Skin Allografts Research Articles

Prolongation of skin graft survival in mice by an azaphenothiazine derivative

Azaphenothiazines are predominantly immunosuppressive compounds. We evaluated the efficacy of an azaphenothiazine derivative, 6-chloroethylureidoethyldiquino[3,2-b;2′,3′-e][1,4]thiazine (DQT) in prolongation of survival of skin allografts between BALB/c and C57Bl/6 mice. The mice were treated intraperitoneally (i.p.) with 100 μg of DQT on alternate days, on days 1–13 of the experiment (7 doses). The effect of DQT on a two-way mixed lymphocyte reaction (MLR) in the human model, as well as its effect on production of TNF α and IL-10 in a whole blood cell culture, stimulated by lipopolysaccharide (LPS), were evaluated. In addition, DQT effects were investigated regarding the proportion of T cell subsets in human peripheral blood lymphocytes (PBMC) by flow cytometry. Lastly, the effect of DQT on expression of signaling molecules involved in pro apoptotic pathways was determined by RT PCR. The results showed that DQT significantly extended skin graft survival. The compound also strongly suppressed two-way MLR in the human model at a concentration range of 2.5–5.0 μM. In addition, DQT inhibited LPS-inducible TNF α, but not IL-10 production. The compound preferentially caused a loss of the CD3-CD8+CD11b + PBMC cell subset, and transformed CD3+CD8+high into CD3+CD8+low cells. Lastly, we demonstrated significant increases in expression of caspases (in particular caspase 8) and of p53 in a culture of Jurkat T cells. We conclude that the immunosuppressive actions of the compound in allograft rejection may be predominantly associated with induction of cell apoptosis and inhibition of TNF α production. The apoptosis could be predominantly selective for the CD3-CD8+CD11b + cell phenotype.

CD8+CD122+PD-1+ Tregs Synergize With Costimulatory Blockade of CD40/CD154, but Not B7/CD28, to Prolong Murine Allograft Survival.

A transplanted organ is always rejected in the absence of any immunosuppressive treatment due to vigorous alloimmunity. However, continuously global immunosuppression with a conventional immunosuppressant may result in severe side effects, including nephrotoxicity, tumors and infections. Tregs have been widely used to inhibit allograft rejection, especially in animal models. However, it's well accepted that administration of Tregs alone is not satisfactory in immune-competent wild-type animals. Therefore, it's imperative to promote Treg therapies under the cover of other approaches, including costimulatory blockade. In the present study, we demonstrated that administration of in vitro-expanded CD8+CD122+PD-1+ Tregs synergized with costimulatory blockade of CD40/CD154, but not B7/CD28, to prolong skin allograft survival in wild-type mice and to reduce cellular infiltration in skin allografts as well. Treg treatment and blockade of CD40/CD154, but not B7/CD28, also exhibited an additive effect on suppression of T cell proliferation in vitro and pro-inflammatory cytokine expression in skin allografts. Importantly, blocking B7/CD28, but not CD40/CD154, costimulation decreased the number of transferred CD8+CD122+PD-1+ Tregs and their expression of IL-10 in recipient mice. Furthermore, it's B7/CD28, but not CD40/CD154, costimulatory blockade that dramatically reduced IL-10 production by CD8+CD122+PD-1+ Tregs in vitro, suggesting that B7/CD28, but not CD40/CD154, costimulation is critical for their production of IL-10. Indeed, infusion of IL-10-deficient CD8+CD122+PD-1+ Tregs failed to synergize with anti-CD154 Ab treatment to further prolong allograft survival. Our data may explain why blocking B7/CD28 costimulatory pathway does not boost IL-10-dependent Treg suppression of alloimmunity. Thus, these findings could be implicated in clinical organ transplantation.

Hyperlipidemia and Allograft Rejection.

