The role of donor hepatocytes and/or splenocytes pre-injection in reducing islet xenotransplantation rejection

OBJECTIVE: To observe the effect of tail vein injection with donor hepatocyte and/or splenocyte on islets xenotransplantation rejection. METHODS: New-born male pigs and BALB/C mice were selected as donors and recipients respectively. Islets xenotransplantation was performed in recipients just after the third time of tail vein injection with donor hepatocytes and/or splenocytes. Macrophage phagocytosis, NK killing activity, T lymphocyte transforming function of spleen cells, antibody forming function of B lymphocytes, and T lymphocyte subsets were taken to monitor transplantation rejection. The effects of this kind of transplantation were indicated as variation of blood glucose and survival days of recipients. RESULTS: Streptozotocin (STZ) succeeded in inducing diabetes mellitus models of mice. Pre-injection of donor hepatocytes, and splenocytes or their mixture via tail vein was effective in preventing donor islets transplantation from rejection, which was demonstrated by the mentioned immunological marks. And each group of transplantation could decrease the blood glucose of recipients and prolong the survival days. Pre-injection of mixture of donor hepatocytes and splenocytes was more effective in preventing rejection than pre-injection of donor hepatocytes or splenocytes separately. CONCLUSION: We propose that pre-injection of donor hepatocytes, splenocytes separately or their mixture before donor islets transplantation is a good way to prevent rejection.

Establishment of Xenotransplantation Model of Human CN-AML with FLT3-ITD^(mut)/NPM1 in NOD/SCID Mice

Summary： Patients with FLT3-ITD^mmutt/NPM1- cytogenetically normal acute myeloid leukemia （CN-AML）, as high-risk molecular group in CN-AML, are associated with a worse prognosis than other CN-AML patients. It is beneficial to generate xenotransplantation model of FLT3-ITD^mut/NPM1- CN-AML to better understand the pathogenesis and therapeutic strategies of such AML subtype. The purpose of present study was to establish the xenotransplantation model in NOD/SCID mice with FLT3-ITD^mut/NPM1- CN-AML primary cells. The FLT3-ITD^mut/NPM1- CN-AML primary cells from 3 of 7 cases were successfully transplanted into NOD/SCID mice, and human CD45 positive cells were detected in the peripheral blood, spleen and bone marrow of mice by using flow cytometry. Infiltration of human leukemia cells in various organs of mice was observed by using immunohistochemistry. Gene analysis confirmed sustained FLT3/ITD mutation without NPM1 mutation in mice. By performing serial transplantation, it was found that characteristics of the leukemia cells in secondary and tertiary genera- tion models remained unchanged. Moreover, in vivo cytarabine administration could extend survival of NOD/SCID mice, which was consistent with clinical observation. In conclusion, we successfully estab- lished xenotransplantation model of human FLT3-ITD^mut/NPM1- CN-AML in NOD/SCID mice. The model was able to present primary disease and suitable to evaluate the curative effects of new drugs or therapy strategies.

Establishment of Reproducible Xenotransplantation Model of T Cell Acute Lymphoblastic Leukemia in NOD/SCID Mice

T cell acute lymphoblastic leukemia(T-ALL) is an aggressive leukemia.However the poor prognosis and low morbidity restrict further analysis of the disease.Therefore there is an increasing demand to develop animal models for identifying novel therapeutic approaches.In this study,we inoculated the anti-mouse CD122 monoclonal antibody conditioned NOD/SCID mice with the leukemia cells from 9 T-ALL patients and 1 cell line via the tail vein.Four of the 9 patients and the cell line were successfully engrafted.Flow cytometry detected high percentage of human CD45 + cells in recipient mice.Immunohistochemistry showed infiltration of human CD45 + cells in different organs.Serial transplantation was also achieved.In vivo drug treatment showed that dexamethasone could extend survival,which was consistent with clinical observation.These results demonstrated that we successfully established 5 xenotransplantation models of T-ALL in anti-mCD122 mAb conditioned NOD/SCID mice,which recapitulated the characteristics of original disease.

