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Prof.Dr. Hans Schmidt of Marburg, Germany, the father of immunobiology, was so puzzled by the alleged lack of immune reactions after fresh cell therapy, that he decided to visit Prof.Dr. Niehans, the father of zellentherapie, in his clinic in Vevey, Switzerland, to see with his own eyes, what happens after the transplantation of fresh cells to patients. H. Schmidt watched nervously the implantations to the patients, and when he did not see any anaphylactic shock, or other serious immune reactions, he admitted that his 20 years' of work in immunobiology was ‘annihilated by just one injection’. This happened in 1950-ies...

No one has been able to explain this puzzle to this date. VI.BIBLIOGRAPHY [143, 95] P. Niehans stated in his book, that he did not know for what reason these considerable quantities of foreign albumins, introduced parenterally into the organism, and repeatedly, fail to produce an anaphylactic reaction. He commented, that professors with experience in the matter of anaphylaxis extended over a number of years, came to his clinic and observed this astonishing fact. He brought up the words of Halsted, that in 1909 stated, that the mechanism of immunization did not work when the organism needed a transplantation, and that an impaired organism, in need of young cells to ensure its recovery, tolerates them extraordinarily well. VI.BIBLIOGRAPHY [18] Perhaps not a purely scientific statement, but it reflects a reality observed daily in the clinical practice of fetal cell transplantation.

Modern immunology has avoided confrontation on this issue by taking an ‘ostrich’ approach: if no immunosuppression is used, then the rejection of the cell xeno-transplants must take place, and if the clinician does not see it, then the clinician is incompetent.

In view of the above it is interesting, and sad, to read the words of K.J. Lafferty, one of the pioneers of islet transplantation in the West, on the right track while he still worked in Australia, when he dared to state that cell xeno-transplantation can be carried out without ‘hyperacute rejection’, and was quickly accused of a scientific fraud by a ‘prince’ of modern immunology, P.B. Medawar. It turned out that K.J. Lafferty was correct and P.B. Medawar was wrong. VI.BIBLIOGRAPHY [170] It is not the first case when ‘Princes’ stopped progress in human history, and also a justification of mandatory retirement of the chiefs of departments at the universities past certain age, regardless of their fame.

Let’s now focus on explaining the reasons, why immunosuppression is not required after fetal precursor cell xeno-transplantation.

Some of the more recent publications indicate that immunologists are beginning to pay attention to the observed clinical data.

Immune responses to all protein antigens require initial processing and presentation of the antigen to T cells by antigen-presenting cells (APC's): dendritic cells, macrophages, and B lymphocytes, are the only cell types, that express also the specialized co-stimulatory molecules, which enable them to activate the naive T cells. VI.BIBLIOGRAPHY [165] T cells recognize foreign antigens only when they are first broken down into short peptides, that are then displayed, in association with major histocompatibility complex (MHC) proteins, on the cell surfaces of APC’s. Thus the role of APC's is to offer antigenic peptides, complexed with MHC proteins, to the available repertoire of T cells. MHC molecules are obligatory components of the antigen complex recognized by T cells. The ability of T cells to recognize specific features of MHC proteins is critical for their survival in the thymus, and for the ability of the immune system to discriminate ’self’ from ‘non-self’.

Human leukocyte antigens (HLA), and genes that encode them, are subdivided into classes I, II, and III. Class I and II molecules are structurally similar cell surface glycoproteins, involved in antigen presentation to T cells. HLA class I proteins are expressed on all somatic cells, while HLA class II proteins are found on only a few cell types. Class II MHC proteins are required for the presentation of the antigen to CD4+ (helper) T cells, whereas CD8+ (cytotoxic) T cells respond to antigens presented by class I MHC molecules.