Advances in the development of immunosuppressive drug regimens have led to impressive survival rates in the year following organ transplantation. However rates of long-term graft dysfunction remain undesirably high. Recently it has been shown that co-morbidities in the patient population may affect graft survival. In mouse models, hyperlipidemia, a co-morbidity present in the majority of cardiac transplant patients, can significantly alter T cell responses to cardiac and skin allografts, and accelerate graft rejection. Here we review recent advances in our understanding of how alterations in lipids affect immune function and graft survival. Recent work in humans has highlighted the importance of controlling low density lipoprotein (LDL) levels in transplant recipients to reduce the development of chronic allograft vasculopathy (CAV). High serum levels of cholesterol containing particles leads to extensive immune system changes to T cell proliferation, differentiation and suppression. Changes in B cell subsets, and the ability of antigen presenting cells to stimulate T cells in hyperlipidemic animals may also contribute to increased organ allograft rejection. Cholesterol metabolism is a critical cellular pathway for proper control of immune cell homeostasis and activation. Increasing evidence in both human, and in mouse models shows that elevated levels of serum cholesterol can have profound impact on the immune system. Hyperlipidemia has been shown to increase T cell activation, alter the development of T helper subsets, increase the inflammatory capacity of antigen presenting cells (APC) and significantly accelerate graft rejection in several models.

Systematic testing and specificity mapping of alloantigen-specific chimeric antigen receptors in T regulatory cells

Chimeric antigen receptor (CAR) technology can be used to engineer the antigen specificity of regulatory T cells (Tregs) and improve their potency as an adoptive cell therapy in multiple disease models. As synthetic receptors, CARs carry the risk of immunogenicity, particularly when derived from nonhuman antibodies. Using an HLA-A*02:01-specific CAR (A2-CAR) encoding a single-chain variable fragment (Fv) derived from a mouse antibody, we developed a panel of 20 humanized A2-CARs (hA2-CARs). Systematic testing demonstrated variations in expression, and ability to bind HLA-A*02:01 and stimulate human Treg suppression in vitro. In addition, we developed a new method to comprehensively map the alloantigen specificity of CARs, revealing that humanization reduced HLA-A cross-reactivity. In vivo bioluminescence imaging showed rapid trafficking and persistence of hA2-CAR Tregs in A2-expressing allografts, with eventual migration to draining lymph nodes. Adoptive transfer of hA2-CAR Tregs suppressed HLA-A2+ cell-mediated xenogeneic graft-versus-host disease and diminished rejection of human HLA-A2+ skin allografts. These data provide a platform for systematic development and specificity testing of humanized alloantigen-specific CARs that can be used to engineer specificity and homing of therapeutic Tregs.

Hematopoietic chimerism and donor-specific skin allograft tolerance after non-genotoxic CD117 antibody-drug-conjugate conditioning in MHC-mismatched allotransplantation

Hematopoietic chimerism after allogeneic bone marrow transplantation may establish a state of donor antigen-specific tolerance. However, current allotransplantation protocols involve genotoxic conditioning which has harmful side-effects and predisposes to infection and cancer. Here we describe a non-genotoxic conditioning protocol for fully MHC-mismatched bone marrow allotransplantation in mice involving transient immunosuppression and selective depletion of recipient hematopoietic stem cells with a CD117-antibody-drug-conjugate (ADC). This protocol resulted in multilineage, high level (up to 50%), durable, donor-derived hematopoietic chimerism after transplantation of 20 million total bone marrow cells, compared with ≤ 2.1% hematopoietic chimerism from 50 million total bone marrow cells without conditioning. Moreover, long-term survival of bone marrow donor-type but not third party skin allografts is achieved in CD117-ADC-conditioned chimeric mice without chronic immunosuppression. The only observed adverse event is transient elevation of liver enzymes in the first week after conditioning. These results provide proof-of-principle for CD117-ADC as a non−genotoxic, highly-targeted conditioning agent in allotransplantation and tolerance protocols.

Deantigenicity of skin homografts in management of major burn cases

Objective The aim was to assess the ability of deantigenicity of skin homograft to decrease its rejection and improve its survival in management of patients with major burns. Background Skin homograft is considered as a good skin substitute in major burn cases. The major problem of the skin homograft is rejection. So, many methods are used to decrease antigenicity of the skin homograft and prolong its survival. Patients and methods This clinical trial included 32 patients with burn who were managed in Plastic Surgery Department, Menoufia University Hospital, from October 2015 to October 2017. Four groups were formed: control group had nine patients without deantigenicity of the homograft; group A had seven patients with freezing of the skin homograft at 4°C; group B had eight patients with skin homograft applied for irradiation; and group C had eight patients, with the recipient patient receiving dexamethasone injection. After escharotomy and coverage with skin homograft, follow-up was done for rejection of the homograft clinically. Results In the control group, the mean percentage of rejection of homograft was 62.22 ± 12.01%. In group A, the mean percentage of rejection was 61.4 ± 10.6%, with P value of 0.9. In group B, the mean percentage of rejection was 21.8 ± 10.69%, with P value of less than 0.001. In group C, the mean percentage of rejection was 48.7 ± 15.07%, with P value of 0.036. Conclusion Local irradiation dose to the skin homograft can minimize its rejection. Systemic corticosteroids produce less effect regarding the rejection of skin homograft when used with the usual dose, but with increasing dose, the rejection decreased. Freezing of the skin homograft in 4°C has no significant effect on the rejection of the skin homograft.

