Epidermal growth factor receptor-dependent DNA repair promotes murine and human hematopoietic regeneration.

Chemotherapy and irradiation cause DNA damage to hematopoietic stem cells (HSCs), leading to HSC depletion and dysfunction and the risk of malignant transformation over time. Extrinsic regulation of HSC DNA repair is not well understood, and therapies to augment HSC DNA repair following myelosuppression remain undeveloped. We report that epidermal growth factor receptor (EGFR) regulates DNA repair in HSCs following irradiation via activation of the DNA-dependent protein kinase-catalytic subunit (DNA-PKcs) and nonhomologous end joining (NHEJ). We show that hematopoietic regeneration in vivo following total body irradiation is dependent upon EGFR-mediated repair of DNA damage via activation of DNA-PKcs. Conditional deletion of EGFR in hematopoietic stem and progenitor cells (HSPCs) significantly decreased DNA-PKcs activity following irradiation, causing increased HSC DNA damage and depressed HSC recovery over time. Systemic administration of epidermal growth factor (EGF) promoted HSC DNA repair and rapid hematologic recovery in chemotherapy-treated mice and had no effect on acute myeloid leukemia growth in vivo. Further, EGF treatment drove the recovery of human HSCs capable of multilineage in vivo repopulation following radiation injury. Whole-genome sequencing analysis revealed no increase in coding region mutations in HSPCs from EGF-treated mice, but increased intergenic copy number variant mutations were detected. These studies demonstrate that EGF promotes HSC DNA repair and hematopoietic regeneration in vivo via augmentation of NHEJ. EGF has therapeutic potential to promote human hematopoietic regeneration, and further studies are warranted to assess long-term hematopoietic effects.

Establishing induced pluripotent stem cell lines from two dominant optic atrophy patients with distinct OPA1 mutations and clinical pathologies.

Dominant optic atrophy (DOA) is an inherited disease that leads to the loss of retinal ganglion cells (RGCs), the projection neurons that relay visual information from the retina to the brain through the optic nerve. The majority of DOA cases can be attributed to mutations in optic atrophy 1 (OPA1), a nuclear gene encoding a mitochondrial-targeted protein that plays important roles in maintaining mitochondrial structure, dynamics, and bioenergetics. Although OPA1 is ubiquitously expressed in all human tissues, RGCs appear to be the primary cell type affected by OPA1 mutations. DOA has not been extensively studied in human RGCs due to the general unavailability of retinal tissues. However, recent advances in stem cell biology have made it possible to produce human RGCs from pluripotent stem cells (PSCs). To aid in establishing DOA disease models based on human PSC-derived RGCs, we have generated iPSC lines from two DOA patients who carry distinct OPA1 mutations and present very different disease symptoms. Studies using these OPA1 mutant RGCs can be correlated with clinical features in the patients to provide insights into DOA disease mechanisms.

PTPσ inhibitors promote hematopoietic stem cell regeneration.

Receptor type protein tyrosine phosphatase-sigma (PTPσ) is primarily expressed by adult neurons and regulates neural regeneration. We recently discovered that PTPσ is also expressed by hematopoietic stem cells (HSCs). Here, we describe small molecule inhibitors of PTPσ that promote HSC regeneration in vivo. Systemic administration of the PTPσ inhibitor, DJ001, or its analog, to irradiated mice promotes HSC regeneration, accelerates hematologic recovery, and improves survival. Similarly, DJ001 administration accelerates hematologic recovery in mice treated with 5-fluorouracil chemotherapy. DJ001 displays high specificity for PTPσ and antagonizes PTPσ via unique non-competitive, allosteric binding. Mechanistically, DJ001 suppresses radiation-induced HSC apoptosis via activation of the RhoGTPase, RAC1, and induction of BCL-XL. Furthermore, treatment of irradiated human HSCs with DJ001 promotes the regeneration of human HSCs capable of multilineage in vivo repopulation. These studies demonstrate the therapeutic potential of selective, small-molecule PTPσ inhibitors for human hematopoietic regeneration.

Dickkopf-1 promotes hematopoietic regeneration via direct and niche-mediated mechanisms.

The role of osteolineage cells in regulating hematopoietic stem cell (HSC) regeneration following myelosuppression is not well understood. Here we show that deletion of the pro-apoptotic genes Bak and Bax in osterix (Osx, also known as Sp7 transcription factor 7)-expressing cells in mice promotes HSC regeneration and hematopoietic radioprotection following total body irradiation. These mice showed increased bone marrow (BM) levels of the protein dickkopf-1 (Dkk1), which was produced in Osx-expressing BM cells. Treatment of irradiated HSCs with Dkk1 in vitro increased the recovery of both long-term repopulating HSCs and progenitor cells, and systemic administration of Dkk1 to irradiated mice increased hematopoietic recovery and improved survival. Conversely, inducible deletion of one allele of Dkk1 in Osx-expressing cells in adult mice inhibited the recovery of BM stem and progenitor cells and of complete blood counts following irradiation. Dkk1 promoted hematopoietic regeneration via both direct effects on HSCs, in which treatment with Dkk1 decreased the levels of mitochondrial reactive oxygen species and suppressed senescence, and indirect effects on BM endothelial cells, in which treatment with Dkk1 induced epidermal growth factor (EGF) secretion. Accordingly, blockade of the EGF receptor partially abrogated Dkk1-mediated hematopoietic recovery. These data identify Dkk1 as a regulator of hematopoietic regeneration and demonstrate paracrine cross-talk between BM osteolineage cells and endothelial cells in regulating hematopoietic reconstitution following injury.

