Objective To summarize the research progress of xenotransplantation.Methods Domestic and international publications about xenotransplantation were summarized and reviewed. Results Hyperacute xenograft rejection was a huge problem for xenotransplantation， but it could be alleviated if the organs or tissues of donor were genetically modified. So far the graft survival time differed greatly due to characteristics of different organ. Conclusions By reviewing the studies of relevant papers about xenotransplantation， a comprehensive understanding of research background and a suitable research direction of xenotransplantation can be supplied. The graft organs or tissues from genetically modified donors are expected to avoid or alleviate hyperacute xenograft rejection.
Objective To study the effect of pravastatin on the survival of islet xenografts.MethodsPigtomouse islet transplantation was performed. The models were divided into 4 groups: group A (control); group B, treated with CsA; group C, treated with pravastatin; group D, treatment with combined CsA with pravastatin. The survival time (ST) of the grafts in each group were recorded. Histological examination was used to detect the inflammation and islet cells in the graft. The infiltrated cells were detected by immunohistochemistry with CD4+, CD8+ and CD68 monoclonal antibody. The serum NO was measured. RTPCR was used in the test of IFNγ mRNA.ResultsThe ST of group A,B,C,D was (6.2±0.82) d, (9.2±1.92) d, (7.2±1.30) d, (11.2±1.76) d respectively, the ST of group D was much longer than that of the other groups (P＜0.05).Compared to that in other groups, less infiltrated cell in group D was found. On the 4th postoperative day, the serum NO in group A was (105.0±19.3) mmol/L,significantly higher than that in group B 〔(88.20±21.04) mmol/L〕, in group C 〔(70.7±17.8) mmol/L)〕 and in group D 〔(56.30±16.4) mmol/L〕. When rejection occurred, the serum NO in group C and D was (83.7±10.6) mmol/L and (71.3±13.8) mmol/L, also lower than that in group A (P＜0.05), the serum NO in group B was (104.7±16.3) mmol/L, compared that in group A, no significance was present (Pgt;0.05）. On the 4th postoperative day, the serum expression of IFNγ mRNA in group D was 23.5±4.6, lower than that in group A (28.8±4.8), and no significance was present compared with that in group B and C. ConclusionPravastatin can abate the role of macrophages, especially combined with Cyclosporine, and can prolong the survival of islet xenograft.
ObjectiveTo investigate the aim antigen coursing the hyperacute rejection of xenotransplantation. MethodsDocuments about hyperacute rejection in xenotransplantation were reviewed and summarized in detail. ResultsPig is thought to be one of the ideal donors of xenotransplantation, but the major obstacle is hyperacute rejection mediated by complement that is activated though human serum. αGal is recognized as the major antigen and its expression is controlled by α1,3 galactosyltransferase. Immunoabsorption of preexsisted antibody, enzymatic digestion of αGal, knockout αGT gene and transgenic technology have been used to solve this problem. Even so, there remain other antigens which can combine with natural antibodies in human serum, such as, 40×103 molecule in erythrocyte, 210×103, 105×103 and 50×103 antigen in pig embryo brain cell, etc. Conclusion αGal is the major antigen which course the hyperacute rejection. Besides αGal, many nonalphagal need further investigation.
ObjectiveTo study the mechanism of reducing the intratumoral microvessel density (MVD) by Ginsenoside Rg3 (Rg3) combined with cytotoxic agent in xenotransplanted human breast infiltrating duct carcinoma in nude mice. MethodsSixteen female nude mice were randomly divided into 4 groups to receive cyclophosphamid (16 mg/kg,qd) combined with Rg3 (10 mg/kg, qd),Rg3(10 mg/kg,qd) alone,cyclophosphamid (16 mg/kg,qd) alone and 0.5% sodium carboxymethyl cellulose (0.5 ml,qd) respectively for 55 days. Breast cancer mass were weighed and sampled for light microscopic observation. The intratumor MVD was examined by immunohistochemical staining. ResultsThe tumor weight of treated group was significantly lower than that of control group. The tumor weight of the Rg3 combined with CTX group was lower than that of Rg3 group. The MVD value of Rg3 group was significantly lower than that of CTX group and control group. The MVD was significantly reduced in the Rg3 combined with CTX group than that in the others.ConclusionRg3 combined with CTX can inhibit the growth of xenotransplanted human breast infiltrating duct carcinoma, and reduce the intratumoral MVD.
