17 research outputs found
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Protamine enhances epidermal growth factor (EGF)-stimulated mitogenesis by increasing cell surface EGF receptor number. Implications for existence of cryptic EGF receptors
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Modulation of the epidermal growth factor receptor by brainâderived growth factor in Swiss mouse 3T3 cells
Incubation of Swiss mouse 3T3 cells at 37°C with bovine brainâderived growth factor (BDGF) decreased the cell surface 125IâEGF binding activity of these cells by 70â80%. This downâmodulation of the EOF receptor by BDGF was time, temperature, and dose dependent Scatchard plot analysis indicated that BDGF binding led to a selective decrease in the number of highâaffinity EGF receptors. The BDGFâinduced downâmodulation of the EGF receptor was completely blocked by protamine, a potent inhibitor of receptor binding and mitogenic activities of BDGF.
BDGF downâmodulated the EGF receptor in phorbol myristic acetate (PMA)âpretreated cells, as well as in control cells. Furthermore, PMAâpretreated cells responded mitogenically to BDGF, whereas PMA itself failed to stimulate the mitogenic response of PMAâpretreated cells. This BDGFâinduced downâmodulation of the EGF receptor in PMAâdesensitized cells suggests that BDGF downâregulates the EGF receptor by a mechanism distinct from that of PMA.
Incubation of cells with compounds which are known to inhibit pinocytosis blocked the downâmodulation induced either by BDGF or by plateletâderived growth factor (PDGF) but had no effect on the PMAâinduced downâmodulation. Incubation of cells with inhibitors of receptor recycling enhanced the BDGFâinduced downâmodulation of the EGF receptor. These results suggest that BDGF and PDGF induce downâmodulation of the EGF receptor by increasing the internalization of cell surface highâaffinity receptors and that the internalization process may not be required for downâmodulation induced by PMA
Cellular growth inhibition by TGF-β1 involves IRS proteins
AbstractIn Mv1Lu cells, insulin partially reverses transforming growth factor-β1 (TGF-β1) growth inhibition in the presence of ι5β1 integrin antagonists. TGF-β1 appears to induce phosphorylation of IRS-2 in these cells; this is inhibited by a TGF-β antagonist known to reverse TGF-β growth inhibition. Stable transfection of 32D myeloid cells (which lack endogenous IRS proteins and are insensitive to growth inhibition by TGF-β1) with IRS-1 or IRS-2 cDNA confers sensitivity to growth inhibition by TGF-β1; this IRS-mediated growth inhibition can be partially reversed by insulin in 32D cells stably expressing IRS-2 and the insulin receptor (IR). These results suggest that growth inhibition by TGF-β1 involves IRS proteins
Inhibitors of clathrin-dependent endocytosis enhance TGFβ signaling and responses
Clathrin-dependent endocytosis is believed to be involved in
TGFβ-stimulated cellular responses, but the subcellular locus at which
TGFβ induces signaling remains unclear. Here, we demonstrate that
inhibitors of clathrin-dependent endocytosis, which are known to arrest the
progression of endocytosis at coated-pit stages, inhibit internalization of
cell-surface-bound TGFβ and promote colocalization and accumulation of
TβR-I and SARA at the plasma membrane. These inhibitors enhance
TGFβ-induced signaling and cellular responses (Smad2
phosphorylation/nuclear localization and expression of PAI-1). Dynasore, a
newly identified inhibitor of dynamin GTPase activity, is one of the most
potent inhibitors among those tested and, furthermore, is a potent enhancer of
TGFβ. Dynasore ameliorates atherosclerosis in the aortic endothelium of
hypercholesterolemic ApoE-null mice by counteracting the suppressed
TGFβ responsiveness caused by the hypercholesterolemia, presumably acting
through its effect on TGFβ endocytosis and signaling in vascular
cells
CRSBP-1/LYVE-1 ligands stimulate contraction of the CRSBP-1-associated ER network in lymphatic endothelial cells
Incidence of and Factors Associated with False Positives in Laboratory Diagnosis of Norovirus Infection by Amplification of the RNA-Dependent RNA Polymerase Gene
<div><p>Background</p><p>Conventional reverse transcription-polymerase chain reaction (RT-PCR) amplification of the RNA-dependent RNA polymerase (RdRp) gene remains a used method for the rapid detection of norovirus (NV) in clinical laboratories. The incidence of and factors associated with false positives in this assay have not been previously evaluated.</p><p>Methods/Principal Findings</p><p>After an NV outbreak caused by the GII.4 Sydney strain in 2012, we reanalysed 250 stool samples positive for NV by RdRp gene detection. True positives were confirmed in 154 (61.6%) samples by successful amplification and sequencing confirmation of the viral protein 1 gene. Of the remaining 96 samples that underwent RT-PCR for the RdRp gene, 34 samples yielded PCR products of the expected length. However, the sequences of the amplicons belonged to the human genome, with 91â97% matched nucleotide sequences, indicating false positives. Multivariate analysis of the clinical features of the patients identified a positive stool culture for bacteria (adjusted odds ratio [aOR] 9.07, 95% adjusted confidence interval [aCI] 2.17â37.92, <i>P</i>â=â.003) and the use of parenteral antibiotics (aOR 5.55, 95% aCI 1.21â24.73, <i>P</i>â=â.027) as significant and independent factors associated with false positives.</p><p>Conclusion</p><p>Conventional RT-PCR targeting the RdRp gene of NV can lead to false positives in patients with bacterial enterocolitis by incidental amplification of DNA from a human source.</p></div