anti p mapkapk 2  (Cell Signaling Technology Inc)

Journal: Cell Death & Disease

Article Title: Involvement of p38 MAPK and MAPKAPK2 in promoting cell death and the inflammatory response to ischemic stress associated with necrotic glioblastoma

doi: 10.1038/s41419-025-07335-3

Figure Lengend Snippet: A LN229 cells were cultured under the ischemic condition for the indicated time and subjected to western blotting. B LN229 vector , LN229 KRAS(G12D), or LN229 PIK3CA(H1047R) cells cultured under regular or ischemic conditions were subjected to western blotting. C LN229 cells treated by indicated p38 inhibitors or the DMSO control were subjected to western blotting. D Viability of LN229 KRAS(G12D) and LN229 PIK3CA(H1047R) cells treated with indicated p38 inhibitors (AMG548, 2 μM; VX745, 2 μM; Scio469, 1 μM) under ischemic conditions. Ordinary one-way ANOVA test. E LN229 KRAS(G12D) or LN229 PIK3CA(H1047R) cells stably transduced by indicated shRNAs targeting p38α or a scrambled shRNA control were subjected to western blotting. F Cells, as indicated in E , were cultured in the ischemic condition for 18 h and subjected to crystal violet cell viability assay. Ordinary one-way ANOVA test. G Viability of LN229 KRAS(G12D) and LN229 PIK3CA(H1047R) cells treated with DMSO or MK25 (10 μM) under ischemic conditions. Student’s t test. H LN229 KRAS(G12D) or LN229 PIK3CA(H1047R) cells stably transduced by indicated shRNAs targeting MK2 or a scrambled shRNA control were subjected to western blotting. I Cells, as indicated in H , were cultured in the ischemic condition for 18 h and subjected to crystal violet cell viability assay. Ordinary one-way ANOVA test.

Article Snippet: The antibodies used for western blotting and immunohistochemistry staining included: ATF4 (Cell Signaling Technology, 11815), p-EIF2a (S51) (Cell Signaling Technology, 3398), β-actin (Cell Signaling Technology, 3700), p-p38 (T180/Y182) (Cell Signaling Technology, 4511), p38 (Cell Signaling Technology, 8690), p-MK2 (T334) (Cell Signaling Technology, 3007), MK2 (Cell Signaling Technology, 12155), p-ERK (Cell Signaling Technology, 4370), p-AKT(S473) (Cell Signaling Technology, 4060), anti-rabbit HRP-conjugated secondary antibody (Cell Signaling Technology, 7074S), anti-mouse HRP-conjugated antibody (Cell Signaling Technology, 7076S).

Techniques: Cell Culture, Western Blot, Plasmid Preparation, Control, Stable Transfection, shRNA, Viability Assay

Journal: Cell Death & Disease

Article Title: Involvement of p38 MAPK and MAPKAPK2 in promoting cell death and the inflammatory response to ischemic stress associated with necrotic glioblastoma

doi: 10.1038/s41419-025-07335-3

Figure Lengend Snippet: A LN229 cells stably transduced by indicated shRNAs targeting IRE1α or a scrambled shRNA control were cultured under the ischemic condition and subjected to western blotting. B LN229 vector , LN229 KRAS(G12D) or LN229 PIK3CA(H1047R) cells transfected by a pool of four siRNA against IRE1α or scrambled siRNA and cultured under regular or ischemic conditions were subjected to western blotting. C , D LN229 cells stably transduced by indicated sgRNAs targeting p38 ( C ) or MK2 ( D ), or an empty vector (EV) were cultured under regular (control) or ischemic conditions and subjected to western blotting. E A diagram illustrating the signaling pathways activated under the ischemic condition.

Article Snippet: The antibodies used for western blotting and immunohistochemistry staining included: ATF4 (Cell Signaling Technology, 11815), p-EIF2a (S51) (Cell Signaling Technology, 3398), β-actin (Cell Signaling Technology, 3700), p-p38 (T180/Y182) (Cell Signaling Technology, 4511), p38 (Cell Signaling Technology, 8690), p-MK2 (T334) (Cell Signaling Technology, 3007), MK2 (Cell Signaling Technology, 12155), p-ERK (Cell Signaling Technology, 4370), p-AKT(S473) (Cell Signaling Technology, 4060), anti-rabbit HRP-conjugated secondary antibody (Cell Signaling Technology, 7074S), anti-mouse HRP-conjugated antibody (Cell Signaling Technology, 7076S).