Objective To investigate the mechanism of hyperacute rejection (HAR) in pig to rhesus monkey vein xenograft. Methods Porcine femoral vein was transplanted into rhesus monkey. Deposits of IgM, IgG, C3 and C4 on the grafts were observed by immunoflurescence. Results Great deal of IgM, C3 and C4 were seen along the endothelium of donor vein, but IgG was not seen. ConclusionIn pig to monkey xenograft model, HAR is intiated by the binding of xenoreactive IgM to donor xenoantigens and followed by the activation of complement via the classical pathway.
Abstract: Objective To assess the feasibility of transferring major histocompatibility complex (MHC) gene into the thymus to mitigate xenograft rejection. Methods By molecular cloning technique, we extracted and proliferated the-H-2K d gene from donor mice (MHC class Ⅰ gene of Balb/c mice) and constructed the expression vector plasmid of pCI-H-2K d. Twenty SD rats were selected as receptors, and by using random number table, they were divided into the experimental group and the control group with equal number of rats in each group. By ultrasoundguided puncture and lipofection method, the pCI-H-2Kd was injected into thymus of SD rats in the experimental group and meanwhile, empty vector plasmid of pCIneo was injected into thymus of SD rats in the control group. Subsequently, we transplanted the donor mice myocardium xenografts into the receptor rats, and observed the xenograft rejection in both the two groups. Results The survival time of the xenotransplanted myocardium in the experimental group was significantly longer than that in the control group (14.61±2.98 d vs. 6.40±1.58 d, t=-7.619,Plt;0.05). Microtome section of transplanted myocardium in the control group showed a relatively large amount of lymphocyte infiltration and necrosis occurred to most part of the transplanted myocardium, while microtome section of experiment group showed no lymphocyte infiltration and most of the cells of the transplanted myocardium were still alive. After mixed lymphocyte culture, the reaction of receptors to donor cells in the experiment group was obviously lower than that in the control group (t=4.758, P=0.000).After the count by flow cytometer, the xenoMHC molecules were expressed in the receptors’ thymus with a transfection efficiency of 60.7%. Conclusion Our findings suggest that xenograft rejection can be mitigated substantially by donor’s MHC gene transferring into receptor’s thymus. This may provide theoretical and experimental evidence for inducing xenotransplantation tolerance.
Objective To develop a new small-caliber vascular xenograft and evaluate the feasibility of xenogenic artery for coronary artery bypass grafting. Methods Canine carotid arteries were decellularized by detergent and enzymatic extraction. All decellularized xenografts were randomly divided into two groups. Heparin-linked group （n=24）： grafts were then covalently linked with heparin. Non-heparin-linked group （n=24）： as control. Xenografts in two groups were implanted in rabbits＇ left and right carotid artery respectively as bypass grafts. Graft patency was checked by ultrasonography after 3 weeks, 3 and 6 months. Grafts were harvested after 3 and 6 months. Microscopic observation and immunohistochemical staining were performed. Results All the cells were removed while the extracellular matrix were well preserved observed. Heparin was successfully linked to the grafts through their whole thickness. There was no obstruction at both sides after implantation of the grafts, while less thrombus was found in the decellularized heparin-linked grafts than in the other side. Smooth muscle cells densely populated the graft wall and endothelial cells covered the lumen at 3 months after implantation. Conclusion Canine common carotid artery treated by detergent and enzymatic extraction and heparin linkage may be a new small-caliber vascular xenograft for coronary artery bypass grafting.
