Eight previously undescribed diterpenoids, along with eleven previously reported analogues, were obtained from the supercritical CO2 extracts of Torreya grandis aril. The structures of these compounds were elucidated based on HRESIMS, NMR, ECD, and single-crystal X-ray diffraction data. In the MTT assay, compound 18 exhibited significant inhibitory effects on two human colon cancer cell lines, HT-29 and HCT 116 cells, with IC50 values of 7.37 μM and 6.55 μM, respectively. It was found that compound 18 induced apoptosis and significantly inhibited the migration of HCT 116 colon cancer cells in a concentration-dependent manner.

OBJECTIVE: GlyH-101 and CaCCinh-A01 are effective blockers of cystic fibrosis transmembrane conductance regulator (CFTR) and Ca2+-activated chloride channels (CaCCs), respectively. Available evidence suggests that GlyH-101 and CaCCinh-A01 can suppress cell proliferation, block invasion and metastasis, and cause several cancer cell types to undergo apoptosis, demonstrating their anti-tumor properties. The aim of this study was to investigate the effect of GlyH-101 and CaCCinh-A01 on HT-29 cell activity and to suggest the possible molecular mechanisms by which inhibitors of CFTR and CaCCs inhibit HT-29 cell activity.
METHODS: Human colon HT-29 cancer cells were treated with GlyH-101 or CaCCinh-A01 or GlyH-101 plus CaCCinh-A01 complex. Cell viability was determined by MTT assay, the apoptosis and cell cycle were determined by flow cytometry, and reactive oxygen species (ROS) leves were determined by 2\',7\'-Dichlorodihydrofluorescein diacetate staining. The expression of proteins related to apoptosis and cell cycle regulation was measured by western blotting.
RESULTS: The proliferative ability of HT-29 cells was dose- and time-dependently reduced by GlyH-101 and CaCCinh-A01. Treatment with GlyH-101 and CaCCinh-A01 resulted in cell necrosis and apoptosis, up-regulated ROS levels, activated the mitochondrial apoptosis pathway, prompted arrest of the cell cycle in S phase, and increased the levels of proteins related to the cell cycle. Additionally, the combination of these two inhibitors had a stronger regulatory effect on HT-29 cell proliferation than either GlyH-101 or CaCCinh-A01 treated alone.
CONCLUSIONS: GlyH-101 and CaCCinh-A01 inhibited cell proliferation through cell cycle arrest and mitochondrial-related pathways in vitro. The combination of these inhibitors could further enhance their anti-proliferative effects. Our findings propose new lead compounds with anti-colon cancer activity, and also provide new evidence for the effectiveness of chloride channels-targeted therapy in anticancer therapy.

Ulcerative colitis (UC) is closely associated with disruption of intestinal epithelial tight junction proteins. A variety of studies have confirmed that resveratrol (RSV), a natural polyphenolic compound, has a potential anti-inflammatory effect and can regulate the expression of tight junction proteins. However, the mechanism by which RSV regulates the expression of tight junction proteins in the intestinal epithelium remains unclear. Therefore, we investigated the potential effect of RSV on tight junction proteins in an HT-29 cell model of inflammation induced by lipopolysaccharide (LPS) and explored its mechanism of action. First, the downregulated expression of the tight junction proteins occludin, ZO-1, and claudin-1 in the HT-29 cell model of inflammation induced by LPS was reversed by incubation with RSV, accompanied by a decrease in the expression of tumor necrosis factor α-converting enzyme (TACE). Additionally, the Notch1 pathway was attenuated and the expression of the inflammatory factors IL-6 and TNF-α was decreased by treatment with RSV. Second, after Jagged-1 was used in combination with RSV to reactivate the Notch1 pathway, the protective effects of RSV against the LPS-induced reductions in the expression of the tight junction proteins occludin, ZO-1, and claudin-1 and the decreases in the levels of the inflammatory factors IL-6 and TNF-α were abolished. These results suggest that RSV might regulate the expression of tight junction proteins by attenuating the Notch1 pathway.

