Journal:

Article Title: Gene Expression in HL60 Granulocytoids and Human Polymorphonuclear Leukocytes Exposed to Candida albicans †

doi: 10.1128/IAI.72.1.414-429.2004

Figure Lengend Snippet: HL60 granulocytoid-C. albicans interaction. GFP-expressing C. albicans were cultured either alone (right column) or with HL60 granulocytoids at the indicated MOIs, as described in Materials and Methods. Photographs were taken 1.5 h later at 400× magnification. Each of the panels in the first column is a superimposition of three images: phase contrast, blue fluorescence (to visualize DAPI staining), and green fluorescence (to visualize GFP-C. albicans). Panels in the second column show the images corresponding to the green fluorescence in the first column. Panels in the last column show green fluorescence images to visualize C. albicans cultured alone at densities corresponding to the indicated MOIs.

Article Snippet: HL60, a human promyelocytic leukemia cell line, was obtained from the American Type Culture Collection and was maintained in RPMI 1640 (Gibco-Invitrogen, Burlington, Ontario, Canada) supplemented with 20% heat-inactivated fetal bovine serum (HyClone, Logan, Utah) (growth medium).

Techniques: Expressing, Cell Culture, Fluorescence, Staining

Journal:

Article Title: Gene Expression in HL60 Granulocytoids and Human Polymorphonuclear Leukocytes Exposed to Candida albicans †

doi: 10.1128/IAI.72.1.414-429.2004

Figure Lengend Snippet: C. albicans-induced mortality in the HL60 granulocytoid population. HL60 granulocytoids were cultured either alone (left column) or with GFP-expressing C. albicans at an MOI of 0.3 (right column) as described in Materials and Methods. Photographs were taken 1.5 h (A) and 6 h (B and C) later. In the first and second rows, images are superimpositions of three photographs: phase contrast, green fluorescence (to visualize GFP-C. albicans), and blue fluorescence (to visualize DAPI staining) at a magnification of 400×. GFP-Candida can be seen engulfed by HL60 granulocytoids at both 1.5 h and 6 h postinfection. The third row shows blue fluorescence images at a magnification of 200×. DAPI-stained blue cells are indicative of cell death. (C) Nuclear morphology visualized by DAPI staining. The arrow points to a typical example of nuclear condensation and the arrowhead points to an example of nuclear fragmentation. A portion of the 400× image was further magnified 4× digitally.

Article Snippet: HL60, a human promyelocytic leukemia cell line, was obtained from the American Type Culture Collection and was maintained in RPMI 1640 (Gibco-Invitrogen, Burlington, Ontario, Canada) supplemented with 20% heat-inactivated fetal bovine serum (HyClone, Logan, Utah) (growth medium).

Techniques: Cell Culture, Expressing, Fluorescence, Staining

Journal:

Article Title: Gene Expression in HL60 Granulocytoids and Human Polymorphonuclear Leukocytes Exposed to Candida albicans †

doi: 10.1128/IAI.72.1.414-429.2004

Figure Lengend Snippet: C. albicans hyphal growth in the presence and absence of dimethyl formamide-induced neutrophils. GFP-Candida were cultured with (dotted line) or without (solid line) HL60 granulocytoids in a Bioptechs ΔTC3 petri dish as described in Materials and Methods. Photographs were taken every 60 min with a 20× objective. Hyphal length was measured for all Candida cells in three microscopic fields for each time point. The number of Candida cells counted for each point on the graph is indicated. The figure shows the change in average hyphal length of C. albicans. The standard error is indicated at each time point.

Article Snippet: HL60, a human promyelocytic leukemia cell line, was obtained from the American Type Culture Collection and was maintained in RPMI 1640 (Gibco-Invitrogen, Burlington, Ontario, Canada) supplemented with 20% heat-inactivated fetal bovine serum (HyClone, Logan, Utah) (growth medium).

Techniques: Cell Culture

Journal:

Article Title: Gene Expression in HL60 Granulocytoids and Human Polymorphonuclear Leukocytes Exposed to Candida albicans †

doi: 10.1128/IAI.72.1.414-429.2004

Figure Lengend Snippet: Viability of HL60 granulocytoids exposed to C. albicans

Article Snippet: HL60, a human promyelocytic leukemia cell line, was obtained from the American Type Culture Collection and was maintained in RPMI 1640 (Gibco-Invitrogen, Burlington, Ontario, Canada) supplemented with 20% heat-inactivated fetal bovine serum (HyClone, Logan, Utah) (growth medium).

