References
Items 145 to 156 of 7892 total
- Banerjee D and Liefshitz A ( 2001) Anticancer research 21 6A 3941--7
Potential of the proteasomal inhibitor MG-132 as an anticancer agent, alone and in combination.
Proteasomal activity is required for normal cellular functions including cell division, where entry and exit from mitosis is strictly regulated by cyclins and cyclin-dependent kinases which are among the important substrates of the proteasomal degradative machinery. Inhibitors of proteasomal activity have been shown to be effective inducers of apoptosis in tumor cells and may be useful as anticancer agents, either alone or in combination with other drugs. We have examined the effect of MG-132, a dipeptide proteasomal inhibitor, on various human cancer cell lines. We have also examined the effect of MG-132 on normal CD34+ enriched primary human peripheral blood stem cells. Our results indicate that MG-312 is a potent anticancer agent with cytotoxic effects on a variety of human cancer cell lines irrespective of their p53 status. MG-132 was found to be more effective in combination with drugs such as doxorubicin and etoposide that act in the S/G2-phase of the cell cycle via a mechanism that involves stabilization of cyclin B1 and increased expression of Bax. Further, MG-132 inhibits CFU-GM colony formation of the CD34+ enriched PBSC population and this inhibition correlates with release of cyt C into the cytosol.Catalog #: Product Name: 73262 (S)-MG132 Catalog #: 73262 Product Name: (S)-MG132 Gudjonsson T et al. (MAR 2002) Genes & development 16 6 693--706Isolation, immortalization, and characterization of a human breast epithelial cell line with stem cell properties.
The epithelial compartment of the human breast comprises two distinct lineages: the luminal epithelial and the myoepithelial lineage. We have shown previously that a subset of the luminal epithelial cells could convert to myoepithelial cells in culture signifying the possible existence of a progenitor cell. We therefore set out to identify and isolate the putative precursor in the luminal epithelial compartment. Using cell surface markers and immunomagnetic sorting, we isolated two luminal epithelial cell populations from primary cultures of reduction mammoplasties. The major population coexpresses sialomucin (MUC(+)) and epithelial-specific antigen (ESA(+)) whereas the minor population has a suprabasal position and expresses epithelial specific antigen but no sialomucin (MUC(-)/ESA(+)). Two cell lines were further established by transduction of the E6/E7 genes from human papilloma virus type 16. Both cell lines maintained a luminal epithelial phenotype as evidenced by expression of the tight junction proteins, claudin-1 and occludin, and by generation of a high transepithelial electrical resistance on semipermeable filters. Whereas in clonal cultures, the MUC(+)/ESA(+) epithelial cell line was luminal epithelial restricted in its differentiation repertoire, the suprabasal-derived MUC(-)/ESA(+) epithelial cell line was able to generate itself as well as MUC(+)/ESA(+) epithelial cells and Thy-1(+)/alpha-smooth muscle actin(+) (ASMA(+)) myoepithelial cells. The MUC(-)/ESA(+) epithelial cell line further differed from the MUC(+)/ESA(+) epithelial cell line by the expression of keratin K19, a feature of a subpopulation of epithelial cells in terminal duct lobular units in vivo. Within a reconstituted basement membrane, the MUC(+)/ESA(+) epithelial cell line formed acinus-like spheres. In contrast, the MUC(-)/ESA(+) epithelial cell line formed elaborate branching structures resembling uncultured terminal duct lobular units both by morphology and marker expression. Similar structures were obtained by inoculating the extracellular matrix-embedded cells subcutaneously in nude mice. Thus, MUC(-)/ESA(+) epithelial cells within the luminal epithelial lineage may function as precursor cells of terminal duct lobular units in the human breast.Dobo I et al. (JAN 2001) The hematology journal : the official journal of the European Haematology Association / EHA 2 6 396--403Comparison of four serum-free, cytokine-free media for analysis of endogenous erythroid colony growth in polycythemia vera and essential thrombocythemia.
