References
Items 1 to 12 of 6063 total
- Magnifico A et al. (MAR 2009) Clinical cancer research : an official journal of the American Association for Cancer Research 15 6 2010--21
Tumor-initiating cells of HER2-positive carcinoma cell lines express the highest oncoprotein levels and are sensitive to trastuzumab.
PURPOSE: The existence of tumor-initiating cells in breast cancer has profound implications for cancer therapy. In this study, we investigated the sensitivity of tumor-initiating cells isolated from human epidermal growth factor receptor type 2 (HER2)-overexpressing carcinoma cell lines to trastuzumab, a compound used for the targeted therapy of breast cancer. EXPERIMENTAL DESIGN: Spheres were analyzed by indirect immunofluorescence for HER2 cell surface expression and by real-time PCR for HER2 mRNA expression in the presence or absence of the Notch1 signaling inhibitor (GSI) or Notch1 small interfering RNA. Xenografts of HER2-overexpressing breast tumor cells were treated with trastuzumab or doxorubicin. The sphere-forming efficiency (SFE) and serial transplantability of tumors were assessed. RESULTS: In HER2-overexpressing carcinoma cell lines, cells with tumor-initiating cell properties presented increased HER2 levels compared with the bulk cell population without modification in HER2 gene amplification. HER2 levels were controlled by Notch1 signaling, as shown by the reduction of HER2 cell surface expression and lower SFE following gamma-secretase inhibition or Notch1 specific silencing. We also show that trastuzumab was able to effectively target tumor-initiating cells of HER2-positive carcinoma cell lines, as indicated by the significant decrease in SFE and the loss of serial transplantability, following treatment of HER2-overexpressing xenotransplants. CONCLUSIONS: Here, we provide evidence for the therapeutic efficacy of trastuzumab in debulking and in targeting tumor-initiating cells of HER2-overexpressing tumors. We also propose that Notch signaling regulates HER2 expression, thereby representing a critical survival pathway of tumor-initiating cells. View PublicationCatalog #: Product Name: 01701 ALDEFLUOR™ Assay Buffer 01700 ALDEFLUOR™ Kit 01705 ALDEFLUOR™ DEAB Reagent Catalog #: 01701 Product Name: ALDEFLUOR™ Assay Buffer Catalog #: 01700 Product Name: ALDEFLUOR™ Kit Catalog #: 01705 Product Name: ALDEFLUOR™ DEAB Reagent Miller CL and Eaves CJ (DEC 1997) Proceedings of the National Academy of Sciences of the United States of America 94 25 13648--53Expansion in vitro of adult murine hematopoietic stem cells with transplantable lympho-myeloid reconstituting ability.
Elucidation of mechanisms that regulate hematopoietic stem cell self-renewal and differentiation would be facilitated by the identification of defined culture conditions that allow these cells to be amplified. We now demonstrate a significant net increase (3-fold, P textless 0.001) in vitro of cells that are individually able to permanently and competitively reconstitute the lymphoid and myeloid systems of syngeneic recipient mice when Sca-1(+)lin- adult marrow cells are incubated for 10 days in serum-free medium with interleukin 11, flt3-ligand, and Steel factor. Moreover, the culture-derived repopulating cells continued to expand their numbers in the primary hosts at the same rate seen in recipients of noncultured stem cells. In the expansion cultures, long-term culture-initiating cells increased 7- +/- 2-fold, myeloid colony-forming cells increased 140- +/- 36-fold, and total nucleated cells increased 230- +/- 62-fold. Twenty-seven of 100 cultures initiated with 15 Sca-1(+)lin- marrow cells were found to contain transplantable stem cells 10 days later. This frequency of positive cultures is the same as the frequency of transplantable stem cells in the original input suspension, suggesting that most had undergone at least one self-renewal division in vitro. No expansion of stem cells was seen when Sca-1+TER119- CD34+ day 14.5 fetal liver cells were cultured under the same conditions. These findings set the stage for further investigations of the mechanisms by which cytokine stimulation may elicit different outcomes in mitotically activated hematopoietic stem cells during ontogeny and in the adult. View PublicationCatalog #: Product Name: 09600 StemSpan™ SFEM 02690 StemSpan™ CC100 02696 StemSpan™ Megakaryocyte Expansion Supplement (100X) 02697 StemSpan™ CC110 09300 10% Bovine Serum Albumin in Iscove's MDM 09500 BIT 9500 Serum Substitute Catalog #: 09600 Product Name: StemSpan™ SFEM Catalog #: 02690 Product Name: StemSpan™ CC100 Catalog #: 02696 Product Name: StemSpan™ Megakaryocyte Expansion Supplement (100X) Catalog #: 02697 Product Name: StemSpan™ CC110 Catalog #: 09300 Product Name: 10% Bovine Serum Albumin in Iscove's MDM Catalog #: 09500 Product Name: BIT 9500 Serum Substitute Tomasz M ( 1995) Chemistry & biology 2 9 575--579Mitomycin C: small, fast and deadly (but very selective).
