Studies on the survival of skin homografts in rabbits treated with alloxan
It is no exaggeration to say that surgery would enter into a new dimension of accomplishment if tissues or organs could be transplanted with the same freedom between two different individuals as between one part and another of a same individual.
The homotransplantation of skin is certainly one of the unsolved problems in medicine and numerous studies for this problem have been designed, conducted and reported. Among all varied homografts, the skin has become the primary choice for
investigation by more researchers because of its ease in operation and observation.
The present situation is that the all skin homografts in any part of the body must be presumed to be impermanent. However, clinically only two types of skin transplantations have been successful; skin homografts between monozygotic twins(Padgett, 1932; Brown, 1937; Schattner, 1944; Converse and Duchet, 1947;
McIndoe and Franceschetti, 1950; Blandford and Garcia, 1953; Cox and Fredricks, 1956), and skin homografts upon recipients with agammaglobulinemia (God and Varco, 1955; Varco et al., 1955; Good et al., 1957).
When the skin is transplanted as a homograft from one individual to another, the circulatory re-establishment of the homograft take place and its vessels become filled with blood from the host in 4 days after the transplantation,. The blood is
seen to circulate freely in the graft; the gross and microscopic appearance of the homograft is comparable to that of an autograft (Medawar, 1944, 1945; Edgerton et al., 1957; Converse and Rapaport, 1956). After a period of a little more than a week, however, the flow of blood in the vessels of the homograft ceases, the vessels become thrombosed, vascular permeability increase, the vessel walls become ruptured, the erythrocytes become extravasated into the perivascular tissue spaces of the graft, and finally the graft become necrotic and is rejected by the host (Scothorne and McGregor, 1953; Ohmori and Kurata, 1960). In this known rejection phenomenon of the homograft a time factor seems to play a role. A lapse of about 7 days usually occurs before the evidences of rejection, such as vascular thromboses
and necrosis occur. This 7 day period is comparable to the average time period during which the body builds up an effective antibody level against a foreign antigen.
The "second-set phenomenon" (Medawar, 1944) seems to be another factor in the immunological reaction. A second-set of skin homografts taken from the same donor and transplanted to the same host sloughs more rapidly than the first-set of homografts (Gibson and Medawar, 1943; Medawar, 1944, 1945, 1946; Woodruff and Simpson, 1955; Allgower et al., 1952; Lehrefeld et al., 1955; Rapaport and Converse, 1958). The shorter survival of second-set grafts may be attributed to an acquired immunity reaction previously developed in the host against the first-set of grafts.
Reserach prior to and subsequent to Medawar's basic studies tends to confirm his acquired immunity hypothesis. Clarification of the nature of this immunological reaction is one of the primary goals of homotransplantation research. Prolonged or
permanent survival of tissue homografts may be possible if this reaction can be prevented, altered, paralyzed or depressed by one or more methods.
Cannon (1957) has shown that approximately 5 to 10% of skin homografts taken from one-day-old chickens and transplanted to one-day-old chickens are permanently successful. The report of Peer et al. (1957) supports the findings of Cannon in that it suggests that neonatal or even young infant tissue apparantly lacks the absolute specific antigenicity characteristic of adult tissues. Billingham et al. (1955), Woodruff and Simpson (1955) have induced tolerance by the injection of cells from the prospective adult donor into newborn animals soon after birth, and found a prolonged or permanent survival of the skin homografts transplanted from the original donor when the animals had reached maturity. Hardin and Werder (1954,1955). Piomelli et al. (1961), using total body irradiation in young mice and rabbit, observed a prolongation of skin homograft survival for 2 to 7 weeks, as did Conway et al. (1955) who irradiated only the recipient site. Levinson and Nechels (1956) obtained a markedly prolonged survival of skin homografts in rats treated with nitrogen mustard. Stark et al. (1960) and Snyder (1964) observed a doubled peeriod of survival of skin homografts in rabbits flllowing regional lymphadenectomy and splenectomy. Billingham et al. (1951) and Woodruff and Llaurado (1956) have reported that adrenocortical hormones, administered either systemically or locally, brought about a significant prolongation of homograft survival in rabbits. Calnan and Kulatilake (1962), Ballantyne et al. (1962,1963) and Converse et al. (1963) have observed a 2 to 3 times longer survival over the control in massive skin homografts in rats. Snyderman et al. (1960) and Gardner et al.
