Butyrate-mediated Resistance to Trichostatin A Accompanied by Elevated Expression of Glucose Transporter 3 (GLUT3) in Human Colorectal Carcinoma HCT116 Cells.

OBJECTIVE: The aim of study was to investigate the correlation of GLUT3 upregulation and butyrate-mediated acquired chemoresistance. METHOD: A butyrate-resistant CRC cell model was established from parental (PT) HCT116 cells by gradually increasing the concentration of sodium butyrate (NaBu), followed by evaluation of resistance to butyrate and trichostatin A (TSA) by the MTT method. The expression of SLC2A3 gene and GLUT3 protein were assessed by semi-quantitative RT-PCR and western blotting, respectively. The correlation of GLUT3 and butyrate-induced acquired chemoresistance was investigated using SLC2A3 silencing. RESULTS: Butyrate-resistant (BR) HCT116 cells were more tolerant to butyrate-induced cell death and also resist to 750 and 1000 nM TSA when compared with HCT116-PT cells (p <0.05). Long-term exposure to butyrate revealed that upregulation of the SLC2A3 gene was significantly increased by more than 20 fold (p < 0.01), and that of GLUT3 was elevated by approximately 2 fold (p < 0.05) in HCT116-BR cells. Silencing of the SLC2A3 gene increased the sensitivity of HCT116-BR cells to the effects of TSA. CONCLUSION: Upregulation of GLUT3 is associated with resistance to butyrate and TSA. GLUT3 is a molecular target for the detection of chemoresistant CRC cells and thus a potential target for diagnostic strategies.

Epigallocatechin gallate triggers apoptosis by suppressing de novo lipogenesis in colorectal carcinoma cells.

The de novo lipogenesis (DNL) pathway has been identified as a regulator of cancer progression and aggressiveness. Downregulation of key lipogenesis enzymes has been shown to activate apoptosis in cancerous cells. Epigallocatechin gallate (EGCG) inhibits cancer cell proliferation without causing cytotoxicity in healthy cells. The present study aimed to investigate the effects of EGCG on the promotion of apoptosis associated with the DNL pathway inhibition in cancer cells, both in vitro and in vivo. We observed that two colorectal cancer cell lines (HCT116 and HT-29) had a higher cytotoxic response to EGCG treatment than hepatocellular carcinoma cells, including HepG2 and HuH-7. EGCG treatment decreased cell viability and increased mitochondrial damage-triggered apoptosis in both HCT116 and HT-29 cancer cells. Additionally, we treated mice transplanted with HCT116 cells with 30 or 50 mg·kg-1 EGCG for 7 days to evaluate the apoptotic effects of EGCG treatment in a xenograft mouse model of cancer. We observed a decrease in intracellular fatty acid levels, which suggested that EGCG-induced apoptosis was associated with a decrease in fatty acid levels in cancer. Suppression of ATP synthesis by EGCG indicated that cell death induction in cancer cells could be mediated by shared components of the DNL and energy metabolism pathways. In addition, EGCG-induced apoptosis suppressed the expression of the phosphorylation protein kinase B and extracellular signal-regulated kinase 1/2 signaling proteins in tumors from xenografted mice. Cytotoxic effects in unaffected organs and tissues of the mouse xenograft model were absent upon EGCG treatment.

Calotropis gigantea stem bark extract induced apoptosis related to ROS and ATP production in colon cancer cells.

