Cytokinesis failure occurs in Fanconi anemia pathway-deficient murine and human bone marrow hematopoietic cells.
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ABSTRACT:
SUBMITTER: Vinciguerra P
PROVIDER: S-EPMC2964984 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Fanconi anemia (FA) is an inherited DNA repair disorder characterized by progressive bone marrow failure (BMF) from hematopoietic stem and progenitor cell (HSPC) attrition. A greater understanding of the pathogenesis of BMF could improve the therapeutic options for FA patients. Using a genome-wide shRNA screen in human FA fibroblasts, we identify transforming growth factor-? (TGF-?) pathway-mediated growth suppression as a cause of BMF in FA. Blocking the TGF-? pathway improves the survival of FA cells and rescues the proliferative and functional defects of HSPCs derived from FA mice and FA patients. Inhibition of TGF-? signaling in FA HSPCs results in elevated homologous recombination (HR) repair with a concomitant decrease in non-homologous end-joining (NHEJ), accounting for the improvement in cellular growth. Together, our results suggest that elevated TGF-? signaling contributes to BMF in FA by impairing HSPC function and may be a potential therapeutic target for the treatment of FA.
| S-EPMC4860147 | biostudies-literature
Project description:BackgroundPatients with bone marrow failure and undiagnosed underlying Fanconi anemia may experience major toxicity if given standard-dose conditioning regimens for hematopoietic stem cell transplant. Due to clinical variability and/or potential emergence of genetic reversion with hematopoietic somatic mosaicism, a straightforward Fanconi anemia diagnosis can be difficult to make, and diagnostic strategies combining different assays in addition to classical breakage tests in blood may be needed.Design and methodsWe evaluated Fanconi anemia diagnosis on blood lymphocytes and skin fibroblasts from a cohort of 87 bone marrow failure patients (55 children and 32 adults) with no obvious full clinical picture of Fanconi anemia, by performing a combination of chromosomal breakage tests, FANCD2-monoubiquitination assays, a new flow cytometry-based mitomycin C sensitivity test in fibroblasts, and, when Fanconi anemia was diagnosed, complementation group and mutation analyses. The mitomycin C sensitivity test in fibroblasts was validated on control Fanconi anemia and non-Fanconi anemia samples, including other chromosomal instability disorders.ResultsWhen this diagnosis strategy was applied to the cohort of bone marrow failure patients, 7 Fanconi anemia patients were found (3 children and 4 adults). Classical chromosomal breakage tests in blood detected 4, but analyses on fibroblasts were necessary to diagnose 3 more patients with hematopoietic somatic mosaicism. Importantly, Fanconi anemia was excluded in all the other patients who were fully evaluated.ConclusionsIn this large cohort of patients with bone marrow failure our results confirmed that when any clinical/biological suspicion of Fanconi anemia remains after chromosome breakage tests in blood, based on physical examination, history or inconclusive results, then further evaluation including fibroblast analysis should be made. For that purpose, the flow-based mitomycin C sensitivity test here described proved to be a reliable alternative method to evaluate Fanconi anemia phenotype in fibroblasts. This global strategy allowed early and accurate confirmation or rejection of Fanconi anemia diagnosis with immediate clinical impact for those who underwent hematopoietic stem cell transplant.
| S-EPMC2663612 | biostudies-literature
Project description:Fanconi anemia is a complex heterogeneous genetic disorder with a high incidence of bone marrow failure, clonal evolution to acute myeloid leukemia and mesenchymal-derived congenital anomalies. Increasing evidence in Fanconi anemia and other genetic disorders points towards an interdependence of skeletal and hematopoietic development, yet the impact of the marrow microenvironment in the pathogenesis of the bone marrow failure in Fanconi anemia remains unclear. Here we demonstrated that mice with double knockout of both Fancc and Fancg genes had decreased bone formation at least partially due to impaired osteoblast differentiation from mesenchymal stem/progenitor cells. Mesenchymal stem/progenitor cells from the double knockout mice showed impaired hematopoietic supportive activity. Mesenchymal stem/progenitor cells of patients with Fanconi anemia exhibited similar cellular deficits, including increased senescence, reduced proliferation, impaired osteoblast differentiation and defective hematopoietic stem/progenitor cell supportive activity. Collectively, these studies provide unique insights into the physiological significance of mesenchymal stem/progenitor cells in supporting the marrow microenvironment, which is potentially of broad relevance in hematopoietic stem cell transplantation.
