Keratinocyte Growth Factor Increases the CD4+Foxp3+ Regulatory T Cell-Pool by an Early Thymus-Independent and a Late Thymus-Dependent Mechanism.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3211-3211
Author(s):  
Marieke Bruinsma ◽  
Peter L. van Soest ◽  
Pieter J.M. Leenen ◽  
Bob Lowenberg ◽  
Jan J. Cornelissen ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Growth Factor ◽  
Regulatory T Cell ◽  
Graft Rejection ◽  
Keratinocyte Growth Factor ◽  
Marrow Graft ◽  
Cell Numbers ◽  
Dependent Mechanism

Abstract Administration of keratinocyte growth factor (KGF) before and shortly after experimental allogeneic bone marrow transplantation (allo-BMT) has been shown to reduce the incidence and severity of graft versus host disease (GVHD) and to inhibit graft rejection. As naturally occurring CD4+Foxp3+ regulatory T cells (Treg) play an important role in the prevention of GVHD and graft rejection, we evaluated the effect of KGF on peripheral and thymic CD4+Foxp3+ regulatory T cell numbers in normal mice and BMT recipients. KGF (5mg/kg/day) was administered subcutaneously to adult B6 mice for 3 consecutive days. KGF enhanced the frequency of CD4+Foxp3+ Treg in peripheral blood and spleen twofold, to more than 15 % of CD4+ T cells, resulting in an increase in absolute numbers of CD4+Foxp3+ Treg within one week. From 2 weeks onwards, the frequency of CD4+Foxp3+ Treg gradually normalized, but the absolute numbers of Treg remained elevated (> 10 weeks) due to an increase of both CD4 and CD8 T cell numbers. In order to assess the relative contribution of thymic output versus peripherally expanded Treg, we next determined the frequency of T-cell receptor excision circles (TREC) in peripheral blood T cells. Analysis of sorted CD4+Foxp3+ Treg and CD4+Foxp3− conventional T cells showed a decline in TREC frequency in both subsets 1 week after KGF administration, suggesting peripheral expansion. In contrast, the TREC frequency in peripheral T cells was significantly higher at 5 and 10 weeks after KGF, indicating a late thymic dependent effect. Normal B6 mice that were adoptively transferred with congenic T cells showed an enhanced, but transient, increase of the frequency of CD4+Foxp3+ Treg within one week, also suggesting that the early effect of KGF on peripheral Treg is independent of thymic output. In addition, we assessed the effects of KGF on Foxp3+ thymocytes. Thymic weight and thymic cellularity of all subsets, including CD4+Foxp3+ thymocytes, were 2 to 3-fold increased one week after KGF administration as compared to control mice. In addition, we found that KGF transiently affected the thymic architecture. The medullary epithelial compartment was virtually absent one week after KGF administration. Normal thymic architecture gradually reappeared after 2–3 weeks. To determine whether KGF-enhanced Treg numbers in BMT recipients would inhibit graft rejection, KGF was administered from day -3 until day -1 to Rag-1−/− mice. Subsequently, the mice were 3Gy irradiated, supplied with 105 B6 CD45.1 congenic T cells and transplanted with a MHC-matched minor-Ag mismatched T cell depleted 129Sv bone marrow graft. As in normal B6 mice, KGF-treatment of BMT-recipients enhanced peripheral CD45.1+CD4+Foxp3+ Treg numbers. This was associated with a reduced rate of bone marrow graft rejection (2 out of 8 KGF-treated mice as compared to 5 out of 7 PBS-treated mice). Taken together, our data show that KGF enhances the peripheral Treg pool by an early, transient and selective thymus-independent mechanism and thereafter by a non-selective thymus-dependent mechanism. The KGF-mediated expansion of the pool of peripheral Treg may contribute to the inhibitory effects of KGF on graft rejection.

scholarly journals Keratinocyte Growth Factor Improves Allogeneic Bone Marrow Engraftment through a CD4+Foxp3+Regulatory T Cell-Dependent Mechanism

