scholarly journals A zebrafish model of dyskeratosis congenita reveals hematopoietic stem cell formation failure resulting from ribosomal protein-mediated p53 stabilization

Blood ◽  
2011 ◽  
Vol 118 (20) ◽  
pp. 5458-5465 ◽  
Author(s):  
Tamara C. Pereboom ◽  
Linda J. van Weele ◽  
Albert Bondt ◽  
Alyson W. MacInnes
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Ribosomal Proteins ◽  
Bone Marrow Failure ◽  
Ribosomal Subunit ◽  
Dyskeratosis Congenita ◽  
Telomere Maintenance ◽  
Zebrafish Model ◽  
Hematopoietic Stem

Abstract Dyskeratosis congenita (DC) is a bone marrow failure disorder characterized by shortened telomeres, defective stem cell maintenance, and highly heterogeneous phenotypes affecting predominantly tissues that require high rates of turnover. Here we present a mutant zebrafish line with decreased expression of nop10, one of the known H/ACA RNP complex genes with mutations linked to DC. We demonstrate that this nop10 loss results in 18S rRNA processing defects and collapse of the small ribosomal subunit, coupled to stabilization of the p53 tumor suppressor protein through small ribosomal proteins binding to Mdm2. These mutants also display a hematopoietic stem cell deficiency that is reversible on loss of p53 function. However, we detect no changes in telomere length in nop10 mutants. Our data support a model of DC whereupon in early development mutations involved in the H/ACA complex contribute to bone marrow failure through p53 deregulation and loss of initial stem cell numbers while their role in telomere maintenance does not contribute to DC until later in life.

scholarly journals ALLOGENEIC HEMATOPOIETIC STEM CELL TRANSPLANTATION FOR ADULT PATIENTS WITH FANCONI ANEMIA.

2016 ◽  
Vol 8 ◽  
pp. 2016054 ◽  
Author(s):  
Hosein Kamranzadeh fumani ◽  
Mohammad Zokaasadi ◽  
Amir Kasaeian ◽  
Kamran Alimoghaddam ◽  
Asadollah Mousavi ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Hematopoietic Stem Cell Transplantation ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Hematopoietic Stem Cell ◽  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
Term Survival ◽  
Hematopoietic Stem

Background & objectives: Fanconi anemia (FA) is a rare genetic disorder caused by an impaired DNA repair mechanism which leads to an increased tendency toward malignancies and progressive bone marrow failure. The only curative management available for hematologic abnormalities in FA patients is hematopoietic stem cell transplantation (HSCT). This study aimed to evaluate the role of HSCT in FA patients.Methods: Twenty FA patients with ages of 16 or more who underwent HSCT between 2002 and 2015 enrolled in this study. All transplants were allogeneic and the stem cell source was peripheral blood and all patients had a full HLA-matched donor.Results: Eleven patients were female and 9 male (55% and 45%). Mean age was 24.05 years. Mortality rate was 50% (n=10) and the main cause of death was GVHD. Survival analysis showed an overall 5-year survival of 53.63% and 13 year survival of 45.96 % among patients.Conclusion: HSCT is the only curative management for bone marrow failure in FA patients and despite high rate of mortality and morbidity it seems to be an appropriate treatment with an acceptable long term survival rate for adolescent and adult group.

scholarly journals Detection of Mutations in Inherited Bone Marrow Failure and Myelodysplastic Syndrome Genes Using Genomic Capture and Massively Parallel Sequencing in Clinical Diagnostics

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1507-1507
Author(s):  
Siobán B. Keel ◽  
Tom Walsh ◽  
Colin Pritchard ◽  
Akiko Shimamura ◽  
Mary-Claire King ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Genetic Testing ◽  
Hematopoietic Stem Cell ◽  
Copy Number ◽  
Bone Marrow Failure ◽  
Copy Number Variants ◽  
Donor Selection ◽  
Hematopoietic Stem ◽  
Pathogenic Mutations

