Search

Experimental and clinical data currently available suggest that of the various possible ways in which the outcome of organ homotransplantation might be improved upon, the one which holds most immediate promise involves devising a selection procedure capable of excluding gross histoincompatibilities. The various approaches to the problem of tissue typing or matching that are presently being investigated are reviewed from the viewpoint of their underlying principles and their potentialities. These methods include direct grafting tests, serologic procedures aimed at matching or identifying pertinent isoantigenic specificities associated with formed elements of the blood, especially leukocytes, and various extracorporeal biologic assays of the capacity of an intended host's immunologically competent cells to react against the transplantation isoantigens of a possible donor. The latter category of tests includes the normallymphocyte transfer test and the irradiated-hamster test.

Based on self-reported levels of alcohol consumption, 473 college students (295 female and 178 male) were placed into at-risk or not-at-risk groups. Using reasons given for drinking as the independent variables, discriminant analysis procedures were conducted separately on the males and females to determine if a function could be found which would discriminate between the groups. For the female group, 11 of 22 reasons defined a discriminant function which accounted for 36% of the variance between the groups (p.001). This function was also able to correctly classify 71% of the holdout sample. For the males, five of the 22 reasons defined a discriminant function which accounted for 36% of the variance between the groups (p.001). This function was able to correctly classify 69% of the holdout sample.

"To supply the defects of Nature" was, according to Ambroise Pare in the sixteenth century, "proper to the duties of a chirurgian." Death, illness, and disability of various kinds frequently result from disease or injury involving an organ of the body. In the case of paired organs, loss or a functional shortcoming of one may be naturally compensated by the other assuming an additional load. In the special case of endocrine glands, life can be sustained by administration of appropriate hormones. It has been argued, however, that the interests of the patient might be better served by means of a living allograft if simple, innocuous means were available to control the host response on an immunologically specific basis. Currently, for the major internal organs and for certain tissues such as bone marrow, the only generally available treatment for loss or failure is allotransplantation with its attendant, and by no means satisfactorily resolved, problems of tissue typing and immunosuppression to assuage host resistance. The modern clinical application of organ transplantation, largely the outcome of interactions between surgeons and basic scientists over the past 25 years, originated from critical analyses of the failures of early empirical attempts at organ and tissue replacement and, with their understanding, the development of means of controlling host resistance. Modern endocrinology, including hormone replacement therapy, also had its origins in transplantation, since the classical criterion for ascribing endocrine function to an organ is that replacement away from its normal vascular and nervous connections must successfully correct specific deficiencies brought about by removal of the organ. Most of the pioneering studies in transplantation biology and endocrinology were conducted within the first 2 decades of the present century, and there was little interaction between the investigators pursuing

Studies conducted upon inbred strains of mice, hamsters and rats have shown that following the interstrain matings the now familiar covert reactivity of pregnant females to the alloantigens of their conceptuses may benefit the latter in two ways; firstly, it exerts a significant influence upon placental weight, and indirectly upon the birth weight of the fetus-allogeneic placentas tending to be heavier than syngeneic placentas, and mothers specifically presensitized against alien paternal tissue antigens gestate fetuses with heavier placentas than normal females. Specifically tolerant mothers, on the other hand, produce smaller, F1 hybrid, fetoplacental (fp) units. The classic notion that the disparity between the birth weights of F1 hybrid and homozygous offspring is due to hybrid vigor has been challenged by the finding that DA and (DA times F1)F1 hybrid blastocysts transferred to the uteri of genetically tolerant (DA times F1)F1 hybrid rats produce fp units of similar weight Maternal immunological reactivity against the fetus qua allograft may make a significant contribution here. Additional support for the premise that maternal reactivity against fetal alloantigens in some way promotes the growth of the fp unit was afforded by the finding that excision of the para-aortic lymph nodes (which drain the uterine horns) from females before interstrain matings resulted in smaller fp units than in females subjected to sham operations. The finding with one rat strain combination that passive immunization of females with serum against their F1 hybrid conceptuses promoted the growth of the latter suggests that a humoral rather than a cellular immunity may be involved. Secondly, in the three species studied, it was observed that genetic disparity between a conceptus and its mother significantly improved its chances of implantation and development to term.

