Journal: Heliyon

Article Title: AP-1 inhibitor induces ferroptosis via the PI3K/AKT pathway in multiple myeloma cells

doi: 10.1016/j.heliyon.2024.e34397

Figure Lengend Snippet: The PI3K/AKT signaling pathway is involved in AP-1 inhibitor-induced ferroptosis. (A) MM cells were exposed to T-5224 (20 μМ) for 48 h. The protein levels of p -AKT(Ser473), total AKT, p-PI3K (Tyr317), and total PI3K were assessed by western blotting. (B) MM cells were exposed to T-5224 (20 μМ) for 48 h. The protein levels of Bcl2 and Bax in MM cells were assessed by western blotting. (C) MM cells were exposed to T-5224 in the presence or absence of Fer-1 (5 μM) for 48 h, after which the protein levels of p -AKT(Ser473), AKT, p-PI3K (Tyr317), and PI3K were assessed by western blotting. (D) MM cells were exposed to T-5224(20 μM) in the presence or absence of 740 Y–P (20 μM) for 48 h. The protein levels of GPX4 and SLC7A11 in MM cells were assessed by western blotting. β-actin served as the protein loading control. The results are presented as the mean ± standard deviation (SD).

Article Snippet: An anti-β-actin antibody was acquired from Proteintech.Anti-total-PI3K and anti-phospho-PI3KCA (Tyr317) antibodies were acquired from Bioss Co., Ltd. (Beijing, China).

Techniques: Western Blot, Control, Standard Deviation

Journal: Frontiers in Cell and Developmental Biology

Article Title: Network Pharmacology and Experimental Validation Reveal the Effects of Chidamide Combined With Aspirin on Acute Myeloid Leukemia-Myelodysplastic Syndrome Cells Through PI3K/AKT Pathway

doi: 10.3389/fcell.2021.685954

Figure Lengend Snippet: CDM combined with ASA inhibited the activation of PI3K/AKT pathway in AML-MDS cells. (A) Western blot analysis of PI3K, p-PI3K AKT, p-AKT. (B) Western blotting of PI3K, p-PI3K, AKT and p-AKT. (C) Western blotting of CDK2, CDK4 and p21. (D) Western blotting of Bcl-2, caspase-3 and cleaved caspase-3. IGF-1 reversed the effect of CDM combined with ASA on AML-MDS cells. β-actin served as a loading control. Data are mean ± SD of three independent experiments. “ ∗ ” indicates a significant difference relative to the control group ( ∗ p < 0.05), “&” indicates a significant difference relative to ASA-treated group (& p < 0.05), “NS” indicates no significant difference relative to the control group.

Article Snippet: PI3K (bsm-33219M), p-PI3K (AB1235888), and Caspase-3 (bs-0081R) from Bioss (Beijing, China).

Techniques: Activation Assay, Western Blot

Journal: Frontiers in Cell and Developmental Biology

Article Title: Network Pharmacology and Experimental Validation Reveal the Effects of Chidamide Combined With Aspirin on Acute Myeloid Leukemia-Myelodysplastic Syndrome Cells Through PI3K/AKT Pathway

doi: 10.3389/fcell.2021.685954

Figure Lengend Snippet: A schematic representation of the proposed pathway responsible for CDM combined with ASA in AML-MDS cells. (A) The predictive pathways of CDM and ASA in AML-MDS through network pharmacology. (B) CDM combined with ASA could inhibited the activation of PI3K/AKT signaling pathways, and then affected the expression of cell cycle and apoptosis-related proteins to induce cell cycle arrest and apoptosis in AML-MDS cells through experimental validation.

Article Snippet: PI3K (bsm-33219M), p-PI3K (AB1235888), and Caspase-3 (bs-0081R) from Bioss (Beijing, China).

