether CD62L–/– T cells were primed in the spleen, we transferred B6 CD62L–/– CD4+ T cells into splenectomized BALB/c recipients (Figure 1B). Unexpectedly, GVHD mortality was paradoxically accelerated in splenectomized recipients of CD62L–/– CD4+ T cells compared with spleen-intact recipients (Figure 1B). Thus, splenic priming was not required, and presumably donor T cells can be primed in other locations less dependent on CD62L for T-cell entry, such as mLN and PP. Impairing CD62L ligands on recipient LN did not ameliorate GVHD
We also tested GVHD induction by WT T cells in recipients that lack glucosyl transferases 2 and 3 (GST2/3–/–), which are necessary to generate ligands for CD62L on HEV.21 There is greatly reduced homing of T cells to pLN and a substantial reduction in homing to mLN in these mice.21 We used GST2/3–/– mice as recipients in the MHC-matched 129/Sv
Although the prior experiments with CD62L–/– T cells and GST2/3–/– recipients excluded an essential role for the engagement of CD62L on donor T cells with PNAd ligands in the host, TN also use CCR7 to enter LN and PP,11,12 and residual homing to LN in GST2/3–/– mice is probably mediated by CCR7.21 To impair both CD62L- and CCR7-mediated T-cell homing, we transplanted into GST2/3–/– recipients CCR7–/– donor CD4 T cells in the BALB/c B6 strain pairing. At day 37 after transplantation, there was equivalent death and liver pathology in GST2/3–/– recipients of either T-cell type (Figure 2C,D). Therefore reducing LN homing mediated by both CD62L and CCR7 did not significantly ameliorate GVHD. LN-, PP-, and/or spleen-deficient recipients developed GVHD That CD62L–/– T cells induced GVHD in splenectomized recipients raised the possibility that even without CD62L or CCR7, donor T cells could gain sufficient access to LN and PP to cause GVHD. Although trafficking studies that used large numbers of transferred cells have clearly shown important roles for these molecules in LN entry one or a few hours after transfer, this has been less well studied in the context of GVHD; studies have visualized T cells only from day 2 and beyond.16,17,20 In our experience, it was technically difficult to track the smaller numbers of cells typically used in a GVHD experiment at very early time points (day 1 and earlier) that reflect direct homing potential. It was not possible to sort enough TEM cells to clearly visualize them after transplantation (B.E.A., unpublished observations, January 2007).
We therefore took a more definitive approach by using recipient mice that lack LN and PP altogether. aly/aly mice are devoid of most lymphoid tissue,22 although they retain nasal-associated lymphoid tissue (NALT) and cryptopatches.22,37–39 aly/+ (with WT phenotype) or aly/aly mice were splenectomized or left intact to create the following types of recipients: WT (aly/+); LN/PP present, no spleen (aly/+Spl–); no LN/PP, spleen present (aly/aly); and no LN/PP or spleen (aly/aly Spl–). We initially used these recipients in the B6bm12
T-cell infiltration and target tissue damage occurred in the majority of T-cell recipients compared with BM controls, regardless of aly genotype and spleen presence (Figure 3B). The severity of histologic GVHD paralleled the weight loss data. In both the colon and skin, aly/+ mice had the highest average pathology scores, followed by aly/aly and then aly/aly Spl– recipients. It is difficult to make comparisons with aly/+ Spl– T-cell recipients because these organs were harvested between days 8 and 10 after BMT compared with day 23 for the other recipients. We could not evaluate the liver for GVHD because aly/aly mice develop spontaneous hepatic inflammation that is difficult to distinguish from GVHD.40 Taken together, these data show that GVHD can develop in the complete absence of LN and PP, as well as in recipients without LN, PP, and spleen. However, that GVHD is markedly reduced in aly/aly Spl– recipients relative to mice that have either LN, PP, or spleen indicates that these are important sites for donor T-cell priming that are redundant among themselves.
