Journal: Toxicology

Article Title: Differential Fmo3 Gene Expression in Various Liver Injury Models Involving Hepatic Oxidative Stress in Mice

doi: 10.1016/j.tox.2014.08.013

Figure Lengend Snippet: Livers were collected from mice (n=6) sacrificed at respective time-points (2,4,8,12,24 & 48h for ANIT; 24 & 48h for CCl4; 6 & 24h for AlOH; and 10 d for BDL). RNA was isolated and cDNA was made using a commercial MMLV-RT kit. The cDNA samples were analyzed by quantitative RT-PCR using Fmo3 mouse-specific primers. Gene expression was normalized to the housekeeping gene β-actin. Fmo3 mRNA expression is presented as mean Fold Change ± SE. One-way ANOVA, t-test or two-way ANOVA was performed, appropriately, followed by the Dunnett's posttest for One-way ANOVA and the Bonferroni posttest for two-way ANOVA. Asterisks (*) represent a statistical difference (p < 0.05) between vehicle-treated and hepatotoxicant-treated or BDL group.

Article Snippet: Membranes were blocked with 5 % non-fat powdered milk in tris buffered saline containing 0.05% tween-20(TBS-T) for 8 h. A rabbit anti-mouse Fmo3 primary antibody (GenScript USA Inc., NJ) (1:5000) was used to detect Fmo3 with β-actin as a loading control.

Techniques: Isolation, Quantitative RT-PCR, Expressing

Journal: Toxicology

Article Title: Differential Fmo3 Gene Expression in Various Liver Injury Models Involving Hepatic Oxidative Stress in Mice

doi: 10.1016/j.tox.2014.08.013

Figure Lengend Snippet: Western immunoblots for Fmo3 were performed using liver microsomes from control and hepatotoxicant-treated or BDL mice. A custom-made rabbit anti-mouse Fmo3 primary antibody, described in Materials and Methods was used to detect Fmo3. Fmo3 protein levels were normalized to β-actin loading control. Microsomal proteins isolated from naïve female mouse liver were used as a positive control (indicated by “+” sign). The data are presented as blots and as mean Fmo3 protein expression (Fold Change) ± SE. One-way ANOVA, t-test or two-way ANOVA was performed, appropriately, followed by the Dunnett's posttest for One-way ANOVA and the Bonferroni posttest for two-way ANOVA. Asterisks (*) represent a statistical difference (p < 0.05) between vehicle-treated and hepatotoxicant-treated or BDL group.

Article Snippet: Membranes were blocked with 5 % non-fat powdered milk in tris buffered saline containing 0.05% tween-20(TBS-T) for 8 h. A rabbit anti-mouse Fmo3 primary antibody (GenScript USA Inc., NJ) (1:5000) was used to detect Fmo3 with β-actin as a loading control.

Techniques: Western Blot, Isolation, Positive Control, Expressing

Journal: Toxicology

Article Title: Differential Fmo3 Gene Expression in Various Liver Injury Models Involving Hepatic Oxidative Stress in Mice

doi: 10.1016/j.tox.2014.08.013

Figure Lengend Snippet: Plasma and livers were collected from mice 72 h following APAP (400 mg/kg) or vehicle treatment. (A) The data are presented as mean plasma ALT (IU/L) ± SE. (B) RNA was isolated from livers andcDNA samples were analyzed by quantitative RT-PCR using Fmo3 mouse-specific primers. Gene expression was normalized to the housekeeping gene β-actin. Fmo3 mRNA expression are presented as mean Fold Change ± SE. Oneway ANOVA was performed followed by the Dunnett's post-test. Asterisks (*) represent a statistical difference (p < 0.05) between vehicle-treated group and APAP-treated group and hash (#) represent a statistical difference (p<0.05) compared with APAP-treated wild-type mice.

Article Snippet: Membranes were blocked with 5 % non-fat powdered milk in tris buffered saline containing 0.05% tween-20(TBS-T) for 8 h. A rabbit anti-mouse Fmo3 primary antibody (GenScript USA Inc., NJ) (1:5000) was used to detect Fmo3 with β-actin as a loading control.