Human milk lymphocytes (ML) can be partially purified and propagated in vitro as a means of assessing their immunological function. When exposed to a variety of stimuli known to activate T lymphocytes, ML respond in a unique manner that indicates a selected population of immunocompetent cells. ML are hyporesponsive to to nonspecific mitogens and respond in a reduced manner to histocompatibility antigens on allogeneic cells. In most cases, they are completely unresponsive to C. albicans although blood lymphocytes from the same patients respond to the antigen. The Kl capsular antigen of E. coli induces significant proliferation in lymphocytes obtained from milk, but fails to stimulate blood lymphocytes. This dichotomy of reactivity does not appear to result from suppressive factors or cells in milk or insufficient adherent cell function. Rather it appears to reflect the accumulation of particular lymphocyte clones in the breast and the local nature of mammary tissue immunity at the T-lymphocyte level.

SUMMARY The case with which rats of one strain can be rendered tolerant of skin grafts from another strain is correlated with their Ag-B genotypes and not with the median survival time of skin grafts exchanged between them. Thus, although adult Lewis recipients reject Ag-B-compatible Fischer skin grafts as promptly as they reject Ag-B-incompatible BN or DA skin grafts, Lewis recipients are much more easily rendered unresponsive of Fischer skin grafts than of BN or DA grafts. Where Ag-B compatibility prevails, tolerance is inducible with relatively low dosages (≤1 million) of cells originating from all components of the lymphohematopoietic tissue system, including the thymus. On the other hand, when donor and host differ at this locus not only are much higher numbers of cells required to induce tolerance, but there are wide disparities in the tolerance-conferring capacity of different cell types. Bone marrow is superior to spleen which, in turn, is better than lymph nodes; thymocytes are practically ineffective. Evidence is also presented which suggests that the facility with which tolerance can be abolished by transfer of isologous cells is a function of the dosage of the inoculum used to induce tolerance. The importance of the Ag-B locus in determining susceptibility to runt disease is also considered.

Using guinea pigs of strains 2 and 13 and their F1 hybrids as experimental subjects, various lines of evidence have been obtained that in this species, as in all others tested, the only significant cellular antigens with which donor lymphocytes engage when normal and immune lymphocyte reactions are incited are radiosensitive leukocytes. Constitutive cells of the skin are unimportant. (a) The intensities of these reactions in irradiated subjects are dependent upon the peripheral leukocyte concentration. When this falls below a certain threshold no reactions are incitable. (b) Highly leukopenic animals are capable of developing immune lymphocyte transfer (ILT) reactions if normal lymphoid cells of their own genetic constitution are mixed with the putative attacking donor cells, as "supplementing antigen," before inoculation. (c) Radiation-chimeric strain 13 animals having F1 hybrid leukocytes in their bloodstream give typical ILT reactions when challenged intradermally with strain 13 anti-2 node cells. Exposure of strain 2 animals to 600 R does not prevent their becoming actively immunized if, 24 hr later, they are injected intradermally with strain 13 lymphocytes. However, this sensitization, revealed by the host's capacity to give delayed hypersensitivity reactions, wanes as leukopenia progresses. On the basis of this and other findings it is argued that the flare-up stage of the NLT reaction in preirradiated hosts is mainly an expression of host sensitivity against the transferred alien cells. Two unexpected observations have been made in the course of this study: (a) F1 hybrid animals developed what appeared to be a strong delayed hypersensitivity after intradermal inoculation with parental strain lymphoid cells or antigenic extracts prepared from them. (b) If strain 13 guinea pigs which had been sensitized against strain 2 tissue antigens by intradermal injection of lymphocytes 7 days beforehand were inoculated intravenously with strain 2 antigenic extract a significant proportion of the animals developed severe delayed necrotizing reactions, recall flares, at some or all of the healed skin inoculation sites.

The possibility that an immunological tolerance of maternal skin homografts may be acquired naturally in guinea pigs as a consequence of the accidental incorporation of maternal cells into the fetuses has been re-investigated. F1 hybrids between two isogenic strains, 2 and 13, were reciprocally backcrossed to strain 13 animals to produce two genetically heterogeneous populations which differed only insofar as those animals which had developed in an F1 hybrid uterine environment had had an opportunity to incorporate strain 2 cellular antigens ante partum. To test for any altered immunological reactivity this might have caused, both populations of animals were challenged with strain 2 skin homografts when they were 30 days of age. Although two of 192 offspring of F1 mothers failed to reject their grafts during the 200 day observation period, the distribution of survival times of the remaining grafts in this group of animals was very similar to that of the grafts transplanted to 167 offspring of strain 13 females. It is concluded that the two exceptional cases are most probably the result of chance antigenic compatibility and that maternally acquired tolerance is a very rare phenomenon in guinea pigs, if it occurs at all.

