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Tissue engineering in urology has shown considerable promise. However, there is still much to understand, particularly regarding the interactions between scaffolds and their host environment, how these interactions regulate regeneration and how they may be enhanced for optimal tissue repair. In this review, we discuss the concept of dynamic reciprocity as applied to tissue engineering, i.e. how bi-directional signaling between implanted scaffolds and host tissues such as the bladder drives the process of constructive remodeling to ensure successful graft integration and tissue repair. The impact of scaffold content and configuration, the contribution of endogenous and exogenous bioactive factors, the influence of the host immune response and the functional interaction with mechanical stimulation are all considered. In addition, the temporal relationships of host tissue ingrowth, bioactive factor mobilization, scaffold degradation and immune cell infiltration, as well as the reciprocal signaling between discrete cell types and scaffolds are discussed. Improved understanding of these aspects of tissue repair will identify opportunities for optimization of repair that could be exploited to enhance regenerative medicine strategies for urology in future studies., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Background: Patients with chronic liver disease have been shown to have impaired immune statuses. Liver transplantation (LT) is the standard treatment for end-stage liver disease patients and immunosuppressive drugs are commonly used to prevent graft rejection. There is an increasing evidence of de novo food allergies post LT., Objective: To investigate the cytokine response of peripheral blood mononuclear cells (PBMCs) of pediatric LT recipients before and six months after transplantation., Method: PBMCs collected before and six months after LT were stimulated with phytohemagglutinin (PHA), beta-lactoglobulin (BLG), tacrolimus (Tac), dexamethasone (Dex), and a combination of BLG and Dex (B+D), BLG and Tac (B+T), BLG and Tac plus Dex (B+T+D). Culture supernatants were measured for IL-5, IFN-γ and IL-10. Blood for liver function tests, complete blood counts, total IgE and specific IgE (sIgE) to cow's milk were recorded., Results: A total of five pediatric LT recipients were enrolled in the study. There were no food allergy cases. Total IgE and sIgE to cow's milk decreased significantly after LT. After transplantation, there was a significant increase in IL-5, IFN-γ and IL-10 in culture supernatants of PHA-stimulated PBMCs. Among different stimulations in post transplantation's PBMCs, the level of IL-5 significantly increased in B+D was suppressed with the combination of B+T+D. The level of IL-10 significantly increased in all conditions containing BLG both before and after transplantation., Conclusion: There was an improvement of the in vitro-cytokine responses after liver transplantations. Immunosuppressive drugs used in post transplantation had an effect on the cytokine responses.

Compared to non-sensitized renal transplant recipients, patients with preformed alloantibodies are at greater risk of cellular and humoral rejection and premature graft failure. We explored the effects of adding B-cell depleting agent (rituximab) to standard rabbit anti-thymocyte globulin (rATG) induction regimen for patients with panel reactive antibody levels >50%. Following induction therapy, 14 recipients were given two doses of rituximab (375 mg/m(2)) within the first month post-transplantation. Their long-term outcomes were compared to a historical control group of 23 recipients who received rATG alone. Graft survival at 5 years was superior with combination therapy compared to induction therapy alone (92.9 versus 48.3%, respectively, p = 0.02). While 30% of the rATG alone group experienced cellular rejection and 26% humoral rejection, none of rituximab plus rATG renal transplant recipients group had rejection. Thus, addition of rituximab to rATG provided superior outcomes to rATG alone. This combination induction therapy should be considered for a high-risk population.

Biointegration refers to the interconnection between a biomedical device and the recipient tissue. In many implant devices, the lack of proper biointegration can cause device failure and potentially serious medical problems. This review summarizes the recent progress in surface chemistry, drug delivery and antifouling methods to improve the biointegration of implants. Much progress has been made as our understanding of biological systems and material properties expands and as new technologies become available. This article addresses methods of enhancing biointegration by means of modifying implant surface chemistry and by drug-delivery approaches.

Islet transplantation is one of the promising ways to treat diabetes. To reduce the immune system response, several methods have been developed, a novel one being the grafting of methoxy polyethylene glycol (mPEG) derivatives onto collagen capsules of islets. In this study, the effects of the first and second generations of activated mPEG on the immunological response of polyethylene glycol (PEG) grafted pancreatic islets were studied. mPEG-Succinimidyl carbonate (mPEG-SC) and mPEG-succinimidyl propionic acid (mPEG-SPA) (with nominal molecular weight 5 kDa), typical of the first and second generations of activated mPEG, were selected, respectively. Both activated mPEGs did not affect the morphology, viability, or functionality of PEGylated islets compared to free islets (naked islets). The amount of IL-2 secreted from lymphocytes co-cultured with mPEG-SPA grafted islets (131.83 ± 15.28 pg/ml) was not significantly different from that with mPEG-SC grafted islets (156.09 ± 27.94 pg/ml). These results indicated that both mPEG-SC and mPEG-SPA had the same effect for camouflaging Langerhans islets, but the former is more suitable due to its easier synthesis process.

