Journal of Vaccine Research 2024, Vol.14, No.3, 120-134 http://medscipublisher.com/index.php/jvr 123 the transplanted organ. These genetic modifications collectively contribute to longer graft survival and improved transplant outcomes (Coe et al., 2020). 4 Key Genetic Modifications in Pig Organs 4.1 Immunomodulatory genes (e.g., GGTA1, CMAH, β4GalNT2) In xenotransplantation, immune rejection is a major barrier. To address this issue, scientists have edited multiple immunomodulatory genes. The GGTA1 gene encodes α1,3-galactosyltransferase (α-Gal), a major xenoantigen. Human immune systems recognize and attack cells expressing α-Gal, leading to hyperacute rejection. Knocking out the GGTA1gene can eliminate α-Gal expression, significantly reducing the risk of hyperacute rejection. The CMAHgene encodes N-glycolylneuraminic acid hydroxylase, producing the Neu5Gc antigen, a key factor in immune response. Knocking out CMAH can reduce human natural antibody binding, thus lowering the potential for immune rejection. Similarly, the β4GalNT2 gene encodes β-1,4-N-acetylgalactosaminyltransferase, producing the Sda antigen, which also plays a role in immune reactions. Eliminating these genes significantly improves graft survival and compatibility(Cooper et al., 2019). Li et al. (2021) successfully used CRISPR/Cas9 technology to knock out GGTA1, CMAH, and β4GalNT2 in pig endothelial cells. These modified cells showed lower antigenicity in vitro, demonstrating the potential of multigene editing in xenotransplantation (Li et al., 2021). By targeting these key immunomodulatory genes, the immune response against pig organs in humans is significantly reduced, making xenotransplantation a more viable and effective solution. 4.2 Anti-inflammatory and anti-apoptotic genes To enhance graft tolerance to the immune system, various anti-inflammatory and anti-apoptotic genes have been introduced. These genes not only reduce immune attacks on the graft but also improve its post-transplant survival. The human heme oxygenase-1 (HO-1) gene, for example, enhances graft survival through its antioxidant and anti-inflammatory properties. Ahrens et al. (2015) integrated HO-1 into GGTA1 knockout pigs, and these transgenic kidneys exhibited significant resistance to rejection and ischemia-reperfusion injury during ex vivo perfusion with human blood. Anti-apoptotic genes such as A20 and Bcl-2 have also been incorporated into pig genomes to inhibit apoptosis and extend graft survival. The A20 gene encodes a protein with anti-inflammatory effects, regulating the NF-κB signaling pathway to reduce inflammation in the graft. Cooper et al. (2019) introduced A20 and other anti-inflammatory genes into transgenic pigs, resulting in significantly improved graft survival in non-human primates. Heat shock protein 70 (HSP70) has also been investigated to enhance graft survival. Hryhorowicz et al. (2017) found that HSP70 expression reduced cellular stress responses and protected cells, demonstrating significant benefits for graft survival. Incorporating anti-inflammatory and anti-apoptotic genes can substantially improve graft survival and functionality, providing robust protection against immune attacks and harsh transplant environments (Hryhorowicz et al., 2017). 4.3 Genes promoting resistance to ischemia-reperfusion injury Ischemia-reperfusion injury (IRI) is a common complication post-transplantation, severely affecting graft function and survival. To mitigate this damage, several genes have been introduced to enhance resistance to IRI. The heme oxygenase-1 (HO-1) gene, for instance, has proven effective in reducing oxidative stress and inflammation. Fischer et al. (2016) used gene editing to integrate HO-1 into multi-gene modified pigs, resulting in organs with improved resistance to IRI post-transplant. The A20 gene plays a crucial role in reducing apoptosis and inflammation. By regulating the NF-κB signaling pathway, A20 reduces inflammatory responses, protecting grafts from IRI. Cooper et al. (2019) introduced A20 along with other anti-inflammatory genes into transgenic pigs, leading to improved graft survival in non-human primates.
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