Human apoptosis antibody array-membranes studying the apoptotic effect of marine bacterial exopolysaccharides in HepG2 cells.

BACKGROUND: Hepatocellular carcinoma (HCC) is considered as the third leading cause of cancer-related deaths, in spite of great advances in its treatment. The carbohydrate polymers, exopolysaccharides (EPSs), showed anticancer activity in diverse cancers. OBJECTIVE: The purpose of this study is to investigate a panel of 43 apoptotic proteins to assess the possible apoptotic induction effect of bacterial EPSs showing promising cytotoxic effects in HepG2 cells in our previous study, in an attempt to introduce exopolysaccharides as new source for cancer treatment. MATERIALS AND METHODS: Apoptosis-related proteins panel were examined through the analysis of Human Apoptosis Antibody Array-Membrane (43 targets). RESULTS: EPS-6 induces apoptosis through upregulation of different pro-apoptotic proteins as cytochrome C (9.52 fold) and tumor necrosis factor-related apoptosis-inducing ligand receptor (TRAIL-R1) (153.49 fold). EPS-RS induces apoptosis through up regulation of second mitochondria-derived activator of caspases (SMAC) (15.75 fold) and the six insulin-like growth factors binding proteins (IGFBP-1 through - 6) (76.81 fold, 7.68 fold, 55.15 fold, 4.9 × 107 fold, 29.69 fold, and 28.92 fold), respectively. CONCLUSION: Our results suggested that EPS-6 and EPS-RS could be considered as promising agents in hepatocellular carcinoma treatment.

Newly isolated marine bacterial exopolysaccharides enhance antitumor activity in HepG2 cells via affecting key apoptotic factors and activating toll like receptors.

Hepatocellular carcinoma (HCC) was accompanied by high incidence of morbidity and mortality worldwide. Apoptosis is a vital biological process playing a critical role in cancer. Besides, toll like receptors were reported to regulator the innate immune response against cancer development. Exopolysaccharides (EPSs) derived from marine bacteria were reported to have a potential biological importance. This work aimed to elucidate the antitumor effects of newly isolated EPSs against HepG2 cells. Moreover, their effects on some apoptotic markers and TLRs were followed. Isolated EPSs were tested for their cytotoxic effects in a previous study and the most promising; MSA1, E4, MGA2, SGA3, and NRC7 EPSs were subjected to molecular analysis to investigate their pro-apoptotic effects, in addition to their effects on TLR2 and TLR-9 using quantitative real time RT-PCR. And the most cytotoxic and pro-apoptotic EPS; MSA1 were subjected to antibody array analysis to investigate a panel of 43 apoptotic proteins. All isolated EPSs produced a positive role in regulating the apoptotic gene and increasing the TLRs expression in different manners. However, the most promising EPS was MSA1. It showed pro-apoptotic effects on gene and protein levels, besides its up-regulation of TLRs.

Antitumor Exopolysaccharides Derived from Novel Marine Bacillus: Isolation, Characterization Aspect and Biological Activity

Objective: Exopolysaccharides gained attention as new source for cancer treatment as recent treatments cause side effects and multidrug resistance. Polysaccharides containing sulfur and uronic acids exhibited antioxidant activity, by restoring cell redox regulation, thus inhibiting cell proliferation and cancer formation. Following this context, our study was performed to assess the cytotoxic activity of exopolysaccharides produced by novel Egyptian marine bacterial strains on HepG2 cells. Methods: Bacteria were isolated, purified and cultured through routine microbiological techniques. 16S rRNA gene amplification and sequence analyses, Fourier Transform Infra-red (FTIR), Identification of monosugars by HPLC molecular weight estimation, sulfur content determination and neutral red uptake assay were utilized. Results: BLAST showed that the isolates were related to the Bacillus sp. FTIR analysis indicated that the four EPSs under study contained sulfur as substituent functional group but with different percentage in each EPS. The highest sulfur percentage (46%) appeared in the EPS-6 that was produced by Bacillus flexus isolated from the Mediterranean Sea. HPLC showed that EPSs contained uronic acids which appeared as glucuronic and galacturonic acid in the low molecular weight EPS-6 (4.296×104 g mol-1). Arabinose appeared besides the glucuronic and galacturonic acid residues. EPS-6 showed the highest cytotoxicity, IC50 (218 µg ml-1) which could be correlated to the presence of sulfure and uronic acids in its structure. Conclusion: The novel Firmicutes from the Egyptian saline habitat produced EPSs of cytotoxic activity on hepatocellular carcinoma.

Differential effects of c-myc and ABCB1 silencing on reversing drug resistance in HepG2/Dox cells.

Multidrug resistance (MDR) in various kinds of cancers represents a true obstacle which hinders the successes of most of current available chemotherapies. ATP-binding cassette (ABC) trasporter proteins have been shown to contribute to the majority of MDR in various types of malignancies. c-myc has recently been reported to participate, at least partly, in MDR to some types of cancers. This study aimed to test whether c-myc could play a role, solely or with coordination with other ABCs, in the resistance of HepG2 cells to doxorubicin (Dox). MDR has been induced in wild-type HepG2 and has been verified both on gene and protein levels. Various assays including efflux assays as well as siRNA targeting ABCB1 and c-myc have been employed to explore the role of both candidate molecules in MDR in HepG2. Results obtained, with regard to ABCB1 silencing on HepG2/Dox cells, have shown that ABCB1-deficient cells exhibited a significant reduction in ABCC1 expression as compared to ABCB1-sufficient cells. However, these cells did not show a significant reduction in other tested ABCs (ABCC5 and ABCC10) while c-myc silencing had no significant effect on any of the studied ABCs. Moreover, silencing of ABCB1 on HepG2 significantly increased fluorescent calcein retention in HepG2 cells as compared to the control cells while downregulation of c-myc did not have any effect on fluorescent calcein retention. Altogether, this work clearly demonstrates that c-myc has no role in MDR of HepG2 to Dox which has been shown to be ABCB1-mediated in a mechanism which might involve ABCC1.