anti p mk2 thr334  (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