Expression of L amino acid transport system 1 and analysis of iodine-123-methyltyrosine tumor uptake in a pancreatic xenotransplantation model using fused high-resolution-micro-SPECT-MRI

BACKGROUND:The specificity in discriminating pancreatitis is limited in the positron emission tomography(PET)using Fluorine-18-fluorodeoxyglucose.Furthermore,PET is not widely available compared to the single photon emission computed tomography(SPECT).Since amino acids play a minor role in metabolism of inflammatory cells,the potential of the SPECT tracer,3-[ 123 I]iodo-L-α-methyltyrosine(123I-IMT),for detecting pancreatic cancer was examined in xenotransplantation models of human pancreatic carcinoma in mice. METHODS: 123 I-IMT was injected to eight mice inoculated with subcutaneous or orthotopic pancreatic tumors.Fused high-resolution-micro-SPECT(Hi-SPECT)and magnetic resonance imaging were performed.The gene expression level of L amino acid transport-system 1(LAT1)was analyzed and correlated with tumor uptake of 123 I-IMT. RESULTS:A high uptake of 123 I-IMT was detected in all tumor-bearing mice.The median tumor-to-background ratio (T/B)was 12.1(2.0-13.2)for orthotopic and 8.4(1.8-11.1)for subcutaneous xenotransplantation,respectively.Accordingly, the LAT1 expression in transplanted Colo357 cells was increased compared to non-malignant controls.CONCLUSIONS:Our mouse model could show a high 123 I-IMT uptake in pancreatic cancer.Fused MRI scans facilitate precise evaluation of uptake in the specific regions of interest.Further studies are required to confirm these findings in tumors derived from other human pancreatic cancer cells.Since amino acids play a minor role in the metabolism of inflammatory cells,the potential for application of 123 I-IMT to distinguish pancreatic tumor from inflammatory pancreatitis warrants further investigation.

Xenotransplantation of embryonic pig pancreas for treatment of diabetes mellitus in non-human primates

Transplantation therapy for diabetes in humans is limited by the low availability of human donor whole pancreas or islets. Outcomes are complicated by immunosuppressive drug toxicity. Xenotransplantation is a strategy to overcome supply problems. Implantation of tissue obtained early during embryogenesis is a way to reduce transplant immunogenicity. Pig insulin is biologically active in humans. In that regard the pig is an appropriate xenogeneic organ donor. Insulin-producing cells originating from embryonic pig pancreas obtained very early following pancreatic primordium formation [embryonic day 28 (E28)] engraft long-term in rhesus macaques. Endocrine cells originating from embryonic pig pancreas transplanted in host mesentery migrate to mesenteric lymph nodes, engraft, differentiate and improve glucose tolerance in rhesus macaques without the need for immune suppression. Transplantation of embryonic pig pancreas is a novel approach towards beta cell replacement therapy that could be applicable to humans.

Response of the xenograft endothelium in the concordant xenotransplantation

To investigate the response of the xenograft endothelium in the concordant hamster to rat cardiac xenotransplantation and the mechanism of acute vascular rejection. Methods： The animals were divided into 5 groups randomly： control group, CsA group, splenectomy group, DO splenectomy＋CsA group and D3 splenectomy＋CsA group. Hamster heart was heterotopicaly transplanted to rat abdominal cavity. The graft survival was monitored by palpation of the rat abdominal wall. The histological and ultrastructural changes of the xenogafts were investigated. NF-κB and P-selectin expression in the xenograft were detected. Heme Oxigenase-1 and Bcl-2 expression were also detected in the xenografts of different groups. Results： The mean survival time of the xenografts in control group, CsA group, splenectomy group, DO splenectomy＋CsA group and D3 splenectomy＋CsA group was 3.4±0.55, 3.8±0,45, 6.4± 1.52, 30 and 7.4 ±1.14 days. The rejected graft showed typical acute vascular rejection in control group, CsA group,splenectomy group and D3 splenectomy＋CsA group. Endothelial cells of the rejected xenograft showed dramatic assembly of ribosomes and expansion of the rough endoplasmic reticulum. However, the endothelium of the long-term survived grafts in DO splenectomy＋CsA group showed normal architecture. NF-κB and P-selectin expression were detected in the rejected xenografts. HO-1 expression was observed in the long-term survived xenografts in DO splenectomy＋CsA group. Conclusion： The endothelial cells of the xenograft might be activated during the acute vascular rejection. Expression of HO-1 might inhibit the upregulation of NF-κB and adhesion molecular which decreases the activation of the endotheliuna of the graft.

