RESUMO
Diamond-Blackfan anemia (DBA) is a congenital disorder characterized by the failure of erythroid progenitor differentiation, severely curtailing red blood cell production. Because many DBA patients fail to respond to corticosteroid therapy, there is considerable need for therapeutics for this disorder. Identifying therapeutics for DBA requires circumventing the paucity of primary patient blood stem and progenitor cells. To this end, we adopted a reprogramming strategy to generate expandable hematopoietic progenitor cells from induced pluripotent stem cells (iPSCs) from DBA patients. Reprogrammed DBA progenitors recapitulate defects in erythroid differentiation, which were rescued by gene complementation. Unbiased chemical screens identified SMER28, a small-molecule inducer of autophagy, which enhanced erythropoiesis in a range of in vitro and in vivo models of DBA. SMER28 acted through autophagy factor ATG5 to stimulate erythropoiesis and up-regulate expression of globin genes. These findings present an unbiased drug screen for hematological disease using iPSCs and identify autophagy as a therapeutic pathway in DBA.
Assuntos
Anemia de Diamond-Blackfan/tratamento farmacológico , Descoberta de Drogas , Células-Tronco Hematopoéticas/metabolismo , Compostos Alílicos/farmacologia , Anemia de Diamond-Blackfan/patologia , Antígenos CD34/metabolismo , Autofagia/efeitos dos fármacos , Proteína 5 Relacionada à Autofagia/metabolismo , Diferenciação Celular/efeitos dos fármacos , Reprogramação Celular , Células Eritroides/efeitos dos fármacos , Células Eritroides/patologia , Eritropoese/efeitos dos fármacos , Teste de Complementação Genética , Globinas/metabolismo , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/efeitos dos fármacos , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/metabolismo , Quinazolinas/farmacologiaRESUMO
Adenylate kinases (AKs) are phosphotransferases that regulate the cellular adenine nucleotide composition and play a critical role in the energy homeostasis of all tissues. The AK2 isoenzyme is expressed in the mitochondrial intermembrane space and is mutated in reticular dysgenesis (RD), a rare form of severe combined immunodeficiency (SCID) in humans. RD is characterized by a maturation arrest in the myeloid and lymphoid lineages, leading to early onset, recurrent, and overwhelming infections. To gain insight into the pathophysiology of RD, we studied the effects of AK2 deficiency using the zebrafish model and induced pluripotent stem cells (iPSCs) derived from fibroblasts of an RD patient. In zebrafish, Ak2 deficiency affected hematopoietic stem and progenitor cell (HSPC) development with increased oxidative stress and apoptosis. AK2-deficient iPSCs recapitulated the characteristic myeloid maturation arrest at the promyelocyte stage and demonstrated an increased AMP/ADP ratio, indicative of an energy-depleted adenine nucleotide profile. Antioxidant treatment rescued the hematopoietic phenotypes in vivo in ak2 mutant zebrafish and restored differentiation of AK2-deficient iPSCs into mature granulocytes. Our results link hematopoietic cell fate in AK2 deficiency to cellular energy depletion and increased oxidative stress. This points to the potential use of antioxidants as a supportive therapeutic modality for patients with RD.
Assuntos
Adenilato Quinase/metabolismo , Células-Tronco Hematopoéticas/fisiologia , Leucopenia/enzimologia , Leucopenia/fisiopatologia , Estresse Oxidativo/fisiologia , Células-Tronco Pluripotentes/fisiologia , Imunodeficiência Combinada Severa/enzimologia , Imunodeficiência Combinada Severa/fisiopatologia , Laranja de Acridina , Adenilato Quinase/deficiência , Animais , Antioxidantes/farmacologia , Apoptose/fisiologia , Compostos Azo , Sequência de Bases , Diferenciação Celular/efeitos dos fármacos , Biologia Computacional , Primers do DNA/genética , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Dados de Sequência Molecular , Naftalenos , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Peixe-ZebraRESUMO
The advent of reprogramming technology has greatly advanced the field of stem cell biology and nurtured our hope to create patient specific renewable stem cell sources. While the number of reports of disease specific induced pluripotent stem cells is continuously rising, the field becomes increasingly more aware that induced pluripotent stem cells are not as similar to embryonic stem cells as initially assumed. Our state of the art understanding of human induced pluripotent stem cells, their capacity, their limitations and their promise as it pertains to the study and treatment of primary immunodeficiencies, is the content of this review.