An Endogenous Vitamin K-Dependent Mechanism Regulates Cell Proliferation in the Brain Subventricular Stem Cell Niche.
Stem Cells. 2012 Jan 30;
Authors: Gely-Pernot A, Coronas V, Harnois T, Prestoz L, Mandairon N, Didier A, Berjeaud JM, Monvoisin A, Bourmeyster N, de Frutos PG, Philippe M, Benzakour O
Abstract
Neural stem cells (NSC) persist in the adult mammalian brain, within the subventricular zone (SVZ). The endogenous mechanisms underpinning SVZ stem and progenitor cell proliferation are not fully elucidated. Vitamin K-dependent proteins (VKDP) are mainly secreted factors that were initially discovered as major regulators of blood coagulation. Warfarin, a widespread anticoagulant, is a vitamin K antagonist that inhibits the production of functional VKDP. We demonstrate that the suppression of functional VKDP production, in vitro, by exposure of SVZ cell cultures to warfarin or in vivo by its intracerebroventricular injection to mice, leads to a substantial increase in SVZ cell proliferation. We identify the anti-coagulant factor, protein S and its structural homolog Gas6, as the two only VKDP produced by SVZ cells and describe the expression and activation pattern of their Tyro3, Axl, Mer tyrosine kinase receptors. Both in vitro and in vivo loss of function studies consisting in either Gas6 gene invalidation or in endogenous protein S neutralization, provided evidence for an important novel regulatory role of these two VKDP in the SVZ neurogenic niche. Specifically, we show that while a loss of Gas6 leads to a reduction in the numbers of stem-like cells and in olfactory bulb neurogenesis, endogenous protein S inhibits SVZ cell proliferation. Our study opens up new perspectives for investigating further the role of vitamin K, VKDP and anticoagulants in neural stem cell biology in health and disease.
PMID: 22290807 [PubMed - as supplied by publisher]
Novel Pathways to Erythropoiesis Induced by Dimerization of Intracellular cMpl in Human Hematopoietic Progenitors.
Stem Cells. 2012 Jan 30;
Authors: Parekh C, Sahaghian A, Kim W, Scholes J, Ge S, Zhu Y, Asgharzadeh S, Hollis R, Kohn D, Ji L, Malvar J, Wang X, Crooks G
Abstract
The cytokine thrombopoietin (Tpo) plays a critical role in hematopoiesis by binding to the extracellular domain, and inducing homodimerization of the intracellular signaling domain of its receptor, cMpl. Mpl homodimerization can also be accomplished by binding of a synthetic ligand to a constitutively expressed fusion protein F36VMpl consisting of a ligand binding domain (F36V) and the intracellular signaling domain of Mpl. Unexpectedly, in contrast to Tpo stimulation, robust erythropoiesis is induced after dimerization of F36VMpl in human CD34+ progenitor cells. The goal of this study was to define the hematopoietic progenitor stages at which dimerization of intracellular Mpl induces erythropoiesis, and the downstream molecular events that mediate this unanticipated effect. Dimerization (in the absence of erythropoietin and other cytokines) in human Common Myeloid Progenitors (CMP) and Megakaryocytic-Erythroid Progenitors (MEP) caused a significant increase in CD34+ cells (p<0.01) and induced all stages of erythropoiesis including production of enucleated red blood cells. In contrast, erythropoiesis was not seen with Tpo stimulation. CD34+ cell expansion was the result of increased cell cycling and survival (p<0.05). Microarray profiling of CD34+ cells demonstrated that a unique transcriptional pattern is activated in progenitors by F36VMpl dimerization. Ligand-inducible dimerization of intracellular Mpl in human myelo-erythroid progenitors induces progenitor expansion and erythropoiesis through molecular mechanisms that are not shared by Tpo stimulation of endogenous Mpl.
PMID: 22290824 [PubMed - as supplied by publisher]
Adipose Tissue-Derived Mesenchymal Stem Cells Improve Revascularization Outcomes to Restore Renal Function in Swine Atherosclerotic Renal Artery Stenosis.