Current advances in xenotransplantation

BACKGROUND: Transplantation of organs is a well-known and accepted life-saving procedure for end-stage kidney, liver, heart and lung diseases. The insufficient number of donor organs limits the application of this technique and leads to unnecessary loss of life. Experimental techniques such as xenotransplantation are extremely important to determine new methods of creating organ availability. DATA SOURCES; A literature search of Pubmed database was conducted and research articles reviewed. RESULTS: Xenotransplantation is a progressive field of research. Human complement regulatory protein ( hDAF) transgenic pigs and new immunosuppressive strategies that reduce xenoreactive αgal antibodies, have decreased rates of acute vascular rejection. Transplantation of α-1, 3-galac-tosyltransferase knock-out pig organs into baboons has resulted in the longest graft survival to date. Coagulation pathways have been identified as having a role in graft rejection. In vitro studies of porcine endogenous retroviruses (PERVs) show encouraging results that zoonosis will be less hindering to xenotransplantation than once thought. CONCLUSIONS: Several recent advances in xenotransplantation research have brought this technique closer to clinical application. The Ethics Committee of the International Xenotransplantation Association has made recommendations to ensure maintenance of ethical standards. Advancement will depend on the development of pig models, novel immunosuppressive strategies to target the innate immune system, and new ways to create donor specific tolerance. Prevention of rejection and transmission of infectious agents remain unresolved issues. In the future, it is feasible that xenotransplantation will be used to resolve this medical dilemma.

Use of zebrafish embryos as avatar of patients with pancreatic cancer:A new xenotransplantation model towards personalized medicine

BACKGROUND The response to chemotherapy treatment of patients with pancreatic ductal adenocarcinoma(PDAC)is difficult to predict and the identification of patients who most likely will benefit from aggressive chemotherapy approaches is crucial.The concept of personalized medicine has emerged in the last years with the objective to tailor the medical treatment to the individual characteristics of each patient,and particularly to the tumor biology of each patient.The need for invivo xenotransplantation models for cancer patients has increased exponentially,and for this reason zebrafish avatars have gained popularity.Preliminary studies were conducted also with PDAC tissue.AIM To develop a simple,not expensive,diffusible zebrafish embryo model as avatar for patients affected by PDAC.METHODS Tumor tissue was taken from the surgical specimen by the histopathologist.After its fragmentation into small pieces,they are stained with CM-Dil.Small pieces of stained tissue were transplanted into the yolk of wt AB zebrafish embryos with a glass capillary needle.Embryos were incubated at 35°C in E3 medium supplemented with 1%Pen/Strep in the presence or absence of drugs for the following days in respect of the treatment plan(Gemcitabine;Gemcitabine and Oxaliplatin;Gemcitabine and nab-Paclitaxel;5-Fluorouracil and Folinic acid and Oxaliplatin and Irinotecan).The response of zebrafish xenografts to the chemotherapy options has been analyzed by monitoring the fluorescent stained area at 2 h post injection(hpi),1 d and 2 d post injection(dpi).In each time point,the mean size of the stained area was measured by ImageJ and it was normalized with respect to the 1 dpi time point mean relative tumor area(RTA).We evaluated the effect of the chemotherapy exposition comparing the mean RTA of each treated subgroup and the control group and evaluating the percentage reduction of the mean RTA by comparing each treated subgroup with the control group.RESULTS Between July 2018 and October 2019,a total of 15 patients with pancreatic cancer were prospectively enrolled.In all cases,it was possible to take a fragment of the tumor from the surgical specimen for the xenotransplantation in the zebrafish embryos.The histological examination confirmed the presence of a PDAC in all cases.In absence of chemotherapy(control group),over time the Dil-stained area showed a statistically significant increase in all cases.A statistically significant reduction of the mean RTA in the treated subgroups for at least one chemotherapy scheme was reported in 6/15(40%)cases.The analysis of the percentage reduction of the RTA in treated subgroups in comparison to the control group revealed the presence of a linear relationship in each subgroup between the percentage reduction of the RTA and the number of cases reporting each percentage threshold considered for the analysis.CONCLUSION Our model seems to be effective for the xenotransplantation of PDAC tissue and evaluation of the effect of each chemotherapy scheme on the xenotransplanted tumor tissue.