INTRODUCTION: The assay of endogenous erythroid colony formation (EEC), a characteristic of polycythemia vera and essential thrombocythemia, is not standardized. In this multicentric study, we tested four semisolid, serum-free, cytokine-free media based on either methylcellulose (M1, M2) or collagen (C1, C2) commercialized for the EEC assay. MATERIALS AND METHODS: Bone marrow mononuclear cells (BMMC) from 73 individuals (62 patients with either polycythemia vera (26), essential thrombocythemia (19), secondary polyglobuly (17) or chronic myeloid leukemia (2) and 11 healthy donors) were grown in parallel in the four media without, or with 0.01 U/ml erythropoietin (EPo). RESULTS: In all four media EEC formation was specific, as it was not observed in cultures of patients with secondary polyglobuly or chronic myeloid leukemia, nor of healthy donors. Analysis of fresh or MGG-stained collagen gel cultures allowed detection of EEC formation significantly more frequently than methylcellulose-based media; addition of 0.01 U/ml of EPo had little or no effect on EEC formation. Collagen-based medium C1 gave better results than the other media tested: the 'C1' EEC assay was positive for 68.2% of polycythemia vera cultures with significantly higher median EEC numbers (6.5/10(5) BMMC for patients with one major criteria of polycythemia vera and 19 and 21/10(5) BMMC for patients with two or three major criteria, respectively). Medium C1 was also better for essential thrombocythemia cultures with 47.4% of positive results but with a low median EEC number (6.7/10(5) BMMC). When associated with the ELISA dosage of serum EPo, the 'C1' EEC assay allowed confirmation or elimination of the diagnosis of polycythemia vera for 91% (20/22) of polyglobulic patients. CONCLUSION: We propose that serum-free collagen-based culture systems be considered to standardize the EEC assay, now part of the new criteria of polycythemia vera.Catalog #: Product Name: 04970 MegaCultâ„¢-C Complete Kit Without Cytokines 04971 MegaCultâ„¢-C Complete Kit with Cytokines 04974 MegaCultâ„¢-C Collagen and Medium with Lipids 04850 MegaCultâ„¢-C Medium with Lipids 04962 MegaCultâ„¢-C Staining Kit for CFU-Mk 04900 MegaCultâ„¢-C Medium Without Cytokines 04901 MegaCultâ„¢-C Medium with Cytokines 04960 MegaCultâ„¢-C Collagen and Medium Without Cytokines 04961 MegaCultâ„¢-C Collagen and Medium with Cytokines Catalog #: 04970 Product Name: MegaCultâ„¢-C Complete Kit Without Cytokines Catalog #: 04971 Product Name: MegaCultâ„¢-C Complete Kit with Cytokines Catalog #: 04974 Product Name: MegaCultâ„¢-C Collagen and Medium with Lipids Catalog #: 04850 Product Name: MegaCultâ„¢-C Medium with Lipids Catalog #: 04962 Product Name: MegaCultâ„¢-C Staining Kit for CFU-Mk Catalog #: 04900 Product Name: MegaCultâ„¢-C Medium Without Cytokines Catalog #: 04901 Product Name: MegaCultâ„¢-C Medium with Cytokines Catalog #: 04960 Product Name: MegaCultâ„¢-C Collagen and Medium Without Cytokines Catalog #: 04961 Product Name: MegaCultâ„¢-C Collagen and Medium with Cytokines Sata M et al. (APR 2002) Nature medicine 8 4 403--9Hematopoietic stem cells differentiate into vascular cells that participate in the pathogenesis of atherosclerosis.
Excessive accumulation of smooth-muscle cells (SMCs) has a key role in the pathogenesis of vascular diseases. It has been assumed that SMCs derived from the outer medial layer migrate, proliferate and synthesize extracellular matrix components on the luminal side of the vessel. Although much effort has been devoted to targeting migration and proliferation of medial SMCs, there is no effective therapy that prevents occlusive vascular remodeling. We show here that in models of post-angioplasty restenosis, graft vasculopathy and hyperlipidemia-induced atherosclerosis, bone-marrow cells give rise to most of the SMCs that contribute to arterial remodeling. Notably, purified hematopoietic stem cells differentiate into SMCs in vitro and in vivo. Our findings indicate that somatic stem cells contribute to pathological remodeling of remote organs, and may provide the basis for the development of new therapeutic strategies for vascular diseases through targeting mobilization, homing, differentiation and proliferation of bone marrow-derived vascular progenitor cells. View PublicationCatalog #: Product Name: 03434 MethoCultâ„¢ GF M3434 Catalog #: 03434 Product Name: MethoCultâ„¢ GF M3434 Samper E et al. (APR 2002) Blood 99 8 2767--75Long-term repopulating ability of telomerase-deficient murine hematopoietic stem cells.
