anti s6k rabbit polyclonal  (Cell Signaling Technology Inc)

Journal: The Journal of Biological Chemistry

Article Title: Metformin attenuates O-GlcNAc modification to improve renal function via AMPK/mTOR signaling in diabetic nephropathy

doi: 10.1016/j.jbc.2025.110909

Figure Lengend Snippet: AMPK/mTOR signaling pathway influences mesangial cell damage via O-GlcNAcylation under high-glucose treatment. A , representative Western blot images of four groups of O-GlcNAc, OGT, and OGA and AMPK/mTOR pathway proteins according to whether or not AMPK agonist and inhibitor were added, n = 3. B , representative quantification of O-GlcNAc, OGT, and OGA in MCs of four groups according to whether or not an AMPK agonist and inhibitor was added, n = 3. C , mRNA expression of OGT and OGA assessed by RT-PCR, n = 3. D–F , representative quantification of p-AMPK, p-mTOR, and p-S6K in MCs of four groups according to whether or not an AMPK agonist and inhibitor was added, n = 3. G , the effect of AMPK agonist and inhibitor treatment on MCs volume under high glucose conditions, n = 3. H , the effect of AMPK agonist and inhibitor treatment on MCs viability under high-glucose conditions, n = 3. Data are presented as mean ± SD. Significance analysis was performed with one-way ANOVA (multiple comparisons). ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; and ∗∗∗∗ p < 0.0001. AMPK, AMP-activated protein kinase; CC, compound C; MC, mesangial cell; Met, Metformin; mTOR, mammalian target of rapamycin; OGA, O-GlcNAcase; OGT, O-GlcNAc transferase.

Article Snippet: The following primary antibodies were used to detect the respective proteins after incubation overnight at 4 °C in 5% BSA: O-GlcNAc (#65292-1-lg, proteintech, 1:1000), OGT (# R25212 , Zen Bio, 1:1000), OGA (# R24971 , Zen Bio, 1:1000), Phospho-AKT (Ser473) (#310021, Zen Bio, 1:1000), AKT (# R23412 , Zen Bio, 1:1000), Phospho-mTOR (Ser2448) (#381557, Zen Bio, 1:1000), mTOR (#380411, Zen Bio, 1:1000), Phospho-AMPK alpha 1/2 (Thr183/Tyr172) (#340763, ZenBio, 1:1000), AMPK alpha (#23314, ZenBio, 1:1000), Phospho-P70 S6K (Thr389) (#AF5899, Beyotime, 1:1000), P70 S6K (#AF0258, Beyotime, 1:1000), and GAPDH (#TA802519, Origene, 1:1000).

Techniques: Western Blot, Expressing, Reverse Transcription Polymerase Chain Reaction