Abstract Accurate and timely diagnosis of inherited bone marrow failure (BMF) and myelodysplastic syndromes (MDS) ensures appropriate clinical management. The correct diagnosis allows appropriate monitoring for both hematopoietic (i.e. clonal evolution and progressive marrow failure) and extra-hematopoietic complications, informs the timing of hematopoietic stem cell transplant, donor selection and transplant regimen planning, and ensures appropriate genetic counseling of family members. Substantial phenotypic overlap among these disorders and the variable expressivity within syndromes complicate their diagnosis based purely on physical exam and standard laboratory testing and provide the rationale for comprehensive genetic diagnostic testing. We report here our initial one-year experience utilizing a targeted capture assay of known inherited BMF/MDS genes for clinical diagnostic purposes at the University of Washington. The assay sequences all exons and 20 base pairs of intronic sequence flanking each exon, as well as several regulatory and intronic regions of specific genes containing known pathogenic variants of 85 known inherited BMF/MDS genes (Zhang M. et al. Haematologica 2016). Between June 2015 and July 2016, 81 individual patients were referred for clinical testing (median age: 15 years-old, range: 0.6-76 years-old). For all samples evaluated, median coverage across the 383kb targeted region was 1887X. This depth of coverage enabled identification of all classes of mutations, including point mutations, small indels, copy number variants, and genomic rearrangements. Pathologic mutations in known inherited BMF/MDS genes were identified in 12 of 82 (14.6%) individuals (median age 13 years-old, range: 1.25-43 years-old) across a broad number of genes and of multiple classes including copy number variants (Table). Among the twelve patients with pathogenic mutations in inherited BMF/MDS genes, genetic testing was consistent with the prior clinical diagnoses of eight patients, including two Fanconi anemia patients subtyped as complementation group A, one of whom demonstrated reversion to wild-type resulting in mosaicism in the peripheral blood. Importantly, four patients carried no specific inherited BMF/MDS diagnosis prior to testing and were found to have pathogenic mutations in RPS10, RTEL1 and RUNX1 (ID 005, 008, 009, 010), suggesting additional diagnostic value to a multiplexed genetic approach in the clinical setting. Detailed clinical information was available for nine of the patients diagnosed with pathogenic mutations, two of whom have or will undergo a sibling or haploidentical hematopoietic stem cell transplantation (009 and 012, respectively) and thus genetic testing informed donor selection. To improve diagnostic accuracy, we are now updating the capture design to include newly discovered inherited BMF/MDS genes and intronic regions to optimize copy number variant detection. We are additionally pursuing CLIA-certified RNA analyses to characterize whether several variants bioinformatically predicted to affect splicing are functionally deleterious. Next-generation sequencing for mutations involved in hereditary marrow failure and MDS may also become increasingly important in the context of precision-medicine in which germline mutations are unexpectedly identified in somatic testing. Disclosures No relevant conflicts of interest to declare.

Late Donor Bone Marrow Failure After Allogeneic Hematopoietic Stem Cell Transplantation

2014 ◽  
Vol 97 (12) ◽  
pp. e75-e77 ◽  
Author(s):  
Mathieu Meunier ◽  
Anne-Claire Manez ◽  
Aliénor Xhaard ◽  
Régis Peffault de Latour ◽  
Flore Sicre de Fontbrune ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Hematopoietic Stem Cell Transplantation ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Hematopoietic Stem Cell ◽  
Bone Marrow Failure ◽  
Hematopoietic Stem ◽  
Donor Bone Marrow

Hematopoietic Stem Cell Differentiation Pathway in Humans Deduced From the Lineage Diversity of PNH-Type Cells in Patients with Bone Marrow Failure.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1489-1489
Author(s):  
Takamasa Katagiri ◽  
Zhirong Qi ◽  
Yu Kiyu ◽  
Naomi Sugimori ◽  
J. Luis Espinoza ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Peripheral Blood ◽  
Blood Cells ◽  
Bone Marrow Failure ◽  
Stem Cell Differentiation ◽  
Refractory Anemia ◽  
Hematopoietic Stem