Techniques: Isolation, Quantitative RT-PCR, Expressing

Journal: Toxicology

Article Title: Differential Fmo3 Gene Expression in Various Liver Injury Models Involving Hepatic Oxidative Stress in Mice

doi: 10.1016/j.tox.2014.08.013

Figure Lengend Snippet: After overnight fasting, groups of wild-type and Nrf2 knockout mice received a single dose of 400 mg/kg APAP or vehicle. Livers were collected 72 h following APAP or vehicle treatments. Western blot for Fmo3 was performed using liver microsomes from control and APAP-treated mice. Equal protein loading (10 μg protein/lane) was confirmed by detection of β-actin. Microsomal proteins isolated from naïve female mouse liver were used as a positive control indicated by “+” sign. The data are presented as blots and as mean Fmo3 protein expression (Fold Change) ± SE (A). FMO activity was measured in liver microsomes from control and APAP-treated mice using methimazole as substrate. Data are presented as mean Specific Activity (μM/min/mg) ± SE (B). Asterisks (*) represent a statistical difference (p < 0.05) between vehicle-treated group and APAP-treated group and hash (#) represent a statistical difference (p<0.05) compared with APAP-treated wild-type mice.

Article Snippet: Membranes were blocked with 5 % non-fat powdered milk in tris buffered saline containing 0.05% tween-20(TBS-T) for 8 h. A rabbit anti-mouse Fmo3 primary antibody (GenScript USA Inc., NJ) (1:5000) was used to detect Fmo3 with β-actin as a loading control.

Techniques: Knock-Out, Western Blot, Isolation, Positive Control, Expressing, Activity Assay

Journal: PLoS ONE

Article Title: RGD-Targeted Ultrasound Contrast Agent for Longitudinal Assessment of Hep-2 Tumor Angiogenesis In Vivo

doi: 10.1371/journal.pone.0149075

Figure Lengend Snippet: Adhesion of different types of microbubbles to bEnd.3 cells or αvβ3-negative MCF-7 cells with and without pre incubation with anti-alpha(v) antibodies.

Article Snippet: To visualize ανβ3 integrin expression on tumor endothelial cells, a double immunostaining procedure was performed using a rabbit anti-mouse alpha(v) primary antibody (BD Biosciences, San Jose, CA) at a 1:50 dilution and a FITC-labeled goat anti-rabbit secondary antibody (BD Biosciences, San Jose, CA) at a 1:100 dilution.

Techniques: Incubation, Standard Deviation

Journal: PLoS ONE

Article Title: RGD-Targeted Ultrasound Contrast Agent for Longitudinal Assessment of Hep-2 Tumor Angiogenesis In Vivo

doi: 10.1371/journal.pone.0149075

Figure Lengend Snippet: Static binding assay on bEnd.3 cells or αvβ3-negative MCF-7 cells was performed with and without pre incubation with anti-alpha(v) antibodies (×200). (a) Non-targeted control MBs (1×10 8 particles/mL). (b) 1×10 6 , (c) 1×10 7 , (d) 1×10 8 , and (e) 1×10 9 particles/mL RGD-MBs. (f) RGD-MBs (1×10 8 particles/mL) incubated with αvβ3-negative MCF-7 cells. (g-i) RGD-MBs (1×10 8 particles/mL) with (g)1 ug/ml, (h) 5 ug/ml, and (i) 25 ug/ml anti-alpha(v) antibodies. (j) Quantitative analysis of the number of attached bubbles per field; (k) Quantitative analysis of the number of attached bubbles per cell. (* P < 0.05, ** P < 0.01).

Article Snippet: To visualize ανβ3 integrin expression on tumor endothelial cells, a double immunostaining procedure was performed using a rabbit anti-mouse alpha(v) primary antibody (BD Biosciences, San Jose, CA) at a 1:50 dilution and a FITC-labeled goat anti-rabbit secondary antibody (BD Biosciences, San Jose, CA) at a 1:100 dilution.

Techniques: Binding Assay, Incubation

Journal: Gastroenterology Research and Practice

Article Title: Differential Effects of Three Techniques for Hepatic Vascular Exclusion during Resection for Liver Cirrhosis on Hepatic Ischemia-Reperfusion Injury in Rats

doi: 10.1155/2018/5309286

Figure Lengend Snippet: Effects of different techniques for hepatic vascular exclusion during resection on TRIF expression after hepatic I/R in rats. TRIF protein expression was determined by immunohistochemistry at different time points (1, 3, 6, and 24 h) after reperfusion in the PR, HH, and WH groups. (a) Representative images of immuno histochemical staining of TRIF in the liver sections from rats in the PR, HH, and WH groups; (b) quantification of IRIF expression in the liver of rats in the PR, HH, and WH groups. The TRIF protein level was significantly higher in the PR group than in the HH and WH groups ( P < 0.05), while the difference between HH group and WH group was not statistically significant.