All genes of human MHC complex are located on the short arm of chromosome 6. There are 3 genes within the class I region: HLA-A, HLA-B, and HLA-C loci, and there are 3 genes within the class II region: HLA-DP, HLA-DQ, and HLA-DR loci. Each of these genes exists in multiple, different allelic forms: 24 alleles of HLA-A and 50 alleles of HLA-B are known. Each person inherits two copies of chromosome 6, and so expresses 6 (3+3) class I and 6 (3+3) class II alleles, but because of extreme polymorphism of these loci, the probability that any two individuals would express identical sets of HLA proteins is very low. VI.BIBLIOGRAPHY [108]

The peptide binding sites of both class I and II MHC proteins are similar, and have similar degenerate specificity, i.e. can bind a fairly wide range of peptides, but the peptides bound by class I MHC proteins are uniformly 8 - 10 aminoacids long, while the peptides bound by class II MHC proteins are 10 - 18 aminoacids long. Both T cell subsets use the same antigen receptors . VI.BIBLIOGRAPHY [108]

Although class I and class II MHC proteins have considerable structural homology, they function differently in antigen presentation. They present antigenic peptides to different T cell types: class I presents to CD8+ cells, and class II presents to CD4+ cells. The two classes of MHC proteins associate with peptides that have been created by different processing pathways within the cell. Class I MHC molecules bind peptides derived from endogenously synthetized proteins, i.e. viral protein in virus-infected cells, while class II MHC molecules bind peptides derived from exogenous proteins, i.e. those in the external medium, that have been internalized by APCs. Thus cells that harbor intracellular infectious agents are recognized and attacked by class I (cytotoxic) T cells, whereas class II (helper) T cells respond to the universe of soluble antigens, providing activating signals for antibody production by B lymphocytes. VI.BIBLIOGRAPHY [108]

It has been speculated that the evolutionary drive behind allo-transplant rejection mediated by cytotoxic T lymphocytes had been the need to eliminate virus infected cells within the organism. In contrast, the defense mechanisms against infection with macroparasites, such as helminths, that are large, multicellular and because of its size cannot be eliminated by phagocytosis, and express both MHC antigens and carbohydrate epitopes, have striking similarities with the findings in the xeno-transplant rejection. VI.BIBLIOGRAPHY [62]

Class I MHC proteins bind peptides already during their synthesis in rough endoplasmic reticulum (RER), and this binding actually helps synthesis and promotes transport of class I molecules from RER to the cell surface. In contrast, class II molecules must be transported from RER to an endosome compartment first, where they lose an invariant chain, and can bind peptides from endocytosed, exogenous protein. Thus class I MHC proteins and class II MHC proteins travel different routes and encounter antigens in different cellular compartments. This difference in routing determines the source of peptide that becomes associated with each class: either endogenously synthetized and transported to the endoplasmic reticulum (endogenous pathway, class I), or exogenous, subsequently endocytosed and degraded in an endosomal compartment (exogenous pathway, class II). VI.BIBLIOGRAPHY [108]

Virtually all somatic cells express class I MHC proteins, and therefore can present endogenous antigens to CD8+ T cells, i.e. when infected by a virus, and killed usually by the lymphocyte as a result. In contrast only a few cell types express class II MHC proteins, and are uniquely able to present exogenous antigens to CD4+ helper T cells, so that they are important for almost all immune responses: ‘antigen-presenting cells’. The best known are macrophages with prodigious phagocytic activity, also producing various regulatory peptides, such as TNF, IL-1, that control lymphocyte proliferation , differentiation, and effector function; and then B-lymphocytes, and dendritic cells derived from a common bone marrow precursor. VI.BIBLIOGRAPHY [108]

Lymphocytes become activated when specific ligands bind to receptors on their surfaces. The required ligands are different for T cells and B cells, but the response itself is similar in many respects for all types of lymphocytes. Only highly polymeric proteins, or polysacharides, are capable to activate B-lymphocytes alone, and they are rare in nature. Usually, the activation of B cells requires the simultaneous presence of a common antigen, binding an individual immunoglobulin on the surface of B cell, and of an activated CD4+ T cell, that interacts with non-immunoglobulin receptors on the surface of B cell to generate a second signal.

Similarly, T cell responses to most antigens require two types of simultaneous stimuli:

1/ an antigen properly displayed by MHC molecule on APC, and

2/ a presence of ‘co-stimulator’: in case of CD4+ T cells a contact with a specific ligand on the surface of APC, or in case of CD8+ T cells the presence of IL-2, secreted by activated CD4+ T cells. VI.BIBLIOGRAPHY [165] Thus all T cells must cooperate with APC's, while B cells depend on CD4+ cells, (CD8+ cells depend on CD4+ cells, too), and these interactions require either surface-to-surface contact or mediation by labile cytokines acting only over extremely short distances, and by an array of adhesion molecules on the surface of the cells with which T lymphocyte interacts, so that all this usually takes place in the secondary lymphoid organs, where lymphocytes, antigens, and APC's, are close to each other at all times. VI.BIBLIOGRAPHY [108, 165]