Telomere length must be tightly regulated in highly proliferative tissues, such as the lymphohematopoietic system. Under steady-state conditions, the levels and functionality of hematopoietic-committed or multipotent progenitors were not affected in late-generation telomerase-deficient mice (mTerc(-/-)) with critically short telomeres. Evaluation of self-renewal potential of mTerc(-/-) day-12 spleen colony-forming units demonstrated no alteration as compared with wildtype progenitors. However, the replating ability of mTerc(-/-) granulocyte-macrophage CFUs (CFU-GMs) was greatly reduced as compared with wildtype CFU-GMs, indicating a diminished capacity of late-generation mTerc(-/-) committed progenitors when forced to proliferate. Long-term bone marrow cultures of mTerc(-/-) bone marrow (BM) cells show a reduction in proliferative capacity; this defect can be mainly attributed to the hematopoietic, not to the stromal, mTerc(-/-) cells. In serial and competitive transplantations, mTerc(-/-) BM stem cells show reduced long-term repopulating capacity, concomitant with an increase in genetic instability compared with wildtype cells. Nevertheless, in competitive transplantations late-generation mTerc(-/-) precursors can occasionally overcome this proliferative impairment and reconstitute irradiated recipients. In summary, our results demonstrate that late-generation mTerc(-/-) BM cells with short telomeres, although exhibiting reduced proliferation ability and reduced long-term repopulating capacity, can still reconstitute myeloablated animals maintaining stem cell function.Dang SM et al. (MAY 2002) Biotechnology and bioengineering 78 4 442--53Efficiency of embryoid body formation and hematopoietic development from embryonic stem cells in different culture systems.
Embryonic stem (ES) cells have tremendous potential as a cell source for cell-based therapies. Realization of that potential will depend on our ability to understand and manipulate the factors that influence cell fate decisions and to develop scalable methods of cell production. We compared four standard ES cell differentiation culture systems by measuring aspects of embryoid body (EB) formation efficiency and cell proliferation, and by tracking development of a specific differentiated tissue type-blood-using functional (colony-forming cell) and phenotypic (Flk-1 and CD34 expression) assays. We report that individual murine ES cells form EBs with an efficiency of 42 +/- 9%, but this value is rarely obtained because of EB aggregation-a process whereby two or more individual ES cells or EBs fuse to form a single, larger cell aggregate. Regardless of whether EBs were generated from a single ES cell in methylcellulose or liquid suspension culture, or aggregates of ES cells in hanging drop culture, they grew to a similar maximum cell number of 28,000 +/- 9,000 cells per EB. Among the three methods for EB generation in suspension culture there were no differences in the kinetics or frequency of hematopoietic development. Thus, initiating EBs with a single ES cell and preventing EB aggregation should allow for maximum yield of differentiated cells in the EB system. EB differentiation cultures were also compared to attached differentiation culture using the same outputs. Attached colonies were not similarly limited in cell number; however, hematopoietic development in attached culture was impaired. The percentage of early Flk-1 and CD34 expressing cells was dramatically lower than in EBs cultured in suspension, whereas hematopoietic colony formation was almost completely inhibited. These results provide a foundation for development of efficient, scalable bioprocesses for ES cell differentiation, and inform novel methods for the production of hematopoietic tissues.Ramalho AC et al. (APR 2002) European cytokine network 13 1 39--45Estradiol and raloxifene decrease the formation of multinucleate cells in human bone marrow cultures.