Immunosuppressive and antiproliferative effects of heparin may be beneficial in the field of solid organ transplantation. The aim of this study was to examine the effect of low-molecular-weight heparin (LMWH) compounds on the development of obliterative airway disease (OAD) in the rat tracheal transplant model. Allogenic heterotopic tracheal transplantations were performed from Brown-Norway into Lewis rats. Recipients were treated either with nadroparin, enoxaparin, parnaparin, or vehicle from day 0 until harvesting at day 7 or 21. Graft rejection was morphometrically assessed to determine the extent of luminal obliteration end epithelial necrosis. All tracheal grafts harvested at day 7 demonstrated nearly equivalent degree of luminal obstruction regardless of treatment regimen. Likewise, at day 21 the extent of airway narrowing and the degree of inflammatory cell infiltration were similar among the groups. Moreover, loss of airway epithelium was not prevented by LMWH treatments. Finally, intragraft mRNA expression for transforming growth factor-β1 and platelet-derived growth factor-A, interleukin-2, interferon-γ, and monocyte chemoattractant protein-1 did not differ between the groups. In contrast with findings in other animal models, treatment with LMWH preparations did not modify the development of OAD in rat tracheal allografts.

The ongoing quandary in kidney transplantation is discovering methods to prolong graft survival. To achieve this, there is a search for optimal methods to use immunosuppressive therapy, where rejection and chronic graft damage is minimized without causing an increased risk of infections, malignancy, or toxicities. The purpose of this review was to discuss the limitations of current immunosuppressant drug monitoring as well as the clinical application of novel methods of monitoring both immunosuppressants and the immune reaction within the allograft., (2010 National Kidney Foundation, Inc. Published by Elsevier Inc. All rights reserved.)

Host antidonor effector T cells represent a major barrier to the successful engraftment of allogeneic donor hematopoietic progenitor and stem cells. Here, administration of a complex of IL-2 and anti-IL 2 antibodies (IAC) significantly enhanced donor chimerism early as well as long-term engraftment following reduced-intensity conditioning (RIC) and allogeneic major histocompatibility complex (MHC)-matched hematopoietic cell transplantion (HCT). Timing of administration of this complex was crucial: administration of IAC post-HCT more efficiently facilitated marrow engraftment than pre-HCT treatment. Donor chimerism persisted to >6 months post-HCT. Importantly, this approach clearly suppressed the emergence of host antidonor CD8 T cells 2 to 3 weeks post-HCT as assessed by tetramer staining. Following in vivo reactivation of IAC-treated and control recipients at >5 months post-HCT with donor antigen, only PBS-treated control marrow allograft recipients responded with tetramer-binding CD8 cells. In total, the present findings support the notion that the transient activation and expansion of host Tregs in situ post-HCT can be explored as a new approach to regulate host alloreactivity posttransplant. Interestingly, direct stimulation of recipient Treg cells in RIC recipients obviated a requirement for exogenous Treg cell transfusion in this model and may represent a viable alternative to, and/or complement the adaptive transfer of Treg populations in clinical HCT.

Background: We evaluated the importance and mechanism of graft and host accommodation in hamster-to-rat cardiac xenotransplantation models., Methods: To evaluate graft accommodation, accommodated hamster grafts (Group 2) were transplanted to naïve host rats treated with FK506, and compared with naïve hamster grafts (Group 1). To evaluate host accommodation, three groups were evaluated: naive hamster hearts were transplanted to naïve hosts treated with FK506 (Group 3: 0.5 mg/kg, Group 4: 1.0 mg/kg) and splenectomy, and compared with accommodating hosts (Group 5) with FK506 0.5 mg/kg and splenectomy. We examined graft survival, histopathology, antihamster antibodies and B-1 cells in blood., Results: Graft survival in Group 2 (3.4+/-0.9 days) was not significantly different from that in Group 1 (2.8+/-0.4 days). Graft survival in Groups 4 and 5 (>30 days) was significantly prolonged compared with that in Group 3 (6.0+/-0.7 days). Histopathology of Groups 1-3 showed humoral rejection, whereas Groups 4 and 5 showed normal histology and expression of protective genes. In Groups 1-3, antihamster immunoglobulin (Ig) M and B-1 cells increased significantly compared to Groups 4 and 5, where IgM and B-1 cells remained low or were reduced., Conclusions: Host accommodation was more important than graft accommodation. Accommodating grafts expressing protective genes were rejected with an elevation of both IgM and B-1 cells. In accommodated hosts, both IgM and B-1 cells decreased, suggesting that B-1 cells may be responsible for the production of antihamster antibodies. These results suggest that sufficient suppression of B-1 cells, resulting in decreased titers of antihamster antibodies, may play an important role in host accommodation.

Background and Objectives: Interleukin-7 (IL-7) has been studied for its possible immunorestorative capacities following stem cell transplantation and has been shown to enhance post-transplant immune recovery predominantly by peripheral T-cell expansion. A major concern of IL-7 is its possible aggravating effect on graft-versus-host and host-versus-graft reactivity., Design and Methods: To study the effect of IL-7 on host-versus-graft reactivity, we applied IL-7 in an experimental transplantation model using RAG-1-/- mice supplied with B6 CD45.1 congenic T cells as recipients of T-cell depleted allogeneic bone marrow grafts., Results: Rejection of minor antigen-mismatched bone marrow was significantly reduced in IL-7 treated recipients compared with PBS treated control mice. Rejection was observed in 2 out of 18 IL-7 treated mice compared with 9 out of 17 PBS treated mice (11% vs. 53%; p=0.012). IL-7 administration resulted in enhanced recovery of peripheral blood CD4+CD25+ regulatory T cells (Treg) with a concomitant increase in peripheral blood Foxp3 mRNA expression. IL-7Ra (CD127) was expressed by the vast majority of CD4+Foxp3+ T cells. The incidence of graft rejection following fully MHC mismatched bone marrow transplantation was not reduced nor enhanced by IL-7 administration., Interpretation and Conclusions: Post-transplant IL-7 administration protects against minor antigen-mismatched bone marrow rejection, which may be due to enhanced Treg recovery.

