It has been well established that the so-called Yamanaka factors, Oct4 (octamer-binding transcription factor 4), Sox2 (SRY (sex determining region Y)-box 2), Klf4 (Kruppel-like factor 4), and c-Myc, together stimulate the generation of induced pluripotent stem cells (iPSC) 3 from diverse types of somatic cells including embryonic fibroblast cells (1-5). Oct4 belongs to the family of POU transcriptional factors and binds to an octamer sequence motif, AGTCAAAT, leading to expression of target genes in developing germ cells (6, 7). Sox2 is a member of Sox transcriptional factor family containing the high mobility group domain and is known to be required for maintenance of diverse types of stem cells throughout life (8). KLF4 is a member of Kruppel-like factor transcriptional factor family that binds to CACCC sequence regulating gene expression during embryonic development (9, 10). c-Myc is a well known basic helixloop-helix transcriptional factor that plays important roles in cell cycle regulation, cellular transformation, and maintenance of pluripotency (11,12).Ahnak is an exceptionally large protein (ϳ700 kDa) and has an extended central region composed of 36 repeat units that serve as a scaffolding motif for interaction of phospholipase C-␥ with PKC in regulation of cell proliferation and migration (13-16). Moreover, the central repeated unit is known to bind to R-Smad protein in response to TGF- and mediate down-regulation of c-Myc expression and cell growth retardation. Specifically, nuclear localization of Ahnak in complex with phospho-Smad3 and binding to promoter of c-Myc were strongly enhanced in response to TGF-, indicating that Ahnak downregulates the expression of c-Myc as Smad3 target genes. Consistently, Ahnak-deficient MEF cells show a significant up-regulation of c-Myc expression (17).There have been efforts to exclude c-Myc from the mix of ectopically expressed transcription factors for iPSC generation mainly because of its potential transforming activity. Exclusion of c-Myc, however, led to a dramatic reduction in efficiency (18 -20). Therefore, developing a method to efficiently utilize endogenous c-Myc activity for iPSC generation could represent an important progress. Here, we present that Ahnak knock-out MEF or wild type MEF expressing shRNA specific to Ahnak can be converted to iPSC at a high efficiency without infection of c-Myc retrovirus. Further dissection of Ahnak pathway may provide novel strategies for regulation of iPSC generation and reducing the potential danger from cellular transformation.
We report that phytosphingosine, a sphingolipid found in many organisms and implicated in cellular signaling, promotes megakaryocytic differentiation of myeloid leukemia cells. Specifically, phytosphingosine induced several hallmark changes associated with megakaryopoiesis from K562 and HEL cells including cell cycle arrest, cell size increase and polyploidization. We also confirmed that cell type specific markers of megakaryocytes, CD41a and CD42b are induced by phytosphingosine. Phospholipids with highly similar structures were unable to induce similar changes, indicating that the activity of phytosphingosine is highly specific. Although phytosphingosine is known to activate p38 mitogen-activated protein kinase (MAPK)-mediated apoptosis, the signaling mechanisms involved in megakaryopoiesis appear to be distinct. In sum, we present another model for dissecting molecular details of megakaryocytic differentiation which in large part remains obscure. [BMB Reports 2015; 48(12): 691-695]
We have previously reported the effects of 2-(trimethylammonium) ethyl (R)-3-methoxy-3-oxo-2-stearamidopropyl phosphate [( R )-TEMOSPho], a synthetic phospholipid, on megakaryocytic differentiation of myeloid leukemia cells. Here, we demonstrate that ( R )-TEMOSPho enhances megakaryopoiesis and plateletogenesis from primary hematopoietic stem cells (HSCs) induced by thrombopoietin (TPO). Specifically, we demonstrate at sub-saturation levels of TPO, the addition of ( R )-TEMOSPho enhances differentiation and maturation of megakaryocytes (MKs) from murine HSCs derived from fetal liver. Furthermore, we show that production of platelets with ( R )-TEMOSPho in combination with TPO is also more efficient than TPO alone and that platelets generated in vitro with these two agents are as functional as those from TPO alone. TPO can thus be partly replaced by or supplemented with ( R )-TEMOSPho, and this in turn implies that ( R )-TEMOSPho can be useful in efficient platelet production in vitro and potentially be a valuable option in designing cell-based therapy.
Reactive oxygen species (ROS) play important roles as second messengers in a wide array of cellular processes including differentiation of stem cells. We identified Nox4 as the major ROS-generating enzyme whose expression is induced during differentiation of embryoid body (EB) into cells of all three germ layers. The role of Nox4 was examined using induced pluripotent stem cells (iPSCs) generated from Nox4 knockout (Nox4−/−) mouse. Differentiation markers showed significantly reduced expression levels consistent with the importance of Nox4-generated ROS during this process. From transcriptomic analyses, we found insulin-like growth factor 2 (IGF2), a member of a gene family extensively involved in embryonic development, as one of the most down-regulated genes in Nox4−/− cells. Indeed, addition of IGF2 to culture partly restored the differentiation competence of Nox4−/− iPSCs. Our results reveal an important signaling axis mediated by ROS in control of crucial events during differentiation of pluripotent stem cells. Graphical Abstract
In this study, we describe a novel kinase inhibitor AX-0085 which can suppress the induction of PD-L1 expression by Interferon-γ (IFN-γ) in lung adenocarcinoma (LUAD) cells. AX-0085 effectively blocks JAK2/STAT1 signaling initiated by IFN-γ treatment and prevents nuclear localization of STAT1. Importantly, we demonstrate that AX-0085 reverses the IFN-γ-mediated repression of T cell activation in vitro and enhances the anti-tumor activity of anti-PD-1 antibody in vivo when used in combination. Finally, transcriptomic analyses indicated that AX-0085 is highly specific in targeting the IFN-γ-pathway, thereby raising the possibility of applying this reagent in combination therapy with checkpoint inhibitor antibodies. It may be particularly relevant in cases in which PD-L1-mediated T cell exhaustion leads to immunoevasive phenotypes.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.