Vascularized Skin Allografts Research Articles

The Positive Impact of Donor Bone Marrow Cells Transplantation into Immunoprivileged Compartments on the Survival of Vascularized Skin Allografts

Using the vascularized skin allograft (VSA) model, we compared the tolerogenic effects of different allogeneic bone marrow transplantation (BMT) delivery routes into immunoprivileged compartments under a 7-day protocol immunosuppressive therapy. Twenty-eight fully MHC mismatched VSA transplants were performed between ACI (RT1a) donors and Lewis (RT11) recipients in four groups of seven animals each, under a 7-day protocol of alfa/beta TCRmAb/CsA (alpha/beta-TCR monoclonal antibodies/Cyclosporine A therapy). Donor bone marrow cells (BMC) (100 × 106 cells) were injected into three different immunoprivileged compartments: Group 1: Control, without cellular supportive therapy, Group 2: Intracapsular BMT, Group 3: Intragonadal BMT, Group 4: Intrathecal BMT. In Group 2, BMC were transplanted under the kidney capsule. In Group 3, BMC were transplanted into the right testis between tunica albuginea and seminiferous tubules, and in Group 4, cells were injected intrathecally. The assessment included: skin evaluation for signs and grade of rejection and immunohistochemistry for donor cells engraftment into host lymphoid compartments. Donor-specific chimerism for MHC class I (RT1a) antigens and the presence of CD4+/CD25+ T cells were assessed in the peripheral blood of recipients. The most extended allograft survival, 50–78 days, was observed in Group 4 after intrathecal BMT. The T cells CD4+/CD25+ in the peripheral blood were higher after intrathecal BMC injection than other experimental groups at each post-transplant time point. Transplantation of BMC into immunoprivileged compartments delayed rejection of fully mismatched VSA and induction of robust, donor-specific chimerism.

Inhibition of Lymphatic Drainage With a Self-Designed Surgical Approach Prolongs the Vascularized Skin Allograft Survival in Rats.

Vascularized composite allotransplantation (VCA) is an emerging treatment for significant tissue defects. However, VCAs usually consist of multiple highly antigenic skin tissues. Previous studies have shown that the lymphatic system in skin plays important roles in the initiation of immune responses during acute rejection, by transporting T cells and antigen-presenting dendritic cells to regional lymph nodes. Therefore, we designed a new surgical treatment to inhibit lymphatic drainage of skin allografts and investigated whether this approach could promote the survival of allografts and suppress immunological events after transplantation. This procedure was achieved by connecting the vascularized allografts to recipient tissues with only an annular plastic holder, allowing the minimum of allograft contact with recipients. Our results showed that the self-designed treatment for inhibiting lymphatic drainage promoted the survival of allografts, reduced the serum concentration of IL-2, and decreased the percentage of CD4CD25 and CD8CD25 from the lymphatic nodes draining the transplantation region. In conclusion, these data suggest that self-designed surgical approach is effective in inhibiting lymphatic drainage of skin allografts, and the lymphatic system may be new therapeutic targets for developing techniques or drugs against acute rejection after VCAs.

Transfusion of ethylene carbodiimide–fixed donor splenocytes prolongs survival of vascularized skin allografts

BackgroundAllograft rejection is a major obstacle to the widespread clinical application of vascularized composite allotransplantation. Recent studies revealed a noncytoreductive strategy to protect allografts by the transfusion of ethylene carbodiimide–fixed donor splenocytes (ECDI-SPs). To determine whether this approach offers advantages in protecting skin allografts, we examined the immunological protection of infusing ECDI-SPs with a 30-d administration of rapamycin on the skin allografts of mice. Materials and methodsC57BL/6 recipient mice received BALB/c donor full-thickness skin or vascularized skin transplants at day 0, along with the infusion of donor ECDI-SPs 7 d before and 1 d after allotransplantation and a 30-d course of rapamycin. Recipients received ECDI-untreated splenocytes or C3H allografts as controls. In vitro allostimulatory activity of ECDI-SPs and donor-specific ex vivo hyporesponsiveness were tested. Production of related cytokines (TGF-β, IL-10, IL-1β, and TNF-α) and expression of CD4+Foxp3+ regulatory T cells (Tregs) were also examined. ResultsTransfusion of ECDI-SPs combined with rapamycin significantly prolonged survival of full-thickness skin (median survival time [MST]: 28 d) and full-thickness skin allografts (MST: 71 d) compared with untreated splenocytes (MSTs: 11 d and 30 d) or C3H allografts (MSTs: 11 d and 38 d). This effect was accompanied by increased production of IL-10 and TGF-β, decreased production of IL-1β and TNF-α, and expansion of Tregs in vitro and in vivo. ConclusionsECDI-SP infusion combined with short-term rapamycin administration provides a promising approach to prolong the skin allograft survival.

