|Application||The Scepter Cell Counter delivers rapid cell counts in a handheld easy to use format. FREE PACK OF 60μm Sensors Cell Counter included.|
|Particle Size||6 µm–36 µm|
|Cell Size Range||4.0–25.0 µm|
|Safety Information according to GHS|
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|Storage and Shipping Information|
|Material Size||1 ea|
|Reference overview||Application||Pub Med ID|
|Nucleolar protein trafficking in response to HIV-1 Tat: rewiring the nucleolus.|
Jarboui, MA; Bidoia, C; Woods, E; Roe, B; Wynne, K; Elia, G; Hall, WW; Gautier, VW
PloS one 7 e48702 2012
The trans-activator Tat protein is a viral regulatory protein essential for HIV-1 replication. Tat trafficks to the nucleoplasm and the nucleolus. The nucleolus, a highly dynamic and structured membrane-less sub-nuclear compartment, is the site of rRNA and ribosome biogenesis and is involved in numerous cellular functions including transcriptional regulation, cell cycle control and viral infection. Importantly, transient nucleolar trafficking of both Tat and HIV-1 viral transcripts are critical in HIV-1 replication, however, the role(s) of the nucleolus in HIV-1 replication remains unclear. To better understand how the interaction of Tat with the nucleolar machinery contributes to HIV-1 pathogenesis, we investigated the quantitative changes in the composition of the nucleolar proteome of Jurkat T-cells stably expressing HIV-1 Tat fused to a TAP tag. Using an organellar proteomic approach based on mass spectrometry, coupled with Stable Isotope Labelling in Cell culture (SILAC), we quantified 520 proteins, including 49 proteins showing significant changes in abundance in Jurkat T-cell nucleolus upon Tat expression. Numerous proteins exhibiting a fold change were well characterised Tat interactors and/or known to be critical for HIV-1 replication. This suggests that the spatial control and subcellular compartimentaliation of these cellular cofactors by Tat provide an additional layer of control for regulating cellular machinery involved in HIV-1 pathogenesis. Pathway analysis and network reconstruction revealed that Tat expression specifically resulted in the nucleolar enrichment of proteins collectively participating in ribosomal biogenesis, protein homeostasis, metabolic pathways including glycolytic, pentose phosphate, nucleotides and amino acids biosynthetic pathways, stress response, T-cell signaling pathways and genome integrity. We present here the first differential profiling of the nucleolar proteome of T-cells expressing HIV-1 Tat. We discuss how these proteins collectively participate in interconnected networks converging to adapt the nucleolus dynamic activities, which favor host biosynthetic activities and may contribute to create a cellular environment supporting robust HIV-1 production.
|Anti-inflammatory effects of benfotiamine are mediated through the regulation of the arachidonic acid pathway in macrophages.|
Shoeb, Mohammad and Ramana, Kota V
Free Radic. Biol. Med., 52: 182-90 (2012) 2012
Benfotiamine, a lipid-soluble analogue of vitamin B1, is a potent antioxidant that is used as a food supplement for the treatment of diabetic complications. Our recent study (U.C. Yadav et al., Free Radic. Biol. Med. 48:1423-1434, 2010) indicates a novel role for benfotiamine in the prevention of bacterial endotoxin, lipopolysaccharide (LPS)-induced cytotoxicity and inflammatory response in murine macrophages. Nevertheless, it remains unclear how benfotiamine mediates anti-inflammatory effects. In this study, we investigated the anti-inflammatory role of benfotiamine in regulating arachidonic acid (AA) pathway-generated inflammatory lipid mediators in RAW264.7 macrophages. Benfotiamine prevented the LPS-induced activation of cPLA2 and release of AA metabolites such as leukotrienes, prostaglandin E2, thromboxane 2 (TXB2), and prostacyclin (PGI2) in macrophages. Further, LPS-induced expression of AA-metabolizing enzymes such as COX-2, LOX-5, TXB synthase, and PGI2 synthase was significantly blocked by benfotiamine. Furthermore, benfotiamine prevented the LPS-induced phosphorylation of ERK1/2 and expression of transcription factors NF-?B and Egr-1. Benfotiamine also prevented the LPS-induced oxidative stress and protein-HNE adduct formation. Most importantly, compared to specific COX-2 and LOX-5 inhibitors, benfotiamine significantly prevented LPS-induced macrophage death and monocyte adhesion to endothelial cells. Thus, our studies indicate that the dual regulation of the COX and LOX pathways in AA metabolism could be a novel mechanism by which benfotiamine exhibits its potential anti-inflammatory response.