Article Snippet: After washing with phosphate-buffered saline (PBS; 0.01 M, pH 7.5), the slides were incubated with rabbit anti-mouse polyclonal TRIF primary antibody (ZSGB-Bio, Beijing, China) overnight at 4°C and rinsed with PBS (0.01 M, pH 7.5) at room temperature (RT).

Techniques: Expressing, Immunohistochemistry, Staining

Journal: American Journal of Physiology - Renal Physiology

Article Title: Isoflurane activates intestinal sphingosine kinase to protect against bilateral nephrectomy-induced liver and intestine dysfunction

doi: 10.1152/ajprenal.00467.2010

Figure Lengend Snippet: Primers used to amplify mRNAs encoding SK1, SK2, GAPDH, and proinflammatory cytokines based on published GenBank sequences for mice

Article Snippet: The slides were stained for SK1 in sequential incubations with rabbit anti-mouse SK1 primary antibody (AP7237, 1:50 dilution; Abgent, San Diego, CA) overnight at 4°C, horseradish peroxidase-conjugated goat anti-rabbit IgG (PI-1000, 1:200 dilution, Vector Laboratories, Burlingame, CA) for 1 h at room temperature, and diaminobenzidine reagent (Vector Laboratories) for 2 min. A rabbit IgG (I-1000, Vector Laboratories) was used at the same concentration as the primary antibody as a negative isotype control for all experiments.

Techniques: Sequencing, Variant Assay

Journal: American Journal of Physiology - Renal Physiology

Article Title: Isoflurane activates intestinal sphingosine kinase to protect against bilateral nephrectomy-induced liver and intestine dysfunction

doi: 10.1152/ajprenal.00467.2010

Figure Lengend Snippet: Plasma alanine aminotransferase (ALT; U/l) after bilateral nephrectomy. A: plasma ALT levels were measured in C57BL/6 mice exposed to 4 h of pentobarbital sodium (PB) or 1.2% isoflurane (Iso) after sham operation or bilateral nephrectomy (BNx); n = 4 for sham groups, n = 8 for BNx groups. B: plasma ALT from C57BL/6 mice treated with the sphingosine kinase (SK) inhibitor SKI-II or from SK1 knockout (SK1KO) mice after sham operation (PB anesthesia) or BNx followed by exposure to 4 h of pentobarbital sodium or isoflurane; n = 4–6/group. In all cases (A and B), plasma ALT was measured 5 h after sham operation or bilateral nephrectomy. Values are means ± SE. #P < 0.05 vs. PB sham mice. *P < 0.05 vs. PB BNx group.

Article Snippet: The slides were stained for SK1 in sequential incubations with rabbit anti-mouse SK1 primary antibody (AP7237, 1:50 dilution; Abgent, San Diego, CA) overnight at 4°C, horseradish peroxidase-conjugated goat anti-rabbit IgG (PI-1000, 1:200 dilution, Vector Laboratories, Burlingame, CA) for 1 h at room temperature, and diaminobenzidine reagent (Vector Laboratories) for 2 min. A rabbit IgG (I-1000, Vector Laboratories) was used at the same concentration as the primary antibody as a negative isotype control for all experiments.

Techniques: Knock-Out

Journal: American Journal of Physiology - Renal Physiology

Article Title: Isoflurane activates intestinal sphingosine kinase to protect against bilateral nephrectomy-induced liver and intestine dysfunction

doi: 10.1152/ajprenal.00467.2010

Figure Lengend Snippet: Isoflurane activates intestinal SK1, but not SK2, mRNA expression. A: representative gel images of RT-PCR (of 4 experiments) of SK1, SK2, and GAPDH from the small intestines of sham mice exposed to 4 h of pentobarbital sodium or 1.2% isoflurane. B: densitometric quantifications of band intensities relative to GAPDH from RT-PCR reactions. *P < 0.05 vs. pentobarbital sodium group.

