Human Liver L02 Cells Research Articles

Effect of functionalized multi-walled carbon nanotubes on L02 cells

To investigate the cytotoxic effect of multi-walled carbon nanotubes (MWCNTs) on human liver L02 cells and its relevant mechanism. MWCNTs, carboxyl modification MWCNTs (MWCNTs-COOH), and hydroxyl modification MWCNTs (MWCNTs-OH) were characterized by transmission electron microscopy, scanning electron microscopy, and X-ray photoelectron spectroscopy. The carbon nanotubes at concentrations of 12.5, 25, 50, 100, and 200 μg/ml were incubated with human liver L02 cells for 24, 48 and 72 hours, respectively. The cell viability was evaluated by water soluble tetrazolium salts assay and the intercellular reactive oxygen species induced by the carbon nanotubes were detected by 2', 7'-dichlorodihydrofluorescein diacetate method. Transmission electron microscope showed that the average outside diameters (10 to 20 nm) and the average length (10 to 30 μm) of the three MWCNTs were similar. Scanning electron microscope indicated that the three MWCNTs had a similar surface topography. X-ray photoelectron spectroscopy demonstrated that the MWCNTs-COOH and MWCNTs-OH had relatively high peak areas at 289 and 286ev, respectively,indicating that they have been modified by carboxyl and hydroxyl groups,respectively. Water soluble tetrazolium salts assay showed that the MWCNTs-COOH was less cytotoxic when compared to MWCNTs which demonstrated to be slightly more cytotoxic than MWCNTs-OH. The capability to induce increase in intracellular reactive oxygen species was in the following order: MWCNTs > MWCNTs-COOH > MWCNTs-OH. Modification of MWCNTs with carboxyl group and hydroxyl group improves the biocompatibility of MWCNTs to some extents. MWCNTs-COOH has better compatibility than MWCNTs at the low concentration,and MWCNTs-OH showed better compatibility than MWCNTs after 48 hours. Different mechanisms may be involved in the interaction between cells and the MWCNTs with different chemical surfaces.

Effects of extract of Polygonum multiflorum on cell cycle arrest and apoptosis of human liver cell line L02

To analyze the chemical constituents of Polygonum multiflorum extract which may cause human liver cell damage and to explore the mechanism. Raw and processed Polygonum multiflorum were extracted by 70% ethanol, then raw and processed Polygonum multiflorum water-eluted material (RW and PW), 50% ethanol-eluted material (R50 and P50) and 95% ethanol-eluted material (R95 and P95) were obtained by absorbing through AB-8 macroporous resin, followed by water, 50% ethanol and 95% ethanol elution in order. The water extracts of raw and processed Polygonum multiflorum (RWE or PWE) were obtained by boiling them in water as usual. Normal human liver L02 cells were treated by different concentrations of eluted Polygonum multiflorum materials for different time, and the cell growth inhibition of each group was determined by methylthiazolyldiphenyl-tetrazolium bromide method. The chemical constituents which had a significant cytotoxicity to L02 cells were analyzed by high-performance liquid chromatography (HPLC). Morphological changes of L02 cells were observed by Giemsa staining and cell cycle distribution was observed by flow cytometry. It was found that 95% ethanol-eluted extracts of raw and processed Polygonum multiflorum showed significant growth inhibition on normal human liver L02 cells, while the other components showed no significant inhibition on cell growth. HPLC analysis showed that the main component in 95% ethanol-eluted extract of raw and processed Polygonum multiflorum was emodin at content of (18.53+/-2.96)% and (10.28+/-1.34)% respectively. Cell cycle analysis showed that 95% ethanol-eluted material of Polygonum multiflorum and emodin had a similar significant effect of S phase arrest and all could induce L02 cell apoptosis. The main part of Polygonum multiflorum causing liver cell damage is the 95% ethanol-eluted extract, and emodin is one of the important chemical constituents leading to liver cell damage.