2009 ◽  
Vol 182 (12) ◽  
pp. 7364-7369 ◽  
Author(s):  
Marieke Bruinsma ◽  
Peter L. van Soest ◽  
Pieter J. M. Leenen ◽  
Bob Löwenberg ◽  
Jan J. Cornelissen ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cell ◽  
Growth Factor ◽  
Regulatory T Cell ◽  
Keratinocyte Growth Factor ◽  
Allogeneic Bone Marrow ◽  
Allogeneic Bone ◽  
Bone Marrow Engraftment ◽  
Dependent Mechanism

scholarly journals Keratinocyte growth factor enhances DNA plasmid tumor vaccine responses after murine allogeneic bone marrow transplantation

Blood ◽  
2009 ◽  
Vol 113 (7) ◽  
pp. 1574-1580 ◽  
Author(s):  
Robert R. Jenq ◽  
Christopher G. King ◽  
Christine Volk ◽  
David Suh ◽  
Odette M. Smith ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Keratinocyte Growth Factor ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone ◽  
Effector Cells ◽  
Cd8 Cells ◽  
T Cell Reconstitution ◽  
Dna Plasmid

Abstract Keratinocyte growth factor (KGF), which is given exogenously to allogeneic bone marrow transplantation (allo-BMT) recipients, supports thymic epithelial cells and increases thymic output of naive T cells. Here, we demonstrate that this improved T-cell reconstitution leads to enhanced responses to DNA plasmid tumor vaccination. Tumor-bearing mice treated with KGF and DNA vaccination have improved long-term survival and decreased tumor burden after allo-BMT. When assayed before vaccination, KGF-treated allo-BMT recipients have increased numbers of peripheral T cells, including CD8+ T cells with vaccine-recognition potential. In response to vaccination, KGF-treated allo-BMT recipients, compared with control subjects, generate increased numbers of tumor-specific CD8+ cells, as well as increased numbers of CD8+ cells producing interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α). We also found unanticipated benefits to antitumor immunity with the administration of KGF. KGF-treated allo-BMT recipients have an improved ratio of T effector cells to regulatory T cells, a larger fraction of effector cells that display a central memory phenotype, and effector cells that are derived from a broader T-cell–receptor repertoire. In conclusion, our data suggest that KGF can function as a potent vaccine adjuvant after allo-BMT through its effects on posttransplantation T-cell reconstitution.

scholarly journals Keratinocyte growth factor (KGF) is required for postnatal thymic regeneration

Blood ◽  
2006 ◽  
Vol 107 (6) ◽  
pp. 2453-2460 ◽  
Author(s):  
Önder Alpdogan ◽  
Vanessa M. Hubbard ◽  
Odette M. Smith ◽  
Neel Patel ◽  
Sydney Lu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cell ◽  
Growth Factor ◽  
Keratinocyte Growth Factor ◽  
Critical Role ◽  
T Cell Development ◽  
Marrow Transplant ◽  
Cell Development ◽  
Allogeneic Bone ◽  
Thymic Regeneration

AbstractKeratinocyte growth factor (KGF) is a member of the fibroblast growth factor family that mediates epithelial cell proliferation and differentiation in a variety of tissues, including the thymus. We studied the role of KGF in T-cell development with KGF-/- mice and demonstrated that thymic cellularity and the distribution of thymocyte subsets among KGF-/-, wildtype (WT), and KGF+/- mice were similar. However, KGF-/- mice are more vulnerable to sublethal irradiation (450 cGy), and a significant decrease was found in thymic cellularity after irradiation. Defective thymopoiesis and peripheral T-cell reconstitution were found in KGF-/- recipients of syngeneic or allogeneic bone marrow transplant, but using KGF-/- mice as a donor did not affect T-cell development after transplantation. Despite causing an early developmental block in the thymus, administration of KGF to young and old mice enhanced thymopoiesis. Exogenous KGF also accelerated thymic recovery after irradiation, cyclophosphamide, and dexamethasone treatment. Finally, we found that administering KGF before bone marrow transplantation (BMT) resulted in enhanced thymopoiesis and peripheral T-cell numbers in middle-aged recipients of an allogeneic BM transplant. We conclude that KGF plays a critical role in postnatal thymic regeneration and may be useful in treating immune deficiency conditions. (Blood. 2006;107:2453-2460)

