Key Spec Table
|Species Reactivity||Key Applications||Host||Format||Antibody Type|
|H||WB, ICC, IHC, IH(P)||Rb||Serum||Polyclonal Antibody|
|Safety Information according to GHS|
|Storage and Shipping Information|
|Storage Conditions||Stable at -20°C in undiluted aliquots for up to 6 months from date of receipt. Avoid repeated freeze/thaw cycles.|
|Material Size||50 µL|
|Anti-Nestin - LV1635955||LV1635955|
|Anti-Nestin - LV1751778||LV1751778|
|Anti-Nestin - NG1853940||NG1853940|
|RABBIT ANTI-NESTIN POLYCLONAL ANTIBODY - LV1519217||LV1519217|
References | 32 Available | See All References
|Reference overview||Species||Pub Med ID|
|Synaptic dysregulation in a human iPS cell model of mental disorders. |
Wen, Z; Nguyen, HN; Guo, Z; Lalli, MA; Wang, X; Su, Y; Kim, NS; Yoon, KJ; Shin, J; Zhang, C; Makri, G; Nauen, D; Yu, H; Guzman, E; Chiang, CH; Yoritomo, N; Kaibuchi, K; Zou, J; Christian, KM; Cheng, L; Ross, CA; Margolis, RL; Chen, G; Kosik, KS; Song, H; Ming, GL
Dysregulated neurodevelopment with altered structural and functional connectivity is believed to underlie many neuropsychiatric disorders, and 'a disease of synapses' is the major hypothesis for the biological basis of schizophrenia. Although this hypothesis has gained indirect support from human post-mortem brain analyses and genetic studies, little is known about the pathophysiology of synapses in patient neurons and how susceptibility genes for mental disorders could lead to synaptic deficits in humans. Genetics of most psychiatric disorders are extremely complex due to multiple susceptibility variants with low penetrance and variable phenotypes. Rare, multiply affected, large families in which a single genetic locus is probably responsible for conferring susceptibility have proven invaluable for the study of complex disorders. Here we generated induced pluripotent stem (iPS) cells from four members of a family in which a frameshift mutation of disrupted in schizophrenia 1 (DISC1) co-segregated with major psychiatric disorders and we further produced different isogenic iPS cell lines via gene editing. We showed that mutant DISC1 causes synaptic vesicle release deficits in iPS-cell-derived forebrain neurons. Mutant DISC1 depletes wild-type DISC1 protein and, furthermore, dysregulates expression of many genes related to synapses and psychiatric disorders in human forebrain neurons. Our study reveals that a psychiatric disorder relevant mutation causes synapse deficits and transcriptional dysregulation in human neurons and our findings provide new insight into the molecular and synaptic etiopathology of psychiatric disorders.
|Different expression patterns of Phactr family members in normal and injured mouse brain. |
J Y Kim,S Y Choi,Y Moon,H J Kim,J H Chin,H Kim,W Sun
Neuroscience 221 2012
Phosphatase and actin regulators (Phactrs) are a novel family of proteins expressed in the brain, and they exhibit both strong modulatory activity of protein phosphatase 1 and actin-binding activity. Phactrs are comprised of four family members (Phactr1-4), but their detailed expression patterns during embryonic and postnatal development are not well understood. We found that these family members exhibit different spatiotemporal mRNA expression patterns. Phactr4 mRNA was found in neural stem cells in the developing and adult brains, whereas Phactr1 and 3 appeared to be expressed in post-mitotic neurons. Following traumatic brain injury which promotes neurogenesis in the neurogenic region and gliogenesis in the injury penumbra, the mRNA expression of phactr2 and 4 was progressively increased in the injury penumbra, and phactr4 mRNA and protein induction was observed in reactive astrocytes. These differential expression patterns of phactrs imply specific functions for each protein during development, and the importance of Phactr4 in the reactive gliosis following brain injury.
|Glioblastoma resistance to anti-VEGF therapy is associated with myeloid cell infiltration, stem cell accumulation, and a mesenchymal phenotype. |
Yuji Piao,Ji Liang,Lindsay Holmes,Amado J Zurita,Verlene Henry,John V Heymach,John F de Groot
Neuro-oncology 14 2012
Vascular endothelial growth factor (VEGF) is a critical regulator of angiogenesis. Inhibiting the VEGF-VEGF receptor (R) signal transduction pathway in glioblastoma has recently been shown to delay progression, but the relative benefit and mechanisms of response and failure of anti-VEGF therapy and VEGFR inhibitors are not well understood. The purpose of our study was to evaluate the relative effectiveness of VEGF sequestration and/or VEGFR inhibition on orthotopic tumor growth and the mechanism(s) of treatment resistance. We evaluated, not only, the effects of anti-VEGF therapy (bevacizumab), anti-VEGFR therapy (sunitinib), and the combination on the survival of mice bearing orthotopic gliomas, but also the differential effects of the treatments on tumor vascularity, cellular proliferation, mesenchymal and stem cell markers, and myeloid cell infiltration using flow cytometry and immunohistochemistry. Bevacizumab significantly prolonged survival compared with the control or sunitinib alone. Both antiangiogenic agents initially reduced infiltration of macrophages and tumor vascularity. However, multitargeted VEGFR inhibition, but not VEGF sequestration, rapidly created a vascular gradient and more rapidly induced tumor hypoxia. Re-infiltration of macrophages was associated with the induction of hypoxia. Combination treatment with bevacizumab and sunitinib improved animal survival compared with bevacizumab therapy alone. However, at the time of tumor progression, a significant increase in CD11b(+)/Gr1(+) granulocyte infiltration was observed, and tumors developed aggressive mesenchymal features and increased stem cell marker expression. Collectively, our results demonstrate a more prolonged decrease in tumor vascularity with bevacizumab than with sunitinib, associated with a delay in the development of hypoxia and sustained reduction of infiltrated myeloid cells.
