Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: (A) (i)Western Blot analysis of the level of eIF2α and eIF2α p[S51] expression and puromycin incorporation assays in U373, MO3.13 and SH-SY5Y cells following siRNA mediated silencing of eIF2Bα, 200 nM ISRIB treatment for 1h, or 1 µM Tg for 1h. (ii) Levels of phosphorylated eIF2α were normalized to levels of total eIF2α and presented as mean ± SD (n=3). (iii) Levels of puromycin were normalized to β-actin and are presented as mean ± SD ( n = 3). p Values derived from a two-way ANOVA test, followed by a Tukey’s multiple analysis, * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, **** p ≤ 0.0001. (B) ELISA analysis of the level of ATF4 expression in in U373, MO3.13 and SH-SY5Y cells following siRNA mediated silencing of eIF2Bα, 200 nM ISRIB treatment for 1h, or 300 nM Tg for 6h. Levels of ATF4 detected by ELISA are presented as mean ± SD (n=3). p Values derived from a one-way ANOVA test, followed by a Tukey’s multiple analysis, * p ≤ 0.05, ** p ≤ 0.01 *** p ≤ 0.001, **** p ≤ 0.0001. (C) Cells were transfected with Cy3 labelled siRNA negative control, Cy3 labelled siRNA targeting EIF2B1, and Cy3 labelled siRNA targeting EIF2B1 coupled with ISRIB 1h (200 nM) treatment. U373-MG and SH-SY5Y cells were fixed in methanol, MO3.13 cells were fixed in 4%PFA and subjected to ICC with anti-G3BP primary antibody and visualized using appropriate secondary antibodies conjugated to Alexa Fluor 488. Mean percentages of U373-MG, MO3.13 and SH-SY5Y cells with G3BPcontaining SGs. Error bars: ±s.d. Data was analysed using one-way ANOVA followed by a Tukey’s multiple analysis. *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001

Article Snippet: The Human ATF4 enzyme-linked immunosorbent assay (ELISA) Kit (Proteintech KE00147), was used to determine levels of ATF4, following manufacturer’s instructions.

Techniques: Western Blot, Expressing, Derivative Assay, Enzyme-linked Immunosorbent Assay, Transfection, Negative Control

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: C-Reactive Protein Decreases Interleukin-10 Secretion in Activated Human Monocyte-Derived Macrophages via Inhibition of Cyclic AMP Production

doi: 10.1161/01.atv.0000241572.05292.fb

Figure Lengend Snippet: Figure 4. A, i, Effect of CRP on intracellular cAMP levels in HMDMs. The cells were incubated with LPS (500 ng/mL for 24 hours) in the absence or presence of CRP (25 g/mL) for 12 hours. The intracellular cAMP levels were measured by ELISA. ANOVA, P0.01; post hoc Tukey test: *P0.001 vs control; **P0.035 vs LPS alone. The results are meanSD of 3 experiments in duplicate. A, ii, Effect of CRP on AC activity in HMDMs. HMDMs on day 7 were pretreated with CRP for 12 hours followed by LPS challenge for 24 hours in serum-free medium. Membrane fractions were used to measure AC activity by the conversion of 32P-ATP to 32P-cAMP. Data are pre- sented as meanSD of 3 experiments in duplicate. The results are expressed as 32P-cAMP counts/mg protein. *P0.05 vs control, **P0.05 vs LPS alone. B, Effect of cAMP agonists on CRP-mediated IL-10 inhibition. Cells were treated with Db-cAMP (50 mol/L) or Br-cAMP (50 mol/L) 1 hour before CRP treatment (12 hours) followed by LPS challenge for 24 hours. IL-10 was measured in superna- tants. The results are meanSD of 3 experiments in duplicate. ANOVA, §P0.01; post hoc Tukey test: P0.001 vs control; *P0.01 vs LPS; **P0.01 vs CRPLPS. C, Effect of CRP on pCREB/CREB levels. Cells were treated with cAMP agonists 30 minutes before cell harvesting following CRPLPS treatment. Cell lysates were prepared as described in Materials and Methods. C, i, Effect of CRP on ratio of pCREB/CREB. Protein (50 g) was used for measurement of CREB and pCREB by ELISA and ratio of pCREB/CREB was cal- culated. ANOVA, P0.05; post hoc Tukey test: §P0.02 vs control; *P0.03 vs LPS alone; **P0.01 vs CRPLPS. The results are meanSD of 3 experiments in duplicate. C, ii, Western blot analysis depicting effect of CRP on pCREB and CREB done in cell lysates as described in Materials and Methods. Lane 1, control; lane 2, LPS; lane 3, CRPLPS; lane 4, CRPLPS Db-cAMP; lane 5, CRPLPSBr-cAMP. §P0.04 vs control, *P0.05 vs LPS alone, **P0.002 vs CRPLPS (n3 experiments for densitometric ratios).

Article Snippet: CREB as well as pCREB ELISA kits and antibodies were from Biosource and Cell Signaling Technology respectively.