Mitomycin C, an important antitumor drug and antibiotic, has an extraordinary ability to crosslink DNA with high efficiency and absolute specificity for the sequence CpG. Recent results have shown how mitomycin C crosslinks DNA, and why the sequence specificity is so complete. This new understanding may allow the design of agents that mimic mitomycin C's economy of structure and can crosslink other sequences. View PublicationCatalog #: Product Name: 73272 Mitomycin C Catalog #: 73272 Product Name: Mitomycin C Li P et al. (DEC 2008) Cell 135 7 1299--310Germline competent embryonic stem cells derived from rat blastocysts.
Rats have important advantages over mice as an experimental system for physiological and pharmacological investigations. The lack of rat embryonic stem (ES) cells has restricted the availability of transgenic technologies to create genetic models in this species. Here, we show that rat ES cells can be efficiently derived, propagated, and genetically manipulated in the presence of small molecules that specifically inhibit GSK3, MEK, and FGF receptor tyrosine kinases. These rat ES cells express pluripotency markers and retain the capacity to differentiate into derivatives of all three germ layers. Most importantly, they can produce high rates of chimerism when reintroduced into early stage embryos and can transmit through the germline. Establishment of authentic rat ES cells will make possible sophisticated genetic manipulation to create models for the study of human diseases. View PublicationCatalog #: Product Name: 72052 CHIR99021 72182 PD0325901 Catalog #: 72052 Product Name: CHIR99021 Catalog #: 72182 Product Name: PD0325901 Zhang CC and Lodish HF (JUN 2005) Blood 105 11 4314--20Murine hematopoietic stem cells change their surface phenotype during ex vivo expansion.
Ex vivo expansion of hematopoietic stem cells (HSCs) is important for many clinical applications, and knowledge of the surface phenotype of ex vivo-expanded HSCs will be critical to their purification and analysis. Here, we developed a simple culture system for bone marrow (BM) HSCs using low levels of stem cell factor (SCF), thrombopoietin (TPO), insulin-like growth factor 2 (IGF-2), and fibroblast growth factor-1 (FGF-1) in serum-free medium. As measured by competitive repopulation analyses, there was a more than 20-fold increase in numbers of long-term (LT)-HSCs after a 10-day culture of total BM cells. Culture of BM side population" (SP) cells� View PublicationCatalog #: Product Name: 09600 StemSpan™ SFEM 28600 L-Calc™ Software Catalog #: 09600 Product Name: StemSpan™ SFEM Catalog #: 28600 Product Name: L-Calc™ Software Wang YI et al. (JUL 2016) Biotechnology and BioengineeringMicrofluidic blood-brain barrier model provides in vivo-like barrier properties for drug permeability screening
Efficient delivery of therapeutics across the neuroprotective blood-brain barrier (BBB) remains a formidable challenge for central nervous system drug development. High-fidelity in vitro models of the BBB could facilitate effective early screening of drug candidates targeting the brain. In this study, we developed a microfluidic BBB model that is capable of mimicking in vivo BBB characteristics for a prolonged period and allows for reliable in vitro drug permeability studies under recirculating perfusion. We derived brain