• Biomolecules & Therapeutics
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• v.15 no.3
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• pp.150-155
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• 2007

In the present study, we have tested the effect of dioleoyl phosphatidic acid (PA) on intracellular $Ca_{2+}$ concentration ($[Ca^{2+}]_{i}$) in two human colon cancer cell lines (HCT116 and HT29). PA and lysophosphatidic acid (LPA), a bioactive lysolipid, increased $[Ca^{2+}]_{i}$ in both HCT116 and HT29 cell lines. Increases of $[Ca^{2+}]_{i}$ by PA and LPA were more robust in HCT116 cells than in HT29 cells. A specific inhibitor of phospholipase C (U73122), however, was not inhibitory to the cell responses. Pertussis toxin, a specific inhibitor of $G_{i/o}$ type G proteins, however, had an inhibitory effect on the responses except for an LPA-induced one in HT29 cells. Ruthenium red, an inhibitor of the ryanodine receptor, was not inhibitory on the responses, however, 2-APB, a specific inhibitor of inositol 1,4,5-trisphosphate receptor, completely inhibited both lipid-induced $Ca^{2+}$ increases in both cell types. Furthermore, by using Ki16425 and VPC32183, two structurally dissimilar specific antagonists for $LPA_{1}/LPA_{3}$ receptors, an involvement of endogenous LPA receptors in the $Ca^{2+}$ responses was observed. Ki16425 completely inhibited the responses but the susceptibility to VPC32183 was different to PA and LPA in the two cell types. Expression levels of five LPA receptors in the HCT116 and HT29 cells were also assessed. Our data support the notion that PA could increase $[Ca^{2+}]_{i}$ in human colon cancer cells, probably via endogenous LPA receptors, G proteins and $IP_{3}$ receptors, thereby suggesting a role of PA as an intercellular lipid mediator.

• Korean Journal of Food Science and Technology
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• v.40 no.2
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• pp.207-214
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• 2008

In Asia Saussurea lappa (SL) has been used as a traditional herbal medicine to treat abdominal pain and tenesmus. Recently, in vitro cell culture studies have shown that SL has anti-ulcer, anti-inflammatory, and anti-tumor properties. To explore its potential chemopreventive and chemotherapeutic effects in colon cancer, we examined whether the hexane extract of SL (HESL) could inhibit the growth of HT-29 human colon cancer cells, and investigated the mechanisms for this effect. The cells were cultured with various concentrations (0-5 ${\mu}g/mL$) of HESL. The results indicated that HESL markedly decreased the numbers of viable HT-29 cells; whereas at the concentration of 5 ${\mu}g/mL$, HESL slightly decreased the viable cell numbers of CCD 1108Sk human skin normal fibroblasts at 72 hr. HESL substantially increased the numbers of cells in the sub G1 phase, and dose-dependently increased apoptotic cell numbers. Western blot analysis of the total cell lysates revealed that HESL increased Bax protein levels, but did not affect Bcl-2 levels. HESL induced the cleavage of poly (ADP-ribose) polymerase and caspases 8, 9, 7, and 3. This study demonstrated that HESL inhibits cell growth and induces apoptosis in HT-29 cells, which may be mediated by its ability to increase Bax levels and activate the caspase pathway. These findings may lead to the development of new therapeutic strategies for colon cancer treatment.

• 한국약용작물학회:학술대회논문집
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• 2011.04a
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• pp.316-317
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• 2011

• Food Science and Biotechnology
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• v.17 no.6
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• pp.1265-1271
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• 2008

Adlay bran is a waste product previously thought to have no commercial value, Its methanolic extract was fractionated using n-hexane (ABM-Hex), ethyl acetate (ABM-EtOAc), 1-butanol (ABM-BuOH), and water (ABM-$H_2O$). The ABM-EtOAc fraction exhibited a strongest inhibition against growth of human lung cancer cell A549 and human colorectal carcinoma cells HT-29 and COLO 205. Inhibition of cell cycle progression at $G_0/G_1$ transition, increase of cells at the sub-$G_1$ phase, and DNA ladders were observed in cells treated with ABM-EtOAc. The ABM-BuOH fraction showed the strongest inhibition of proinflammatory cytokines tumor necrosis factor (TNF)-$\alpha$ and interlukin (IL)-$1{\beta}$ in stimulated RAW 264.7 macrophages. Further, ABM-EtOAc and ABM-BuOH inhibited cyclooxygenase (COX)-2 expression in A549 and HT-29 carcinoma cells, while COX-l expression was not affected. These results reveal that both ABM-EtOAc and ABM-BuOH may aid the prevention of cancers and the applications in cancer chemotherapy.