Abstract Abstract 1489 Poster Board I-512 The hematopoietic stem cell (HSC) differentiation pathway in humans remains largely unknown due to the lack of an appropriate in vivo assay allowing the growth of HSCs as well as of clonal markers that enable the tracing of their progenies. Small populations of blood cells deficient in glycosylphosphatidylinositol-anchored proteins (GPI-APs) such as CD55 and CD59 are detectable in approximately 50% of patients with aplastic anemia (AA) and 15% of patients with refractory anemia (RA) of myelodysplastic syndrome defined by the FAB classification. Such blood cells with the paroxysmal nocturnal hemoglobinuria (PNH) phenotype (PNH-type cells) are derived from single PIGA mutant HSCs and their fate depends on the proliferation and self-maintenance properties of the individual HSCs that undergo PIG-A mutation by chance (Blood 2008;112:2160, Br J Haematol 2009 in press) Analyses of the PNH-type cells from a large number of patients on the diversity of lineage combination may help clarify the HSC differentiation pathway in humans because PIG-A mutant HSCs in patients with bone marrow failure appear to reflect the kinetics of healthy HSCs. Therefore, different lineages of peripheral blood cells were examined including glycophorin A+ erythrocytes (E), CD11b+ granulocytes (G), CD33+ monocytes (M), CD3+ T cells (T), CD19+ B cells (B), and NKp46+ NK cells (Nk) from 527 patients with AA or RA for the presence of CD55−CD59− cells in E and G, and CD55−CD59−CD48− cells in M,T, B, Nk with high sensitivity flow cytometry. Two hundred and twenty-eight patients (43%) displayed 0.003% to 99.1% PNH-type cells in at least one lineage of cells. The lineage combination patterns of PNH-type cells in these patients included EGM in 71 patients (31%), EGMTBNk in 43 (19%), EG in 37 (16%), T alone 14 (6%), EGMBNk in 11 (5%), G alone in 10 (4%), GM in 10 (4%), EGMNk in 7 (3%), EGMT in 7 (3%), EGMB in 6 (3%), EM in 5 (2%), EGMTB in 3 (1%), EGNk in 1 (0.4%), EGMTNk in 1 (0.4%), GMTB in 1 (0.4%), and GT in 1 (0.4%) (Table). All patterns included G or M, except for 14 patients displaying PNH-type T cells alone. No patients showed TB or TBNk patterns suggestive of the presence of common lymphoid progenitor cells. Peripheral blood specimens from 123 patients of the 228 patients possessing PNH-type cells were examined again after 3 to 10 months and all patients showed the same combination patterns as those revealed by the first examination. PIG-A gene analyses using sorted PNH-type cells from 3 patients revealed the same mutation in G and Nk for 1 patient and in G and T for 2 patients. These findings indicate that human HSCs may take a similar differentiation pathway to that of murine HSCs, the ‘myeloid-based model’ that was recently proposed by Kawamoto et al. (Nature 2008; 10:452), though the cases with PNH-type T cells alone remain to be elucidated. Table. Lineages of cells containing PNH-type cells in patients with AA or RA. The number in the parenthesis denotes the proportion of patients showing each combination pattern in the total patients possessing PNH-type cells. (+ ; presence of PNH-type cells) Disclosures No relevant conflicts of interest to declare.

DNA Damage Accumulation, Accelerated Hematopoietic Stem Cell Aging, and Partial Reversal by Antioxidant Treatment in a Mouse Model of Dyskeratosis Congenita.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 498-498
Author(s):  
Baiwei Gu ◽  
JianMeng Fan ◽  
Monica Bessler ◽  
Philip Mason
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Cell Function ◽  
Bone Marrow Failure ◽  
Dyskeratosis Congenita ◽  
Telomere Maintenance ◽  
Cell Aging ◽  
Female Mice ◽  
Stem Cell Aging ◽  
Population Doublings