Rats have been shown to be capable of displaying the various kinds of delayed cutaneous hypersensitivity reactions attributable to transplantation immunity previously described in guinea pigs, hamsters, rabbits, dogs, and man. The rat is unlike most other species in that much larger numbers of lymphoid cells are needed to incite these cutaneous reactions. With direct, transfer, or normal lymphocyte transfer reactions, the cutaneous responses were greater when donor and recipient differed at the Ag-B histocompatibility locus than when donor and recipient shared the same Ag-B alleles. An experiment was performed in which adult rats of a genetically defined backcross population, resulting from matings between DA and (DA x Lewis) F1 hybrid rats, were inoculated intradermally with lymph node cells from DA rats sensitized against tissues from Lewis-strain donors. Some of the R2 animals gave a biphasic transfer reaction with peak reactivities occurring first at 48 hr and recurring at 96–120 hr, while the others lacked this second component. Hemagglutination tests revealed that the R2 rats giving the biphasic response possessed the Ag-B,1 antigen, which is also present in Lewis rats, whereas rats which gave monophasic reactions were homozygous for the Ag-B,4 antigenic determinant which is present in the DA strain. This suggested that the recall flare at 96–120 hr reflects proliferative activity on the part of the inoculated cells confronted by the disparate Ag-B isoantigen in the host's dermis. Skin homografts from R1 animals bearing the Ag-B,1 antigen were uniformly rejected by DA hosts in 11 days or less, while grafts from backcross animals homozygous for the Ag-B,4 antigen usually lived longer, being rejected in 9 to 27 days. Evidence is also presented which suggests that specific isoantibodies may act synergistically with immune lymphocytes to bring about cutaneous inflammatory reactions in the rat.

Summary It has been shown that when grafts of adult chickens' skin are transplanted to the chorioallantoic membranes of 9-10-day-old chick embryos they heal in soundly, become vascularized and evoke a characteristic cellular response in the underlying extraembryonic mesenchymal tissue of their host. These grafts also bring about enlargement of, and certain lesions in, the hosts' spleens. It is argued that these changes are the result of an immunologic reaction on the part of certain cellular ingredients present in the skin grafts against the foreign cellular “transplantation” antigens of the embryonic hosts. Experiments involving the use of exsanguinated and perfused donors, and determinations of the minimal amount of whole blood required to produce lesions in the hosts' spleens, have shown that the immunologic competence of chickens's kin cannot simply be ascribed to the activity of certain leukocytic ingredients of the blood in its vessels, but must be regarded as residing mainly, if not entirely, in fixed cells of the skin which are almost certainly lymphocytes.

The intracutaneous inoculation of lymph node cell suspensions from normal hamsters into normal, homologous hosts causes the development of delayed inflammatory reactions, normal lymphocyte transfer (NLT) reactions, the intensities of which reflect the antigenic disparity between donor and recipient. The immunogenetic situations in which they occur indicate that these reactions are due to sensitization in situ on the part of inoculated immunologically competent cells against alien antigens of the host. They are graft versus host reactions. If hamsters are sensitized by cellular or solid tissue homografts and then challenged intracutaneously with lymphoid or epidermal cells from the donor strain, direct reactions are provoked. A positive correlation exists between the development of transplantation immunity and the capacity to give direct reactions. The sensitivity responsible for direct reactions can be suppressed by irradiation or administration of cortisone, and it is transferable by means of viable lymphoid tissues or cells but not by means of serum. Intracutaneous inoculation of viable node or splenic cells from specifically sensitized hamsters into hosts of the donor strain incites transfer reactions, the intensities of which depend upon the number of cells transferred and the level of sensitivity in the animal that provided them. These reactions are not incitable by thymocytes, by killed lymphoid cells, or by serum. Transfer reactions differ from NLT reactions only on a quantitative and chronologic basis. However, a sharp experimental distinction can be drawn between them. For example, if normal MHA node cells and node cells from MHA hamsters sensitized against CB antigens are injected into the skins of MHA's tolerant of CB tissue, only the sensitized cells incite reactions. Both direct and transfer reactions are highly specific immunologically. The inability of node cells to incite NLT or transfer reactions in heavily irradiated hosts and other findings sustain the thesis that it is host cells of hematologic origin, rather than indigenous skin cells, that contribute the antigenic stimulus required for these reactions. All the findings presented are consistent with the hypothesis that the various delayed inflammatory reactions described are manifestations of immunological responses on the part of immunologically competent cells against alien transplantation antigens.

