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Attenuation of PKC? enhances metabolic activity and promotes expansion of blood progenitors.

ABSTRACT: A finely tuned balance of self-renewal, differentiation, proliferation, and survival governs the pool size and regenerative capacity of blood-forming hematopoietic stem and progenitor cells (HSPCs). Here, we report that protein kinase C delta (PKC?) is a critical regulator of adult HSPC number and function that couples the proliferative and metabolic activities of HSPCs. PKC?-deficient mice showed a pronounced increase in HSPC numbers, increased competence in reconstituting lethally irradiated recipients, enhanced long-term competitive advantage in serial transplantation studies, and an augmented HSPC recovery during stress. PKC?-deficient HSPCs also showed accelerated proliferation and reduced apoptosis, but did not exhaust in serial transplant assays or induce leukemia. Using inducible knockout and transplantation models, we further found that PKC? acts in a hematopoietic cell-intrinsic manner to restrict HSPC number and bone marrow regenerative function. Mechanistically, PKC? regulates HSPC energy metabolism and coordinately governs multiple regulators within signaling pathways implicated in HSPC homeostasis. Together, these data identify PKC? as a critical regulator of HSPC signaling and metabolism that acts to limit HSPC expansion in response to physiological and regenerative demands.

SUBMITTER: Rao TN

PROVIDER: S-EPMC6293338 | biostudies-literature | 2018 Dec

REPOSITORIES: biostudies-literature

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Attenuation of PKCδ enhances metabolic activity and promotes expansion of blood progenitors.

Rao Tata Nageswara TN Gupta Manoj K MK Softic Samir S Wang Leo D LD Jang Young C YC Thomou Thomas T Bezy Olivier O Kulkarni Rohit N RN Kahn C Ronald CR Wagers Amy J AJ

The EMBO journal 20181116 24

A finely tuned balance of self-renewal, differentiation, proliferation, and survival governs the pool size and regenerative capacity of blood-forming hematopoietic stem and progenitor cells (HSPCs). Here, we report that protein kinase C delta (PKCδ) is a critical regulator of adult HSPC number and function that couples the proliferative and metabolic activities of HSPCs. PKCδ-deficient mice showed a pronounced increase in HSPC numbers, increased competence in reconstituting lethally irradiated r ...[more]

PMID: 30446598

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Project description:The unlimited expansion of human progenitor cells in vitro could unlock many prospects for regenerative medicine. However, it remains an important challenge as it requires the decoupling of the mechanisms supporting progenitor self-renewal and expansion from those mechanisms promoting their differentiation. This study focuses on the expansion of human pluripotent stem (hPS) cell-derived pancreatic progenitors (PP) to advance novel therapies for diabetes. We obtained mechanistic insights into PP expansion requirements and identified conditions for the robust and unlimited expansion of hPS cell-derived PP cells under GMP-compliant conditions through a hypothesis-driven iterative approach. We show that the combined stimulation of specific mitogenic pathways, suppression of retinoic acid signaling, and inhibition of selected branches of the TGFβ and Wnt signaling pathways are necessary for the effective decoupling of PP proliferation from differentiation. This enabled the reproducible, 2000-fold, over 10 passages and 40-45 d, expansion of PDX1+/SOX9+/NKX6-1+ PP cells. Transcriptome analyses confirmed the stabilization of PP identity and the effective suppression of differentiation. Using these conditions, PDX1+/SOX9+/NKX6-1+ PP cells, derived from different, both XY and XX, hPS cell lines, were enriched to nearly 90% homogeneity and expanded with very similar kinetics and efficiency. Furthermore, non-expanded and expanded PP cells, from different hPS cell lines, were differentiated in microwells into homogeneous islet-like clusters (SC-islets) with very similar efficiency. These clusters contained abundant β-cells of comparable functionality as assessed by glucose-stimulated insulin secretion assays. These findings established the signaling requirements to decouple PP proliferation from differentiation and allowed the consistent expansion of hPS cell-derived PP cells. They will enable the establishment of large banks of GMP-produced PP cells derived from diverse hPS cell lines. This approach will streamline SC-islet production for further development of the differentiation process, diabetes research, personalized medicine, and cell therapies.

Project description:The immense regenerative power of hematopoietic tissue stems from the activation of the immature stem cells and the progenitor cells. After partial damage, hematopoiesis is reconstituted through a period of intense regeneration when blood cell production originates from erythro-myeloid progenitors in the virtual absence of stem cells. Since the damaged hematopoiesis can also be reconstituted from transplanted hematopoietic cells, we asked whether this also leads to the transient state when activated progenitors initially execute blood cell production. We first showed that the early reconstitution of hematopoiesis from transplanted cells gives rise to extended populations of developmentally advanced but altered progenitor cells, similar to those previously identified in the bone marrow regenerating from endogenous cells. We then identified the cells that give rise to these progenitors after transplantation as LSK CD48- cells. In the submyeloablative irradiated host mice, the transplanted LSK CD48- cells preferably colonized the spleen. Unlike the endogenous hematopoiesis reconstituting cells, the transplanted whole bone marrow cells and sorted LSK CD48- cells had greater potential to differentiate to B-lymphopoiesis. Separate transplantation of the CD150- and CD150+ subsets of LSK CD48- cells suggested that CD150- cells had a greater preference to B-lymphopoiesis than CD150+ cells. In the intensively regenerating hematopoiesis, the CD71/Sca-1 plot of immature murine hematopoietic cells revealed that the expanded populations of altered myeloid progenitors were highly variable in the different places of hematopoietic tissues. This high variability is likely caused by the heterogeneity of the hematopoiesis supporting stroma. Lastly, we demonstrate that during the period when active hematopoiesis resumes from transplanted cells, the hematopoietic tissues still remain highly permissive for further engraftment of transplanted cells, particularly the stem cells. Thus, these results provide a rationale for the transplantation of the hematopoietic stem cells in successive doses that could be used to boost the transplantation outcome.

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Attenuation of PKC? enhances metabolic activity and promotes expansion of blood progenitors.

Publications

Attenuation of PKCδ enhances metabolic activity and promotes expansion of blood progenitors.

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