A Mouse Model of Vascularized Skin Transplantation.

Allogeneic skin tissues are highly antigenic and induce intensive immune rejection after composite tissue allotransplantation. Mouse models have advantages in mechanistic studies of immune rejection. However, due to technical challenge in vascular anastomosis with suture technique, mouse vascularized skin allotransplantation models are not widely used in studies of immune rejection. Therefore, the authors propose vascular anastomosis through cuff technique during allotransplantation of mouse donor free groin skin flaps to either recipient inguinal or cervical site. Free groin skin flaps from BALB/c or C57BL/6 donor mice were transplanted to the groin (inguinal vascularized skin transplantation [IVST]) or to the neck of recipient sites (cervical vascularized skin transplantation [CVST]) of C57BL/6 mice. A nonsuture cuff technique was utilized to anastomose the donor vessels with either femoral vessels in IVST recipients or common carotid arteries and external jugular veins in CVST mice. Immunosuppressant drugs were used in the allogeneic skin group. The overall success rate was higher in the CVST (88.5%) when compared with the IVST (78.9%). Total operation time in CVST mice lasted 96 minutes (95% confidence interval, 92-101 minutes) that was shorter than that for IVST mice (136 minutes, 95% confidence interval, 127-176 minutes; P < 0.01). Complications, such as hindlimb necrosis and self-mutilation, were observed in IVST mice. Rapamycin (3 mg/kg, daily) significantly prolonged vascularized skin allografts survival with median survival time of 80 days. All syngeneic grafts survived for more than 80 days. We developed a novel mouse vascularized skin transplantation model that is feasible for the study of clinically relevant skin rejection and tolerance.

Primary Vascularization of Allografts Governs Their Immunogenicity and Susceptibility to Tolerogenesis

We investigated the influence of allograft primary vascularization on alloimmunity, rejection, and tolerance in mice. First, we showed that fully allogeneic primarily vascularized and conventional skin transplants were rejected at the same pace. Remarkably, however, short-term treatment of mice with anti-CD40L Abs achieved long-term survival of vascularized skin and cardiac transplants but not conventional skin grafts. Nonvascularized skin transplants triggered vigorous direct and indirect proinflammatory type 1 T cell responses (IL-2 and IFN-γ), whereas primarily vascularized skin allografts failed to trigger a significant indirect alloresponse. A similar lack of indirect alloreactivity was also observed after placement of different vascularized organ transplants, including hearts and kidneys, whereas hearts placed under the skin (nonvascularized) triggered potent indirect alloresponses. Altogether, these results suggest that primary vascularization of allografts is associated with a lack of indirect T cell alloreactivity. Finally, we show that long-term survival of vascularized skin allografts induced by anti-CD40L Abs was associated with a combined lack of indirect alloresponse and a shift of the direct alloresponse toward a type 2 cytokine (IL-4, IL-10)-secretion pattern but no activation/expansion of Foxp3(+) regulatory T cells. Therefore, primary vascularization of allografts governs their immunogenicity and tolerogenicity.