Stem Cells. 2012 Jan 30;
Authors: Eirin A, Zhu XY, Krier JD, Tang H, Jordan KL, Grande JP, Lerman A, Textor SC, Lerman LO
Abstract
Background: Reno-protective strategies are needed to improve renal outcomes in patients with atherosclerotic renal artery stenosis (ARAS). Adipose tissue-derived mesenchymal stem cells (MSC) can promote renal regeneration, but their potential for attenuating cellular injury and restoring kidney repair in ARAS has not been explored. We hypothesized that replenishment of MSC as an adjunct to percutaneous transluminal renal angioplasty (PTRA) would restore renal cellular integrity and improve renal function in ARAS pigs. Methods and Results: Four groups of pigs (n=7 each) were studied after 16 weeks of ARAS, ARAS 4 weeks after PTRA and stenting with or without adjunct intra-renal delivery of MSC (10×10(6) cells), and controls. Stenotic kidney blood flow (RBF) and glomerular filtration rate (GFR) were measured using multidetector CT. Renal microvascular architecture (micro-CT), fibrosis, inflammation, and oxidative stress were evaluated ex-vivo. Four weeks after successful PTRA, mean arterial pressure fell to a similar level in all revascularized groups. Stenotic kidney GFR and RBF remained decreased in ARAS (p=0.01 and p=0.02) and ARAS+PTRA (p=0.02 and p=0.03) compared to normal, but rose to normal levels in ARAS+PTRA+MSC (p=0.34 and p=0.46 vs. normal). Interstitial fibrosis, inflammation, microvascular rarefaction, and oxidative stress were attenuated only in PTRA+MSC-treated pigs. Conclusions: A single intra-renal delivery of MSC in conjunction with renal revascularization restored renal hemodynamics and function, and decreased inflammation, apoptosis, oxidative stress, microvascular loss, and fibrosis. This study suggests a unique and novel therapeutic potential for MSC in restoring renal function when combined with PTRA in chronic experimental renovascular disease.
PMID: 22290832 [PubMed - as supplied by publisher]
Nadph Oxidase 2 Regulates Bone Marrow Microenvironment Following Hindlimb Ischemia: Role in Reparative Mobilization of Progenitor Cells.
Stem Cells. 2012 Jan 30;
Authors: Urao N, McKinney RD, Fukai T, Ushio-Fukai M
Abstract
Bone marrow (BM) microenvironment, which is regulated by hypoxia and proteolytic enzymes, is crucial for stem/progenitor cell function and mobilization involved in postnatal neovascularization. We demonstrated that NADPH oxidase2 (Nox2)-derived reactive oxygen species (ROS) are involved in post-ischemic mobilization of BM cells and revascularization. However, role of Nox2 in regulating BM microenvironment in response to ischemic injury remains unknown. Here we show that hindlimb ischemia of mice increases ROS production in both the endosteal and central region of BM tissue in situ, which is almost completely abolished in Nox2 knockout (KO) mice. This Nox2-dependent ROS production is mainly derived from Gr-1(+) myeloid cells in BM. In vivo injection of hypoxyprobe reveals that endosteum at the BM is hypoxic with high expression of HIF-1α in basal state. Following hindlimb ischemia, hypoxic areas and HIF-1α expression are expanded throughout the BM, which is inhibited in Nox2 KO mice. This ischemia-induced alteration of Nox2-dependent BM microenvironment is associated with an increase in VEGF expression and Akt phosphorylation in BM tissue, thereby promoting Lin(-) progenitor cell survival and expansion, leading to their mobilization from BM. Furthermore, hindlimb ischemia increases proteolytic enzymes MT1-MMP expression and MMP-9 activity in BM, which is inhibited in Nox2 KO mice. In summary, Nox2-dependent increase in ROS play a critical role in regulating hypoxia expansion and proteolytic activities in BM microenvironment in response to tissue ischemia. This in turn promotes progenitor cell expansion and reparative mobilization from BM, leading to post-ischemic neovascularization and tissue repair.
PMID: 22290850 [PubMed - as supplied by publisher]
SFRP1 and 2 Dose-Dependently Regulate Midbrain Dopamine Neuron Development In vivo and in Embryonic Stem Cells.
Stem Cells. 2012 Jan 30;
Authors: Kele J, Andersson ER, Villaescusa JC, Cajanek L, Parish CL, Bonilla S, Toledo EM, Bryja V, Rubin JS, Shimono A, Arenas E
Abstract
Secreted Frizzled related proteins (sFRPs) are a family of proteins that modulate Wnt signaling, which in its turn regulates multiple aspects of ventral midbrain (VM) and dopaminergic (DA) neuron development. However, it is not known which Wnt signaling branch and what aspects of midbrain DA neuron development are regulated by sFRPs. Here, we show that sFRP1 and sFRP2 activate the Wnt/PCP/Rac1 pathway in DA cells. In the developing VM, sFRP1 and sFRP2 are expressed at low levels and sFRP1-/- or sFRP2-/- mice had no detectable phenotype. However, compound sFRP1-/-; sFRP2-/- mutants revealed a Wnt/PCP phenotype similar to that previously described for Wnt5a-/- mice. This included an antero-posterior shortening of the VM, a lateral expansion of the Shh domain and DA lineage markers (Lmx1a and Th), as well as an accumulation of Nurr1+ precursors in the VM. In vitro experiments showed that, while very high concentrations of SFRP1 had a negative effect on cell survival, low/medium concentrations of sFRP1 or sFRP2 promoted the DA differentiation of progenitors derived from primary VM cultures or mouse embryonic stem cells (mESC), mimicking the effects of Wnt5a. We thus conclude that the main function of sFRP1 and sFRP2 is to enhance Wnt/PCP signaling in DA cells and to regulate Wnt/PCP-dependent functions in midbrain development. Moreover, we suggest that low-medium concentrations of sFRPs may be used to enhance the DA differentiation of ES cells and improve their therapeutic application.