DNA-dependent protein kinase catalytic subunit modulates the stability of c-Myc oncoprotein

BackgroundC-Myc is a short-lived oncoprotein that is destroyed by ubiquitin-mediated proteolysis. Dysregulated accumulation of c-Myc commonly occurs in human cancers. Some of those cases with the dysregulated c-Myc protein accumulation are attributed to gene amplification or increased mRNA expression. However, the abnormal accumulation of c-Myc protein is also a common finding in human cancers with normal copy number and transcription level of c-Myc gene. It seems that the mechanistic dysregulation in the control of c-Myc protein stabilization is another important hallmark associated with c-Myc accumulation in cancer cells. Here we report a novel mechanistic pathway through which DNA-dependent protein kinase catalytic subunit (DNA-PKcs) modulates the stability of c-Myc protein.ResultsFirstly, siRNA-mediated silencing of DNA-PKcs strikingly downregulated c-Myc protein levels in HeLa and HepG2 cells, and simultaneously decreased cell proliferation. The c-Myc protein level in DNA-PKcs deficient human glioma M059J cells was also found much lower than that in DNA-PKcs efficient M059K cells. ATM deficiency does not affect c-Myc expression level. Silencing of DNA-PKcs in HeLa cells resulted in a decreased stability of c-Myc protein, which was associated the increasing of c-Myc phosphorylation on Thr58/Ser62 and ubiquitination level. Phosphorylation of Akt on Ser473, a substrate of DNA-PKcs was found decreased in DNA-PKcs deficient cells. As the consequence, the phosphorylation of GSK3 β on Ser9, a negatively regulated target of Akt, was also decreased, and which led to activation of GSK 3β and in turn phosphorylation of c-Myc on Thr58. Moreover, inhibition of GSK3 activity by LiCl or specific siRNA molecules rescued the downregulation of c-Myc mediated by silencing DNA-PKcs. Consistent with this depressed DNA-PKcs cell model, overexpressing DNA-PKcs in normal human liver L02 cells, by sub-chronically exposing to very low dose of carcinogen 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), increased c-Myc protein level, the phosphorylation of Akt and GSK3 β, as well as cell proliferation. siRNA-mediated silencing of DNA-PKcs in this cell model reversed above alterations to the original levels of L02 cells.ConclusionA suitable DNA-PKcs level in cells is necessary for maintaining genomic stability, while abnormal overexpression of DNA-PKcs may contribute to cell proliferation and even oncogenic transformation by stabilizing the c-Myc oncoprotein via at least the Akt/GSK3 pathway. Our results suggest DNA-PKcs a novel biological role beyond its DNA repair function.

Study on the structure–activity relationships of phoxalone and induction of apoptosis in H446 tumor cells

A novel macrolide with dioxa and double ring pentadecanone, named as phoxalone, isolated from the culture broth of a myxobacterium Sorangium cellulosum WXNXJ-C, was well investigated. The chemical structure of Phoxalone was elucidated by spectroscopic analysis including UV, IR, 1H NMR, MALDI-TOF-MS, and LC/MS spectra. The results of cytotoxic bioactivities on human non-small lung cancer H446, in vitro, showed that not only did phoxalone express higher antitumor bioactivity than many antitumor drugs, but it also displayed less cytotoxicity to normal human liver L02 cell lines (IC50, 286 μg mL−1). Further cytotoxic bioactivity study indicated that phoxalone could induce H446 cell line apoptosis in vitro. More investigation results of flow-cytometric analysis suggested that phoxalone arrested the mitosis of H446 cell line at G2/M phase. Hence, phoxalone and its derivatives or analogs would reveal huge research value and fascinating foreground.

Phoxalone, a novel macrolide from Sorangium cellulosum: structure identification and its anti-tumor bioactivity in vitro

A novel macrolide, isolated from the myxobacteria Sorangium cellulosum WXNXJ-C, was identified as 1,7,12,13-tetrahydroxy-14-methoxy-8,10-dimethyl-6-phenyl-5,15-dioxa-bicyclo[9.3.1]pentadecan-4-one, "Phoxalone". It had minimum IC(50) values of 0.24, 6.9, 10.3, 0.98 and 4 microg/ml, respectively, against tumour cell lines: B16, Bel7402, H446, MCF-7, and SGC7901. In addition, it had less cytotoxicity to normal human liver L02 cell lines (286 microg/ml, 24 h). A cytotoxic bioactivity study on H446 cell line in vitro suggested that Phoxalone arrested the mitosis in the G2/M phase.