Techniques: Positive Control

Journal:

Article Title: Gene Expression in HL60 Granulocytoids and Human Polymorphonuclear Leukocytes Exposed to Candida albicans †

doi: 10.1128/IAI.72.1.414-429.2004

Figure Lengend Snippet: Candida colony formation after 5 h of coculture with HL60 or HL60 granulocytoids. C. albicans was cultured alone or with HL60 or HL60-derived granulocytoids for 5 h at the indicated MOIs as described in Materials and Methods. The figure shows percent killing by HL60 granulocytoids (solid bars) or undifferentiated HL60 (hatched bars). The results presented are the means of three independent experiments. Bars represent the standard error.

Article Snippet: HL60, a human promyelocytic leukemia cell line, was obtained from the American Type Culture Collection and was maintained in RPMI 1640 (Gibco-Invitrogen, Burlington, Ontario, Canada) supplemented with 20% heat-inactivated fetal bovine serum (HyClone, Logan, Utah) (growth medium).

Techniques: Cell Culture, Derivative Assay

Journal:

Article Title: Gene Expression in HL60 Granulocytoids and Human Polymorphonuclear Leukocytes Exposed to Candida albicans †

doi: 10.1128/IAI.72.1.414-429.2004

Figure Lengend Snippet: Quantitative RT-PCR analysis. Relative RNA levels of HNP1, N.E., HBEGF, and PAC1 measured by quantitative RT-PCR in RNA from HL60 granulocytoids or HL60 granulocytoids exposed to different MOIs (0.1, 0.5, and 5) of C. albicans for 1 h. RNA levels were measured relative to the amount of ACTB mRNA as described in Materials and Methods. Results are presented as the increase over expression in uninfected HL60 granulocytoids. Bars represent the standard error.

Article Snippet: HL60, a human promyelocytic leukemia cell line, was obtained from the American Type Culture Collection and was maintained in RPMI 1640 (Gibco-Invitrogen, Burlington, Ontario, Canada) supplemented with 20% heat-inactivated fetal bovine serum (HyClone, Logan, Utah) (growth medium).

Techniques: Quantitative RT-PCR, Over Expression

Journal:

Article Title: Gene Expression in HL60 Granulocytoids and Human Polymorphonuclear Leukocytes Exposed to Candida albicans †

doi: 10.1128/IAI.72.1.414-429.2004

Figure Lengend Snippet: Expression of cell surface markers on 4-day dimethyl formamide-treated HL60 cells. Expression of cell surface markers was detected by binding of fluorescently tagged antibodies recognizing the markers in question. The figure shows the number of cells binding antibody (y axis) and the intensity of fluorescence (x axis) determined by flow cytometry. Specific binding was determined by comparing the binding profile of anti-CD11b, anti-CD116, anti-CD14, anti-mannose receptor, and anti-CD16b (filled profile) to the binding profile of the corresponding isotypic negative controls (described in Materials and Methods).

Article Snippet: HL60, a human promyelocytic leukemia cell line, was obtained from the American Type Culture Collection and was maintained in RPMI 1640 (Gibco-Invitrogen, Burlington, Ontario, Canada) supplemented with 20% heat-inactivated fetal bovine serum (HyClone, Logan, Utah) (growth medium).

Techniques: Expressing, Binding Assay, Fluorescence, Flow Cytometry

Journal:

Article Title: Gene Expression in HL60 Granulocytoids and Human Polymorphonuclear Leukocytes Exposed to Candida albicans †

doi: 10.1128/IAI.72.1.414-429.2004

Figure Lengend Snippet: Scatter plot analysis of microarray data. (A) Comparison of the levels of expression of 7,000 genes in HL60 granulocytoids (HL60 granulocytoid single) with levels in pooled RNA from three extractions (HL60 granulocytoid pooled). (B) HL60 granulocytoids exposed to Candida at an MOI of 0.5 for 1 h (HL60 granulocytoid+candida single) with pooled RNA from independent extractions of neutrophils exposed to Candida at an MOI of 0.5:1 for 1 h (HL60 granulocytoid+candida pooled). (C) HL60 granulocytoid single compared to HL60 granulocytoid+candida single. (D) HL60 granulocytoid pooled compared to HL60 granulocytoid+candida pooled.