Ablation of interferon regulatory factor 4 in T cells induces "memory" of transplant tolerance that is irreversible by immune checkpoint blockade.

Achieving transplant tolerance remains the ultimate goal in the field of organ transplantation. We demonstrated previously that ablation of the transcription factor interferon regulatory factor 4 (IRF4) in T cells induced heart transplant acceptance by driving allogeneic CD4+ T cell dysfunction. Herein, we showed that heart-transplanted mice with T cell-specific IRF4 deletion were tolerant to donor-specific antigens and accepted the subsequently transplanted donor-type but not third-party skin allografts. Moreover, despite the rejection of the primary heart grafts in T cell-specific Irf4 knockout mice under immune checkpoint blockade, the establishment of donor-specific tolerance in these mice was unhindered. By tracking alloantigen-specific CD4+ T cells in vivo, we revealed that checkpoint blockade restored the expression levels of the majority of wild-type T cell-expressed genes in Irf4-deficient T cells on day 6 post-heart grafting, indicating the initial reinvigoration of Irf4-deficient T cells. Nevertheless, checkpoint blockade did not restore cell frequency, effector memory cell generation, and IFN-γ/TNF-α production of Irf4-/- alloreactive T cells at day 30 post-heart grafting. Hence, targeting IRF4 represents a potential therapeutic strategy for driving intrinsic T cell dysfunction and achieving alloantigen-specific transplant tolerance.

The impact of skin allograft on inpatient outcomes in the treatment of major burns 20–50% total body surface area — A propensity score matched analysis using the nationwide inpatient sample

BackgroundHuman cadaveric skin (allograft) is used in treating major burns both as temporizing wound coverage and a means of testing wound bed viability following burn excision. There is limited information on outcomes, and clinicians disagree on indications for application in intermediate-sized burns. This study aims to improve understanding of allograft use in 20–50% total body surface burns by assessing current utilization and evaluating inpatient outcomes. MethodsDischarge data from the Nationwide Inpatient Sample (NIS), Healthcare Cost and Utilization Project (HCUP), Agency for Healthcare Research and Quality assessed 3557 major burn patients (>second degree depth and 20–50% TBSA) undergoing operative treatment. Outcomes were evaluated with propensity score matching. The primary outcome was mortality with secondary outcomes including complications, length of stay, total burn operations, and charges. ResultsAfter matching, 771 allografted patients were paired with 1774 controls. Covariate mean standard differences were all <11% after matching. The average treatment effect (ATE) of allograft on inpatient mortality was an increase of 2.8% (95% CI 0.2–5.3%, p=0.041). Allograft ATEs were all significantly higher for secondary outcomes: composite complication index increased 0.13 (95% CI 0.07–0.20, p<0.001), length of stay 8.4days (95% CI 6.1–1.9 days, p<0.001), total burn operations 1.6 (95% CI 1.4–1.9, p<0.001), and total charges $139,476 [$100,716–178,236, p<0.001). ConclusionsAllograft use in major burns 20–50% TBSA was associated with a significant increase in inpatient mortality. There was a notable correlation with increased inpatient complications, longer length of stay, more burn operations, and greater total charges. Better studies are needed to justify the use of this costly and limited resource in the intermediate sized major burn population.

The features and treatment of Xixia "May 17th" explosion accident

The features and treatment of Xixia "May 17th" explosion accident

Mast Cell Degranulation Exacerbates Skin Rejection by Enhancing Neutrophil Recruitment.