Dendritic cells, which are found in the T cell areas of lymphnodes, are the most efficient inducers of naive T lymphocytes. Their responsibility is to detect viruses that do not induce MHC class II and co-stimulatory molecules on macrophages. VI.BIBLIOGRAPHY [165]

Rejection reaction:

According to the leading theory in ‘discordant’ organ xeno-transplantation in the un-manipulated recipient, a rapid and violent rejection reaction, destroys the graft within minutes to a few hours. In ‘concordant’ organ xeno-transplantation, where a ‘hyperacute xenograft rejection’ does not take place, or in discordant xeno-transplantation where all anti-donor antibodies, or complement, were removed from the recipient, a delayed form of rejection: ‘acute vascular xenograft rejection’, occurs 2 - 5 days later, which causes a failure of the transplanted xeno-organ. VI.BIBLIOGRAPHY [39]

Hyperacute rejection is due to natural immunity. It is initiated by the specific binding of xeno-reactive natural antibodies, usually IgM, to blood vessels of the donor organ. But in some species combination the hyperacute rejection depends primarily upon the activation of the specific alternative complement pathway. VI.BIBLIOGRAPHY [39]

In a given species combination the hyperacute rejection occurs nearly every time, but not all the time. Lack of hyperacute rejection may be due to the variable level of expression of xeno-antigens, VI.BIBLIOGRAPHY [39], or due to occasional lack of action by complement, for whatever reasons. For example, organ xeno-transplant of liver appears to be resistant to hyperacute rejection for reasons unknown. VI.BIBLIOGRAPHY [31]

Besides liver, several tissues which require neovascularization by the recipient, such as skin grafts, pancreatic islets, etc., are resistant to hyperacute rejection, but not so bone marrow cells. VI.BIBLIOGRAPHY [31] Isolated islets from discordant species apparently do not suffer a high rate of hyperacute rejection either. VI.BIBLIOGRAPHY [37] Naturally occurring cytotoxic xenoreactive antibodies do not cause hyperacute rejection of the transplanted fetal porcine islet cell clusters. VI.BIBLIOGRAPHY [58] Porcine fetal pancreatic islet cells carry determinants for human natural antibodies but are neither sensitive to the direct cytotoxic effect of antibodies, nor are they sensitive to antibody- dependent cellular cyto-toxicity destruction mediated by natural antibodies, and such islet cells can reconstitute streptozotocine induced DM, indicating that fetal porcine islet-cell clusters can survive in vivo. VI.BIBLIOGRAPHY [12] Discordant cell xeno-transplants do not have a high rate of primary non-function, in any case not any higher than cell allo-transplants. VI.BIBLIOGRAPHY [35]

In neo-vascularized cell xeno-transplants, such as skin or pancreatic islets, the hyper-acute rejection is prevented by the absence of initial vascularization, which means a lack of endothelial cells. VI.BIBLIOGRAPHY [57, 59, 60, 150] Rejection of these tissues appears several days after implantation and is assumed to be mediated mainly by the cellular mechanism. VI.BIBLIOGRAPHY [62]

Even if the hyper-acute rejection would somehow be avoided, an ‘acute vascular xeno-transplant rejection’ takes place, for which the complement system is not necessary. Since the ‘acute vascular xeno-transplant rejection’ is unavoidable, the treatment appears futile and the best approach is prevention. VI.BIBLIOGRAPHY [39]

Sometimes, when anti-donor antibodies and / or complement are depleted from the recipient for a period of time before xeno-transplantation, acute vascular rejection does not occur, and the xeno-transplant continues to function even after antibodies and complement have been restored. An ‘accomodation’ has taken place. If accomodation could be achieved, the continuous immunosuppression of patients could be avoided. VI.BIBLIOGRAPHY [39, 33]

Micro-environment of liver, that includes phagocytosis of antigen by Kupffer cells, plays a unique role in the success of the induction of immune unresponsiveness to islet xeno-transplants by the pre-treatment with intra-hepatic transplantation of small number of islets VI.BIBLIOGRAPHY [3], or to cardiac graft by intra-portal transplantation of donor splenocytes. VI.BIBLIOGRAPHY [5] The same applies to thymus as intrathymic implantation of splenocytes with ALS leads to indefinite cardiac allograft survival, and prolonged xeno-transplant survival. VI.BIBLIOGRAPHY [44].