|NAD(P)H:quinone oxidoreductase 1 protects lungs from oxidant-induced emphysema in mice.|
Potts-Kant, Erin N, et al.
Free Radic. Biol. Med., 52: 705-15 (2012) 2012
Emphysema is currently a leading cause of mortality with no known effective therapy to attenuate progressive loss of lung function. Previous work supports that activation of nuclear factor erythroid 2-related factor 2 (Nrf2) is protective to the lung through induction of hundreds of antioxidant genes. In models of lung injury, the expression of NAD(P)H:quinone oxidoreductase 1 (NQO1) is upregulated in a manner dependent on Nrf2 and human emphysema is associated with reduced levels of NQO1. However, the functional role of NQO1 in emphysema remains unknown. In this study, we demonstrate the protective role of NQO1 in the development of emphysema using mouse models. NQO1-deficient animals demonstrated premature age-related emphysema and were more susceptible to both elastase and inhaled lipopolysaccharide models of emphysema. The absence of NQO1 was associated with enhanced markers of oxidant stress. Treatment of NQO1-deficient animals with the antioxidant N-acetylcysteine reversed the NQO1-dependent emphysematous changes. In vitro studies utilizing either inhibition or induction of NQO1 demonstrated a potent antioxidant role of NQO1 in macrophages, suggesting a role for macrophage-derived oxidants in the pathogenesis of emphysema. These novel findings support a functional role for NQO1 in protecting the lung from development of emphysema.
|Retinoic acid-induced differentiation increases the rate of oxygen consumption and enhances the spare respiratory capacity of mitochondria in SH-SY5Y cells.|
Xun, Zhiyin, et al.
Mechanisms of ageing and development, (2012) 2012
Retinoic acid (RA) is used in differentiation therapy to treat a variety of cancers including neuroblastoma. The contributing factors for its therapeutic efficacy are poorly understood. However, mitochondria (MT) have been implicated as key effectors in RA-mediated differentiation process. Here we utilize the SH-SY5Y human neuroblastoma cell line as a model to examine how RA influences MT during the differentiation process. We find that RA confers an approximately sixfold increase in the oxygen consumption rate while the rate of glycolysis modestly increases. RA treatment does not increase the number of MT or cause measurable changes in the composition of the electron transport chain. Rather, RA treatment significantly increases the mitochondrial spare respiratory capacity. We propose a competition model for the therapeutic effects of RA. Specifically, the high metabolic rate in differentiated cells limits the availability of metabolic nutrients for use by the undifferentiated cells and suppresses their growth. Thus, RA treatment provides a selective advantage for the differentiated state.
|IL-23 Dampens the Allergic Response to Cryptococcus neoformans through IL-17-Independent and -Dependent Mechanisms.|
Szymczak, Wendy A, et al.