| S-EPMC5451333 | biostudies-literature
Project description:Hematopoietic stem cell (HSC) attrition is considered the key event underlying progressive BM failure (BMF) in Fanconi anemia (FA), the most frequent inherited BMF disorder in humans. However, despite major advances, how the cellular, biochemical, and molecular alterations reported in FA lead to HSC exhaustion remains poorly understood. Here, we demonstrated in human and mouse cells that loss-of-function of FANCA or FANCC, products of 2 genes affecting more than 80% of FA patients worldwide, is associated with constitutive expression of the transcription factor microphthalmia (MiTF) through the cooperative, unscheduled activation of several stress-signaling pathways, including the SMAD2/3, p38 MAPK, NF-κB, and AKT cascades. We validated the unrestrained Mitf expression downstream of p38 in Fanca-/- mice, which display hallmarks of hematopoietic stress, including loss of HSC quiescence, DNA damage accumulation in HSCs, and reduced HSC repopulation capacity. Importantly, we demonstrated that shRNA-mediated downregulation of Mitf expression or inhibition of p38 signaling rescued HSC quiescence and prevented DNA damage accumulation. Our data support the hypothesis that HSC attrition in FA is the consequence of defects in the DNA-damage response combined with chronic activation of otherwise transiently activated signaling pathways, which jointly prevent the recovery of HSC quiescence.
| S-EPMC7269568 | biostudies-literature
Project description:Inherited bone marrow failure syndromes (IBMFSs) are a heterogeneous group of disorders characterized by defective hematopoiesis, impaired stem cell function, and cancer susceptibility. Diagnosis of IBMFS presents a major challenge due to the large variety of associated phenotypes, and novel, clinically relevant biomarkers are urgently needed. Our study identified nuclear interaction partner of ALK (NIPA) as an IBMFS gene, as it is significantly downregulated in a distinct subset of myelodysplastic syndrome-type (MDS-type) refractory cytopenia in children. Mechanistically, we showed that NIPA is major player in the Fanconi anemia (FA) pathway, which binds FANCD2 and regulates its nuclear abundance, making it essential for a functional DNA repair/FA/BRCA pathway. In a knockout mouse model, Nipa deficiency led to major cell-intrinsic defects, including a premature aging phenotype, with accumulation of DNA damage in hematopoietic stem cells (HSCs). Induction of replication stress triggered a reduction in and functional decline of murine HSCs, resulting in complete bone marrow failure and death of the knockout mice with 100% penetrance. Taken together, the results of our study add NIPA to the short list of FA-associated proteins, thereby highlighting its potential as a diagnostic marker and/or possible target in diseases characterized by hematopoietic failure.