Objective To evaluate tissue regeneration, body reaction, and biological safety of xenogeneous bladder acellular matrix (BAM) that can be used to repair rabbit bladder. Methods Porcine BAM was prepared through physical, chemical, and enzymatic methods, and the effects of acellularization and the structure were observed with HE staining and scanning electron microscope (SEM). Eighteen New Zealand white rabbits (weighing, 2.5-3.0 kg) undergoing partial cystectomy were randomly divided into 2 groups. After partial (about 30%) cystectomy, the porcine BAM was used to replace partial rabbit bladder in the experimental group (n=12), and the incision was directly sutured as control group (n=6). The survival condition of animals was observed after operation. At 15 days, 1, 2, 3, and 6 months after operation, the blood routine, renal function, and electrolyte were tested by collecting the blood samples. At 1, 2, 3, and 6 months after operation, maximum bladder capacity, bladder leak point pressure, and bladder compliance were measured through urodynamic studies. Then gross observation was performed for regeneration of bladder, and the specimens of the bladder were harvested for HE staining and immunohistochemical staining. The surrounding organs and local lymphoid tissues were harvested for gross observation and HE staining. Results Cell components were completely removed in the porcine BAM, showing three-dimensional porous structure under SEM. All the animals survived during the experiment. At 15 days after operation, white blood cell count increased, and then returned to normal level in 2 groups, showing no significant difference between 2 groups (P gt; 0.05). The tests of renal function and electrolyte suggested no significant difference between 2 groups (P gt; 0.05). The level of serum creatinine showed a tendency of increase, but it remained within normal range at 6 months after operation. The maximum bladder capacity and compliance in experimental group were significantly higher than those in control group at 3 and 6 months after operation (P lt; 0.05), but no significant difference in bladder leak point pressure at each time point between 2 groups (P gt; 0.05). The urothelial regeneration, smooth muscle regeneration, and blood vessel regeneration were seen by histological observation in 2 groups. In the 2 groups, chronic inflammatory cells infiltration could be observed at 1 month postoperatively, and then chronic inflammatory cells decreased significantly (P lt; 0.05), until complete disappearance. There was no significant difference in score of chronic inflammatory cell infiltration between 2 groups at 3 and 6 months after operation (P gt; 0.05). The α-smooth muscle actin expression was significantly increased with time passing in 2 groups (P lt; 0.05), and it was significantly higher in control group than in experimental group at each time point (P lt; 0.05). In addition, gross and HE staining observations showed no abnormalities in surrounding organs and local lymphoid tissues. Conclusion No immune rejection response occurs when porcine BAM is used for xenotransplantation. It is indicated that porcine BAM is relative safety for xenotransplantation.
Objective To observe the systemic and local immune response after repair of nerve defect with acellular nerve xenograft laden with allogenic adipose-derived stem cells (ADSCs) in rhesus monkey so as to evaluate the safety of the proposed material for nerve reconstruction. Methods Bilateral tibial nerves were taken from a healthy adult male landrace (weighing 48 kg) to prepare acellular nerve xenograft by chemical extraction. ADSCs were isolated from a healthy adult male rhesus monkey (weighing 4.5 kg), and were seeded into the acellular nerve grafts. The radial nerve defect models with 25 mm in length were established in 10 healthy adult female rhesus monkeys (weighing 3-5 kg), and they were divided into cell-laden group (n=5) and non-cell-laden group (n=5) randomly. Defect was repaired with acellular nerve xenograft laden with allogenic ADSCs in cell-laden group, with acellular nerve xenograft only in non-cell-laden group. The blood samples were taken from peripheral vein preoperatively and at 14, 60, and 90 days after operation for lymphocyte analysis; at 5 months after operation, the grafts were harvested to perform histological examination for local immune response and nerve regeneration. The nerve autograft in rhesus monkey was used as control. Results In cell-laden group and non-cell-laden group, no significant difference was found in the count of lymphocytes and T lymphocytes, the percentage of T lymphocytes, CD8+ T lymphocytes, as well as the ratio of CD4+ T lymphocytes to CD8+ T lymphocytes between pre- and post-operation (P gt; 0.05); in cell-laden group, the percentage of CD4+ T lymphocytes at 14 days was significantly lower than that at 60 and 90 days postoperatively (P lt; 0.05). The percentage of CD4+ T lymphocytes in cell-laden group was significantly lower than that in non-cell-laden group at 14 days (P lt; 0.05), but no significant difference was found in the other indexes at the other time between 2 groups (P gt; 0.05). At 5 months after operation, mild adhesion was found on the surface of nerve xenografts; the epineurium of nerve xenografts was thicker than that of nerve autografts; and neither necrosis nor fibrosis was found. CD3+, CD4+, CD8+, CD68+, and CD163+ T lymphocytes were scattered within the grafts, in which regenerative axons were revealed. CD3+, CD4+, CD8+, CD68+, and CD163+ T lymphocytes were comparable in cell-laden group, non-cell-laden group, and autograft group. Conclusion Repair of nerve defect with acellular nerve xenograft elicits neither systemic nor local immune response in rhesus monkeys. Implantation of allogenic ADSCs might result in transient depression of CD4+ T lymphocytes proliferation early after surgery, no immune response can be found.
Objective To review the common methods of isolation and purification of porcine islets and research progress. Methods Domestic and abroad literature concerning the isolation and purification of porcine islets was reviewed and analyzed thoroughly. Results The efficacy of the isolation and purification depends on the selection of donor, the procurement and cryopreservation of high-quality donor pancreas, and the selection and improvement of the operation. Conclusion The shortage of transplanted islets could be resolved by the establishment of standardized and optimal process, which may also promote the development of porcine islet xenograft.