A humanized mouse model to study human immune response in xenotransplantation

BACKGROUND: A major barrier to the clinical application of xenotransplantation as a treatment option for patients is T cell-mediated rejection. Studies based on experimental rodent models of xenograft tolerance or rejection in vivo have provided useful information about the role of T cell immune response in xenotransplantation. However not all observations seen in rodents faithfully recapitulate the human situation This study aimed to establish a humanized mouse model of xenotransplantation, which mimics xenograft rejection in the context of the human immune system. METHODS: NOD-SCID IL2rγ -/- mice were transplanted with neonatal porcine islet cell clusters (NICC) followed by reconstitution of human peripheral blood mononuclear cells (PBMC). Human leukocyte engraftment and islet xenograft rejection were confirmed by flow cytometric and histological analyses. RESULTS: In the absence of human PBMC, porcine NICC transplanted into NOD-SCID IL2rγ -/- mice revealed excellent graft integrity and endocrine function. Human PBMC demonstrated a high level of engraftment in NOD-SCID IL2rγ -/- mice. Reconstitution of NICC recipient NOD-SCID IL2rγ -/- mice with human PBMC led to the rapid destruction of NICC xenografts in a PBMC number-dependent manner. CONCLUSIONS: Human PBMC-reconstituted NOD-SCID IL2rγ -/- mice provide an ideal model to study human immune responses in xenotransplantation. Studies based on this humanized mouse model will provide insight for improving the outcomes of clinical xenotransplantation.

An improved model of heterotopic cardiac xenotransplantation from guinea－pig to rat

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Is the near coming xenotransplantation era relieving us from needing to look for more non-living organ donors?

Despite organ transplantation being the most successful treatment for end-stage organ dysfunction,the number of annual solid organ transplantations is much lower than that required to satisfy the demand of patients on waiting lists.The explanation for this phenomenon is the relative scarcity of non-living organ donors due to several factors,such as:(1)Late arrival of patients with a neurocritical condition to an emergency service;(2)lack of detection of those patients as possible organ donors by health professionals dedicated to procurement or by clinicians at emergency and intensive care units,for instance;(3)late transfer of the patient to an intensive care unit to try to recover their health and to provide hemodynamic,ventilatory,and metabolic support;(4)lack of confirmation of the physiological status of the possible donor;(5)late or incorrect positive diagnosis of the subject’s death,either due to brain or cardiac death;(6)difficulty in obtaining legal authorization,either by direct relatives or by the authority,for the extraction of organs;and(7)deficient retrieval surgery of the organs actually donated.The recent reports of relatively successful xenotransplants from genetically modified pigs open the possibility to fix this mismatch between supply and demand,but some technical(organ rejection and opportunistic infections),and economic issues,still remain before accepting a progressive replacement of the organ sources for transplantation.An approximate economic cost analysis suggests that the hypothetical acquisition cost of any genetically modified pig derived organ is high and would not even satisfy the solid organ demand of the wealthiest countries.

Multiple gene modifications of pigs for overcoming obstacles of xenotransplantation

Xenotransplantation,involving animal organ transplantation into humans to address the human organ shortage,has been studied since the 17th century.Early attempts to obtain organs from animals such as goats,dogs,and non-human primates proved unsuccessful.In the 1990s,scientists agreed that pigs were the most suitable donor animals for xenotransplantation.However,immune rejection between pig and human has hindered the application.To overcome these challenges,researchers developed genetically modified pigs that deactivate xenoreactive antigen genes and express human protective genes.These advances extended xenograft survival from days to years in non-human primates,resulting in the first human heart xenotransplant trial.Using genetically engineered pigs for the organ shortage is promising.This review provides an overview of potential incompatibilities of immunogenicity and functional proteins related to xenotransplantation between humans and pigs.Furthermore,it elucidates possible approaches for multiplex gene modification to breed better-humanized pigs for clinical xenotransplantation.

