Notch-Induced hIL-6 Production Facilitates the Maintenance of Self-Renewal of hCD34+ Cord Blood Cells Through the Activation of Jak-PI3K-STAT3 Pathway
Department of Molecular Medicine, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, South Korea. American Journal Of Pathology
(Impact Factor: 4.59).
11/2011; 180(1):351-64. DOI: 10.1016/j.ajpath.2011.09.030
Ex vivo expansion of CD34(+) stem cells in contact culture between hCD34(+)CD38(-)Lin(-) cord blood stem cells and human delta-like-expressing AFT024 feeder cells revealed increased amounts of stemness-related proteins such as HoxB4, GATA2, Bmi-1, and p21 and anti-apoptotic proteins such as Bcl-2, Bcl-xL, Mcl-1, and phospho-Bad, when compared with control or noncontact culture. Production of human IL-6 (hIL-6) was markedly elevated in the culture, but was profoundly inhibited by treatment with γ-secretase inhibitor. In addition, Notch-induced activation of STAT3 was directly involved in gene expression of hIL-6 and soluble hIL-6Rα, indicating the close linkage between Notch signaling and hIL-6 production. Furthermore, depletion of soluble hIL-6 (with hIL-6-specific antibodies) and inhibition of IL-6-mediated signals (with a Jak1 inhibitor and wortmannin) severely affected the maintenance of self-renewal of hCD34(+) cord blood cells. It was also observed that the ex vivo expanded CD34(+) cord blood cells were induced to reconstitute human immune cells in nonobese diabetic mice with severe combined immunodeficiency when compared with freshly isolated CD34(+) cord blood cells. Together, these results strongly demonstrate that Notch signaling in the "cell-to-cell contact" between hCD34(+) cord blood and delta-like-expressing AFT024 feeder cells facilitates maintenance of self-renewal of hCD34(+) cord blood cells through direct regulation of hIL-6 production.
Available from: Marc F Diederich
- "In this specific case, inhibition of P-AKT-S473 is likely a secondary effect resulting from primary inactivation of upstream kinases including the serine/threonine PI3K since phosphorylation/activation of p85 regulatory subunit was inhibited by 13c. Besides, it was also reported that Notch signaling induces activation of STAT3 in the self-renewal phase of CD34 + hematopoietic cells through the JAK1/PI3K/AKT pathway , suggesting a role for PI3K/AKT in STAT3 activation. Similarly P-STAT3-Y705 inhibition should result from upstream inhibition of kinases JAK2 and Src, since P-JAK2 and P-STAT3 were inhibited at the same concentration (50 lM 13c) as soon as 6 h in HEL cells. "
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ABSTRACT: Constitutive activity of kinases has been reported in many types of cancers, so that inhibition of "onco-kinases" became a validated anti-cancer strategy. We found that the polyphenol 13c, a tri-vanillate derivative, inhibited kinase phosphorylation in leukemia cells. P-JAK2, P-Src and P-PI3Kp85 inhibition occurred independently of phosphatase involvement in JAK2V617F expressing HEL cells while 13c inhibited Bcr-Abl expression without inhibition of phosphorylation in chronic myelogenous cell lines (K562, MEG-01). In correlation with kinase inhibition, 13c abolished constitutive P-STAT3/P-STAT5 expression, down-regulated Mcl-1 and c-Myc gene expression and induced apoptosis. Altogether, polyphenol 13c displays potential antitumor activities by affecting onco-kinases and STAT activities.
Cancer letters 06/2013; 340(1). DOI:10.1016/j.canlet.2013.06.023 · 5.62 Impact Factor
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ABSTRACT: Clinical trials over the last 15 years have demonstrated that cell and gene therapies for cancer, monogenic and infectious disease are feasible and can lead to long-term benefit for patients. However, these trials have been limited to proof-of-principle and were conducted on modest numbers of patients or over long periods of time. In order for these studies to move towards standard practice and commercialization, scalable technologies for the isolation, ex vivo manipulation and delivery of these cells to patients must be developed. Additionally, regulatory strategies and clinical protocols for the collection, creation and delivery of cell products must be generated. In this article we review recent progress in hematopoietic cell and gene therapy, describe some of the current issues facing the field and discuss clinical, technical and regulatory approaches used to navigate the road to product development.
Cytotherapy 08/2012; 14(7):775-90. DOI:10.3109/14653249.2012.694420 · 3.29 Impact Factor
Available from: onlinelibrary.wiley.com
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ABSTRACT: Notch signalling has been implicated in haematopoietic stem cell self-renewal. Although several studies have tested the effect of activating or inhibiting the Notch signalling pathway in stem cells, no study has yet determined the functional differences associated with expressing Notch1. The aims of this study were to characterise the expression of human cell surface Notch1 in cord blood (CB) CD34+ cells and to study the function of Notch in CD34+ cells in vitro.
A monoclonal antibody against the extracellular domain of Notch1 was developed, and Notch1 expression in CB CD34+ cells was assessed by flow cytometry. CB CD34+ cells were sorted on the basis of their Notch1 expression and cultured in serum-free media. Single sorted CD34+CD38- Notch1+/- cells were cultured for 8 weeks on murine stroma monolayers and assayed for stem cell activity and lineage potential using a cobblestone area-forming cell (CAFC) assay.
Cell surface Notch1 expression was characterised in various primitive CD34+ cell compartments including a small subpopulation of CD34+CD38-cells. We found the CD34+CD38-Notch1+ population to be enriched for stem cell activity. Moreover, CD34+CD38-Notch1+ but not Notch1- cells demonstrated multi-lineage potential.
These data show that Notch1 is expressed on a functionally distinct sub-population of CD34+ cells that is highly enriched for stem cell activity and multi-lineage potential and could suggest that Notch1 could be used as a novel stem cell marker. This article is protected by copyright. All rights reserved.
European Journal Of Haematology 09/2013; 92(1). DOI:10.1111/ejh.12200 · 2.07 Impact Factor
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