The use of nonmyeloablative conditioning prior to bone marrow transplantation is an important component of transplantation-based therapies for nonmalignant blood diseases. In this study, treatment of recipient mice with granulocyte colony-stimulating factor (G-CSF) prior to low-dose total body irradiation (LD-TBI) enhanced long-term engraftment of freshly isolated congenic marrow 1.5- to 2-fold more than treatment with LD-TBI alone. This combined regimen was also evaluated in a mouse model of X-linked chronic granulomatous disease (X-CGD), where neutrophils have a defective NADPH oxidase due to genetic deletion of the gp91(phox) subunit. Long-term engraftment of male X-CGD bone marrow cells cultured ex vivo for retroviral transduction of gp91(phox) was enhanced by approximately 40% when female X-CGD recipients were pretreated with G-CSF prior to 300 cGy. These data confirm that sequential treatment with G-CSF and LD-TBI prior to transplantation increases long-term engraftment of donor marrow, and they extend this approach to transplantation of murine donor marrow cultured ex vivo for gene transfer. Additional studies showed that the administration of G-CSF prior to LD-TBI did not alter early homing of donor marrow cells. However, the combined regimen significantly decreased the content of long-term repopulating cells in recipient marrow compared with LD-TBI alone, as assessed in competitive assays, which may contribute to the enhanced engraftment of donor marrow cells. Disclosure of potential conflicts of interest is found at the end of this article.

The combination of 8-methoxypsoralen (8-MOP) and long wave ultraviolet radiation (UV-A) has immunomodulatory effects and might abolish both graft-vs-host and host-vs-graft reactions after allogeneic hematopoietic stem cell transplantation. In the present study, we have confirmed the sensitivity of T lymphocytes to 8-MOP treatment plus UV-A exposure as evidenced by the abrogation of the alloreactivity in mixed lymphocyte cultures as well as the inhibition of the response to phytohemagglutinin A. However, the clonogenic capacity of the bone marrow hematopoietic progenitors was inhibited with UV-A doses lower than the doses needed to inhibit T-lymphocytes alloreactivity. Moreover, long-term bone marrow cultures showed that 8-MOP plus UV-A treatment had detrimental effects on the more immature bone marrow stem cells. These data were confirmed when murine bone marrow graft was treated with 8-MOP, exposed to UV-A, then transplanted into semiallogeneic recipient mice. The treated cells could not maintain their clonogenic capacity in vivo resulting in death of all animals. Taken together, these data show that ex vivo 8-MOP plus UV-A treatment of the marrow graft cannot be used to prevent post-bone marrow transplantation alloreactivity.

A stem cell transplant candidate whose best-matched related donor had a mismatch for A68 by virtue of the patient's maternal human leukocyte antigen A/B (HLA-A/B) recombinant haplotype, was referred to Wilford Hall Medical Center for transplantation. She was determined prior to transplant to have a high-titered antibody to that HLA type and to several crossreactive antigens (A2 and A24). When she initially failed to engraft, plasmapheresis was tried three times with no effect on the antibody titer. Subsequently, plasmapheresis followed by intravenous immunoglobulin treatment and retransplant completely eliminated the donor-specific antibody. Engraftment occurred, and several weeks after the second transplant antibodies to A68 could no longer be detected although the antibodies to other HLA types were still present. This case report should serve to remind stem cell transplant programs of the importance of checking for the presence of recipient antibodies to donor-mismatched antigens and suggests that patients with such antibodies be treated prior to transplant.

The combined effects of RAD and Neoral were tested in a rat orthotopic small-bowel transplantation model. Seven groups (n = 6) were involved in this study, and each one was included in three rejection models for the evaluation of host-vs.-graft disease (HVG) (LBN-F1 to LEW), graft-vs.-host disease (GVH) (LEW to LBN-F1), and combined HVG and GVH immune responses (BN to LEW). Both drugs were administered orally throughout the study. Low doses of RAD (1.0-2.5 mg/kg/day) combined with Neoral (2.0-5.0 mg/kg/day) produced strong synergistic effects in the prolongation of small-bowel graft survival in HVG (combination index, CI = 0.095, 0.1212), GVH (CI = 0.027, 0.020), and combined HVG and GVH immune responses (CI = 0.070, 0.301). The combination therapy of RAD and Neoral produces a strong synergistic effect toward the inhibition of HVG, GVH, and combined HVG and GVH immune responses in a rat small-bowel transplantation model., (Copyright 2003 Wiley-Liss, Inc.)

