Project description:Inherited mitochondrial DNA (mtDNA) diseases transmit maternally and cause severe phenotypes. Since no effective treatment or genetic screening is available, nuclear genome transfer between patients’ and healthy eggs to replace mutant mtDNAs holds promises. Since polar body contains very few mitochondria and share same genomic material as oocyte, here we perform polar body transfer to prevent the transmission of inherited mtDNA variants. We compare the value of different germline genome transfer (spindle-chromosome, pronuclear, first and second polar body) in a mouse model. Reconstructed embryos support normal fertilization and produce live offspring. Strikingly, genetic analysis confirms F1 generation after polar body transfer possesses minimal donor mtDNA carry-over compared with spindle-chromosome (low/medium carry-over) and pronuclear (medium/high carry-over) transfer. Moreover, mtDNA genotype remains stable in F2 generation of progeny after polar body transfer. Our preclinical model demonstrates polar body transfer holds great potential in preventing the transmission of inherited mtDNA diseases. The objective of the present study was to detect genomic aberrations between PB1 and its counterpart, spindle-chromosome complex in human MII oocyte, PB2 and female pronucleus in human zygote at a single-cell level.
Project description:Inherited mitochondrial DNA (mtDNA) diseases transmit maternally and cause severe phenotypes. Since no effective treatment or genetic screening is available, nuclear genome transfer between patients’ and healthy eggs to replace mutant mtDNAs holds promises. Since polar body contains very few mitochondria and share same genomic material as oocyte, here we perform polar body transfer to prevent the transmission of inherited mtDNA variants. We compare the value of different germline genome transfer (spindle-chromosome, pronuclear, first and second polar body) in a mouse model. Reconstructed embryos support normal fertilization and produce live offspring. Strikingly, genetic analysis confirms F1 generation after polar body transfer possesses minimal donor mtDNA carry-over compared with spindle-chromosome (low/medium carry-over) and pronuclear (medium/high carry-over) transfer. Moreover, mtDNA genotype remains stable in F2 generation of progeny after polar body transfer. Our preclinical model demonstrates polar body transfer holds great potential in preventing the transmission of inherited mtDNA diseases.
Project description:Although polar body transfer (PBT) has the potential to prevent the transmission of inherited mitochondrial DNA (mtDNA) diseases, the PBT technique is still at an early stage, as no human data are publicly available for PBT. Here, we investigated the comparative values of first and second PBT (PB1T, PB2T), spindle-chromosome and pronuclear transfer (ST, PNT), modified ST and PNT (mST and mPNT) to explore the efficiency and safety of these approaches. A comparative analysis confirmed that PB1T, mST, PB2T and mPNT could be used to donate mtDNA without resulting in significant heteroplasmy or alterations in the methylation profile and gene expression. Importantly, PB1T produced reconstructed embryos and embryonic stem cells (ESCs) in every generation with undetectable donor mtDNA. However, donor mtDNA seems to have a tendency to be amplified in generations of mPNT-ESCs with up to 3% heteroplasmy. These results suggest that PB1T holds great potential in eliminating mtDNA variants.
Project description:Although polar body transfer (PBT) has the potential to prevent the transmission of inherited mitochondrial DNA (mtDNA) diseases, the PBT technique is still at an early stage, as no human data are publicly available for PBT. Here, we investigated the comparative values of first and second PBT (PB1T, PB2T), spindle-chromosome and pronuclear transfer (ST, PNT), modified ST and PNT (mST and mPNT) to explore the efficiency and safety of these approaches. A comparative analysis confirmed that PB1T, mST, PB2T and mPNT could be used to donate mtDNA without resulting in significant heteroplasmy or alterations in the methylation profile and gene expression. Importantly, PB1T produced reconstructed embryos and embryonic stem cells (ESCs) in every generation with undetectable donor mtDNA. However, donor mtDNA seems to have a tendency to be amplified in generations of mPNT-ESCs with up to 3% heteroplasmy. These results suggest that PB1T holds great potential in eliminating mtDNA variants.
Project description:Here we report the derivation of human ESCs from the polar body thansfer reconstructed embryos. We choose three cell lines from all the cell lines and compare the DNA methylation state. We use human methylation chip to compare genomic DNA methylation level among three PB1 transfer human ES cell lines and three PB2 transfer human ES cell lines.
Project description:Currently there is no treatment for mitochondrial disease, a group of devastating inherited disorders caused by mutations in mitochondrial DNA (mtDNA). Here we report a strategy to prevent the germline transmission of mitochondrial diseases. This technique is based on the specific elimination of mutated mtDNA through the use of mitochondria targeted nucleases. Our approaches represent a potential therapeutic avenue for preventing the transgenerational transmission of human mitochondrial diseases caused by mutations in mtDNA. A total of 4 samples were analyzed. Test samples were compared to sex-matched reference samples.