Article Snippet: HL60, a human promyelocytic leukemia cell line, was obtained from the American Type Culture Collection and was maintained in RPMI 1640 (Gibco-Invitrogen, Burlington, Ontario, Canada) supplemented with 20% heat-inactivated fetal bovine serum (HyClone, Logan, Utah) (growth medium).

Techniques: Microarray, Comparison, Expressing

Journal:

Article Title: Gene Expression in HL60 Granulocytoids and Human Polymorphonuclear Leukocytes Exposed to Candida albicans †

doi: 10.1128/IAI.72.1.414-429.2004

Figure Lengend Snippet: Graphic representation of RNA levels. Panel A shows genes whose expression levels were higher in HL60 granulocytoids exposed to C. albicans at an MOI of 0.1 than in HL60 granulocytoids exposed to C. albicans at an MOI of 5. Panel B shows genes whose expression levels remained unaffected in HL60 granulocytoids on exposure to C. albicans (internal controls). Panel C shows genes whose expression levels were lower in HL60 granulocytoids exposed to C. albicans at an MOI of 0.1 than in HL60 granulocytoids exposed to C. albicans at an MOI of 5.

Article Snippet: HL60, a human promyelocytic leukemia cell line, was obtained from the American Type Culture Collection and was maintained in RPMI 1640 (Gibco-Invitrogen, Burlington, Ontario, Canada) supplemented with 20% heat-inactivated fetal bovine serum (HyClone, Logan, Utah) (growth medium).

Techniques: Expressing

Journal: International Journal of Biological Sciences

Article Title: Mitochondrial Dysfunction by FADDosome Promotes Gastric Mucosal Injury in Portal Hypertensive Gastropathy

doi: 10.7150/ijbs.90835

Figure Lengend Snippet: NOX2 enhances FADDosome-induced gastric epithelial mitochondrial fission and dysfunction in PHG. (A) Two-dimensional hierarchical clustering results for the enzyme NADPH oxidase (NOX) family were presented for the comparison between PHG patients and healthy volunteers (n = 3 per group). The fold changes and P values of the indicated mRNAs in PHG tissues relative to those in normal (Uninvolved) tissues from the microarray experiment were also presented. (B) NOX1, NOX2 and NOX4 IHC staining (brown) showed that NOX2, but not NOX1 or NOX4, was upregulated in the gastric mucosal samples of PHG patients, and the NOX1, NOX2 and NOX4 areas were also analyzed. n = 6 per group. * P < 0.05. (C) Representative images of IHC staining (brown) for NOX1, NOX2 and NOX4 in mouse models revealed that Nec-1 (an inhibitor of RIPK1) repressed NOX2 expression in mice with PVL. n = 6 per group. (D) The NOX1, NOX2 and NOX4 areas detected by IHC staining in (C) were presented. n = 6 per group. * P < 0.05 versus SO mice; # P < 0.05 versus mice with PVL without Nec-1 treatment. (E) The colocalization of TOMM20 (red) and NOX2 (green) determined by confocal imaging in GES-1 cells confirmed that TNF-α increased the mitochondrial localization of NOX2 (white arrows indicate colocalization). Nuclei (blue) were counterstained with DAPI. (F) The levels of and interaction between NOX2 and TOMM20 were determined by western blotting, and IP showed that Nec-1 decreased the epithelial expression of total and mitochondrial NOX2 in the primary epithelial cells of mice with PVL. TEM also revealed that Nec-1 partly normalized the abnormal or damaged mitochondria in the mice with PVL. n = 6 per group. * P < 0.05 versus SO mice; # P < 0.05 versus mice with PVL without Nec-1 treatment. (G) The ATP concentration in the mouse models verified that Nec-1 and GSK2795039 (GSK, a NOX2 inhibitor) improved the production of ATP in mice with PVL. n = 6 per group. * P < 0.05. (H) Representative 4-HNE IHC staining (brown) and quantification analysis in SO mice and mice with PVL suggested that GSK2795039 (GSK) reduced the level of 4-HNE in mice with PVL. n = 6 per group. * P < 0.05 versus SO mice; # P < 0.05 versus mice with PVL without GSK2795039 treatment. (I) Representative MitoSox staining (red) and the MitoSox intensity in mouse model revealed that GSK2795039 alleviated mitochondrial ROS (mtROS) accumulation in the primary epithelial cells of mice with PVL. n = 6 in each group. * P < 0.05 versus SO mice; # P < 0.05 versus mice with PVL without GSK2795039 treatment. (J) Related protein levels in primary epithelial cells from mouse models treated with the indicated agents ( shFADD , Nec-1 or BAY) showed interactions among TAK1, NF-κBp65, FADD and NOX2 signaling. (K) GES-1 cells were transfected with either the control vector or shFADD and then treated with TNF-α or vehicle, and the NOX2 luciferase reporter activities were presented (left panel). * P < 0.05 versus the vehicle group and TNF-α-treated cells transfected with shFADD . The NOX2 luciferase reporter activities in GES-1 cells transfected with pcDNA3.1-p65 -vector or pcDNA3.1 -vector were also shown (right panel), * P < 0.05.