Adult islet allo-transplant rejection is dependent on the activity of CD8+ cells, with CD4+ acting as ‘helper’. VI.BIBLIOGRAPHY [59, 61] But CD4+ T cells alone play a pivotal role in the allo- transplant response to the purified islet tissue, xeno-transplant response, and expression of the autoimmune disease process. VI.BIBLIOGRAPHY [13]

Strong T cell immunity against allo-antigens is partially due to their similarities to self-antigens, since T cells are selected in the thymus for their affinity to self-antigens. Therefore, T cells might actually have weaker affinity for more discordant xeno-antigens. Other molecular interactions involved in allogeneic responses, besides receptor-antigen binding, may not operate efficiently when there are species differences. VI.BIBLIOGRAPHY [34]

Xeno-transplants are essentially always rejected faster than allo-transplants, if similar types of tissues are transplanted under similar circumstances. But, after the depletion of CD4+ T cells just the opposite happens: the xenogeneic reaction becomes weaker than allogeneic. VI.BIBLIOGRAPHY [31] The rejection of pig islet xeno-transplant is a CD4+ dependent process, with only a minor participation of CD8+ cells. VI.BIBLIOGRAPHY [11, 59, 61] In reality,helper and cytotoxic T cell responses to xeno-transplants are generally weaker, than to allo-transplants. VI.BIBLIOGRAPHY [31, 34] Complete rejection of islets across discordant species barrier is slower than across a closely related barrier and may be occurring by a different rejection process. VI.BIBLIOGRAPHY [1]

Reduction of immunogenicity of donor tissues prior to transplantation by elimination of donor-type hematopoietic APC's has been based on the theory that donor-derived APC's serve as major stimulators for triggering the rejection of tissue allo-transplants. Such treatment does not eliminate transplant antigen, however. The transplanted tissue retains its antigenic components, but the loss of APC’s eliminates tissue immunogenicity, that is, the capacity to activate an immune response in the host. One cannot assume that antigenicity, as the capacity to be recognized by the immune system, and immunogenicity, as the capacity to activate a response in the immune system, are regulated by the same process: antigen recognition. Antigenicity is involved in both these processes, indeed. However, immunogenicity requires the provision of a co-stimulatory signal in conjunction with antigen recognition by the specific immunocyte, and a co-stimulatory activity provided by active APC’s. VI.BIBLIOGRAPHY [170]

It appears that the capacity of APC's to stimulate T cells against xenogeneic cells is generally deficient: in vitro the T cell reactivity to xenogeneic APC's tends to decrease as the phylogenetic disparity increases between the stimulating APC and responding T cell, yet in vivo rat islet xeno-transplants are rejected more vigorously than islet allo-transplants. Thus the mechanism of islet allo-transplant and xeno-transplant rejection differ in their dependence on donor type APC. VI.BIBLIOGRAPHY [61]

In the xeno-transplant rejection, a progressive cellular infiltrate, consisting mainly of eosinophilic granulocytes and macrophages with only a small number of T lymphocytes was detected. VI.BIBLIOGRAPHY [64] Thus non-T cells or T cells lacking the conventional T-cell phenotype, but expressing CD4+ antigen, are the major cellular components mediating xenogeneic rejection. VI.BIBLIOGRAPHY [12] The xeno-transplant rejection is different from the allo-transplant rejection in that the major cellular components are macrophage-like CD4+ cells and not T cells VI.BIBLIOGRAPHY [57, 62]. The absence of immunoglobulins and/or complement depositions in the early rejection phase may indicate a pure cellular cytotoxic mechanism exerted by a macrophage-like cell. VI.BIBLIOGRAPHY [12] But, it is important to realize here, that macrophages should not be normally capable of activating T lymphocytes in the absence of microbial infection. VI.BIBLIOGRAPHY [165]