Background: Copolymer 1 (Cop 1) was previously shown to prevent graft-versus-host disease and interfere in various manifestations of immune rejection. In this study, we tested whether Cop 1 can also hinder the reactivity of host against graft and inhibit graft rejection., Methods: Cop 1 was tested in two transplantation systems: skin and thyroid grafting assays. The effect of Cop 1 on T cell reactivity was investigated by proliferation and cytokine secretion of spleen and lymph node cells from transplanted mice, as well as the T cell lines generated therefrom., Results: Cop 1 treatment prolonged skin graft survival and inhibited the functional deterioration of thyroid grafts, in various strain combinations, across minor and major histocompatibility barriers, similarly to the potent immunosuppressive drug FK506. Cop 1 inhibited the proliferation of graft-specific T cell lines, as well as their interleukin (IL)-2 and interferon-gamma (IFN-gamma) secretion, when incubated in vitro with the stimulating allogeneic cells. Spleen and lymph node cells from Cop 1-treated mice, as well as the T cell lines generated from them, demonstrated a pronounced inhibition of proliferation and secretion of T helper (Th)1 cytokines in response to graft cells. In addition, cells from Cop 1-treated mice secreted high amounts of Th2 cytokines in response to Cop 1 and small but significant amounts of Th2 cytokines, mainly IL-10, in response to allograft cells., Conclusions: Cop 1 treatment inhibited the Th1 response to graft and induced a Th2 cytokines secretion in response to both Cop 1 and graft cells, leading to improved survival and function of the transplanted grafts.

Background: We aimed to identify the polymorphic epitopes that mitigate graft-versus-host disease (GvHD) and host-versus-graft response (HvGR) toward rat small bowel allografts in rats., Methods: We tailored class I major histocompatibility complex (MHC) allochimeric antigens encoding 10 al-helical (alpha(1h)l58-80-RT1.Aa) or 4 (alpha(1h)l/u62-69-RT1.Aa) polymorphic amino acids. In the GvHD model, ACI (RT1a) donors were pretreated (day -14) with an intrathymic injection of alpha(1h)l58-80-RT1.Aa, alpha(1h)l/u62-69-RT1.Aa, or RT1.Al protein, with or without simultaneous intravenous injection of anti-T-cell receptor R73 monoclonal antibodies. Wistar-Furth (WF; RT1u) donors were tested with a similar protocol. In the HvGR model, ACI recipients were treated with a protocol designed to induce transplantation tolerance toward WF heart allografts: a portal vein injection of alpha(1h)l/u62-69-RT1.Aa protein and cyclosporine (4 mg/kg, intramuscular; days 0-6)., Results: GvHD was prevented in all (ACI x LEW) F1 recipients (RT1a/l) by pretreating ACI donors with R73 monoclonal antibody and recipient RT1.Al or alpha(1h)l58-80-RT1.Aa protein. Similarly, pretreatment of WF donors with RT1.Aa protein also prevented GvHD in (ACI x WF) F1 recipients. However, in a combined GvHD/HvGR model, ACI recipient perioperative treatment designed to prevent HvGR only modestly prolonged WF small bowel allograft survival (27.7+/-5.3 days compared to 17.4+/-4.6 days in the cyclosporine-alone group). In contrast, application of the two protocols significantly prolonged WF allograft survival (55.6+/-34.6 days), with two of seven recipients surviving more than 100 days., Conclusion: Simultaneous inhibition of GvHD and HvGR significantly prolongs small bowel allograft survival.

Conditioning regimens have been intensified to a level at which organ toxicties are dose-limiting, which restricts the application of hematopoietic stem cell transplants to relatively young patients in otherwise good clinical condition. Studies done in a canine model have demonstrated that stable allogeneic mixed donor/host hematopoietic chimerism can be established by the administration of a sublethal dose of 2.0 Gy total body irradiation followed by immunosuppression with mycophenolate mofetil and cyclosporine after major histocompatibility complex-identical marrow transplantation. Both host-versus-graft and graft-versus-host reactions are controlled with mycophenolate mofetil and cyclosporine, which results in a stable state of graft/host tolerance manifested by stable mixed donor/ host hematopoietic chimerism. Current efforts are directed at replacing pretransplant radiation by anti-T-cell reagents, such as antibodies to T cells, or by purine antagonists, such as pentostatin (Nipent; SuperGen, San Ramon, CA). Given the minimal toxicity of this approach in dogs, a clinical study was initiated that uses an almost identical conditioning regimen. Thus far, 26 patients have been treated. Results to date indicate that this is a well-tolerated procedure that can be performed entirely in an outpatient setting. All patients have shown primary engraftment with persistence of mixed or full donor chimerism present through at least 2 months after transplant. Three patients experienced nonfatal graft rejection between 2 and 3 months after transplant, with a return to baseline peripheral counts over the subsequent 1 to 2 months. Acute graft-versus-host disease developed in 10 of 24 evaluable patients, occurring only after discontinuation of mycophenolate mofetil, and was controlled with additional immunosuppression in all cases. Overall, this novel nonmyeloablative conditioning regimen has been well tolerated and has encouraged us to investigate these transplants in other clinical settings, including using HLA-matched unrelated donors.

Traditional approaches to allogeneic stem cell transplantation have relied on the use of toxic high-dose conditioning therapy to achieve allogeneic engraftment and control of underlying disease. Preclinical observations have shown that, for engraftment purposes, conditioning regimens can be reduced in intensity, resulting in reduced treatment toxicities. In preclinical canine studies, the use of potent pre- and postgrafting immunosuppression allowed for reduction in conditioning regimens and development of stable mixed chimerism. If these newer approaches using attenuated conditioning regimens can be successfully applied to human transplantation, an improved safety profile will allow potentially curative treatment of patients not currently offered such therapy. Mixed chimerism per se could prove curative of disease manifestation for various nonmalignant disturbances of the hematopoietic and immune systems. For patients with malignancy, infusion of additional donor lymphocytes may be needed to effectively treat underlying disease.

