Journal: Cell Biochemistry and Biophysics

Article Title: Interaction of L-SIGN with Hepatitis C Virus Envelope Protein E2 Up-Regulates Raf–MEK–ERK Pathway

doi: 10.1007/s12013-012-9505-4

Figure Lengend Snippet: a Expression of L-SIGN on NIH3T3 and NIH3T3/L-SIGN. Cells were incubated with mouse anti-L-SIGN mAb ( thick lines ) or isotype control ( dotted lines ). Expression of L-SIGN was detected by flow cytometry using FITC-conjugated goat anti-mouse IgG. The results are representative of three experiments. b Confocal microscopy was carried out to characterize cellular localization of L-SIGN on NIH3T3 and NIH3T3/L-SIGN. Cells were stained with mouse anti-L-SIGN mAb and FITC-conjugated goat anti-mouse IgG. The results are reproducible in two experiments, and representative fields are shown

Article Snippet: Horseradish peroxidase-conjugated goat anti-rabbit IgG and alkaline phosphatase-conjugated goat anti-rabbit or anti-mouse IgG were from Vector Lab (Burlingame, CA, USA).

Techniques: Expressing, Incubation, Flow Cytometry, Confocal Microscopy, Staining

Journal: Cell Biochemistry and Biophysics

Article Title: Interaction of L-SIGN with Hepatitis C Virus Envelope Protein E2 Up-Regulates Raf–MEK–ERK Pathway

doi: 10.1007/s12013-012-9505-4

Figure Lengend Snippet: a Binding of HCV E2 protein to NIH3T3/L-SIGN. Cells were treated with E2 ( thick lines ) or left untreated ( dotted lines ) and the E2 binding was detected with mouse anti-E2 mAb and FITC-conjugated goat anti-mouse IgG by flow cytometry. b Inhibition of HCV E2 binding to NIH3T3/L-SIGN by antibodies against L-SIGN or DC-SIGN. Cells were incubated with mouse anti-L-SIGN mAb at a concentration of 4 μg/ml (II), 10 μg/ml (III) or 10 μg/ml anti-DC-SIGN mAb, and 10 μg/ml anti-L-SIGN mAb (IV) before the E2 incubation. The E2 binding was detected with goat anti-E2 Ab and FITC-conjugated rabbit anti-goat IgG in the presence or absence (I) of the antibody incubation. The percentage of marker-positive cells is indicated in each case. Data are representative of three experiments

Article Snippet: Horseradish peroxidase-conjugated goat anti-rabbit IgG and alkaline phosphatase-conjugated goat anti-rabbit or anti-mouse IgG were from Vector Lab (Burlingame, CA, USA).

Techniques: Binding Assay, Flow Cytometry, Inhibition, Incubation, Concentration Assay, Marker

Journal: Asian Journal of Chemistry

Article Title: Adsorption Behaviour of Reduced Graphene Oxide for Removal of Heavy Metal Ions

doi: 10.14233/ajchem.2014.17024

Figure Lengend Snippet: Fig. 1. (a) FT-IR spectra of graphene oxide and reduced graphene oxide, (b) Raman spectra of graphite, graphene oxide and reduced graphene oxide, (c) TGA curves of graphene oxide and reduced graphene oxide and (d) AFM image of reduced graphene oxide

Article Snippet: Raman spectra were obtained using a confocal microprobe Raman system (Renishaw, RM 2000).

Techniques:

Journal: Respiratory Research

Article Title: RAGE inhibition alleviates lipopolysaccharides-induced lung injury via directly suppressing autophagic apoptosis of type II alveolar epithelial cells

doi: 10.1186/s12931-023-02332-6

Figure Lengend Snippet: LPS-induced autophagic death in A549 cells depended on RAGE activation. A Treated A549 cells with siRAGE and detected the RAGE mRNA expression. The expression of Ager genes was decreased by 50%. B Detected the immunoblots of RAGE as well as Beclin1, LC3 II/I and cleaved Caspase 3 in A549 cells, each experiment was repeated more than three times. C The cell survival rate of A549 cells in response to siRAGE pre-treatment followed by LPS stimulation. Data were obtained from CCK-8 experiments and performed more than three times. D Immunofluorescence of cleaved Caspase3 to detect the apoptosis of A549 cells, spinning disk confocal microscopy at × 40 magnification (scale bar 50 μm). E Detected the immunoblots of STAT3 and phosphorylation STAT3 (p-STAT3) in A549 cells, each experiment was repeated more than three times. *Indicates the significant difference compared with the control group. # Indicates the significant difference compared with LPS group. P < 0.05, differences in characteristics between groups were analyzed using the Kruskal–Wallis test with Dunn’s post hoc tests

