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Our focus here is on organotropic effect of cell transplants, i.e. the stimulation of the same organ of the recipient by implanted cell transplant, heart by cardiomyoblasts, kidney by epithelial cells of nephron, liver by hepatocytes, etc., as was postulated by Paracelsus already at the beginning of 16th century.

The direct stimulation by fetal cell transplantation is recognizable as an increase of cell count of growing organs or tissues, intensification of metabolism, or of a certain specific function, such as accentuation of anabolism. Such direct stimulation can be triggered only by implanted cell transplant of the corresponding organ, but not by transplantation of other cell types.

Since it is rare that only an individual organ is malfunctioning, on the contrary usually the whole organ systems are diseased, it is necessary to treat all involved organs by corresponding cell transplants. The list of cell transplants necessary for treatment of each patient has to be individually designed in terms of cell composition, dosage, date of implantation.

In 1916 Murphy and Danchakoff incubated chicken’s eggs for 7 days, then opened the shells, and placed a small piece of spleen, liver, bone marrow, kidney, on the chorio-allantoic membrane, then re-sealed the egg and continued the incubation. On the 17th day the shells were re-opened and spleens were found to be markedly enlarged, kidneys moderately enlarged, while liver not at all. Subsequently an implantation of tissues of peripheral muscles, bone, cartilage and sarcoma, were done in the same way but no change of size of spleen was observed. They observed the presence of an effect that was limited to the tissue of the corresponding organs, i.e. organospecificity. This study was repeated in 1956. Implantation of spleen pulp caused a considerable progress in tissue differentiation of embryonic spleen, while application of gelatin, or a drop of milk, inhibited a differentiation, and implantation of splenic homogenate, or of splenic tissue subjected to 60º heat for one hour, had no effect whatsoever, thus proving that effect of an accelerated development of spleen was dependent upon an implantation of intact live splenic cells. VI.BIBLIOGRAPHY [324]

Subsequently P. Weiss and A.C. Taylor carried out classical experiments on the in-vivo influence of homologous organ pulp on the growth rate of embryonic organs. VI.BIBLIOGRAPHY [325]

In 1909 Halsted succeeded in transplanting parathyroid glands from one dog into the spleen or muscles of another dog, but only after he had previously produced a hypofunction, and therefore a ‘need’ in the recipient animal, by a partial removal of the parathyroid glands before the transplantation. The more extensive damage of an organ or tissue, the higher proportion of the transplanted cells ‘home’ into the same organ or tissue. This has been known in cell therapy as ‘Halsted principle’. VI.BIBLIOGRAPHY [143]

Demonstration of the organ-specific effect of a cell transplant required a proper experimental set-up that included an induced damage of the recipient organ. Eventually the organospecificity was proven by multiple experiments in 50- and 60-ies with liver (Walter, Allman, Mahler; Marshak, Walker), heart (Tumanishvili, Jandieri, Svanidze; Walter, Allman, Mahler), lacrimal glands (Teir), skin (McJunkin, Matsui), thymus (Roberts, White), Langerhans islets of pancreas (DuBois and Gonet).

Kalb demonstrated organ-specific stimulating factors in the blood serum of rats 4 to 5 days after implantations of liver tissue.

All described effects were independent of the species used for an experiment, but the results were always limited to homologous organs. VI.BIBLIOGRAPHY [326]

In clinical practice it was the field of endocrinology where the organospecificity was recognized first. Already in 1771 Hunter and then in 1849 Berthold demonstrated the organotropic effect of implanted testis in the castrated cock. Then in 1889 Brown-Sequard reported to the French Academy of Sciences about self-treatment by extract prepared by himself from rooster testes. In 1906 Morris transplanted human ovaries into a castrated woman, and menstruation re-started 4 months later. In 1912 Herrmann and Sutten transplanted the testis from a human cadaver to a castrated male. In 1925 Comolli removed one parathyroid gland during thyroidectomy and transplanted it to another patient suffering from tetany. In 1931 Dolby transplanted pancreas from still-born babies into diabetic patients.

In 1930’s the interest in glandular transplantation waned because of rapid development of hormone therapy. Soon the drawbacks of hormonal therapy were recognized, i.e. that the effect of hormones is only temporary, and over-dosage, or continuous application, cause trouble because of atrophy of the gland from inactivity.