The resurgent landscape of xenotransplantation of pig organs in nonhuman primates

Organ shortage is a major bottleneck in allotransplantation and causes many wait-listed patients to die or become too sick for transplantation.Genetically engineered pigs have been discussed as a potential alternative to allogeneic donor organs.Although xenotransplantation of pig-derived organs in nonhuman primates(NHPs)has shown sequential advances in recent years,there are still underlying problems that need to be completely addressed before clinical applications,including(i)acute humoral xenograft rejection;(ii)acute cellular rejection;(iii)dysregulation of coagulation and inflammation;(iv)physiological incompatibility;and(v)cross-species infection.Moreover,various genetic modifications to the pig donor need to be fully characterized,with the aim of identifying the ideal transgene combination for upcoming clinical trials.In addition,suitable pretransplant screening methods need to be confirmed for optimal donor-recipient matching,ensuring a good outcome from xenotransplantation.Herein,we summarize the understanding of organ xenotransplantation in pigs-to-NHPs and highlight the current status and recent progress in extending the survival time of pig xenografts and recipients.We also discuss practical strategies for overcoming the obstacles to xenotransplantation mentioned above to further advance transplantation of pig organs in the clinic.

Cytokine production during the inhibition of acute vascular rejection in a concordant hamster-to-rat cardiac xenotransplantation model

Background The aim of the current study was to investigate the role of interleukin （IL）-2, interferon （IFN）-y, IL-4 and IL-10 in a concordant hamster-to-rat cardiac xenotransplantation model. Methods A hamster-to-rat cardiac transplantation was performed using SD rats as recipients of Golden Syrian hamster hearts. A total of 60 SD rats were divided into four groups and treated as follows： control group （n=15）; splenectomy group （n=15）; CsA group （n=15）; CsA ＋ splenectomy group （n=-15）. Levels of IL-2, IFN-γ, IL-4 and IL-10 were measured by enzyme linked immunosorbent assay （ELISA）. Sera were harvested at different time points in each group： day 1, and 3 as well as the day the xenograft stopped beating in the control group and CsA group; day 1, 3, 7, 14 and 30 in the splenectomy group and CsA＋splenectomy group. The expression of P-selectin and intercellular adhesion molecule-1 （ICAM-1） was examined by immunohistochemical analysis of the xenogralt alter cardiac xenotransplantation. Results Serum levels of IL-2 and IFN-γ were upregulated in untreated （day 3） and splenectomy-treated animals （day 7） compared to CsA ＋ splenectomy treated animals （day 7）. IL-10 was upregulated in long-term survival recipients following splenectomy ＋ CsA. Neither P-selectin nor ICAM-1 expression was detected in long-term survival xenografts. Conclusions Serum IL-2 and IFN-γ were elevated following acute vascular rejection. Serum IL-10 was correlated to immunosuppression and protective effects in long-term survival rats following concordant cardiac xenotransplantation.

Pig islets for islet xenotransplantation： current status and future perspectives

Objective To review the current status and progress on pig islet xenotransplantation.Data sources Data used in this review were mainly from English literature of Pubmed database.The search terms were ＂pig islet＂ and ＂xenotransplantation＂.Study selection The original articles and critical reviews selected were relevant to this review＇s theme.Results Pigs are suggested to be an ideal candidate for obtaining available islet cells for transplantation.However,the potential clinical application of pig islet is still facing challenges including inadequate yield of high-quality functional islets and xenorejection of the transplants.The former can be overcome mainly by selection of a suitable pathogen-free source herd and the development of isolation and purification technology.While the feasibility of successful preclinical pig islet xenotranplantation provides insights in the possible mechanisms of xenogeneic immune recognition and rejection to overwhelm the latter.In addition,the achievement of long-term insulin independence in diabetic models by means of distinct islet products and novel immunotherapeutic strategies is promising.Conclusions Pig islet xenotransplantation is one of the prospective treatments to bridge the gap between the needs of transplantation in patients with diabetes and available islet cells.Nonetheless,further studies and efforts are needed to translate obtained findings into tangible applications.

Successful xenotransplantation of microencapsulated newborn pig parathyroid cells in the treatment of hypoparathyroidism in rats