• Radiation Oncology Journal
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• v.20 no.4
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• pp.367-374
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• 2002

Purpose : To investigate the growth inhibitory effects, and the underlying mechanism of human colon cancer cell (HT-29) death, induced by a new synthetic bile acid derivative (HS-1200). Materials and Methods : Human colon cancer cells (HT-29), in exponential growth phase, were treated with various concentrations of a new synthetic bile acid derivative (HS-1200). The growth inhibitory effects on HT-29 cells were examined using a frypan blue exclusion assay. The extent of apoptosis was determined using agarose gel electrophoresis, TUNEL assays and Hoechst staining. The apoptotic cell death was also confirmed by Western blotting of PARP, caspase-3 and DNA fragmentation factor (DFF) analysis. To investigate the involvement of mitochondria, we employed immunofluorescent staining of cytochrome c and mitochondrial membrane potential analyses. Results : The dose required for the half maximal inhibition $(IC_{50})$ of the HT-29 cell growth was $100\~150\;{\mu}M$ of HS-1200. Several changes, associated with the apoptosis of the HT-29 cells, were reveal by the agarose gel eletrophoresis, TUNEL assays and Hoechst staining, following their treatment with $100\;{\mu}M$ of HS-1200. HS-1200 treatment also induced caspase-3, PARP and DFF degradations, and the western blotting showed the processed caspase-3 p20, PARP p85 and DFF p30 and p11 cleaved products. Mitochondrial events were also demonstrated. The cytochrome c staining indicated that cytochrome c had been released from the mitochondria in the HS-1200 treated cells. The mitochondrial membrane potential $(\Delta\Psi_m)$ was also prominently decreased in the HS-1200 treated cells. Conclusion : These findings suggest that the HS-1200 - induced apoptosis of human colon cancer cells (HT-29) is mediated via caspase and mitochondrial pathways.

• Korean Journal of Microbiology
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• v.51 no.3
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• pp.241-248
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• 2015

Although studies on probiotics have been performed mostly with viable microbes, the beneficial functions of dead or heat-killed form of probiotic strains have also been examined. In this study, live and heat-killed forms of Lactobacillus acidophilus CBT LA1 were investigated in vitro and in vivo to evaluate the properties necessary for gut barrier protection. Cell surface hydrophobicity (CSH), autoaggregation (AA), cell adhesion, and lipopolysaccharide (LPS)-binding properties were evaluated. In addition, the suppressive effect on LPS-induced interleukin (IL)-8 expression was investigated in HT-29 cells. To identify optimal conditions for CBT LA1 to adhere to HT-29 cells, CBT LA1 cells were heat-treated at 80, 85, 90, 95, 100, or $121^{\circ}C$ for 10 min; cells treated at $80^{\circ}C$ for 10 min showed the highest adhesion. Heat-killed bacteria at $80^{\circ}C$ showed higher levels of LPS-binding, CSH, AA, adhesion to HT-29, and suppression of IL-8 expression than did live CBT LA1. In vivo imaging was performed to evaluate the ability of live or heat-killed CBT LA1 to remove LPS from the intestine in a rat model of infection. At 16 h after infection, fluorescence from FITC-conjugated LPS had mostly disappeared from the intestine of the rats administered with live or heat-killed CBT LA1; the effect was greater with heat-killed CBT LA1 at $80^{\circ}C$. These results suggest that heat-killed CBT LA1 as well as its live form can be applied as a pharmabiotic for protection of the gut barrier.

• The Korean Journal of Physiology and Pharmacology
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• v.15 no.6
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• pp.423-429
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• 2011

B13 analogues are being considered as therapeutic agents for cancer cells, since B13 is a ceramide analogue and inhibits ceramidase to promote apoptosis in cancer cells. B13 sulfonamides are assumed to have biological activity similar to B13, since they are made by bioisosterically substituting the carboxyl moiety of B13 with sulfone group. Twenty B13 sulfonamides were evaluated for their in vitro cytotoxicities against human colon cancer HT-29 and lung cancer A549 cell lines using MTT assays. Replacement of the amide group with a sulfonamide group increased cytotoxicity in both cancer cell lines. The sulfonamides with long alkyl chains exhibited activities two to three times more potent than that of B13 and compound (15) had the most potent activity with $IC_{50}$ values of 27 and $28.7{\mu}M$ for HT-29 and A549, respectively. The comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) were used to carry out QSAR molecular modeling of these compounds. The predictive CoMSIA models for HT-29 and A549 gave cross-validated q2 values of 0.703 and 0.830, respectively. From graphical analysis of these models, we suppose that the stereochemistry of 1,3-propandiol is not important for activity and that introduction of a sulfonamide group and long alkyl chains into B13 can increase cytotoxicity.

