iPSC modeling of severe aplastic anemia reveals impaired differentiation and telomere shortening in blood progenitors

Abstract

Aplastic Anemia (AA) is a bone marrow failure (BMF) disorder, resulting in bone marrow hypocellularity and peripheral pancytopenia. Severe aplastic anemia (SAA) is a subset of AA defined by a more severe phenotype. Although the immunological nature of SAA pathogenesis is widely accepted, there is an increasing recognition of the role of dysfunctional hematopoietic stem cells in the disease phenotype. While pediatric SAA can be attributable to genetic causes, evidence is evolving on previously unrecognized genetic etiologies in a proportion of adults with SAA. Thus, there is an urgent need to better understand the pathophysiology of SAA, which will help to inform the course of disease progression and treatment options. We have derived induced pluripotent stem cell (iPSC) from three unaffected controls and three SAA patients and have shown that this in vitro model mimics two key features of the disease: (1) the failure to maintain telomere length during the reprogramming process and hematopoietic differentiation resulting in SAA-iPSC and iPSC-derived-hematopoietic progenitors with shorter telomeres than controls; (2) the impaired ability of SAA-iPSC-derived hematopoietic progenitors to give rise to erythroid and myeloid cells. While apoptosis and DNA damage response to replicative stress is similar between the control and SAA-iPSC-derived-hematopoietic progenitors, the latter show impaired proliferation which was not restored by eltrombopag, a drug which has been shown to restore hematopoiesis in SAA patients. Together, our data highlight the utility of patient specific iPSC in providing a disease model for SAA and predicting patient responses to various treatment modalities.

Disrupted alternative splicing for genes implicated in splicing and ciliogenesis causes PRPF31 retinitis pigmentosa

“…Hereafter, all patients and derived cells are referred to as RP11 accompanied by M (moderate), S (severe) and VS (very severe). Three unaffected controls are referred to as WT1 (wild type), WT2 and WT3 (Supplementary Data 1 ) 25 , 26 . Dermal skin fibroblasts were reprogrammed to iPSCs using a non-integrative RNA-based Sendai virus (Supplementary Figure 1A ).…”

Laminin γ3 plays an important role in retinal lamination, photoreceptor organisation and ganglion cell differentiation

“…Retinal organoids were generated as described previously with some modifications 37 . Briefly, hESCs (H9; WiCell Inc.) and hiPSCs derived from adult fibroblast (SB-Ad3 45 ) were expanded in mTESR™1 (StemCell Technologies, 05850) on growth factor-reduced Matrigel (BD Biosciences, San Jose, CA) coated plates at 37 °C and 5% CO 2 . In contrast to the protocol described by Mellough et al 37 , the medium was supplemented with 10 µM Rock inhibitor (Y-27632 dihydrochloride; Chemdea, Ridgewood, USA) for the first 2 days of differentiation and with 10% foetal calf serum (FCS; Life Technologies, UK), T 3 (40 ng/ml; Sigma-Aldrich, UK), Taurine (0.1 mM; Sigma-Aldrich) and retinoic acid (0.5 µM; Sigma-Adrich) after day 18 of differentiation.…”

Acquired Aplastic Anemia as a Clonal Disorder of Hematopoietic Stem Cells

“…Peripheral granulocyte and mononuclear cell telomere shortening are found in approximately 35% of AA patients indicating a defect in the HSPCs [52,53]. Furthermore, to evaluate the telomeres and their impact on dysfunctional hematopoietic stem cells in the AA phenotype, in in vitro model mimics two typical features of the AA were built [54]. Not only did AA patients have shorter telomere length compared to the health control, but also AA patients with somatic mutations had shorter telomere lengths, compared with patients without mutations [55].…”