Conventional chemotherapeutic agents for colorectal cancer (CRC) cause systemic side effects and eventually become less efficacious owing to the development of drug resistance in cancer cells. Therefore, new therapeutic regimens have focused on the use of natural products. The anticancer activity of several parts of Calotropis gigantea has been reported; however, the effects of its stem bark extract on inhibition of cancer cell proliferation have not yet been examined. In this study, the anticancer activity of C. gigantea stem bark extract, both alone and in combination with 5-fluorouracil (5-FU), was evaluated. A crude ethanolic extract was prepared from dry, powdered C. gigantea barks using 95% ethanol. This was then partitioned to obtain dichloromethane (CGDCM), ethyl acetate, and water fractions. Quantitative analysis of the constituent secondary metabolites and calotropin was performed. These fractions exhibited cytotoxicity in HCT116 and HT-29 cells, with CGDCM showing the highest potency in both the cell lines. A combination of CGDCM and 5-FU significantly enhanced the cytotoxic effect. Moreover, the resistance of normal fibroblast, HFF-1, cells to this combination demonstrated its safety in normal cells. The combination significantly enhanced apoptosis through the mitochondria-dependent pathway. Additionally, the combination reduced adenosine triphosphate production and increased the production of reactive oxygen species, demonstrating the mechanisms involved in the induction of apoptosis. Our results suggest that CGDCM is a promising anti-cancer agent and may enhance apoptosis induction by 5-FU in the treatment of CRC, while minimizing toxicity toward healthy cells.

Microarray Analysis of Gene Expression Involved in Butyrate-Resistant Colorectal Carcinoma HCT116 Cells.

BACKGROUND: Resistance to chemotherapeutic agents is usually found in cancer stem cells (CSCs) and cancer stem-like cells that are often regarded as the target for cancer monitoring. However, the different patterns of their transcriptomic profiling is still unclear. OBJECTIVE: This study aims to illustrate the transcriptomic profile of CSCs and butyrate-resistant colorectal carcinoma cells (BR-CRCs), by comparing them with parental colorectal cancer (CRC) cells in order to identify distinguishing transcription patterns of the CSCs and BR-CRCs. METHODS: Parental CRC cells HCT116 (HCT116-PT) were cultured and induced to establish the butyrate resistant cell model (HCT116-BR). Commercial enriching of the HCT116-CSCs were grown in a tumorsphere suspension culture, which was followed firstly by the assessment of butyrate tolerance using MTT and PrestoBlue. Then their gene expression profiling was analyzed by microarray. RESULTS: The results showed that both butyrate-resistant HCT116 cells (HCT116-BR) and HCT116-CSCs were more tolerant a butyrate effects than HCT116-PT cells. Differentially expressed gene profiles exhibited that IFI27, FOXQ1, PRF1, and SLC2A3 genes were increasingly expressed in CSCs, and were dramatically overexpressed in HCT116-BR cells when compared with HCT116-PT cells. Moreover, PKIB and LOC399959 were downregulated both in HCT116-CSCs and HCT116-BR cells. CONCLUSION: Our findings shed light on the transcriptomic profiles of chemoresistant CRC cells. This data should be useful for further study to provide guidelines for clinical prognosis to determine the guidelines for CRC treatment, especially in patients with chemoresistance and designing novel anti-neoplastic agents.

Changes in the NS1 gene of avian influenza viruses isolated in Thailand affect expression of type I interferon in primary chicken embryonic fibroblast cells.

The non-structural protein 1 (NS1) of avian influenza virus was defined as one of the virulent factors. To understand the effect of NS1 protein of influenza virus H5N1 isolated in Thailand on type I (α/ß) interferon (IFN) synthesis, five reverse genetic viruses were constructed and used as models. The viruses were generated using NS genomic segment from A/Peurto Rico/8/1934 (H1N1) and four avian influenza viruses isolated from the first outbreak in Thailand. All the viruses have the rest of the genome from A/Peurto Rico/8/1934 (H1N1). The constructed viruses were named (1) NS1 PR8/34, (2) NS1 wild type, (3) NS1 L15FD53G, (4) NS1 N171I and (5) NS1 E71K, respectively. The type I (α/ß) IFN gene expression in control and infected primary chicken embryonic fibroblast cells were analyzed by quantitative polymerase chain reaction. The results show that the inhibition of IFN-ß gene expression by NS1 wild type infected cells is stronger than NS1 N171I, NS1 E71K, NS1 PR8/34 and NS1 L15FD53G, respectively. The data suggest that the difference of amino acid sequence of NS1 protein contributes to the IFN-ß antagonist. In contrast, the difference of the NS1 protein does not influence in the IFN-α antagonistic activity.