Estrogen (E2) deficiency is responsible for increased bone turnover in the postmenopausal period, and it can be prevented by estrogen replacement therapy. The way estrogen acts on bone cells is not fully understood. Human bone marrow cell cultures may be a reliable model for studying the action of steroids on osteoclastogenesis in vitro. We examine the effects of estradiol and Raloxifene, a selective estrogen receptor modulator, on human primary bone marrow cells cultured for 15 days. 17beta-estradiol and Raloxifene significantly decreased the number of tartrate-resistant acid phosphatase multinucleate cells from osteoclast precursors on day 15. Estrogen receptor alpha (ER-alpha) mRNA was present in bone marrow mononuclear cells cultured for 5 days, but there was no estrogen receptor beta (ER-beta) mRNA, suggesting that this effect was mediated by ER-alpha. 15-day cultures no longer contained ER-alpha mRNA, suggesting that estrogen acts on early events of osteoclast differentiation. Finally, 10-8 M 17beta-estradiol has no effect on the release of IL-6 and IL-6-sr into the medium of marrow mononuclear cells cultured for 5 or 15 days. Osteoclast apoptosis was not affected by estradiol or Raloxifene after 15 days of culture under our conditions. In conclusion, we have shown that both estradiol and Raloxifene inhibit osteoclast differentiation in human bone marrow mononuclear cultures. The biological effect that can mimic in vivo differentiation could be mediated through ER-alpha.Fontana JA and Rishi AK (APR 2002) Leukemia 16 4 463--72Classical and novel retinoids: their targets in cancer therapy.
Retinoids are important mediators of cellular growth and differentiation. Retinoids modulate the growth of both normal and malignant cells through their binding to retinoid nuclear receptors and their subsequent activation. While retinoids have demonstrated therapeutic efficacy in the treatment of acute promyelocytic leukemia, their spectrum of activity remains limited. Other agents such as histone deacetylase inhibitors may significantly increase retinoid activity in a number of malignant cell types. The novel retinoids N-(4-hydroxyphenyl) retinamide (4-HPR) and 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalene carboxylic acid (CD437; AHPN) induce apoptosis in a wide variety of malignant cells. Their mechanism(s) of action remain unclear, although a number of potential targets have been identified. Whether the retinoid receptors are involved in 4-HPR and CD473/AHPN mediated apoptosis remains unclear. Both 4-HPR and CD437/AHPN display significant potential as therapeutic agents in the treatment of a number of premalignant and malignant conditions.Catalog #: Product Name: 72722 CD437 Catalog #: 72722 Product Name: CD437 Marchetti S et al. (MAY 2002) Journal of cell science 115 Pt 10 2075--85Endothelial cells genetically selected from differentiating mouse embryonic stem cells incorporate at sites of neovascularization in vivo.
Large scale purification of endothelial cells is of great interest as it could improve tissue transplantation, reperfusion of ischemic tissues and treatment of pathologies in which an endothelial cell dysfunction exists. In this study, we describe a novel genetic approach that selects for endothelial cells from differentiating embryonic stem (ES) cells. Our strategy is based on the establishment of ES-cell clones that carry an integrated puromycin resistance gene under the control of a vascular endothelium-specific promoter, tie-1. Using EGFP as a reporter gene, we first confirmed the endothelial specificity of the tie-1 promoter in the embryoid body model and in cells differentiated in 2D cultures. Subsequently, tie-1-EGFP ES cells were used as recipients for the tie-1-driven puror transgene. The resulting stable clones were expanded and differentiated for seven days in the presence of VEGF before puromycin selection. As expected, puromycin-resistant cells were positive for EGFP and also expressed several endothelial markers, including CD31, CD34, VEGFR-1, VEGFR-2, Tie-1, VE-cadherin and ICAM-2. Release from the puromycin selection resulted in the appearance of alpha-smooth muscle actin-positive cells. Such cells became more numerous when the population was cultured on laminin-1 or in the presence of TGF-beta1, two known inducers of smooth muscle cell differentiation. The hypothesis that endothelial cells or their progenitors may differentiate towards a smooth muscle cell phenotype was further supported by the presence of cells expressing both CD31 and alpha-smooth muscle actin markers. Finally, we show that purified endothelial cells can incorporate into the neovasculature of transplanted tumors in nude mice. Taken together, these results suggest that application of endothelial lineage selection to differentiating ES cells may become a useful approach for future pro-angiogenic and endothelial cell replacement therapies.Dahl C et al. (APR 2002) Journal of immunological methods 262 1-2 137--43The establishment of a combined serum-free and serum-supplemented culture method of obtaining functional cord blood-derived human mast cells.
