Anti cleaved poly adenosine diphosphate ribose polymerase parp

Seventy-two hours post-transfection, total protein was extracted from SH-SY5Y and SK-N-SH cells. Western blotting was performed to detect the expression of target proteins. The primary antibodies, including anti-extracellular signal-regulated protein kinase 1/2 (ERK1/2), anti-phospho (p)-ERK1/2 (Thr202/Tyr204), anti-c-Jun N-terminal kinase (JNK), anti-p-JNK (Thr183/Tyr185), anti-p38, anti-p-p38 (Thr180/Tyr182), anti-phosphatidylinositol 3-kinase (PI3K), anti-p-PI3K (Tyr458/Tyr199), anti-Akt, anti-p-Akt (Ser473), anti-mammalian target of rapamycin (mTOR), and anti-p-mTOR (Ser2448) antibodies were purchased from Cell Signaling Technology (CST, Danvers, MA, USA). Anti-PCNP, Anti-B-cell lymphoma-2 (Bcl-2), anti-Bcl-2-associated X protein (Bax), anti-B-cell lymphoma-extra large (Bcl-xl), anti-Bcl-xl/Bcl-2-associated death promoter (Bad), anti-cleaved caspase-3, anti-cleaved caspase-8, anti-cleaved caspase-9, anti-Cleaved poly adenosine diphosphate-ribose polymerase (PARP), and anti-GAPDH antibodies were purchased from ProteinTech (Chicago, IL, USA). The horseradish peroxidase-conjugated secondary antibody was purchased from Cell Signaling Technology. The results were normalized to the level of GAPDH. The reaction was visualized using an enhanced chemiluminescence system (Thermo Fisher Scientific, Rockford, IL, USA). The bands were semi-quantified with Image J software.

Total protein was extracted from human HCC cells. Western blotting was used to determine the expression levels of relevant proteins. The primary antibodies include anti-Cyclin E1, anti-Cyclin D1, anti-cyclin-dependent kinase (CDK) 2, anti-CDK4, anti-p27, anti-p21, anti-AKT, anti-phospho (p)-AKT (Ser473), anti-glycogen synthase kinase-3 beta (Gsk‐3β), anti-p-Gsk-3β (Ser9), anti-β-catenin, anti-p-β-catenin (Ser552), anti-beclin-1, anti-p62, anti-LC3A/B, anti-Smad2, anti-p-Smad2 (Ser465/467), anti-Smad3, anti-p-Smad3 (Ser423/425), and anti-transforming growth factor-beta (TGF‐β) antibodies, as well as the horseradish peroxidase-conjugated secondary antibody obtained from Cell Signaling Technology (CST, Danvers, MA, USA). Anti-B-cell lymphoma-2 (Bcl-2), anti-B-cell lymphoma-extra large (Bcl-xl), anti-Bcl-2-associated X protein (Bax), anti-Bcl-xl/Bcl-2-associated death promoter (Bad), anti-cleaved caspase (cas)-3, anti-cleaved cas-9, anti-cleaved poly adenosine diphosphate‐ribose polymerase (PARP), and anti-β-actin antibodies were obtained from ProteinTech (Chicago, IL, USA). The protein bands were detected with the enhanced chemiluminescence system (Thermo, Rockford, IL, USA) and semiquantified by ImageJ software.