New insights into xenotransplantation for cartilage repair: porcine multi-genetically modified chondrocytes as a promising cell source
| Autor: | Markus Huber-Lang, Jörg Fiedler, Angelika Schnieke, Rolf E. Brenner, Rebecca Halbgebauer, Konrad Fischer, Jochen Seissler, Hanna Tritschler |
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| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
Cartilage
Articular CD55 antigen Swine medicine.medical_treatment Gene Expression Epitope Animals Genetically Modified Gene Knockout Techniques DDC 570 / Life sciences xenotransplantation complement Biology (General) CD46 Cells Cultured TNFAIP3 α-1 3-Gal-epitope Xenografts CD55 Antigens Chemistry heme oxygenase-1 General Medicine Komplementierung medicine.anatomical_structure porcine chondrocytes Bone Diseases CD55 CD59 Neu5Gc-epitope QH301-705.5 Transgene Xenotransplantation Transplantation Heterologous CD59 Antigens Chondrocyte Article Membrane Cofactor Protein Chondrocytes ddc:570 medicine Animals Humans ddc:610 Tumor Necrosis Factor alpha-Induced Protein 3 Complement System Proteins Molecular biology Heterotransplantation Complement system Transplantation DDC 610 / Medicine & health |
| Zdroj: | Cells, Vol 10, Iss 2152, p 2152 (2021) Cells Volume 10 Issue 8 |
| Popis: | Transplantation of xenogenic porcine chondrocytes could represent a future strategy for the treatment of human articular cartilage defects. Major obstacles are humoral and cellular rejection processes triggered by xenogenic epitopes like α-1,3-Gal and Neu5Gc. Besides knockout (KO) of genes responsible for the biosynthesis of respective epitopes (GGTA1 and CMAH), transgenic expression of human complement inhibitors and anti-apoptotic as well as anti-inflammatory factors (CD46, CD55, CD59, TNFAIP3 and HMOX1) could synergistically prevent hyperacute xenograft rejection. Therefore, chondrocytes from different strains of single- or multi-genetically modified pigs were characterized concerning their protection from xenogeneic complement activation. Articular chondrocytes were isolated from the knee joints of WT, GalTKO, GalT/CMAH-KO, human CD59/CD55//CD46/TNFAIP3/HMOX1-transgenic (TG), GalTKO/TG and GalT/CMAHKO/TG pigs. The tissue-specific effectiveness of the genetic modifications was tested on gene, protein and epitope expression level or by functional assays. After exposure to 20% and 40% normal human serum (NHS), deposition of C3b/iC3b/C3c and formation of the terminal complement complex (TCC, C5b-9) was quantified by specific cell ELISAs, and generation of the anaphylatoxin C5a by ELISA. Chondrocyte lysis was analyzed by Trypan Blue Exclusion Assay. In all respective KO variants, the absence of α -1,3-Gal and Neu5Gc epitope was verified by FACS analysis. In chondrocytes derived from TG animals, expression of CD55 and CD59 could be confirmed on gene and protein level, TNFAIP3 on gene expression level as well as by functional assays and CD46 only on gene expression level whereas transgenic HMOX1 expression was not evident. Complement activation in the presence of NHS indicated mainly effective although incomplete protection against C3b/iC3b/C3c deposition, C5a-generation and C5b-9 formation being lowest in single GalTKO. Chondrocyte viability under exposure to NHS was significantly improved even by single GalTKO and completely preserved by all other variants including TG chondrocytes without KO of xenoepitopes. publishedVersion |
| Databáze: | OpenAIRE |
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