|Girdin maintains the stemness of glioblastoma stem cells. |
A Natsume,T Kato,S Kinjo,A Enomoto,H Toda,S Shimato,F Ohka,K Motomura,Y Kondo,T Miyata,M Takahashi,T Wakabayashi
Oncogene 31 2012
Glioblastomas (GBMs) are the most common and aggressive type of brain tumor. GBMs usually show hyperactivation of the PI3K-Akt pathway, a pro-tumorigenic signaling cascade that contributes to pathogenesis. Girdin, an actin-binding protein identified as a novel substrate of Akt, regulates the sprouting of axons and the migration of neural progenitor cells during early postnatal-stage neurogenesis in the hippocampus. Here, we show that Girdin is highly expressed in human glioblastoma (GBM). Stable Girdin knockdown in isolated GBM stem cells resulted in decreased expression of stem cell markers, including CD133, induced multilineage neural differentiation, and inhibited in vitro cell motility, ex vivo invasion, sphere-forming capacity and in vivo tumor formation. Furthermore, exogenous expression of the Akt-binding domain of Girdin, which competitively inhibits its Akt-mediated phosphorylation, diminished the expression of stem cell markers, SOX2 and nestin, and migration on the brain slice and induced the expression of neural differentiation markers glial fibrillary acidic protein/βIII Tubulin. Our results reveal that Girdin is required for GBM-initiating stem cells to sustain the stemness and invasive properties.
|Osteoblastic expansion induced by parathyroid hormone receptor signaling in murine osteocytes is not sufficient to increase hematopoietic stem cells. |
Laura M Calvi,Olga Bromberg,Yumie Rhee,Jonathan M Weber,Julianne N P Smith,Miles J Basil,Benjamin J Frisch,Teresita Bellido
Blood 119 2012
Microenvironmental expansion of hematopoietic stem cells (HSCs) is induced by treatment with parathyroid hormone (PTH) or activation of the PTH receptor (PTH1R) in osteoblastic cells; however, the osteoblastic subset mediating this action of PTH is unknown. Osteocytes are terminally differentiated osteoblasts embedded in mineralized bone matrix but are connected with the BM. Activation of PTH1R in osteocytes increases osteoblastic number and bone mass. To establish whether osteocyte-mediated PTH1R signaling expands HSCs, we studied mice expressing a constitutively active PTH1R in osteocytes (TG mice). Osteoblasts, osteoclasts, and trabecular bone were increased in TG mice without changes in BM phenotypic HSCs or HSC function. TG mice had progressively increased trabecular bone but decreased HSC function. In severely affected TG mice, phenotypic HSCs were decreased in the BM but increased in the spleen. TG osteocytes had no increase in signals associated with microenvironmental HSC support, and the spindle-shaped osteoblastic cells that increased with PTH treatment were not present in TG bones. These findings demonstrate that activation of PTH1R signaling in osteocytes does not expand BM HSCs, which are instead decreased in TG mice. Therefore, osteocytes do not mediate the HSC expansion induced by PTH1R signaling. Further, osteoblastic expansion is not sufficient to increase HSCs.
|Spatio-temporal dynamics, differentiation and viability of human neural stem cells after implantation into neonatal rat brain. |
Kallur T, Farr TD, Böhm-Sturm P, Kokaia Z, Hoehn M
The European journal of neuroscience 34 382-93. doi 2011
Neural stem cells (NSCs) have attracted major research interest due to their potential use in cell replacement therapy. In patients, human cells are the preferred choice, one source of human NSCs being the brain of fetuses. The aims of the present study were to explore the long-term differentiation, mobility and viability of NSCs derived from the human fetal striatum in response to intracerebral implantation. To investigate long-term spatio-temporal and functional dynamics of grafts in vivo by magnetic resonance imaging, these cells were labeled with superparamagnetic iron oxide (SPIO) nanoparticles prior to implantation. SPIO-labeling of human NSCs left the quantitative profile of the proliferation, cell composition and differentiation capacity of the cells in vitro unaltered. Also after transplantation, the phenotypes after long-term cell differentiation were not significantly different from naïve cells. Upon transplantation, we detected a hypointensity corresponding to the striatal graft location in all animals and persisting for at least 4 months. The hypointense signal appeared visually similar both in location and in volume over time. However, quantitative volumetric analysis showed that the detectable, apparent graft volume decreased significantly from 3 to 16 weeks. Finally, the human NSCs were not proliferating after implantation, indicating lack of tumor formation. These cells are thus a promising candidate for translationally relevant investigations for stem cell-based regenerative therapies.© 2011 The Authors. European Journal of Neuroscience © 2011 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.