We extended the investigation of the roles of LN/PP to a second MHC-mismatched strain pairing, BALB/c
Because most human allogeneic stem cell transplants are MHC-matched, we investigated requirements for CD4 T-cell priming in the MHC-matched 129/Sv
aly/aly mice have defects in NF
Thus in 4 different strain pairings, CD4+ TN did not require priming in LN and PP to induce GVHD. Interestingly, depending on the model studied, manifestations of GVHD were ameliorated, equivalent or more severe in the absence of recipient LN/PP, suggesting that the relative importance of LN/PP to the GVHD syndrome is model-dependent. MHCII expression on hematopoietic cells was required for T-cell priming in aly/aly Spl– recipients
Occurrence of GVHD in aly/aly Spl– recipients suggested that parenchymal tissues alone could activate TN donor T cells when priming in SLT is prevented. To address this, we created B6/MHC II–/– Memory cells constitutively expressing CD62L did not cause GVHD
We addressed the hypothesis that TEM would induce GVHD if more efficiently recruited into LN by taking advantage of transgenic mice we created in which T cells constitutively express CD62L. These transgenic mice (L
Because we could not use CD62L as a marker to sort L
Compared with TN, TEM are much less effective at causing GVHD.2–5 We tested 2 related hypotheses to explain why TN cause GVHD and TEM do not: (1) donor T cells need to be primed in recipient LN to cause GVHD, and (2) donor TEM cells lack the expression of LN-homing molecules, in particular CD62L and CCR7, which reduces their entry to and priming in recipient LN and dampens their ability to cause GVHD. Our results argue against both of these hypotheses. With regard to the first, although CD62L and CCR7 are critically important for T-cell entry into LN, TN lacking CD62L or CCR7 caused severe GVHD. Furthermore, recipients with defective or completely absent LN still developed GVHD. Thus, restricting TN homing to sites accessible by TEM did not impair their ability to cause GVHD. With respect to the second hypothesis, we found that TEM with constitutive expression of CD62L still had very limited capacity to cause GVHD. These data formally exclude the hypothesis that priming in LN is required for GVHD induction and suggest that, at least as a consequence of reduced CD62L-expression, reduced LN homing by TEM does not by itself account for why TN cells cause GVHD but TEM do not. We took multiple approaches to suppress TN cell entry into LN. CD62L-deficiency had no effect on GVHD, while inhibiting WT donor T cells with CD62L blocking antibody delayed GVHD only slightly. We further inhibited T-cell migration to LN by infusing CCR7–/– T cells into GST2/3–/– mice, resulting in simultaneous inhibition of both chemokine and CD62L-mediated T-cell entry, yet GVHD was essentially unaffected and in some cases even exacerbated (Figure 2). Given the lack of impact of these pathways on GVHD-induction by TN, we reason that the absence of CCR7 and CD62L on TEM in turn could not solely explain their inability to cause GVHD.
Dutt et al20 also found that CD62L–/– and WT T cells initiated comparable GVHD but concluded that the ability to traffic to mLN was nonetheless critical for GVHD, because recipients of CD62L–/–β7–/– T cells had improved survival and, in particular, little gut GVHD. Yet, in several of the models we tested, gut GVHD was evident in the absence of LN/PP. To reconcile these results, we first note that even in the recipients of CD62L–/–β7–/– T cells, weight loss and histologic changes were consistent with GVHD. In addition, the apparent difference in the conclusions from the 2 studies could be ascribed to the interpretation of the effect of β7 deficiency. We suggest that genetic deletion of β7 on donor T cells ameliorated GVHD by reducing the entry of activated T cells not only into mLN but also into intestinal tissue. Thus, if priming can occur alternatively in the spleen, as our data would indicate, the Experiments exploiting genetic deficiency of CD62L, CCR7, and GST2/3 were limited by how well the particular gene deficiencies restrict T-cell trafficking as well as their potential to affect T-cell entry into target tissues. To eliminate T-cell priming in LN/PP completely, we induced GVHD in recipient animals lacking all LN/PP. In all strain pairs tested, recipients deficient in all LN/PP, but with an intact spleen, still developed GVHD. GVHD was ameliorated (but present) in 2 models, exacerbated in one model, and equivalent in a fourth. Taken together, these data suggest that priming in LN/PP is not required to induce GVHD but may be required for maximal GVHD in certain genetic mismatches or experimental conditions. Somewhat surprisingly, we found that GVHD developed in splenectomized aly/aly recipients that lacked essentially all SLT, although the disease was reduced compared with intact controls. BM chimera experiments proved that BM-derived APCs were still required to initiate GVHD in the absence of spleen, LN, and PP. These experiments did not address where T cells were primed. Potential sites include the liver,43 bone marrow,44 NALT or cryptopatches, or other inducible gut- or lung-associated lymphoid tissue.45 Indeed, recent studies showed