Following the publication of Liu et al.(2024),an error was identified in Figure 4B,in which the image representing the lung from the E529G group was inadvertently duplicated with the image of the lung from the WT grou...Following the publication of Liu et al.(2024),an error was identified in Figure 4B,in which the image representing the lung from the E529G group was inadvertently duplicated with the image of the lung from the WT group during figure preparation.展开更多
Background:Pig organ xenotransplantation is a potential solution for the severe organ shortage in clinic,while immunogenic genes need to be eliminated to improve the immune compatibility between humans and pigs.Curren...Background:Pig organ xenotransplantation is a potential solution for the severe organ shortage in clinic,while immunogenic genes need to be eliminated to improve the immune compatibility between humans and pigs.Current knockout strategies are mainly aimed at the genes causing hyperacute immune rejection(HAR)that occurs in the first few hours while adaptive immune reactions orchestrated by CD4 T cell thereafter also cause graft failure,in which process the MHCⅡmolecule plays critical roles.Methods:Thus,we generate a 4-gene(GGTA1,CMAH,β4GalNT2,and CIITA)knockout pig by CRISPR/Cas9 and somatic cell nuclear transfer to compromise HAR and CD4 T cell reactions simultaneously.Results:We successfully obtained 4KO piglets with deficiency in all alleles of genes,and at cellular and tissue levels.Additionally,the safety of our animals after gene editing was verified by using whole-genome sequencing and karyotyping.Piglets have survived for more than one year in the barrier,and also survived for more than 3 months in the conventional environment,suggesting that the piglets without MHCⅡcan be raised in the barrier and then gradually mated in the conventional environment.Conclusions:4KO piglets have lower immunogenicity,are safe in genomic level,and are easier to breed than the model with both MHCⅠandⅡdeletion.展开更多
Because of their physiological similarity to humans, pigs provide an excellent model for the study of obesity. This study evaluated diet-induced adiposity in genetically lean pigs and found that body weight and energy...Because of their physiological similarity to humans, pigs provide an excellent model for the study of obesity. This study evaluated diet-induced adiposity in genetically lean pigs and found that body weight and energy intake did not differ between controls and pigs fed the high-fat （HF） diet for three months. However, fat mass percentage, adi- pocyte size, concentrations of total cholesterol （TC）, triglyceride （TG）, high-density lipoprotein cholesterol （HDL-C） and low-density lipoprotein cholesterol （LDL-C）, insulin, and leptin in plasma were significantly higher in HF pigs than in controls. The HF diet increased the expression in backfat tissue of genes responsible for cholesterol synthesis such as Insig-1 and Insig-2. Lipid metabolism-related genes including sterol regulatory element binding protein lc （SREBP-lc）, fatty acid synthase 1 （FASN1）, diacylglycerol O-acyltransferase 2 （DGAT2）, and fatty acid binding protein 4 （FABP4） were significantly up-regulated in backfat tissue, while the expression of proliferator-activated receptor-α（PPAR-α） and carnitine palmitoyl transferase 2 （CPT2）, both involved in fatty acid oxidation, was reduced. In liver tissue, HF feeding significantly elevated the expression of SREBP-lc, FASN1, DGAT2, and hepatocyte nuclear factor-4α （HNF-4α） mRNAs. Microarray analysis further showed that the HF diet had a significant effect on the expression of 576 genes. Among these, 108 genes were related to 21 pathways, with 20 genes involved in adiposity deposition and 26 related to immune response. Our results suggest that an HF diet can induce genetically lean pigs into obesity with body fat mass expansion and adipose-related inflammation.展开更多