|MEK-ERK Signaling Dictates DNA-Repair Gene MGMT Expression and Temozolomide Resistance of Stem-Like Glioblastoma Cells via the MDM2-p53 Axis. |
Sato A, Sunayama J, Matsuda K, Seino S, Suzuki K, Watanabe E, Tachibana K, Tomiyama A, Kayama T, Kitanaka C
Stem cells (Dayton, Ohio) 29 1942-51. doi 2011
Overcoming the resistance of glioblastoma cells against temozolomide, the first-line chemotherapeutic agent of choice for newly diagnosed glioblastoma, is a major therapeutic challenge in the management of this deadly brain tumor. The gene encoding O(6) -methylguanine DNA methyltransferase (MGMT), which removes the methyl group attached by temozolomide, is often silenced by promoter methylation in glioblastoma but is nevertheless expressed in a significant fraction of cases and is therefore regarded as one of the most clinically relevant mechanisms of resistance against temozolomide. However, to date, signaling pathways regulating MGMT in MGMT-expressing glioblastoma cells have been poorly delineated. Here in this study, we provide lines of evidence that the mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK)-extracellular signal-regulated kinase (ERK)--murine double minute 2 (MDM2)-p53 pathway plays a critical role in the regulation of MGMT expression, using stem-like glioblastoma cells directly derived from patient tumor samples and maintained in the absence of serum, which not only possess stem-like properties but are also known to phenocopy the characteristics of the original tumors from which they are derived. We show that, in stem-like glioblastoma cells, MEK inhibition reduced MDM2 expression and that inhibition of either MEK or MDM2 resulted in p53 activation accompanied by p53-dependent downregulation of MGMT expression. MEK inhibition rendered otherwise resistant stem-like glioblastoma cells sensitive to temozolomide, and combination of MEK inhibitor and temozolomide treatments effectively deprived stem-like glioblastoma cells of their tumorigenic potential. Our findings suggest that targeting of the MEK-ERK-MDM2-p53 pathway in combination with temozolomide could be a novel and promising therapeutic strategy in the treatment of glioblastoma. STEM CELLS 2011;29:1942-1951.Copyright © 2011 AlphaMed Press.
|Ginsenoside Rd attenuates mitochondrial dysfunction and sequential apoptosis after transient focal ischemia. |
Ye R, Zhang X, Kong X, Han J, Yang Q, Zhang Y, Chen Y, Li P, Liu J, Shi M, Xiong L, Zhao G
We previously found that ginsenoside Rd (Rd), one of the major active ingredients in Panax ginseng, protects neuronal cells from hydrogen peroxide and oxygen-glucose deprivation, an in vitro model of cerebral ischemia. In this study, we examined the protective effects of Rd in an animal model of focal cerebral ischemia. Rats administered with Rd or vehicle were subjected to transient middle cerebral artery occlusion (MCAO). Rd (50 mg/kg) significantly reduced the infarct volume by 52.8%. This reduction of injury volume was associated with an improvement in neurological function and was sustained for at least 2 weeks after the induction of ischemia. To evaluate the underlying mechanisms of Rd against stroke, brain tissues were assayed for mitochondrial enzyme activities, mitochondrial membrane potential (MMP), production of reactive oxygen species (ROS), energy metabolites, and apoptosis. Rd markedly protected mitochondria as indicated by preserved respiratory chain complex activities and aconitase activity, lowered mitochondrial hydrogen peroxide production, and hyperpolarized MMP. Microdialysis results illustrated that Rd significantly decreased the accumulation of lactate, the end product of anaerobic glycolysis, and increased pyruvate, the end product of aerobic glycolysis, hence inducing a lower lactate/pyruvate ratio. Additionally, in vitro studies further exhibited that Rd protected isolated mitochondria from calcium-induced damage by attenuating mitochondrial swelling, preserving MMP and decreasing ROS production. Moreover, Rd treatment reduced mitochondrial release of cytochrome c (CytoC) and apoptosis-inducing factor (AIF), thereby minimizing mitochondria-mediated apoptosis following ischemia. In conclusion, these findings demonstrated that Rd exerts neuroprotective effects in transient focal ischemia, which may involve an integrated process of the mitochondrial protection, energy restoration and inhibition of apoptosis.Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights reserved.
|Doublecortin-positive cells in the adult primate cerebral cortex and possible role in brain plasticity and development. |
Jocelyne Bloch,Mélanie Kaeser,Yalda Sadeghi,Eric M Rouiller,D Eugene Redmond,Jean-François Brunet
The Journal of comparative neurology 519 2011
We have demonstrated that cortical cell autografts might be a useful therapy in two monkey models of neurological disease: motor cortex lesion and Parkinson's disease. However, the origin of the useful transplanted cells obtained from cortical biopsies is not clear. In this report we describe the expression of doublecortin (DCX) in these cells based on reverse-transcription polymerase chain reaction (RT-PCR) and immunodetection in the adult primate cortex and cell cultures. The results showed that DCX-positive cells were present in the whole primate cerebral cortex and also expressed glial and/or neuronal markers such as glial fibrillary protein (GFAP) or neuronal nuclei (NeuN). We also demonstrated that only DCX/GFAP positive cells were able to proliferate and originate progenitor cells in vitro. We hypothesize that these DCX-positive cells in vivo have a role in cortical plasticity and brain reaction to injury. Moreover, in vitro these DCX-positive cells have the potential to reacquire progenitor characteristics that confirm their potential for brain repair.
|Expression of embryonic stem cell markers SOX2 and nestin in dermatofibrosarcoma protuberans and dermatofibroma. |
Klaus Sellheyer,Paula Nelson,Rajiv M Patel
Journal of cutaneous pathology 38 2011
Dermatofibrosarcoma protuberans (DFSP) was recently proposed to originate from a nestin-positive stem cell. In postnatal skin, nestin and another embryonic stem cell marker, SOX2, display a similar expression pattern with immunoreactivity in the hair follicle papilla and scattered cells of the perifollicular connective tissue sheath. The distribution pattern differs only in early embryogenesis, when nestin but not SOX2 is also expressed throughout the entire interfollicular dermis. We speculated that DFSP would not only be nestin-positive but also SOX2-positive.