Correlation of Chimerism With Graft Size and Revascularization in Vascularized and Nonvascularized Skin Allografts

The high antigenicity of skin components of composite tissue allografts is the most challenging factor in achieving long-term viability after composite tissue allografts transplantation. The vascularization pattern of vascularized skin allografts (VSA) and nonvascularized skin allografts (NVSA) differs significantly and these differences may alter host immune responses. We hypothesized that vascularized grafts contribute to better engraftment and long-term survival. We have tested our hypothesis by transplantation of different sizes (2 x 2 cm, 4 x 4 cm, 6 x 6 cm) of VSA and NVSA across major histocompatibility complex barrier between LBN (RT1(1+n)) and Lewis (RT1(1)) rats. Correlation of revascularization process with development of donor-specific chimerism was tested. We found that the highest chimerism levels (8%-12.2% in VSA groups; 2.53%-3.92% in NVSA groups) were reached as early as 7 days in both VSA and NVSA. Chimerism decreased in both groups during 100-day follow-up and higher chimerism was found only in VSA in late posttransplant period. Revascularization, assessed by the presence of CD31 positive vessels, was significantly higher in VSA compared with NVSA and to controls (P < 0.05). A direct correlation was seen between increased skin diameter and donor chimerism in VSA, whereas inverse correlation was tested in NVSA. We have confirmed that allograft size and vascularization pattern contribute to donor chimerism development and maintenance.

Immunologic Responses in Vascularized and Nonvascularized Skin Allografts

The skin is a protective interface between internal organs and the environment; it is the largest tissue of the human body. However, the skin does not serve merely as a physical barrier. It is also an active immune organ, traversed by a network of lymphatic and blood vessels. This immunologic structure contains immunologic cells such as T lymphocytes, Langerhans cells (LCs), dendritic cells, and keratinocytes. Langerhans cells represent the cutaneous counterpart of dendritic cells. LCs not only act as professional antigen-presenting cells to induce antigen-specific T cells for adaptive immune responses, but they also initiate a cascade of innate immune responses by antigenic stimulus such as transplant tissue. In transplantation immunology, either donor or recipient LCs fulfill an important mission in rejection or acceptance of donor tissue. Vascularized or nonvascularized skin allografts may create an immunologic response through different pathways. In both transplant models, skin diameter may change antigenic load, thereby determining rejection or acceptance response. This article discusses the effects of the cellular component in the skin immune system on immunologic responses of vascularized or nonvascularized skin allografts and describes the differences between the two immunologic cascades.

Co-infusion of donor bone marrow with host mesenchymal stem cells treats GVHD and promotes vascularized skin allograft survival in rats

We investigated the effect of autologous mesenchymal stem cells (MSC) on multiple unmodified donor bone marrow (BM) infusions and vascularized skin graft outcome. BM-derived rat MSC were examined for phenotype and function. MSC/MSC-conditioned-medium suppressed IFN-γ production by T cells and modified DC function. Infusions of MSC with one-time BM improved vascularized skin graft survival, while with one–two-times BM reversed graft versus host disease (GVHD). Mixed chimerism was enhanced in recipients given two–four-times BM with MSC infusions. Interestingly, four-times BM infusions with MSC delayed GVHD onset, reduced host tissue damage and enhanced vascularized skin allograft survival compared to four-times BM alone. These data demonstrate that, the co-infusion of MSC with unmodified BM limit the toxicity of allogeneic BM transplantation, enhance mixed chimerism and improve vascularized skin graft survival. These findings provide insights for the development of autologous MSC-based BM transplantation and prevention of graft rejection or treatment of autoimmunity.

