Novel improvements to the differentiation of human iPS and ES cells for the derivation of pancreatic beta cells

Type 1 diabetes is caused by pancreatic β cell destruction due to autoimmunity. The prospect of utilizing hESC or hiPSC derived [beta] cells as cell replacement therapy is a promising potential cure, but proof of principle in animal models has yet to be achieved. In this study, we sought to improve the D'Amour et al., 2006 protocol for deriving [beta] cells from hESCs and hiPSCs by targeting the duration of the developmental stages in hiPSCs, testing small molecules for their effect on the generation of pancreatic hormone progenitors in both hESCs and hiPSCs, and transplanting differentiated hiPSCs into immune-suppressed mice to evaluate their ability to secrete insulin in vivo. We found that by optimizing the duration of the developmental stages of the protocol for the hiPSCs, we could generate higher levels of hormone- producing cells by the end of the protocol. We also found that the addition of the Src family kinase inhibitor, PP2, improved the generation of NGN3+ hormone progenitors and the eventual yield of hormone-producing cells. Though we were able to improve the in vitro yields of endocrine progenitors and the eventual yield of hormone-producing cells, after transplantation, the cells produced teratomas in about 50% of the mice that received them and no C- peptide release was found, up to four months after transplantation. While our studies have found novel ways to improve the differentiation protocol, further studies are needed to test methods for reducing teratoma formation and improve endocrine progenitor survival after transplantatio

Novel improvements to the differentiation of human iPS and ES cells for the derivation of pancreatic beta cells

Type 1 diabetes is caused by pancreatic β cell destruction due to autoimmunity. The prospect of utilizing hESC or hiPSC derived [beta] cells as cell replacement therapy is a promising potential cure, but proof of principle in animal models has yet to be achieved. In this study, we sought to improve the D'Amour et al., 2006 protocol for deriving [beta] cells from hESCs and hiPSCs by targeting the duration of the developmental stages in hiPSCs, testing small molecules for their effect on the generation of pancreatic hormone progenitors in both hESCs and hiPSCs, and transplanting differentiated hiPSCs into immune-suppressed mice to evaluate their ability to secrete insulin in vivo. We found that by optimizing the duration of the developmental stages of the protocol for the hiPSCs, we could generate higher levels of hormone- producing cells by the end of the protocol. We also found that the addition of the Src family kinase inhibitor, PP2, improved the generation of NGN3+ hormone progenitors and the eventual yield of hormone-producing cells. Though we were able to improve the in vitro yields of endocrine progenitors and the eventual yield of hormone-producing cells, after transplantation, the cells produced teratomas in about 50% of the mice that received them and no C- peptide release was found, up to four months after transplantation. While our studies have found novel ways to improve the differentiation protocol, further studies are needed to test methods for reducing teratoma formation and improve endocrine progenitor survival after transplantatio

C9orf72 BAC Transgenic Mice Display Typical Pathologic Features of ALS/FTD.

Noncoding expansions of a hexanucleotide repeat (GGGGCC) in the C9orf72 gene are the most common cause of familial amyotrophic lateral sclerosis and frontotemporal dementia. Here we report transgenic mice carrying a bacterial artificial chromosome (BAC) containing the full human C9orf72 gene with either a normal allele (15 repeats) or disease-associated expansion (∼100-1,000 repeats; C9-BACexp). C9-BACexp mice displayed pathologic features seen in C9orf72 expansion patients, including widespread RNA foci and repeat-associated non-ATG (RAN) translated dipeptides, which were suppressed by antisense oligonucleotides targeting human C9orf72. Nucleolin distribution was altered, supporting that either C9orf72 transcripts or RAN dipeptides promote nucleolar dysfunction. Despite early and widespread production of RNA foci and RAN dipeptides in C9-BACexp mice, behavioral abnormalities and neurodegeneration were not observed even at advanced ages, supporting the hypothesis that RNA foci and RAN dipeptides occur presymptomatically and are not sufficient to drive neurodegeneration in mice at levels seen in patients. Neuron 2015 Dec 2; 88(5):892-901