|Complement Receptor 2 Is Expressed in Neural Progenitor Cells and Regulates Adult Hippocampal Neurogenesis. |
Moriyama M, Fukuhara T, Britschgi M, He Y, Narasimhan R, Villeda S, Molina H, Huber BT, Holers M, Wyss-Coray T
J Neurosci 31 3981-3989. 2011
Injury and inflammation are potent regulators of adult neurogenesis. As the complement system forms a key immune pathway that may also exert critical functions in neural development and neurodegeneration, we asked whether complement receptors regulate neurogenesis. We discovered that complement receptor 2 (CR2), classically known as a coreceptor of the B-lymphocyte antigen receptor, is expressed in adult neural progenitor cells (NPCs) of the dentate gyrus. Two of its ligands, C3d and interferon-α (IFN-α), inhibited proliferation of wild-type NPCs but not NPCs derived from mice lacking Cr2 (Cr2(-/-)), indicating functional Cr2 expression. Young and old Cr2(-/-) mice exhibited prominent increases in basal neurogenesis compared with wild-type littermates, whereas intracerebral injection of C3d resulted in fewer proliferating neuroblasts in wild-type than in Cr2(-/-) mice. We conclude that Cr2 regulates hippocampal neurogenesis and propose that increased C3d and IFN-α production associated with brain injury or viral infections may inhibit neurogenesis.Full Text Article
|Secreted factors from brain endothelial cells maintain glioblastoma stem-like cell expansion through the mTOR pathway. |
Galan-Moya, Eva Maria, et al.
EMBO Rep., 12: 470-6 (2011) 2011
Glioma stem-cells are associated with the brain vasculature. However, the way in which this vascular niche regulates stem-cell renewal and fate remains unclear. Here, we show that factors emanating from brain endothelial cells positively control the expansion of long-term glioblastoma stem-like cells. We find that both pharmacological inhibition of and RNA interference with the mammalian target of rapamycin (mTOR) pathway reduce their spheroid growth. Conversely, the endothelial secretome is sufficient to promote this mTOR-dependent survival. Thus, interfering with endothelial signals might present opportunities to identify treatments that selectively target malignant stem-cell niches.
|Forest trees influence distribution of the mineral weathering bacterial communities from the Scleroderma citrinum mycorrhizosphere. |
Calvaruso C, Turpault MP, Leclerc E, Ranger J, Garbaye J, Uroz S, Frey-Klett P
Appl Environ Microbiol 2010
In acidic forest soils, availability of inorganic nutrients is a tree-growth limiting factor. A hypothesis to explain sustainable forest development proposes that tree roots select soil microbes involved in central biogeochemical processes such as mineral weathering that may contribute to nutrient mobilization and tree nutrition. Here we showed, by combining soil analyses to cultivation-dependent analyses of the culturable bacterial communities associated to the widespread mycorrhizal fungus Scleroderma citrinum, a significant enrichment of bacterial isolates with efficient mineral weathering potentials around the oak and beech mycorrhizal roots compared to bulk soil. Such a difference did not exist in the rhizosphere of Norway spruce. The mineral weathering ability of the bacterial isolates was assessed using a microplaque assay that measures the pH and the amount of iron released from biotite. Using this microplate assay, we demonstrated that the bacterial isolates harbouring the most efficient mineral weathering potential belonged to the Burkholderia genus. Notably, previous work revealed that oak and beech harboured very similar pH in the 5- to 10-cm horizon in both rhizosphere and bulk soil environments. In the spruce rhizosphere in contrast, pH was significantly lower than in bulk soil. Because the production of proton is one of the main mechanisms responsible for mineral weathering, our results suggest that certain tree species have developed indirect strategies for mineral weathering in nutrient-poor soils, which lie on the selection of bacterial communities with efficient mineral weathering potentials.
|Human tooth germ stem cells preserve neuro-protective effects after long-term cryo-preservation. |
ME Yalvac, M Ramazanoglu, M Tekguc, OF Bayrak, AK Shafigullina, II Salafutdinov, NL Blatt, AP Kiyasov, F Sahin, A Palotas, AA Rizvanov
Current neurovascular research 7 49-58 2010
The use of mesenchymal stem cells (MSCs) has been shown to be promising in chronic disorders such as diabetes, Alzheimer's dementia, Parkinson's disease, spinal cord injury and brain ischemia. Recent studies revealed that human tooth germs (hTG) contain MSCs which can be easily isolated, expanded and cryo-preserved. In this report, we isolated human tooth germ stem cells (hTGSCs) with MSC characteristics from third molar tooth germs, cryo-preserved them at -80( degrees )C for 6 months, and evaluated for their surface antigens, expression of pluri-potency associated genes, differentiation capacity, karyotype, and proliferation rate. These characteristics were compared to their non-frozen counterparts. In addition, neuro-protective effects of cryo-preserved cells on neuro-blastoma SH-SY5Y cells were also assessed after exposure to stress conditions induced by hydrogen-peroxide (oxidative stress) and paclitaxel (microtubule stabilizing mitotic inhibitor). After long term cryo-preservation hTGSCs expressed surface antigens CD29, CD73, CD90, CD105, and CD166, but not CD34, CD45 or CD133, which was typical for non-frozen hTGSCs. Cryo-preserved hTGSCs were able to differentiate into osteo-, adipo- and neuro-genic cells. They also showed normal karyotype after high number of population doublings and unchanged proliferation rate. On the other hand, cryo-preserved cells demonstrated a tendency for lower level of pluri-potency associated gene expression (nanog, oct4, sox2, klf4, c-myc) than non-frozen hTGSCs. hTGSCs conditioned media increased survival of SH-SY5Y cells exposed to oxidative stress or paclitaxel. These findings confirm that hTGSCs preserve their major characteristics and exert neuro-protection after long-term cryo-preservation, suggesting that hTGSCs, harvested from young individuals and stored for possible use later as they grow old, might be employed in cellular therapy of age-related degenerative disorders.,
|Vein tissue expression of matrix metalloproteinase as biomarker for hemodialysis arteriovenous fistula maturation. |
Lee ES, Shen Q, Pitts RL, Guo M, Wu MH, Yuan SY
Vasc Endovascular Surg 44 674-9. Epub 2010 Aug 18. 2010
Failure of arteriovenous fistula (AVF) maturation is attributed to impaired vein remodeling. The purpose of this study is to identify whether vein matrix metalloproteinase (MMP) expression and activity is associated with AVF maturation. Patients with renal insufficiency undergoing surgery had their vein segments harvested and snap-frozen at time of AVF construction. Expression of MMP-2, MMP-9, membrane type-1 MMP (MT1-MMP), tissue inhibitor of metallopreoteinases type 2 (TIMP-2), and TIMP-4 were measured using zymography and Western blotting techniques. Of 14 patients enrolled, 9 had successful maturation and 5 had failure of AVF maturation. Significantly higher levels of MT1-MMP (an MMP-2 activator; P = .01), TIMP-2 (an MMP-2 inhibitor; P = .03), MMP-2 latent (P = .02), and MMP-2 total (P = .03) were associated with AVF maturation. There was a trend toward higher levels of TIMP-4 in the successful group (P = .18). These data demonstrate a positive relationship between MMP-2 expression in veins and AVF maturation. MMP-2 could serve as a potential preoperative marker to predict maturation.