Rapamycin-conditioned, alloantigen-pulsed dendritic cells promote indefinite survival of vascularized skin allografts in association with T regulatory cell expansion

Clinically-applicable protocols that promote tolerance to vascularized skin grafts may contribute to more widespread use of composite tissue transplantation. We compared the properties of alloantigen (Ag)-pulsed, rapamycin (Rapa)-conditioned and control bone marrow-derived host myeloid dendritic cells (DCs) and their potential, together with transient immunosuppression (anti-lymphocyte serum+cyclosporine), to promote long-term, vascularized skin graft survival in Lewis rats across a full MHC barrier. Both types of DCs expressed low levels of CD86, but Rapa DC expressed lower levels of MHC II and CD40 and were less stimulatory in MLR. While both Rapa and control DCs produced low levels of IL-12p70 and moderate levels of IL-6 and IL-10 following TLR ligation, Rapa DC secreted significantly lower levels of IL-6 and IL-10 in response to LPS. Donor Ag-pulsed Rapa DC, but not control DC, induced long-term skin graft survival (median survival time >133 days) when administered 7 and 14 days post-transplant. Circulating T cells in hosts with long-surviving grafts were hyporesponsive to donor alloAg stimulation, but proliferated in response to third-party stimulation and produced IFN-γ and IL-10. When recipients of long-surviving grafts were challenged with skin grafts, donor but not third-party grafts were prolonged, suggesting underlying regulatory mechanisms. Both flow cytometry and immunohistochemical analysis revealed that donor Ag-pulsed Rapa DC infusion expanded CD4+ Foxp3+ Treg in recipients' spleens, graft-associated lymph nodes and the graft. These data demonstrate for the first time that pharmacologically-modified, donor Ag-pulsed host DC administered post-transplant can promote indefinite vascularized skin graft survival, associated with Treg expansion.

Prolonged Survival of Vascularized Skin Grafts Across a Full MHC Mismatch Using Donor Antigen-Pulsed Tolerogenic GM+Rapa Dendritic Cells (102.22)

Abstract We compared the tolerogeneic properties of BM-derived GM+IL-4 and GM+Rapa host DCs and their potential to induce long-term acceptance of vascularized skin allograft in rats across a full MHC mismatch with transient immunosuppression. Results show that both DCs were CD11b+ and expressed low levels of CD86 and produced low levels of IL-12p70 following TLR activation. GM+Rapa DC produced lower levels of anti- and pro-inflammatory cytokines in response to LPS. Both DCs had low T cell stimulatory capacity in vitro. Interestingly, donor Ag-pulsed GM+Rapa DC (GM+Rapa DCp) induced long-term survival of the vascularized skin grafts and this was significant compare to GM+IL-4 DC and control groups. PBMC from long-term graft survivors that were previously treated with donor Ag-pulsed GM+Rapa DC were hyporesponsive to donor Ag, but proliferated in response to the third-party Ag and produced IL-10 in addition to IFN-gamma. Recipients of the long-term surviving allografts were challenged with full-thickness tail skin grafts. While the third-party grafts were rejected fast the donor graft had delayed rejection, suggesting that this treatment induced a regulatory mechanism that prevented the acute rejection of donor skin grafts. Further experiments demonstrated presence of CD4+ Foxp3+ T cells in the vascularized skin and secondary lymphoid tissues. Taken together, these results demonstrate that the donor Ag-pulsed tolerogenic GM+Rapa DC have ability to induce regulatory T cells and prevent vascularized skin graft rejection across a full MHC mismatch.

Allogeneic bone marrow cells co-infused with stromal stem cells for induction of vascularized skin allograft survival

Allogeneic bone marrow cells co-infused with stromal stem cells for induction of vascularized skin allograft survival

Trafficking of Donor-Derived Bone Marrow Correlates With Chimerism and Extension of Composite Allograft Survival Across MHC Barrier

