Lamin b1 12586

The nucleus and cytoplasmic fractions of CCA cells were prepared as previously described^{25 (link)}. Cell lysates of the entire cells or nucleus fractions were collected by digesting with RIPA buffer and protease inhibitors (Sigma-Aldrich, USA). Protein concentration was determined by BCA kit (Thermo Fisher, USA). A total of 30 μg protein was loaded on 10% or 15% SDS-PAGE gels and transferred onto nitrocellulose membranes. The membranes were incubated with specific first antibodies and corresponding second antibody. The specific antibodies were listed below: Cleaved Caspase-3 #9664, PCNA #13110; MMP-3 #14351, ZEB1 #70512, E-Cadherin #3195, Vimentin #5741, Oct-4 #2750, LIN28A #3695, Nanog #3580, Sox2 #3579, β-Catenin #8480, Phospho-β-Catenin (Ser33/37/Thr41) # 9561, Phospho-GSK-3β (Ser9) #5558, GSK-3β#12456, c-Myc #9402, Survivin #2803, Lamin B1 #12586, and GAPDH #5174 were purchased from Cell Signaling Technology with a work concentration at 1: 1000. BCL9L #ab233736 was purchased from Abcam with a work concentration at 1: 500. The second antibodies were goat anti-rabbit IgG HRP-linked antibody (Cell signaling #7074; 1:4000) and sheep anti-mouse IgG-HRP (GE/Amershan #NXA931; 1: 5000).

CSR (purity > 99%, determined by HPLC) was purchased from Extrasynthese (Genay Cedex, France). Indomethacin, crystal violet, phosphate‐buffered saline (PBS) and dimethylsulfoxide (DMSO) were purchased from Sigma Chemical Co. (St. Louis, MO, USA). CCK-8 (Cell Counting Kit-8) was purchased from TransGen Biotech (Beijing, China). Recombinant human IL-6 and recombinant human soluble IL-6 receptor α (sIL-6R) were purchased from PeproTech (Rocky Hill, NY, USA). Janus kinase 2 (JAK2, ab39636) and phospho-JAK2 (Tyr1007/1008, ab32101) monoclonal antibodies, HRP-conjugated secondary antibodies (ab7090, ab97040), and Annexin V-FITC Apoptosis Staining / Detection Kit (#ab14085) were purchased from Abcam (Cambridge, CB2 0AX, UK). B-cell lymphoma 2 (Bcl-2, sc-7382) and GAPDH (sc-365062) monoclonal antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Myeloid cell leukemia 1 (Mcl-1, #94296), caspase-9 (#9508), caspase-3 (#9662), STAT3 (#12640), phospho-STAT3 (Tyr705, #9131) and lamin B1 (#12586) monoclonal antibodies were obtained from Cell Signaling Technology (Boston, MA, USA). All other chemicals used were purchased from Sigma and were of analytical grade.