Article Snippet: In some experiments, the mice were treated for 2 weeks after the operation with the following reagents or drugs according to the experimental needs: for NF-κB inhibition, the mice were intraperitoneally injected with Bay11708 (BAY, Calbiochem, La Jolla, CA, USA) at 200 μg/per mouse daily for 2 weeks; for TNF-α inhibition, the mice were intraperitoneally injected with infliximab (IFX, Janssen Biotech, Horsham, PA, USA) at 5 mg/kg per day for 2 weeks; for RIPK1 inhibition, the mice were intraperitoneally injected with 2 mg/kg necrostatin-1 (Nec-1, HY-15760, MedChemExpress, NJ, USA) per day for 2 weeks; for above experiments of the inhibitors, the mice in the control group (vehicle group) were intraperitoneally injected with an equal volume of phosphate-buffered saline (PBS) daily for 2 weeks; for Drp1 inhibition, the mice were intraperitoneally injected with 20 mg/kg Mdivi-1 (dissolved in DMSO, HY-15886, MedChemExpress), and the control animals (vehicle) were injected with an equal volume of DMSO; for ROS scavenging, the mice were intraperitoneally injected with Mito-TEMPO (MT, dissolved in PBS, 10 mg/kg, HY-112879, MedChemExpress) every other day for 2 weeks, and the mice in the control group (vehicle) were intraperitoneally injected with an equal volume of PBS; for NOX2 blockade, 50 mg/kg GSK2795039 (GSK, dissolved in 20% DMSO, 20% Tween 80 and 60% polyethylene glycol 200, HY-18950, MedChemExpress) was administered by intraperitoneal injection daily for 2 weeks; for pan-caspase inhibition, the pan-caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp (OMe) fluoromethylketone (Z-VAD-FMK, dissolved in 10% DMSO, 40% PEG300, 5% Tween-80 and 45% saline, 10 mg/kg daily; HY-16658B; MedChemExpress) was administered by intraperitoneal injection for 2 weeks, and the mice in the control group (vehicle) were intraperitoneally injected with an equal volume of the abovementioned solvents daily for 2 weeks.

Techniques: Comparison, Microarray, Immunohistochemistry, Expressing, Imaging, Western Blot, Concentration Assay, Staining, Transfection, Control, Plasmid Preparation, Luciferase

Journal: International Journal of Biological Sciences

Article Title: Mitochondrial Dysfunction by FADDosome Promotes Gastric Mucosal Injury in Portal Hypertensive Gastropathy