Article Snippet: The slides were stained for SK1 in sequential incubations with rabbit anti-mouse SK1 primary antibody (AP7237, 1:50 dilution; Abgent, San Diego, CA) overnight at 4°C, horseradish peroxidase-conjugated goat anti-rabbit IgG (PI-1000, 1:200 dilution, Vector Laboratories, Burlingame, CA) for 1 h at room temperature, and diaminobenzidine reagent (Vector Laboratories) for 2 min. A rabbit IgG (I-1000, Vector Laboratories) was used at the same concentration as the primary antibody as a negative isotype control for all experiments.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction

Journal: American Journal of Physiology - Renal Physiology

Article Title: Isoflurane activates intestinal sphingosine kinase to protect against bilateral nephrectomy-induced liver and intestine dysfunction

doi: 10.1152/ajprenal.00467.2010

Figure Lengend Snippet: Isoflurane increases intestinal SK1, but not SK2, protein expression. A: representative immunoblot images (of 4 experiments) of SK1, SK2, and β-actin from the small intestines of sham mice exposed to 4 h of pentobarbital sodium or 1.2% isoflurane. B: densitometric quantifications of band intensities relative to β-actin from immunoblot images. *P < 0.05 vs. pentobarbital sodium group.

Article Snippet: The slides were stained for SK1 in sequential incubations with rabbit anti-mouse SK1 primary antibody (AP7237, 1:50 dilution; Abgent, San Diego, CA) overnight at 4°C, horseradish peroxidase-conjugated goat anti-rabbit IgG (PI-1000, 1:200 dilution, Vector Laboratories, Burlingame, CA) for 1 h at room temperature, and diaminobenzidine reagent (Vector Laboratories) for 2 min. A rabbit IgG (I-1000, Vector Laboratories) was used at the same concentration as the primary antibody as a negative isotype control for all experiments.

Techniques: Expressing, Western Blot

Journal: American Journal of Physiology - Renal Physiology

Article Title: Isoflurane activates intestinal sphingosine kinase to protect against bilateral nephrectomy-induced liver and intestine dysfunction

doi: 10.1152/ajprenal.00467.2010

Figure Lengend Snippet: Isoflurane increases SK1 in small intestinal crypts. Shown are representative immunohistochemistry images (of 4 experiments) for SK1 (diaminobenzidine stain, dark brown) from sham mice exposed to 4 h of pentobarbital sodium (A) or 1.2% isoflurane (C). Small intestinal crypts are magnified (boxed area in A and C) after pentobarbital sodium (B) or isoflurane (D) exposure. SM designates smooth muscle layer. Arrows highlight areas of small intestinal crypts with increased staining for SK1.

Article Snippet: The slides were stained for SK1 in sequential incubations with rabbit anti-mouse SK1 primary antibody (AP7237, 1:50 dilution; Abgent, San Diego, CA) overnight at 4°C, horseradish peroxidase-conjugated goat anti-rabbit IgG (PI-1000, 1:200 dilution, Vector Laboratories, Burlingame, CA) for 1 h at room temperature, and diaminobenzidine reagent (Vector Laboratories) for 2 min. A rabbit IgG (I-1000, Vector Laboratories) was used at the same concentration as the primary antibody as a negative isotype control for all experiments.

Techniques: Immunohistochemistry, Staining

Journal: American Journal of Physiology - Renal Physiology

Article Title: Isoflurane activates intestinal sphingosine kinase to protect against bilateral nephrectomy-induced liver and intestine dysfunction

doi: 10.1152/ajprenal.00467.2010

Figure Lengend Snippet: Isoflurane exposure increases SK1 activity and SK1-phosphate (S1P) formation. A: relative SK1 activity (fold-over pentobarbital sodium group) from the intestines of sham mice exposed to 4 h of pentobarbital sodium or 1.2% isoflurane (n = 6/group). B: formation of S1P (fold-over pentobarbital sodium group) from the intestines of sham mice exposed to 4 h of pentobarbital sodium or 1.2% isoflurane (n = 4/group). *P < 0.05 vs. pentobarbital sodium group.

Article Snippet: The slides were stained for SK1 in sequential incubations with rabbit anti-mouse SK1 primary antibody (AP7237, 1:50 dilution; Abgent, San Diego, CA) overnight at 4°C, horseradish peroxidase-conjugated goat anti-rabbit IgG (PI-1000, 1:200 dilution, Vector Laboratories, Burlingame, CA) for 1 h at room temperature, and diaminobenzidine reagent (Vector Laboratories) for 2 min. A rabbit IgG (I-1000, Vector Laboratories) was used at the same concentration as the primary antibody as a negative isotype control for all experiments.

Techniques: Activity Assay