Melanocytes may be regarded as unicellular, pigment-secreting glands, largely though not entirely confined to the epidermis, of which they are a constant though often un-recognized cellular component. Their numerical incidence, and their branching form, with numerous dichotomizing processes, are such as to form a reticular system within the epidermis. Evidence is evaluated that they are a race or lineage of cells sui generis and form a self-maintaining system within the epidermis. Their product-melanin granules-is secreted directly into the cytoplasm of neighboring Malpighian cells upon which the processes of the melanocytes terminate in the form of small swellings or end-caps. The several types of branched cells demonstrable within the epidermis by various techniques are discussed in relation to their identity as living melanocytes, or as effete cells of this lineage which, having lost or discharged their pigment, participate in the general out-ward movement of epidermal cells to be lost from the skin su...

The experiments reported herein provide ample evidence that mice, like most other mammalian species, are capable of displaying readily observable and reproducible delayed cutaneous hypersensitivity reactions indicative of transplantation immunity. By employing a variety of genetically defined strains, it has been shown that a genetic requirement for the development of a positive normal lymphocyte transfer reaction in mice is a difference between host and cell donor at the H-2 locus. By contrast, the immune lymphocyte transfer reaction consistently reflected the full range of histoincompatibility, both inclusive and exclusive of the H-2. It was incidentally discovered that erythema regularly accompanied delayed cutaneous reactions in the skins of female mice, whereas no local redness accompanied their counterparts in male skins. The influence of cutaneous erythema on the scoring of delayed skin reactions is discussed.

I FEEL GREATLY privileged to be the recipient of the 1971 Adair Award, partly on account of the scientific eminence of my predecessors, but principally because of the distinction of Dr. Fred Lyman Adair himself, as a scholar, physician, pioneer, and leaader of his own specialty. The theme of my lecture is an analysis of mammalian reproductive activity from the viewpoint of an immunogeneticist, with special emphasis upon nature’s striking success in ensuring exemption from rejection for her own, all-important, histoincompatible organismic grafts. I believe that no apology is needed for inclusion of a brief introductory account of the basic principles governing the success of the conventional grafts of the experimentalist and the surgeon.

Free tail skin grafts or suspensions of viable epidermal cells have been placed in the atraumatized uterus of isologous rat hosts and allowed to "implant" of their own accord to study the possible uniqueness of this site for other than nature's transplants, i.e. conceptuses, and its response to unnatural grafts. Despite the presence of an intact endometrial epithelium, free skin grafts heal-in rapidly, provided that a state of estrogen excess is established at the time of transplantation. In the absence of estrogen most of the grafts failed to implant. Once established, the grafts survive indefinitely without further estrogen. However, if at any stage a state of continual estrus is established, skin epidermis migrates centrifugally from the graft perimeter invasively replacing the native uterine epithelium. The results of an analysis of the modus operandi of this estrogen-facilitated epidermal migration in utero sustain the view that the hormone acts upon the uterine stroma rather than upon the epidermal cells. When grafts of lingual mucosa or vaginal "skin" were placed in the uteri of rats maintained in chronic estrus, migration of epidermis took place even more vigorously than from tail skin. These epithelia conserved their distinctive histologic specificities indefinitely when growing as heterotypic recombinants on the alien mesenchymal stroma of the uterus. Monodisperse suspensions of epidermal cells appear to "implant" and establish small epidermal plaques in the uterus only at sites predestined to accept conceptuses. That the endocrinologic parameters for the establishment of skin grafts in the uterus are similar to those for blastocysts is suggested by the finding that both kinds of graft can become established in the same uterine horn in the absence of exogenous hormones.