microvascular endothelial cells (BMECs) from human induced pluripotent stem cells (hiPSCs) and cocultured them with rat primary astrocytes on the two sides of a porous membrane on a pumpless microfluidic platform for up to 10 days. The microfluidic system was designed based on the blood residence time in human brain tissues, allowing for medium recirculation at physiologically relevant perfusion rates with no pumps or external tubing, meanwhile minimizing wall shear stress to test whether shear stress is required for in vivo-like barrier properties in a microfluidic BBB model. This BBB-on-a-chip model achieved significant barrier integrity as evident by continuous tight junction formation and in vivo-like values of trans-endothelial electrical resistance (TEER). The TEER levels peaked above 4000 $$ textperiodcentered cm(2) on day 3 on chip and were sustained above 2000 $$ textperiodcentered cm(2) up to 10 days, which are the highest sustained TEER values reported in a microfluidic model. We evaluated the capacity of our microfluidic BBB model to be used for drug permeability studies using large molecules (FITC-dextrans) and model drugs (caffeine, cimetidine, and doxorubicin). Our analyses demonstrated that the permeability coefficients measured using our model were comparable to in vivo values. Our BBB-on-a-chip model closely mimics physiological BBB barrier functions and will be a valuable tool for screening of drug candidates. The residence time-based design of a microfluidic platform will enable integration with other organ modules to simulate multi-organ interactions on drug response. Biotechnol. Bioeng. 2016;9999: 1-11. textcopyright 2016 Wiley Periodicals, Inc. View PublicationCatalog #: Product Name: 85850 mTeSR™1 Catalog #: 85850 Product Name: mTeSR™1 U. Rajamani et al. (MAY 2018) Cell stem cell 22 5 698--712.e9Super-Obese Patient-Derived iPSC Hypothalamic Neurons Exhibit Obesogenic Signatures and Hormone Responses.
The hypothalamus contains neurons that integrate hunger and satiety endocrine signals from the periphery and are implicated in the pathophysiology of obesity. The limited availability of human hypothalamic neurons hampers our understanding of obesity disease mechanisms. To address this, we generated human induced pluripotent stem cells (hiPSCs) from multiple normal body mass index (BMI; BMI ≤ 25) subjects and super-obese (OBS) donors (BMI ≥ 50) with polygenic coding variants in obesity-associated genes. We developed a method to reliably differentiate hiPSCs into hypothalamic-like neurons (iHTNs) capable of secreting orexigenic and anorexigenic neuropeptides. Transcriptomic profiling revealed that, although iHTNs maintain a fetal identity, they respond appropriately to metabolic hormones ghrelin and leptin. Notably, OBS iHTNs retained disease signatures and phenotypes of high BMI, exhibiting dysregulated respiratory function, ghrelin-leptin signaling, axonal guidance, glutamate receptors, and endoplasmic reticulum (ER) stress pathways. Thus, human iHTNs provide a powerful platform to study obesity and gene-environment interactions. View PublicationCatalog #: Product Name: 07930 CryoStor® CS10 Catalog #: 07930 Product Name: CryoStor® CS10 Lin F et al. (AUG 2015) Hepatology (Baltimore, Md.) 62 2 505--20The camKK2/camKIV relay is an essential regulator of hepatic cancer.
