Establishment of iPSC lines and zebrafish with loss-of-function AHDC1 variants: Models for Xia-Gibbs syndrome.

Xia-Gibbs syndrome (XGS) is a syndromic form of intellectual disability caused by heterozygous AHDC1 variants, but the pathophysiological mechanisms underlying this syndrome are still unclear. In this manuscript, we describe the development of two different functional models: three induced pluripotent stem cell (iPSC) lines with different loss-of-function (LoF) AHDC1 variants, derived by reprogramming peripheral blood mononuclear cells from XGS patients, and a zebrafish strain with a LoF variant in the ortholog gene (ahdc1) obtained through CRISPR/Cas9-mediated editing. The three iPSC lines showed expression of pluripotency factors (SOX2, SSEA-4, OCT3/4, and NANOG). To verify the capacity of iPSC to differentiate into the three germ layers, we obtained embryoid bodies (EBs), induced their differentiation, and confirmed the mRNA expression of ectodermal, mesodermal, and endodermal markers using the TaqMan hPSC Scorecard. The iPSC lines were also approved for the following quality tests: chromosomal microarray analysis (CMA), mycoplasma testing, and short tandem repeat (STR) DNA profiling. The zebrafish model has an insertion of four base pairs in the ahdc1 gene, is fertile, and breeding between heterozygous and wild-type (WT) animals generated offspring in a genotypic proportion in agreement with Mendelian law. The established iPSC and zebrafish lines were deposited on the hpscreg.eu and zfin.org platforms, respectively. These biological models are the first for XGS and will be used in future studies that investigate the pathophysiology of this syndrome, unraveling its underlying molecular mechanisms.

Induced Pluripotent Stem Cell for the Study and Treatment of Sickle Cell Anemia.

Sickle cell anemia (SCA) is a monogenic disease of high mortality, affecting millions of people worldwide. There is no broad, effective, and safe definitive treatment for SCA, so the palliative treatments are the most used. The establishment of an in vitro model allows better understanding of how the disease occurs, besides allowing the development of more effective tests and treatments. In this context, iPSC technology is a powerful tool for basic research and disease modeling, and a promise for finding and screening more effective and safe drugs, besides the possibility of use in regenerative medicine. This work obtained a model for study and treatment of SCA using iPSC. Then, episomal vectors were used for reprogramming peripheral blood mononuclear cells to obtain integration-free iPSC. Cells were collected from patients treated with hydroxyurea and without treatment. The iPSCP Bscd lines were characterized for pluripotent and differentiation potential. The iPSC lines were differentiated into HSC, so that we obtained a dynamic and efficient protocol of CD34+CD45+ cells production. We offer a valuable tool for a better understanding of how SCA occurs, in addition to making possible the development of more effective drugs and treatments and providing better understanding of widely used treatments, such as hydroxyurea.