Alantolactone induces apoptosis in THP-1 cells through STAT3, survivin inhibition, and intrinsic apoptosis pathway.

Cancer is the second foremost cause of mortality in the world, and THP-1 cells play an important role in cancer progression. Alantolactone (ALT), a sesquiterpene lactone compound derived from Inula helenium, has a number of biological activities including antibacterial, antifungal, and anticancer. The current study was conducted to investigate the effects of ALT on THP-1 cells and its underlying molecular mechanisms. THP-1 cells were cultured and treated with ALT (20, 40 µM) for 12 hr, and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, cell morphology, live/dead, and apoptosis assays were performed. The gene expressions at the protein level were checked through Western blot. Results show that ALT decreased cell viability and increased cell death and apoptosis. We found that ALT inhibited STAT3 and survivin expression. Furthermore, ALT induced mitochondrial-dependent apoptosis through a decrease in B-cell lymphoma-2 (Bcl-2) and Bcl-xL and increase in Bax expression, resulting in the release of cytochrome c (Cyt-c) from mitochondria. Cyt-c release from mitochondria further increased cleaved (cl) caspase-3 and cl-PARP expression and led the cells to apoptosis. Therefore, ALT might be a good therapy for the progression due to THP-1 cells.

Improving pathological early diagnosis and differential biomarker value for hepatocellular carcinoma via RNAscope technology.

BACKGROUND: The diagnostic and prognostic values of glypican3 (GPC3) and glutamine synthetase (GS) proteins in hepatocellular carcinoma (HCC) have been reported, but their specificity and sensitivity remain low. Here, we applied RNAscope to improve HCC early pathological and differential diagnosis by estimating GPC3 and GS mRNAs. METHODS: We performed RNAscope and immunohistochemistry (IHC) to detect GPC3 and GS biomarkers on the tissue sections of 194 cases, including high- and low-grade liver dysplastic nodules; highly, moderately, and poorly differentiated HCCs; intrahepatic cholangiocarcinomas (ICCs); metastatic HCC; and carcinomas from other organs. RESULTS: The results showed that all the cases that were negative for GPC3 by RNAscope were also negative for this protein by IHC. The use of RNAscope assay improved the GPC3 and GS specificity and sensitivity by 20-30%. Hence, HCC shows early recognition and upgrades the metastatic HCC differentiation by 23% compared with IHC (p = 0.0001, 0.0064). Meanwhile, all liver cirrhosis, cholangiocytes and non-HCC samples were negative for GPC3 and GS except lymphocytes in lymphomas, and 2 (8.3%) out of the 24 ICC samples but not in the cancer cells. CONCLUSION: RNAscope for GPC3 and GS panel was highly specific and sensitive for the pathological identification of dysplastic nodules, early stages of HCCs, and would differentiate them from HCCs and metastatic tumors compared with IHC.

Ezrin as a possible diagnostic and/or prognostic biomarker in mice lymphatic metastatic hepatocellular carcinoma in vivo.

Hepatocellular carcinoma (HCC) ranks in the top of cancers leading to death. Early diagnosis is the big challenge in the case of HCC. Our in vitro study showed that Ezrin expression in lymphatic metastasis hepatocellular carcinoma (LNM-HCC) was associated with the metastatic rate. Here we aim to evaluate Ezrin expression as diagnostic and/or prognostic biomarker of LNM-HCC in mice. Chinese inbred 615 mice, Hca-F and Hca-P cell lines were used in the study. Histological changes were determined by Hematoxylin and Eosin, while Ezrin expression was assessed by qRT-PCR, western blot, immunohistochemistry, and enzyme-linked immunosorbent assay. Ezrin expression in this study gives credit to our in vitro study which Ezrin expression was positively correlated with LNM-HCC and negatively with Annexin7 (A7) expression. The highest histological changes were observed in high metastatic primary/secondary tumors combined with high Ezrin expression. Ezrin and A7 are higher in total primary tumors than in total secondary tumors (P = 0.0001, P = 0.021), respectively. Ezrin expression was enhanced in Hca-P A7 down-regulated primary/secondary tumors (P = 0.004), whereas, Ezrin expression was suppressed in Hca-F A7 upregulated primary/secondary tumors. Serum ELISA indicated differential expression of Ezrin among the study groups (P ≤ 0.0001). Ezrin expression was higher in NC-Hca-F than NC-Hca-P (P ≤ 0.0001), suppressed in Hca-F A7 upregulation (P ≤ 0.0001) and in enhanced in Hca-P A7 down-regulation (P = 0.0001). In conclusion, Ezrin level may serve as a differential diagnostic and/or prognostic biomarker for high and low LNM-HCC and may be beneficial in the diagnosis of HCC disease. © 2017 BioFactors, 43(5):662-672, 2017.

Pathway-related molecules of VEGFC/D-VEGFR3/NRP2 axis in tumor lymphangiogenesis and lymphatic metastasis.

Precondition for tumor lymphatic metastasis is that tumor cells induce formation of original and newborn lymphatic vessels and invade surrounding lymphatic vessels in tumor stroma, while some pathway-related molecules play an important role in mechanisms associated with proliferation and migration of lymphatic endothelial cells (LECs) and tumor cells. In lymphangiogenesis and lymphatic metastasis, the pathway-related molecules of VEGFC/D-VEGFR3/NRP2 axis, such as Furin-like enzyme, CNTN1, Prox1, LYVE-1, Podoplanin, SOX18, SDF1 and CXCR4, are direct constitutors as a portion of VEGFC/D-VEGFR3/NRP2 axis, and their biological activities rely on this ligand-receptor system. These axis-related signal molecules could gradually produce waterfall-like cascading effects, mediate differentiation and maturation of LECs, remodel original and neonatal lymphatic vessels, as well as ultimately promote tumor cell chemotaxis, migration, invasion and metastasis to lymphoid tracts. This review summarizes the structure and function features of pathway-related molecules of VEGFC/D-VEGFR3/NRP2 axis, the expression changes of these molecules in different anatomic organs or histopathologic types or development stages of various tumors, the characteristics of transduction, implementation, integration of signal networks, the interactive effects on biological behaviors between tumor cells and lymphatic endothelial cells, and their molecular mechanisms and significances in tumor lymphangiogenesis and lymphatic metastasis.