OBJECTIVE To investigate the effects of celastrol （CSL） on drug resistance of liver cancer cells. METHODS Human liver cancer lenvatinib （Len）-resistant cells Huh7/Len were constructed and divided into control group， CSL low-， medium- and high-concentration groups （1， 2.5， 5 μmol/L）， and CSL high-concentration+Zn27 [focal adhesion kinase （FAK） inhibitor] group （5 μmol/L CSL+2 nmol/L Zn27）， with 6 holes in each group. The proliferation （by absorbance） and cloning ability， apoptotic rate， the number of invasion cells and migration cells， the level of reactive oxygen species（ROS） as well as the protein expressions of phosphorylated FAK （p-FAK）， phosphorylated mitogen-activated protein kinase kinase （p-MEK）， phosphorylated extracellular signal-regulated kinase （p-ERK）， B-cell lymphoma-2 （Bcl-2）， Bcl-2 associated X protein （Bax） and caspase-3 were detected. RESULTS Compared with control group， cell absorbance， clone count， invasion count and migration count ， and the protein expressions of p-FAK， p-MEK， p-ERK and Bcl-2 were significantly reduced in the CSL low- ， medium- ， high- concentration groups； the apoptosis rate， ROS level， and protein expressions of Bax and caspase-3 were significantly increased， in a concentration-dependent manner （P＜0.05）. Compared with CSL high-concentration group， the changes of above indexes were all reversed significantly in CSL high-concentration+Zn27 group （P＜0.05）. CONCLUSIONS CSL can enhance oxidative stress， promote cell apoptosis， inhibit malignant progression and chemotherapy resistance of liver cancer cells， and its mechanism may be related to the inhibition of the FAK/MEK/ERK signaling pathway.

BACKGROUND: Bladder cancer, characterized by a high potential of tumor recurrence, has high lifelong monitoring and treatment costs. To date, tumor cells with intrinsic softness have been identified to function as cancer stem cells in several cancer types. Nonetheless, the existence of soft tumor cells in bladder tumors remains elusive. Thus, our study aimed to develop a micro-barrier microfluidic chip to efficiently isolate deformable tumor cells from distinct types of bladder cancer cells. METHODS: The stiffness of bladder cancer cells was determined by atomic force microscopy (AFM). The modified microfluidic chip was utilized to separate soft cells, and the 3D Matrigel culture system was to maintain the softness of tumor cells. Expression patterns of integrin β8 (ITGB8), protein kinase B (AKT), and mammalian target of rapamycin (mTOR) were determined by Western blotting. Double immunostaining was conducted to examine the interaction between F-actin and tripartite motif containing 59 (TRIM59). The stem-cell-like characteristics of soft cells were explored by colony formation assay and in vivo studies upon xenografted tumor models. RESULTS: Using our newly designed microfluidic approach, we identified a small fraction of soft tumor cells in bladder cancer cells. More importantly, the existence of soft tumor cells was confirmed in clinical human bladder cancer specimens, in which the number of soft tumor cells was associated with tumor relapse. Furthermore, we demonstrated that the biomechanical stimuli arising from 3D Matrigel activated the F-actin/ITGB8/TRIM59/AKT/mTOR/glycolysis pathways to enhance the softness and tumorigenic capacity of tumor cells. Simultaneously, we detected a remarkable up-regulation in ITGB8, TRIM59, and phospho-AKT in clinical bladder recurrent tumors compared with their non-recurrent counterparts. CONCLUSIONS The ITGB8/TRIM59/AKT/mTOR/glycolysis axis plays a crucial role in modulating tumor softness and stemness. Meanwhile, the soft tumor cells become more sensitive to chemotherapy after stiffening, that offers new insights for hampering tumor progression and recurrence.
Animals ; Mice ; Humans ; Proto-Oncogene Proteins c-akt/metabolism* ; Actins/metabolism* ; Neoplasm Recurrence, Local ; TOR Serine-Threonine Kinases/metabolism* ; Urinary Bladder Neoplasms ; Glycolysis ; Cell Line, Tumor ; Cell Proliferation ; Mammals/metabolism* ; Tripartite Motif Proteins/metabolism* ; Intracellular Signaling Peptides and Proteins/metabolism* ; Integrin beta Chains