Am. J. Pathol., 180: 1547-59 (2012) 2012
The cytokines IL-23 and IL-17 have been implicated in resistance to cryptococcal disease, but it is not clear whether IL-23-mediated production of IL-17 promotes fungal containment following pulmonary challenge with Cryptococcus neoformans. We used mice lacking IL-23 (IL-23p19(-/-)) or IL-17RA (IL-17RA(-/-)), and wild type (WT) C57BL/6 mice to examine the IL-23/IL-17 axis after intranasal infection with the C. neoformans strain 52D. The absence of IL-23 or IL-17RA had no effect on pulmonary or brain fungal burden at 1 or 6 weeks after infection. However, survival of IL-23p19(-/-) mice was reduced compared to IL-17RA(-/-) mice. IL-I7 production by CD4 T cells and natural killer T (NKT) cells was impaired in IL-23p19(-/-) lungs, but was not completely abolished. Both IL-23p19(-/-) and IL-17RA(-/-) mice exhibited impaired neutrophil recruitment, increased serum levels of IgE and IgG2b, and increased deposition of YM1/YM2 crystals in the lung, but only IL-23p19(-/-) mice developed persistent lung eosinophilia. Although survival of IL-17RA(-/-) and WT mice was similar after 17 weeks of infection, only surviving IL-17RA(-/-) mice exhibited cryptococcal dissemination to the blood. These data demonstrate that IL-23 dampens the allergic response to cryptococcal infection through IL-17-independent suppression of eosinophil recruitment and IL-17-dependent regulation of antibody production and crystal deposition. Furthermore, IL-23, and to a lesser extent IL-17, contribute to disease resistance.
|Overexpression of the lung cancer-prognostic miR-146b microRNAs has a minimal and negative effect on the malignant phenotype of A549 lung cancer cells.|
Patnaik, Santosh Kumar, et al.
PLoS ONE, 6: e22379 (2011) 2011
Expression levels of miR-146b-5p and -3p microRNAs in human non-small cell lung cancer (NSCLC) are associated with recurrence of the disease after surgery. To understand this, the effect of miR-146b overexpression was studied in A549 human lung cancer cells.
|Oncogenes induce senescence with incomplete growth arrest and suppress the DNA damage response in immortalized cells.|
Sherman, Michael Y, et al.
Aging Cell, 10: 949-61 (2011) 2011
Activation of the Her2 (ErbB2) oncogene is implicated in the development of breast, ovary and other cancers. Here, we show that expression of NeuT, a mutant-activated rodent isoform of Her2, in immortalized breast epithelial cells, while promoting senescence-associated morphological changes, up-regulation of senescence-associated ?-galactosidase activity, and accumulation of the cyclin-dependent kinase inhibitor p21, failed to trigger the major senescence end-point, i.e. permanent growth arrest. Similar senescence-associated phenotype with incomplete growth arrest, which we dubbed senescence with incomplete growth arrest (SWING), could also be triggered by the expression of the Ras oncogene. SWING phenotype was stable, and persisted in tumor xenografts established from NeuT-transduced cells. Furthermore, a significant population of cells in SWING state was found in tumors in the MMTV/NeuT transgenic mouse model. SWING cells showed downregulation of histone H2AX, critical for repair of double-stranded DNA breaks, and impaired activation of Chk1 kinase. Overall, SWING cells were characterized by increased DNA instability and hypersensitivity to genotoxic stresses. We propose that the SWING state could be a stage in the process of cancer development.
|Reduction of infarct size by intravenous injection of uncultured adipose derived stromal cells in a rat model is dependent on the time point of application.|
van Dijk, A, et al.
Stem Cell Res, 7: 219-29 (2011) 2011
Stem cell therapy is a promising tool to improve outcome after acute myocardial infarction (AMI), but needs to be optimized since results from clinical applications remain ambiguous. A potent source of stem cells is the stromal vascular fraction of adipose tissue (SVF), which contains high numbers of adipose derived stem cells (ASC). We hypothesized that: 1) intravenous injection can be used to apply stem cells to the heart. 2) Uncultured SVF cells are easier and safer when cultured ASCs. 3) Transplantation after the acute inflammation period of AMI is favorable over early injection. For this, AMI was induced in rats by 40min of coronary occlusion. One or seven days after AMI, rats were intravenously injected with vehicle, 5×10(6) uncultured rat SVF cells or 1×10(6) rat ASCs. Rats were analyzed 35 days after AMI. Intravenous delivery of both fresh SVF cells and cultured ASCs 7 days after AMI significantly reduced infarct size compared to vehicle. Similar numbers of stem cells were found in the heart, after treatment with fresh SVF cells and cultured ASCs. Importantly, no adverse effects were found after injection of SVF cells. Using cultured ASCs, however, 3 animals had shortness of breath, and one animal died during injection. In contrast to application at 7 days post AMI, injection of SVF cells 1 day post AMI resulted in a small but non-significant infarct reduction (p=0.35). Taken together, intravenous injection of uncultured SVF cells subsequent to the acute inflammation period, is a promising stem cell therapy for AMI.