| S-EPMC7260023 | biostudies-literature
Project description:IntroductionFanconi anemia (FA) is an inherited disorder characterized by bone marrow failure, congenital malformations, and predisposition to malignancies. Alterations in hematopoietic stem cells (HSC) have been reported, but little is known regarding the bone marrow (BM) stroma. Thus, the characterization of Mesenchymal Stromal Cells (MSC) would help to elucidate their involvement in the BM failure.MethodsWe characterized MSCs of 28 FA patients (FA-MSC) before and after treatment (hematopoietic stem cell transplantation, HSCT; or gene therapy, GT). Phenotypic and functional properties were analyzed and compared with MSCs expanded from 26 healthy donors (HD-MSCs). FA-MSCs were genetically characterized through, mitomycin C-test and chimerism analysis. Furthermore, RNA-seq profiling was used to identify dysregulated metabolic pathways.ResultsOverall, FA-MSC had the same phenotypic and functional characteristics as HD-MSC. Of note, MSC-GT had a lower clonogenic efficiency. These findings were not confirmed in the whole FA patients' cohort. Transcriptomic profiling identified dysregulation in HSC self-maintenance pathways in FA-MSC (HOX), and was confirmed by real-time quantitative polymerase chain reaction (RT-qPCR).DiscussionOur study provides a comprehensive characterization of FA-MSCs, including for the first time MSC-GT and constitutes the largest series published to date. Interestingly, transcript profiling revealed dysregulation of metabolic pathways related to HSC self-maintenance. Taken together, our results or findings provide new insights into the pathophysiology of the disease, although whether these niche defects are involved in the hematopoietic defects seen of FA deserves further investigation.
| S-EPMC11307092 | biostudies-literature
Project description:FancD2 plays a central role in the human Fanconi anemia DNA damage response (DDR) pathway. Fancd2(-/-) mice exhibit many features of human Fanconi anemia including cellular DNA repair defects. Whether the DNA repair defect in Fancd2(-/-) mice results in radiologic changes in all cell lineages is unknown. We measured stress of hematopoiesis in long-term marrow cultures and radiosensitivity in clonogenic survival curves, as well as comet tail intensity, total antioxidant stores and radiation-induced gene expression in hematopoietic progenitor compared to bone marrow stromal cell lines. We further evaluated radioprotection by a mitochondrial-targeted antioxidant GS-nitroxide, JP4-039. Hematopoiesis longevity in Fancd2(-/-) mouse long-term marrow cultures was diminished and bone marrow stromal cell lines were radiosensitive compared to Fancd2(+/+) stromal cells (Fancd2(-/-) D0 = 1.4 ± 0.1 Gy, ñ = 5.0 ± 0.6 vs. Fancd2(+/+) D0 = 1.6 ± 0.1 Gy, ñ = 6.7 ± 1.6), P = 0.0124 for D0 and P = 0.0023 for ñ, respectively). In contrast, Fancd2(-/-) IL-3-dependent hematopoietic progenitor cells were radioresistant (D0 = 1.71 ± 0.04 Gy and ñ = 5.07 ± 0.52) compared to Fancd2(+/+) (D0 = 1.39 ± 0.09 Gy and ñ = 2.31 ± 0.85, P = 0.001 for D0). CFU-GM from freshly explanted Fancd2(-/-) marrow was also radioresistant. Consistent with radiosensitivity, irradiated Fancd2(-/-) stromal cells had higher DNA damage by comet tail intensity assay compared to Fancd2(+/+) cells (P < 0.0001), slower DNA damage recovery, lower baseline total antioxidant capacity, enhanced radiation-induced depletion of antioxidants, and increased CDKN1A-p21 gene transcripts and protein. Consistent with radioresistance, Fancd2(-/-) IL-3-dependent hematopoietic cells had higher baseline and post irradiation total antioxidant capacity. While, there was no detectable alteration of radiation-induced cell cycle arrest with Fancd2(-/-) stromal cells, hematopoietic progenitor cells showed reduced G2/M cell cycle arrest. The absence of the mouse Fancd2 gene product confers radiosensitivity to bone marrow stromal but not hematopoietic progenitor cells.