doi: 10.7150/ijbs.90835

Figure Lengend Snippet: The alteration of glycolysis associated with dysfunction of the mitochondrial electron transport chain contributes to oxidative stress in the PHG. (A) Western blotting showed that the protein levels of LDHA and HK2 were increased in mice with PVL but were decreased by GSK2795039 (GSK, a NOX2 inhibitor) in mice with PVL. The ratios of the normalized LDHA/β-actin and HK2/β-actin densitometric units and the relative level of lactic acid were also analyzed. n = 6 per group. *P < 0.05 versus SO mice; #P < 0.05 versus mice with PVL not treated with GSK2795039. (B) Oxygen consumption rate (OCR) analysis of primary epithelial cells isolated from mouse models suggested that increased nonmitochondrial oxygen consumption and reduced maximum respiration were present in mice with PVL but were reversed by GSK2795039. n = 6 in each group. * P < 0.05 versus SO mice; # P < 0.05 versus mice with PVL without GSK2795039 treatment. (C) Examination of the glycolytic rate (for the extracellular acidification rate [ECAR]) showed that GSK2795039 treatment reduced basal and compensatory glycolysis in mice with PVL. n = 6 in each group. * P < 0.05 versus SO mice; # P < 0.05 versus mice with PVL without GSK2795039 treatment. (D) Two-dimensional hierarchical clustering results of the genes of electron transport chain (ETC)-related elements between PHG patients (as PHG) and healthy volunteers (as Uninvolved) (n = 3 per group). (E) Western blotting analysis (left panel) of protein levels of representative ETC complex subunits in the primary epithelial cells of PHG patients and healthy volunteers (as Uninvolved) revealed that the levels of the mitochondrial complex I, II, III, IV and V subunits were decreased in PHG patients. The activity of the antioxidant enzyme SOD and the ratio of reduced (GSH) to oxidized (GSSG) states (GSH/GSSG) detected by assay kits (right panel) were found to be decreased in PHG. n = 6 per group. * P < 0.05. (F) Western blotting analysis of representative ETC complex subunits from primary epithelial cells isolated from mouse models suggested that Nec-1 (an inhibitor of RIPK1) mitigated the defects in ETC subunit expression and decreased ROS levels in mice with PVL. The ratios of the normalized NDUFA9/β-actin, SDHA/β-actin, Cyt b/β-actin, COX I/β-actin and ATP5A/β-actin densitometric units and the levels of ROS were also analyzed. n = 6 per group. *P < 0.05 versus SO mice; #P < 0.05 versus mice with PVL without Nec-1 treatment. (G) SOD activity and the GSH/GSSG ratio, as detected by assay kits, were inhibited in mice with PVL but were restored by Nec-1 treatment. n = 6 per group. *P < 0.05 versus SO mice; #P < 0.05 versus mice with PVL without Nec-1 treatment. (H) Western blotting analysis and quantification of protein levels for representative ETC complex subunits from primary epithelial cells isolated from mouse models showed that Mdivi-1 reversed the decrease in mitochondrial complex I, II, III, IV and V subunit levels and reduced ROS levels in mice with PVL. The ratios of the normalized NDUFA9/β-actin, SDHA/β-actin, Cyt b/β-actin, COX I/β-actin and ATP5A/β-actin densitometric units and the levels of ROS were also presented. n = 6 per group. *P < 0.05 versus SO mice; #P < 0.05 versus mice with PVL without Mdivi-1 treatment. (I) Decreases in SOD activity and the GSH/GSSG ratio detected by assay kits were found in mice with PVL but were reversed by Mdivi-1 treatment. n = 6 per group. *P < 0.05 versus SO mice; #P < 0.05 versus mice with PVL without Mdivi-1 treatment.

Article Snippet: In some experiments, the mice were treated for 2 weeks after the operation with the following reagents or drugs according to the experimental needs: for NF-κB inhibition, the mice were intraperitoneally injected with Bay11708 (BAY, Calbiochem, La Jolla, CA, USA) at 200 μg/per mouse daily for 2 weeks; for TNF-α inhibition, the mice were intraperitoneally injected with infliximab (IFX, Janssen Biotech, Horsham, PA, USA) at 5 mg/kg per day for 2 weeks; for RIPK1 inhibition, the mice were intraperitoneally injected with 2 mg/kg necrostatin-1 (Nec-1, HY-15760, MedChemExpress, NJ, USA) per day for 2 weeks; for above experiments of the inhibitors, the mice in the control group (vehicle group) were intraperitoneally injected with an equal volume of phosphate-buffered saline (PBS) daily for 2 weeks; for Drp1 inhibition, the mice were intraperitoneally injected with 20 mg/kg Mdivi-1 (dissolved in DMSO, HY-15886, MedChemExpress), and the control animals (vehicle) were injected with an equal volume of DMSO; for ROS scavenging, the mice were intraperitoneally injected with Mito-TEMPO (MT, dissolved in PBS, 10 mg/kg, HY-112879, MedChemExpress) every other day for 2 weeks, and the mice in the control group (vehicle) were intraperitoneally injected with an equal volume of PBS; for NOX2 blockade, 50 mg/kg GSK2795039 (GSK, dissolved in 20% DMSO, 20% Tween 80 and 60% polyethylene glycol 200, HY-18950, MedChemExpress) was administered by intraperitoneal injection daily for 2 weeks; for pan-caspase inhibition, the pan-caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp (OMe) fluoromethylketone (Z-VAD-FMK, dissolved in 10% DMSO, 40% PEG300, 5% Tween-80 and 45% saline, 10 mg/kg daily; HY-16658B; MedChemExpress) was administered by intraperitoneal injection for 2 weeks, and the mice in the control group (vehicle) were intraperitoneally injected with an equal volume of the abovementioned solvents daily for 2 weeks.

