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 mode...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.展开更多
OBJECTIVE: To observe the effect of tail vein injection with donor hepatocyte and/or splenocyte onislets xenotransplantation rejection.METHODS: New-born male pigs and BALB/C mice were selected as donors and recipients...OBJECTIVE: To observe the effect of tail vein injection with donor hepatocyte and/or splenocyte onislets 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 withdonor hepatocytes and/or splenocytes. Macrophage phagocytosis, NK killing activity, T lymphocytetransforming function of spleen cells, antibody forming function of B lymphocytes, and T lymphocytesubsets were taken to monitor transplantation rejection. The effects of this kind of transplantation wereindicated as variation of blood glucose and survival days of recipients.RESULTS: Streptozotocin (STZ) succeeded in inducing diabetes mellitus models of mice. Pre-injectionof donor hepatocytes, and splenocytes or their mixture via tail vein was effective in preventing donor isletstransplantation from rejection, which was demonstrated by the mentioned immunological marks. And eachgroup 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 rejectionthan pre-injection of donor hepatocytes or splenocytes separately.CONCLUSION: We propose that pre-injection of donor hepatocytes, splenocytes separately or theirmixture before donor islets transplantation is a good way to prevent rejection.展开更多
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 e...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.展开更多
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...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.展开更多
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 rej...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.展开更多
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 tec...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.展开更多
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 chemothe...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.展开更多
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...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.展开更多
AIM: To identify the decreasing effect of xenotransplantion in combination with privileged sites on rejection and death of biological semipermeable membrane-(BSM) encapsulated implanted islets.METHODS: After the BSM e...AIM: To identify the decreasing effect of xenotransplantion in combination with privileged sites on rejection and death of biological semipermeable membrane-(BSM) encapsulated implanted islets.METHODS: After the BSM experiment in vitro, BSMencapsulated SD rat's islet-like cell clusters (ICCs) were xenotransplanted into normal dog's brain. Morphological changes were observed under light and transmission electron microscope. The islets and apoptosis of implanted B cells were identified by insulin-TUNEL double staining.RESULTS: The BSM used in our study had a favorable permeability, some degree of rigidity, lighter foreign body reaction and toxicity. The grafts consisted of epithelioid cells and loose connective tissue. Severe infiltration of inflammatory cells was not observed. The implanted ICCs were identified 2 mo later and showed typical apoptosis.CONCLUSION: BSM xenotransplantation in combination with the privileged site can inhibit the rejection of implanted heterogeneous ICCs, and death of implanted heterogeneous B cells is associated with apoptosis.展开更多
To lay background for studying rejection mechanisms in xenotransplantation and developing the strategies for intervention, class I genes of swine leukocyte antigens (SLA) of three Chinese pig strains Bm, Gz and Yn wer...To lay background for studying rejection mechanisms in xenotransplantation and developing the strategies for intervention, class I genes of swine leukocyte antigens (SLA) of three Chinese pig strains Bm, Gz and Yn were cloned and sequenced. The eDNA of the class I loci P1 and P14 were amplified by RT-PCR and subjected to insert into sequencing vectors. All six allelic sequences we examined, each two for one Chinese strain, are not identical to those reported, which allows these novel sequences receiving their accession numbers AY102467- AY102472 from GenBank. This study further reveals that the homologies of MHC class I genes in their primary structures and the deduced amino acids between Chinese pigs (SLA) and human (HLA-A*0201) are better than those between pigs and mice (H-2Db/H-2Kb). The comparison also indicates that the amino acid residues critical for recognition by human KIRs are altered in the swine class I molecules. The amino acids responsible for binding human CD8 coreceptor are largely conserved although there are two critical residues substituted. A functional test indicated that the human T cells specific for the prokaryotically expressed SLA P1protein could respond quite well in vitro to the class I-positive swine chondrocytes and PBMCs in presence of human APCs. This implies that, due to the substitution of two critical residues, the inaccessibility of human CD8 coreceptor to swine class I molecule might be contributable to the indirect pathway that the human T cells have to use for recognizing the SLA class I xenogeneic antigens.展开更多