| S-EPMC3970166 | biostudies-literature
Project description:Fanconi anemia (FA) is an inherited DNA repair deficiency syndrome. FA patients undergo progressive bone marrow failure (BMF) during childhood, which frequently requires allogeneic hematopoietic stem cell transplantation. The pathogenesis of this BMF has been elusive to date. Here we found that FA patients exhibit a profound defect in hematopoietic stem and progenitor cells (HSPCs) that is present before the onset of clinical BMF. In response to replicative stress and unresolved DNA damage, p53 is hyperactivated in FA cells and triggers a late p21(Cdkn1a)-dependent G0/G1 cell-cycle arrest. Knockdown of p53 rescued the HSPC defects observed in several in vitro and in vivo models, including human FA or FA-like cells. Taken together, our results identify an exacerbated p53/p21 "physiological" response to cellular stress and DNA damage accumulation as a central mechanism for progressive HSPC elimination in FA patients, and have implications for clinical care.
| S-EPMC3392433 | biostudies-literature
Project description:Fanconi anemia (FA) is a severe hereditary disorder with defective DNA damage response and repair. It is characterized by phenotypes including progressive bone marrow failure (BMF), developmental abnormalities, and increased occurrence of leukemia and cancer. Recent studies in mice have suggested that the FA proteins might counteract aldehyde-induced genotoxicity in hematopoietic stem cells. Nearly half of the Japanese population carries a dominant-negative allele (rs671) of the aldehyde-catalyzing enzyme ALDH2 (acetaldehyde dehydrogenase 2), providing an opportunity to test this hypothesis in humans. We examined 64 Japanese FA patients, and found that the ALDH2 variant is associated with accelerated progression of BMF, while birth weight or the number of physical abnormalities was not affected. Moreover, malformations at some specific anatomic locations were observed more frequently in ALDH2-deficient patients. Our current data indicate that the level of ALDH2 activity impacts pathogenesis in FA, suggesting the possibility of a novel therapeutic approach.
| S-EPMC3953058 | biostudies-literature
Project description:PurposeThe inherited bone marrow failure syndromes (IBMFS) are a heterogeneous group of genetic disorders that share the inability of the bone marrow to produce an adequate number of blood cells. The 4 most frequent syndromes are Fanconi anemia (FA), dyskeratosis congenita (DC), Diamond-Blackfan anemia (DBA), and Shwachman-Diamond syndrome (SDS). All 4 syndromes have been associated with various physical abnormalities. As part of a genotype/phenotype/cancer susceptibility study, we determined the prevalence of ophthalmic manifestations in these 4 syndromes.DesignCross-sectional study of a patient cohort.ParticipantsSeventy-five patients with an IBMFS and 121 of their first-degree relatives were seen in the National Eye Institute, National Institutes of Health, from 2001 to 2007. The patient group included 22 with FA, 28 with DC, 19 with DBA, and 6 with SDS.MethodsEvery participant underwent a complete ophthalmic evaluation and digital facial photography with an adhesive paper ruler on the patient's forehead for an internal measure of scale. Interpupillary distance (IPD), inner canthal distance (ICD), outer canthal distance (OCD), palpebral fissure length (PFL), and corneal diameter (CD) were measured. Thirteen of the 22 patients with FA underwent axial length (AL) measurements by A-scan ultrasonography.Main outcome measuresType and prevalence of ophthalmic manifestations.ResultsNinety-five percent of patients with FA had at least 1 abnormal parameter, and 25% of patients had at least 4 abnormal parameters. Eighty-two percent of patients had small palpebral fissures, 69% of patients had simple microphthalmia, 64% of patients had small OCD, 55% of patients had microcornea, 28% of patients had ptosis, and 6% of patients had epicanthal folds. In patients with DC, abnormalities of the lacrimal drainage system (29%) were the most prevalent findings, followed by retinal abnormalities (pigmentary changes, retinal neovascularization, retinal detachment, exudative retinopathy) in 21%, cicatricial entropion with trichiasis and blepharitis in 7% each, and sparse eyelashes and congenital cataract in 3.5% each. No significant ophthalmic abnormalities were seen in patients with DBA or SDS.ConclusionsSyndrome-specific ocular findings are associated with FA and DC and may antedate diagnosis of the specific syndrome. Early recognition of these abnormalities is important for optimal management.
| S-EPMC2830377 | biostudies-literature