Journal: Molecular Medicine Reports

Article Title: Establishment and evaluation of a simulated high-altitude hypoxic brain injury model in SD rats

doi: 10.3892/mmr.2019.9939

Figure Lengend Snippet: Study design of the experimental procedure. SD, Sprague-Dawley; MDA, malondialdehyde; LDH, lactate dehydrogenase; SOD, superoxide dismutase; T-GSH, total glutathione; ELISA, enzyme-linked immunosorbent assay; H&E, hematoxylin and eosin; IHC, immunohistochemistry; HIF-1α, hypoxia inducible factor-1α.

Article Snippet: The total extraction sample kit (cat. no. AR0101-30), BCA protein assay kit (cat. no. AR0146), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) protein sample buffer 2X (denaturation; cat. no. AR0131), broad spectrum protease inhibitor (cat. no. AR1182-1), broad spectrum phosphatase inhibitor (cat. no. AR1183), terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) apoptosis detection kit I-POD (cat. no. MK1025) and primary antibodies against Bcl-2-associated X protein (Bax; cat. no. BA0315), apoptotic protease activating factor-1 (Apaf-1; cat. no. BA2373), hypoxia inducible factor (HIF)-1α (cat. no. PB0245) and cytochrome c (cyto-c; cat. no. A03529) were obtained from Wuhan Boster Biological Technology, Ltd. (Wuhan, China).

Techniques: Enzyme-linked Immunosorbent Assay, Immunohistochemistry

Journal: Molecular Medicine Reports

Article Title: Establishment and evaluation of a simulated high-altitude hypoxic brain injury model in SD rats

doi: 10.3892/mmr.2019.9939

Figure Lengend Snippet: Immunohistochemistry for caspase-3 and HIF-1α protein expression and Nissl's and TUNEL staining in the CA1 region in the hippocampus following hypoxia. (A) Nissl-stained neurons in the CA1 region of the hippocampus of SD rats. (B) TUNEL-positive apoptotic cells in the CA1 region of the hippocampus of SD rats. (C) Caspase-3 expression examined using immunohistochemical analysis and quantified. (D) HIF-1α expression examined using immunohistochemical analysis and quantified. Scale bar, 50 µm. Original magnification, ×200. The values are presented as the mean ± standard deviation. **P<0.01 with comparisons shown by lines. HIF-1α, hypoxia inducible factor-1α; TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling; SD, Sprague-Dawley; IOD, integrated optical density.

Article Snippet: The total extraction sample kit (cat. no. AR0101-30), BCA protein assay kit (cat. no. AR0146), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) protein sample buffer 2X (denaturation; cat. no. AR0131), broad spectrum protease inhibitor (cat. no. AR1182-1), broad spectrum phosphatase inhibitor (cat. no. AR1183), terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) apoptosis detection kit I-POD (cat. no. MK1025) and primary antibodies against Bcl-2-associated X protein (Bax; cat. no. BA0315), apoptotic protease activating factor-1 (Apaf-1; cat. no. BA2373), hypoxia inducible factor (HIF)-1α (cat. no. PB0245) and cytochrome c (cyto-c; cat. no. A03529) were obtained from Wuhan Boster Biological Technology, Ltd. (Wuhan, China).

Techniques: Immunohistochemistry, Expressing, TUNEL Assay, Staining, Immunohistochemical staining, Standard Deviation

Journal: Molecular Medicine Reports

Article Title: Establishment and evaluation of a simulated high-altitude hypoxic brain injury model in SD rats

doi: 10.3892/mmr.2019.9939

Figure Lengend Snippet: Hypoxia-activated mitochondrial apoptosis pathway. Western blot analysis for (A) caspase-3 and cleaved caspase-3, (B) Apaf-1, HIF-1α, Bax and cyto-c protein level in each group, n=6. (C) Quantified western blot analysis results. The values are presented as mean ± standard deviation. **P<0.01 with comparisons shown by lines. Apaf-1, apoptotic protease activating factor-1; HIF-1α, hypoxia inducible factor-1α; Bax, Bcl-2-associated X protein; cyto-c, cytochrome c.

Article Snippet: The total extraction sample kit (cat. no. AR0101-30), BCA protein assay kit (cat. no. AR0146), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) protein sample buffer 2X (denaturation; cat. no. AR0131), broad spectrum protease inhibitor (cat. no. AR1182-1), broad spectrum phosphatase inhibitor (cat. no. AR1183), terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) apoptosis detection kit I-POD (cat. no. MK1025) and primary antibodies against Bcl-2-associated X protein (Bax; cat. no. BA0315), apoptotic protease activating factor-1 (Apaf-1; cat. no. BA2373), hypoxia inducible factor (HIF)-1α (cat. no. PB0245) and cytochrome c (cyto-c; cat. no. A03529) were obtained from Wuhan Boster Biological Technology, Ltd. (Wuhan, China).

