BACKGROUND: The primary limitation to kidney transplantation is organ shortage. Recent progress in gene editing and immunosuppressive regimens has made xenotransplantation with porcine organs a possibility. However, evidence in pig-to-human xenotransplantation remains scarce, and antibody-mediated rejection (AMR) is a major obstacle to clinical applications of xenotransplantation. METHODS: We conducted a kidney xenotransplantation in a brain-dead human recipient using a porcine kidney with five gene edits (5GE) on March 25, 2024 at Xijing Hospital, China. Clinical-grade immunosuppressive regimens were employed, and the observation period lasted 22 days. We collected and analyzed the xenograft function, ultrasound findings, sequential protocol biopsies, and immune surveillance of the recipient during the observation. RESULTS: The combination of 5GE in the porcine kidney and clinical-grade immunosuppressive regimens prevented hyperacute rejection. The xenograft kidney underwent delayed graft function in the first week, but urine output increased later and the single xenograft kidney maintained electrolyte and pH homeostasis from postoperative day (POD) 12 to 19. We observed AMR at 24 h post-transplantation, due to the presence of pre-existing anti-porcine antibodies and cytotoxicity before transplantation; this AMR persisted throughout the observation period. Plasma exchange and intravenous immunoglobulin treatment mitigated the AMR. We observed activation of latent porcine cytomegalovirus toward the end of the study, which might have contributed to coagulation disorder in the recipient. CONCLUSIONS 5GE and clinical-grade immunosuppressive regimens were sufficient to prevent hyperacute rejection during pig-to-human kidney xenotransplantation. Pre-existing anti-porcine antibodies predisposed the xenograft to AMR. Plasma exchange and intravenous immunoglobulin were safe and effective in the treatment of AMR after kidney xenotransplantation.
Transplantation, Heterologous/methods* ; Kidney Transplantation/methods* ; Heterografts/pathology* ; Immunoglobulins, Intravenous/administration & dosage* ; Graft Survival/immunology* ; Humans ; Animals ; Sus scrofa ; Graft Rejection/prevention & control* ; Kidney/pathology* ; Gene Editing ; Species Specificity ; Immunosuppression Therapy/methods* ; Plasma Exchange ; Brain Death ; Biopsy ; Male ; Aged

OBJECTIVE: To prepare decellularized nerve grafts from alpha-1, 3-galactosyltransferase (GGTA1) gene-edited pigs and explore their biocompatibility for xenotransplantation. METHODS: The sciatic nerves from wild-type pigs and GGTA1 gene-edited pigs were obtained and underwent decellularization. The alpha-galactosidase (α-gal) content in the sciatic nerves of GGTA1 gene-edited pigs was detected by using IB4 fluorescence staining and ELISA method to verify the knockout status of the GGTA1 gene, and using human sciatic nerve as a control. HE staining and scanning electron microscopy observation were used to observe the structure of the nerve samples. Immunofluorescence staining and DNA content determination were used to evaluate the degree of decellularization of the nerve samples. Fourteen nude mice were taken, and subcutaneous capsules were prepared on both sides of the spine. Decellularized nerve samples of wild-type pigs ( n=7) and GGTA1 gene-edited pigs ( n=7) were randomly implanted in the subcutaneous capsules. Blood was drawn at 1, 3, 5, and 7 days after implantation to detect neutrophil counting. RESULTS: IB4 fluorescence staining and ELISA detection showed that GGTA1 gene was successfully knocked out in the nerves of GGTA1 gene-edited pigs. HE staining showed that the structure of the decellularized nerve from GGTA1 gene-edited pigs was well preserved; the nerve basement membrane tube structure was visible under scanning electron microscopy; no cell nuclei was observed, and the extracellular matrix components was retained in the nerve grafts by immunofluorescence staining; and the DNA content was significantly reduced when compared with the normal nerves ( P<0.05). In vivo experiments showed that the number of neutrophils in the two groups were similar at 1, 3, and 7 days after implantation, with no significant difference ( P>0.05); only at 5 days, the number of neutrophils was significantly lower in the GGTA1 gene-edited pigs than in the wild-type pigs ( P<0.05). CONCLUSION The decellularized nerve grafts from GGTA1 gene-edited pigs have well-preserved nerve structure, complete decellularization, retain the natural nerve basement membrane tube structure and components, and low immune response after xenotransplantation through in vitro experiments.
Animals ; Transplantation, Heterologous ; Galactosyltransferases/genetics* ; Sciatic Nerve/immunology* ; Swine ; Tissue Engineering/methods* ; Humans ; Graft Rejection/prevention & control* ; Gene Editing ; Mice ; Mice, Nude ; Heterografts/immunology* ; Animals, Genetically Modified ; Tissue Scaffolds ; Decellularized Extracellular Matrix