Involvement of Donor T-Cell Cytotoxic Effector Mechanisms in Preventing Allogeneic Marrow Graft Rejection

Blood ◽  
1998 ◽  
Vol 92 (6) ◽  
pp. 2177-2181 ◽  
Author(s):  
Paul J. Martin ◽  
Yoshiki Akatsuka ◽  
Michael Hahne ◽  
George Sale
Keyword(s):  
T Cells ◽  
T Cell ◽  
Graft Rejection ◽  
Fas Ligand ◽  
Conditioning Regimen ◽  
Marrow Graft ◽  
Hematopoietic Stem ◽  
Cd8 Cells ◽  
Allogeneic Marrow ◽  
Cytotoxic Effector

Abstract Donor CD8 cells play a pivotal role in preventing allogeneic marrow graft rejection, possibly by generating cytotoxic effectors needed to eliminate recipient T cells remaining after the pretransplant conditioning regimen or by producing cytokines needed to support the growth and differentiation of hematopoietic stem cells. In the present study, we assessed the role of donor T-cell cytotoxic effector function as a mechanism for eliminating recipient CD8 cells that cause marrow graft rejection in mice. The ability to prevent rejection was minimally affected by the presence of a defect in Fas ligand binding or by the absence of granzyme B but was severely affected by the absence of perforin. Doubly mutant perforin-deficient, Fas ligand-defective CD8 cells were completely unable to prevent rejection. Our results indicate first that recipient CD8 effectors responsible for causing marrow graft rejection are sensitive to cytotoxicity mediated by both perforin- and Fas-ligand-dependent mechanisms, and second that donor T cells must have at least one functional cytotoxic mechanism to prevent allogeneic marrow graft rejection. © 1998 by The American Society of Hematology.

scholarly journals Keratinocyte growth factor augments immune reconstitution after autologous hematopoietic progenitor cell transplantation in rhesus macaques.

Blood ◽  
2007 ◽  
Vol 110 (1) ◽  
pp. 441-449 ◽  
Author(s):  
Ruth Seggewiss ◽  
Karin Loré ◽  
F. Javier Guenaga ◽  
Stefania Pittaluga ◽  
Joseph Mattapallil ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Growth Factor ◽  
Immune Reconstitution ◽  
Progenitor Cell ◽  
Rhesus Macaques ◽  
Keratinocyte Growth Factor ◽  
Hematopoietic Progenitor Cell ◽  
Thymic Epithelial Cells ◽  
T Cell Reconstitution

Opportunistic infections contribute to morbidity and mortality after peripheral blood progenitor cell (PBPC) transplantation and are related to a deficient T-cell compartment. Accelerated T-cell reconstitution may therefore be clinically beneficent. Keratinocyte growth factor (KGF) has been shown to protect thymic epithelial cells in mice. Here, we evaluated immune reconstitution after autologous CD34+ PBPC transplantation in rhesus macaques conditioned with myeloablative total body irradiation in the absence or presence of single pretotal body irradiation or repeated peritransplant KGF administration. All KGF-treated animals exhibited a well-preserved thymic architecture 12 months after graft. In contrast, thymic atrophy was observed in the majority of animals in the control group. The KGF-treated animals showed higher frequencies of naive T cells in lymph nodes after transplantation compared with the control animals. The animals given repeated doses of KGF showed the highest levels of T-cell receptor excision circles (TRECs) and the lowest frequencies of Ki67+ T cells, which suggest increased thymic-dependent reconstitution in these animals. Of note, the humoral response to a T-cell–dependent neo-antigen was significantly higher in the KGF-treated animals compared with the control animals. Thus, our findings suggest that KGF may be a useful adjuvant therapy to augment T-cell reconstitution after human PBPC transplantation.