Techniques: Activation Assay, Expressing

Journal: Brain

Article Title: MMP13 inhibition rescues cognitive decline in Alzheimer transgenic mice via BACE1 regulation

doi: 10.1093/brain/awy305

Figure Lengend Snippet: MMP13-mediated BACE1 regulation involves PI3K signalling and is unrelated to BACE1 transcription and protein degradation. (A) BACE1 protein levels in SH-SY5Y cells treated with 5 μM CL82198 (CL) for 48 h in the absence (CTRL) or presence of the RTK inhibitor 341610 (4 μM). (B) SH-SY5Y cells were transfected with either control or MMP13 vector for 48 h in the absence or presence of 5 μM CL82198 (CL) and 4 μM 341610. (C) SH-SY5Y cells were transfected with either control or MMP13 vector for 48 h in the absence or presence of 4 μM 341610, and PI3K activity was measured by ELISA. (D) p-Akt protein in SH-SY5Y cells transfected with either control or MMP13 vector for 48 h in the absence or presence of 4 μM 341610. (E) BACE1 protein levels in SH-SY5Y cells treated with 5 μM CL82198 (CL) for 48 h in the absence (CTRL) or presence of the PI3K inhibitor LY294002 (LY, 5 and 10 μM). (F) SH-SY5Y cells were transfected with either control or MMP13 vector for 48 h in the absence or presence of 5 μM CL82198 (CL) and LY294002 (LY, 5 and 10 μM). (G) Relative BACE1 mRNA levels in SH-SY5Y cells treated with 5 μM CL82198 (CL) for 48 h in the absence (CTRL) or presence of LY294002 (LY, 5 and 10 μM). (H) Relative Bace1 mRNA levels in HT22 cells treated with vehicle (CTRL) or Mmp13 shRNA-1 for 72 h. (I and J) BACE1 protein levels in SH-SY5Y cells treated with 5 μM CL82198 (CL) for 48 h in the absence (CTRL) or presence of 1 μM MG132 (G) or 100 μM CQ (H). (K) BACE1 protein levels in SH-SY5Y cells treated with 5 μM CL82198 (CL) for 48 h in the absence (CTRL) or presence of the transcriptional inhibitor actinomycin D (ActD, 0.1 μM) or the protein synthesis inhibitor cycloheximide (CHX, 5 μM). All values were normalized to CTRL (1.0) within each experiment. The error bars are the SEM. n.s. = no significant difference; *P < 0.05, **P < 0.01, ***P < 0.001 (ANOVA, n = 3 or 4).

Article Snippet: The monoclonal or polyclonal antibodies used were BACE1 (Abcam, 1:1000); APP-full length and β-CTF, α-CTF (A8717, Sigma, 1:3000); sAPPα (6E10, Covance, 1:1000); sAPPβ (Covance, 1:500); ADAM10 (Abcam, 1:1000); phospho-eIF4B (Ser422) (Cell Signaling Technology, 1:1000); eIF4B (Proteintech, 1:2000); MMP13 (Santa Cruz, 1:500); PSEN1 (Proteintech, 1:500); neprilysin (Proteintech, 1000); IDE (Proteintech, 1:1000); phospho-Akt (Ser473) (Proteintech, 1:500); Akt (Proteintech, 1:1000); PI3K (phosphatidylinositide 3-kinase; Proteintech, 1:500); GAPDH (Proteintech, 1:10 000); β-actin (Proteintech, 1:2000).

Techniques: Transfection, Plasmid Preparation, Activity Assay, Enzyme-linked Immunosorbent Assay, shRNA

Journal: Brain

Article Title: MMP13 inhibition rescues cognitive decline in Alzheimer transgenic mice via BACE1 regulation