Recombinant adeno-associated virus （rAAV） vectors have been extensively used for experimental gene therapy of inherited human diseases. Several advantages, such as simple vector construction, high targeting frequenc...Recombinant adeno-associated virus （rAAV） vectors have been extensively used for experimental gene therapy of inherited human diseases. Several advantages, such as simple vector construction, high targeting frequency by homologous recombination, and applica- bility to many cell types, make rAAV an attractive approach for targeted genome editing. Combined with cloning by somatic cell nuclear transfer （SCNT）, this technology has recently been successfully adapted to generate gene-targeted pigs as models for cystic fibrosis, hereditary tyrosinemia type 1, and breast cancer. This review summarizes the development of rAAV for targeted genome editing in mammalian cells and provides strategies for enhancing the rAAV-mediated targeting frequency by homologous recombination. We discuss current development and application of the rAAV vectors for targeted genome editing in porcine primary fibroblasts, which are subse- quently used as donor cells for SCNT to generate cloned genetically designed pigs and provide positive perspectives for the generation of gene-targeted pigs with rAAV in the future.展开更多
Objective: TO investigate the killing differences of human blood group antibodies IgG and IgM on PBMC and RBC in peripheral blood of genetically modified pigs, and provide theoretical basis for the selection of pig bl...Objective: TO investigate the killing differences of human blood group antibodies IgG and IgM on PBMC and RBC in peripheral blood of genetically modified pigs, and provide theoretical basis for the selection of pig blood group for clinical application of porcine xenotransplantation. Methods: Serum samples were collected from 20 healthy subjects, 20 patients with end‑stage renal disease and 20 brain dead organ donors, and divided into 4 groups on the basis of ABO blood group (A: n=20;B: n=17;AB: n=7;O: n=16). Flow cytometry was used to detect antibody binding or complement dependent cytotoxicity test (CDC) between human serum and O blood group Wild‑type (WT), α1, 3‑galactosyltransferase gene‑knockout(GTKO), cytidine monophosphate‑N‑acetylneuraminic acid hydroxylase gene‑knockout (GTKO/β4GalNT2KO), β‑1, 4N‑acetylgalactosaminyltransferase gene‑knockout(GTKO/CMAHKO) and mononuclear cells (PBMC) and red blood cells (RBC) of Triple knockout (TKO/hCD55) porcine peripheral blood, respectively. Results: There was no significant difference in binding and killing of human serum antibodies with blood group A, B, AB and O on PBMC of WT pigs, GTKO pigs, GTKO/β4GalNT2KO pigs, GTKO/CMAHKO pigs and TKO/hCD55 pigs, respectively. There was no significant difference in RBC binding with RBC of WT pig, GTKO pig, GTKO/β4GalNT2KO pig, GTKO/CMAHKO pig and TKO/hCD55 pig. Conclusion: The selection of recipients of pigs with type O blood group can be done without considering blood group.展开更多
[Objective] The genetic background of four types of fragrance pigs was studied using microsatellite molecular markers,in order to fully understand the genetic resources of miniature pigs in China.[Method] Using 27 pai...[Objective] The genetic background of four types of fragrance pigs was studied using microsatellite molecular markers,in order to fully understand the genetic resources of miniature pigs in China.[Method] Using 27 pairs of microsatellite loci jointly recommended by Food and Agricultural Organization（FAO）and International Society for Animal Genetics（ISAG）,we detected the genotypes of 200 fragrance individuals belonging to four types（Jiuyang fragrance pig,Jianbai fragrance pig,Congjiang fragrance pig and Huanjiang fragrance pig）,and analyzed their Inter-and intra-breed genetic variations.[Result]The 23 loci detected in the test were high polymorphic;the mean heterozygosity（H） of Jiuyang fragrance pig,Jianbai fragrance pig,Congjiang fragrance pig and Huanjiang fragrance pig were 0.683 6,0.667 9,0.697 3 and 0.702 2,and their mean polymorphism information contents（PIC） were 0.6263,0.6063,0.6420 and 0.6415,respectively.[Conclusion]Four types of fragrance pigs detected in the test all had high intra-breed genetic variability.展开更多
文摘Following the publication of Liu et al.(2024),an error was identified in Figure 4B,in which the image representing the lung from the E529G group was inadvertently duplicated with the image of the lung from the WT group during figure preparation.