|Caloric Restriction Reverses Hepatic Insulin Resistance and Steatosis in Rats with Low Aerobic Capacity. |
Bowman TA, Ramakrishnan SK, Kaw M, Lee SJ, Patel PR, Golla VK, Bourey RE, Haram PM, Koch LG, Britton SL, Wisløff U, Lee AD, Najjar SM
Rats selectively bred for low aerobic running capacity exhibit the metabolic syndrome, including hyperinsulinemia, insulin resistance, visceral obesity, and dyslipidemia. They also exhibit features of nonalcoholic steatohepatitis, including chicken-wire fibrosis, inflammation, and oxidative stress. Hyperinsulinemia in these rats is associated with impaired hepatic insulin clearance. The current studies aimed to determine whether these metabolic abnormalities could be reversed by caloric restriction (CR). CR by 30% over a period of 2-3 months improved insulin clearance in parallel to inducing the protein content and activation of the carcinoembryonic antigen-related cell adhesion molecule 1, a main player in hepatic insulin extraction. It also reduced glucose and insulin intolerance and serum and tissue (liver and muscle) triglyceride levels. Additionally, CR reversed inflammation, oxidative stress, and fibrosis in liver. The data support a significant role of CR in the normalization of insulin and lipid metabolism in liver.
|Maturation of tumor vasculature by interferon-beta disrupts the vascular niche of glioma stem cells. |
R F Williams,T L Sims,L Tracey,A L Myers,C Y C Ng,H Poppleton,A C Nathwani,A M Davidoff
Anticancer research 30 2010
The vascular niche necessary for cancer stem cell maintenance is a potential target for cancer therapy.
|Systems-level metabolic flux profiling elucidates a complete, bifurcated TCA cycle in Clostridium acetobutylicum. |
Amador-Noguez D, Feng XJ, Fan J, Roquet N, Rabitz H, Rabinowitz JD
J Bacteriol 2010
Obligatory anaerobic bacteria are major contributors to the overall metabolism of soil and the human gut. The metabolic pathways of these bacteria remain, however, poorly understood. Here we directly map, using isotope tracers, mass spectrometry, and quantitative flux modeling, the metabolic pathways of Clostridium acetobutylicum, a soil bacterium whose major fermentation products include the biofuels butanol and hydrogen. While genome annotation suggested the absence of most TCA cycle enzymes, our results demonstrate that this bacterium has a complete, albeit bifurcated, TCA cycle: oxaloacetate flows to succinate both through citrate/alpha-ketoglutarate and via malate/fumarate. Our investigations also yielded insights into the pathways utilized for glucose catabolism and amino acid biosynthesis and revealed that the organism's one-carbon metabolism is distinct from that of model microbes, involving reversible pyruvate decarboxylation and use of pyruvate as the one-carbon donor for biosynthetic reactions. This study represents the first in vivo characterization of the TCA cycle and central metabolism of C. acetobutylicum. Our results establish a role for the full TCA cycle in an obligatory anaerobic organism, and demonstrate the importance of complementing genome annotation with isotope tracer studies for determining the metabolic pathways of diverse microbes.
|Large Litter Rearing Enhances Leptin Sensitivity and Protects Selectively Bred Diet-Induced Obese Rats from Becoming Obese. |
Patterson CM, Bouret SG, Park S, Irani BG, Dunn-Meynell AA, Levin BE
Because rearing rats in large litters (LLs) protects them from becoming obese, we postulated that LL rearing would protect rats selectively bred to develop diet-induced obesity (DIO) from becoming obese by overcoming their inborn central leptin resistance. Male and female DIO rats were raised in normal litters (NLs; 10 pups/dam) or LLs (16 pups/dam) and assessed for anatomical, biochemical, and functional aspects of leptin sensitivity at various ages when fed low-fat chow or a 31% fat high-energy (HE) diet. LL rearing reduced plasma leptin levels by postnatal day 2 (P2) and body weight gain by P8. At P16, LL DIO neonates had increased arcuate nucleus (ARC) binding of leptin to its extracellular receptors and at P28 an associated increase of their agouti-related peptide and alpha-MSH axonal projections to the paraventricular nucleus. Reduced body weight persisted and was associated with increased ARC leptin receptor binding and sensitivity to the anorectic effects of leptin, reduced adiposity, and enhanced insulin sensitivity in LL DIO rats fed chow until 10 wk of age. The enhanced ARC leptin receptor binding and reduced adiposity of LL DIO rats persisted after an additional 5 wk on the HE diet. Female LL DIO rats had similar reductions in weight gain on both chow and HE diet vs. normal litter DIO rats. We postulate that LL rearing enhances DIO leptin sensitivity by lowering plasma leptin levels and thereby increasing leptin receptor availability and that this both enhances the ARC-paraventricular nucleus pathway development and protects them from becoming obese.