Recent evidences indicate an important role of tissue inflammatory responses by innate immune cells in allograft acceptance and survival. Here we investigated the role of mast cells (MC) in an acute male to female skin allograft rejection model using red MC and basophil (RMB) mice enabling conditional MC depletion. Kinetic analysis showed that MCs markedly accelerate skin rejection. They induced an early inflammatory response through degranulation and boosted local synthesis of KC, MIP-2, and TNF. This enhanced early neutrophil infiltration compared to a female-female graft-associated repair response. The uncontrolled neutrophil influx accelerated rejection as antibody-mediated depletion of neutrophils delayed skin rejection. Administration of cromolyn, a MC stabilizer and to a lesser extent ketotifen, a histamine type I receptor antagonist, and absence of MCPT4 chymase also delayed graft rejection. Together our data indicate that mediators contained in secretory granules of MC promote an inflammatory response with enhanced neutrophil infiltration that accelerate graft rejection.

A STUDY OF USING ABORTED BOVINE FETAL SKIN AND SKIN ALLOGRAFT FOR REPAIRING CRITICAL SKIN DEFECT IN DOGS

The present study was designed to compare the potential repair efficacy of using either skin xenograft or allograft on the treatment of induced skin defects. The study was conducted on twelve stray dogs. Their weight and age ranges were 12-18 kg and 10-15 months, respectively. Animals were randomly allocated into two equal groups. In the first group, a 10 cm of full-thickness skin piece was harvested aseptically from a 4-month-aborted fetus. Along with, in the experimental animals, comparable skin defects were created at the lateral aspect of the forelimb. The grafts were fixed firmly to the wound bed by suturing. In the second group, the skin grafts (10 cm) were harvested from the lower abdomen of dogs and were firmly fixed over induced wounded beds at the forelimb of other dogs. In the first group, a few days postoperatively, the operative site demonstrated dryness, scar tissue formations, delayed wound healing, and local inflammation. However, after one-month post-surgery, there was a separation of the grafted tissue from the wound bed and signs of tissue rejection. In first group, histopathological examination revealed extensive infiltration of the inflammatory cells and separation of the grafted tissue from the subdermal layer. In the second group, the repaired site exhibited inflammatory signs and skin ulcers during the first postoperative week. After one month of the operation, formations of granulation tissue and restoration of normal skin coloration without hair formation were revealed on the allograft site. Complete skin healing and union of the graft with the recipient host were determined after two months of surgery. Histopathological examination demonstrated fibrocystic hyperplasia of the grafted tissue one month post-operatively, while deposition of collagen fibers was the main feature at the second month. In conclusion, unlike using of skin from the aborted bovine fetuses, the study specified the successful outcomes of using skin allograft in tissue repairing in dogs.

Bacterial contamination of human skin allografts and antimicrobial resistance: a skin bank problem

BackgroundBacterial contamination remains the major problem in skin banks, even after antimicrobial treatment, and results in high rates of tissue discarding. This study aimed to analyze bacterial contamination in 32 human skin allografts from the skin bank of Dr. Roberto Corrêa Chem from the Hospital Complex Santa Casa de Misericórdia de Porto Alegre. These samples were already discarded due to microbial contamination. The identification of the bacteria isolated from skin allografts was performed by matrix assisted laser desorption ionization–time of flight. The antimicrobial susceptibility of the isolates to six different classes of antimicrobials was determined using the disk-diffusion agar method, and the evaluation of the inhibitory potential was determined by the minimal inhibitory concentration (50/90) of antimicrobials already used in the skin bank and those that most isolates were susceptible to.ResultsA total of 21 (65.6%) skin samples were contaminated with Gram-positive bacteria: 1 (4.7%) with Paenibacillus sp., 12 (61.9%) with Bacillus sp., 6 (28.5%) with Staphylococcus sp., and 2 (9.5%) with Bacillus sp. and Staphylococcus sp. Several resistance profiles, including multiresistance, were found among the isolates. Most of the isolates were susceptible to at least one of the antimicrobials used in the skin bank. All isolates were susceptible to amikacin, gentamicin, and tetracycline, which demonstrated the best inhibitory activities against the isolates and were considered as potential candidates for new antimicrobial treatments.ConclusionsBacillus, Paenibacillus, and Staphylococcus were isolated from the skin allografts, thus demonstrating the predominance of Gram-positive bacteria contamination. Other factors not related to the resistance phenotype may also be involved in the persistence of bacterial isolates in the skin allografts after antibiotic treatment. Gentamicin, amikacin, and tetracycline can be considered as an option for a more effective treatment cocktail.

C5aR1 regulates migration of suppressive myeloid cells required for costimulatory blockade-induced murine allograft survival.