Objective: 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 ...Objective: 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, D0 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, D0 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 D0 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 D0 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 endothelium of the graft.展开更多
Patients with FLT3-ITDmut /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 gen...Patients with FLT3-ITDmut /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-ITDmut /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-ITDmut /NPM1 CN-AML primary cells. The FLT3-ITDmut /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 generation 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 established xenotransplantation model of human FLT3-ITDmut /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.展开更多
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 no...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.展开更多
目的:研究溶质载体家族6成员9(solute carrier family 6 member 9,SLC6A9)表达对结直肠癌细胞增殖、迁移和5-氟尿嘧啶(5-fluorouracil,5-FU)药物敏感性的影响。方法:TCGA数据库分析、实时荧光定量PCR和Western blot分析检测SLC6A9在结...目的:研究溶质载体家族6成员9(solute carrier family 6 member 9,SLC6A9)表达对结直肠癌细胞增殖、迁移和5-氟尿嘧啶(5-fluorouracil,5-FU)药物敏感性的影响。方法:TCGA数据库分析、实时荧光定量PCR和Western blot分析检测SLC6A9在结肠癌组织、正常结肠细胞系(NCM460)和结直肠癌细胞系(SW620、HCT116、HT29、Lovo和SW480)中的表达。将SCL6A9过表达质粒及阴性对照(SLC6A9 OE、Vector)转染HT29细胞,将SCL6A9小干扰RNA及阴性对照(SLC6A9 siRNA1#、siRNA2#和Scramble)转染SW620细胞。划痕愈合实验和Transwell实验检测各组细胞的迁移、侵袭能力。Western blot和细胞免疫荧光检测EMT相关蛋白E-cadherin、Vimentin的表达水平。利用CCK-8法和构建裸鼠移植瘤模型检测SLC6A9过表达对结直肠癌细胞5-FU药物敏感性的影响。结果:与正常结肠组织和NCM460细胞相比,SLC6A9在结肠癌组织和结直肠癌细胞系中低表达(均P<0.05)。SLC6A9过表达引起E-cadherin蛋白表达增加,Vimentin蛋白水平降低,抑制结直肠癌细胞的迁移、侵袭(P<0.05)。SLC6A9低表达引起E-cadherin蛋白表达降低,Vimentin蛋白水平增加,促进结直肠癌细胞的迁移、侵袭能力(P<0.05)。SLC6A9过表达提高了5-FU的药物敏感性,并使肿瘤生长缓慢,质量减轻(P<0.05)。而SLC6A9低表达降低了5-FU的药物敏感性(P<0.05)。结论:SLC6A9过表达能够抑制结直肠癌细胞的迁移、侵袭和EMT进程,并增强5-FU对结直肠癌细胞的药物敏感性。展开更多
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...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.展开更多
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...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.展开更多
基金supported in part by the National Natural Science Foundation of China (No. 81025011 and No.81090414)
文摘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.
基金This work was supported by Natural Science Foundation of Shandong province, China (No. Y99C07).
文摘OBJECTIVE: To observe the effect of tail vein injection with donor hepatocyte and/or splenocyte onislets 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 withdonor hepatocytes and/or splenocytes. Macrophage phagocytosis, NK killing activity, T lymphocytetransforming function of spleen cells, antibody forming function of B lymphocytes, and T lymphocytesubsets were taken to monitor transplantation rejection. The effects of this kind of transplantation wereindicated as variation of blood glucose and survival days of recipients.RESULTS: Streptozotocin (STZ) succeeded in inducing diabetes mellitus models of mice. Pre-injectionof donor hepatocytes, and splenocytes or their mixture via tail vein was effective in preventing donor isletstransplantation from rejection, which was demonstrated by the mentioned immunological marks. And eachgroup 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 rejectionthan pre-injection of donor hepatocytes or splenocytes separately.CONCLUSION: We propose that pre-injection of donor hepatocytes, splenocytes separately or theirmixture before donor islets transplantation is a good way to prevent rejection.
基金supported in part by a BMBF grant(TOMCAT)given to H.K.the Molecular Imaging North Competence Center(MOIN-CC)
文摘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.
文摘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.
基金supported by grants from the National Health and Medical Research Council of Australiathe National Natural Science Foundation of China(81271712)Hunan Provincial Natural Science Foundation of China(11JJ4078)
文摘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.
基金This study was supported by Ontario Research Develop- ment Challenge Fund.
文摘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.
文摘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.
文摘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.
基金Supported by the Natural Science Foundation of Shaanxi Province, No. 98SZ-064
文摘AIM: To identify the decreasing effect of xenotransplantion in combination with privileged sites on rejection and death of biological semipermeable membrane-(BSM) encapsulated implanted islets.METHODS: After the BSM experiment in vitro, BSMencapsulated SD rat's islet-like cell clusters (ICCs) were xenotransplanted into normal dog's brain. Morphological changes were observed under light and transmission electron microscope. The islets and apoptosis of implanted B cells were identified by insulin-TUNEL double staining.RESULTS: The BSM used in our study had a favorable permeability, some degree of rigidity, lighter foreign body reaction and toxicity. The grafts consisted of epithelioid cells and loose connective tissue. Severe infiltration of inflammatory cells was not observed. The implanted ICCs were identified 2 mo later and showed typical apoptosis.CONCLUSION: BSM xenotransplantation in combination with the privileged site can inhibit the rejection of implanted heterogeneous ICCs, and death of implanted heterogeneous B cells is associated with apoptosis.