Techniques: Western Blot, Standard Deviation

Journal: Molecular Medicine Reports

Article Title: Establishment and evaluation of a simulated high-altitude hypoxic brain injury model in SD rats

doi: 10.3892/mmr.2019.9939

Figure Lengend Snippet: Schematic view of HIF-1α and the intrinsic triggering pathway of the apoptotic signaling cascade. Hypoxia stimulates the HIF-1α and mitochondrial apoptotic signaling pathway and triggers the release of cyto-c from the mitochondrial matrix to the cytoplasm, thereby binding to the Apaf-1 protein, inducing its conformational change and activating it. Thus, the apoptosome consisting of cyto-c/Apaf-1/caspase-9 is yielded and is able to cleave or activate the executioner caspase-3. The activated caspase-3 complex then results in apoptosis. C9, caspase-9; C3, caspase-3; HIF-1α, hypoxia inducible factor-1α; cyto-c, cytochrome c; Apaf-1, apoptotic protease activating factor-1; CARD, caspase recruitment domain containing; Bak, Bcl2 antagonist/killer; Bax, Bcl-2-associated X protein; Bcl-2, Bcl2, apoptosis regulator; CBP, CREB binding protein; HIF-1β, hypoxia inducible factor-1β; HNF4, hepatocyte nuclear factor 4α.

Article Snippet: The total extraction sample kit (cat. no. AR0101-30), BCA protein assay kit (cat. no. AR0146), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) protein sample buffer 2X (denaturation; cat. no. AR0131), broad spectrum protease inhibitor (cat. no. AR1182-1), broad spectrum phosphatase inhibitor (cat. no. AR1183), terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) apoptosis detection kit I-POD (cat. no. MK1025) and primary antibodies against Bcl-2-associated X protein (Bax; cat. no. BA0315), apoptotic protease activating factor-1 (Apaf-1; cat. no. BA2373), hypoxia inducible factor (HIF)-1α (cat. no. PB0245) and cytochrome c (cyto-c; cat. no. A03529) were obtained from Wuhan Boster Biological Technology, Ltd. (Wuhan, China).

Techniques: Binding Assay

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Dishevelled Associated Activator Of Morphogenesis (DAAM) Facilitates Invasion of Hepatocellular Carcinoma by Upregulating Hypoxia-Inducible Factor 1α (HIF-1α) Expression

doi: 10.12659/MSM.924670

Figure Lengend Snippet: DAAM2 decreased VHL and up-regulated HIF-1α in a ubiquitin-dependent pathway. ( A ) Expressions of DAAM1 and DAAM2 in different HCC cell lines. ( B ) Cell proliferation ( left ) and invasion ( right ) were detected with MTT assay and transwell assay, respectively, after silencing DAAM1 or DAAM2 expression by siRNA. DAAM1 was silenced in HepG2 while DAAM2 was knocked down in HuH7. N.s. represented not significant and ** meant P<0.01 compared with the control group. ( C ) Representative images of HuH7 cells transfected with scrambled siRNA, siDAAM2-1 and siDAAM2-2 in transwell assay. ( D ) Expression of VHL and HIF-1α was detected with Western blot 48 hr after knocking down DAAM1 or DAAM2. DAAM1 was silenced in HepG2 while DAAM2 was knocked down in HuH7 ( E ). In HuH7 and HepG2, expression of VHL and HIF-1α was detected 48 hr after transfection of empty vector or plasmid carrying DAAM2, in the presence or absence of 10 μM G132 for 6 hr. ( F ) In the 117 patients with HCC, patients with high DAAM2 expression had lower VHL IHC score and higher HIF-1α IHC score.

Article Snippet: Primary antibodies were as follows: DAAM1 (Cat. 71499, Abcam, Cambridge, Massachusetts, United States), DAAM2 (Cat. NBP2–47496, Novus Biologicals, Centennial, Colorado, United States), VHL (Cat. 556347, BD Pharmingem), Hypoxia-inducible factor 1α (HIF-1α) (Cat. NB100-105, Novus), and β-actin (Cat. sc-8432, Santa Cruz Biotechnology, Santa Cruz, California, United States).

Techniques: MTT Assay, Transwell Assay, Expressing, Transfection, Western Blot, Plasmid Preparation

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Dishevelled Associated Activator Of Morphogenesis (DAAM) Facilitates Invasion of Hepatocellular Carcinoma by Upregulating Hypoxia-Inducible Factor 1α (HIF-1α) Expression

doi: 10.12659/MSM.924670

Figure Lengend Snippet: HIF-1α was required in DAAM2-induced invasion of HCC. ( A ) MG132 (10μM) incubation for 6 hr had no influence on proliferation of HuH7. Proliferation was assessed by MTT assay. ( B ) Invasion of HuH7 was detected after overexpressing DAAM2. 10 μM MG132 or 10 μM HIF-1α inhibitor BAY87-2243 was used to incubate HuH7. ** Represented P<0.01 compared with control group, and ## represented P<0.01 between indicated groups. Analyzed data were from three independent experiments. ( C ) Representative images in transwell assay of B . ( D ) Western blot showed that HIF-1α was knocked down while DAAM2 was overexpressed in HuH7. ( E ) HIF-1α knockdown extensively decreased the DAAM2-induced invasion. The invasion was detected with transwell assay. ## Represented P<0.01 between indicated groups. ( F ) Representative images in transwell assay of ( E ).

Article Snippet: Primary antibodies were as follows: DAAM1 (Cat. 71499, Abcam, Cambridge, Massachusetts, United States), DAAM2 (Cat. NBP2–47496, Novus Biologicals, Centennial, Colorado, United States), VHL (Cat. 556347, BD Pharmingem), Hypoxia-inducible factor 1α (HIF-1α) (Cat. NB100-105, Novus), and β-actin (Cat. sc-8432, Santa Cruz Biotechnology, Santa Cruz, California, United States).

Techniques: Incubation, MTT Assay, Transwell Assay, Western Blot