Abstract Abstract 498 X-linked dyskeratosis congenita (DC) caused by mutations in DKC1, encoding the protein dyskerin, is the most common form of DC, a severe inherited bone marrow failure (BMF) syndrome associated with a predisposition to malignancy. Dyskerin is a component of small nucleolar ribonucleoprotein particles(snoRNPs) that modify specific residues in nascent ribosomal RNA(rRNA) molecules and also forms part of the telomerase complex responsible for synthesizing the telomere repeats at the ends of chromosomes. Strong evidence suggests that compromised telomerase function is the major cause of DC but defects in ribosome biogenesis may contribute to the disease. Excessive telomere shortening resulting in premature cellular senescence is thought to be the primary cause of bone marrow failure in dyskeratosis congenita. Our previous data showed that, in mice, cells expressing a Dkc1 mutation (Dkc1Δ15) had a telomerase dependent but telomere length independent growth defect. Here we show that the growth rate of Δ15 MEF cells was lower when cultured at both ambient oxygen (21%) and low (3%) oxygen. In 21% oxygen both Δ15 and WT cells stopped growing and entered senescence after 8-10 population doublings, with the Δ15 cells growing more slowly than the WT cells. In 3% oxygen Δ15 cells grew more slowly and entered senescence earlier than WT cells. Further investigations reveal that both γ-H2AX foci number and reactive oxygen species (ROS) levels in Δ15 cells were significantly higher than in WT cells with increased passage number even when cultured in low oxygen. Increased levels of γ-H2AX and p53 in Dkc1Δ15 mice, particularly in older mice, were also detected in liver, spleen and bone marrow. To study the effect of the mutation on stem cell function during aging, we carried out competitive repopulation experiments using the CD45.1/CD45.2 congenic system. Irradiated mice were injected with a 1:1 mixture of Dkc1Δ15 and Dkc1+ bone marrow from old (77-88w) or young (10w) animals. Old Dkc1Δ15cells are less able to compete with age matched WT cells in primary recipients, making up only 20% of cells after 12 weeks compared with 40% for the young cells. Moreover, serial transplantation results show that, in secondary recipients, BM cells from old Dkc1Δ15 mice were not detectable while Dkc1Δ15 cells from young mice still comprise 10-30% of the bone marrow after 12 weeks. These results strongly indicate the Dkc1Δ15 mutation causes decay of stem cell function with age. Because of the association with ROS we asked whether treatment with an antioxidant could rescue the growth disadvantage of Δ15 cells. We grew primary MEF cells from Dkc1Δ15/+ female mice in the presence or absence of 100 M N-acetyl cysteine (NAC), a clinically approved antioxidant. These cultures consist in early passages of 50% cells expressing WT and 50% expressing Δ15 dyskerin, reflecting random X-chromosome inactivation, Without NAC the WT cells almost completely outgrew the Δ15 cells after 11 population doublings but in the presence of NAC the Δ15 cells are still clearly present after 15 population doublings, suggesting that NAC at least partially rescues the growth disadvantage of dyskerin mutant cells. More impressively, the growth disadvantage of the Δ15 cells is also rescued in vivo in Dkc1Δ15/+ female mice given the NAC (1mg/ml) in their drinking water. Although the precise mechanism will be the subject of further investigation, these results point to a functional link between increased oxidative stress, defective telomere maintenance and stem cell aging in the pathogenesis of BMF in dyskeratosis congenita. Disclosures: Bessler: Alexion: Membership on an entity's Board of Directors or advisory committees.

Antioxidant Treatment Rescues An Accelerated Aging Phenotype In Dyskerin Mutated Bone Marrow Stem Cells.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2614-2614
Author(s):  
Baiwei Gu ◽  
Jian-meng Fan ◽  
Monica Bessler ◽  
Philip J Mason
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Cell Function ◽  
Bone Marrow Failure ◽  
Dyskeratosis Congenita ◽  
Untreated Group ◽  
Telomere Maintenance ◽  
Antioxidant Treatment ◽  
Transplantation Experiments

