ldn 27219 powder  (Tocris)

Journal: Gastroenterology Report

Article Title: Targeting transglutaminase 2: pathways to celiac disease therapies

doi: 10.1093/gastro/goaf086

Figure Lengend Snippet: Effects of different TG2 inhibitors on the cross-linking activity of recombinant TG2, cell viability, and extracellular TG2 activity. (A) Fluorometric quantitation of the cross-linking of biotin-conjugated gliadin peptide P56-88 by recombinant TG2 ( n = 3, three technical replicates per experiment). (B) Fluorometric quantitation of cell viability after 24 h of incubation with the corresponding substances using the resazurin-based assay ( n = 3, two technical replicates per experiment). (C) Fluorometric quantitation of extracellular cross-linking of the TG2 substrate 5BP by Caco-2 cells ( n = 3, with three technical replicates per experiment). All data are shown as mean ± SD. Statistical significance was tested by using univariate ANOVA (A) or Student’s t -test with Welch correction. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. 5BP = 5-(biotinamido)-pentylamine, ZnCl 2 = zinc chloride, AA = ascorbic acid.

Article Snippet: LDN27219 , , GTP antagonism, stabilization of closed (inactive) conformation of TG2 , HY-16693, MedChemExpress, Monmouth Junction, NJ, USA.

Techniques: Activity Assay, Recombinant, Quantitation Assay, Incubation, Resazurin Assay

Journal: Gastroenterology Report

Article Title: Targeting transglutaminase 2: pathways to celiac disease therapies

doi: 10.1093/gastro/goaf086

Figure Lengend Snippet: Effect of IFN-γ on the expression of ERp57, TRX and TG2 in Caco-2 cells. Analysis of relative expression of (A) ERp57, (B) TRX, and (C) TG2 by Western blotting after stimulation with different concentrations of IFN-γ over 48 h. No significant change in the expression of ERp57 and TRX. Dose-dependent increase in TG2 expression. All data are shown as mean ± SD. Statistical significance was tested by using Student’s t -test. n = 4. * P < 0.05.

Article Snippet: LDN27219 , , GTP antagonism, stabilization of closed (inactive) conformation of TG2 , HY-16693, MedChemExpress, Monmouth Junction, NJ, USA.

Techniques: Expressing, Western Blot

Journal: Gastroenterology Report

Article Title: Targeting transglutaminase 2: pathways to celiac disease therapies

doi: 10.1093/gastro/goaf086

Figure Lengend Snippet: SiRNA knockdown of ERp57, TRX, and TG2, and the effect on cellular/extracellular TG2 activity. Caco-2 cells were incubated with specific siRNAs or mock siRNA for 48 h. The efficacy of the siRNA knockdown was determined by using Western blotting. Expression of ERp57, TRX, and TG2 was normalized against the housekeeping protein GAPDH. (A–C) SiRNA treatment significantly reduced expression of (A) ERp57, (B) TRX, and (C) TG2. Incubation with mock siRNA did not affect protein expression ( n = 4). (D) Treatment with ERp57-siRNA und TRX-siRNA did not show any off-target effects on TG2 expression ( n = 3). (E) Cellular expression of ERp57, TRX, and TG2 determined by fluorometry after siRNA knockdown. Protein levels of all three target proteins were reduced by siRNA-mediated knockdown. (F) Cellular TG2 activity was determined by fluorometry after siRNA treatment. SiRNA knockdown of TRX and TG2 reduced cellular TG2 activity ( n = 5). (G) Extracellular expression of ERp57, TRX, and TG2 was investigated in unpermeabilized Caco-2 cells by fluorometry. Protein levels of all three target proteins were significantly reduced by siRNA-mediated knockdown. (H) Extracellular TG2 activity was investigated in unpermeabilized Caco-2 cells by fluorometry. SiRNA knockdown of TRX and TG2 significantly reduced extracellular TG2 activity ( n = 5). All data are shown as mean ± SD. Statistical significance was tested by using Student’s t -test with Welch correction where appropriate. * P < 0.05, ** P < 0.01, *** P < 0.001. GAPDH = glyceraldehyde 3-phosphate dehydrogenase.

Article Snippet: LDN27219 , , GTP antagonism, stabilization of closed (inactive) conformation of TG2 , HY-16693, MedChemExpress, Monmouth Junction, NJ, USA.

Techniques: Knockdown, Activity Assay, Incubation, Western Blot, Expressing

Journal: Gastroenterology Report

Article Title: Targeting transglutaminase 2: pathways to celiac disease therapies

doi: 10.1093/gastro/goaf086

Figure Lengend Snippet: TG2 is expressed in the lamina propria and the epithelium of CD biopsies. (A) Fluorescence staining revealed TG2 to be nearly absent in the epithelium of control biopsies and low levels were expressed in the lamina propria (arrows). In CD biopsies, TG2 expression was increased in the epithelium (arrowheads) and the lamina propria (arrows). (B) Detailed images of the duodenal mucosa reveal high levels of TG2 in the lamina propria (arrows) and the epithelium (arrowheads) of CD patients. (C) TG2 expression was significantly increased in the epithelium and the lamina propria of CD patients. All data are shown as median. Statistical significance was tested by using Student’s t -test. n = 5. * P < 0.05; scale: 20 µm.

Article Snippet: LDN27219 , , GTP antagonism, stabilization of closed (inactive) conformation of TG2 , HY-16693, MedChemExpress, Monmouth Junction, NJ, USA.

Techniques: Fluorescence, Staining, Control, Expressing