Fig 1: JNK knockdown alleviates CESCs degeneration by inhibiting phosphorylation of Raptor. After transfection of si-JNK/si-NC in TL-induced CESCs, (A): mRNA level of JNK in CESCs after transfection of si-JNK was detected by qRT-PCR; (B): changes in total Raptor and p-Raptor protein levels were detected by WB; (C): cell activity was detected by MTT assay; (D): apoptosis was detected by Annexin V-FITC/PI double staining assay; (E): mRNA expressions of cartilage markers ACAN, COL-2A, and Sox9 were detected by RT-qPCR; (F): WB was used to detect the protein levels of autophagy markers LC3-II, Beclin-1, and p62. Cell experiment was conducted 3 times. Cell experiment was conducted 3 times. Data was expressed as mean ± standard deviation. One-way ANOVA was used for data comparison among multiple groups and Tukey’s test was used for the post hoc test. * P < .05, ** P < .01, *** P < .001.
Fig 2: Raptor expression in ERα-positive, ERα-negative, and tamoxifen-resistant breast cancer cells. Raptor expression was higher in ERα-positive than in ERα-negative cell lines by Western blot analysis (a). Comparing ERα-positive with ERα-negative cells shows raptor in the ERα-positive but not in the ERα-negative nuclei as observed by in situ immunofluorescence (b). Raptor presence increased in the nucleus (N) in MCF7 tamoxifen-resistant cells (TamR) compared with MCF7 parental cells (Control), and changes in mTORC1 and Akt signaling were observed (c)
Fig 3: The mechanism underlying the RPTOR-promoted NSCLC-BM via the SPHK2/S1P/STAT3 signaling pathway. RPTOR regulated the transcriptional activation of the ceramidase SPHK2 by binding to the transcription factor YY1. SPHK2 catalyzed the degradation of ceramide to S1P. The latter, in turn, activated the STAT3 signaling pathway by binding to S1PR1 (solid lines), altering the microenvironment and enhancing the permeability of the BBB (dotted lines), resulting in the BM of NSCLC
Fig 4: The SASP is inhibited by rapamycin in endothelial cells (ECs) subjected to SIPS. a Real-time PCR analysis of SASP factors using cDNA derived from human coronary artery endothelial cells (HCAECs) at 72 h in Fig. 1a. The results are shown after normalization to values obtained from control HCAECs (value = 1). Results are presented as means ± SD from three independent experiments. ap < 0.05 vs. the Ctr and bp < 0.05 vs. the H2O2-treated HCAECs. b Immunoblotting for the levels of mTOR, RICTOR, RAPTOR, and MAPKAPK2 was performed using cell lysates and mTOR immunoprecipitates from HCAECs at 72 h in Fig. 1a. β-ACTIN was used as a loading control. The histogram shows mean densitometric readings for the proteins normalized to β-ACTIN or mTOR from three independent experiments. ap < 0.01 vs. the Ctr and bp < 0.01 vs. the H2O2-treated HCAECs. Control (Ctr): untreated cells. SIPS, stress-induced premature senescence. SASP, senescence-associated secretory phenotype
Fig 5: PAI reduces autophagic flux by promoting the mTOR complex pathways. (A) LM8 cells were loaded with Cyto-ID and treated with PAI (50%) or rapamycin (positive control) for 18 h and analyzed by flow cytometry. (B) LM8 cells were treated with PAI (50%) for the indicated time and then analyzed for the expression of p70-S6 kinase, Rictor, Raptor, p62, and LC3-I/II, and their phosphorylated forms by western blotting analysis using specific antibodies. GAPDH was used as a loading control. See Supplementary Figures 3 , 4 for examples of uncropped images and quantification for each antibody.
Supplier Page from Abcam for Anti-Raptor antibody [EP539Y]