UNLABELLED Hepatic cancer is one of the most lethal cancers worldwide. Here, we report that the expression of Ca(2+) /calmodulin-dependent protein kinase kinase 2 (CaMKK2) is significantly up-regulated in hepatocellular carcinoma (HCC) and negatively correlated with HCC patient survival. The CaMKK2 protein is highly expressed in all eight hepatic cancer cell lines evaluated and is markedly up-regulated relative to normal primary hepatocytes. Loss of CaMKK2 function is sufficient to inhibit liver cancer cell growth, and the growth defect resulting from loss of CaMKK2 can be rescued by ectopic expression of wild-type CaMKK2 but not by kinase-inactive mutants. Cellular ablation of CaMKK2 using RNA interference yields a gene signature that correlates with improvement in HCC patient survival, and ablation or pharmacological inhibition of CaMKK2 with STO-609 impairs tumorigenicity of liver cancer cells in vivo. Moreover, CaMKK2 expression is up-regulated in a time-dependent manner in a carcinogen-induced HCC mouse model, and STO-609 treatment regresses hepatic tumor burden in this model. Mechanistically, CaMKK2 signals through Ca(2+) /calmodulin-dependent protein kinase 4 (CaMKIV) to control liver cancer cell growth. Further analysis revealed that CaMKK2 serves as a scaffold to assemble CaMKIV with key components of the mammalian target of rapamycin/ribosomal protein S6 kinase, 70 kDa, pathway and thereby stimulate protein synthesis through protein phosphorylation. CONCLUSION The CaMKK2/CaMKIV relay is an upstream regulator of the oncogenic mammalian target of rapamycin/ribosomal protein S6 kinase, 70 kDa, pathway, and the importance of this CaMKK2/CaMKIV axis in HCC growth is confirmed by the potent growth inhibitory effects of genetically or pharmacologically decreasing CaMKK2 activity; collectively, these findings suggest that CaMKK2 and CaMKIV may represent potential targets for hepatic cancer. View PublicationCatalog #: Product Name: 73862 STO-609 Catalog #: 73862 Product Name: STO-609 D. K. Mahajan and S. N. London (dec 1997) Fertility and sterility 68 6 967--76Mifepristone (RU486): a review.
OBJECTIVE To review the literature concerning the mechanism of action and pharmacodynamics of mifepristone (RU486), potential new uses of RU486, and its current use not only as an abortifacient but also as therapy for endometriosis, leiomyoma, breast cancer, and meningioma. DATA IDENTIFICATION AND SELECTION Studies that relate to RU486 were identified through a MEDLINE search. CONCLUSION(S) RU486 is an 11 beta-dimethyl-amino-phenyl derivative of norethindrone with a high affinity for P and glucocorticoid receptors. The receptor binding is not followed by transcription of P-dependent genes. Mifepristone effectively blocks P receptors in the placenta, resulting in the termination of pregnancy. In addition, it has been used in the treatment of leiomyomata, endometriosis, advanced breast cancer, and meningioma. It is a powerful tool to study the molecular action of P and in the future may be used as an estrogen-free contraceptive. Through an online search of MEDLINE, the authors reviewed the literature on the development of mifepristone (RU-486); RU-486's mechanism of action, pharmacodynamics, and distribution; the physiologic action of RU-486; potential new uses for RU-486; and its current use as both an abortifacient and therapy for endometriosis, leiomyoma, breast cancer, and meningioma. RU-486 is an 11beta-dimethyl-amino-phenyl derivative of norethindrone with a high affinity for P and glucocorticoid receptors. Receptor binding is not followed by the transcription of P-dependent genes. RU-486 effectively blocks P receptors in the placenta, resulting in the termination of pregnancy. It has also been used to treat leiomyomata, endometriosis, advanced breast cancer, and meningioma. The following therapeutic uses of RU-486 are discussed: the termination of early pregnancy, treatment with RU-486 in combination with prostaglandins, the termination of second-trimester pregnancy, cervical ripening, labor induction, postcoital contraception, uterine leiomyomata, endometriosis, breast cancer, and meningioma. View PublicationCatalog #: Product Name: 100-0564 Mifepristone Catalog #: 100-0564 Product Name: Mifepristone de Meester C et al. ( 2014) Cardiovascular research 101 1 20--29Role of AMP-activated protein kinase in regulating hypoxic survival and proliferation of mesenchymal stem cells.