PMID: 22290867 [PubMed - as supplied by publisher]
Authors: Zhao C, Xiu Y, Ashton J, Xing L, Morita Y, Jordan CT, Boyce BF
Abstract
RelB and NF-κB2 are the main effectors of NF-κB non-canonical signaling and play critical roles in many physiological processes. However, their role in hematopoietic stem/progenitor cell (HSPC) maintenance has not been characterized. To investigate this, we generated RelB/NF-κB2 double-knockout (dKO) mice and found that dKO HSPCs have profoundly impaired engraftment and self-renewal activity after transplantation into wild-type recipients. Transplantation of wild-type bone marrow cells into dKO mice to assess the role of the dKO microenvironment showed that wild-type HSPCs cycled more rapidly, were more abundant, and had developmental aberrancies: increased myeloid and decreased lymphoid lineages, similar to dKO HSPCs. Notably, when these wild-type cells were returned to normal hosts, these phenotypic changes were reversed, indicating a potent but transient phenotype conferred by the dKO microenvironment. However, dKO bone marrow stromal cell numbers were reduced, and bone-lining niche cells supported less HSPC expansion than controls. Further, increased dKO HSPC proliferation was associated with impaired expression of niche adhesion molecules by bone-lining cells and increased inflammatory cytokine expression by bone marrow cells. Thus, RelB/NF-κB2 signaling positively and intrinsically regulates HSPC self-renewal and maintains stromal/osteoblastic niches and negatively and extrinsically regulates HSPC expansion and lineage commitment through the marrow microenvironment.
PMID: 22290873 [PubMed - as supplied by publisher]
Intratumoral Heterogeneity in the Self-Renewal and Tumorigenic Differentiation of Ovarian Cancer.
Stem Cells. 2012 Jan 20;
Authors: Abelson S, Shamai Y, Berger L, Shouval R, Skorecki K, Tzukerman M
Abstract
Resistance to anticancer therapy has been attributed to inter-individual differences in gene expression pathways among tumors, and to the existence within tumors of cancer stem cells with self-renewal capacity. In previous studies we have demonstrated that the human embryonic stem cells (hESC)-derived cellular microenvironment in immunocompromised mice, enables functional distinction of heterogeneous tumor cells, including cells which do not grow into a tumor in conventional direct tumor xenograft platform. In the current study, we use clonally expanded subpopulations derived from ovarian clear cell carcinoma of a single tumor, to demonstrate striking intratumoral phenotypic heterogeneity that is dynamically dependent on the tumor growth microenvironment. Each of six clonally expanded subpopulations displays a different level of morphologic and tumorigenic differentiation, wherein growth in the hESC-derived microenvironment favors growth of CD44,+ aldehyde dehydrogenase positive pockets of self-renewing cells that sustain tumor growth through a process of tumorigenic differentiation into CD44- aldehyde dehydrogenase negative derivatives. Strikingly, these derivative cells display microenvironment-dependent plasticity with the capacity to restore self-renewal and CD44 expression. Such intratumoral heterogeneity and plasticity at the level of the key properties of self-renewal and tumorigenic differentiation, suggests that a paradigm shift is needed in the approach to anti-cancer therapy, with the aim of turning malignant growth into a chronic manageable disorder, based on continual monitoring of these tumor growth properties. The hESC-based in vivo model renders intratumoral heterogeneity in the self renewal and tumorigenic differentiation amenable to biological analysis as well as anticancer therapy testing.
PMID: 22267284 [PubMed - as supplied by publisher]
Mechanism of Action of HOXB4 on the Haematopoietic Differentiation of Embryonic Stem Cells.
Stem Cells. 2012 Jan 20;
Authors: Forrester LM, Jackson M
Abstract
Pluripotent stem cells can be differentiated into haematopoietic lineages in vitro and hold promise for the future treatment of haematological disease. Differentiation strategies involving defined factors in serum-free conditions have been successful in producing hematopoietic progenitors and some mature cell types from mouse and human embryonic stem cells and induced pluripotent cells. However, these precisely defined protocols are relatively inefficient and have not been used successfully to produce haematopoietic stem cells capable of multi-lineage long term reconstitution of the haematopoietic system. More complex differentiation induction strategies including co-culture with stromal cells derived from sites of haematopoietic activity in vivo and enforced expression of reprogramming transcription factors, such as HOXB4 have been required to increase the efficiency of the differentiation procedure and to produce these most potent haematopoietic stem cells. We review the studies that have used HOXB4 to improve haematopoietic differentiation from pluripotent cells focusing on studies that have provided some insight into its mechanism of action. A better understanding of the molecular pathways involved in the action of HOXB4 might lead to more defined culture systems and safer protocols for clinical translation.