OBJECTIVE: To evaluate the immunogenicity of self-made sheep skin acellular dermal matrix (ADM).
 METHODS: Both cross-linked and non-crosslinked ADMs were made after the sheep skin ADM was ready. A total of 160 healthy mice were then randomly divided into 4 groups. Group A, cross-linked ADM was implanted under skin; Group B, non-crosslinked ADM was implanted under skin; Group C, allosome skin was embedded under skin; Control group, without any implantation. Any visible changes in implantation region and any apparent rejection was recorded. Any symptoms of inflammatory reaction in implantation region were also observed by microscope. The levels of IL-4 and IFN-γ in serum were measured on the 3rd day, 1 week, 2 weeks and 3 weeks after the operation, respectively.
 RESULTS: Visual study: wound healing in implantation regions were obvious, the immunologic rejection was not significant, and the sheep ADM was absorbed gradually by ambient tissues. Microscope results: the connection between the sheep ADM and surrounding tissues was very tight with infiltration of a few inflammatory cells into the depth of sheep ADM. The structure of sheep ADM was gradually destroyed by inflammatory cells. The levels of IL-4 and IFN-γ were not significant difference between the group of A, B or C and the control group on the 3rd day after the operation. The inflammatory reaction in implantation region of the control group degraded after 3 days, concomitant with a decrease in the levels of IL-4 and IFN-γ. But the levels of IL-4 and IFN-γ in the Group A, B or C remained at high level, suggesting the elevation of IL-4 and IFN-γ levels were related to the immunologic reaction by sheep ADM. The values of IL-4/IFN-γ were significant difference between the group A, B or C and the control group at the end of the 1st week, 2nd week, and 3rd week. However, there were not significantly different among the Group A, B and C.
 CONCLUSION Sheep skin ADMs do not cause apparent rejection from mice after the transplantation because of the low immunogenicity. The cross-linked and non-crosslinked ADMs do not obviously affect the levels of IL-4 and IFN-γ.
Acellular Dermis ; Animals ; Graft Rejection ; blood ; Heterografts ; immunology ; Interferon-gamma ; blood ; Interleukin-4 ; blood ; Mice ; Random Allocation ; Sheep ; Skin Transplantation ; Wound Healing

The development of highly immunodeficient mouse strains has allowed the reconstitution of functional human immune system components in mice. New-generation humanized mice generated in this manner have been extensively used for modeling viral infections that are exclusively human tropic. Epstein-Barr virus (EBV)-infected humanized mice reproduce cardinal features of EBV-associated B-cell lymphoproliferative disease and EBV-associated hemophagocytic lymphohistiocytosis (HLH). Erosive arthritis morphologically resembling rheumatoid arthritis (RA) has also been recapitulated in these mice. Low-dose EBV infection of humanized mice results in asymptomatic, persistent infection. Innate immune responses involving natural killer cells, EBV-specific adaptive T-cell responses restricted by human major histocompatibility and EBV-specific antibody responses are also elicited in humanized mice. EBV-associated T-/natural killer cell lymphoproliferative disease, by contrast, can be reproduced in a distinct mouse xenograft model. In this review, recent findings on the recapitulation of human EBV infection and pathogenesis in these mouse models, as well as their application to preclinical studies of experimental anti-EBV therapies, are described.
Animals ; Disease Models, Animal ; Epstein-Barr Virus Infections/complications/immunology/*virology ; Herpesvirus 4, Human/*physiology ; Heterografts ; Humans ; Killer Cells, Natural/pathology/virology ; Lymphoproliferative Disorders/etiology ; Mice ; Mice, SCID ; T-Lymphocytes/pathology/virology