Techniques: Incubation, Enzyme-linked Immunosorbent Assay, Control, Activity Assay, Membrane, Inhibition, Cell Harvesting, Western Blot

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: C-Reactive Protein Decreases Interleukin-10 Secretion in Activated Human Monocyte-Derived Macrophages via Inhibition of Cyclic AMP Production

doi: 10.1161/01.atv.0000241572.05292.fb

Figure Lengend Snippet: Figure 5. Effect of transfection in THP-1 with indicated CREB plasmids on CRP- mediated IL-10 inhibition. THP-1 cells were grown in 12-well plates and trans- fected with indicated CREB plasmids as detailed in Materials and Methods. Forty- eight hours after transfection, THP-1 cells were pretreated with CRP (25 g/mL) for 12 hours followed by LPS (500 ng/mL) for 24 hours. A, Superna- tants were used for IL-10 measurement *P0.05 vs control, **P0.05 vs LPS alone, €P0.01 vs control in transfected cells. B, Cells were used for the mea- surement of total and pCREB. The results are expressed as pCREB/CREB ratio. *P0.03 vs control, **P0.045 vs LPS alone, €P0.029 vs control in transfected cells.

Article Snippet: CREB as well as pCREB ELISA kits and antibodies were from Biosource and Cell Signaling Technology respectively.

Techniques: Transfection, Inhibition, Control

Journal: Basic and Clinical Neuroscience

Article Title: Datumetine Preferentially Upregulates N-methyl-D-aspartate Receptor Signalling Pathways in Different Brain Regions of Mice

doi: 10.32598/bcn.2021.3397.1

Figure Lengend Snippet: List and manufacturer details of ELISA kits used

Article Snippet: EM0951 , Mouse CREB (cyclic AMP response element binding protein) ELISA Kit , Wuhan Fine Biotech Co., Ltd. Wuhan China..

Techniques: Enzyme-linked Immunosorbent Assay, Binding Assay

Journal: Basic and Clinical Neuroscience

Article Title: Datumetine Preferentially Upregulates N-methyl-D-aspartate Receptor Signalling Pathways in Different Brain Regions of Mice

doi: 10.32598/bcn.2021.3397.1

Figure Lengend Snippet: Graphical representation of cyclic AMP response element binding protein (CREB) level in different brain regions A) Hippocampal level of CREB was significantly ( *** P<0.001) increased in MK-80+datumetine mice than Veh and datumetine mice. B) PFC level of CREB was increased in Datumetine mice while MK-801+datumetine mice showed reduced CREB than Veh, though no significant difference was observed. C) Cerebellar CREB level was reduced in all treated mice compared to Veh mice, though not significant.

Article Snippet: EM0951 , Mouse CREB (cyclic AMP response element binding protein) ELISA Kit , Wuhan Fine Biotech Co., Ltd. Wuhan China..

Techniques: Binding Assay

Journal: Current Research in Physiology

Article Title: Yeast supplementation potentiates fluoxetine's anti-depressant effect in mice via modulation of oxido-inflammatory, CREB, and MAPK signaling pathways

doi: 10.1016/j.crphys.2024.100132

Figure Lengend Snippet: Yeast and/or fluoxetine prevent UCMS-induced CREB and MAPK activities dysregulation in mice. Bars represent the mean ± S.E.M. ∗ p < 0.0001 compared to normal control group, # p < 0.0001 compared to negative control group and β p < 0.05 compared to fluoxetine (20 mg/kg) group. VEH = vehicle; FLX = fluoxetine; CREB = cAMP response element-binding protein; MAPK = mitogen-activated protein kinase; UCMS = unpredictable chronic mild stress.

Article Snippet: CREB levels were measured using a CREB ELISA kit (Wuhan Fine Biotech Co., Ltd, China).

Techniques: Control, Negative Control, Binding Assay

Journal: International Journal of Molecular Sciences

Article Title: Extracts of Sideritis scardica and Clinopodium vulgare Alleviate Cognitive Impairments in Scopolamine-Induced Rat Dementia

doi: 10.3390/ijms25031840

Figure Lengend Snippet: Effect of S. scardica , C. vulgare extracts and their combination on the brain concentrations of BDNF and pCREB in healthy rats and rats with scopolamine-induced memory impairment. The BDNF and pCREB concentrations were assessed in the cortex ( A , C ) and hippocampus ( B , D ) of the rats with enzyme-linked immunosorbent assay (ELISA). Results are presented as mean values ± SEM ( n = 6 animals per group). Statistical analysis involved one-way analysis of variance (ANOVA), followed by Tukey’s multiple comparison test. Significance vs. saline-treated group: ### p < 0.001 significance vs. Sco-treated group * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: The following kits were used: Rat BDNF ELISA Kit (cat. no. E-EL-R1235); Rat Phospho cAMP response element binding protein (P-CREB) ELISA Kit (cat. no. SL1344Ra), purchased from Elabscience (Houston, TX, USA).

Techniques: Enzyme-linked Immunosorbent Assay, Comparison, Saline