We proposed to evaluate differences between recipient’s immune response to vascularized skin and combined vascularized skin/bone allografts, under a 7-day αβ-TCR plus cyclosporine (CsA) treatment protocol. Thirty-six transplantations were performed in six groups: group I (isograft control-vascularized skin graft; n = 6); group II (isograft control-combined vascularized skin/bone graft; n = 6); group III (allograft rejection control group-vascularized skin graft; n = 6); group IV (allograft rejection control-combined vascularized skin/bone graft; n = 6); group V (allograft treatment-vascularized skin graft; n = 6); and group VI (allograft treatment-combined vascularized skin/bone graft; n = 6). Isograft transplantations were performed between Lewis rats and allografts were transplanted across the MHC barrier from Brown Norway to Lewis rats. In the allograft treatment group, a combined αβ-TCR+CsA protocol was applied for 7 days. All groups were compared clinically, immunologically and histologically. Statistical significance was determined with two-tailed Student’s t test. Indefinite graft survival was achieved in the isograft control group (>300 days). Allograft rejection controls rejected within 5 to 9 days posttransplant; chimerism levels were undetectable (<.5%). Allografts under the αβ-TCR+CsA protocol had significantly extended survival when skin was combined with bone (61–125 days) compared to vascularized skin allografts (43–61 days). Lymphoid macrochimerism was significantly higher in group VI than group V. Histology confirmed skin and bone viability. Combined vascularized skin/bone allografts had higher and sustained levels of donor-specific chimerism and extended allograft survival.

Studies of the immediately vascularized skin allograft.

In order to evaluate the effect of the nature of revascularization on survival rates of skin allografts, a rat model was devised which provided simultaneous and immediate establishment of arterial and venous blood supply to a composite skin-renal allograft. For 5 days after transplantation the skin portion of the composite grafts appeared normal, with minimal inflammation in 11 recipients. From 6 to 11 days, in 6 rats, active rejection developed with a marked inflammatory reaction of the skin-kidney interface. In 10 rats in which the composite grafts remained in the secondary hosts for 12--21 days rejection of the skin was complete. The renal portion of all composite grafts appeared normal in all three panels and all recipients of composite grafts rejected subsequent orthotopic skin grafts in an accelerated manner, with median survival time of 8.2 +/- 0.3 days compared with 11.5 +/- 0.7 days in untreated Fisher leads to Lewis controls. These results demonstrate that the skin on the composite graft retained its immunogenicity, that immediately vascularized skin allografts between AgB compatible rats will be rejected in a normal time and that anatomical factors are not sufficient to account for the difference in survival times between skin and solid organs.

Strategies to develop chimerism in vascularized skin allografts across MHC barrier

In this study, we investigated the effects of 7-day-protocols of alphabeta-T-cell receptor monoclonal antibody (alphabeta-TCRmAb), cyclosporine A (CsA), and tacrolimus (FK-506) immunosuppressive monotherapies, and their combinations on the survival of vascularized skin allografts (VSA). Forty-two transplantations of VSA across a strong MHC barrier were performed between ACI (RT1a) donors and Lewis (RT1(l)) recipients in seven groups. Isograft and allograft rejection controls received no treatment. Treatment groups received a 7-day protocol of alphabeta-TCRmAb, CsA, or FK-506 monotherapy, or a combination of alphabeta-TCRmAb/CsA and alphabeta-TCRmAb/FK-506. VSA transplants were evaluated on a daily basis. Donor-specific chimerism was determined by flow cytometry (FC). The combined protocols of alphabeta-TCRmAb/FK-506 and alphabeta-TCRmAb/CsA significantly prolonged VSA survivals compared to monotherapy groups ( P < 0.005). FC analysis revealed 15.82% of donor-specific chimerism on day 7 under the alphabeta-TCRmAb/CsA protocol and a gradual chimerism decline on day 63 posttransplant. The significant extension of VSA survival achieved under 7-day protocols of combined therapies was directly associated with the presence of donor-specific chimerism.

Vascularized bone-marrow allotransplantation in rats prolongs the survival of simultaneously grafted alloskin.

