Human Skin Allografts Research Articles

Severe combined immunodeficient mice engrafted with human splenocytes have functional human T cells and reject human allografts.

Previous studies have shown that human hemopoietic cells can be adoptively transferred into immunodeficient C.B-17 scid/scid (SCID) mice that lack autologous T and B lymphocytes, to generate chimeric animals. The future development of novel immunomodulatory drugs in transplantation will depend increasingly on experimental animal models to investigate the properties of the agents on human cells before starting clinical trials. However, in previous models of SCID mice engrafted with human PBLs, human T cells have been found either to be in an unresponsive state, unable to respond to mitogenic stimulations in vitro, or to mediate skin graft rejection only when HLA-primed in vivo before their adoptive transfer into SCID mice. In addition, T cells and other leukocyte subsets engraft quite poorly in the lymphoid tissues of the animals. In an attempt to develop a useful model for transplantation research, we have inoculated SCID mice with fresh human splenocytes from cadaveric organ donors (hu-Spl-SCID mice). In this model, various leukocyte subsets engraft effectively in different lymphoid compartments. In addition, human T cells retain their proliferative responses to mitogens and to alloantigens when tested 3 wk after engraftment into SCID mice. Finally, mice engrafted with unprimed human spleen cells acutely reject human foreskin allografts. Treatment of hu-Spl-SCID mice with OKT3, an immunosuppressive mAb directed against the human CD3 complex associated with the TCR, prevents the rejection of most human skin allografts, indicating a major role for human T cells in this phenomenon. Thus, this hu-Spl-SCID model may be useful for the study of immunosuppressive therapies in a preclinical in vivo setting.

Indefinite survival of human skin allografts in patients with long-term immunosuppression.

The transplantation of a sufficient quantity of good quality skin is of great clinical importance. Severe burn patients lack adequate skin donor sites to cover their wounds with autografts. Patients with severe injuries and burns of areas such as the entire face are presently reconstructed with multiple full- or split-thickness skin grafts. The final result is a patchwork appearance of skin with inferior qualities to normal full-thickness facial skin. Before structures such as hands should be transplanted, each individual tissue component should be evaluated to determine whether the tissue will survive and function with immunosuppression. Skin is obviously an important part of the hand. One hypothesis tested is the ability of transplanted human skin to survive indefinitely with long-term immunosuppression. The early detection of rejection of transplanted organs is paramount in reversing the rejection phenomena and hence saving the transplanted organ. Another hypothesis tested is the ability of a synchronously placed, same-donor, same-recipient skin graft to be used to detect rejection of a transplanted organ. As the skin and kidney from the same donor have similar antigens and the transplantation of both these tissues is to the same recipient and immunological system, it is hypothesized that there will be a correlation of rejection between the transplanted skin and the transplanted organ. To test the above hypotheses a small skin graft was placed on renal transplant patients synchronously with renal transplantation. The skin allografts were followed by direct observation and biopsy at regular intervals and at the time of suspected rejection. The patients were treated with the usual renal transplant immunosuppressant drugs.(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of biological wound dressings on the healing process

Three major biological dressings are available for the temporary closure of wounds: partial-thickness cadaveric human allograft skin, several forms of partial-thickness antibiotic-treated porcine xenograft skin, and human amnion. Generally, biological dressings reduce pain, close the wound to contamination and fluid loss, and prepare the wound bed for permanent closure, usually with autografts. The three types of biological dressings differ in their performance, with allograft skin being clearly superior in its wound maintenance and preparation characteristics, while porcine xenograft presents serious difficulties in incorporation into the wound bed and antigenic challenge to the recipient, and amnion is excessively fragile and tends to allow wound desiccation. The most serious potential liability of biological wound dressings is transmission of infection; however, the actual incidence of such transmission is extremely low. The advantages of physiological coverage provided by biological wound dressings greatly outweighs the chance for harm in the case of human allograft.

The use of intermingled autograft and parental allograft skin in the treatment of major burns in children

Large burns continue to pose the problem of providing sufficient autologous skin cover. The experience of this unit using cultured keratinocytes as a substitute for split-skin grafts has been disappointing; at the same time, we have been obliged to abandon human allograft skin from cadavers and other patients because of the possibility of infection with HIV. Our favoured method for resurfacing large-area burns in children is to use widely meshed autologous skin overlaid with meshed allograft from a parent (to minimise the risk of HIV transmission). We report our experience using this technique in 10 children. The fate of the intermingled grafts has been followed clinically, and in some cases histologically and by Y-chromosome identification. There has generally been long-term persistence of the parental skin without rejection, and allograft dermis appears to contribute to the final cover. Evidence suggests, however, that cellular elements of the parental skin do not survive.

DC electrical properties of frozen, excised human skin.