In vitro the interactions of CD4+ or CD8+ T cells, with their respective ligands on MHC proteins, may be weak when bound antigens are from a different and discordant species. Also, some lymphokines produced by stimulating lymphocytes of one species do not function well with receptors for these lymphokines expressed on T cells of another species. A theory was proposed whereby MHC bound xeno-antigens of the donor species may be so different in structure from the MHC bound xeno-antigens of the recipient species, that the T cell repertoire of the recipient will not contain receptors capable of directly recognizing those MHC bound xeno-antigens of the donor. And then, apparently some animal species are likely to elicit more defective human T-cell responses than others. VI.BIBLIOGRAPHY [21] All this is remarkable in view that the cell populations mediating xenogeneic responses, and the target antigens recognized by xeno-active T cells, are largely the same as those found in allogeneic responses. VI.BIBLIOGRAPHY [34]

Tolerance: The induction of ’tolerant state’, in which the recipient's immune system regards donor antigens as ‘self’ prior to fetal cell xeno-transplantation, would be essential for a full clinical success of this therapeutic method. VI.BIBLIOGRAPHY [150] There are three known mechanisms for induction of ‘tolerance’: anergy, suppression, and deletion. Tolerance among T cells could be either ‘central’, occurring at the time of T cell development in the thymus, or ‘peripheral’, occurring after T cell moved from the thymus to the periphery. VI.BIBLIOGRAPHY [30]

In the thymus there is a positive selection of thymocytes followed by a negative selection of self-reactive cells, that leads to the death of undesirable cells by apoptosis. The same can happen in the periphery. VI.BIBLIOGRAPHY [108]

MHC is important for ‘determinant selection’, the multistage process by which each individual's immune system selects the specific immunogens (and the specific epitopes from within complex immunogens) to which it will respond. Any given allelic form of MHC protein can bind only a finite range of peptides. Thus, an individual with a particular subset of class II genes may lack a class II MHC protein capable of binding a peptide from a particular foreign protein, so that no part of that protein can be presented to helper T cell, in which case no immune response would occur. VI.BIBLIOGRAPHY [108] Xenogeneic class II MHC protein/antigen complex is the principal target recognized by xeno-specific helper T cells. VI.BIBLIOGRAPHY [34] Immuno-alteration occurs either via a loss of class II MHC antigen cells, or via a loss of class I MHC antigen cells, which should be expected to directly stimulate CD8+ cell. There is much evidence that xeno-transplant rejection occurs via indirect presentation of antigens, and therefore the direct stimulation of the CD8+ cell by a class I MHC antigen would not be a major factor in xeno-transplant immunogenicity. VI.BIBLIOGRAPHY [37]

A lymphocyte is considered ‘anergic’ if an encounter with the antigen, that its clonally distributed surface receptor specifically recognizes as non-self, does not fully activate the cell. It can be induced in T cells, when antigen is presented without an adequate co-stimulation, as T cells require additional ‘co-stimulatory’ signal from APC's in addition to those provided by the MHC-antigenic peptide complex presented to the T cell receptor, as well as when the milieu of the regional lymph-node is lacking. The anergic T cells cannot produce IL-2, so that they cannot respond when they encounter an antigen, even if properly presented by APC. In view of the great phylogenetic disparity between humans and rabbits, it seems that many co-stimulatory, adhesive, and cytokine mediated interactions between the two species may be defective, resulting in greater ease of tolerance induction. VI.BIBLIOGRAPHY [30, 165]

In some systems studied, the delivery of ‘minimal signals’ results in ‘anergy’, or non-response of the T cells. Stimulation with the specific antigen, in the absence of other signals, can result in development of anergy (or non-response to that antigen on a normal antigen-presenting cell) at later times. Providing one signal to a macrophage, without other signals that are needed to activate that macrophage, results in a transcription of certain genes without their translation, that then makes those macrophages normally become non-responsive to the later delivery of the other signals. VI.BIBLIOGRAPHY [33]. Macrophages cannot generate specific immune response on their own, but have to be linked to the target cell by accessory factors, such as antibodies or complement. VI.BIBLIOGRAPHY [62]

