OBJECTIVE:To knock out PDK1 by using Vav-Cre and observe the effects of PDK1 knock out on the ratio, number and differentiation of neutrophil.METHODS:The PDK1 expression level of Vav-Cre;PDK1lox/lox mouse in the bone marrow cells was analyzed by RT-PCR. The effect of PDK1 on hematopoietic progenitor was observed by CFU-C assay, and the effect of PDK1 on the ratio and number of neutrophil was detected by flow cytometric analysis.RESULTS:The RNA expression level of bone marrow cells of Vav-Cre;PDK1lox/lox mouse was dramatically lower than that of the control mouse. The number of functional GMP was lower in Vav-Cre;PDK1lox/lox mouse in contrast to controls. The percentage and number of neutrophil were lower, but the percentage of premyelocyte/myelocytes was more than 2 times in Vav-Cre;PDK1lox/lox group compared with control groups.CONCLUSION:PDK1 affects the process of the differention of hematopoietic stem cells to GMP, the neutrophil differention and maturation.

3-Phosphoinositide-dependent protein kinase 1 (PDK1) is a pivotal regulator in the phosphoinositide 3-kinase (PI3K)-Akt signaling pathway that have been shown to play key roles in the functional development of B and T cells via activation of AGC protein kinases during hematopoiesis. However, the role of PDK1 in HSCs has not been fully defined. Here we specifically deleted the PDK1 gene in the hematopoietic system and found that PDK1-deficient HSCs exhibited impaired function and defective lineage commitment abilities. Lack of PDK1 caused HSCs to be less quiescent and to produce a higher number of phenotypic HSCs and fewer progenitors. PDK1-deficient HSCs were also unable to reconstitute the hematopoietic system. Notably, HSC function was more dependent on PDK1 than on mTORC2, which indicates that PDK1 plays a dominant role in the Akt-mediated regulation of HSC function. PDK1-deficient HSCs also exhibited reduced ROS levels, and treatment of PDK1-deficient HSCs with L-butathioninesulfoximine in vitro elevated the low ROS level and promoted colony formation. Therefore, PDK1 appears to contribute to HSC function partially via regulating ROS levels.

OBJECTIVE:To explore the role of PDK1 in T-ALL development through establishing the Notch1-induced T-ALL mouse model by using Mx1-cre; LoxP system to knock-out PDK1.METHODS:Cell cycle and apoptosis of leukemic cells were detected by flow cytometry, and relative expression of tumor-related genes and transcription factors of leukemic cells were determined by quantitative real-time PCR.RESULTS:Notch1-induced T-ALL mouse model with inducible knock-out of PDK1 was established successfully. Compared to T-ALL control mouse model, PDK1 knock-out mice showed a significant longer survival time (P<0.01). There was no difference of cell cycle between control and PDK1 knock-out mice, and the apoptosis rate of leukemic cells in PDK1 knock-out mice was higher than that of control mice (P<0.001). PDK1 knock-out resulted in decreased expression of tumor-related genes and transcription factors, such as c-Myc and NF-κB (P<0.01), and increased expression level of P53 (P<0.01).CONCLUSION:PDK1 knock-out can inhibit the development of T-ALL, and its mechanism may be the leukemia progression inhibited by regulating the apoptosis and expression of multiple related genes and transcription factors.

Hematopoietic stem cells (HSCs) exist as a rare population with two essential properties of self-renewal and differentiation. HSCs can give rise to all hematopoietic progenitor and mature cells. While critical for a full understanding of the hematopoietic process and HSC-related clinical applications, the mechanisms of self-renewal and differentiation of HSCs remain elusive. The PI3K-Akt signaling pathway plays essential roles in the regulation of hematopoiesis. 3-Phosphoinositide-dependent protein kinase 1 (PDK1) activates multiple AGC kinases including Akt and is a pivotal regulator in this pathway. PDK1 phosphorylates Akt at its T308 residue and regulates the functional development of B and T cells during hematopoiesis. However, the role of PDK1 in HSCs has not been fully defined.

Although great efforts have been made to improve available therapies, the mortality rate of acute myeloid leukemia (AML) remains high due to poor treatment response and frequent relapse after chemotherapy. Leukemia stem cells (LSCs) are thought to account for this poor prognosis and relapse. Phosphoinositide-dependent kinase 1 (PDK1) is a critical regulator of the PI3K/Akt pathway and has been shown to be frequently activated in leukemia. However, the role of PDK1 in the regulation of LSCs in AML is still not clear. Using a PDK1 conditional deletion MLL-AF9 murine AML model, we revealed that the deletion of PDK1 prolonged the survival of AML mice by inducing LSC apoptosis. This was accompanied by the increased expression of the pro-apoptotic genes Bax and p53 and the reduced expression of Stat5, which has been shown to be constitutively activated in leukemia. Thus, our findings suggest that PDK1 plays an essential role in maintaining LSCs. Further delineating the function of PDK1 in LSCs may provide a new strategy for the improved treatment of AML relapse.