Article Snippet: Anti-RAGE (1:1000, #6996S; Cell Signaling Technology, Inc., MA, USA), anti-STAT3 (1:1000, #12640; Cell Signaling Technology, Inc., MA, USA), anti-phosphorylated STAT3 (1:2000, #9145; Cell Signaling Technology, Inc., MA, USA), anti-cleaved caspase 3 (1:1000, #9664; Cell Signaling Technology, Inc., MA, USA), anti-LC3II/I (1:1000, #4108; Cell Signaling Technology, Inc., MA, USA), anti-Beclin1 (1:1000, #3495; Cell Signaling Technology, Inc., MA, USA) and anti-GADPH (1:1000, #2118; Cell Signaling Technology, Inc., MA, USA) were used as primary antibodies.

Techniques: Activation Assay, Expressing, Western Blot, CCK-8 Assay, Immunofluorescence, Confocal Microscopy, Phospho-proteomics, Control

Journal: International Journal of Biological Sciences

Article Title: Metformin Alleviates Liver Metabolic Dysfunction in Polycystic Ovary Syndrome by Activating the Ethe1/Keap1/PINK1 Pathway

doi: 10.7150/ijbs.104778

Figure Lengend Snippet: Metformin improves mitochondrial function in the liver of PAMH-induced PCOS mice. Malondialdehyde (MDA) (A) , superoxide dismutase (SOD) (B) , glutathione (GSH) (C) , GSH/GSSG (D) , ATP (E) , and mitochondrial DNA (mtDNA) (F) in liver tissues from the control, PAMH-induced, and metformin treatment groups. (G, H) Reactive oxygen species levels in liver tissues labeled with dihydroethidium (DHE) as observed under a fluorescence microscope (G) . Mean fluorescence intensity of DHE in each group (H) . (I-M) Mitochondrial respiratory chain complexes: Complex I (NADH dehydrogenase) (I) , Complex II (succinate-coenzyme Q reductase) (J) , Complex III (ubiquinol-cytochrome c oxidoreductase) (K) , Complex IV (cytochrome C oxidase) (L) , and Complex V (ATP synthase) (M) . (N-Q, S, V) Markers related to mitochondrial autophagy (Mfn2, DRP1, LC3, P62, TOMM20, VDAC1, Nrf2, Keap1, PINK1, and Parkin) in total protein extraction from liver tissue based on western blotting (N, P) . Protein relative expression levels were calculated in the control group (orange bars), PAMH group (pink bars), and PAMH+Met group (purple bars) and also shown (O, Q, S, V) . (R, T, U) Expression of p62 and LC3 in each group. All error bars are mean values ± SD, p- values were determined by unpaired two-tailed Student's t test ( n = 3) in independent biological experiments. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.