Pentostatin Facilitates Fully MHC-Disparate Murine Mini-Transplantation.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3539-3539
Author(s):  
Jacopo Mariotti ◽  
Kaitlyn Ryan ◽  
Paul Massey ◽  
Nicole Buxhoeveden ◽  
Jason Foley ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Immune Suppression ◽  
Graft Rejection ◽  
Mixed Chimerism ◽  
Post Transplant

Abstract Abstract 3539 Poster Board III-476 Pentostatin has been utilized clinically in combination with irradiation for host conditioning prior to reduced-intensity allogeneic hematopoietic stem cell transplantation (allo-HSCT); however, murine models utilizing pentostatin to facilitate engraftment across fully MHC-disparate barriers have not been developed. To address this deficit in murine modeling, we first compared the immunosuppressive and immunodepleting effects of pentostatin (P) plus cyclophosphamide (C) to a regimen of fludarabine (F) plus (C) that we previously described. Cohorts of mice (n=5-10) received a three-day regimen consisting of P alone (1 mg/kg/d), F alone (100 mg/kg/d), C alone (50 mg/kg/d), or combination PC or FC. Combination PC or FC were each more effective at depleting and suppressing splenic T cells than either agent alone (depletion was quantified by flow cytometry; suppression was quantified by cytokine secretion after co-stimulation). The PC and FC regimens were similar in terms of yielding only modest myeloid suppression. However, the PC regimen was more potent in terms of depleting host CD4+ T cells (p<0.01) and CD8+ T cells (p<0.01), and suppressing their function (cytokine values are pg/ml/0.5×106 cells/ml; all comparisons p<0.05) with respect to capacity to secrete IFN-g (13±5 vs. 48±12), IL-2 (59±44 vs. 258±32), IL-4 (34±10 vs. 104±12), and IL-10 (15±3 vs. 34±5). Next, we evaluated whether T cells harvested from PC-treated and FC-treated hosts were also differentially immune suppressed in terms of capacity to mediate an alloreactive host-versus-graft rejection response (HVGR) in vivo when transferred to a secondary host. BALB/c hosts were lethally irradiated (1050 cGy; day -2), reconstituted with host-type T cells from PC- or FC-treated recipients (day -1; 0.1 × 106 T cells transferred), and challenged with fully allogeneic transplant (B6 donor bone marrow, 10 × 106 cells; day 0). In vivo HVGR was quantified on day 7 post-BMT by cytokine capture flow cytometry: absolute number of host CD4+ T cells secreting IFN-g in an allospecific manner was ([x 106/spleen]) 0.02 ± 0.008 in recipients of PC-treated T cells and 1.55 ± 0.39 in recipients of FC-treated cells (p<0.001). Similar results were obtained for allospecific host CD8+ T cells (p<0.001). Our second objective was to characterize the host immune barrier for engraftment after PC treatment. BALB/c mice were treated for 3 days with PC and transplanted with TCD B6 bone marrow. Surprisingly, such PC-treated recipients developed alloreactive T cells in vivo and ultimately rejected the graft. Because the PC-treated hosts were heavily immune depleted at the time of transplantation, we reasoned that failure to engraft might be due to host immune T cell reconstitution after PC therapy. In an experiment performed to characterize the duration of PC-induced immune depletion and suppression, we found that although immune depletion was prolonged, immune suppression was relatively transient. To develop a more immune suppressive regimen, we extended the C therapy to 14 days (50 mg/Kg) and provided a longer interval of pentostatin therapy (administered on days 1, 4, 8, and 12). This 14-day PC regimen yielded CD4+ and CD8+ T cell depletion similar to recipients of a lethal dose of TBI, more durable immune depletion, but again failed to achieve durable immune suppression, therefore resulting in HVGR and ultimate graft rejection. Finally, through intensification of C therapy (to 100 mg/Kg for 14 days), we were identified a PC regimen that was both highly immune depleting and achieved prolonged immune suppression, as defined by host inability to recover T cell IFN-g secretion for a full 14-day period after completion of PC therapy. Finally, our third objective was to determine with this optimized PC regimen might permit the engraftment of MHC disparate, TCD murine allografts. Indeed, using a BALB/c-into-B6 model, we found that mixed chimerism was achieved by day 30 and remained relatively stable through day 90 post-transplant (percent donor chimerism at days 30, 60, and 90 post-transplant were 28 ± 8, 23 ± 9, and 21 ± 7 percent, respectively). At day 90, mixed chimerism in myeloid, T, and B cell subsets was observed in the blood, spleen, and bone marrow compartments. Pentostatin therefore synergizes with cyclophosphamide to deplete, suppress, and limit immune reconstitution of host T cells, thereby allowing engraftment of T cell-depleted allografts across MHC barriers. Disclosures: No relevant conflicts of interest to declare.