that CD8 responses to influenza infection do not require secondary lymphoid organs.45,46 Although GVHD initiation is not accompanied by the same infection-induced inflammatory signals, secondary lymphoid architecture may not be absolutely required when antigen is ubiquitous, as is the case with GVHD. We unexpectedly observed that altering the trafficking and priming site of donor T cells affected the nature of GVHD. In 3 MHC-mismatched models, LN/PP-intact but splenectomized recipients developed more rapidly lethal GVHD than normal recipients. There are several nonexclusive explanations for this finding. The spleen may be a source of radiation-resistant recipient regulatory cells, which can suppress GVHD.47 If the spleen traps alloreactive cells,48,49 in its absence donor T cells may have enhanced ability to migrate to target organs and LN. Finally, T cells activated in the spleen versus LN may develop differently. For example, there may be altered cytokine skewing or expression of alternate homing receptors,6,50–52 which in some contexts could affect GVHD induction. These results are relevant to a related article that was submitted and published while our manuscript was under review.53 In parallel with our studies, but using a single MHC-mismatched model of GVHD, Beilhack et al53 also used a variety of strategies to restrict trafficking and priming sites of GVHD-inducing lymphocytes. These researchers found that preventing priming in the LN/PP had no effect, from which they concluded that priming site did not alter GVHD. In contrast, we clearly found effects of absence of LN/PP alone on GVHD in 3 models—decreased GVHD in 2 and augmented skin GVHD in the third. Mice lacking spleen, leading therefore to priming in LN/PP and elsewhere, also had more severe and different GVHD. Thus, we conclude that priming site does affect the character of GVHD. The different conclusions could be attributable to different methods, as we documented GVHD by clinical assessment, weight loss, and quantitative pathologic assessment, whereas Beilhack et al53 relied on bioluminescence imaging and in some cases survival curves. Alternatively or in addition, these outcomes are model-dependent, and by studying more and different models, we uncovered the dependence of GVHD on priming site that was not found in the model used by Beilhack et al.53 These authors also concluded that there was no GVHD in mice lacking LN/PP (B6.LTa KO mice) that had also been splenectomized. However, we found, in both MHC-matched and MHC-mismatched models, that mice completely lacking SLT do get GVHD. Again, differences between our positive result and their negative result could be attributable to methods, model systems, or both.
Although our studies restricting the trafficking and priming venues of TN cells suggested that the reason TEM are unable to mediate GVHD is not the inability to traffic to particular sites, a complementary aspect of the work used the opposite approach—to restore homing capacity to TEM cells using our L
Considering our results in context,16,20,54–56 it is no longer reasonable to think that any one site, or even SLT per se, is required or critical for the induction of GVHD. However, because altering the priming site modulates disease quality or severity, we suggest that the priming site does qualitatively affect the nature of GVHD. This for example would account for the predominant effect on gut GVHD of
Contribution: B.E.A. designed and executed experiments, analyzed data, and wrote the manuscript. A.A. contributed unique reagents and insights. P.A.T. and B.R.B designed and carried out experiments. J.M.M., D.J., A.J.D., and A.P.-M. interpreted and scored histopathologic sections. W.D.S. and M.J.S. designed experiments, analyzed data, and wrote the manuscript. Conflict-of-interest disclosure: The authors declare no compet-ing financial interests. Correspondence: Mark J. Shlomchik, Department of Laboratory Medicine, Box 208035, Yale University School of Medicine, New Haven, CT 06520-8035; e-mail: mark.shlomchik{at}yale.edu.
We thank Cuiling Zhang, Catherine Matte-Martone, William Tan, and Srividhya Venkatesan for outstanding technical support. We thank Drs Steve Rosen, Thomas Tedder, Uta Hopken, Martin Lipp, Jeff Browning, Evangelia Notidis, and Fadi Lakkis for gifts of reagents. We thank Yale Animal Resources Center for expert animal care. This work was supported by National Institutes of Health grants R01-HL066279 (M.J.S. and W.D.S.), T32-HL07974 (B.E.A.), R01-AI34495, and R37HL56067 (B.R.B.). W.D.S is supported by a Clinical Scholar Award from the Leukemia & Lymphoma Society.
Submitted September 4, 2007; accepted February 16, 2008.
Prepublished online as Blood First Edition Paper, February 19, 2008
DOI: 10.1182/blood-2007-09-107953
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 USC section 1734.
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Abstract
Introduction
4β7 on donor T cells can ameliorate GVHD. Incubating donor T cells with antibodies against CD62L and CD49d (
P) and control transgenic mice with a normal protease site were described.
P < .05 for BM controls versus CD4
B signaling, which could have contributed to our findings, independent of the absence of LN/PP.