Costimulatory blockade-induced murine cardiac allograft survival requires intragraft accumulation of CD11b+ Ly6Clo Ly6G- regulatory myeloid cells (Mregs) that expand regulatory T cells (Tregs) and suppress effector T cells (Teffs). We previously showed that C5a receptor (C5aR1) signaling on T cells activates Teffs and inhibits Tregs, but whether and/or how C5aR1 affects Mregs required for transplant survival is unknown. Although BALB/c hearts survived >60days in anti-CD154 (MR1)-treated or cytotoxic T-lymphocyte associated protein 4 (CTLA4)-Ig-treated wild-type (WT) recipients, they were rejected at ~30days in MR1-treated or CTLA4-Ig-treated recipients selectively deficient in C5aR1 restricted to myeloid cells (C5ar1fl/fl xLysM-Cre). This accelerated rejection was associated with ~2-fold more donor-reactive T cells and ~40% less expansion of donor-reactive Tregs. Analysis of graft-infiltrating mononuclear cells on posttransplant day 6 revealed fewer Ly6Clo monocytes in C5ar1fl/fl xLysM-Cre recipients. Expression profiling of intragraft Ly6Clo monocytes showed that C5aR1 deficiency downregulated genes related to migration/locomotion without changes in genes associated with suppressive function. Cotransfer of C5ar1fl/fl and C5ar1fl/fl xLysM-Cre myeloid cells into MR1-treated allograft recipients resulted in less accumulation of C5ar1-/- cells within the allografts, and in vitro assays confirmed that Ly6Chi myeloid cells migrate to C5a/C5aR1-initiated signals. Together, our results newly link myeloid cell-expressed C5aR1 to intragraft accumulation of myeloid cells required for prolongation of heart transplant survival induced by costimulatory blockade.

Skin Grafting on 3D Bioprinted Cartilage Constructs In Vivo.

Background:Three-dimensional (3D) bioprinting of cartilage is a promising new technique. To produce, for example, an auricle with good shape, the printed cartilage needs to be covered with skin that can grow on the surface of the construct. Our primary question was to analyze if an integrated 3D bioprinted cartilage structure is a tissue that can serve as a bed for a full-thickness skin graft.Methods:3D bioprinted constructs (10 × 10 × 1.2 mm) were printed using nanofibrillated cellulose/alginate bioink mixed with mesenchymal stem cells and adult chondrocytes and implanted subcutaneously in 21 nude mice.Results:After 45 days, a full-thickness skin allograft was transplanted onto the constructs and the grafted construct again enclosed subcutaneously. Group 1 was sacrificed on day 60, whereas group 2, instead, had their skin-bearing construct uncovered on day 60 and were sacrificed on day 75 and the explants were analyzed morphologically. The skin transplants integrated well with the 3D bioprinted constructs. A tight connection between the fibrous, vascularized capsule surrounding the 3D bioprinted constructs and the skin graft were observed. The skin grafts survived the uncovering and exposure to the environment.Conclusions:A 3D bioprinted cartilage that has been allowed to integrate in vivo is a sufficient base for a full-thickness skin graft. This finding accentuates the clinical potential of 3D bioprinting for reconstructive purposes.

Regulation of human T cell responses by dNP2-ctCTLA-4 inhibits human skin and microvessel graft rejection

Manipulation of human T cell functioning by delivery of macromolecules such as DNA, RNA, or protein is limited, unless the human T cells have been stimulated or electropermeabilized. To achieve successful adaptation and survival of a grafted organ, the alloreactive T cells that induce graft rejection must be regulated. Corticosteroids, calcineurin inhibitors, and mTOR inhibitors, which are systemic immunosuppressants, are currently used for transplantation, with significant side effects. In this study, we demonstrated that a cell-permeable peptide (CPP), dNP2, could efficiently deliver proteins into human CD4 and CD8 T cells. We confirmed regulatory functioning of the cytoplasmic domain of CTLA-4 conjugated with dNP2 (dNP2-ctCTLA-4) in human T cell activation, proliferation, and chemokine receptor expression. We utilized a human skin allograft system in SCID/beige mice to examine whether dNP2-ctCTLA-4 could inhibit allograft rejection by controlling T cell responses. The grafted skin tissue inflammation, allogeneic T cell infiltration, and blood cytokine level was markedly reduced by dNP2-ctCTLA-4, resulting in successful transplantation. In addition, it also inhibited T cell alloresponses against microvessels formed form Bcl-2-transduced human umbilical vein endothelial cells implanted into Balb/c Rag1−/−/IL-2Rγ−/- double knockout (DKO) mice, assessed as reduced T cell infiltration and granzyme B expression. These results collectively suggest that dNP2 peptide conjugation offers a valuable tool for delivering macromolecules like proteins into human T cells, and dNP2-ctCTLA-4 is a novel agent that shows potential in controlling human T cell responses to allow successful adaptation of grafted tissues.