Techniques: Western Blot, Isolation, Activity Assay, Expressing

Journal: Current Issues in Molecular Biology

Article Title: Development of a Novel Anti-CD44 Variant 4 Monoclonal Antibody C 44 Mab-108 for Immunohistochemistry

doi: 10.3390/cimb45030121

Figure Lengend Snippet: Immunohistochemical analysis using C 44 Mab-108 and C 44 Mab-46 against oral squamous cell carcinoma (OSCC) tissues. After antigen retrieval, the sections were incubated with 10 µg/mL of C 44 Mab-108 ( A , B , I , J ), 1 µg/mL of C 44 Mab-46 ( C , D , K , L ), and without the primary antibody (control) ( E , F , M , N ) followed by treatment with the Envision+ kit. The color was developed using 3,3′-diaminobenzidine tetrahydrochloride (DAB), and the sections were counterstained with hematoxylin. ( G , H , O , P ) Hematoxylin and eosin (HE) staining. ( Q , R ) Blocking of the C 44 Mab-108 reactivity to OSCC tissue by the CD44 peptide (aa 271–290) containing the C 44 Mab-108 epitope. After antigen retrieval, sections were incubated with C 44 Mab-108 (10 μg/mL) or C 44 Mab-108 (10 μg/mL) plus human CD44 peptide (aa 271–290, 10 μg/mL) followed by treatment with the Envision+ kit. The color was developed using DAB, and sections were counterstained with hematoxylin. Scale bar = 100 µm.

Article Snippet: The formalin-fixed paraffin-embedded (FFPE) OSCC tissue microarray (Product Code: OR601c, US Biomax Inc., Rockville, MD, USA) and the esophageal tissue microarray (Product Code: BC02011, US Biomax Inc.) were deparaffinized in xylene (Sigma-Aldrich Corp.) and rehydrated.

Techniques: Immunohistochemical staining, Incubation, Staining, Blocking Assay

Journal: Journal of Cancer

Article Title: CCT8 promotes cell migration and tumor metastasis in lung adenocarcinomas

doi: 10.7150/jca.87983

Figure Lengend Snippet: CCT8 is highly expressed in lung adenocarcinoma cancer. (A) The expression level of CCT8 in different cancers was analyzed using TIMER2.0, *p < 0.05; **p < 0.01; and ***p < 0.001. (B) Overall survival plots of lung cancer (n= 982) in Kaplan-Meier Plotter stratified by CCT8 mRNA expression. (C) Overall survival plots of LUAD (n = 504) and LUSC (n = 495) in TCGA stratified by CCT8 mRNA expression.

Article Snippet: After being rinsed in PBS for 5 min, the slides were blocked with a solution of 3% bovine serum albumin (BSA) at room temperature for 60 min, and incubated at 4 °C overnight with the primary antibodies: anti-CCT8 antibody (1:1000 dilution; 67539-1-Ig, Proteintech, Wuhan, China).

Techniques: Expressing

Journal: Journal of Cancer

Article Title: CCT8 promotes cell migration and tumor metastasis in lung adenocarcinomas

doi: 10.7150/jca.87983

Figure Lengend Snippet: Correlation between CCT8 expression and clinicopathological characteristics

Article Snippet: After being rinsed in PBS for 5 min, the slides were blocked with a solution of 3% bovine serum albumin (BSA) at room temperature for 60 min, and incubated at 4 °C overnight with the primary antibodies: anti-CCT8 antibody (1:1000 dilution; 67539-1-Ig, Proteintech, Wuhan, China).

Techniques: Expressing

Journal: Journal of Cancer

Article Title: CCT8 promotes cell migration and tumor metastasis in lung adenocarcinomas

doi: 10.7150/jca.87983

Figure Lengend Snippet: Association between CCT8 protein expression and overall survival (OS). (A) The intensity of staining was scored as follows: 0, no expression; 1, mild expression; 2, intermediate expression; and 3, strong expression. (B) Representative microphotographs of CCT8 immunoreactivities in lung adenocarcinoma and adjacent tissue. (C) Quantitative analysis of the expression of CCT8 in lung adenocarcinoma and adjacent tissue of the tissue microarray. (D) Association between OS and CCT8 protein expression. The OS of patients with high CCT8 expression was lower than that of patients with low CCT8 expression.