‘Suppressive’ cell populations, or antibodies, may inhibit the responses of donor-reactive T or B cells. There are two subsets of CD4+ T cell with different function VI.BIBLIOGRAPHY [11]:TH1 subset, T-helpers with a type 1 response, that is pro-inflammatory, indirectly involved in the rejection of allo-transplants and xeno-transplants, producing IL-2, IFN-? and TFN-?; and TH2 subset, T-helpers with a type 2 response, that is tolerogenic, associated with allo-transplant and xeno-transplant acceptance, producing IL-4, IL-5, IL-6 and Il-10. VI.BIBLIOGRAPHY [30] While naive CD8+ cells emerging from thymus are predestined to become cytotoxic cells, the fate of CD4+ cells is decided only during their first encounter with the antigen: they become either inflammatory TH1 cells, leading to the cell-mediated immunity, or helper TH2 cells providing humoral immunity. VI.BIBLIOGRAPHY [165] Peripheral xenogeneic immune unresponsiveness induced by intraportal implantation of cultured rat islet cells in diabetic mice, is transferrable via splenocytes to a second diabetic recipient, and T lymphocytes are the mediators. These T lymphocytes appear to be anergic, and they suppress TH1 lymphocytes either by releasing IL-4 and IL-10, which suppression is reversible by IL-2, or alternatively by competing with them for ligands on the APC membrane. VI.BIBLIOGRAPHY [7]

Clonal ‘deletion’ of T or B cells with reactivity to donor is clinically the most promising. If there are no lymphocytes with reactivity to donor, then no specific response to donor antigens could be induced regardless of what immunologic stimuli would be encountered. T cell tolerance can be induced across allogeneic histocompatibility barriers by transplanting pluripotent hematopoietic stem cells, that induce intrathymic deletional tolerance, VI.BIBLIOGRAPHY [30] as well as across concordant xenogeneic barriers VI.BIBLIOGRAPHY [38]. Tolerance can be induced across concordant xenogeneic barrier also by intrahepatic or intrathymic islet transplantation together with anti-lymphocyte serum.VI.BIBLIOGRAPHY [8, 44] Pre-treatment by liver hematopoietic cells induced neonatal tolerance to cardiac organ allo-transplants, VI.BIBLIOGRAPHY [4] and the same applies to intrathymic splenocytes. VI.BIBLIOGRAPHY [43]

B cell tolerance could be achieved either by eliminating any cells that express an autoreactive antibody (clonal deletion), or by permitting such cells to survive in a functionally inactive state (clonal anergy). Clonal deletion takes place among early B lymphocytes in the marrow: contact of surface immunoglobulins with a self antigen at this stage transmits a signal that arrests further development of the early B cell and causes the subsequent death of the arrested cell. Several forms of clonal anergy have been observed among mature B cells in the periphery. A contact of surface immunoglobulins with a non-self antigen in the absence of appropriate CD4+ T cell, or when helper cell is anergic, may be crucial in initiating various forms of B cell anergy. At other times, the affected B cells fail to express surface immunoglobulins, or if they are expressed, they fail to transmit an effective signal to the cell upon binding the non-self antigen. VI.BIBLIOGRAPHY [108]

Immunologically incompetent embryos and neonates readily accept grafts from xenogeneic donors, which proves that cells of different species are not innately incompatible. On the contrary, in the absence of immune response, transplant and host cells of like histogenicity from widely divergent species show remarkable affinities. The ability to reject foreign transplants depends on the maturation of the ability to respond immunologically. VI.BIBLIOGRAPHY [32, 166]

A comprehensive picture of immunological research on cell xeno-transplantation, as presented here, is applicable to organ xeno-transplantation, but not to the BCRO fetal precursor cell xeno-transplantation when such cell xeno-transplants are prepared by described in this book method.

Experiments on avoidance of immunosuppression:

In 1965 insulin producing Langerhans islets were isolated in guinea pigs. VI.BIBLIOGRAPHY [123] Later on the same was accomplished with several other vertebrates, fetal, newborn and adult, and also with human fetuses. VI.BIBLIOGRAPHY [171, 176, 177, 178, 188] Lately an industrial method of isolation of islets from human adult cadavers was described.

Experiments with auto-, iso-, allo-, and xeno-transplantation of microfragments of fetal or adult pancreas, of isolated islets, either subjected to tissue culture or without culture, of cultured dispersed islet cells, have been carried out. Immunological reactions have been of major concern: they are minimal in case of auto- and iso-transplantation, but allegedly should be very serious with allo- and xeno-transplantation.