Rhododendron molle G. Don is one example of traditional Chinese medicine with important medicinal value. In this study, the effects of methanol extract of R. molle leaves (RLE) on colorectal cancer HT-29 cells and its potential molecular mechanism were investigated. MTT analysis showed that RLE could significantly inhibit the cell viability and migration of HT-29 cells in a concentration-dependent manner. Cell cycle analyses via flow cytometer suggested that RLE induced DNA fragmentation, indicative of apoptosis, and arrest at the S phase in HT-29 cells. Quantitative real-time PCR (qRT-PCR) analysis showed that RLE could upregulate the mRNA expression of p53 and p21 in HT-29 cells, which would result in HT-29 cells being blocked in S phase. Meanwhile, RLE could upregulate the expression of Bax, and downregulate the expression of Bcl-2, which would induce cell apoptosis. Further western blot analysis showed that the protein expression changes of Bax and P53 were basically consistent with the results of qRT-PCR. In addition, GC-MS analysis detected 17 potential anticancer components in R. molle. These results indicate that R. molle has significant anticancer activity, which provides some useful information for further study and clinical application for R. molle.

We designed specific small interfering RNA (siRNA) targeting vascular endothelial growth factor (VEGF) mRNA and synthesized oligo fragments, then siRNA was obtained by in vitro transcription and transfected into cultured human colon carcinoma cell line HT-29 with lipofectamine. We also analyzed the effect of the siRNA on proliferation of HT-29 cells by methyl thiazolyl tetrazolium (MTT) assay and expression level of VEGF mRNA of transfected cells by RT-PCR as well as amounts of secreted VEGF protein in the supernatant by enzyme linked immunosorbent assay (ELISA). Two groups of siRNA targeting human VEGF effectively inhibited proliferation of HT-29 cells after transfection. The secretion of VEGF protein also notably decreased, but the control scramble siRNA showed no effect.

Atractylodes macrocephala is known to exhibit multi-arrays of biologic activity in vitro. However, detail of its anti-tumor activity is lacking. In this study, the effects of atractylenolide I (AT-I), a bio-active compound present in Atractylodes macrocephala rhizome was studied in the human colorectal adenocarcinoma cell line HT-29. The results showed that AT-I induced apoptosis of human colon cancer cells through activation of the mitochondria-dependent pathway. The IC50 of AT-I was 277.6 μM, 95.7 μM and 57.4 μM, after 24, 48 and 72 h of incubation with HT-29, respectively. TUNEL and Annexin V-FITC/PI double stain assays showed HT-29 DNA fragmentation after cell treatment with various AT-I concentrations. Western blotting analysis revealed activation of both initiator and executioner caspases, including caspase 3, caspase 7, and caspase 9, as well as PARP, after HT-29 treatment with AT-I via downregulation of pro-survival Bcl-2, and upregulation of anti-survival Bcl-2 family proteins, including Bax, Bak, Bad, Bim, Bid and Puma. The studies show for the first time that AT-I is an effective drug candidate towards the HT-29 cell.

This study aimed to investigate the antioxidant activity and release behavior of anthocyanin (ANC) loaded within FA-g-MD wall (ANC-FA-g-MD microcapsule) in vitro. The microencapsulation of ANC was prepared by spray drying and displayed a biphasic release profile. The combination of ANC and FA-g-MD (0.0625-1 mg/mL) showed a higher antioxidant activity than that of both individuals. A possible intermolecular interaction between ANC and FA-g-MD was studied by UV-vis spectra. Intracellular reactive oxygen species (ROS), 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) test, and protein expression of quinone oxidoreductase 1(NQO1), glutathione reductase (GSR) and γ-glutamate cysteine ligase catalytic subunit (γ-GCLC) were measured through human colon cancer cells (HT-29). After a 24-hour incubation of the HT-29, the combinations (0-60 μg/mL) exhibited a high potential to diminish the ROS level. And the distinct upregulated expressions of GCLC and NQO1 of HT-29 were detected after treatment with combinations compared to those of single ones. These results suggested that the ANC-FA-g-MD microcapsules exerts enhanced antioxidant effect with capability of the modulation of GCLC and NQO1.

