Journal: Shock (Augusta, Ga.)

Article Title: RESVERATROL DOWNREGULATES BIOMARKERS OF SEPSIS VIA INHIBITION OF PROTEASOME’S PROTEASES

doi: 10.1097/SHK.0000000000001080

Figure Lengend Snippet: Measurement of Proteasome Activities. Dose-dependent inhibition of purified human proteasome or immunoproteasome protease activities using resveratrol (0.1 μM – 100 μM), and ONX-0914 (0.01 μM – 80 μM). Proteasome activities were measured at excitation 345 and emission 445 wavelengths after addition of peptide fluorogenic substrates using Cytation 3 Plate Multi-mode reader. Data are shown as means ± SE (n=3) and each reaction was performed in triplicates. Results were plotted as percentage inhibition compared with vehicle (DMSO) control.

Article Snippet: Purified human erythrocyte 20S proteasome and peripheral blood monocyte 20S immunoproteasomes (R&D Systems) were assayed for different peptidase activities using fluorogenic tri-and tetra-peptide substrates coupled to AMC.

Techniques: Inhibition, Purification, Control

Journal: Shock (Augusta, Ga.)

Article Title: RESVERATROL DOWNREGULATES BIOMARKERS OF SEPSIS VIA INHIBITION OF PROTEASOME’S PROTEASES

doi: 10.1097/SHK.0000000000001080

Figure Lengend Snippet: Measurement of Cellular Proteasome Activities. Dose dependent effect of resveratrol on cellular proteasome activity. THP1 cells at 1×104 cells/well were treated with different concentrations of resveratrol (0.1 μM – 100 μM) and incubated at 37°C and 5% CO2, for 30 min. The relative luminescence units (RLU) of assays were read in a Cytation 3 Plate Multi-mode reader. Data are shown as means ±SE (n=3) and each reaction was performed in triplicates. Results were plotted as percentage inhibition compared with vehicle (DMSO) control.

Article Snippet: Purified human erythrocyte 20S proteasome and peripheral blood monocyte 20S immunoproteasomes (R&D Systems) were assayed for different peptidase activities using fluorogenic tri-and tetra-peptide substrates coupled to AMC.

Techniques: Activity Assay, Incubation, Inhibition, Control

Journal: Shock (Augusta, Ga.)

Article Title: RESVERATROL DOWNREGULATES BIOMARKERS OF SEPSIS VIA INHIBITION OF PROTEASOME’S PROTEASES

doi: 10.1097/SHK.0000000000001080

Figure Lengend Snippet: Effects of resveratrol (80 μM) or ONX-0914 (0.25 μM) on proteasome subunits gene expression using CD14+ human blood monocytes. CD14+ monocytes (2×106 cells/well) were treated for Vehicle, LPS, Res, Res+LPS, ONX and ONX+LPS conditions, as described in the methods section. Gene expression of proteasome subunits PSMB5 (X), PSMB6 (Y), PSMB7 (Z), PSMB8 (LMP7), PSMB9 (LMP2) and PSMB10 (LMP10) were analyzed by qRT-PCR. Data are shown as means ±SE (n=3). *Statistically significant difference of LPS treatment vs veh, Res, Res+LPS, ONX and ONX+LPS (p≤ 0.05).

Article Snippet: Purified human erythrocyte 20S proteasome and peripheral blood monocyte 20S immunoproteasomes (R&D Systems) were assayed for different peptidase activities using fluorogenic tri-and tetra-peptide substrates coupled to AMC.

Techniques: Gene Expression, Quantitative RT-PCR

Journal: British journal of cancer

Article Title: Inactivation of the WASF3 gene in prostate cancer cells leads to suppression of tumorigenicity and metastases.

doi: 10.1038/sj.bjc.6605850

Figure Lengend Snippet: Figure 7 Knockdown of WASF3 expression suppresses MMP9 expression levels and inhibits MMP9 release from prostate cancer cell lines. (A) RT–PCR analysis of PC3 and DU145 cells showing knockdown of WASF3 also show down-regulation of MMP9, but not MMP2 expression levels. Cont ¼ parental cells. (B) Secretion of the pro- and active form of MMP9 from PC3 and DU145 cells were measured in the supernatants of actively growing cells using the Fluorokine MAP assay procedure. Cells from each clone showing knockdown of WASF3 also showed reduced MMP9 levels compared with parental (control) and non-knockdown cells that did not.

Article Snippet: After 24 h, the media were collected in tubes and centrifuged for 10 min at 10 000 g. The pro- and active MMP-9 levels released into the media were measured using a Fluorokine MAP human MMP9 kit (R&D Systems, Minneapolis, MN, USA) according to the manufacturer’s instructions.

Techniques: Knockdown, Expressing, Reverse Transcription Polymerase Chain Reaction, Control