COA 533001
- Document Type:
- Certificate of Analysis
- Product Catalog Number:
- 533001
5.33001
17 Results
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The role of histone acetylation in regulating early gene expression patterns during early embryonic stem cell differentiation.
- We have examined the role of histone acetylation in the very earliest steps of differentiation of mouse embryonic stem cells in response to withdrawal of leukemia inhibitory factor (LIF) as a differentiation signal. The cells undergo dramatic changes in morphology and an ordered program of gene expression changes representing differentiation to all three germ layers over the first 3-5 days of LIF withdrawal. We observed a global increase in acetylation on histone H4 and to a lesser extent on histone H3 over this time period. Treatment of the cells with trichostatin A (TSA), a histone deacetylase inhibitor, induced changes in morphology, gene expression, and histone acetylation that mimicked differentiation induced by withdrawal of LIF. We examined localized histone acetylation in the regulatory regions of genes that were transcriptionally either active in undifferentiated cells, induced during differentiation, or inactive under all treatments. There was striking concordance in the histone acetylation patterns of specific genes induced by both TSA and LIF withdrawal. Increased histone acetylation in local regions correlated best with induction of gene expression. Finally, TSA treatment did not support the maintenance or progression of differentiation. Upon removal of TSA, the cells reverted to the undifferentiated phenotype. We concluded that increased histone acetylation at specific genes played a role in their expression, but additional events are required for maintenance of differentiated gene expression and loss of the pluripotent state.
- Document Type:
- Reference
- Product Catalog Number:
- Multiple
- Product Catalog Name:
- Multiple
-
Activity of PXD101, a histone deacetylase inhibitor, in preclinical ovarian cancer studies.
- Histone deacetylase inhibitors represent a promising new class of anticancer agents. In the current investigation, we examined the activity of PXD101, a potent histone deacetylase inhibitor, used alone or in combination with clinically relevant chemotherapeutics (docetaxel, paclitaxel, and carboplatin), in preclinical in vitro and in vivo models of ovarian cancer. In vitro activity was examined in ovarian cancer and multidrug-resistant cell lines grown in monolayer culture, and in primary clinical ovarian cancer specimens grown in three-dimensional organoid culture. PXD101 was found to inhibit in vitro cancer cell growth at sub- to low micromolar IC(50) potency, exhibited synergistic activity when used in combination with relevant chemotherapeutics, and effectively inhibited the growth of multidrug-resistant cells. In vivo, PXD101 displayed single-agent antitumor activity on human A2780 ovarian cancer s.c. xenografts which was enhanced via combination therapy with carboplatin. In support of these findings, PXD101 was shown to increase the acetylation of alpha-tubulin induced by docetaxel and the phosphorylation of H2AX induced by carboplatin. Taken together, these results support the clinical evaluation of PXD101 used alone or in combination therapy for the treatment of ovarian cancer.
- Document Type:
- Reference
- Product Catalog Number:
- 3301
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Inhomogeneous disappearance of myofilament-related cytoskeletal proteins in stunned myocardium of guinea pig.
- The decrease in Ca2+ responsiveness of myofilaments in stunned myocardium implies that there may be structural changes in proteins composing the contractile machinery. To elucidate the lesion in stunned myocardium, isolated guinea pig hearts were subjected to global ischemia at 37 degrees C and reperfused. SDS-PAGE revealed that the contents of desmin, alpha-actinin, and spectrin decreased in the myofibrillar fraction isolated from hearts reperfused after 60-minute ischemia compared with nonischemic control hearts. To examine the change of cytoskeletal proteins in stunned myocardium, immunohistochemical studies with antibodies against these proteins were performed after 15 minutes of ischemia. In stunned myocardium, the staining was largely intact, but there were some lesions where desmin was not stained and alpha-actinin and spectrin were only weakly identified. The percentage of normally stained areas in the myocardium (percent stained area), quantified by image processing, was significantly lower in stunned myocardium (79.6 +/- 3.6%, mean +/- SEM) than in nonischemic control myocardium (96.5 +/- 0.7%). Percent recovery of developed pressure significantly correlated with percent stained area (r = .82, P < .001). In hearts subjected to 15-minute ischemia but not reperfused, or in hearts reperfused with Ca(2+)-free solution after 15-minute ischemia, staining by the antibodies remained intact, suggesting that the change of the cytoskeletal proteins is mediated by Ca2+ overload during reperfusion. In hearts treated with the protease inhibitor leupeptin (50 mumol/L) or calpain inhibitor I (100 mumol/L), both developed pressure and staining were well preserved. These results indicate that contractile dysfunction in stunned myocardium has a strong correlation with the disappearance of cytoskeletal proteins that may be mediated by a Ca(2+)-dependent intracellular protease activated during reperfusion. The disruption of cytoskeletal proteins is a possible mechanism for stunning, although it may be a secondary effect of protease activation.
- Document Type:
- Reference
- Product Catalog Number:
- 3301
-
Nanostructured ordering of fluorescent markers and single proteins on substrates.
- Highly ordered hexagonal nanopatterns of gold clusters on glass substrates were used as anchoring points for the specific attachment of fluorescence dyes and proteins labeled with fluorescence dyes. Thiol- or disulfide-containing linker molecules were used for the binding to the gold dots. In order to ensure specific binding on the gold dots only, the surface area in between the dots was protected against unspecific adsorption. For the attachment of polar low-molecular-weight fluorescence dyes, an octadecyltrichlorosilane self-assembled monolayer was prepared on the surface in between the gold dots, whereas a layer prepared from star-shaped poly(ethylene oxide-stat-propylene oxide) prepolymers was used to prevent unspecific adsorption of proteins between the gold dots. Fluorescence microscopy proved the specific binding of the dyes as well as of the proteins. Scanning force microscopy studies show that each gold dot is only capable of binding one protein at a time.
- Document Type:
- Reference
- Product Catalog Number:
- 3301