Article Snippet: The PVDF membranes were incubated with anti-Ethe1 (1:2000, 27786-1-AP, Proteintech), anti-ETFDH polyclonal (1:2000, 11109-1-AP, Proteintech), NRF2,NFE2L2 polyclonal antibody (1:5000, 16396-1-AP, Proteintech), anti-Keap1 (1:5000, 10503-2-AP, Proteintech), PARK2/Parkin polyclonal antibody (1:2000, 14060-1-AP, Proteintech), PINK1 polyclonal antibody (1:500, 23274-1-AP, Proteintech), MFN2 polyclonal antibody (1:5000, 12186-1-AP, Proteintech), DRP1 (C-terminal) polyclonal antibody (1:2000, 12957-1-AP, Proteintech), anti-TOMM20 antibody (1:2000, ab186735, Abcam), VDAC1/Porin polyclonal antibody (1:2000, 55259-1-AP, Proteintech), P62, SQSTM1 polyclonal antibody (1:10000, 18420-1-AP, Proteintech), anti-LC3B antibody (1:2000, ab192890, Abcam), anti-βactin (1:5000; AC026, Abclonal), COXIV polyclonal antibody (1:5000, 11242-1-AP, Proteintech), and Histone H3 polyclonal antibody (1:5000, 17168-1-AP, Proteintech) at 25 °C for 2 h. After incubation with the corresponding secondary antibody (horseradish peroxidase-conjugated goat anti-rabbit (1:5000, ZB2301, Zsbio) and horseradish peroxidase-conjugated goat anti-mouse (1:5000, ZB2305, Zsbio)) for 1 h at 20-25 °C, the blots were visualized using SuperSignal-enhanced chemiluminescent substrate solution (WBULS0100, Millipore).

Techniques: Control, Labeling, Fluorescence, Microscopy, Protein Extraction, Western Blot, Expressing, Two Tailed Test

Journal: International Journal of Biological Sciences

Article Title: Metformin Alleviates Liver Metabolic Dysfunction in Polycystic Ovary Syndrome by Activating the Ethe1/Keap1/PINK1 Pathway

doi: 10.7150/ijbs.104778

Figure Lengend Snippet: Metformin improves mitophagy in damaged hepatocytes. (A, B) Intracellular ATP levels of control (orange bars), DHEA- and FFA-induced (pink bars), and Met-treated (purple bars) cells. (C, D, G, H) Expression of Ethe1 and key molecules related to mitochondrial autophagy (Mfn2, DRP1, PINK1, Parkin, and TOMM20) in each group (colors as described for A and B ). (F, F) MtDNA copy number for each group. (I, K, L) Co-localization of mitochondria with LC3 and Mito-Tracker Red, PINK1, and Mito-Tracker Red under confocal microscopy in the control, DHEA- and FFA-induced, and Met-added cells. (I) Mander's co-localization coefficients (K, L) . (J) Mean fluorescence intensity of JC-1 in each group. All error bars are mean values ± SD, p- values were determined by unpaired two-tailed Student's t test ( n = 3) in independent biological experiments. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.

Article Snippet: The PVDF membranes were incubated with anti-Ethe1 (1:2000, 27786-1-AP, Proteintech), anti-ETFDH polyclonal (1:2000, 11109-1-AP, Proteintech), NRF2,NFE2L2 polyclonal antibody (1:5000, 16396-1-AP, Proteintech), anti-Keap1 (1:5000, 10503-2-AP, Proteintech), PARK2/Parkin polyclonal antibody (1:2000, 14060-1-AP, Proteintech), PINK1 polyclonal antibody (1:500, 23274-1-AP, Proteintech), MFN2 polyclonal antibody (1:5000, 12186-1-AP, Proteintech), DRP1 (C-terminal) polyclonal antibody (1:2000, 12957-1-AP, Proteintech), anti-TOMM20 antibody (1:2000, ab186735, Abcam), VDAC1/Porin polyclonal antibody (1:2000, 55259-1-AP, Proteintech), P62, SQSTM1 polyclonal antibody (1:10000, 18420-1-AP, Proteintech), anti-LC3B antibody (1:2000, ab192890, Abcam), anti-βactin (1:5000; AC026, Abclonal), COXIV polyclonal antibody (1:5000, 11242-1-AP, Proteintech), and Histone H3 polyclonal antibody (1:5000, 17168-1-AP, Proteintech) at 25 °C for 2 h. After incubation with the corresponding secondary antibody (horseradish peroxidase-conjugated goat anti-rabbit (1:5000, ZB2301, Zsbio) and horseradish peroxidase-conjugated goat anti-mouse (1:5000, ZB2305, Zsbio)) for 1 h at 20-25 °C, the blots were visualized using SuperSignal-enhanced chemiluminescent substrate solution (WBULS0100, Millipore).