Desymmerization of symmetric FTY720 by substitution of different alkyl groups for one of the prochiral hydroxymethyl groups was performed. The size of the alkyl groups and the absolute configuration at quaternary carbon were important on immunosuppressive activity.

We hypothesized that the adherence of leukocytes to a vascular graft surface is mediated by B2 integrins. We studied integrin expression and monoclonal antibody inhibition of peripheral blood leukocyte (PBL) binding in vitro and CD11b expression in vivo. Human PBL were grown on ePTFE in culture and labeled with monoclonal antibodies to CD11a, -b, -c, and/or CD18. Percentage of monoclonal antibody binding and inhibition of leukocyte adherence was studied for up to 30 min using flow cytometry. ePTFE segments were implanted subcutaneously in SH-1 mice and PBL harvested 4 days later. PBL binding of monoclonal antibodies against CD11b was measured using flow cytometry. CD18 was constitutively expressed and CD11a decreased over time. CD11b increased from 41 to 62% and CD11c increased transiently (P < 0.003, P < 0.005). Inhibiton of PBL adherence was greatest by CD11b (34%) and CD11b + CD18 (57%) (P < 0.005, P < 0.0025). Implanted ePTFE caused a fourfold increase in PBL monoclonal antibody binding of CD11b (P < 0.001) compared to sham, Staphylococcus aureus alone, and combination of ePTFE and S. aureus. In conclusion, leukocyte integrins play a central role in the interaction between PBLs and ePTFE as measured by binding of monoclonal antibodies and inhibition of PBL adherence. This is also true in vivo since PBL increase CD11b expression four times when ePTFE is compared to sham. These observations indicate a potential role in vivo for B2 integrins in vascular prosthetic infections.

The induction of donor-specific unresponsiveness in allograft recipients would lessen the need for chronic immunosuppression and its concomitant morbidities. In view of recognized interactions between the immune and neuroendocrine systems, we hypothesized that manipulating prolactin (PRL) levels might enhance the immunosuppressive effects of donor-specific blood transfusions. Bromocriptine (BR) and domperidone (DOM), administered via osmotic pumps, were used to inhibit or increase pituitary PRL secretion, respectively, in male LEW rats treated with donor-specific transfusions (DST, Day -1), cyclosporine (CsA, 5 mg/kg, Days -1 to +1), and receiving ACI heart allografts. Neither compound had direct effects on lymphoid cells in vitro. BR had no effects on graft survival in rats treated with either BT or CsA (BR-DST, 7.0 +/- 0.7; BR-CsA, 9.2+/-3.1; CsA, 11.3 +/- 3.9 days). DOM-DST-CsA also did not affect graft survival (8.7 +/- 3.1 days). BR and CsA, similarly, had no effects in rats receiving a nonspecific transfusion (8.8 +/- 1.1 days). In contrast, BR administration in rats treated with DST and CsA unequivocally prolonged graft survival (17.0 +/- 1.4 days; P < 0.01 vs all controls), suggesting that hypoprolactinemia increased the tolerogenic effects of DST. Spleen and lymph node cells harvested from BR-DST-CsA rats on Postoperative Day 8 showed impaired responses to mitogenic or allogeneic challenges. Cytotoxic antibody levels at Day 5 were low in all groups receiving CsA. Possible mechanisms are discussed.

In the present experiments, a multimodality regimen was developed that included an anti-T cell receptor R73 monoclonal antibody and the pharmacologic agents brequinar (BQR), cyclosporine (CsA), and sirolimus (rapamycin; RAPA) to prolong the survival of small bowel (SB) allografts. BQR was the most potent single drug: the 4.0 or 8.0 mg/kg/day BQR doses delivered every second day (q.o.d.) per gavage for 28 days prolonged the survival of Brown Norway (BN; RT1n) SB allografts in Lewis (LEW; RT1l) recipients from a mean survival time of 10.6 +/- 1.9 days in untreated controls to 29.2 +/- 5.8 days, respectively (both P < 0.001). When treatment was extended to 56 days, 8.0 mg/kg/q.o.d BQR produced a mean survival time of 83.8 +/0 33.8 days (P < 0.001), with 2/5 hosts surviving more than 100 days. In a host-versus-graft model, BQR (8.0 mg/kg/q.o.d) delivered for 28 days with CsA (2.0 mg/kg/day) and RAPA (0.04 mg/kg/day) delivered intravenously for 14 days prolonged the survival of BN SB grafts in LEW recipients to 54.4 +/- 21.0 days (P < 0.001). Extending triple-drug therapy to 42 days induced the prolongation of SB allograft survival to greater than 100 days in 5/7 recipients. Although pretransplant perfusion of the grafts with R73 mAb was ineffective alone, the combination of graft perfusion and a 28-day course of BQR (8.0 mg/kg/q.o.d) in the GVH model indefinitely prolonged LEW graft in F1 recipients. Alternatively, indefinite survival of SB allografts ( > 100 days; P < 0.001) was achieved by the combination of a 14-day course of a triple-drug regimen using each agent at subtherapeutic doses, namely BQR (2.0 mg/kg/q.o.d.), CsA (2.0 mg/kg/day), and RAPA (0.04 mg/kg/day). The state of transplantation tolerance is these hosts was documented by the acceptance of donor-type but not third-party heart allografts.