Objective To study the effect of xenotransplantation with pig parathyroid cells,which was prepared using cell microencapsulation technique,on the treatment of hypoparathyroidism in rats without immunosuppressor. Methods Parathyroid cells were isolated from 10 healthy newborn pigs and encapsulated in alginate-polylysine-alginate (APA) membranes. Thirty-two aparathyroid Wistar rats were randomly allocated to microcapsule,non-microcapsule,empty microcapsule,and control groups. Each rat was injected intraperitoneally with encapsulated porcine parathyroid cells,free porcine parathyroid cells,empty capsules or 0.9% NaCl,respectively. Total serum calcium and parathyroid hormone levels were monitored continuously for 40 weeks. And then,the transplant beds were retrieved and subjected to morphologic and electron microscopic examination. Results In those animals xenotransplanted with microencapsulated porcine parathyroid cells,the calcium and PTH levels were consistently within the normal range during the 40 weeks. In contrast,no therapeutic effects were observed in rats in the non-microcapsule group. Furthermore,neither empty capsules nor 0.9% NaCl were shown to have any effect on the recipient’s serum calcium or PTH levels. After 40 weeks,electron microscopic examination demonstrated that the parathyroid cells within the microcapsules had survived well in vivo . Conclusions Xenotransplantation of microencapsulated newborn pig parathyroid cells can successfully treat hypoparathyroidism in rats without using immunosuppressive drugs. The results of this study show the possible clinical use of microencapsulated porcine parathyroid cells.

Experimental study on the vitrification and xenotransplantation of human ovarian tissue

Background:Cryopreservation of ovarian tissue is a promising method for preserving fertility.Transmission electron microscopy(TEM)is an evaluation system for cryo-injury during the cooling and warming process which is very laborious and needs to be optimized.Objective:In this study,we evaluated that serum 17β-oestradiol(E2)may be used as an indicator of vitrified ovarian tissue.Methods:Immunodeficient nude mice were used as hosts for xenografting of vitrified-warmed human ovarian tissues.A total of 54 mice were divided into two group:vitrified ovarian xenotransplant(VOX)group(n?45)and non-transplant control group(n?9).The transplanted mice were grouped into vitrified/warmed grafted-4 weeks(VOX-4w,n?15),vitrified/warmed grafted-6weeks(VOX-6w n?15)and vitrified/warmed grafted-12 weeks(VOX-12w n?15)according to the time after transplantation.The viable and functional recovery of grafted ovarian tissue was assessed by light microscopy,transmission electron microscopy,and hormone(E2)assays.Results:Serum E2 concentration was significantly higher in VOX-6w(group(21.07 pg/ml)than that of VOX-12w group(15.59 pg/ml).VOX-12w group showed a lower value(12.61 pg/ml)for E2 concentration.The trend for E2 concentration was consistent with the morphological identification of the grafts.Conclusion:In vivo serum hormone E2 released by cortical biopsies can be used as a functional marker for xenotransplanted vitrified-warmed human ovarian tissue reserve.

EXPERIMENTAL INVESTIGATION OF THREE IMMUNO-SUPPRESSIVE DRUGS IN XENOTRANSPLANTATION

Organ transplantation between phylogenetically disparate animals are subject to hyperacute rejection that is thought to be mediated by the reaction of natural and

A bioartificial transgenic porcine whole liver expressing human proteins alleviates acute liver failure in pigs

Background:Preventing heterologous protein influx in patients is important when using xenogeneic bioartificial livers(BALs)to treat liver failure.The development of transgenic porcine livers synthesizing human proteins is a promising approach in this regard.Here,we evaluated the safety and efficacy of a transgenic porcine liver synthesizing human albumin(h ALB)and coagulation factor VII(h FVII)within a bioartificial system.Methods:Tibetan miniature pigs were randomly subjected to different interventions after surgeryinduced partially ischemic liver failure.Group A(n=4)was subjected to basic treatment;group B(n=4)was to standard medical treatment and wild-type porcine BAL perfusion,and group C(n=2)was to standard medical treatment and transgenic BAL perfusion.Biochemical parameters,coagulation status,survival time,and pathological changes were determined.Expressions of h ALB and h FVII were detected using immunohistochemistry and enzyme-linked immunosorbent assays.Results:The survival time in group A was 9.75±1.26 days;this was shorter than that in both perfused groups,in which all animals reached an endpoint of 12 days(P=0.006).Ammonia,bilirubin,and lactate levels were significantly decreased,whereas albumin and fibrinogen levels were increased after perfusion(all P<0.05).h ALB and h FVII were detected in transgenic BAL-perfused pig serum and ex vivo in the liver tissues.Conclusions:The humanized transgenic pig livers could synthesize and secrete h ALB and h FVII ex vivo in a whole organ-based bioartificial system,while maintaining their metabolism,detoxification,transformation,and excretion functions,which were comparable to those observed in wild-type porcine livers.Therefore,the use of transgenic bioartificial whole livers is expected to become a new approach in treating acute liver failure.