FTY720 Promotes Engraftment of Allogeneic Donor Bone Marrow Stem Cells but Impairs Thymopoiesis Resulting in Delayed Graft Rejection.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3530-3530
Author(s):  
Patricia A Taylor ◽  
Ryan M Kelly ◽  
Michael J Ehrhardt ◽  
Bruce R. Blazar
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Graft Rejection ◽  
Absolute Number ◽  
Cell Lineages ◽  
Single Positive ◽  
High Level

Abstract Abstract 3530 Poster Board III-467 FTY720 (FTY), a sphingosine-1-phosphate receptor agonist, inhibits lymphocyte egress from lymphoid tissues although the complete mechanism of its immunomodulatory effects is not fully understood. We previously published that FTY inhibited but did not prevent graft-versus-host disease by multiple mechanisms. Using the same dose and schedule (3 mg/kg orally d0-28) we evaluated FTY for its effect on allogeneic bone marrow (BM) engraftment in sublethally-irradiated mice. C57BL/6 mice were irradiated with 5.0 Gy total body irradiation (TBI) on day -1, and received 107 T-cell depleted BALB/c BM cells on day 0. At 5 wks, FTY-treated mice had a mean 84% ± 4% (mean ± SEM, n=47) donor chimerism in peripheral blood leukocytes (PBL) versus 5% ± 2% in water-treated controls (n=38, p<0.001). However, engraftment promotion was transient in most mice. PBL phenotyping at 3 months revealed that mean donor chimerism decreased to 22% ± 6%. Of the 32 mice that were >90% donor at 5 wks, only 6 were >50% donor at 3 months indicating that even high level donor chimeras were subject to delayed graft rejection. We found that although FTY promoted robust donor engraftment in the NK, myeloid and B cell lineages in BM, spleen, and lymph nodes by the first week after transplantation, thymopoiesis was severely impaired at 1 month resulting in near absent donor (and also host) thymic T cell production. FTY-treated mice had very low thymocyte cellularity (<7×106, n=10). Most thymocytes (65-85%) were host CD4 or CD8 single positive T cells. We hypothesized that upon cessation of FTY, which prevents thymocyte egress, the mature host single positive T cells were released into the periphery and mediated delayed graft rejection. Consistent with this hypothesis, the in vivo depletion of host T cells but not host NK cells, at the time of cessation of FTY treatment, abrogated the loss of the donor graft indicating that host T cells were responsible for delayed graft rejection. Also consistent with our hypothesis, and demonstrating the immune competence of the host T cells retained in the thymus, the adoptive transfer of thymocytes from FTY-treated engrafted mice into lethally-irradiated C57BL/6 recipients mediated donor BALB/c BM rejection. To further examine the mechanism of early and robust albeit transient engraftment promotion in some cell lineages, but near absent thymopoiesis, we evaluated the absolute number of donor lin−Sca-1+cKit+ stem cells in the BM at 1 month. For these experiments, an engrafted control was deemed to be a more useful comparator than water-treated mice that rejected their graft. To ensure an engrafted control using the same TBI and allogeneic cell dose parameters, control mice were given peri-transplant injections of anti-CD4 and anti-CD8, a strategy that depletes host T cells and results in durable high level donor chimeras. Consistent with reports that FTY supports migration and bone marrow homing of stem cells, FTY-treated mice had a 4.9-fold increase in the absolute number of donor lin−Sca-1+cKit+ stem cells in the BM compartment compared to anti-CD4/8-treated mice. We hypothesized that the lack of donor thymopoiesis was the result of common lymphoid progenitors being trapped in the BM compartment and unable to migrate to and/or enter the thymus. Consistent with this hypothesis, FTY-treated mice had 125-fold fewer donor-type linlocKithiCD25− early thymic progenitors (ETPs) compared to anti-CD4/8-treated control mice. In contrast to FTY-treated mice, anti-CD4/8-treated mice had evidence of vigorous donor thymopoiesis. Collectively these data indicate that although FTY supports donor stem cell migration/homing to the BM and early donor NK, myeloid and B cell engraftment, the block in donor thymopoiesis and retention of thymic host T cells result in only transient engraftment in most sublethally-irradiated mice. These data have important implications in the use of FTY in BMT and further warrant examination of thymopoiesis in patients receiving FTY for immune suppression. Disclosures: No relevant conflicts of interest to declare.

