Phospho jak2 tyr1007 1008

Myricitrin was obtained from Aladdin Industrial Corporation (Shanghai, China). Monoclonal and polyclonal antibodies against iNOS, COX-2, JNK, phospho-JNK (Thr183/Tyr185), p38, phospho-p38 (Thr180/Tyr182), ERK1/2, phospho-ERK1/2 (Thr202/Tyr204), JAK1, phospho-JAK1 (Tyr1022/1023), JAK2, phospho-JAK2 (Tyr1007/1008), STAT1, phospho-STAT1 (Tyr701), STAT3, phospho-STAT3 (Tyr705), phospho-STAT3 (Ser727), TBP, gp91^phox, Na/K ATPase-α1, and GAPDH were purchased from Cell Signaling Technology (Beverly, MA, USA). Antibody to p47^phox was obtained from Santa Cruz Biotechnology (CA, USA). All secondary antibodies used for western blotting were purchased from LI-COR Biosciences (Lincoln, NE, USA). LPS (from Escherichia coli 0111:B4), NAC, and DAPI were obtained from Sigma-Aldrich (St. Louis, MO, USA). CCK-8 was purchased from KeyGen Biotech (Nanjing, JS, China). CM-H2DCFDA was obtained from Invitrogen (Carlsbad, CA, USA). Ruxolitinib and apocynin were purchased from Selleck Chemicals (Houston, TX, USA). All ELISA kits were purchased from R&D Systems China Co. Ltd. (Shanghai, China).

This has been performed as previously described [32 (link)]. Antibodies directed against alpha smooth muscle actin (α-SMA), Twist-1, Vimentin (RV202), N-cadherin, transforming growth factor beta 1 (TGF-β1), VEGF-A (ab46154) and IL-6 were purchased from Abcam (Cambridge, MA); Lin28B (D4H1), AKT (C73H10), p-AKT (Thr308), STAT3, pSTAT3-Tyr705 (D3A7), Snail (C15D3), E-cadherin (24E10), EpCam (UV1D9), JAK-2 (D2E12), phospho-JAK-2 (TYR1007/1008), Cyclin D1 (2922), Akt, phospho-Akt (193H12), NF-κB, ERK1/2 (137F5), phospho-ERK1/2, MMP-9, HIF-1α, Glyceraldehydes-3-phosphate dehydrogenase (GAPDH, 14C10) and β-Actin from Cell Signaling (Danvers, MA); ZEB-1 (4C4) from Abnova (Taipei, Taiwan), ALDH1A1 from Sigma (CA, USA) and IL-8 (EpR1116(2)) from Origene (Rockville, MD, USA).

For western blot analysis, differentially treated cells were washed with cold PBS and then lysed in lysis buffer containing protease and phosphatase inhibitor. Insoluble cell fragments were removed by centrifugation. Cells were separated by SDS-PAGE electrophoresis and next transferred to PVDF membranes. Membranes were incubated overnight with the primary antibodies against phospho-AKT (Cell Signaling Technologies), ERK-1/2 (Cell Signaling Technologies), phosphor-ERK-1/2 (Cell Signaling Technologies), STAT3 (Cell Signaling Technologies), phospho-STAT3 (Cell Signaling Technologies), phospho-IGF-I (Cell Signaling Technologies, Danvers, MA, USA), JAK1, phospho-Jak1 (Tyr1022/1023) (Cell Signaling Technologies), JAK2 (Cell Signaling Technologies), phospho-Jak2 (Tyr1007/1008) (Cell Signaling Technologies), IGF-I Receptor β (Cell Signaling Technologies), AKT (Cell Signaling Technologies), epithelial-mesenchymal transition (EMT) antibody sampler kit (Cell Signaling Technologies). Fluorescent secondary anti-mouse (Thermo Scientific, Waltham, USA) and anti-rabbit (Thermo Scientific, Waltham, USA) antibodies were used and were detected using Odyssey Sa infrared imager (LI-COR Biosciences, Lincoln, USA).