Journal: Open Medicine

Article Title: ADAR1 plays a protective role in proximal tubular cells under high glucose conditions by attenuating the PI3K/AKT/mTOR signaling pathway

doi: 10.1515/med-2024-1037

Figure Lengend Snippet: Detect the expression characteristics of ADAR1 in proximal renal tubular cells of db/db mice with spontaneous type 2 diabetes mellitus. (a) Detect FBG, (b) weight, (c) food intake, and (d) water intake of db/db mice with spontaneous type 2 diabetes mellitus and normal db/m mice. (e)–(i) IHC assays were presented for detecting the expression of (f) ADAR1, (g) p-PI3K, (h) p-AKT1, and (i) p-mTOR. (j)–(n) Western blot was presented for detecting the expression of (k) ADAR1, (l) p-PI3K, (m) p-AKT1, and (n) p-mTOR. Bar, 20 μm. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Primary antibodies and corresponding dilution rate are as follows: ADAR1 (1:100, sc-271854; SANTA CRUZ, USA), PI3K (1:1,000, AF6241; Affinity, China), phospho-PI3K (1:500, AF3241; Affinity, China), phospho-AKT (1:500, 80455-1-RR; Proteintech, China), AKT (1:1,000, 10176-2-AP; Proteintech, China), phospho-mTOR (1:500, 67778-1-Ig; Proteintech, China), mTOR (1:1,000, 28273-1-AP; Proteintech, China), cyclin D1 (1:1,000, 26939-1-AP; Proteintech, China), Ki-67 (1:1,000, 28074-1-AP; Proteintech, China), and β-actin (1:2,000, 20536-1-AP; Proteintech, China).

Techniques: Expressing, Western Blot

Journal: Open Medicine

Article Title: ADAR1 plays a protective role in proximal tubular cells under high glucose conditions by attenuating the PI3K/AKT/mTOR signaling pathway

doi: 10.1515/med-2024-1037

Figure Lengend Snippet: ADAR1 regulates SOD and MDA. (a) ADAR1-overexpression plasmids regulated ADAR1 mRNA expression level. The effects of different concentrations of (b) ADAR1 inhibitor 8-azaadenosine, (c) PI3K inhibitor LY294002, and (d) mTOR inhibitor rapamycin on the proliferation of HK-2 cells were screened by CCK8. (e) and (f) Detect SOD activity and MDA content under high-glucose conditions, ADAR1 inhibitor 8-azaadenosine, and ADAR1-overexpression. *** p < 0.001.

Article Snippet: Primary antibodies and corresponding dilution rate are as follows: ADAR1 (1:100, sc-271854; SANTA CRUZ, USA), PI3K (1:1,000, AF6241; Affinity, China), phospho-PI3K (1:500, AF3241; Affinity, China), phospho-AKT (1:500, 80455-1-RR; Proteintech, China), AKT (1:1,000, 10176-2-AP; Proteintech, China), phospho-mTOR (1:500, 67778-1-Ig; Proteintech, China), mTOR (1:1,000, 28273-1-AP; Proteintech, China), cyclin D1 (1:1,000, 26939-1-AP; Proteintech, China), Ki-67 (1:1,000, 28074-1-AP; Proteintech, China), and β-actin (1:2,000, 20536-1-AP; Proteintech, China).

Techniques: Over Expression, Expressing, Activity Assay

Journal: Open Medicine

Article Title: ADAR1 plays a protective role in proximal tubular cells under high glucose conditions by attenuating the PI3K/AKT/mTOR signaling pathway

doi: 10.1515/med-2024-1037

Figure Lengend Snippet: ADAR1 affects PI3K/AKT/mTOR signaling pathway under the high-glucose environment. (a)–(j) Expressions of (b) ADAR1, (c) PI3K, (d) p-PI3K, (e) AKT1, (f) p-AKT1, (g) mTOR, (h) p-mTOR, (i) cyclin D1, and (j) Ki67 were measured by western blot in HK-2 cells or ADAR1-overexpressed HK-2 cells under the high-glucose environment. *** p < 0.001.

Article Snippet: Primary antibodies and corresponding dilution rate are as follows: ADAR1 (1:100, sc-271854; SANTA CRUZ, USA), PI3K (1:1,000, AF6241; Affinity, China), phospho-PI3K (1:500, AF3241; Affinity, China), phospho-AKT (1:500, 80455-1-RR; Proteintech, China), AKT (1:1,000, 10176-2-AP; Proteintech, China), phospho-mTOR (1:500, 67778-1-Ig; Proteintech, China), mTOR (1:1,000, 28273-1-AP; Proteintech, China), cyclin D1 (1:1,000, 26939-1-AP; Proteintech, China), Ki-67 (1:1,000, 28074-1-AP; Proteintech, China), and β-actin (1:2,000, 20536-1-AP; Proteintech, China).

Techniques: Western Blot

Journal: Open Medicine

Article Title: ADAR1 plays a protective role in proximal tubular cells under high glucose conditions by attenuating the PI3K/AKT/mTOR signaling pathway

doi: 10.1515/med-2024-1037

Figure Lengend Snippet: ADAR1 affects PI3K/AKT/mTOR signaling pathway under high-glucose environment and LY294002 treatment. (a)–(h) Expressions of (b) ADAR1 (c) PI3K, (d) p-PI3K, (e) AKT1, (f) p-AKT1, (g) mTOR, and (h) p-mTOR were measured by western blot in HK-2 cells or ADAR1-overexpressed HK-2 cells under high-glucose environment and LY294002 treatment. ** p < 0.01; *** p < 0.001.