AIMS: Mesenchymal stem cells (MSCs) are widely used for cell therapy, particularly for the treatment of ischaemic heart disease. Mechanisms underlying control of their metabolism and proliferation capacity, critical elements for their survival and differentiation, have not been fully characterized. AMP-activated protein kinase (AMPK) is a key regulator known to metabolically protect cardiomyocytes against ischaemic injuries and, more generally, to inhibit cell proliferation. We hypothesized that AMPK plays a role in control of MSC metabolism and proliferation. METHODS AND RESULTS: MSCs isolated from murine bone marrow exclusively expressed the AMPKα1 catalytic subunit. In contrast to cardiomyocytes, a chronic exposure of MSCs to hypoxia failed to induce cell death despite the absence of AMPK activation. This hypoxic tolerance was the consequence of a preference of MSC towards glycolytic metabolism independently of oxygen availability and AMPK signalling. On the other hand, A-769662, a well-characterized AMPK activator, was able to induce a robust and sustained AMPK activation. We showed that A-769662-induced AMPK activation inhibited MSC proliferation. Proliferation was not arrested in MSCs derived from AMPKα1-knockout mice, providing genetic evidence that AMPK is essential for this process. Among AMPK downstream targets proposed to regulate cell proliferation, we showed that neither the p70 ribosomal S6 protein kinase/eukaryotic elongation factor 2-dependent protein synthesis pathway nor p21 was involved, whereas p27 expression was increased by A-769662. Silencing p27 expression partially prevented the A-769662-dependent inhibition of MSC proliferation. CONCLUSION: MSCs resist hypoxia independently of AMPK whereas chronic AMPK activation inhibits MSC proliferation, p27 being involved in this regulation. View PublicationCatalog #: Product Name: 72922 A769662 Catalog #: 72922 Product Name: A769662 L. Chicaybam et al. ( 2016) Frontiers in bioengineering and biotechnology 4 99An Efficient Electroporation Protocol for the Genetic Modification of Mammalian Cells.
Genetic modification of cell lines and primary cells is an expensive and cumbersome approach, often involving the use of viral vectors. Electroporation using square-wave generating devices, like Lonza's Nucleofector, is a widely used option, but the costs associated with the acquisition of electroporation kits and the transient transgene expression might hamper the utility of this methodology. In the present work, we show that our in-house developed buffers, termed Chicabuffers, can be efficiently used to electroporate cell lines and primary cells from murine and human origin. Using the Nucleofector II device, we electroporated 14 different cell lines and also primary cells, like mesenchymal stem cells and cord blood CD34+, providing optimized protocols for each of them. Moreover, when combined with sleeping beauty-based transposon system, long-term transgene expression could be achieved in all types of cells tested. Transgene expression was stable and did not interfere with CD34+ differentiation to committed progenitors. We also show that these buffers can be used in CRISPR-mediated editing of PDCD1 gene locus in 293T and human peripheral blood mononuclear cells. The optimized protocols reported in this study provide a suitable and cost-effective platform for the genetic modification of cells, facilitating the widespread adoption of this technology. View PublicationCatalog #: Product Name: 04034 MethoCult™ H4034 Optimum 22000 STEMvision™ Catalog #: 04034 Product Name: MethoCult™ H4034 Optimum Catalog #: 22000 Product Name: STEMvision™ Azari H et al. (JAN 2011) Journal of visualized experiments : JoVE 49Neural-colony forming cell assay: an assay to discriminate bona fide neural stem cells from neural progenitor cells.
The neurosphere assay (NSA) is one of the most frequently used methods to isolate, expand and also calculate the frequency of neural stem cells (NSCs). Furthermore, this serum-free culture system has also been employed to expand stem cells and determine their frequency from a variety of tumors and normal tissues. It has been shown recently that a one-to-one relationship does not exist between neurosphere formation and NSCs. This suggests that the NSA as currently applied, overestimates the frequency of NSCs in a mixed population of neural precursor cells isolated from both the embryonic and adult mammalian brain. This video practically demonstrates a novel collagen based semi- solid assay, the neural-colony forming cell assay (N-CFCA), which has the ability to discriminate stem from progenitor cells based on their long-term proliferative potential, and thus provides a method to enumerate NSC frequency. In the N-CFCA, colonies ≥2 mm in diameter are derived from cells that meet all the functional criteria of a NSC, while colonies textless 2mm are derived from progenitors. The N-CFCA procedure can be used for cells prepared from different sources including primary and cultured adult or embryonic mouse CNS cells. Here we use cells prepared from passage one neurospheres generated from embryonic day 14 mice brain to perform N-CFCA. The cultures are replenished with proliferation medium every seven days for three weeks to allow the plated cells to exhibit their full proliferative potential and then the frequency of neural progenitor and bona fide neural stem cells is calculated respectively by counting the number of colonies that are textless 2mm and the ones that are ≥2mm in reference to the number of cells that were initially plated. View PublicationItems 1 to 12 of 6063 total
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