|Gene expression profiling of human neural progenitor cells following the serum-induced astrocyte differentiation. |
Shinya Obayashi, Hiroko Tabunoki, Seung U Kim, Jun-ichi Satoh, Shinya Obayashi, Hiroko Tabunoki, Seung U Kim, Jun-ichi Satoh, Shinya Obayashi, Hiroko Tabunoki, Seung U Kim, Jun-Ichi Satoh
Cellular and molecular neurobiology 29 423-38 2009
Neural stem cells (NSC) with self-renewal and multipotent properties could provide an ideal cell source for transplantation to treat spinal cord injury, stroke, and neurodegenerative diseases. However, the majority of transplanted NSC and neural progenitor cells (NPC) differentiate into astrocytes in vivo under pathological environments in the central nervous system, which potentially cause reactive gliosis. Because the serum is a potent inducer of astrocyte differentiation of rodent NPC in culture, we studied the effect of the serum on gene expression profile of cultured human NPC to identify the gene signature of astrocyte differentiation of human NPC. Human NPC spheres maintained in the serum-free culture medium were exposed to 10% fetal bovine serum (FBS) for 72 h, and processed for analyzing on a Whole Human Genome Microarray of 41,000 genes, and the microarray data were validated by real-time RT-PCR. The serum elevated the levels of expression of 45 genes, including ID1, ID2, ID3, CTGF, TGFA, METRN, GFAP, CRYAB and CSPG3, whereas it reduced the expression of 23 genes, such as DLL1, DLL3, PDGFRA, SOX4, CSPG4, GAS1 and HES5. Thus, the serum-induced astrocyte differentiation of human NPC is characterized by a counteraction of ID family genes on Delta family genes. Coimmunoprecipitation analysis identified ID1 as a direct binding partner of a proneural basic helix-loop-helix (bHLH) transcription factor MASH1. Luciferase assay indicated that activation of the DLL1 promoter by MASH1 was counteracted by ID1. Bone morphogenetic protein 4 (BMP4) elevated the levels of ID1 and GFAP expression in NPC under the serum-free culture conditions. Because the serum contains BMP4, these results suggest that the serum factor(s), most probably BMP4, induces astrocyte differentiation by upregulating the expression of ID family genes that repress the proneural bHLH protein-mediated Delta expression in human NPC.
|Immunohistochemical markers for quantitative studies of neurons and glia in human neocortex. |
Lise Lyck, Ishar Dalmau, John Chemnitz, Bente Finsen, Henrik Daa Schrøder, Lise Lyck, Ishar Dalmau, John Chemnitz, Bente Finsen, Henrik Daa Schrøder
The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society 56 201-21 2008
Reproducible visualization of neurons and glia in human brain is essential for quantitative studies of the cellular changes in neurological disease. However, immunohistochemistry in human brain specimens is often compromised because of prolonged fixation. To select cell lineage-specific antibodies for quantitative studies of neurons and the major types of glia, we used 29 different antibodies, different epitope retrieval methods, and different detection systems to stain tissue arrays of formalin-fixed human brain. The screening pointed at CD45/leukocyte common antigen (LCA), CD68(KP1), 2',3' cyclic nucleotide phosphatase (CNPase), glial fibrillary acidic protein (GFAP), HLA-DR, Ki67, neuronal nuclei (NeuN), p25alpha-antigen, and S100beta as candidates for future cell counting purposes, because these markers visualized specific neuronal and glial cell bodies. However, significant negative correlation between staining result and formalin fixation was observed by blinded scoring of staining for CD45/LCA, CNPase, GFAP, and NeuN in brain specimens fixed by immersion and stored up to 10 years in 4% formalin solution at room temperature, independent of donor sex and postmortem interval. In contrast, improved preservation of NeuN and CNPase staining, and full preservation of GFAP and CD45/LCA staining in tissue fixed by perfusion and stored for up to 3 years in 0.1% paraformaldehyde solution at 4C, indicated that immunohistochemistry can be performed in well-preserved biobank material.Full Text Article
|Differentiation of human embryonic stem cells to regional specific neural precursors in chemically defined medium conditions. |
Erceg, Slaven, et al.
