Journal: American Journal of Physiology - Heart and Circulatory Physiology

Article Title: Angiopoietin-1 prevents severe bleeding complications induced by heparin-like drugs and fibroblast growth factor-2 in mice

doi: 10.1152/ajpheart.00373.2015

Figure Lengend Snippet: PPS did not affect the expression of metalloproteinases or VEGF-A in the intestine of mice infected with rAd-LacZ. A: representative pictures of the Western blots done with samples derived from the small intestinal of mice infected with rAd-LacZ vectors and treated with PPS (60 mg/kg) or phosphate-buffered saline (PBS). Only the intestinal expression of ICAM-1 was significantly increased in mice treated with rAd-LacZ + PPS relative to the control mice infected with rAd-LacZ (*P < 0.05; n = 6 samples per group). B: the densitometric analysis corresponding to the Western blots. Results are expressed in arbitrary optical density units as ratio of the beta-actin expression (means ± SE). Samples from 6 different mice were included in each group.

Article Snippet: Recombinant adenoviral (rAd) vectors. rAd-vectors carrying either the Escherichia coli LacZ gene (rAd-Lac-Z), a 700-bp cDNA sequence encoding a secreted form of human recombinant FGF-2 (rAd-FGF-2), or a genetically modified form of angiopoietin-1 (Ang-1*) from Regeneron Pharm (Tarrytown, NY), were generated, amplified, and purified as previously described ( 13 , 21 , 44 , 50 ).

Techniques: Expressing, Infection, Western Blot, Derivative Assay, Saline, Control

Journal: PLoS Biology

Article Title: A Lack of Premature Termination Codon Read-Through Efficacy of PTC124 (Ataluren) in a Diverse Array of Reporter Assays

doi: 10.1371/journal.pbio.1001593

Figure Lengend Snippet: (A) Incubation with G418 for 24 hours leads to a dose-dependent increase in enzyme activity of stably transfected PTC-β-Gal. (B) 24-hour incubation with PTC124 at concentrations in excess of its reported effective concentration range of 0.1–30 µM has no effect. All data points represent mean ± standard deviation (n = 5).

Article Snippet: The cysteine 320 codon of the lac Z gene in the pSV-β-galactosidase vector (Promega) was replaced by a premature TGA stop codon using the QuickChange site-directed mutagenesis system (Stratagene) with the following primers: pSV- lac Z-TGA.F 5′ GAT TGA AGC AGA AGC ATG AGA TGT CGG TTT CCG CG 3′ and pSV- lac Z-TGA.R 5′ CGC GGA AAC CGA CAT CGC AGG CTT CTG CTT CAA TC 3′ .

Techniques: Incubation, Activity Assay, Stable Transfection, Transfection, Concentration Assay, Standard Deviation

Journal:

Article Title: Analysis of dofA , a fruA -Dependent Developmental Gene, and Its Homologue, dofB , in Myxococcus xanthus

doi: 10.1128/JB.184.24.6803-6810.2002

Figure Lengend Snippet: Bacterial strains and plasmids used in this study

Article Snippet: pGEM-T easy , Ap r lacZ cloning vector , Promega.

Techniques: Plasmid Preparation, Clone Assay

Journal: Free Radical Biology & Medicine

Article Title: Opposite effects of HIF-1α and HIF-2α on the regulation of IL-8 expression in endothelial cells

doi: 10.1016/j.freeradbiomed.2011.08.023

Figure Lengend Snippet: HIF-2α overexpression increases SP-1 activity. (A) SP-1 transcription factor binding site found in the promoter of IL-8 using the Alibaba 2.1 program. (B) Cells were cotransfected with the pSP-1-luc vector containing the SP-1 binding site driving luciferase expression (500 ng) and a plasmid containing the LacZ gene (100 ng) as an internal control and after 24 h transduced with AdHIF-1α, AdHIF-2α, or AdGFP for the next 48 h. As shown by measurement of luciferase activity, HIF-2α augmented SP-1 activity. (C) Real-time PCR was performed to examine the mRNA level of a TP gene containing binding sites for SP-1 after 48 h transduction with AdHIF-1α, AdHIF-2α, or AdGFP. TP mRNA was upregulated only by HIF-2α (a similar tendency of increase in TP was noted in three independent experiments). (D) Real-time PCR and (E) ELISA were performed to examine the mRNA and protein level of IL-8, respectively, after 48 h transduction with AdHIF-2α or AdGFP together with stimulation with 10 μM SP-1 inhibitor, mithramycin A. Note that the effect of HIF-2α on IL-8 was partially reversed by mithramycin A, particularly at the mRNA level. (F) Cells were transfected for 48 h with the pCMV-SP-1 vector (500 ng) or control Bluescript plasmid (100 ng). As shown by ELISA, SP-1 overexpression tended to increase IL-8 production. (G) SP-1 transcription factor binding site found in the promoter of IL-8 was mutated at guanine residues (in red in (A)). Such mutation did not change HIF-2α-induced IL-8 promoter activity. Each bar represents the mean ± SD of three to six independent experiments performed in duplicate. * p < 0.05, comparing AdGFP vs AdHIF-1α or AdHIF-2α; # p < 0.05, comparing AdGFP control vs AdGFP stimulated or AdHIF-2α control vs AdHIF-2α stimulated.

Article Snippet: The pCMV-lacZ vector containing the β-galactosidase gene driven by the CMV promoter was purchased from Promega and was cotransfected into cells together with one of the above-described reporter plasmids as an internal control.

Techniques: Over Expression, Activity Assay, Binding Assay, Plasmid Preparation, Luciferase, Expressing, Transduction, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Transfection, Mutagenesis