Skin allografts in rats were rejected, even if they were treated with a sub-optimal dose of FK506. However, skin allografts in rats which simultaneously received vascularized limb allografts were rejected more slowly. In this study, the authors examined the reasons why vascularized limb allografts have the ability to induce immunologic tolerance. For investigating which tissue components provide such tolerance, they compared the mean survival times (MST) of skin allografts on the backs of rats (used as an indicator of rejection), which were simultaneously engrafted with vascularized skin allografts, muscle allografts, joint allografts, and limb allografts. In the cases of limb allografts (MST=32.1 days) (these allografts contain bone-marrow cells), the skin allografts survived longer than vascularized skin allografts (MST=23.6 days) or muscle allografts (MST=22.6 days). For ascertaining whether the ability to induce immunologic tolerance of limb allografts was due to bone-marrow cells or not, the authors compared the MST of skin allografts in rats that were simultaneously engrafted with non-irradiated limb allografts, irradiated limb allografts, and bone-marrow cells. The allografted skin in the rats that received non-irradiated limbs (MST=32.1 days) survived longer than irradiated limb allografts (MST=20.3 days) and bone-marrow cells (MST=18.4 days). Limb allografts are, as it were, vascularized bone-marrow allografts. These results suggested that vascularized bone-marrow allografts can facilitate the induction of immunologic tolerance.

The allogeneic response to cultured human skin equivalent in the hu-PBL-SCID mouse model of skin rejection.

Engineered tissues have been proposed for the treatment of a variety of conditions including the partial or complete replacement of human organs. To determine the basis for the rejection of these tissues, we analyzed the immune response to allogeneic human skin equivalent (HSE, also called Apligraf) in the humanized SCID mouse (hu-PBL-SCID). Two models of hu-PBL-SCID were used for these studies. In one model, human skin or HSE was transplanted onto humanized mice so that graft survival could be analyzed. In the other model, skin grafts were allowed to heal on naive mice before humanization. This model was used to analyze the immunologic response to the vascularized skin allograft. Humanization was performed by adoptive transfer of human PBL into SCID mice by i.p. injection. Both human foreskin and HSE successfully engrafted onto naive SCID mice and remained stable for more than 6 months. In contrast, human foreskin was rejected by 21 days posttransplant in hu-PBL-SCID, whereas HSE consistently engrafted for more than 28 days. Treatment of HSE grafts with interferon-y for 5 days to induce maximal MHC class II molecule expression before grafting failed to induce rejection. HSE also engrafted onto hu-PBL-SCID mice that were exposed to alloantigen by prior injection with interferon-gamma-treated keratinocytes identical to those used to generate the HSE. In addition, we determined that humanization of SCID mice following engraftment and vascularization of human foreskin resulted in marked CD3+ T cell infiltrates and a lymphocyte-induced vasculitis. In contrast, the response in vascularized HSE was associated with minimal CD3+ T cell infiltration in the absence of vasculitis or morphological features of rejection. These results support the use of HSE and other allogeneic engineered tissues in humans provided that such tissues are limited in their antigen presenting capabilities. In addition, our findings suggest a critical function for the donor endothelial cell in rejection.

Immunosuppression by Anti-ICAM-1 and Anti-LFA-1 Monoclonal Antibodies of Free and Vascularized Skin Allograft Rejection

Immunosuppression by anti-adhesion molecule antibody of free or vascularized skin allograft rejection was investigated in rats. Lewis (LEW, RT1 1) rats were used as donors and Fisher (F344, RT1 1v1) rats as the recipients. When F344 rats were treated intraperitoneally (i.p.) with anti-intercellular adhesion molecule-1 (ICAM-1) mAb (1A29) (3 mg/kg/day) and anti-leukocyte function-associated antigen,-1 (LFA-1) mAb (WT. 1) (3 mg/kg/day) one day prior to grafting and daily after grafting for nine days, free skin graft survival was prolonged only slightly compared with that in control rats which were injected i.p. with a daily dose of 6 mg/kg of anti-TNP mAbs (H1-6-2) one day prior to grafting and daily after grafting for nine days. (Mean survival time [MST] of the free skin graft was 11.2 ± 0.6 days in the control group and 13.4 ± 0.3 days in the 1A29 + WT-1 treated group [p<0.01], respectively.) On the other hand, the vascularized graft survival was prolonged significantly in anti-ICAM-1/LFA-1 mAbs-treated F344 rats as compared with that in control rats. (The mean vascularized graft survival time was 14.2 ± 0.7 days in the control group and 21.5 ± 1.9 days in 1A29 + WT-1 treated group [p<0.002].) Our results suggest that interaction with ICAM-1 and LFA-1 is more important in the rejection of vascularized skin allografts than that of free skin allografts.