Article Snippet: After being rinsed in PBS for 5 min, the slides were blocked with a solution of 3% bovine serum albumin (BSA) at room temperature for 60 min, and incubated at 4 °C overnight with the primary antibodies: anti-CCT8 antibody (1:1000 dilution; 67539-1-Ig, Proteintech, Wuhan, China).

Techniques: Expressing, Staining, Microarray

Journal: Journal of Cancer

Article Title: CCT8 promotes cell migration and tumor metastasis in lung adenocarcinomas

doi: 10.7150/jca.87983

Figure Lengend Snippet: CCT8 promotes cell migration. Stable PC9 cells, which were infected with lentivirus encoding empty vector or CCT8, and stable H1299 cells, which were infected with lentivirus expressing sgC or sgCCT8, were subjected to immunoblot analysis for detection of CCT8 protein expression (A), colony formation (B), transwell assay (C), and wound-healing assay for testing of cell migration (D).

Article Snippet: After being rinsed in PBS for 5 min, the slides were blocked with a solution of 3% bovine serum albumin (BSA) at room temperature for 60 min, and incubated at 4 °C overnight with the primary antibodies: anti-CCT8 antibody (1:1000 dilution; 67539-1-Ig, Proteintech, Wuhan, China).

Techniques: Migration, Infection, Plasmid Preparation, Expressing, Western Blot, Transwell Assay, Wound Healing Assay

Journal: Journal of Cancer

Article Title: CCT8 promotes cell migration and tumor metastasis in lung adenocarcinomas

doi: 10.7150/jca.87983

Figure Lengend Snippet: CCT8 interacts with and activates AKT signaling to accelerate cell migration and tumor metastasis. (A) PC9 stable cells, which were infected with lentivirus encoding empty vector or CCT8, were subjected to qPCR analysis for mRNA levels of CDH1, CDH2, MMP2, MMP9 and VEGFA. (B) PC9 cells and H1299 cells were subjected to anti-CCT8 (or anti-normal rabbit IgG) immunoprecipitation (IP), and coprecipitated endogenous AKT was detected by immunoblot analysis (IB). (C) PC9 stable cells, which were infected with lentivirus encoding empty vector or CCT8, and H1299 stable cells, which were infected with lentivirus expressing sgC or sgCCT8, were subjected to immunoblot analysis for protein levels of phosphorylated AKT (p-AKT), total AKT (t-AKT) and CCT8, Ecadherin (E-cad). (D-G) Stable PC9 cells were treated with MK2206 for 24 h and then subjected to immunoblot analysis for protein levels of phosphorylated AKT (p-AKT), total AKT (t-AKT) and CCT8, E-cadherin (E-cad) (D), colony formation (E), ranswell assay (F) and wound-healing assay (G). (H-I) Stable cells were injected into the tail vein of nude mice (4 mice per group). Two weeks after cell inoculation, these mice were treated with MK2206 (120 mg/kg daily for two weeks) or DMSO daily for two weeks. Lungs were dissected (H) and then subjected to H&E staining for histological analysis (left), and the lung lesions were counted (right) (I). (J) A working model showing that CCT8 activates AKT to promote tumor metastasis. **, P < 0.01; ns, not significant.

Article Snippet: After being rinsed in PBS for 5 min, the slides were blocked with a solution of 3% bovine serum albumin (BSA) at room temperature for 60 min, and incubated at 4 °C overnight with the primary antibodies: anti-CCT8 antibody (1:1000 dilution; 67539-1-Ig, Proteintech, Wuhan, China).

Techniques: Migration, Infection, Plasmid Preparation, Immunoprecipitation, Western Blot, Expressing, Wound Healing Assay, Injection, Staining

Journal: Journal of Cancer

Article Title: CCT8 promotes cell migration and tumor metastasis in lung adenocarcinomas

doi: 10.7150/jca.87983

Figure Lengend Snippet: Univariate analyses of the factors correlated with overall survival of cancer patients

Article Snippet: After being rinsed in PBS for 5 min, the slides were blocked with a solution of 3% bovine serum albumin (BSA) at room temperature for 60 min, and incubated at 4 °C overnight with the primary antibodies: anti-CCT8 antibody (1:1000 dilution; 67539-1-Ig, Proteintech, Wuhan, China).

Techniques: Expressing