doi: 10.1093/brain/awy305

Figure Lengend Snippet: Mmp13 knockdown ameliorates Alzheimer’s disease-associated pathology in APP/PS1 mice. (A) Representative western blots and bar plot summary of MMP13 and PI3K expression in the prefrontal cortex of control (n = 13) and Alzheimer’s disease patients (n = 13). Data in (i and ii) and in (iii–v) were from samples provided by Sydney Brain Bank and NIH NeuroBiobank, respectively. (B) Immunohistochemical images and bar plot summary of neuritic plaques in the hippocampi of wild-type (WT) and APP/PS1 mice (AD) treated with vehicle (CTRL) and Mmp13 shRNA-1 (shMMP13-1). Neuritic plaques were probed with the amyloid-β-specific monoclonal antibody 6E10 (n = 6). (C) Representative western blots and bar plot summary of MMP13, APP, ADAM10, BACE1, PSEN1, βCTF, αCTF, sAPPβ and sAPPα expression in the hippocampi of wild-type (n = 8), wild-type with Mmp13 shRNA-1 (WT-shMMP13-1, n = 8), Alzheimer’s disease (n = 8) and Alzheimer’s disease with Mmp13 shRNA-1 mice (AD-shMMP13-1, n = 8). The soluble fractions were used to detect sAPPβ and sAPPα with sAPPβ and 6E10 antibodies. (D) Representative western blots and bar plot summary of MMP13 and BACE1 expression in the hippocampi of wild-type (n = 6), wild-type with Mmp13 shRNA-2 (WT-shMMP13-2, n = 6), Alzheimer’s disease (n = 6) and Alzheimer’s disease with Mmp13 shRNA-2 mice (AD-shMMP13-2, n = 6). (E) ELISAs were used to measure soluble and insoluble amyloid-β40/42 levels in the brain homogenates of wild-type (n = 6), WT-shMMP13-1 (n = 6), Alzheimer’s disease (n = 6) and AD-shMMP13-1 (n = 6) mice. (F) Representative western blots and bar plot summary of p-eIF4B, eIF4B, neprilysin and insulin-degrading enzyme (IDE) levels in the hippocampi of wild-type (n = 8), WT-shMMP13-1 (n = 8), Alzheimer’s disease (n = 8) and AD-shMMP13-1 (n = 8) mice. All values were normalized to CTRL or wild-type (1.0) within each experiment. The error bars are the SEM. n.s. = no significant difference; *P < 0.05, **P < 0.01, ***P < 0.001 (ANOVA).

Article Snippet: The monoclonal or polyclonal antibodies used were BACE1 (Abcam, 1:1000); APP-full length and β-CTF, α-CTF (A8717, Sigma, 1:3000); sAPPα (6E10, Covance, 1:1000); sAPPβ (Covance, 1:500); ADAM10 (Abcam, 1:1000); phospho-eIF4B (Ser422) (Cell Signaling Technology, 1:1000); eIF4B (Proteintech, 1:2000); MMP13 (Santa Cruz, 1:500); PSEN1 (Proteintech, 1:500); neprilysin (Proteintech, 1000); IDE (Proteintech, 1:1000); phospho-Akt (Ser473) (Proteintech, 1:500); Akt (Proteintech, 1:1000); PI3K (phosphatidylinositide 3-kinase; Proteintech, 1:500); GAPDH (Proteintech, 1:10 000); β-actin (Proteintech, 1:2000).

Techniques: Western Blot, Expressing, Immunohistochemical staining, shRNA

Journal: Brain

Article Title: MMP13 inhibition rescues cognitive decline in Alzheimer transgenic mice via BACE1 regulation

doi: 10.1093/brain/awy305

Figure Lengend Snippet: Schematic diagram depicting the possible mechanisms through which MMP13 regulates BACE1 translation. PI3K signalling downstream of MMP13 promotes eIF4B phosphorylation, which in turn facilitates the 5′UTR-dependent BACE1 translation. Consequently, increased BACE1 protein levels enhance amyloid-β production and impair cognitive function. The inhibition of MMP13 by CL82198 results in reduced BACE1 and amyloid-β accumulation in the brain, which contributes to the improved learning and memory functions in APP/PS1 mice. AD = Alzheimer;s disease; WT = wild-type.

Article Snippet: The monoclonal or polyclonal antibodies used were BACE1 (Abcam, 1:1000); APP-full length and β-CTF, α-CTF (A8717, Sigma, 1:3000); sAPPα (6E10, Covance, 1:1000); sAPPβ (Covance, 1:500); ADAM10 (Abcam, 1:1000); phospho-eIF4B (Ser422) (Cell Signaling Technology, 1:1000); eIF4B (Proteintech, 1:2000); MMP13 (Santa Cruz, 1:500); PSEN1 (Proteintech, 1:500); neprilysin (Proteintech, 1000); IDE (Proteintech, 1:1000); phospho-Akt (Ser473) (Proteintech, 1:500); Akt (Proteintech, 1:1000); PI3K (phosphatidylinositide 3-kinase; Proteintech, 1:500); GAPDH (Proteintech, 1:10 000); β-actin (Proteintech, 1:2000).

Techniques: Inhibition