The effect of rapamycin (RAPA) as graft pretreatment was evaluated in orthotopic small bowel and kidney allotransplantation (Tx) in the rat. In the small bowel Tx model, six groups were involved, each including three combinations for evaluation of host-versus-graft (HVG) [Lewis (LEW) x Brown Norway (BN) (LBN)-F1-->Lewis], graft-versus-host (GVH) (LEW-->F1), and combined HVG and GVH immune responses (BN-->LEW). RAPA graft pretreatment alone (16 micrograms/ml x 3 ml) was able to induce a modest but significant prolongation of survival in all three combination models compared with controls (P < 0.05). The same was observed for low dose CsA treatment (2 mg/kg/day x 14 days) of the recipient only (P < 0.05). Combination of graft pretreatment with RAPA and CsA recipient treatment produced a marked prolongation of survival especially in HVG response. Recipients treatment with one 48-microgram bolus of RAPA i.v. immediately after graft revascularization failed to achieve any prolongation of survival for the GVH or combined HVG and GVH responses. This seems to exclude a "carry-over" effect of RAPA from graft to recipient. RAPA efficacy was also clearly confirmed in the kidney graft pretreatment model as compared to recipient treatment with an equivalent RAPA dose. These results demonstrate that graft RAPA pretreatment prolongs SB survival after Tx in the rat for HVG, GVH, and bidirectional immune responses. Intragraft interaction with passenger leukocytes or APC function appears as one of the possible mechanisms. RAPA graft pretreatment potentiates low dose CsA recipient treatment suggesting a possible use in clinical organ Tx.

The effect of lithium chloride (LiCl) administration on the stress-induced depression of cell-mediated immunity was studied in mice. Two models of stress-induced depression of immunity were used: (1) keeping the animals at a temperature of 4 degrees C twice for 24 h at a 24-h interval, and (2) keeping them in the dark for 96 h. Both kinds of stress significantly decreased the reactivity of cell donors in the graft-versus-host (GVH) reaction and recipients in the host-versus-graft (HVG) reaction. Treatment with LiCl of the cell donors in GVH reaction or the recipients in HVG reaction daily for three weeks before stress application completely abolished the immunosuppressive effect of "cooling" stress, but not "darkness" stress. The LiCl is thus shown as a potential immunomodulator protecting mice from some forms of the stress-induced depression of cell-mediated immunity.

Allograft transplantation with concomitant chemotherapy has proven successful in the treatment of malignant bone tumors. However, these chemotherapeutic agents may delay tissue healing, resulting in clinical complications. To clarify the effects of cisplatin on the healing of bone grafts, we studied the incorporation of stably fixed massive diaphyseal femoral syngeneic and allogeneic grafts in rats treated with cisplatin. These data were compared with those of historical controls from animals that did not receive cisplatin. Rats that were to receive a fresh syngeneic graft or frozen allogeneic graft were given cisplatin every 4 weeks starting 9 weeks preoperatively and continuing until the time of death. The total bone area of the graft in animals that received cisplatin was smaller than that of the graft in untreated control rats that did not receive cisplatin. The area of the frozen allograft did not increase between 2 and 4 months. Revascularization was incomplete in cisplatin-treated groups at 2 months, but by 4 months, vessel ingrowth in fresh syngeneic grafts approached control values. Frozen allografts remained poorly revascularized at 4 months. Host-graft union was poor at 2 months in cisplatin-treated rats compared with controls. In cisplatin-treated rats, the host-graft union of the frozen allograft remained inferior at 4 months while that of the syngeneic graft improved. Allogeneic cortical bone grafts are incorporated more slowly and incompletely than syngeneic grafts, and this handicap is exacerbated by the administration of cisplatin.

Rapamycin, a novel macrocyclic immunosuppressive agent, suppresses murine T cell activation in vitro by mechanisms distinct from cyclosporin A (CsA). This study was designed to examine rapamycin and CsA in the host vs graft popliteal lymph node (PLN) model, an in vivo system of T cell-dependent lymphocyte activation. The PLN procedure was modified by using irradiated CTLL-2 cells of C57BL/6 origin, instead of primary mouse splenocytes, as the allogeneic stimulus in C3H/HeN recipient mice. PLN cell proliferation was determined by [3H]-thymidine uptake. We found that the host lymphocyte proliferative response to CTLL-2 cells (H-2b) is greater than the response to mouse Balb/c splenocytes (H-2d). Rapamycin (ip or po) produced a dose-related inhibition of the in vivo mixed lymphocyte reaction. By contrast, the effects of CsA and FK-506 were not dose related within the same dose range (0.006-12 mg/kg). These data indicate that rapamycin is an effective immunosuppressive agent and confirm its ability to affect the allogeneic T cell response in vivo. Furthermore, the pharmacological data suggest that this PLN model utilizing irradiated CTLL-2 cells as an allogeneic stimulus provides a reproducible system to examine mixed lymphocyte reactions in vivo.

Using the experimental model of chicken-rabbit penetrating corneal grafting, micro-whole-blood 3H-TdR incorporation and ELISA the authors dynamically observed the functional state of cellular and humoral immunities for 10 weeks after operation. The results showed, (1) In both the homogroup and the hetero-group, the transformational degree of lymphocytes in peripheral blood increased, most obviously in the 3rd or 4th week postoperatively. (2) In the hetero-group, specific anti-corneal antibody appeared obviously in the serum during 2-8 weeks postoperatively. Combined with morphologic observation, it has proved that: (1) The immune rejection is mainly cellular mediated immunoresponse, meanwhile, the humoral immunoresponse also participates in it, and the humoral immunoresponse participates in the heterokeratoplasty more obviously. (2) Applying the micro-whole-blood 3H-TdR incorporation is capable of determining the immunologic state of the body after corneal transplantation. (3) Attention should be focused on the 3-4 week period postoperatively when immunosuppressive-agent is administered.