Techniques: Stable Transfection, shRNA, Control, Cell Culture, Western Blot, Plasmid Preparation, Transfection, Protein-Protein interactions

Journal: Cell Death & Disease

Article Title: Involvement of p38 MAPK and MAPKAPK2 in promoting cell death and the inflammatory response to ischemic stress associated with necrotic glioblastoma

doi: 10.1038/s41419-025-07335-3

Figure Lengend Snippet: A Kaplan–Meier survival curves comparing mice intracranially injected with LN229 vector , LN229 KRAS(G12D) , or LN229 PIK3CA(H1047R) cells. B H&E-stained tumor sections derived from LN229 vector , LN229 KRAS(G12D) or LN229 PIK3CA(H1047R) terminal tumors. C Quantification of the necrotic area presented as the percentage of total tumor area in H&E-stained tumor sections from LN229 vector , LN229 KRAS(G12D) or LN229 PIK3CA(H1047R) tumors. Ordinary one-way ANOVA test. D Immunofluorescent staining of mouse neutrophil marker, Ly6G, hypoxia marker, pimonidazole, and DAPI on paraffin-embedded sections of LN229 KRAS(G12D) - or LN229 PIK3CA(H1047R) -derived tumors upon reaching endpoints. E Chromogenic staining of p-MK2 on paraffin-embedded sections of LN229 KRAS(G12D) - or LN229 PIK3CA(H1047R) -derived endpoint tumors. In B , D , and E , N denotes tumor necrosis, and CT denotes cellular tumor. In D , the right images depict magnified views of the corresponding outlined areas from the left images. F Growth curve of LN229 KRAS(G12D) or LN229 PIK3CA(H1047R) cells cultured under regular conditions. (n = 3). G The size of tumors derived from LN229 KRAS(G12D) or LN229 PIK3CA(H1047R) cells transduced by an empty vector (EV) or p38 gRNA (KO-p38) was monitored using bioluminescence imaging. (n = 10 mice for each group, except n = 9 for EV of each tumor type at the last time point). Multiple unpaired t-test. H Kaplan-Meier survival curves comparing mice intracranially injected with p38α-depleted (KO-p38) LN229 KRAS(G12D) or LN229 PIK3CA(H1047R) cells. Log-rank test. I Quantification of the necrotic area presented as the percentage of total tumor area in H&E-stained tumor sections from LN229 KRAS(G12D) cells transduced by EV or KO-p38. Student’s t test.

Article Snippet: The antibodies used for western blotting and immunohistochemistry staining included: ATF4 (Cell Signaling Technology, 11815), p-EIF2a (S51) (Cell Signaling Technology, 3398), β-actin (Cell Signaling Technology, 3700), p-p38 (T180/Y182) (Cell Signaling Technology, 4511), p38 (Cell Signaling Technology, 8690), p-MK2 (T334) (Cell Signaling Technology, 3007), MK2 (Cell Signaling Technology, 12155), p-ERK (Cell Signaling Technology, 4370), p-AKT(S473) (Cell Signaling Technology, 4060), anti-rabbit HRP-conjugated secondary antibody (Cell Signaling Technology, 7074S), anti-mouse HRP-conjugated antibody (Cell Signaling Technology, 7076S).