基金supported by the grants from National Natural Science Foundation of China(No.39993430-2,30000157)
文摘To lay background for studying rejection mechanisms in xenotransplantation and developing the strategies for intervention, class I genes of swine leukocyte antigens (SLA) of three Chinese pig strains Bm, Gz and Yn were cloned and sequenced. The eDNA of the class I loci P1 and P14 were amplified by RT-PCR and subjected to insert into sequencing vectors. All six allelic sequences we examined, each two for one Chinese strain, are not identical to those reported, which allows these novel sequences receiving their accession numbers AY102467- AY102472 from GenBank. This study further reveals that the homologies of MHC class I genes in their primary structures and the deduced amino acids between Chinese pigs (SLA) and human (HLA-A*0201) are better than those between pigs and mice (H-2Db/H-2Kb). The comparison also indicates that the amino acid residues critical for recognition by human KIRs are altered in the swine class I molecules. The amino acids responsible for binding human CD8 coreceptor are largely conserved although there are two critical residues substituted. A functional test indicated that the human T cells specific for the prokaryotically expressed SLA P1protein could respond quite well in vitro to the class I-positive swine chondrocytes and PBMCs in presence of human APCs. This implies that, due to the substitution of two critical residues, the inaccessibility of human CD8 coreceptor to swine class I molecule might be contributable to the indirect pathway that the human T cells have to use for recognizing the SLA class I xenogeneic antigens.
文摘Objective: 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, D0 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, D0 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 D0 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 D0 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 endothelium of the graft.
基金supported by the National Natural Science Foundation of China (No. 81200380)
文摘Patients with FLT3-ITDmut /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-ITDmut /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-ITDmut /NPM1 CN-AML primary cells. The FLT3-ITDmut /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 generation 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 established xenotransplantation model of human FLT3-ITDmut /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.
基金supported by the National Key Research and Development Program of China(2022YFA1105404,2021YFF0702601 and 2021YFA0805300)the Research Unit of Generation of Large Animal Disease Models,Chinese Academy of Medical Sciences(2019I2M-5-025)+3 种基金the Science and Technology Program of Guangzhou(202201010409)the Key Research&Development Program of Hainan Province(ZDYF2021SHFZ052)the Major Science and Technology Project of Hainan Province(ZDKJ2021030)the 2020 Research Program of Sanya Yazhou Bay Science and Technology City(202002011)
文摘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.
文摘目的:研究溶质载体家族6成员9(solute carrier family 6 member 9,SLC6A9)表达对结直肠癌细胞增殖、迁移和5-氟尿嘧啶(5-fluorouracil,5-FU)药物敏感性的影响。方法:TCGA数据库分析、实时荧光定量PCR和Western blot分析检测SLC6A9在结肠癌组织、正常结肠细胞系(NCM460)和结直肠癌细胞系(SW620、HCT116、HT29、Lovo和SW480)中的表达。将SCL6A9过表达质粒及阴性对照(SLC6A9 OE、Vector)转染HT29细胞,将SCL6A9小干扰RNA及阴性对照(SLC6A9 siRNA1#、siRNA2#和Scramble)转染SW620细胞。划痕愈合实验和Transwell实验检测各组细胞的迁移、侵袭能力。Western blot和细胞免疫荧光检测EMT相关蛋白E-cadherin、Vimentin的表达水平。利用CCK-8法和构建裸鼠移植瘤模型检测SLC6A9过表达对结直肠癌细胞5-FU药物敏感性的影响。结果:与正常结肠组织和NCM460细胞相比,SLC6A9在结肠癌组织和结直肠癌细胞系中低表达(均P<0.05)。SLC6A9过表达引起E-cadherin蛋白表达增加,Vimentin蛋白水平降低,抑制结直肠癌细胞的迁移、侵袭(P<0.05)。SLC6A9低表达引起E-cadherin蛋白表达降低,Vimentin蛋白水平增加,促进结直肠癌细胞的迁移、侵袭能力(P<0.05)。SLC6A9过表达提高了5-FU的药物敏感性,并使肿瘤生长缓慢,质量减轻(P<0.05)。而SLC6A9低表达降低了5-FU的药物敏感性(P<0.05)。结论:SLC6A9过表达能够抑制结直肠癌细胞的迁移、侵袭和EMT进程,并增强5-FU对结直肠癌细胞的药物敏感性。
基金the National Basic Research Program of China(2015CB554100)the National Key Research and Development Program of China(2017YFC1103703)+2 种基金the National Natural Science Foundation of China(81870446,81671838,81670593,81900571)Natural Science Foundation of Shaanxi Province(2016JM8026,2020JQ451)Academic Assistant Program of Xijing Hospital(XJZT18MJ29,XJZT18MJ27)。
文摘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.
基金the Science and Technology Development Fund Project of Shenzhen,China(JCYJ 20140415162338852)Scientific Research Project of Shenzhen health family planning,China(201401038)Natural Science Foundation of Guangdong Province,China(2015A030313889).
文摘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.