Summary The principle of immunologic tolerance has been employed to study the isoantigen—usually referred to as the Y-factor—that is responsible for the rejection of male skin isografts by female mice of many different inbred strains. Various lines of evidence are presented that lend strong support to the view that this antigen is determined by a locus on the Y chromosome and that it behaves in all respects like the tissue transplantation antigens responsible for the destruction of tissue homografts. Tolerance of male skin isografts, like tolerance of skin homografts, can be abolished by transfer of normal or sensitized lymphoid cells from a genetically appropriate donor. Exactly the same Y antigen appears to be present in all male mice, irrespective of their strain of origin, as evidenced by the fact that C57 female mice could be made tolerant of subsequent male skin isografts by inoculation at birth with male cells derived from any one of four different and unrelated donor strains. The tolerance-responsive period of C57 female mice to this fairly weak antigen was found to be of the order of 17 days. Because of the very high degree of toleranceresponsiveness of C57 females to the Y antigen, this simple tolerance system has been employed to investigate several fundamental aspects of transplantation immunology. For example, it has been found possible to render female mice tolerant of male skin isografts by repeated inoculations with high dosages of a cell-free extract of male lymphoid cells. Highly suggestive evidence has also been obtained that persistence of tolerance of the Y antigen does not require persistence of the antigen.

A study has been made of the effect of prior intravenous injections of living homologous dissociated epidermal cells and blood-cells on the survival of skin homografts subsequently transplanted from the cell donors in rabbits. These treatments failed to elicit an immune response resulting in the accelerated breakdown of the skin homografts, even when a massive dosage of homologous blood was administered. On the contrary, in a high proportion of animals injected with their intended donors’ epidermal cells it was found that the host’s capacity to react against the skin homografts was abrogated to an extent that actually increased their survival by a factor of 2 or 3. This prolongation of survival is not merely the outcome of the non-specific treatment of a host with epidermal cells as such, since pre-treatment of animals with suspensions of their own epidermal cells was ineffective in prolonging the survival of homografts. This modification in the host’s response was not obtained by the administration of epidermal cell suspensions by other routes; when the cells were injected intradermally a state of immunity was elicited, and their intraperitoneal injection was without demonstrable effect. The injection of living cells was found to be obligatory since cells killed by freezing and thawing before their injection had no effect on the course of the homograft reaction. The injection of homologous blood gave results comparable with those obtained with epidermal cell suspensions, although they were not as consistent. Evidence is presented suggesting that a state of immunity towards grafts of homologous skin may be evoked by prior intradermal injections of epidermal cell washings not containing intact living cells. Possible explanations and affinities of the induced abrogation of a host’s immunological response to skin homografts are discussed, as well as its possible implications.

F1 hybrid hamsters derived from genetically disparate strains develop a severe and often lethal cutaneous disorder when inoculated intracutaneously with immunologically competent lymphoid cells from either parental strain, The disease is characterized clinically by extensive epidermal necrolysis, and histologically by a complete dissolution of the dermal-epidermal junction. The requisites for elicitation of this syndrome were determined to be: (a) the parental strains must differ from each other at a major histocompatibility locus, and (b) the donor inoculum must contain immunologically competent parental strain cells. In addition it was found that specifically sensitized cells surpassed normal unsensitized ones in their ability to elicit the disease, and that the disease can be transferred adoptively from affected to normal F1 hosts by means of lymphoid cells. On the basis of these observations, it was concluded that the disease was immunologic in nature, and graft-versus-host in type. However, a series of critical studies failed to demonstrate that the epidermolysis had an immunogenetically specific basis, thus invalidating the provisional assumption that this lesion resulted from a direct immunologic attack upon parenchymal cells of the epidermis and dermis. With the aid of radiation chimeras, it was clearly established that typical epidermolysis could be induced in skin of the same genetic constitution as the attacking donor lymphoid cells. This paradox was taken into account by the possibility that, amid the intense local cutaneous graft-versus-host reactions, "skin-specific" antigenic determinants are bared which incite a quasi autoimmune response that in turn is responsible for the epidermolytic lesions.