The Microvasculature is the Critical Target of the Immune Response in Vascularized Skin Allograft Rejection

Vascularized first set human skin allografts were rejected largely by a process of extensive and progressive microvascular damage leading to ischemia and infarction. Microvascular injury was associated with a cellular immune response. However, vessel damage was at least in part immunologically nonspecific because vessels of the graft bed (host tissue) were damaged as well as those of the graft itself. We conclude that the microvascular endothelium is the critical target of the immune response in vacularized skin allografts in man, and that this sequence of events--primary vascular damage followed by ischemic infarction--may have significance in a variety of experimental and clinical settings.

THE IMMEDIATELY VASCULARIZED SKIN ALLOGRAFT

A model for immediate vascularization of skin was devised to examine one of the possible explanations for the differential survival rates of transplants of freely grafted skin and organs, i.e., the increased vulnerability of skin to early ischemic necrosis prior to revascularization. Female Fischer (F) rat skin was transplanted beneath the kidney capsule of tolerant female Lewis (LEW) recipients. This skin healed in and initially appeared to be normal grossly and microscopically. In 27 rats, after 30 days, the composite grafts were excised, and immediately transplanted by means of vascular anastomosis into normal LEW recipients (syngeneic to the kidney portion of the graft and allogeneic to the skin). For 5 days after transplantation of the composite graft, the skin appeared to be normal with minimal or no inflammation in a panel of 11 recipients. From the 6th to 11th day, an active rejection reaction developed at the skin-kidney interface in a panel of six rats. In 10 rats, in which the composite grafts remained in their secondary hosts for 12 to 21 days, rejection of the skin was complete. The renal portion of all composite grafts appeared to be normal histologically. All recipients of composite grafts rejected subsequent orthotopic F skin grafts in an accelerated manner, with median survival times of 8.2 +/- 0.3 days compared to 11.5 +/- 0.7 days in untreated F leads to LEW controls, demonstrating that the skin on the composite graft was fully immunogenic. These results demonstrate that immediately vascularized skin allografts between rats compatible at the major Ag-B1 locus will still be rejected within 2 weeks compared to survivals of up to 48 weeks in renal allografts in the same histocompatibility combinations, suggesting that anatomical factors are not sufficient to account for the differences in survival times between skin and solid organs.

Rejection of first-set skin allografts in man. the microvasculature is the critical target of the immune response.

Recent reports of microvascular injury in delayed hypersensitivity skin reactions prompted us to reexamine the pathogenesis of first-set skin allograft rejection in man using morphologic techniques that allowed both extensive vessel sampling and unequivocal evaluation of microvascular endothelium. We here report that widespread microvascular damage is a characteristic, early consequence of the cellular immune response to first-set human skin allografts and is qualitatively similar to, but substantially more extensive than, that occurring in delayed hypersensitivity reactions. Microvascular damage in invariably preceded significant epithelial necrosis and affected initially and primarily those venules, arterioles, and small veins enveloped by lymphocytes. Vessels of both the allograft itself and the underlying graft bed (recipient tissue) were equally affected. These data suggest that endothelial cells of the microvasculature are the critical target of the immune response in first-set vascularized skin allograft rejection in man and that rejection can be attributed largely to ischemic infarction resulting from extensive microvascular damage. Other mechanisms, such as direct cellular contacts between infiltrating lymphocytes and epithelium, apparently played only a minor role. The findings presented here indicate that the rejection of first-set vascularized skin allografts, though induced by immunologically specific mechanisms, is primarily effected by final pathways that are relatively nonspecific and that may cause damage to both foreign and host vessels and cells. Rather than contradicting studies demonstrating the exquisite specificity of allograft rejection in other systems, these findings provide a further example of the heterogeneity of the cellular immune response. Recognition of the critical role of immunologically mediated microvascular injury may prove important both for an understanding of the biology of allograft rejection and for strategies aimed at prolonging allograft survival.