Requirements for the promotion of allogeneic engraftment by anti-CD154 (anti-CD40L) monoclonal antibody under nonmyeloablative conditions

Blood ◽  
2001 ◽  
Vol 98 (2) ◽  
pp. 467-474 ◽  
Author(s):  
Patricia A. Taylor ◽  
Christopher J. Lees ◽  
Herman Waldmann ◽  
Randolph J. Noelle ◽  
Bruce R. Blazar
Keyword(s):  
Monoclonal Antibody ◽  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Immune Suppression ◽  
Regulatory T Cell ◽  
Cd4 Cells ◽  
Major Goal ◽  
Cd8 Cells

The promotion of alloengraftment in the absence of global immune suppression and multiorgan toxicity is a major goal of transplantation. It is demonstrated that the infusion of a single modest bone marrow dosage in 200 cGy-irradiated recipients treated with anti-CD154 (anti-CD40L) monoclonal antibody (mAb) resulted in chimerism levels of 48%. Reducing irradiation to 100 or 50 cGy permitted 24% and 10% chimerism, respectively. In contrast, pan–T-cell depletion resulted in only transient engraftment in 200 cGy-irradiated recipients. Host CD4+ cells were essential for alloengraftment as depletion of CD4+ cells abrogated engraftment in anti-CD154–treated recipients. Strikingly, the depletion of CD8+ cells did not further enhance engraftment in anti-CD154 mAb–treated recipients in a model in which rejection is mediated by both CD4+ and CD8+ T cells. However, anti-CD154 mAb did facilitate engraftment in a model in which only CD8+ T cells mediate rejection. Furthermore, CD154 deletional mice irradiated with 200 cGy irradiation were not tolerant of grafts, suggesting that engraftment promotion by anti-CD154 mAb may not simply be the result of CD154:CD40 blockade. Together, these data suggest that a CD4+regulatory T cell may be induced by anti-CD154 mAb. In contrast to anti-CD154 mAb, anti-B7 mAb did not promote donor engraftment. Additionally, the administration of either anti-CD28 mAb or anti-CD152 (anti–CTLA-4) mAb or the use of CD28 deletional recipients abrogated engraftment in anti-CD154 mAb–treated mice, suggesting that balanced CD28/CD152:B7 interactions are required for the engraftment-promoting capacity of anti-CD154 mAb. These data have important ramifications for the design of clinical nonmyeloablative regimens based on anti-CD154 mAb administration.

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