Techniques: Injection, Plasmid Preparation, Staining, Derivative Assay, Marker, Cell Culture, Imaging

Journal: Cell Death & Disease

Article Title: Involvement of p38 MAPK and MAPKAPK2 in promoting cell death and the inflammatory response to ischemic stress associated with necrotic glioblastoma

doi: 10.1038/s41419-025-07335-3

Figure Lengend Snippet: A A diagram describing the steps for analyzing and comparing differentially expressed genes in ischemic LN229 cells and the GBM PNZ relative to control LN229 cells and the GBM CT area, respectively. B , C Differentially expressed genes when comparing cells under the ischemic condition treated by VX745 ( B ) or MK25 ( C ) to those treated by a DMSO control. D Venn diagram showing overlapping genes in between indicated conditions. E , F Ingenuity Pathway Analysis identified upstream regulators under each indicated culture condition. G LN229 KRAS(G12D) or LN229 PIK3CA(H1047R) cells cultured under a regular (control) or the ischemic condition, treated by indicated inhibitors or DMSO were subjected to q-RT-PCR for TNFα mRNA. Ordinary two-way ANOVA test. H A diagram illustrating the involvement of the p38-MK2 and UPR signaling pathways in mediating ischemic stress-induced inflammatory response and cell death.

Article Snippet: The antibodies used for western blotting and immunohistochemistry staining included: ATF4 (Cell Signaling Technology, 11815), p-EIF2a (S51) (Cell Signaling Technology, 3398), β-actin (Cell Signaling Technology, 3700), p-p38 (T180/Y182) (Cell Signaling Technology, 4511), p38 (Cell Signaling Technology, 8690), p-MK2 (T334) (Cell Signaling Technology, 3007), MK2 (Cell Signaling Technology, 12155), p-ERK (Cell Signaling Technology, 4370), p-AKT(S473) (Cell Signaling Technology, 4060), anti-rabbit HRP-conjugated secondary antibody (Cell Signaling Technology, 7074S), anti-mouse HRP-conjugated antibody (Cell Signaling Technology, 7076S).

Techniques: Control, Cell Culture, Reverse Transcription Polymerase Chain Reaction, Protein-Protein interactions

Journal: Cell Death & Disease

Article Title: Protein phosphatase 4 maintains the survival of primordial follicles by regulating autophagy in oocytes

doi: 10.1038/s41419-024-07051-4

Figure Lengend Snippet: A – C Signaling studies in Ppp4c fl/fl and Ppp4c fl/fl ;GCre+ ovaries at PD4, showing levels of p-mTOR S2448 , p-AKT S473 , p-MAPKAPK-2 T334 , PPP4C, p-rpS6 S240/244 , P62 and LC3B. Level of β-actin was detected as internal control. D , E Western blots showing levels of p-mTOR S2448 , p-AKT S473 and p-MAPKAPK-2 T334 in Ppp4c fl/fl and Ppp4c fl/fl ;GCre+ ovaries at PD7. Mice were treated with (FA) or without (NT) fasting for 2 days before sacrifice. Level of β-actin was detected as internal control. F Western blot analysis of p62 in soluble lysates of PD7 ovaries. Level of β-actin was detected as internal control. G Western blot analysis of p62 in insoluble lysates of PD7 ovaries. Level of Ub-H2A was detected as internal control. H – J Signaling studies in Ppp4c fl/fl and Ppp4c fl/fl ;GCre+ primordial follicle oocytes at PD7, showing levels of p-mTOR S2448 , p-rpS6 S240/244 , p-MAPKAPK-2 T334 and LC3B. Level of β-actin was detected as internal control. The ovary lysates or oocytes were collected at least from three mice of each genotype. Each experiment was repeated at least 3 times.

Article Snippet: Mouse anti-γH2AX (#80312), rabbit anti-Caspase-3 (#9579), rabbit anti-p-AKT S473 (#4060), rabbit anti-p-mTOR S2448 (#5536), rabbit anti-p-MAPKAPK-2 T334 (#3007), rabbit anti-p-S6K T389 (#9234), rabbit anti-p-rpS6 S240/244 (#5364), rabbit anti-GAPDH (#5174), mouse anti-β-actin (#3700) and rabbit anti-α-tubulin (#2144) were purchased from Cell Signaling Technology, Inc. Rabbit anti-p62 (#nbp1-48320) was purchased from Novus.