PMID: 22267295 [PubMed - as supplied by publisher]
Efficient Stage Specific Differentiation of Human Pluripotent Stem Cells Towards Retinal Photoreceptor Cells.
Stem Cells. 2012 Jan 20;
Authors: Mellough CB, Sernagor E, Moreno-Gimeno I, Steel DH, Lako M
Abstract
Recent successes in the stem cell field have identified some of the key chemical and biological cues which drive photoreceptor derivation from human embryonic stem cells (hESC) and human induced pluripotent stem cells (hiPSC), however the efficiency of this process is variable. We have designed a three-step photoreceptor differentiation protocol combining previously published methods that direct the differentiation of hESC and hiPSC towards a retinal lineage which we further modified with additional supplements selected on the basis of reports from eye field and retinal development. We report that hESC and hiPSC differentiating under our regime over a 60 day period sequentially acquire markers associated with neural, retinal field, retinal pigmented epithelium and photoreceptor cells, including mature photoreceptor markers OPN1SW and RHODOPSIN with a higher efficiency than previously reported. In addition, we report the ability of hESC and hiPSC cultures to generate neural and retinal phenotypes under minimal culture conditions which may be linked to their ability to endogenously up-regulate the expression of a range of factors important for retinal cell type specification. However, cultures that were differentiated with full supplementation under our photoreceptor-induction regime achieve this within a significantly shorter time frame and show a substantial increase in the expression of photoreceptor-specific markers in comparison to cultures differentiated under minimal conditions. Interestingly, cultures supplemented only with B27 and/or N2 displayed comparable differentiation efficiency to those under full supplementation, indicating a key role for B27 and N2 during the differentiation process. Furthermore, our data highlight an important role for Dkk1 and Noggin in enhancing the differentiation of hESC and hiPSC towards retinal progenitor cells and photoreceptor precursors during the early stages of differentiation, while suggesting that further maturation of these cells into photoreceptors may not require additional factors and can ensue under minimal culture conditions.
PMID: 22267304 [PubMed - as supplied by publisher]
Induction of Osteogenesis in Mesenchymal Stem Cells by Activated Monocytes/Macrophages Depends on Oncostatin M Signaling.
Stem Cells. 2012 Jan 20;
Authors: Guihard P, Danger Y, Brounais B, David E, Brion R, Delecrin J, Richards CD, Chevalier S, Rédini F, Heymann D, Gascan H, Blanchard F
Abstract
Bone resorption by osteoclasts and bone formation by osteoblasts are tightly coupled processes implicating factors in TNF, bone morphogenetic protein and Wnt families. In osteoimmunology, macrophages were described as another critical cell population regulating bone formation by osteoblasts but the coupling factors were not identified. Using a high throughput approach, we identified here Oncostatin M (OSM), a cytokine of the IL-6 family, as a major coupling factor produced by activated circulating CD14(+) or bone marrow CD11b(+) monocytes/macrophages that induces osteoblast differentiation and matrix mineralization from human mesenchymal stem cells (MSC) while inhibiting adipogenesis. Upon toll-like receptors (TLRs) activation by lipopolysaccharide or endogenous ligands, OSM was produced in classically activated inflammatory M1 and not M2 macrophages, through a cyclooxygenase-2 and prostaglandin-E2 regulatory loop. Stimulation of osteogenesis by activated monocytes/macrophages was prevented using neutralizing antibodies or siRNA to OSM, OSM receptor subunits gp130 and OSMR or to the downstream transcription factor STAT3. The induced osteoblast differentiation program culminated with enhanced expression of C/EBPδ (CCAAT-enhancer-binding protein δ), Cbfa1 and alkaline phosphatase. Overexpression of OSM in the tibia of mice has led to new bone apposition with no sign of bone resorption. Two other cytokines had also a potent role in bone formation induced by monocytes/macrophages and TLRs activation: IL-6 and Leukemia inhibitory factor. We propose that during bone inflammation, infection or injury, the IL-6 family signaling network activated by macrophages and TLR ligands stimulates bone formation that is largely uncoupled from bone resorption and is thus an important target for anabolic bone therapies.
PMID: 22267310 [PubMed - as supplied by publisher]
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