No abstract

We have shown that both xenografts and allografts of rat skin can be acutely and severely damaged by antisera specifically reactive with graft antigens, and that in appropriate circumstances the phenomenon can be elicited routinely (1, 2). A matter of paramount importance among the variables that influence the occurrence of this form of immunologically mediated tissue damage is the interval of time between the placement of the grafts and the administration of antiserum (3). Contrary to widely held views on the special vulnerability of grafts that are healing into place, it has been found that antiserum injected at the time of grafting or within 5-6 d thereafter has little detectable influence on the course or time of survival of the transplanted skin. This period of insusceptibility is rapidly succeeded by a state of sensitivity to humoral antibody that, in immunosuppressed hosts, reaches a peak of intensity at about 14-16 d after grafting and persists at decreasing levels for an additional 3 wk. Grafts that survive beyond that period of time regain their resistance to antiserum, and this state is maintained for the duration of survival of the grafts.We have analyzed these changes in the responses of xenografts of skin to antisera, and we have found that the initial state of insensitivity differs substantively from that observed in long-standing grafts. We describe here the results of our studies on freshly placed grafts, and in a succeeding paper (4) we report on the mechanisms involved in the acquired resistance of grafts that have survived for relatively long periods of time. Materials and MethodsAnimals. B6AF1 and CAFa mice were purchased from The Jackson Laboratory, Bar Harbor, Maine. Lewis (LE) and (Le × BN)F1 hybrid rats (LBN) were obtained from Microbiological Associates, Walkersville, Md. CD rats were purchased from the Charles River Breeding Laboratories, Wilmington, Mass.Antisera. Rabbit anti-mouse thymocyte serum (RAMTS) 1 was prepared as described (5) or was obtained from Microbiological Associates. 13 pools of antisera were used, the least potent of which extended the mean survival time of rat skin grafts to 32.5 :t: ! 1 d. Rabbit anti-rat *

Rat skin that survives for long periods of time on immunosuppressed mice becomes resistant to anti-graft serum and remains so for as long as it survives. When long-standing grafts are removed and placed on new immunosuppressed mice, they remain resistant to antiserum for as long as they survive. The acquired resistance to antiserum seems, therefore, to be due to changes in the grafts rather than to changes in their hosts. Furthermore, it was found that the acquisition of resistance is correlated with replacement of graft endothelium by host cells, as demonstrated by the use of immunofluorescent techniques in conjunction with mouse anti-rat serum and rat anti-mouse serum. Evidently, humoral antibodies are able to cause acute damage to skin grafts, and presumably to grafts to other organized tissues, only if they react with antigens of graft endothelium. Long-term grafts that are retransplanted to their original donors or to rats syngeneic with those donors are in most cases rejected, whereas 14-d-old grafts similarly regrafted are in no case rejected. Apparently, the responses of the secondary recipients to the mouse endothelial antigens in long-term grafts lead to destruction of the entire grafts. When long-standing rat skin xenografts are removed and placed on untreated mice syngeneic with the primary hosts, they are in every case rejected, although they survive slightly longer than skin taken directly from rat donors. Rejection is accompanied by a mononuclear infiltrate and is qualitatively indistinguishable from the rejection of freshly prepared rat skin. Clearly, sensitized cells are more efficient than humoral antibody in destroying grafted tissues.

We have described briefly the acute destruction of rat skin xenografts after injections of their hosts with antisera specifically reactive with graft antigens (1). The rat skin had been grafted to mice whose immune responses were suppressed by removal of the thymus and treatment with rabbit antimouse lymphocyte serum. When these xenografts had healed completely into their beds and could be fully expected to survive for one or more additional weeks, the mice were injected with antirat serum that had been prepared in mice or in rabbits. Signs of graft damage were evident as early as 10 rain after the injection of serum, and in most animals so injected the grafts were completely destroyed within 24-48 h. Studies in which fixed sections of grafts were examined microscopically revealed the early occurrence of edema and infiltration of the grafts with polymorphonuclear leukocytes followed by extreme dilatation and congestion of small vessels, hemorrhage, and necrosis (2). Both gross and microscopic aspects of these reactions were similar to those described for Arthus reactions and other forms of immune vasculitis in rabbits and guinea pigs (3, 4). Reactions of the latter types have been the subject of intensive investigation in recent years, and a good deal has been learned of the pathways leading to tissue damage in these reactions (5). We have looked, therefore, for similar mechanisms in the destruction of skin grafts that is mediated by humoral antibodies, and the experiments to be described here were designed primarily to evaluate the roles of granulocytes and the plasma complement system, agents which are known to be involved in a variety of lesions that are initiated through immunological reactions. Materials and MethodsExperimental System.--The basic test system has been described in detail (2). Adult mice were thymectomized 1-2 wk before receiving grafts of rat ear skin. Rabbit antimouse lympho-