The effect of rapamycin (RAPA) on both host-versus-graft (HVG) and graft-versus-host (GVH) immune responses was examined in small bowel transplant models using strongly histoincompatible donor-recipient combinations. Normal Wistar Furth (WFu; RT-1u) recipients rejected Buffalo (BUF; RT-1b) small bowel allografts within a mean survival time (MST) of 10.5 +/- 0.5 days. Administration of RAPA (0.8 mg/kg) by continuous intravenous infusion for 14 days via an osmotic pump prolonged graft survival to 25.0 +/- 4.6 days (P = 0.01). In a second strain combination, the 12.5 +/- 2.2 day survival of Brown Norway (BN; RT-1n) small bowel allografts in Lewis (RT-1l) recipients was prolonged to 21.6 +/- 2.0 and 28.5 +/- 2.8 days by 14 days of i.v. RAPA at doses of 0.8 and 1.6 mg/kg, respectively. In this model RAPA is five times more effective than cyclosporine, which at 4.0 mg/kg prolongs BN small bowel allografts in Lewis recipients to 21.6 +/- 6.3. To isolate HVG and GVH immune responses, (BN x Lewis)F1 hybrid rats served as the graft donor or host, respectively. In the HVG model, (BN x Lewis)F1 small bowel allografts, which were rejected by normal Lewis recipients at 12.2 +/- 3.6 days, were prolonged to 40.8 +/- 5.8 days (P = 0.001) by RAPA (0.8 mg/kg x 14 days). In the GVH model, the ability of Lewis small bowel allografts to produce severe GVH disease in untreated (BN x Lewis)F1 recipients at 12.3 +/- 2.8 days was delayed to 21.3 +/- 5.2 days by 0.8 mg/kg RAPA (P = 0.025). Thus, RAPA protects small bowel allografts more effectively against HVG than GVH immune responses.

In mice immunized with staphylococcal vaccine the arresting of graft-versus-host reaction under the influence of small doses of staphylococcal vaccine, hyperimmune antistaphylococcal serum, cyclophosphamide, antilymphocytic serum has been demonstrated. Small doses of staphylococcal vaccine stimulated the production of antibodies to staphylococci and dermal extract in the animals, previously immunized with this vaccine, with the simultaneous suppression of cell-mediated immune reactions to both antigens. Immunosuppressing agents have been found to inhibit humoral and cell-mediated immune response to microbial antigen and dermal extract. No influence of vermox and levamisole on the outcome of the graft-versus-host reaction has been registered; the latter preparation has been found to intensify cell-mediated immune reactions to microbial and tissue antigens.

Objective: To present a case in which acute failure of a liver graft was treated with an infusion of prostaglandin E1 (PGE1)., Clinical Features: A nine-year-old girl underwent liver transplantation for Wilson's disease. Her course was complicated by primary non-function of the graft as evidenced by poor bile production, rising bilirubin and liver enzyme levels and a severe coagulopathy., Intervention and Outcome: PGE1 was infused and there was recovery of graft function. The need for urgent re-transplantation was avoided., Conclusions: PGE1 may have an important role in liver transplantation considering the logistic difficulties of obtaining donor livers for urgent transplantation in Australia.

Cyclosporine (CS) is a relatively new immunosuppressive agent which is selective in its action in that it acts on lymphocytes, T and B cells, and only indirectly if at all on other haemopoietic cells. Numerous studies over the last decade have shown quite clearly that, in vitro, CS inhibits lymphocyte activation at a relatively early stage by preventing the production and release of lymphokines, probably by inhibition of lymphokine gene transcription. However, studies in vivo have produced clearly different findings--although CS in vivo is completely effective in preventing the development of mature effector function (i.e., T-dependent antibody synthesis by B cells or T cell-mediated cytotoxicity) and the consequences of that effector function (e.g., transplant rejection), there is clear evidence that lymphocytes can be activated to at least the stages of cell division and clonal expansion under cover of therapeutic levels of CS. The contradictions between in vitro and in vivo findings are discussed in terms of the possible mechanism of action of CS in vivo.

The successful use of a bone allograft may be negated by the host's immune response. This investigation assessed the efficacy of combining freeze-dried cortical allografts in three to six weeks azathioprine-immunosuppressed dogs. Forty-eight of 94 adult mongrel dogs were initiated for this study, and 46 of 94 were previously published and recompiled. The dogs were divided into five groups and followed for six months: Group I consisted of bilateral fresh autografts as an external control; Group II assessed the effect of freeze-drying on autogenous bone; Group III compared fresh autografts with fresh allografts; Group IV assessed the effect of freeze-drying on allografts; and Group V assessed the combined effect of placing freeze-dried allografts in immunosuppressed hosts. Biweekly roentgenograms were made to evaluate the time to union and the incidence of graft fatigue failure. Mechanical graft strength was assessed by rapid torsional loading to failure at the time of sacrifice. Biologic repair was assessed with the use of tetracycline and microradiographic techniques. The incorporation and repair of a fresh cortical autograft is better than that of a freeze-dried autograft because of fractures, nonunion, or delayed union of graft-host junctions; freeze-dried autografts have increased peripheral and internal resorption, yet an increased peripheral bony callus maintains normal graft strength; freeze-dried and fresh allografts are similar in roentgenographic characteristics, mechanical strength, and in the mechanism of graft incorporation; the use of three or six weeks azathioprine therapy did not improve the fate of freeze-dried allografts.

