phosphor creb  (Danaher Inc)

Journal: Biology

Article Title: Amphiregulin Upregulation in Visfatin-Stimulated Colorectal Cancer Cells Reduces Sensitivity to 5-Fluororacil Cytotoxicity

doi: 10.3390/biology13100821

Figure Lengend Snippet: JNK1/2 and p38 signaling affects CREB and STAT3 phosphorylation and subsequent 5-FU-initiated cytotoxicity in Visfatin-stimulated HCT-116 CRC cells. ( A ) HCT-116 CRC cells were pretreated with DMSO (vehicle), SP600125 (a JNK1/2 inhibitor at 20 μM), or SB203580 (a p38 inhibitor at 10 μM) for 30 min and then either maintained as controls or treated with Visfatin (50 ng/mL) for 4 h. The phosphorylation of CREB and STAT3 was determined using Western blot analysis. ( B ) HCT-116 CRC cells were pretreated with DMSO (vehicle), PD98059 (an ERK1/2 inhibitor at 25 μM), SP600125 (a JNK1/2 inhibitor at 20 μM), or SB203580 (a p38 inhibitor at 10 μM) for 30 min and then treated with vehicle (PBS) or Visfatin (50 ng/mL) for 1 h. Subsequently, the cells were then either maintained as controls or treated with 5-FU (10 μM) for 24 h. The viability of the treated cells was examined using the MTT assay. The results in ( A ) were obtained from three independent experiments, all of which exhibited similar outcomes. The data in ( A , B ) were calculated from the results of three independent experiments and are presented as the mean ± SEM. The results were considered statistically significant when p < 0.05. *, compared to untreated control cells. #, compared to cells treated with ( A ) DMSO and Visfatin and ( B ) 5-FU alone. ( B ) **, compared to cells treated with DMSO and 5-FU.

Article Snippet: For antibody detection, the following primary antibodies were used, all procured from Cell Signaling Technology (Beverly, MA, USA): AREG (#42953, 1:800 dilution), phosphor-JNK1/2 (#4671, 1:1000 dilution), JNK1 (#3708, 1:1000 dilution), phosphor-p38 (#4511, 1:1000 dilution), p38 (#9212, 1:1000 dilution), phosphor-CREB (#9196, 1:500 dilution), CREB (#9197, 1:1000 dilution), phosphor-STAT3 (#9145, 1:1500 dilution), STAT3 (#9139, 1:1000 dilution), and β-actin (#3700, 1:2000 dilution).

Techniques: Phospho-proteomics, Western Blot, MTT Assay, Control

Journal: Frontiers in Neuroscience

Article Title: Cocaine amphetamine-regulated transcription peptide inhibits apoptosis in oxygen–glucose deprived neural stem cells

doi: 10.3389/fnins.2024.1424719

Figure Lengend Snippet: The effect of KG501 (a specific inhibitor p-CREB) on proliferation and apoptosis levels in OGD NSCs. (A) CART55-102(0.8 nM) treatment protects against loss of the structure of NSCs (Nestin, red) and enhances Nestin (Red) expression and KG501 inhibits the protection of structure in cultured oxygen and glucose deprived NSCs. Scale bar = 100 pm. (B) CART55-102(0.8 nM) treatment enhances Sox-2(Green) expression and KG501 inhibits the protection of neural stem cells in cultured oxygen and glucose deprived. Scale bar = 50 μm (C) CART55-102(0.8 nM) treatment increases the proliferation of oxygen and glucose deprived neural stem cells and KG-501 inhibits the proliferation of NSCs in cultured oxygen and glucose deprived. Scale bar = 100um. (D) Quantitative analysis of fluorescence results using ImageJ software. Data was presented as mean ± SEM, *** p < 0.001, ### p < 0.001. (E) The protein levels Bcl-2 and Bax were measured by Western blot in different groups. (F) Quantitative analysis of Bcl-2 expression. Blot images were cropped for comparison. All values are expressed as mean ± SEM of the band intensity of Bcl-2 normalized to Tubulin from 3 independent experiments. Compared with the OGD, after treatment CART55-102 (0.8 nM), the levels of Bcl-2 significantly increased. But the levels of Bcl-2 were significantly decreased after pre-treatment with KG501 in the CART treatment group. ** p < 0.001, *** p < 0.01, ## p < 0.01, ### p < 0.001. (G) Quantitative analysis of Bax expression. Blot images were cropped for comparison. All values are expressed as meant ± SEM of the band intensity of Bax normalized to Tubulin from 3 independent experiments. Compare with OGD, after treatment CART55-102(0.8 nM), the levels of Bax significantly decreased. But the levels of Bax were significantly increased after pre-treatment with KG501 in the CART treatment group. *** p < 0.001, ### p < 0.001. (H) The protein levels of p-CREB were measured by Western blot in different groups. (I) Quantitative analysis p-CREB expression. Blot images were cropped for comparation. All values are expressed as mean ± SEM of the band intensity of p-CREB normalized to 3 independent experiments. Compare with the OGD, after treatment CART55-102(0.8 nM), the levels of p-CREB significantly increased. But the levels of p-CREB were significantly decreased after pre-treatment with KG501 in the CART treatment group. *** p < 0.001, ## p < 0.01.

Article Snippet: Antibodies against Nestin, Sox, Synaptophysin CREB and phosphor-CREB (phosphor S133) were purchased from Abcam (Cambridge, MA, United States).

Techniques: Expressing, Cell Culture, Fluorescence, Software, Western Blot, Comparison