PLoS ONE, 3: e2122 (2008) 2008
|Mesenchymal stem cells derived from human exocrine pancreas express transcription factors implicated in beta-cell development. |
Reto M Baertschiger, Domenico Bosco, Philippe Morel, Veronique Serre-Beinier, Thierry Berney, Leo H Buhler, Carmen Gonelle-Gispert, Reto M Baertschiger, Domenico Bosco, Philippe Morel, Veronique Serre-Beinier, Thierry Berney, Leo H Buhler, Carmen Gonelle-Gispert
Pancreas 37 75-84 2008
OBJECTIVES: Transplantation of in vitro generated islets or insulin-producing cells represents an attractive option to overcome organ shortage. The aim of this study was to isolate, expand, and characterize cells from human exocrine pancreas and analyze their potential to differentiate into beta cells. METHODS: Fibroblast-like cells growing out of human exocrine tissue were characterized by flow cytometry and by their capacity to differentiate into mesenchymal cell lineages. During cell expansion and after differentiation toward beta cells, expression of transcription factors of endocrine pancreatic progenitors was analyzed by reverse transcription polymerase chain reaction. RESULTS: Cells emerged from 14/18 human pancreatic exocrine fractions and were expanded up to 40 population doublings. These cells displayed surface antigens similar to mesenchymal stem cells from bone marrow. A culture of these cells in adipogenic and chondrogenic differentiation media allowed differentiation into adipocyte- and chondrocyte-like cells. During expansion, cells expressed transcription factors implicated in islet development such as Isl1, Nkx2.2, Nkx6.1, nestin, Ngn3, Pdx1, and NeuroD. Activin A and hepatocyte growth factor induced an expression of insulin, glucagon, and glucokinase. CONCLUSIONS: Proliferating cells with characteristics of mesenchymal stem cells and endocrine progenitors were isolated from exocrine tissue. Under specific conditions, these cells expressed little insulin. Human pancreatic exocrine tissue might thus be a source of endocrine cell progenitors.
|Nestin expression in adult and developing human kidney. |
Eugenio Bertelli, Marì Regoli, Luciano Fonzi, Rossella Occhini, Susanna Mannucci, Leonardo Ermini, Paolo Toti
The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society 55 411-21 2007
Nestin is considered a marker of neurogenic and myogenic precursor cells. Its arrangement is regulated by cyclin-dependent kinase 5 (CDK5), which is expressed in murine podocytes. We investigated nestin expression in human adult and fetal kidney as well as CDK5 presence in adult human podocytes. Confocal microscopy demonstrated that adult glomeruli display nestin immunoreactivity in vimentin-expressing cells with the podocyte morphology and not in cells bearing the endothelial marker CD31. Glomerular nestin-positive cells were CDK5 immunoreactive as well. Western blotting of the intermediate filament-enriched cytoskeletal fraction and coimmunoprecipitation of nestin with anti-CDK5 antibodies confirmed these results. Nestin was also detected in developing glomeruli within immature podocytes and a few other cells. Confocal microscopy of experiments conducted with antibodies against nestin and endothelial markers demonstrated that endothelial cells belonging to capillaries invading the lower cleft of S-shaped bodies and the immature glomeruli were nestin immunoreactive. Similar experiments carried out with antibodies raised against nestin and alpha-smooth muscle actin showed that the first mesangial cells that populate the developing glomeruli expressed nestin. In conclusion, nestin is expressed in the human kidney from the first steps of glomerulogenesis within podocytes, mesangial, and endothelial cells. This expression, restricted to podocytes in mature glomeruli, appears associated with CDK5.
|Functional engraftment of human ES cell-derived dopaminergic neurons enriched by coculture with telomerase-immortalized midbrain astrocytes. |
Roy, Neeta S, et al.
Nat. Med., 12: 1259-68 (2006) 2006
To direct human embryonic stem (HES) cells to a dopaminergic neuronal fate, we cocultured HES cells that were exposed to both sonic hedgehog and fibroblast growth factor 8 with telomerase-immortalized human fetal midbrain astrocytes. These astrocytes substantially potentiated dopaminergic neurogenesis by both WA09 and WA01 HES cells, biasing them to the A9 nigrostriatal phenotype. When transplanted into the neostriata of 6-hydroxydopamine-lesioned parkinsonian rats, the dopaminergic implants yielded a significant, substantial and long-lasting restitution of motor function. However, although rich in donor-derived tyrosine hydroxylase-expressing neurons, the grafts exhibited expanding cores of undifferentiated mitotic neuroepithelial cells, which can be tumorigenic. These results show the utility of recreating the cellular environment of the developing human midbrain while driving dopaminergic neurogenesis from HES cells, and they demonstrate the potential of the resultant cells to mediate substantial functional recovery in a model of Parkinson disease. Yet these data also mandate caution in the clinical application of HES cell-derived grafts, given their potential for phenotypic instability and undifferentiated expansion.
|Nestin immunoreactivity of Purkinje cells in Creutzfeldt-Jakob disease. |
Yuji Mizuno, Eisaku Ohama, Junko Hirato, Yoichi Nakazato, Hitoshi Takahashi, Masamitsu Takatama, Toshiyuki Takeuchi, Koichi Okamoto
Journal of the neurological sciences 246 131-7 2006
Nestin, an intermediate filament protein, is mainly expressed in neural progenitor/stem cells in the central nervous system. Recently, we reported that nestin is expressed in Purkinje cells in patients with Creutzfeldt-Jakob disease (CJD). In this study, we examined a total of 19 CJD cerebella to analyze the intensity and pattern of nestin immunoreactivity of Purkinje cells in different pathological stages of degeneration in the cerebellar cortex. The results showed that the Purkinje cells were immunoreactive with nestin regardless of the severity of degenerative cerebellar cortex. Furthermore, we noted several different types of nestin immunoreactivity, indicated by diffuse and fine, coarse, and inclusion-like immunostainings within Purkinje cell bodies as well as dot-like staining outside of the cell bodies. In contrast, on examination of cerebella from non-CJD patients, 6 of 30 cases showed nestin immunoreactivity to a lesser extent. Thus, nestin-positive Purkinje cells are more common in CJD cerebella than in non-CJD cerebella. Although the mechanism of nestin expression in Purkinje cells is not yet understood, we suggest that such nestin-positive Purkinje cells are being reactivated to survive the cell death.
|Transplantation of cultured neural cells from human fetuses into the brain of rats exposed to acute hypoxia. |
Aleksandrova, M A, et al.