Summary An investigation has been made of the possibility of conferring tolerance in respect of skin homografts from any donor in a heterogeneous rat population by inoculation of newborn animals with pooled cell suspensions prepared from multiple, randomly selected donors. Subjects for these experiments were Wistar rats, from a large, closed, but noninbred colony, whose heterogeneity was such that the median survival time of skin homografts exchanged between them was about 12 days, although a small proportion of the animals (10%) were genetically tolerant, accepting their grafts for longer than 50 days. Subcutaneous injection of newborn animals with 15,000,000 to 20,000,000 pooled splenic cells from standard panels of 10 donors was almost ineffective in prolonging the lives of test skin homografts, but following intravenous inoculation, the grafts on about 30% of the subjects lived for longer than 50 days—an arbitrary criterion of a high degree of tolerance. Similar dosages of pooled newborn or infant donor splenic cells conferred a high degree of tolerance upon about 60% of the subjects and pooled adult bone marrow cells were even more effective since more than 80% of the animals were highly tolerant. Evidence is presented that if newborn Wistar rats are injected with cells from newborn or adult spleens or from adult bone marrow prepared from the derived inbred Lewis strain, instead of from pooled Wistar donors, then the proportion that became tolerant of Lewis grafts (60–70%) does not differ significantly according to the type of cell inoculated, so that within the dosage limits employed, cells from infant and adult spleen, and from adult bone marrow, must be considered as potentially equally effective in conferring tolerance. It is argued that the different results obtained with the different tissues when pooled cells from heterogeneous donors were employed are expressions of the varying extents to which immunologic interactions occurred on the part of immunologically competent, lymphoid-type cells present in the inoculums. These reactions may have led to the destruction of all cells of some donor genotypes before they had had an opportunity to induce tolerance in respect of some important transplantation antigens of the population. The high degree of efficacy of pooled marrow is attributable to its very low content of immunologically competent cells, whereas that of infant spleen probably depends upon the fact that here the majority of the abundant lymphoid cells are too immature to interact, or may even become tolerant of many of the “foreign” antigens present. Evidence is presented that the “polyvalent” tolerance induced by pooled donor cells is highly colony-specific; although it extends to the majority of members of the colony, and also to members of the derived Lewis inbred strain, it is completely ineffective in respect of skin homografts derived from unrelated chocolate B.N. inbred strain donors. It is suggested on the basis of these findings that the probability of being able to confer polyvalent tolerance in a truly wild mammalian population is extremely remote because of the large number of different histocompatibility genes present that would have to be represented in the pooled, tolerance-conferring inoculums. During the course of this study evidence has been obtained that, contrary to the accepted view, the duration of the tolerance-responsive period in the rat closely resembles that characteristic of the mouse. A preliminary account of “runt disease”—the outcome of the reaction of inoculated adult lymphoid tissue cells against a homologous infant host—as it expresses itself in rats is given.

The epidermolytic syndrome that can be obtained at will in F1 hybrid hamsters by the cutaneous inoculation of adequate doses of parental strain lymphoid cells has been investigated to determine whether the cutaneous lesions are due to an autoimmune process arising from the severe, initial GVH reactions in the skin. It was amply demonstrated that inoculation of donor cells into the skin was of crucial importance to the development of epidermolysis. Parental strain lymphoid cells in similar doses delivered by any other route into normal F1 hybrids failed absolutely to incite the acute syndrome. If "immune lymphocyte transfer" reactions incited by donor cells in the host's skin were surgically removed at timed intervals after inoculation, only complete excision within 24 hr prevented the appearance of epidermolysis in F1 hybrid hosts, indicating that inoculated donor cells must remain within the confines of the skin for approximately 24 hr in order to evoke the disease, persistence for longer periods of time being unnecessary for the subsequent course of the disease. However, reconstitution experiments involving the intramuscular inoculation of suspensions containing mixtures of donor cells and host lymphoid cells, in the presence or absence of epidermal cells, unequivocally indicated that no intimate exposure of lymphoid cells to putative skin-specific antigens was essential. Similarly, the elicitation of generalized epidermolysis in F1 hybrids irradiated with 300 r and then inoculated intravenously with donor cells casts further doubt on the pathogenic importance of the skin as a source of tissue-specific antigen. The results of subsequent experiments indicated that host leukocytes, rather than parenchymal cells of the dermis or the epidermis, were important contributors of the transplantation antigenic stimulus. Moreover, a series of experiments, using (CB x MHA)F1 hybrid hosts that had been lethally irradiated and reconstituted with bone marrow cells from ALS-treated MHA donors, indicated that from 6 to 10 wk after reconstitution—when direct and immune lymphocyte transfer reactions showed a virtual absence of native F1 leukocytes from the circulation—donor cells obtained from specifically sensitized MHA donors were completely ineffective in inducing epidermolysis, while equivalent lymphoid cell inocula derived from CB donors evoked the cutaneous disease irrespective of the time elapsed since reconstitution. To explain these findings it is postulated that in hamsters, the primary targets in graft-versus-host disease incited by the intracutaneous inoculation of donor cells are leukocytes originating in bone marrow or lymph node, or both.