Abstract Abstract 2614 X-linked Dyskeratosis Congenita (DC) is due to mutations in the DKC1 gene, which encodes the protein dyskerin. Dyskerin is a highly conserved nucleolar protein that, as part of a specialized nucleolar RNP, catalyzes the pseudouridylation of specific residues in newly synthesized ribosomal RNAs and spliceosomal snRNAs. Dyskerin also associates with telomerase and is involved in telomere maintenance. In addition to the well known effect of telomere homeostasis on cancer, it is evident that telomere maintenance may also be important in replicative aging because of telomere shortening due to the limited expression of telomerase activity in dividing somatic cells. Accumulating evidence suggests that dysfunctional telomeres resulting in premature cellular senescence is the primary cause of bone marrow failure in dyskeratosis congenita. It is important to determine the mechanism whereby Dkc1 mutations lead to premature cellular senescence in bone marrow. We have produced a line of mice containing a mutation, Dkc1Δ15, which is a copy of a pathogenic human mutation. Male Dkc1Δ15 mice showed a decrease in the proportion of B and T lymphocytes in peripheral blood and reduced body weight with age but no overt bone marrow failure syndrome phenotypes. Our previous competitive bone marrow transplantation experiments showed that the Dkc1Δ15 mutation caused decay of stem cell function with age. Bone marrow from older Dkc1Δ15 mice was markedly inefficient in repopulation studies compared with bone marrow from age matched wild type mice. We also found that N-acetyl cysteine (NAC) could at least partially rescue the growth disadvantage of dyskerin mutant spleen cells or fibroblasts which was associated with accumulation of DNA damage and reactive oxygen species. To determine if NAC, or other antioxidants might be useful therapeutically it is important to determine their effects on stem cell function, which is defective in DC. To this end we established a cohort of mice that were given NAC in their drinking water (1mg/ml) from 3-weeks of age and maintained on NAC for 1 year. We found that long term NAC treatment did not show significant side effects on the mice. They had slightly increased neutrophils, but no difference in life span and body weight compared with the untreated group. Impressively, old male Dkc1Δ15 mice showed corrected B and T cell proportions in peripheral blood after treatment with NAC. Competitive bone marrow transplantation experiments were carried out in which a 1:1 mixture of BM cells from mutant and WT mice was used to repopulated lethally irradiated recipient mice. These experiments showed that, when taken from NAC treated animals, old Dkc1Δ15 BM cells could compete with age matched WT cells with 40–45% of Dkc1Δ15 cells in primary recipients compared with only 20% for the untreated group. Moreover, after secondary transplantation, cells from the NAC treated group still represent 15–20% of Dkc1Δ15 cells in recipients while those from the untreated group could not be detected. These results strongly suggest that NAC treatment can partially restore the bone marrow repopulating ability of Dkc1Δ15 stem cells. Together with our previous results these data suggest that a pathogenic Dkc1 mutation, through its effect on telomerase, initiates stem cell aging before telomeres are short and that increased oxidative stress might play a role in this process. Moreover the effects of the mutation may be prevented or delayed by antioxidant treatment, although the precise mechanism will be the subject of future investigation. Disclosures: Bessler: Alexion Pharmaceutical Inc: Consultancy; Novartis: Membership on an entity's Board of Directors or advisory committees; Taligen: Consultancy.

Outcome of Pediatric Allogeniec Hematopoietic Stem Cell Transplantation for Nonmalignant Disease: A Single Center Experience

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 4558-4558
Author(s):  
Ibraheem Abosoudah ◽  
Asem Lashin ◽  
Fawwaz Yassin ◽  
Hassan Al trabolsi ◽  
Mohamed Bayoumy
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Hematopoietic Stem Cell Transplantation ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Hematopoietic Stem Cell ◽  
Graft Failure ◽  
Bone Marrow Failure ◽  
Hematopoietic Stem ◽  
Nonmalignant Disease

Abstract Abstract 4558 Background: The aim of this study was to determine the outcome of pediatric allogeneic hematopoietic stem cell transplantation (Allo-HSCT) for nonmalignant disease in our center. Method: Data were retrospectively collected for all patients (aged 0–18 years) who received allogeneic HSCT between May, 2005 and December, 2011. Outcomes according to the type of transplant, diagnosis, and transplant-related complications are reported. Result: Allo-HSCT was performed in 17 patients, (11 male; 6 female). Bone marrow Failure (7;41%) was the commonest nonmalignant disease followed by B-thalassemia major (4;24%) and others (6;35%). Only HLA-matched family donors were used. Most patients were conditioned with Busulfan/Cyclophosphamide/ATG. GVHD prophylaxis comprised mainly of cyclosporine and methotrexate. The median time to neutrophil engraftment was 20 (9–27) days. Two patients experienced graft failure. Four patients had Grade (I–III) aGVHD and no patient had grade (IV). Four patients (23%) had chronic GVHD. No patient had sinusoidal obstruction syndrome (SOS). Five patients had reactivation of CMV infection, which was treated and resolved in all of them. Only one patient died of sepsis. Overall survival (OS) and event free survival (EFS) were 94%, and 88% respectively. Conclusion: In this cohort of patients, bone marrow failure was the main reason for transplantation. The early results of HSCT were promising and consistent with published international data. Similarly, graft failure remains a concern. Disclosures: No relevant conflicts of interest to declare.

Sign in / Sign up

Export Citation Format

Share Document