• Asian Pacific Journal of Cancer Prevention
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• v.17 no.5
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• pp.2637-2641
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• 2016

Tumor necrosis factor ($TNF-{\alpha}$), an inflammatory cytokine that plays an important role in the control of cell proliferation, differentiation, and apoptosis, has previously been used in anti-cancer therapy. However, the therapeutic applications of $TNF-{\alpha}$ are largely limited due to its general toxicity and anti-apoptotic influence. To overcome this problem, the present study focused on the effect of active constituents isolated from a medicinal plant on $TNF-{\alpha}$-induced apoptosis in human colon adenocarcinoma (HT-29) cells. Nimbolide from Azadirachta indica was evaluated for cytotoxicity by methyl tetrazolium 3-[4,5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide (MTT) assay and phase contrast microscopy. Effects on apoptotic signaling proteins were investigated using Western blot analysis. Nimbolide showed cytotoxicity against HT-29 cells that was significantly different from the control group (p<0.01), a concentration of $10{\mu}M$ significantly inducing cell death (p<0.01). In combination with $TNF-{\alpha}$, nimbolide significantly enhanced-induced cell death. In apoptotic pathway, nimbolide activated c-Jun N-terminal kinase (JNK) phosphorylation, BH3 interacting-domain death agonist (Bid) and up-regulated the death receptor 5 (DR5) level. In the combination group, nimbolide markedly sensitized $TNF-{\alpha}$-induced JNK, Bid, caspase-3 activation and the up-regulation of DR5. Our findings overall indicate that nimbolide may enhance $TNF-{\alpha}$-mediated cellular proliferation inhibition through increasing cell apoptosis of HT-29 cells by up-reglation of DR5 expression via the JNK pathway.

• Biomolecules & Therapeutics
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• v.19 no.2
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• pp.211-217
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• 2011

Panax ginseng CA Meyer, a herb from the Araliaceae, has traditionally been used as a medicinal plant in Asian countries. Ginseng extract fermented by ginsenoside-${\beta}$-glucosidase treatment is enriched in ginsenosides such as Rh2 and Rg3. Here we show that a fermented ginseng extract, BST204, has anti-proliferative and anti-invasive effects on HT-29 human colon cancer cells. Treatment of HT-29 cells with BST204 induced cell cycle arrest at $G_1$ phase without progression to apoptosis. This cell cycle arrest was accompanied by up-regulation of tumor suppressor proteins, p53 and p21$^{WAF1/Cip1}$, down-regulation of the cyclin-dependent kinase/cyclins, Cdk2, cyclin E, and cyclin D1 involved in $G_1$ or $G_1/S$ transition, and decrease in the phosphorylated form of retinoblastoma protein. In addition, BST204 suppressed the migration of HT-29 cells induced by 12-O-tetradecanoylphorbol-13-acetate, which correlated with the inhibition of metalloproteinase-9 activity and extracellular signal-regulated kinase activity. The effects of BST204 on the proliferation and the invasiveness of HT-29 cells were similar to those of Rh2. Taken together, the results suggest that fermentation of ginseng extract with ginsenoside-${\beta}$-glucosidase enhanced the anti-proliferative and the anti-invasive activity against human colon cancer cells and these anti-tumor effects of BST204 might be mediated in part by enriched Rh2.

• Natural Product Sciences
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• v.15 no.4
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• pp.185-191
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• 2009

Intestinal epithelial cells (IEC) play an important role in the mucosal immune system. IEC-derived mediators of inflammatory cascades play a principal role in the development of colon inflammation. The aim of this study was to investigate the inhibitory effect of aqueous extracts of Schizandra chinensis fruits (SC-Ex) on the production of inflammatory mediators by the human colonic epithelial cells. HT-29 cells were stimulated with dextran sulfate sodium in the presence or absence of SC-Ex to examine the cytoprotection and production of IL-8 and reactive oxygen species (ROS). It was shown that dextran sulfate sodium (DSS) caused the reduction of cell viability and production of IL-8 and ROS in DSS-treated HT-29 cells. We observed that the treatment of SC-Ex protected significantly cell proliferation from DSS-induced damage in dose-dependent manner. SC-Ex (10 and 100 ${\mu}g$/ml) also suppressed DSS-induced production of IL-8 mRNA and protein. Moreover, DSS-induced ROS production was inhibited markedly by the treatment of 100 ${\mu}g$/ml SC-Ex. These results suggest that SC-Ex has the protective effects on DSS-induced cell damage and the release of inflammatory mediators in the intestinal epithelial cells.