基金National Key Research and Development Program,Grant/Award Number:2019YFA0903800,2021YFA0805701,2021YFA0805905 and 2022YFA1103603CAS Project for Young Scientists in Basic Research,Grant/Award Number:YSBR-012+2 种基金STI 2030-Major Project,Grant/Award Number:2023ZD0407503National Natural Science Foundation of China,Grant/Award Number:32071456 and 82241224Strategic Priority Research Program of the Chinese Academy of Sciences,Grant/Award Number:XDA16030000。
文摘Background:Pig organ xenotransplantation is a potential solution for the severe organ shortage in clinic,while immunogenic genes need to be eliminated to improve the immune compatibility between humans and pigs.Current knockout strategies are mainly aimed at the genes causing hyperacute immune rejection(HAR)that occurs in the first few hours while adaptive immune reactions orchestrated by CD4 T cell thereafter also cause graft failure,in which process the MHCⅡmolecule plays critical roles.Methods:Thus,we generate a 4-gene(GGTA1,CMAH,β4GalNT2,and CIITA)knockout pig by CRISPR/Cas9 and somatic cell nuclear transfer to compromise HAR and CD4 T cell reactions simultaneously.Results:We successfully obtained 4KO piglets with deficiency in all alleles of genes,and at cellular and tissue levels.Additionally,the safety of our animals after gene editing was verified by using whole-genome sequencing and karyotyping.Piglets have survived for more than one year in the barrier,and also survived for more than 3 months in the conventional environment,suggesting that the piglets without MHCⅡcan be raised in the barrier and then gradually mated in the conventional environment.Conclusions:4KO piglets have lower immunogenicity,are safe in genomic level,and are easier to breed than the model with both MHCⅠandⅡdeletion.
基金Project supported by the National Key Research and Development Program of China(Nos.2018YFD0500400 and 2018YFD0501100)the National Basic Research Program(973)of China(No.2013CB127304)+1 种基金the China Agriculture Research System(No.CARS-36)and the National Natural Science Foundation of China(No.31402086)
文摘Because of their physiological similarity to humans, pigs provide an excellent model for the study of obesity. This study evaluated diet-induced adiposity in genetically lean pigs and found that body weight and energy intake did not differ between controls and pigs fed the high-fat （HF） diet for three months. However, fat mass percentage, adi- pocyte size, concentrations of total cholesterol （TC）, triglyceride （TG）, high-density lipoprotein cholesterol （HDL-C） and low-density lipoprotein cholesterol （LDL-C）, insulin, and leptin in plasma were significantly higher in HF pigs than in controls. The HF diet increased the expression in backfat tissue of genes responsible for cholesterol synthesis such as Insig-1 and Insig-2. Lipid metabolism-related genes including sterol regulatory element binding protein lc （SREBP-lc）, fatty acid synthase 1 （FASN1）, diacylglycerol O-acyltransferase 2 （DGAT2）, and fatty acid binding protein 4 （FABP4） were significantly up-regulated in backfat tissue, while the expression of proliferator-activated receptor-α（PPAR-α） and carnitine palmitoyl transferase 2 （CPT2）, both involved in fatty acid oxidation, was reduced. In liver tissue, HF feeding significantly elevated the expression of SREBP-lc, FASN1, DGAT2, and hepatocyte nuclear factor-4α （HNF-4α） mRNAs. Microarray analysis further showed that the HF diet had a significant effect on the expression of 576 genes. Among these, 108 genes were related to 21 pathways, with 20 genes involved in adiposity deposition and 26 related to immune response. Our results suggest that an HF diet can induce genetically lean pigs into obesity with body fat mass expansion and adipose-related inflammation.