(1961,1962) have experienced a prolonged survival of skin homografts in the advanced state of various malignant tumor patients, and the chemotherapeutic agents for carcinoma, such as nitrogen mustard, 6-mercaptopurine, cyclophosphomide, 5-flourouracil, methotrexate, thioguanine and azoserine have been proved effective in prolonging the survival of skin homografts in animals by Levinson and Necheles (1956), McLaren (1961) and Sutton et al. (1991, 1963). Schatten et al. (1958) reported 10.5 days survival (control 7.l days) of skin homografts in hypophysectomized and hypothyroid rats and stated that hypometabolism of the host cell may result in a less intense antibody response. Heslop et al. (1954) have showed 50 to 100% longer survival of skin homografts than that of control in pregnant rabbits, and Dammin et al. (1957) in uremic patients, and Smiddy et al. (1960) in uremic rabbits, have showed a significant prolongation of survival of the skin homografts. Besides the above mentioned, the skin homografts in animal experiments treated with antihistaminics (Forster and Hanrahan, 1948; Conway et al., 1954), anticoagulants (Conway et al., 1953), sodium salicylate (Conway et al., 1955), and RNA (Ashley et al., 1960) have shown also a significant prolongation of their survival.
In light of above review of accumulated knowledge of the problem of homotransplantation of skin, an investigation of the effect of hyperglycemia was undertaken, in order to clarify a clinical impression of a prolongation of survival time of skin homografts on a severely burned diabetic patient.
Materials and Methods
Five group experiments were performed. In all the donors were adult male rabbits and the hosts were laboratory bred, unrelated, adult albino rabbits (1800-2100 Gm), both males and females. The graft survival times were determined by hardening and discoloration on gross examination and simultaneous serial biopsy. Throughout the experiment periodic determinations of the blood sugar level (Folin-Wu method) were carried out. The following description of grafting applies to all of the experimental groups. Each host received 4-6 homografts (2x2 cm) of full thickness skin from a single donor, and one autograft. All grafts were stured in place on freshly prepared hostbeds made by removing skin down to the panniculus carnosus muscle over dorsum of trunk. For 24 hours slight pressure dressings were applied with 3-4 turns of elastic bandage, and they were then removed and daily inspection of the grafts was undertaken. All the host animals in this experiment were kept in standard type individual cages.
1. Total of 40 homografts on 10 normal rabbits (control group), blood sugar ranging between 100 and 120mg%, showed a mean survival of 7.04±0.26 days. All auto grafts performed in each group were successful and to be considered as permanent survival.
2. Alloxan 40mg/kg was injected intravenously to each of 5 rabbits and a permanent hyperglycemic state was confirmed. Blood sugar of 160 mg% or more in 24 and 48 hours was maintained following injection. A total of 20 homografts on these 5 hyperglycemic rabbits showed a mean survival of 11.25±0.44 days, and this is about 60% prolongation of the survival comparing with the control group.
Additional 28 homografts on 7 hyperglycemic rabbits, 3 with average blood sugar of 200mg% after 80mg/kg alloxan injection, and 4 with average blood sugar of 300mg% after 120mg/kg alloxan, showed mean survivals of 13.08±0.22 days and 18.63±2.02 days respectively. This demonstrated a definite prolongation of sruvival of the homografts and the survival rate seemed to be related with the blood sugar level.
3. Protamine-zinc insulin 1 unit per kg. of body weight was injected intramuscularly to 7 rabbits daily from 5 days prior to grafting and until the day of complete rejection of all homografts. Their blood sugar levels were between 50 and 80 mg%. Total of 38 homografts on these 7 hypoglycemic rabbits showed
approximatly 14% shorter survival time than the control.
4. A total of 30 homografts on 5 alloxan diabetic rabbits which were treated with daily insulin injection ( 1 unit/kg PZI) demonstrated no significant changes in survival comparing with that of the control.
5. A total of 7 rabbits were fed 15-20 gm/kg of sugar daily in divided dose, in addition to the regular laboratory diet. The feeding was started 10 days prior to grafting and ended when experiments were completed. Total of 34 homografts on the 7
sugar fed rabbits (blood sugar maintained about 180 mg%) show an average survival of 10.24±0.48 days, and statistically, this was a significant prolongation of survival.
To summarize, there was a definite prolongation in survival of the skin homografts made in alloxan hyperglycemic rabbits, and the time of survival was clearly related to the severity of the hyperglycemia. Also it is suggested that the prolonged survival of skin homografts was not due to a certain specific action of
the alloxan which might directly effect the homografts, but rather this was closely related to the blood sugar level. Much more remains to be investigated along these lines.