Subcutaneous (sc) immunization of mice with allogeneic spleen cells can induce delayed-type hypersensitivity (DTH) to histocompatibility antigens. Intravenous immunization with irradiated allogeneic spleen cells, on the other hand, induces suppressor T (Ts) lymphocytes. These Ts cells are capable of suppressing the host-versus-graft (HvG) DTH reactivity which normally arises after sc immunization. Moreover they can suppress the development of antihost DTH effector T cells during graft-versus-host (GvH) reactions. These models for HvG and GvH DTH reactivity were used to study the influence of 2'-deoxyguanosine (dGuo) on the induction, further development, and expression of Ts cells in vivo. It was found that administration of dGuo inhibits the proliferation-dependent induction and further development of Ts cells, but not the suppression mediated by already activated Ts cells.

Suppression of cell-mediated immune responses by essential fatty acids (EFA) was demonstrated in mice maintained on a standard laboratory diet for rodents. Daily oral administration of EFA at doses ranging from 125 to 750 mg/kg body weight significantly suppressed local host-versus-graft and graft-versus-host reactions as measured by popliteal lymph node assay. Studies employing immune sera directed against E-type prostaglandin demonstrated that n-6 EFA-induced suppression was mediated through prostaglandin E1. In titration experiments the effect of n-6 EFA on the reactions was dose dependent with enhancement of the responses at low concentrations and suppression at high concentrations.

The antitumor activity of hexamethylmelamine (HMM) was tested in various mouse tumor models in the presence or absence of host-vs-tumor graft responses. The drug was moderately active against Sarcoma-180 growing in different strains of non-sensitized mice. Strong protection was afforded when recipients were preimmunized with irradiated tumor cells 15 days before tumor challenge followed by HMM treatment. The drug did not show antitumor activity against two radiation-induced lymphomas of congenic mice of B10 background, inoculated into H-2 compatible hosts, or into mice incompatible for subregions of H-2. In this model HMM increased mortality of allogeneic mice presumably through impairment of host-vs-lymphoma graft resistance. In conclusion this study shows that synergistic or antagonistic effects can be obtained by combining chemotherapy with antitumor immune responses.

Subcutaneous (s.c.) immunization of mice with allogeneic spleen cells can induce delayed-type hypersensitivity (DTH) to both major and minor histocompatibility antigens. Intravenous immunization with allogeneic spleen cells, however, induces a poor state of DTH. Furthermore, i.v. immunization with allogeneic spleen cells, especially if they have been irradiated, induces suppressor T lymphocytes. These suppressor T cells are capable of suppressing the host-vs-graft (HvG) DTH reactivity that normally arises after s.c. immunization. Moreover, they can suppress the development of anti-host DTH effector T cells during graft-vs-host (GvH) reactions. These models for HvG and GvH DTH reactivity were used to study the influence of 2'-deoxyguanosine (dGuo) and guanosine (Guo) on the generation of DTH-reactive T cells and suppressor T cells in vivo. It was found that daily i.p. administration of 0.01 mg dGuo to mice immunized i.v. partially prevented the generation of suppressor T cell activity, whereas daily administration of 0.1 or 1 mg dGuo resulted in a complete abolition. Administration of dGuo has no effect on the anti-host DTH reactivity by spleen cells from nonsuppressed donors except for when a daily dose of 10 mg is administered. This dose proved to be toxic for precursors of DTH effector T cells. Daily i.p. injection of Guo had no effect on the generation of suppressor T cells nor on the generation of DTH effector T cells. The effect of dGuo was found to be due to a direct effect on suppressor T cells and not to the induction of contrasuppressor cells. These data suggest a differential sensitivity of DTH-reactive T cells and suppressor T cells for dGuo. Because suppressor T cells and DTH-reactive T cells require proliferation for expressing maximal functional activity in the systems used, both cell types probably have different enzyme activities involved in the purine metabolism and similar deoxycytidine kinase activities, but have different nucleotidase (5'NT) activities, those in suppressor T cells being the lowest. If so, suppressor T cells will accumulate deoxyguanosine triphosphate, which causes an inhibition of the ribonucleotide reductase activity and thus of the DNA synthesis by these cells.

5-azacytidine [5-AzCR] and 5-aza-2'-deoxycytidine [5-AzCdR] can produce an immunosuppressive or an immunostimulatory effect on the graft-versus-host or host-versus-graft reactions in the regional popliteal lymph node of the mouse. The result depends on the dose of the drug and the time between its administration and induction of the cell-mediated immune response. Syngeneic spleen cells are capable of inducing regional lymphadenomegaly if injected into the footpads of recipients treated 1 day previously or concurrently with 5-AzCR or 5-AzCdR. Spleen cells from allogeneic and syngeneic donors treated with 5-AzCR or 5-AzCdR in vivo or in vitro can elicit greater regional lymphadenomegaly than the untreated cells. Both drugs not only seem to influence the proliferative activity of effector and regulatory populations of immunocompetent cells, but they probably also change their recognition ability and immunogenic properties.