Graft-versus-host disease (GVHD) is a prevalent and potential complication of hematopoietic stem cell transplantation. An animal model, xenogeneic GVHD (X-GVHD), that mimics accurately the clinical presentation of GVHD would provide a tool for investigating the mechanism involved in disease pathogenesis. Murine models indicated that inhibiting IL-21 signaling was a good therapy to reduce GVHD by impairing T cell functions. We sought to investigate the effect of exogenous human IL-21 on the process of X-GVHD. In this study, human IL-21 was expressed by hydrodynamic gene delivery in BALB/c-Rag2⁻/⁻ IL-2RΓc⁻/⁻ (BRG) immunodeficient mice which were intravenously transplanted human peripheral blood mononuclear cells (hPBMCs). We found that human IL-21 exacerbated X-GVHD and resulted in rapid fatality. As early as 6 days after hPBMCs transplanted to BRG mice, a marked expansion of human CD19⁺ B cells, but not T cells, was observed in spleen of IL-21-treated mice. Compared with control group, IL-21 induced robust immunoglobulin secretion, which was accompanied by increased accumulation of CD19⁺ CD38(high) plasma cells in spleen. In addition, we demonstrated that B-cell depletion was able to ameliorate X-GVHD. These results are the first to find in vivo expansion and differentiation of human B cells in response to IL-21, and reveal a correlation between the expansion of B cells and the exacerbation of xenogeneic GVHD. Our findings show evidence of the involvement of B cells in X-GVHD and may have implications in the treatment of the disease.
Animals ; B-Lymphocytes ; immunology ; metabolism ; pathology ; Cell Differentiation ; Cell Proliferation ; DNA-Binding Proteins ; deficiency ; Female ; Graft vs Host Disease ; blood ; genetics ; immunology ; metabolism ; Heterografts ; immunology ; Humans ; Immunoglobulin G ; metabolism ; Immunoglobulin M ; metabolism ; Interleukins ; genetics ; immunology ; Male ; Mice ; Mice, Inbred BALB C ; Plasmids ; genetics

The Present study investigated not only the feasibility of organotypic spheroid culture system taken from human malignant gliomas, but also the similarities and differences between surgical specimens and cultured spheroids using light microscopy, electron microscopy, and flow cytometric examination. Surgically resected tumor specimens from eighteen human malignant gliomas were minced and explanted into agarose-coated culture wells. After three to five days, these microtumor fragments emerged as spheroids and could be maintained as organotypic spheroids for more than eight weeks. Measurements of the spheroids showed that they decreased during the initial two to three weeks and afterwards remained unaltered over a specific period of time. This growth pattern of the spheroids was consistent with the condition of tumors in vivo suggesting the linkage of cell proliferation and loss. Light microscopic and electron microscopic studies of the spheroids demonstrated that morphological structures were similar to those of the original tumor tissue in vivo and histopathologic characteristics of the original tumor were maintained over a long culture period. The spheroids contained connective tissues, blood vessels, macrophages, and neutrophils maintaining a three-dimensional architectural resemblance to the original tumors. Of three pairs of the surgical and spheroid specimen examined by the flow cytometry, one showed a change of ploidy pattern and two contained increased fractions of proliferating cells. It is concluded that this microtumor spheroid system can maintain the characteristics of the original tumors, and may serve as an alternative to the in vivo xenograft model for the research of brain tumor biology, invasion and immunology while providing a valuable technique for the evaluation of new therapies, such as biological response modifiers.
Allergy and Immunology ; Biology ; Blood Vessels ; Brain Neoplasms ; Cell Proliferation ; Connective Tissue ; Flow Cytometry ; Glioma* ; Heterografts ; Humans* ; Immunologic Factors ; Macrophages ; Microscopy ; Microscopy, Electron ; Neutrophils ; Ploidies

There have been reported some important findings in immunology of psoriasis, such as lower than normal level of IgM, defective humoral immune, responsiveness to newly administered antigens during cytotoxic drug therapy, lower than normal incidence of active sensitization with DNCB, prolonged skin heterograft survival and presence of anti-IgG antibody in peripheral lymphocytes. These findings are strongly suggestive of some humoral as well as cellular immunne dysfunction in patients with psoriasis. The present study was undertaken to evaluate the therapeutic effectiveness of BCG, a potent nonspecific immune stimulator, in patierts with psoriasis. Total 41 cases of more than 2 years' duration of illness and with widespread involvement and having had history of various previous treatment were slelcted at the Department of Dermatology, National Medical Center during October, 1973 and October, 1974, Liquid BCG(manufactured by NIH, Korea) was administered intradermally in a starting dose of 0.1 ml(0.01 mg in dry weight) weekly. The dosage of BCG was increased to 0.3 or 0.5ml after 3 or 5 injections and this dosage was maintained until some clinical changes were observed. The mean injection time to each patient was 20. 7.The results were as follows: 1. Eighteen cases showed no therapeutic response or aggravation of the lesions. 2. Six cases showed good therapeutic effect, resulting in only a few small lesions remained on elbows, knees or scalp areas. 3. Fourteen cases showed excellent therapeutic effect, that is complete disappearance of the lesions. During 2 to 6 months'follow up period, 7 cases showed relapse and the remainders are still under observation. The mechanism of influence of BCG on the clinical course of psoriatic patients is not determined, however, possible mechanism was discussed.
Allergy and Immunology ; Dermatology ; Dinitrochlorobenzene ; Drug Therapy ; Elbow ; Heterografts ; Humans ; Immunoglobulin M ; Incidence ; Knee ; Lymphocytes ; Mycobacterium bovis* ; Psoriasis* ; Recurrence ; Scalp ; Skin