Using rats of the Lewis and BN (Ag-B locus incompatible) isogenic strains, a comparative study has been made of the capacity to prevent or mitigate the development of runt disease with: (a) lymph node cell suspensions from normal adult BN rats, (b) node cells, or (c) serum from donors sensitized against Lewis tissue antigens, or (d) heterologous anti-lymphocyte serum (ALS) raised in rabbits against rat thymocytes. Following a standard intravenous or intraperitoneal inoculation of 20 x 106 Lewis node cells into neonatal BN hosts, there are cutaneous manifestations of runt disease within 11–15 days and death invariably takes place within 20 days. However, complete protection is afforded by administration of a similar number of normal BN node cells via a different vein, or admixed with the otherwise harmful Lewis node cells. However, timing of the administration was crucially important—precedence or delay by as little as 4 hr resulted in a great impairment of protection. When the inoculations of the two cell suspensions were separated by 24 hr, no protection was afforded. These and other observations suggested that a necessary condition for protection of the hosts by unsensitized isologous cells requires that they establish a prompt and intimate confrontation with the homologous target cells. At the same dosage level, suspensions of node cells from sensitized isologous donors were much more effective therapeutically, saving the lives of 92% of treated subjects when administered after a delay of 3 days, and of 19% when the delay was 4 or 5 days. Of the various immunotherapeutic agents studied, daily injections of 0.2 ml of isoantiserum gave the best results, and could totally reverse the course of the disease even when initiated at age 10–13 days and subjects already presented symptoms. ALS, although inferior to isoantiserum at the dosage levels tested, proved to be superior to sensitized isologous cells as a protective agent, since the initiation of daily injections after delays of 6 or 8 days were still effective. The observations that delayed treatments of infant rats with isoantisera or ALS resulted in complete recoveries sustain the thesis that the lesions responsible for the fatal outcome of runt diseases are not inflicted at a very early stage. The efficacy of both isoantisera and ALS as a means of inhibiting the progression of homologous disease also suggests that they may have therapeutic value in situations where this condition is encountered.

With the BN Lewis rat strain combination it has been shown that the median survival time of skin homografts on three-day-old hosts (10.0 ± 0.5 days) does not differ significantly from that of similar grafts on adult animals. This rapid attainment of immunologic maturity has provided the basis for a sensitive test for the induction of tolerance of skin homografts. The putative tolerance-conferring stimulus is injected intravenously at birth and the hosts are challenged with donor strain skin three days later. With this system it has been found that whereas an inoculum of 100,000 bone marrow cells elicited weak sensitization, as evidenced by a significant curtailment of graft survival, increasing the dosage to one million or more cells induced tolerance, as shown by prolongation of graft survival. The proportion of hosts displaying tolerance and its degree, increased with the cell dosage. When test-grafting with skin was delayed until the hosts were 50–60 days old, the minimal number of cells required to confer tolerance was seven million. With the present assay it has been shown that the tolerance induced by low dosage cellular inocula wanes fairly rapidly, though the presence of a skin homograft from the third day onwards may prolong its duration. A previous finding that, with the present strain combination, bone marrow cells are significantly superior to either spleen or lymph node cells in conferring tolerance of skin homografts has been confirmed. It has been shown that adult thymocytes are only capable of inducing feeble degrees of tolerance over a wide dosage range (5 - 50 × 106) and that thymocytes from infant donors are ineffective. Viable suspensions of epidermal cells and x-irradiated or frozen-thawed suspensions of bone marrow cells failed to confer tolerance, though it was induced fitfully by means of a microsomal lipoprotein extract of BN lymphoid tissue. Finally, it was shown that the survival of BN test grafts on three-day-old Lewis hosts was significantly prolonged if they were born of Lewis mothers specifically presensitized against BN antigens. This constitutes evidence of a passive transfer of the isoantibodies responsible for the phenomenon of immunological enhancement.