DC current-voltage relationships and sodium ion transport measurements for human allograft skin immersed in saline buffers have been determined using a four terminal potentiometric method and diffusion cells of our own design. About three-fourths of the skin samples were deemed suitable for study on the basis of their high resistivities and similar j-V characteristics. Most of these samples yielded sodium ion permeability coefficients less than or equal to those reported for human skin in vivo. The current-voltage relationship in these tissues was time dependent, highly nonlinear, and slightly asymmetric with respect to the sign of the applied potential. Skin resistance decreased as current or voltage increased. For current densities less than 15 microA/cm2 and exposure times of 10-20 min, this decrease was almost completely reversible; at higher current densities, both reversible and irreversible effects were observed. The overall dependence of current on voltage was nearly exponential and was satisfactorily described by an equation of the form j approximately sinh V. Diffusion potentials, sodium ion membrane transference numbers, and sodium ion flux enhancement factors during iontophoresis were measured for skin immersed both in normal saline solutions and in saline solutions of differing concentrations. The sign of the diffusion potentials and the value of the sodium ion transference number (0.51 in normal saline at pH 7.4) indicated a weak permselectivity of the skin for transport of sodium ion versus chloride. At a current density of 71 microA/cm2 and transmembrane potentials in the range of 1.1-1.6 V, the flux enhancement for sodium ion was three to five times greater than that predicted for an uncharged homogeneous membrane according to electrodiffusion theory. For transmembrane potentials less than 0.17 V, agreement of this theory with the data was better but still incomplete.

Electrical analysis of fresh, excised human skin: a comparison with frozen skin.

Samples of human allograft skin prepared without freezing ("fresh skin") were found to have electrical and sodium ion transport properties which differed only slightly from those of skin which had been similarly treated but stored frozen ("frozen skin"). The fresh skin samples were less permeable to sodium ions during passive diffusion and less conductive than frozen skin at low current levels. They were more permselective for sodium versus chloride during constant-current iontophoresis and showed slightly more asymmetry in their current-voltage properties. Overall, the electrical behavior of the two tissues was similar enough to support the use of frozen tissue in iontophoresis studies. However, caution should be exercised when considering the use of frozen skin for applications, such as those based on electroosmosis, where the observed differences could have a major impact on the results.

Treatment of Burns in Children

Decreases in mortality from major thermal injury over the last 20 years have been due to advances in resuscitation, control of infection, support of the hypermetabolic response, and early closure of the burn wound. Of these advances in burn care, early wound closure has progressed the most in the last five years. The restoration of the protective functions of the skin is of primary importance to the recovery of the burn patient. Biologic dressings (pigskin, amnion, human skin allograft) when applied to fully debrided, relatively uncontaminated wounds have been shown to adhere to the wound surface, reduce the wound colony counts, limit fluid and protein loss, reduce pain, and increase the rate of epithelialization over that obtained with application of topical antimicrobial agents.

T cell subsets in allograft rejection. In situ characterization of T cell subsets in human skin allografts by the use of monoclonal antibodies.

We have provided evidence that both major T cell subsets, T4-positive (helper/inducer) and T8-positive (cytotoxic/suppressor), infiltrate human skin allografts. Overall, and in the graft dermis and graft bed, T4-positive cells were predominant (1.5 to 3 times more numerous than T8-positive cells). In contrast, T8-positive cells were relatively more numerous in the epidermis and hair follicles. Rejection probably proceeded by two apparently independent pathways: 1) direct contact killing of graft epithelial cells, presumably by immunologically specific T8-positive cytotoxic cells, and 2) injury of microvascular endothelium of both the graft and graft bed with secondary graft infarction. Although important in first set skin allograft rejection, the mechanism of the second type of killing is uncertain. T4-positive cells were probably involved, as evidenced by their greater numbers; furthermore, studies in mice have shown that transfused helper/inducer cells are able to effect first-set skin graft rejection. It remains to be determined whether T4-positive cells act alone or cooperate with other cells to destroy vessels and bring about graft rejection. Langerhans cells were recognized in epithelial and dermal compartments of both allografts and autografts by their reactivity with anti-T6 and anti-Ia antibodies. We could not determine whether such cells in allografts were of host or donor origin.

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.

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.

Clinical Skin Banking

A simplified procedure for the harvest and preservation of human allograft skin for burn treatment is outlined. In establishing a skin bank to serve the San Diego County Regional Burn Treatment Center, our primary goals were to deliver a low-cost product, monitored to be free of significant bacterial contamination. Our own innovations include the use of Falcon 2070 tubes as packaging material, and the use of microwave energy for the safe, rapid thawing of liquid nitrogen stored tissue.

Allotransplantation of cultured human skin

Human skin allografts cultured for approximately 6 weeks on a solid medium were transplanted to non-related human recipients with skin defects of various etiology. Although based on this study no answer could be given to the question whether these grafts survived or were replaced by cells of the recipient, it was observed that in most cases the grafts were not rejected and remained in situ several months up to over a year.

The effect of culturing human skin allografts before transplantation

Summerlin (1973) reported that mouse and human skin allografts behaved as autografts after a period in a tissue culture medium. In man, he used 2× 2 cm pieces of skin cultured for 4 to 6 weeks and reported success in 4 patients with granulating wounds. To be of value clinically, however, homograft skin must be available in sizes much larger than Summerlin's and the following experiments were devised to see if his technique worked equally for large grafts.