Infusion of donor hepatic non-parenchymal cells prolongs survival of skin allografts in mice:role of microchimerism and IL-4

BACKGROUND: In the mouse skin allograft model, specific immune tolerance to the donor was induced by injection of donor hepatic non-parenchymal cells (NPCs). This markedly prolonged the survival time of the allograft. The mechanism of the induction of immune tolerance with donor hepatic NPCs is thought to be related to microchimerism and the IL-4 level. This work aimed at exploring the way of inducing immune tolerance and understanding the mechanism. METHODS: C57BL/6 (B6) mice were primed by intravenous injection of 2 X10(7) NPCs from C3H mice. Cyclophosphamide (200 mg/kg) was injected intraperitoneally 48 hours later. Eighteen days after the NPC injection, skin from C3H mice was transplanted to B6 mice and the survival of the grafts was assessed. The immune reaction of splenocytes from the treated B6 mice to donor-specific T-cells was measured by H-3-TdR incorporation. Microchimerism in the spleen was determined by flow cytometric analysis sytem (FCAS) analysis, and the serum level of IL-4 was assayed by ELISA at designed times. RESULTS: The survival time of the skin graft was markedly prolonged from 10 days to 70 days in controls. Microchimerism. in the spleen was found as early as day I post-NPC injection, then it increased steadily, and there was a positive relationship between graft survival and the quantity of microchimerism. The ELISA results showed that NPC infusion enhanced IL-4 production, especially in the mice with longer graft survival. CONCLUSION: Donor NPC infusion pre-transplant can prolong the survival of the skin graft and microchimerism and high levels of IL-4 may be involved.

Identification of the miniature pig inbred line by skin allograft

Skin grafting has been used as one of the most reliable tests to determine the genetic stability of laboratory animal such as mice and rats inbred line, but no identification of swine inbred lines by skin grafting has been reported. At present, Wuzhishan miniature pig （WZSP） inbred line has acquired the F24 individuals in China. In order to verify whether WZSP inbred line had D^en cultivated successfully, allogeneic skin grafts and related research were performed on F20 individuals of WZSP inbreeding population, compared with a control group of autologous transplantation. We observed the transplant recipients＇ wounds, detected peripheral blood-related indicators interleukin-2, 4 and 10, CD4~ and CD8~ lymphocytes, and conducted hematoxylin-eosin （HE） and Masson＇s staining of skin to judge whether the immune rejection reactions occurred within 28 days after transplantation. Chr. 7 genomic heterozygosity of 48 WZSP individuals from F20 to F22 was analyzed by high-density single nucleotide polymorphism （SNP） chips （60 000 SNPs）. The result showed that there were no significant differences in graft skin, the plasma interleukin-2, 4, 10, CD4~ and CD8~, HE and Masson＇s staining results between the allograft and autograft groups, and no immune rejection occurred on the allograft group. We found that 11 genes in Chr. 7 of major histocompatibility complex （MHC） I and MHC II were homozygous which confirmed that immune antibody of the allograft and autograft groups were highly identical and also provided a theoretical basis to no immune rejection occurred on the allograft in the inbred WZSP. The result proved that the WZSP inbred line had been cultivated successfully for the first time in the world. The test methods also provide a scientific basis for the identification of swine and mammal inbred lines.

Effects of combined radiation and thermal burn injury on the survival of skin allograft and immune function in rats

Abstract Objectives To investigate the effects of combined radiation and thermal burn injury on the survival of skin allografts and to analyze the relationship between the prolongation of allograft survival and the changes of immune functions of the thymocytes and splenocytes in rats. Methods Wistar rats were irradiated with 3, 4, 5, 6 and 8 Gy of gamma rays. Thirty minutes after radiation, 15% TBSA Ⅲ degree burn was inflicted to the rats. Twenty four hours after the burn injury, allografts were used to cover the burn wounds. In the 8 Gy group, 1 hour before skin grafting, the bone marrow cells (4×10 8) from the same donor were also transplanted. All rats were carefully observed after injury. The rats with single radiation injury of 5 Gy gamma rays, with single burn injury and with combined radiation burn injury were killed 3, 7, 10, 15 and 30 days after skin grafting for immunological assay and pathological study. Results All the allografts in the single burn group were rejected in 10 days. In the combined injury groups, the survival rates of the allografts in rats undergoing 3 and 4 Gy radiation were 20% and 30%, respectively. In the combined injury groups undergoing 5, 6 and 8 Gy radiation, the 10 day survival rates of the allografts were 69%, 88% and 100% respectively, and the 30 day survival rates in the three groups were 36%, 42% and 100% separately. The grafted allogenic skin, with normal epithelial cells and good vascularity, healed well with the recipient's skin. Hairs grew well from the allografts 30 days after grafting. Three, 7 and 15 days after allografting, in the single burn group, the proliferative activities of the thymocytes were 90%, 185% and 130% of the preinjury level, and the antibody forming capacities of the splenocytes were 200%, 171% and 300% of the preinjury level, respectively; in the combined injury groups, the proliferative activities were 6%, 99% and 91% of the preinjury level, and the forming capacities were 2%, 36% and 90% of the preinjury level. Conclusions The survival rate of allograft in rats undergoing combined radiation and thermal burn injury rises with the increase in radiation dosage. The allograft covering single bun injury is severely rejected by immune reaction. The prolongation of the survival of allograft in combined injury group mainly results from radiation that suppresses immune functions.

A Review of Skin Banking Guidelines and Standards Worldwide:Towards the Harmonization of Guidelines for Skin Banking in Therapeutic Applications for the Regions under the Asia Pacific Burn Association(APBA)

Currently,there are no harmonized guidelines which govern skin banking in the Asia Pacific region.Therefore,skin banks are either unregulated or rely on their nation’s legislation or international accreditation to uphold their quality standards.A new set of skin banking guidelines was devel-oped through a comprehensive review and collation of best international practices for the Asia Pacific Burn Association(APBA)members,from donor screening and testing,to skin recovery,processing,storage and distribution,and quality assurance.National regulatory requirements reviewed include the European directives,Australia’s Therapeutic Goods Administration and Sin-gapore’s tissue banking standards.Further technical and quality management recommendations are referenced from the American Association of Tissue Banks(AATB),the United States Food and Drug Administration standards and guidance documents,various relevant European guides,Japanese Society of Tissue Transplantation guidelines and the Asia Pacific Association of Surgical Tissue Banking.Adapted mainly from the AATB standards,the new Asia Pacific Burn Association Guidelines for Skin Banking in Therapeutic Applications offer a comprehensive manual,address-ing:governance and contracts;staff responsibilities;quality management;facilities,equipment and supplies management;donor consent and testing;and recommendations of good practices pertaining to skin recovery,processing,storage and distribution.Besides complementing current generic regulations,they provide technical specifications of major aspects unaddressed in most legislations.This inaugural set of new regional skin banking guidelines would be a start for regional members of the APBA to adopt,and will hopefully culminate in a set of standards so that,in the long run,skin allografts from this region can be of similar quality,which can simplify import process and facilitate the exchange of allografts between members.