The present contribution presents a sequel to our previous investigations into problems which arose when attempts were made to use skin grafting to differentiate between monozygotic and dizygotic twins in cattle. In each of three trials grafts from a dam elicited similar reactions when transplanted to each of her one-egg twin offspring. On the basis of this finding the asymmetry of response which the majority of two-egg twins show towards grafts from their dam may be taken as evidence of their dizygotic origin.It has been shown that mutual graft exchange between unlike-sexed twins can be used as a reliable method for distinguishing reproductively normal females from freemartins. The complete success of this method provides strong evidence that homograft tolerance and the freemartin condition have a common anatomical origin.Grafts from a dam to her tetrazygotic offspring lived for upwards of 70 days, whereas grafts from a dam to her offspring of single birth rarely live for as long as 14 days. It is argued that this great prolongation in the life of their grafts is the outcome of the quadruplets having previously exchanged cells in foetal life through anastomoses of their circulations. Each must be incapable of reacting against a very wide spectrum of antigens since it will accept grafts from all its brothers and sisters.Evidence is presented which suggests that cells from her foetus not infrequently gain access to a dam and sensitize her to subsequent grafts of her offspring’s skin. A first pregnancy may be sufficient to elicit this sensitization. On the other hand, if maternal cells gain access to the foetus, this must occur very rarely for in none of seventeen calves tested with grafts of their dam’s skin was there the slightest evidence that they were tolerant.New-born calves react as vigorously as adult cattle against homografts, even when they have not received colostrum. Since new-born calves possess no serum globulins, including antibodies, this provides strong evidence that immune globulins play no essential part in the homograft reaction.

Pigment spread is the natural or experimentally procured (through grafting) progressive encroachment of pigmentation from black or red skin areas into juxtaposed white skin areas, or from black skin areas into red skin areas in spotted guinea pigs and other mammals. So far as spread from black into white skin is concerned, it had previously been shown that migration of epidermal melanocytes into skin lacking homologues of these cells was responsible. However, since red skin already has its own complement of phenotypically "red" melanocytes, the intriguing possibility remained that when black pigment encroaches upon red, rather than melanocyte migration being responsible, phaeomelanin (red)-producing melanocytes are transformed into eumelanin (black)-producing cells by some kind of serially transmissible factor derived from contiguous eumelanotic melanocytes. By utilizing two isogenic strains (Nos. 2 and 13) of spotted guinea pigs and their F1 hybrids, the mechanism underlying the spread of pigment from black into red skin has been analyzed, employing cellular transplantation antigens as melanocyte "markers." The findings demonstrate unequivocally that a physical migration of pigment cells is responsible. By comparing the extents of pigment spread from black ear skin grafts, or from epidermal cell suspensions prepared therefrom, from parental strain or from F1 hybrid donors in white host skin areas of F1 hybrid guinea pigs, it has been possible to evaluate the influence of the intimate contact of melanocytes with alien transplantation antigens on their survival and migratory behavior. No evidence was forthcoming that pigment spread takes place less readily when the cells responsible are confronted by epidermal cells bearing foreign antigens than when they are confronted by cells of their own antigenic constitution. These findings are contrary to expectation if the phenomena of allogeneic inhibition or contact-induced cytotoxicity apply to normal cells in in vivo situations.

IN mice and rats there is one exception to the rule that grafts are always accepted between members of an isogenic population. This applies when females are challenged with skin and other tissues from male donors1–6. The ability of females to reject these grafts varies from strain to strain. For example, while virtually 100 per cent of C57BL/6 (hereafter C57) females and about 75 per cent of A strain females reject male skin isografts, CBA and AU strain females usually accept such grafts. This interstrain diversity occurs despite the fact that the specificity of the Y antigen or “Y factor” is apparently the same in male mice of all stocks7. The basis for this variability stems from two factors: the first is the genotype of the female which determines her capacity to react against male skin and the second is the genetic background of the male which influences the expression of the Y antigen8. Thus the complete penetrance of the Y factor in C57 mice is due to the females of this strain being relatively strong reactors against male skin isografts, and is not because the antigen is strongest in this strain. Indeed, that the antigen is weaker in C57 males than in CBA males follows from the fact that (CBA × C57)F1 hybrid females reject grafts from CBA males more frequently than from C57 males8.