Adhesion to epithelial cells is considered important for Lactobacillus to exert probiotic effects. In this study, we found that trypsin treatment decreased the adhesion ability of Lactobacillus plantarum AR326 and AR269, which exhibit good adhesion ability, and surface proteins extracts increased the adhesion of the strains with poor adhesion ability. By SDS-polyacrylamide gel electrophoresis and mass spectrometry analysis, the main component of the surface proteins was detected and identified as a protein of approximately 37 kDa. It was 100% homologous with glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from L. plantarum WCFS1. The adhesion of AR326 and AR269 was decreased significantly by blocking with the anti-GAPDH antibody, and GAPDH restored the adhesion of AR326 and AR269 treated with trypsin. In addition, purified GAPDH significantly increased the adhesion of the strains with poor adhesion ability. These results indicated that GAPDH mediates the adhesion of these highly adhesive lactobacilli to epithelial cells and can be used to improve the adhesion ability of probiotics or other bacteria of interest.

UNASSIGNED: Proanthocyanidin-rich longan flower extract (LFP) has been previously shown to inhibit the proliferation and anchorage-independent growth in soft agar of two colorectal carcinoma (CRC) cells in vitro. In this report, we further examined the effects of LFP in a CRC spheroid model.
UNASSIGNED: A liquid-overlay assay employing HT-29 spheroids was used to evaluate the effects of LFP on cancer cell tumorigenesis, viability, and apoptosis. Associated effects on signaling path ways (epidermal growth factor receptor [EGFR], Akt) and apoptotic regulators were measured using Western blot.
UNASSIGNED: Treatment with LFP up to 200 μg/ml inhibited tumor growth in a dose-dependent manner and induced prominent apoptosis as measured by annexin V staining. Cells treated with LFP showed decreased EGFR and Akt phosphorylation with decreased expression of B-cell lymphoma 2.
UNASSIGNED: The ability of LFP to induce apoptosis in CRC spheroids warrants further investigation of its composition and identification of tumor-active components.

This study aims to investigate the effect of long non-coding RNA (lncRNA) Gas5 on proliferation, migration, invasion and apoptosis of colorectal cancer (CRC) HT-29 cell line.
CRC and normal tissues were collected and prepared from a total of 126 CRC patients, and normal intestinal epithelial cell line FHC and CRC cell lines (HCT-8, HT-29, HCT-116 and SW-480) were prepared. Gas5 expression was detected by quantitative reverse transcriptase-polymerase chain reaction. HT-29 cell line exhibiting the lowest Gas5 expression was selected for further experimentation and divided into blank, negative control and pcNDA-Gas5 groups. The cell counting kit-8 assay was used to test cell proliferation. Flow cytometry was applied to examine cell apoptosis. Transwell assay was performed to detect the migration and invasion of HT-29 cells. The mRNA and protein expression of factors in the classical proliferation (Akt/Erk) and apoptosis (caspase-9/caspase-3) pathways were detected.
Gas5 expression was lower in CRC tissues compared to the adjacent normal tissues, and is also lower in CRC cell lines than FHC cell line. Gas5 expression was associated with tumor size and TNM staging. Gas5 expression, distant metastasis, tumor differentiation and TNM staging were independent CRC prognostic factors. The results showed that elevated Gas5 expression inhibited proliferation, migration and invasion, but promoted apoptosis of CRC cells. Meanwhile, elevated Gas5 expression inhibited mRNA expression of Akt and Erk and protein expression of p-Akt and p-Erk, which promoted Casp9 mRNA and pho-Casp9 protein expression but inhibited Casp3 mRNA and pho-Casp3 protein expression.
The findings indicated that overexpression of lncRNA Gas5 can inhibit the proliferation, migration and invasion but promote apoptosis of CRC cells.