Techniques: Control, Expressing, Confocal Microscopy, Fluorescence, Two Tailed Test

Journal: International Journal of Biological Sciences

Article Title: Metformin Alleviates Liver Metabolic Dysfunction in Polycystic Ovary Syndrome by Activating the Ethe1/Keap1/PINK1 Pathway

doi: 10.7150/ijbs.104778

Figure Lengend Snippet: Metformin alleviates FFA- and DHEA-induced hepatocyte injury by regulating the Ethe1/Keap1/PINK1 pathway through ETFDH. (A, B) Expression of Keap1 in cells and Nrf2 in the nucleus and cytoplasm of control, DHEA-induced, and Met-treated cells (A) . Gray value levels (B) . (C-D) Expression of Keap1 in cells and Nrf2 in the nucleus and cytoplasm from the DHEA-induced, si-Ethe1, and Met-treated cells (C) . Relative gray value levels (D) . (F) Experimental design flowchart for Nrf2 and PINK1 functions. (E, G, H) CO-IP experiments verified the interaction between Ethe1 and ETFDH as well as the interaction between Ethe1 and Keap1 (E) in control group (orange bars), DHEA group (pink bars), and DHEA+Met group (purple group) (G, H) . (I, J) TG concentrations (I) . Representative Oil Red O staining imagines for each group (J) . (K, L-P) Each group of cells was labeled with Mitosox TM Red, DCFH-DA, and JC-1 probes, and the co-localization of LC3 and PINK1 with Mito-Tracker Red in cells from different groups was observed under a confocal microscope (K) . Mean fluorescence intensity of Mitosox TM Red, DCFH-DA, and JC-1 (L-N) . Mander's co-localization coefficients (O, P) . (Q-T) Expression of molecules related to mitochondrial autophagy (LC3, DRP1, PINK1, Parkin, and TOMM20) (Q) in the DHEA/FFAs+Met+si NC group (orange bars), DHEA/FFAs+Met+si Nrf2 group (pink bars), and DHEA/FFAs+Met+si PINK1 group (purple bars) (R-T) . All error bars are mean values ± SD, p- values were determined by unpaired two-tailed Student's t test ( n = 3) in independent biological experiments. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.

Article Snippet: The PVDF membranes were incubated with anti-Ethe1 (1:2000, 27786-1-AP, Proteintech), anti-ETFDH polyclonal (1:2000, 11109-1-AP, Proteintech), NRF2,NFE2L2 polyclonal antibody (1:5000, 16396-1-AP, Proteintech), anti-Keap1 (1:5000, 10503-2-AP, Proteintech), PARK2/Parkin polyclonal antibody (1:2000, 14060-1-AP, Proteintech), PINK1 polyclonal antibody (1:500, 23274-1-AP, Proteintech), MFN2 polyclonal antibody (1:5000, 12186-1-AP, Proteintech), DRP1 (C-terminal) polyclonal antibody (1:2000, 12957-1-AP, Proteintech), anti-TOMM20 antibody (1:2000, ab186735, Abcam), VDAC1/Porin polyclonal antibody (1:2000, 55259-1-AP, Proteintech), P62, SQSTM1 polyclonal antibody (1:10000, 18420-1-AP, Proteintech), anti-LC3B antibody (1:2000, ab192890, Abcam), anti-βactin (1:5000; AC026, Abclonal), COXIV polyclonal antibody (1:5000, 11242-1-AP, Proteintech), and Histone H3 polyclonal antibody (1:5000, 17168-1-AP, Proteintech) at 25 °C for 2 h. After incubation with the corresponding secondary antibody (horseradish peroxidase-conjugated goat anti-rabbit (1:5000, ZB2301, Zsbio) and horseradish peroxidase-conjugated goat anti-mouse (1:5000, ZB2305, Zsbio)) for 1 h at 20-25 °C, the blots were visualized using SuperSignal-enhanced chemiluminescent substrate solution (WBULS0100, Millipore).

Techniques: Expressing, Control, Co-Immunoprecipitation Assay, Staining, Labeling, Microscopy, Fluorescence, Two Tailed Test