BACKGROUND: Serum-free cultures supplemented with stem cell factor (SCF) and IL-6 is reported to support the extensive growth of less functional human cord blood-derived mast cells. OBJECTIVE: To obtain more functional mast cells from cord blood, we developed a culture system combining a serum-free condition for 0-8 weeks of culture, and followed by a serum-supplemented culture condition and examined the function of the cells compared to the cells cultured continuously in serum-free condition. METHODS: Human cord blood progenitors were purified with anti-CD133 antibody. They were cultured in a serum-free medium StemSpan supplemented with SCF at 100 ng/ml and IL-6 at 50 ng/ml for 8 weeks. Then, an aliquot of the cultured cells were cultured in the above condition but further supplemented with 10% fetal calf serum (FCS). RESULTS: The addition of FCS after 8 weeks of culture significantly increased the amount of histamine per mast cell (3.8 pg/cell) when compared to the serum-free condition (0.7 pg/cell). The cells cultured with FCS after 8 weeks expressed more FcvarepsilonRI alpha and released textgreater30% of the histamine content upon anti-IgE stimulation than those cultured without serum. CONCLUSION: It is uncertain why FCS enhanced the functional maturation of mast cells when added after week 8 of culture but suppressed mast cell development when added at day 0 of culture. Yet, the present method combining a serum-free culture system with a serum-supplemented culture system seems to be beneficial for most of the laboratories to obtain functional human mast cells.Catalog #: Product Name: 09600 StemSpanâ„¢ SFEM Catalog #: 09600 Product Name: StemSpanâ„¢ SFEM van der Kolk DM et al. (MAY 2002) Blood 99 10 3763--70Expression and activity of breast cancer resistance protein (BCRP) in de novo and relapsed acute myeloid leukemia.
Overexpression of the breast cancer resistance protein (BCRP) efflux pump in human cancer cell lines results in resistance to a variety of cytostatic agents. The aim of this study was to analyze BCRP protein expression and activity in acute myeloid leukemia (AML) samples and to determine whether it is up-regulated due to clonal selection at relapse/refractory disease. BCRP protein expression was measured flow cytometrically with the monoclonal antibodies BXP-34 and BXP-21 in 20 paired samples of de novo and relapsed/refractory AML. BXP-34/immunoglobulin G1 ratios were observed of 1.6 +/- 0.5 (mean +/- SD, range 0.8-2.7) and BXP-21/immunoglobulin G2a ratios of 4.9 +/- 3.0 (range 1.1-14.5) in the patient samples versus 9.8 +/- 6.8 and 6.5 +/- 2.4, respectively, in the MCF-7 cell line. BCRP activity was determined flow cytometrically by measuring mitoxantrone accumulation in absence and presence of the inhibitor fumitremorgin C. Mitoxantrone accumulation, expressed as mean fluorescence intensity (MFI), varied between 44 and 761 MFI (227 +/- 146 MFI) and correlated inversely with BCRP expression (r = -0.58, P textless.001). Addition of fumitremorgin C showed a small increase in mitoxantrone accumulation (11 +/- 29 MFI, n = 40) apart from the effect of PSC833 and MK-571. No consistent up-regulation of BCRP expression or activity was observed at relapse/refractory disease; some cases showed an increase and other cases a decrease at relapse. Relatively high BCRP expression correlated with immature immunophenotype, as determined by expression of the surface marker CD34 (r = 0.54, P =.001). In conclusion, this study shows that BCRP protein is expressed at low but variable levels in AML, especially in immature CD34(+) cells. BCRP was not consistently up-regulated in relapsed/refractory AML.N. L. Urizar et al. (may 2002) Science (New York, N.Y.) 296 5573 1703--6A natural product that lowers cholesterol as an antagonist ligand for FXR.
Extracts of the resin of the guggul tree (Commiphora mukul) lower LDL (low-density lipoprotein) cholesterol levels in humans. The plant sterol guggulsterone [4,17(20)-pregnadiene-3,16-dione] is the active agent in this extract. We show that guggulsterone is a highly efficacious antagonist of the farnesoid X receptor (FXR), a nuclear hormone receptor that is activated by bile acids. Guggulsterone treatment decreases hepatic cholesterol in wild-type mice fed a high-cholesterol diet but is not effective in FXR-null mice. Thus, we propose that inhibition of FXR activation is the basis for the cholesterol-lowering activity of guggulsterone. Other natural products with specific biologic effects may modulate the activity of FXR or other relatively promiscuous nuclear hormone receptors.Catalog #: Product Name: 100-0252 (Z)-Guggulsterone Catalog #: 100-0252 Product Name: (Z)-Guggulsterone Items 145 to 156 of 7892 total
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