In mid-80-ies a theory of the rejection of cell allo- and xeno-transplants was presented, whereby class II MHC antigens carrying group of cells, so called ‘leukocytes- passengers’, were responsible for an induction of a rejection reaction. Dendritic cells, lymphocytes, macrophages and capillary endothelial cells, belong to this group. VI.BIBLIOGRAPHY [127, 170] Several experiments proved that the survival of cell allo- and xeno-transplants in the body of the recipient can be substantially prolonged if class II MHC antigens carrying cells are removed from the transplant. This can be accomplished by sufficiently long tissue culture of pancreatic islets, during which endocrine cells survive, while class II MHC antigen carrying cells die. VI.BIBLIOGRAPHY [122, 128]

Alteration, and/or elimination of ‘leukocytes-passengers’, is caused by tissue culture at 24 - 26 degree C VI.BIBLIOGRAPHY [1, 2], together with 95% O2 VI.BIBLIOGRAPHY [170], or with anti-lymphocytic serum, or with anti-thymocytic globulin, or with monoclonal antibodies against dendritic cells, or with immunotoxin, or with growth-transforming factor. VI.BIBLIOGRAPHY [122, 128, 36, 169, 174, 181] Tissue culture of pancreatic islets causes death of majority or all exocrine cells, and cessation of their enzymatic activity, while the islet cells proliferate, and differentiate, and thereby increase their hormone-producing capability. VI.BIBLIOGRAPHY [129]

Also UV light exposure, gamma irradiation, exposure to various chemicals, etc., have been used in an attempt to lower the antigenicity of cells and prolongation of the survival of the transplants. More recently diffuse capsules of semi-permeable membranes (alginate-polylysine, agar gel, etc.), permitting passing through of substances with molecular weight of less that 100,000 D, i.e. glucose or insulin, but blocking cell-destroying antibodies, have been used to encase islet cells. They protect islet cells for a few weeks or months, but ultimately lymphoid infiltration surrounds the capsules, followed by vascularized connective tissue, that becomes a complete barrier between implanted cells and the recipient. VI.BIBLIOGRAPHY [36]

New, less toxic and less dangerous to the recipient, methods of immunosuppression have been tried, such as treatment with anti-T cell monoclonal antibodies, or antilymphocytic serum VI.BIBLIOGRAPHY [1, 2, 3, 8,9], or pre-transplantation injection of donor-specific splenocytes. Also, pre-treatment plasmapheresis has been tested.

Let’s return yet again to the subject of ‘organospecificity’. The theoretical basis of fetal cell transplantation is the absence of antigenic differences of corresponding cells of humans and animals from different taxonomic groups, i.e. organospecificity. This is applicable to pancreatic islets, as well as all other organs and tissues. VI.BIBLIOGRAPHY [127] It appears that U.S. cell transplantologists came here to the same conclusion as pioneers of cell therapy 50 years earlier.

Antibodies reacting to the cell surface of viable rat islet cells are present in serum of children with IDDM. Human diabetic serums yielding positive immunofluorescence with rat islet cells were equally reactive with beta-cell suspensions from ob/ob mouse. In vitro physiologic characteristics of isolated islets from human pancreas seem to be similar to those of rats or mice. Thus organo-specific, non-species-specific, antibodies reactive with the cell surface of viable rat islet cells are present in serum from many children with IDDM, and the cell-surface immunoreactions caused by human diabetic serums are specific for islet cells of any species, i.e. organo-specific. VI.BIBLIOGRAPHY [116]

A typical characteristic of antibodies to the surface of islet cells is the organo-specificity, and due to this phenomenon such antibodies can be identified with the help of islet cells of not only human origin, but also from other animals, specifically rats and mice. Antibodies to the surface of islet cells do not cross-react with rat hepatocytes and splenocytes. There is significant physico/chemical and immunochemical similarity between the basic protein of an auto-antigen of islet cells of man and of rat. VI.BIBLIOGRAPHY [117]

Antibody assays and immuno-fluorescence studies of the membrane protein antigens of isolated hepatocytes from man, rat and rabbit demonstrate a complete organo=specificity of such surface antigens. VI.BIBLIOGRAPHY [118]

Rat islet cells were perfused with an immunoglobulin fraction of diabetic patients, and in every case there was an inhibition of the glucose-induced insulin release, which indicates the presence of antibodies interfering directly with rat beta-cell function. Immunoprecipitation of lysates of mouse pancreatic islets and rat insulinoma cells, and immunofluorescence of mouse and rat islet cells by sera of diabetic children support the fact, that islet cell autoantibodies are ‘cell-specific’, but ‘not species-specific’. VI.BIBLIOGRAPHY [119]