|MicroRNAs 10a and 10b are potent inducers of neuroblastoma cell differentiation through targeting of nuclear receptor corepressor 2.|
Foley, N H, et al.
Cell Death Differ., 18: 1089-98 (2011) 2011
MicroRNAs function as negative regulators of posttranscriptional gene expression, having major roles in cellular differentiation. Several neuroblastoma cell lines can be induced to undergo differentiation by all-trans-retinoic acid (ATRA) and are used for modeling signaling pathways involved in this process. To identify miRNAs contributing to differentiation, we profiled 364 loci following ATRA treatment of neuroblastoma cell lines and found miR-10a and miR-10b to be highly overexpressed in SK-N-BE, LAN5 and SHSY-5Y. Ectopic overexpression of these miRNAs led to a major reprogramming of the transcriptome and a differentiated phenotype that was similar to that induced by ATRA in each of these cell lines. One of the predicted downregulated miR-10a/b targets was nuclear receptor corepressor 2 (NCOR2), a corepressor of gene transcription, which is known to suppress neurite outgrowth. NCOR2 was experimentally validated as a direct target of miR-10a/b, and siRNA-mediated inhibition of this mRNA alone resulted in neural cell differentiation. Moreover, induction of differentiation could be blocked by ectopic upregulation of NCOR2 using an expression construct lacking the miR-10a/b 3' untranslated region target site. We conclude that miR-10a/b has major roles in the process of neural cell differentiation through direct targeting of NCOR2, which in turn induces a cascade of primary and secondary transcriptional alterations, including the downregulation of MYCN.
|Bardet-Biedl syndrome highlights the major role of the primary cilium in efficient water reabsorption.|
Marion, Vincent, et al.
Kidney international, (2011) 2011
Studies of the primary cilium, now known to be present in all cells, have undergone a revolution, in part, because mutation of many of its proteins causes a large number of diseases, including cystic kidney disease. Bardet-Biedl syndrome (BBS) is an inherited ciliopathy characterized, among other dysfunctions, by renal defects for which the precise role of the cilia in kidney function remains unclear. We studied a cohort of patients with BBS where we found that these patients had a urinary concentration defect even when kidney function was near normal and in the absence of major cyst formation. Subsequent in vitro analysis showed that renal cells in which a BBS gene was knocked down were unciliated, but did not exhibit cell cycle defects. As the vasopressin receptor 2 is located in the primary cilium, we studied BBS-derived unciliated renal epithelial cells and found that they were unable to respond to luminal arginine vasopressin treatment and activate their luminal aquaporin 2. The ability to reabsorb water was restored by treating these unciliated renal epithelial cells with forskolin, a receptor-independent adenylate cyclase activator, showing that the intracellular machinery for water absorption was present but not activated. These findings suggest that the luminal receptor located on the primary cilium may be important for efficient transepithelial water absorption.Kidney International advance online publication, 26 January 2011; doi:10.1038/ki.2010.538.
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Newsletters / Publications
|Cellutions - The Newsletter for Cell Biology Researchers Vol 1: 2012|
|Cellutions - The newsletter for Cell Biology Researchers Volume 3: 2011|
|Cellutions Newsletter: 2011, Vol. 1|
|Cellutions Newsletter: 2011, Vol. 2|