Thioredoxin downregulates VCAM-1 and ICAM-1 expression in HUVECs.

<p>(<b>A</b>) <i>Top</i>, Immunoblot of Trx1 in HUVECs infected by GFP adenovirus (Ad-GFP), Trx adenovirus (Ad-Trx), and TD adenovirus (Ad-TD). <i>Bottom</i>, Trx1 activity in Ad-GFP, Ad-Trx, and Ad-TD cells, determined by insulin reduction-based assay. (<b>B</b>) Reactive oxygen species production in HUVECs. After 2 h stimulation of ox-LDL (100 µg/ml), ROS production was analyzed by measuring the mean fluorescence intensity using flow cytometry. (<b>C</b>, <b>D</b>) Immunoblot of VCAM-1 and ICAM-1 in Ad-GFP, Ad-Trx, and Ad-TD cells under basal conditions and after 4 h ox-LDL (100 µg/ml) stimulation. Relative VCAM-1 and ICAM-1 expression was determined by densitometric analysis. In all of the histograms, each value represents the mean ±SEM (<i>n</i> = 3 independent measurements). *<i>p</i><0.05, compared with unstimulated Ad-GFP cells; ^#<i>p</i><0.05, compared with ox-LDL-stimulated Ad-GFP cells. (<b>E</b>) U937 monocyte adhesion assay in ox-LDL-stimulated Ad-GFP, Ad-Trx, and Ad-TD cells. U937 cells that adhered to HUVECs were observed under a fluorescent microscope at 200× magnification. (<b>F</b>) The intensity of fluorescence-labeled adherent U937 monocytes was measured with a fluorometer (excitation wavelength, 485 nm; emission wavelength, 530 nm). The intensity was normalized to that of control cells in each group. The data are expressed as mean ±SEMs (<i>n</i> = 5). *<i>p</i><0.05, compared with ox-LDL-treated Ad-GFP group; ^#<i>p</i><0.05, compared with unstimulated Ad-GFP group. (<b>G</b>) <i>Top</i>, Immunoblot of Trx1, VCAM-1, and ICAM-1 in wildtype HUVECs (Wt) and HUVECs transfected by negative control siRNA (NC) or Trx siRNA (si-Trx). <i>Bottom</i>, Relative Trx, VCAM-1, and ICAM-1 expression was determined by densitometric analysis. In all of the histograms, each value represents the mean ±SEM (<i>n</i> = 3 independent measurements). *<i>p</i><0.05, compared with Wt cell.</p

Role of SIS3 in VCAM-1 and ICAM-1 expression in HUVECs stimulated with ox-LDL.

<p>(<b>A</b>) Overexpression of Trx suppressed ox-LDL-induced VCAM-1 expression. The pretreatment of Ad-Trx cells with SIS3 for 1 h completely reversed the inhibitory effect of Trx on VCAM-1 expression. (<b>B</b>) The overexpression of Trx inhibited ox-LDL-induced ICAM-1 expression. The pretreatment of Ad-Trx cells with SIS3 reversed the inhibitory effect of Trx on ICAM-1 expression. The relative expression of VCAM-1 and ICAM-1 was determined by densitometric analysis. The data from three separate experiments are expressed as mean ±SEM. *<i>p</i><0.05, compared with unstimulated Ad-GFP cells; ^#<i>p</i><0.05, compared with ox-LDL-stimulated Ad-GFP cells; ^$<i>p</i><0.05, compared with ox-LDL-stimulated Ad-Trx cells.</p

Role of Trx in nuclear translocation of pSmad3.

<p>(<b>A</b>) <i>Top</i>, Nuclear pSmad3 and Trx proteins in Ad-GFP, Ad-Trx, and Ad-TD cells. <i>Bottom</i>, Cytosolic Trx expression in Ad-GFP, Ad-Trx, and Ad-TD cells. The cells were treated with or without ox-LDL (100 µg/ml) stimulation for 6 h as indicated. <i>Left column</i>, Quantitative nuclear pSmad3 protein is shown. <i>Right column</i>, Nuclear and cytosolic Trx expression was normalized to that of the unstimulated Ad-GFP group. The numbers above the columns indicate the relative expression ratio of nuclear Trx to cytosolic Trx. The data from three separate experiments are expressed as mean ±SEM. *<i>p</i><0.05, compared with unstimulated Ad-GFP cells; ^#<i>p</i><0.05, compared with ox-LDL-stimulated Ad-GFP cells. (<b>B</b>) Phosphorylated Smad3 expression in Ad-GFP, Ad-Trx, and Ad-TD cells was assessed by immunofluorescent analysis.</p

Smad3 is a newly recognized interaction partner of Trx.

<p>Reciprocal immunoprecipitation was performed using an anti-Trx antibody. Smad3 and pSmad3 were co-immunoprecipitated with Trx. Rabbit IgG served as a negative control. (<b>A, B</b>) Immunoblot with anti-Trx, anti-Smad3, or anti-pSmad3 antibody in wildtype HUVECs under basal or ox-LDL-stimulated conditions. The data from three separate experiments are expressed as mean ±SEM. *<i>p</i><0.05, compared with unstimulated HUVECs. (<b>C, D</b>) Immunoblot with anti-Trx, anti-Smad3, or anti-pSmad3 antibody in Ad-GFP (1, 2, 5, 6), Ad-Trx (3, 7), and Ad-TD (4, 8) cells under basal or ox-LDL-stimulated conditions. The data from three separate experiments are expressed as mean ±SEM. *<i>p</i><0.05, compared with unstimulated Ad-GFP cells; ^#<i>p</i><0.05, compared with unstimulated Ad-Trx cells; ^$<i>p</i><0.05, compared with unstimulated Ad-TD cells.</p

Effect of Trx on the expression of pSmad3 and Smad3 in HUVECs stimulated with ox-LDL and nLDL.

<p>(<b>A</b>) Immunoblot of pSmad3 and Smad3 expression in ox-LDL-stimulated Ad-GFP, Ad-Trx, and Ad-TD HUVECs. (<b>B</b>) Immunoblot of pSmad3 and Smad3 expression in nLDL-treated Ad-GFP, Ad-Trx, and Ad-TD HUVECs. Relative pSmad3 and Smad3 content was determined by densitometric analysis. The data from three separate experiments are expressed as mean ±SEM. *<i>p</i><0.05, compared with unstimulated Ad-GFP cells (Smad3/β-actin); ^#<i>p</i><0.05, compared with ox-LDL-stimulated Ad-GFP cells (pSmad3/Smad3); ^&<i>p</i><0.05, unstimulated cells compared with ox-LDL stimulated cells (Smad3/β-actin).</p

Variation rule of branch fracture propagation pressure.

<p>(a) Δσ=0 MPa, K_<b>IC</b>=0.8 MPa m^0.5; (b) Δσ=0 MPa, K_<b>IC</b>=1.6 MPa m^0.5; (c) Δσ=5 MPa, K_<b>IC</b>=0.8 MPa m^0.5; (d) Δσ=5 MPa, K_<b>IC</b>=1.6 MPa m^0.5; (e) Δσ=10 MPa, K_<b>IC</b>=0.8 MPa m^0.5; (f) Δσ=10 MPa, K_<b>IC</b>=1.6 MPa m^0.5.</p

Production performance of oil well under different fracturing patterns.

<p>Production performance of oil well under different fracturing patterns.</p

Classification of the FPP.

<p>Classification of the FPP.</p

Flow chart of the solution to the combined stress field.

<p>Flow chart of the solution to the combined stress field.</p

Stress and strain field at the fracture tip.

<p>Stress and strain field at the fracture tip.</p