In 1927 Niehans began transplantation of the anterior lobe of the pituitary from calves to the patients with nanismus, with the height increase up to 32 cm. In 1928 he transplanted the anterior lobe of pituitary from sheep to women with primary amenorrhea with good to a very good results. In 1929 he began to transplant the posterior lobe, and a stalk of pituitary, into the patients with diabetes insipidus, and adrenal glands into patients with polyarthritis. And then in 1931 came the famous case when he saved the life of a female patient with a post-thyroidectomy tetany by the first cell transplantation of minced tissue of parathyroid.

It was confirmed that a hormonal effect lasted only a short time, and the real effect of cell transplantation did not take place until some weeks later, that was due to something else, not hormones. The insufficiency of an endocrine gland could be eliminated only by an implantation of tissues from the same gland: a tetany due to the removal of parathyroids responded only to the implantation of parathyroids, i.e. following the rules of organospecificity.

This was tested in experimental animals with thyroid gland insufficiency (Goos, Sturm, Kment ), anterior lobe of pituitary deficiency (Maischein). But animal models of human hypo-endocrinopathies were not adequate, as they did not correspond to human illnesses. So human testing began in those endocrine diseases, where the hormone production could be tested by urinary excretion of various fractions of 17-keto-steroids and corticoids: pituitary gland, sex glands, adrenal cortex. Implantation of tissue fragments of any of the three endocrine glands caused a dramatic increase in the excretion of the respective fractions of 17-ketosteroids, with a maximum after 8 to 14 days, and then gradual disappearance of the effect during several months. The implantation of any other non-endocrine tissues caused no change in 17-keto-steroid excretion.

Implantation of placenta caused an increase of all 17-keto-steroid fractions to levels that exceeded even those after an implantation of anterior lobe of pituitary. Schubert showed a change of vaginal epithelium from an atrophic state to normal in post-menopausal women after an implantation of placenta that, due to a lyophilization, was free of any estrogen or gonadotropin.

Niehans stressed the implantation of hypothalamus in the treatment of diseases with disturbed neurosecretory regulations. If transplants of anterior lobe of pituitary were implanted after the hypothalamic cells, a marked increase of excretion of all fractions of 17-keto-steroids was observed. VI.BIBLIOGRAPHY [327]

Cell transplantation is a vastly different approach to medical treatment and cannot be immediately understood by the mind accustomed to deal with (chemical) drug therapy.

The therapeutic effect of drugs of chemical origin is not as broad as those of any of the 200+ known types of cells transplanted into a diseased body with insufficient quantity or quality of a particular cell type(s).

Compare any publication about the treatment of diabetes with insulin, or oral anti-diabetics, with those discussing various aspects of treatment of diabetes mellitus by fetal precursor cell transplantation.

For example the effect of different site of implantation of fetal precursor cell transplants on the course of experimental diabetes was tested in the search for a way to decrease the immune response to cell transplantation by implanting cell transplants into various ‘immunologically privileged’ regions: intrahepatic, intrasplenic, intraportal, intrapancreatic, sub-capsular (kidney), intraperitoneal, intraomental, intramuscular, intracerebral, intra-spinal cord, intraocular (anterior chamber), intratesticular, intrathymic, etc. VI.BIBLIOGRAPHY [37] The immunoprivileged areas are unusual in two ways:

1/ the communication between the privileged site and the body is atypical in that the extracellular fluid in these sites does not pass through lymphatics, and

2/ certain humoral factors affecting immune system are produced in the privileged sites, such as TGF, which leaves the sites along with antigens, and induces TH1 rather than TH2 responses. So immunoprivileged sites do not prevent the interaction of antigens with T cells, but instead of eliciting a destructive immune reaction, such interaction induces tolerance. VI.BIBLIOGRAPHY [165]

Another issue is that after transplantation of fetal islet cells their function is slow to develop, because further growth and differentiation of endocrine tissue is necessary for the therapeutic effect to take place. About 2 - 3 months has to pass for that to happen. Compare it with the transplanted adult islets, that reverses diabetes in 1 - 2 weeks. VI.BIBLIOGRAPHY [170, 181]

Concept of critical mass of the transplanted endocrine tissue of islets necessary to accomplish insulin-independence represented an enormous step forward. VI.BIBLIOGRAPHY [130] When a sufficient Beta-cell mass was transplanted, the subsequent replication of Beta-cells was normal, and the transplanted Beta-cell mass remained unchanged. If an insufficient Beta-cell mass was transplanted, a limitation in Beta-cell replication was found after transplantation, and the Beta-cell mass of the graft declined progressively.