Bull. Exp. Biol. Med., 137: 262-5 (2004) 2004
Neural stem cells of human brain were cultured for a long time and successfully transplanted into the brain of rats exposed to acute hypoxia. Stem and committed cells, neuroblasts, and astrocytes were revealed in transplants by immunohistochemical assay. The transplants and brain tissue were not separated with a glial barrier. Human neuroblasts widely migrated into regions of neuronal degeneration in the host brain.
|Differentiation of human embryonic stem cells to dopaminergic neurons in serum-free suspension culture. |
Thomas C Schulz, Scott A Noggle, Gail M Palmarini, Deb A Weiler, Ian G Lyons, Kate A Pensa, Adrian C B Meedeniya, Bruce P Davidson, Nevin A Lambert, Brian G Condie
Stem cells (Dayton, Ohio) 22 1218-38 2004
The use of human embryonic stem cells (hESCs) as a source of dopaminergic neurons for Parkinson's disease cell therapy will require the development of simple and reliable cell differentiation protocols. The use of cell cocultures, added extracellular signaling factors, or transgenic approaches to drive hESC differentiation could lead to additional regulatory as well as cell production delays for these therapies. Because the neuronal cell lineage seems to require limited or no signaling for its formation, we tested the ability of hESCs to differentiate to form dopamine-producing neurons in a simple serum-free suspension culture system. BG01 and BG03 hESCs were differentiated as suspension aggregates, and neural progenitors and neurons were detectable after 2-4 weeks. Plated neurons responded appropriately to electrophysiological cues. This differentiation was inhibited by early exposure to bone morphogenic protein (BMP)-4, but a pulse of BMP-4 from days 5 to 9 caused induction of peripheral neuronal differentiation. Real-time polymerase chain reaction and whole-mount immunocytochemistry demonstrated the expression of multiple markers of the midbrain dopaminergic phenotype in serum-free differentiations. Neurons expressing tyrosine hydroxylase (TH) were killed by 6-hydroxydopamine (6-OHDA), a neurotoxic catecholamine. Upon plating, these cells released dopamine and other catecholamines in response to K+ depolarization. Surviving TH+ neurons, derived from the cells differentiated in serum-free suspension cultures, were detected 8 weeks after transplantation into 6-OHDA-lesioned rat brains. This work suggests that hESCs can differentiate in simple serum-free suspension cultures to produce the large number of cells required for transplantation studies.
|Early acquisition of typical metabolic features upon differentiation of mouse neural stem cells into astrocytes. |
J F Brunet, L Grollimund, J-Y Chatton, S Lengacher, P J Magistretti, J G Villemure, L Pellerin
Glia 46 8-17 2004
Specific metabolic features, such as glutamate reuptake, have been associated with normal functions of mature astrocytes. In this study, we examined whether these characteristics are acquired together with classical phenotypic markers of differentiated astrocytes. Differentiation of E14 mouse neurospheres into astrocytes was induced by the addition of fetal bovine serum (FBS). Degree of differentiation was assessed by reverse transcription-polymerase chain reaction (RT-PCR) and immunofluorescence for both GFAP and nestin. Neural stem cells expressed nestin but not GFAP, while differentiated astrocytes were immunopositive for GFAP but displayed low levels of nestin expression. A strong increase in the expression of the glutamate transporter GLAST and the monocarboxylate transporter MCT1 accompanied phenotypic changes. In addition, active glutamate transport appeared in differentiated astrocytes, as well as their capacity to increase aerobic glycolysis in response to glutamate. Leukemia inhibitory factor (LIF) and ciliary neurotrophic factor, but not interleukin-6, triggered the expression of phenotypic and morphological characteristics of astrocytes. In addition, exposure to LIF led to the appearance of metabolic features typically associated with astrocytes. Altogether, our results show that acquisition of some specific metabolic features by astrocytes occurs early in their differentiation process and that LIF represents a candidate signal to induce their expression.
|Cancerous stem cells can arise from pediatric brain tumors |
Hemmati, Houman D, et al.
Proc Natl Acad Sci USA, 100:15178-15183 (2003) 2003
|Cryopreservation of human brain tissue allowing timely production of viable adult human brain cells for autologous transplantation. |
Jean François Brunet, Luc Pellerin, Pierre Magistretti, Jean Guy Villemure
Cryobiology 47 179-83 2003
BACKGROUND: Autologous transplantation is an attractive approach to treat some neurological diseases. A major obstacle is the capacity to produce cells for transplantation at the appropriate time. We describe a cryopreservation procedure for adult human brain tissue allowing the generation of cells in vitro. METHODS: Neurological resections were dissected to separate white and grey matter. Fractions were frozen in a specific cryopreservation medium containing a selected serum and stored in liquid nitrogen. Tissue was thawed, cells were mechanically dissociated, expanded in culture and characterized by immunochemistry. RESULTS: Adult human brain tissue cryopreserved for up to two years was successfully used to generate brain cells that could be maintained in culture for up to 100 days. Cells expressed a variety of neuroectodermal markers including GFAP, S100beta, and neurofilament. CONCLUSION: A successful procedure for cryopreservation of adult human brain tissue has been established that might facilitate future autologous transplantation strategies.
|Coexpression of nestin in neural and glial cells in the developing human CNS defined by a human-specific anti-nestin antibody. |
Messam, C A, et al.
Exp. Neurol., 161: 585-96 (2000) 2000
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