Article Snippet: Primary antibodies and corresponding dilution rate are as follows: ADAR1 (1:100, sc-271854; SANTA CRUZ, USA), PI3K (1:1,000, AF6241; Affinity, China), phospho-PI3K (1:500, AF3241; Affinity, China), phospho-AKT (1:500, 80455-1-RR; Proteintech, China), AKT (1:1,000, 10176-2-AP; Proteintech, China), phospho-mTOR (1:500, 67778-1-Ig; Proteintech, China), mTOR (1:1,000, 28273-1-AP; Proteintech, China), cyclin D1 (1:1,000, 26939-1-AP; Proteintech, China), Ki-67 (1:1,000, 28074-1-AP; Proteintech, China), and β-actin (1:2,000, 20536-1-AP; Proteintech, China).

Techniques: Western Blot

Journal: Open Medicine

Article Title: ADAR1 plays a protective role in proximal tubular cells under high glucose conditions by attenuating the PI3K/AKT/mTOR signaling pathway

doi: 10.1515/med-2024-1037

Figure Lengend Snippet: ADAR1 affects PI3K/AKT/mTOR signaling pathway under high-glucose environment and rapamycin treatment. (a)–(h) Expressions of (a) ADAR1 (c) PI3K, (d) p-PI3K, (e) AKT1, (f) p-AKT1, (g) mTOR, and (h) p-mTOR were measured by western blot in HK-2 cells or ADAR1-overexpressed HK-2 cells under high-glucose environment and rapamycin treatment. *** p < 0.001.

Article Snippet: Primary antibodies and corresponding dilution rate are as follows: ADAR1 (1:100, sc-271854; SANTA CRUZ, USA), PI3K (1:1,000, AF6241; Affinity, China), phospho-PI3K (1:500, AF3241; Affinity, China), phospho-AKT (1:500, 80455-1-RR; Proteintech, China), AKT (1:1,000, 10176-2-AP; Proteintech, China), phospho-mTOR (1:500, 67778-1-Ig; Proteintech, China), mTOR (1:1,000, 28273-1-AP; Proteintech, China), cyclin D1 (1:1,000, 26939-1-AP; Proteintech, China), Ki-67 (1:1,000, 28074-1-AP; Proteintech, China), and β-actin (1:2,000, 20536-1-AP; Proteintech, China).

Techniques: Western Blot

Journal: Current Oncology

Article Title: Overcoming of Microenvironment Protection on Primary Chronic Lymphocytic Leukemia Cells after Treatment with BTK and MDM2 Pharmacological Inhibitors

doi: 10.3390/curroncol28040223

Figure Lengend Snippet: Ibrutinib plus nutlin-3 combination mediates mitochondria-dependent apoptosis through inhibition of phospho-BAD. Patients’ derived B-CLL cells were grown in vitro with stroma or with CpG-ODN and CD40L and exposed to ibrutinib (10 µM), nutlin-3 (10 µM), ibrutinib+nutlin-3 (10 µM) for 24 h or left untreated as control. In ( A ), mitochondrial activity, evaluated using MitoTracker ® Red staining on B-CLL cells stimulated with CpG-ODN and CD40L, was analyzed by flow cytometry and expressed as fold of modulation with respect to the MFI value of the untreated B-CLL cultures set at 1. Results are reported as mean ± SD of five independent experiments. Statistical analysis was performed by ANOVA followed by Bonferroni post-hoc test. Asterisks indicate: *, p < 0.05; **, p < 0.01; ***, p < 0.001. In ( B ), Western blotting analyses of pro-caspase 9 and pro-caspase 3 protein levels are shown. In C, Western blotting analyses of BAD phosphorylation in ser136, total BAD, BAX, Akt phosphorylation in ser473 and total Akt protein levels are shown. In ( B , C ), results are shown for representative B-CLL patients and β-actin levels are shown as loading control. In ( D ), schematic representation of early molecular events that drive the mitochondria-dependent apoptosis following the combination treatment ibrutinib plus nutlin-3.

Article Snippet: The following antibodies were used for immunoblotting onto nitrocellulose filters: anti c-MYC, anti-phospho p53-Ser15, anti-phospho p53-Ser392, BAD, phospho BAD-Ser136 and p14ARF from Cell Signaling (Danvers, MA, USA); anti-p53, anti-MDM2, anti-p21, anti-PUMA, anti-procaspase 9 and anti-procaspase 3 from Santa Cruz Biotechnology (Santa Cruz, CA, USA); anti-BAX and anti-Akt from BD (BD Biosciences); phospho Akt-Ser473 from Merck Millipore (KGaA, Darmstadt, Germany) and anti β-actin, from Sigma-Aldrich (Merck KGaA).

Techniques: Inhibition, Derivative Assay, In Vitro, Control, Activity Assay, Staining, Flow Cytometry, Western Blot, Phospho-proteomics