Techniques: Control, Western Blot

Journal: Cell Death & Disease

Article Title: Protein phosphatase 4 maintains the survival of primordial follicles by regulating autophagy in oocytes

doi: 10.1038/s41419-024-07051-4

Figure Lengend Snippet: A – C Signaling studies in Ppp4c fl/fl and Ppp4c fl/fl ;GCre+ ovaries at PD4, showing levels of p-mTOR S2448 , p-AKT S473 , p-MAPKAPK-2 T334 , PPP4C, p-rpS6 S240/244 , P62 and LC3B. Level of β-actin was detected as internal control. D , E Western blots showing levels of p-mTOR S2448 , p-AKT S473 and p-MAPKAPK-2 T334 in Ppp4c fl/fl and Ppp4c fl/fl ;GCre+ ovaries at PD7. Mice were treated with (FA) or without (NT) fasting for 2 days before sacrifice. Level of β-actin was detected as internal control. F Western blot analysis of p62 in soluble lysates of PD7 ovaries. Level of β-actin was detected as internal control. G Western blot analysis of p62 in insoluble lysates of PD7 ovaries. Level of Ub-H2A was detected as internal control. H – J Signaling studies in Ppp4c fl/fl and Ppp4c fl/fl ;GCre+ primordial follicle oocytes at PD7, showing levels of p-mTOR S2448 , p-rpS6 S240/244 , p-MAPKAPK-2 T334 and LC3B. Level of β-actin was detected as internal control. The ovary lysates or oocytes were collected at least from three mice of each genotype. Each experiment was repeated at least 3 times.

Article Snippet: Mouse anti-γH2AX (#80312), rabbit anti-Caspase-3 (#9579), rabbit anti-p-AKT S473 (#4060), rabbit anti-p-mTOR S2448 (#5536), rabbit anti-p-MAPKAPK-2 T334 (#3007), rabbit anti-p-S6K T389 (#9234), rabbit anti-p-rpS6 S240/244 (#5364), rabbit anti-GAPDH (#5174), mouse anti-β-actin (#3700) and rabbit anti-α-tubulin (#2144) were purchased from Cell Signaling Technology, Inc. Rabbit anti-p62 (#nbp1-48320) was purchased from Novus.

Techniques: Control, Western Blot

Journal: Cell Death & Disease

Article Title: Protein phosphatase 4 maintains the survival of primordial follicles by regulating autophagy in oocytes

doi: 10.1038/s41419-024-07051-4

Figure Lengend Snippet: A , B Signaling studies in Ppp4c fl/fl and Ppp4c fl/fl ;GCre+ ovaries at PD4, showing levels of p-mTOR S2448 , p-rpS6 S240/244 and P62. Level of β-actin was detected as internal control. Ovaries were treated with 300 nm rapamycin (Rap) or not (NT) for 6 h before adding lysis buffer. C Representative images of p62 foci in untreated (NT) or rapamycin (Rap) treated primordial follicle oocytes of Ppp4c fl/fl and Ppp4c fl/fl ;GCre+ females at PD5. Scale bars: 5 μm. D Quantification of p62 foci in ( C ) was analyzed with ImageJ software. E Representative images of ULK1 foci in untreated (NT) or rapamycin (Rap) treated primordial follicle oocytes of Ppp4c fl/fl and Ppp4c fl/fl ;GCre+ females at PD5. Scale bars: 5 μm. F Quantification of ULK1 foci in E was analyzed with ImageJ software. Data are presented as mean ± s.e.m. *** P < 0.001. Each experiment was repeated at least 3 times.

Article Snippet: Mouse anti-γH2AX (#80312), rabbit anti-Caspase-3 (#9579), rabbit anti-p-AKT S473 (#4060), rabbit anti-p-mTOR S2448 (#5536), rabbit anti-p-MAPKAPK-2 T334 (#3007), rabbit anti-p-S6K T389 (#9234), rabbit anti-p-rpS6 S240/244 (#5364), rabbit anti-GAPDH (#5174), mouse anti-β-actin (#3700) and rabbit anti-α-tubulin (#2144) were purchased from Cell Signaling Technology, Inc. Rabbit anti-p62 (#nbp1-48320) was purchased from Novus.

Techniques: Control, Lysis, Software