基金supported by the grants from the Danish National Researeh Infrastructure Programme to the Danish Genetieally Modified Animal Resource(DAG- MAR)as well as from the"Sino一Danish Breast Caneer Research Centre"under the ausPiees of the Danish National Researeh Foundation(Grundforskningsfonden)the National Natural Seience Foundation of China
文摘Recombinant adeno-associated virus （rAAV） vectors have been extensively used for experimental gene therapy of inherited human diseases. Several advantages, such as simple vector construction, high targeting frequency by homologous recombination, and applica- bility to many cell types, make rAAV an attractive approach for targeted genome editing. Combined with cloning by somatic cell nuclear transfer （SCNT）, this technology has recently been successfully adapted to generate gene-targeted pigs as models for cystic fibrosis, hereditary tyrosinemia type 1, and breast cancer. This review summarizes the development of rAAV for targeted genome editing in mammalian cells and provides strategies for enhancing the rAAV-mediated targeting frequency by homologous recombination. We discuss current development and application of the rAAV vectors for targeted genome editing in porcine primary fibroblasts, which are subse- quently used as donor cells for SCNT to generate cloned genetically designed pigs and provide positive perspectives for the generation of gene-targeted pigs with rAAV in the future.
基金Key Science and Technology Project of Hainan Provincial Science and Technology Department (ZDKT2019009)Hainan Clinical Medical Center construction project.
文摘Objective: TO investigate the killing differences of human blood group antibodies IgG and IgM on PBMC and RBC in peripheral blood of genetically modified pigs, and provide theoretical basis for the selection of pig blood group for clinical application of porcine xenotransplantation. Methods: Serum samples were collected from 20 healthy subjects, 20 patients with end‑stage renal disease and 20 brain dead organ donors, and divided into 4 groups on the basis of ABO blood group (A: n=20;B: n=17;AB: n=7;O: n=16). Flow cytometry was used to detect antibody binding or complement dependent cytotoxicity test (CDC) between human serum and O blood group Wild‑type (WT), α1, 3‑galactosyltransferase gene‑knockout(GTKO), cytidine monophosphate‑N‑acetylneuraminic acid hydroxylase gene‑knockout (GTKO/β4GalNT2KO), β‑1, 4N‑acetylgalactosaminyltransferase gene‑knockout(GTKO/CMAHKO) and mononuclear cells (PBMC) and red blood cells (RBC) of Triple knockout (TKO/hCD55) porcine peripheral blood, respectively. Results: There was no significant difference in binding and killing of human serum antibodies with blood group A, B, AB and O on PBMC of WT pigs, GTKO pigs, GTKO/β4GalNT2KO pigs, GTKO/CMAHKO pigs and TKO/hCD55 pigs, respectively. There was no significant difference in RBC binding with RBC of WT pig, GTKO pig, GTKO/β4GalNT2KO pig, GTKO/CMAHKO pig and TKO/hCD55 pig. Conclusion: The selection of recipients of pigs with type O blood group can be done without considering blood group.
基金Supported by Annual Experimental Animal Research Topics of Guizhou Science and Technology Department(2004JN0029)
文摘[Objective] The genetic background of four types of fragrance pigs was studied using microsatellite molecular markers,in order to fully understand the genetic resources of miniature pigs in China.[Method] Using 27 pairs of microsatellite loci jointly recommended by Food and Agricultural Organization（FAO）and International Society for Animal Genetics（ISAG）,we detected the genotypes of 200 fragrance individuals belonging to four types（Jiuyang fragrance pig,Jianbai fragrance pig,Congjiang fragrance pig and Huanjiang fragrance pig）,and analyzed their Inter-and intra-breed genetic variations.[Result]The 23 loci detected in the test were high polymorphic;the mean heterozygosity（H） of Jiuyang fragrance pig,Jianbai fragrance pig,Congjiang fragrance pig and Huanjiang fragrance pig were 0.683 6,0.667 9,0.697 3 and 0.702 2,and their mean polymorphism information contents（PIC） were 0.6263,0.6063,0.6420 and 0.6415,respectively.[Conclusion]Four types of fragrance pigs detected in the test all had high intra-breed genetic variability.