Deletion of the Imprinted Gene Grb10 Promotes Hematopoietic Stem Cell Self-Renewal and Regeneration.

Imprinted genes are differentially expressed by adult stem cells, but their functions in regulating adult stem cell fate are incompletely understood. Here we show that growth factor receptor-bound protein 10 (Grb10), an imprinted gene, regulates hematopoietic stem cell (HSC) self-renewal and regeneration. Deletion of the maternal allele of Grb10 in mice (Grb10m/+ mice) substantially increased HSC long-term repopulating capacity, as compared to that of Grb10+/+ mice. After total body irradiation (TBI), Grb10m/+ mice demonstrated accelerated HSC regeneration and hematopoietic reconstitution, as compared to Grb10+/+ mice. Grb10-deficient HSCs displayed increased proliferation after competitive transplantation or TBI, commensurate with upregulation of CDK4 and Cyclin E. Furthermore, the enhanced HSC regeneration observed in Grb10-deficient mice was dependent on activation of the Akt/mTORC1 pathway. This study reveals a function for the imprinted gene Grb10 in regulating HSC self-renewal and regeneration and suggests that the inhibition of Grb10 can promote hematopoietic regeneration in vivo.

Uma leitura psicanalítica sobre o corpo e as ressonâncias da violência sexual na infância complementar / A psychoanalytic reading of the body and the resonances of childhood sexual violence / Une lecture psychanalytique du corps et les résonances de la violence sexuelle dans l'enfance

Este estudo baseou-se na teoria psicanalítica e, a partir da construção de um caso clínico, teve o objetivo de analisar o corpo enquanto ló cus de expressão do sofrimento psíquico após a vivência de uma experiência traumática como a violência sexual incestuosa na infância. A paciente Lis, aos dezoito anos, foi encaminhada para acompanhamento psicológico após realizar a denúncia de violência física, psicológica e sexual contra o pai biológico. Os atendimentos perduraram por dois anos e a dimensão corporal emergiu como aspecto privilegiado de análise. As manifestações corporais da paciente desvelavam as experiências emocionais excluídas do seu discurso e se destacavam em suas formações sintomáticas. Ante a dificuldade significativa de Lis em discriminar, organizar e acolher seus afetos e suas experiências, o corpo se constituiu como principal via de comunicação entre analista e analisanda. A análise do caso sinalizou que o corpo ancorava marcas do excesso pulsional traumático e aparecia como palco de experiências primitivas. Dessa forma, a pesquisa apontou para a indissociabilidade entre o campo analítico e a problemática do corpo, testemunha dos não ditos e do que escapa ao discurso simbólico do sujeito.

This study was based on the psychoanalytical theory and, through the construction of a clinical case, aimed to analyze the body as a locus of expression of the psychic suffering after undergoing a traumatic experience such as incestuous sexual violence in childhood. The patient Lis, eighteen years old, was referred to psychological counseling after reporting physical, psychological and sexual violence against her biological father. The consultations lasted for two years and the body dimension emerged as a privileged aspect of the analysis. T he patient's body manifestations revealed emotional experiences excluded from her discourse, which stood out in her symptomatic formations. Faced with Lis's significant difficulty in discriminating, organizing and welcoming her affections and experiences, the body became the main channel of communication between analyst and analysand. The analysis of the case signaled that the body anchored marks of the traumatic drive excess and appeared as the stage for primitive experiences. Thusly, the research pointed to the inseparability between the analytical field and the problematic of the body, a witness of the unsaid and of what escapes the subject's symbolic discourse.

Cette étude était basée sur la théorie psychanalytique, et de la construction d'un cas clinique, avait comme objectif d'analyser le corps comme locus d'expression de la souffrance psychique après avoir vécu une expérience traumatique que telle que la violence sexuelle incestueuse dans l'enfance. La patiente Lis, à dix-huit ans, a été orientée vers un conseil psychologique après avoir signalé des violence physique, psychologiques et sexuelles contre le père biologique. Les consultations ont duré deux ans et la dimension corporelle est apparue comme un aspect privilégié de l'analyse. Les manifestations du corps du patient ont révélé les expériences émotionnelles de son discours et se sont distinguées dans leurs formations symptomatiques. Face à l'importante difficulté de Lis à discriminer, organiser et accueillir ses affections et ses expériences, le corps était le principal canal de communication entre l'analyste et l'analysant. L'analyse de cas a signalé que le corps a ancré les marques de l'excès des pulsions traumatique et est apparu comme une scène des expériences primitives. Ainsi, la recherche a mis en évidence l'inséparabilité entre le camp analytique et la problématique du corps, témoin du non-dit et qui échappe au discours symbolique du sujet.