Heme oxygenase-2 suppresses acute inflammation and improves the survival of skin allografts

The heme oxygenase (HO) system is an important regulatory arm of the intrinsic cytoprotective and anti-inflammatory system. HO-2 plays an important role in regulating inflammation following injury. The aim of this study was to evaluate the effect of HO-2 overexpression on inflammatory responses. A skin transplantation model, involving the application of skin grafts from wild-type or HO-2 overexpressing mice to BALB/c mice, was used for investigation. HO-2 overexpression suppressed the production of pro-inflammatory cytokines (IL-1β, TNF-α, and IL-6) in macrophages compared to that in macrophages obtained from control mice. HO-2 overexpression also significantly prolonged the survival of allografted skin. Our findings suggest that HO-2 attenuates inflammatory responses and effectively prolongs skin graft survival.

Cadaver skin allograft may improve mortality rate for burns involving over 30% of total body surface area: a propensity score analysis of data from four burn centers.

Cadaver skin is used for temporary wound covering, but there is insufficient evidence regarding its clinical usefulness in patients with major burns. We aimed to analyze the effect of cadaveric skin allograft on mortality rates in patients with burns involving > 30% of total body surface area (TBSA). Our study included 1282 patients with > 30% of TBSA burned admitted to four hospitals in Korea between June 1, 2008 and December 31, 2016. Of these, 698 patients underwent cadaver skin allograft (cadaver group), and 584 were treated with conventional treatment (non-cadaver group). We corrected the differences between the two groups using propensity score matching, and generated 474 propensity score-matched pairs. Overall 90-day in-hospital mortality rate among all patients was 35.3% (453/1282). There was a significant difference in 90-day in-hospital mortality between the two groups for both unmatched [cadaver vs. conventional, 31.7 vs. 39.7%; difference, 8.0; 95% confidence interval (CI) 2.8-13.3] and propensity-matched groups (37.8 vs. 47.3%; difference, 9.5; 95% CI 3.2-15.8). Logistic regression analyses showed a significant association between cadaver skin allograft and lower 90-day in-hospital mortality in the propensity-matched groups (odds ratio, 0.42; 95% CI 0.29-0.62). Patients with major burns who underwent cadaver skin allograft had a lower mortality rate compared to those who did not. Cadaver skin allograft may improve the survival of patients with major burns, especially in the early phase of injury.

Mouse Model Established by Early Renal Transplantation After Skin Allograft Sensitization Mimics Clinical Antibody-Mediated Rejection.

Antibody-mediated rejection (AMR) is the main barrier to renal graft survival, and mouse renal AMR models are important to study this process. Current mouse models are established by priming the recipient to donor skin for over 7 days before kidney transplantation. The robustness of AMR in these cases is too strong to mimic clinical AMR and it is unclear why altering the priming times ranging from 7 to 91 days fails to reduce the AMR potency in these models. In the present study, we found that the donor-recipient combination and skin graft size were determinants of donor-specific antibody (DSA) development patterns after skin transplantation. DSA-IgG was sustained for over 100 days after skin challenge, accounting for an identical AMR robustness upon different skin priming times over 7 days. However, decreasing the skin priming time within 7 days attenuated the robustness of subsequent renal allograft AMR in C3H to Balb/c mice. Four-day skin priming guaranteed that recipients develop acute renal AMR mixed with a high ratio of graft-infiltrating macrophages, renal grafts survived for a mean of 6.4 ± 2.1 days, characterized by typical AMR histological changes, such as glomerulitis, peritubular capillary (PTC) dilation, and capillaritis, deposition of IgG and C3d in PTCs, but less prevalence of microthrombus, whereas the cellular rejection histological change of tubulitis was absent to mild. With this scheme, we also found that the renal AMR model can be developed using common mouse strains such as C57BL/6 and Balb/c, with mean prolonged renal graft survival times of 14.4 ± 5.0 days. Finally, we proved that donor-matched skin challenge after kidney transplantation did not strongly affect DSA development and kidney graft outcome. These findings may facilitate an understanding and establishment of mouse renal allograft AMR models and promote AMR-associated studies.