When normoglycemia was restored by transplantation, the replication of transplanted Beta-cells was similar to that of pancreatic Beta-cells of normal animals. When islet transplants were sufficient to restore normoglycemia, the Beta-cell mass of the graft on days 10 and 30 post-transplantation was similar to the originally transplanted Beta-cell mass, and on day 30 the grafted Beta-cells had an increased size when compared to other transplanted groups.

When an insufficient islet tissue was transplanted, an initial increase in replication was found after 10 days of hyperglycemia, however, after 18 and 30 days of severe hyperglycemia, this increased replication was not maintained, that was an indication that the capacity of transplanted Beta-cells to grow was limited.

In chronically hyperglycemic animals, despite the increased Beta-cell replication by day 10, there was a continuous decline in Beta-cell mass of the grafts: increased Beta-cell destruction ‘over-powered’ the Beta-cell replicative capacity. After 30 days, Beta-cell mass was significantly lower, and despite the persistent hyperglycemia the replication was no longer increased. A significant portion of the fall in Beta-cell mass was due to increased cell death.

When hyperglycemia persists after transplantation of an insufficient Beta-cell mass, both limited Beta-cell replication and accelerated cell death are found, leading to a progressive reduction in Beta-cell mass and failure of transplant. In contrast, when sufficient Beta-cell mass is transplanted, a balance among Beta-cell replication, Beta-cell hypertrophy, and Beta-cell death is obtained, and Beta-cell mass remains reasonably constant.

In normal recipients the transplanted Beta-cell mass dramatically diminished at days 10 and 30, despite normal replication of Bet-cells. This reduction was confirmed by a similar decrease in the insulin content of the graft. This was a result of down-regulation in order to protect from the effect of excessive Beta-cell mass causing hypoglycemia. VI.BIBLIOGRAPHY [47]

Thus a critical islet mass, higher than predicted, must be transplanted to achieve normoglycemia, and late failures depend on the number of initially transplanted islets. Islets exposed to sustained hyperglycemia have shown an impairment of Beta-cell function, limited replicative response and reduced Beta-cell mass.

Outcome of islet cell transplantation was improved when islets were transplanted to insulin treated diabetic mice with long-term normoglycemia and normal glucose tolerance: in such case transplanted Beta-cells maintained a normal insulin content. Also, insulin treatment reduced significantly the Beta-cell mass needed to cure DM in transplanted mice: only 10 - 30% of normal islet tissue was necessary to maintain normoglycemia, as compared when normoglycemia had to be restored in hyperglycaemic mice not treated by insulin.

When insulin implants were removed 14 days after transplantation, transplanted islets were already vascularized and able to sense and respond more appropriately to changes of blood glucose. Islets transplanted to insulin-treated mice were able to increase their Beta-cell mass when insulin was withdrawn; this capacity of Beta-cell mass to adopt to changes in functional demand is essential to maintain normoglycemia. On day 60 the transplanted Beta-cell mass was similar to the initially transplanted mass. - Even in those non-insulin-treated mice that eventually achieved normoglycaemia, there was an abnormal glucose tolerance, suggesting that Beta-cell function was impaired in the islets that had been previously exposed to sustained hyperglycemia. VI.BIBLIOGRAPHY [45]

The functioning of transplanted pancreatic islets was proved by the fact, that after removal of an organ into which islets were transplanted, the hyperglycemia, glycosuria and other symptoms of diabetes mellitus, recurred. VI.BIBLIOGRAPHY [134]

Timing of fetal precursor cell transplantation is extremely important: the sooner after the onset of disease it is carried out, the better are the results. When cultured islet cells were implanted under the kidney capsule of syngeneic mice already 3 weeks after the induction of diabetes mellitus, the capillaries of retina and kidneys maintained normal thickness of basal membrane, however, when implantation was delayed 7 months after the induction of diabetes, the basal membrane of glomerular capillaries became thicker, although the basal membrane of retinal capillaries maintained normal thickness. VI.BIBLIOGRAPHY [133]

Morphometric study of kidneys before and 4 months after transplantation showed decreased percentage of damaged glomeruli, widening of lumen of glomerular capillaries, and decrease of mesangial accumulation of class C immunoglobulins and complement. VI.BIBLIOGRAPHY [132] In long-term diabetic rats with nephropathy, 6 weeks after the transplantation of islets the phagocytosis and clearance capacity of the mesangial cells was restored and the mesangial enlargement reversed. VI.BIBLIOGRAPHY [172] In rats with induced diabetes mellitus the transplantation of islets reduced within two weeks significantly the mesangial thickening and mesangial staining for IgG, IgM, and complement C3. VI.BIBLIOGRAPHY [173]