NK Cells Regulate CD8 T Cell-Mediated Allogeneic Rejection in Immunocompetent Recipients across an MHC Class I Mismatched Barrier

Non-T natural killer (NK) cells are a population of granular lymphoid cells TcR- CD3− CD122+ (NK1.1+ in C57BL/6 mice and DX5+ in Balb/c mice). These innate lymphoid cells mature in the bone marrow and are responsible for killing viral-infected and transformed cells in a non-MHC restricted manner, without the need of prior sensitization due to a recognition mechanism based on the array of germline-encoded inhibitory and activating receptors expressed on the cell surface of NK cells [1]. NK cells modulate CD8 T cell expansion during homeostatic proliferation in response to lymphopenic conditions by killing activated T cells and competing for IL-15 with the proliferating CD8 effector T cells [2]. This NK cell behavior may respond to an adaptive evolutionary mechanism to prevent aggressive anti-viral immune responses during the acute phase of infection and thus attenuate immunopathology [3]. We set up an experimental mouse model of alloreactivity across an MHC class I barrier to evaluate the role of NK cells in CD8 T cell-mediated rejection of bm1 skin allografts in naïve B6 recipients. In this transplant setting, host CD8 T cells are the main effectors T cells involved in the rejection of class I MHC mismatched skin allografts preferentially through the direct pathway of antigen presentation with limited help from CD4 T cells primed by the indirect pathway of antigen presentation. As opposed to previous studies done in lymphopenic immunodeficient recipients [2], depletion of NK cells was performed in immunocompetent C57BL/6 recipients with or without CD8 T depletion, the former displaying homeostatic proliferation in response to lymphopenia due to antibody-mediated elimination of CD8 T cells. We observed that depletion of NK cells in both CD8-repleted and CD8-depleted naïve C57BL/6 immunocompetent mice accelerated significantly bm1 graft rejection compared to non-NK-depleted controls (p<0.0005) (Figure 1). Our data claims for precaution in solid organ transplantation under inducing protocols of extensive depletion of leukocytes, such as ATG or alentuzumab. The immunotherapeutic interventions that include the removal of NK cells may accelerate graft rejection and promote aggressive CD8 T cell cytotoxic responses, which could be refractory to current immunosuppression. It also suggests that enhancing NK-cell functional activity combined with CD8 T cell depletion under tolerogenic regimen that tolerizes CD4 T cells at the time of transplantation may contribute to allogeneic donor-specific tolerance induction by elimination of the direct pathway of antigen presentation and by regulating the recovery of CD8 T cells.Bibliographic

Skin Allografting Activates Anti-tumor Immunity and Suppresses Growth of Colon Cancer in Mice

INTRODUCTION: The tumor cells could escape from the immune elimination through the immunoediting mechanisms including the generation of immunosuppressive or immunoregulative cells. By contrast, allograft transplantation could activate the immune system and induce a strong allogenic response. The aim of this study was to investigate the efficacy of allogenic skin transplantation in the inhibition of tumor growth through the activation of allogenic immune response. METHODS: Full-thickness skin transplantation was performed from C57BL/6 (H-2b) donors to BALB/c (H-2d) recipients that were receiving subcutaneous injection of isogenic CT26 colon cancer cells (2 × 106 cells) at the same time. The tumor size and pathological changes, cell populations and cytokine profiles were evaluated at day 14 post-transplantation. RESULTS: The results showed that as compared to non-transplant group, the allogenic immune response in the skin-grafting group inhibited the growth of tumors, which was significantly associated with increased numbers of intra-tumor infiltrating lymphocytes, increased populations of CD11c+MHC-classII+CD86+ DCs, CD3+CD4+ T cells, CD3+CD8+ T cells, and CD19+ B cells, as well as decreased percentage of CD4+CD25+Foxp3+ T cells in the spleens. In addition, the levels of serum IgM and IgG, tumor necrosis factor (TNF)-α and interferon (IFN)-γ were significantly higher within the tumor in skin transplant groups than that in non-transplant group. CONCLUSIONS: Allogenic skin transplantation suppresses the tumor growth through activating the allogenic immune response, and it may provide a new immunotherapy option for the clinical refractory tumor treatment.