HT-29 Human Colon Cancer Cells Research Articles

Phytochemical screening, antioxidant, anti-diabetic and cytotoxic activity of leaves of Pandanus canaranus Warb

Pandanus, a traditional medicinal plant is used for the ailment of several diseases in India. The extract prepared from leaves was tested for its secondary metabolites and also to phenolics, alkaloids, saponins and flavonoids. Subsequently, the antioxidant activity was observed using DPPH, reducing power, FRAP, metal chelating activity and total antioxidant assays. It is evident that more secondary metabolites were present in ethyl acetate and methanol solvents enriched in free radicals scavenging ability compared with other solvents. Further, the extracts were tested and observed for potential presence of anti-diabetic activity with strong inhibition of α-amylase and α-glucosidase which suggests that the absorption and digestion of carbohydrates would possibly reduce the blood glucose levels and hence help management of type-2 diabetes. Though the ethyl acetate and methanol extract exhibited cytotoxicity and apoptosis activity on human HT-29 colon cancer cell line via MTT, AO/EtBr and DAPI staining assays that inhibits cancer cell line which in turn prevents cancer.

Comparison of anticarcinogenic properties of Viburnum opulus and its active compound p-coumaric acid on human colorectal carcinoma.

Resistance to therapeutic agents and the highly toxic side effects of synthetic drugs has spurred new research in the treatment of colon cancer, which has high morbidity and mortality ratios. This study aims to clarify the molecular mechanisms of the anticarcinogenic properties of methanol extract of Viburnum opulus L. (EVO)and its main active compound, trans-p -coumaric acid ( p -CA), on human colon cancer cells (DLD-1, HT-29, SW-620, Caco-2) and healthy colon epithelial cells (CCD-18Co). The effects of EVO on controlled cell death (apoptosis) and the cell division cycle were determined by flow cytometry. Alteration in mRNA and protein expressions of switch genes in colorectal carcinoma (APC, MLH1, TP53, SMAD4, KRAS, and BRAF) were determined by qRT-PCR and Western blot, respectively. Our results show that EVO possesses a strong reducing capacity and free-radical scavenging activity. HPLC analyses prove that p -CAis the main compound of EVO. EVO and p -CA inhibit the proliferation of human colon cancer cells DLD-1 and HT-29 in a dose-dependent manner. EVO increases apoptosis of DLD-1 cells and halts the cell cycle in the G2 stage in HT-29 cells. mRNA and protein expressions of p53 and SMAD-4 are upregulated, while BRAFs are downregulated. The results were directly proportional to p -CA. EVO and p -CA up- and downregulate switch genes and protein expressions of DLD-1 cells, which alter the expression of 186 other genes. This is the first study of pharmacological exploration of V.opulus in human colon cancer. Its antiproliferative effects may be due to the presence of p -CA.

The Effects of Doxorubicin, Ethanol Extract and Flavonoid-rich Fraction of Euphorbia Splendida Mobayen on the PARP Level, and APC Gene Expression in HT-29 Human Colon Cancer Cell Line

Background: Natural products derived from various sources are being used to develop chemotherapeutic drugs. Euphorbiaceae is widely used to treat different types of cancers. Colorectal cancer (CRC) is the second and third cause of cancer in women and men, respectively. CRC is strongly associated with the deregulation of the Adenomatous polyposis coli (APC) gene and Poly [ADP-ribose] polymerase (PPAR) protein. Objectives: The current study aimed to examine the effect of doxorubicin, ethanol extract, and the flavonoid-rich fraction of Euphorbia Splendida Mobayen on colon cancer HT-29 cell line death, APC gene expression, and PPAR concentration. Methods: Following treatment of cells by Euphorbia ethanol extract, Euphorbia flavonoid-rich fraction, and doxorubicin, cell viability assay was used to investigate the viability status of the HT-29 cell line. Total RNA was isolated from the cell line and converted into cDNA. The expression level of the APC gene was determined by quantitative real-time PCR. Poly (ADP-ribose) polymerase (PPAR) protein was detected by the ELIZA method. Results: We found that Euphorbia ethanol extract, Euphorbia flavonoid-rich fraction, and doxorubicin can stimulate dose-dependent cell death in the HT-29 cell line, increase ACP gene expression (P = 0.001, P = 0.041, P = 0.019), and decrease PARP level (P = 0.001, P = 0.001, P = 0.001, respectively). Conclusions: The findings indicated that doxorubicin, ethanol extract, and the flavonoid-rich fraction of Euphorbia Splendida Mobayen had cytotoxic effects on human colon cancer HT-29 cell line by possibly stimulating apoptosis.

Proteomic Approach for Searching for Universal, Tissue-Specific, and Line-Specific Markers of Extracellular Vesicles in Lung and Colorectal Adenocarcinoma Cell Lines.

Tumor-derived extracellular vesicles (EVs), including exosomes, contain proteins that mirror the molecular landscape of producer cells. Being potentially detectible in biological fluids, EVs are of great interest for the screening of cancer biomarkers. To reveal universal, tissue-specific, and line-specific markers, we performed label-free mass spectrometric profiling of EVs originating from the human colon cancer cell lines Caco-2, HT29, and HCT-116, as well as from the lung cancer cell lines NCI-H23 and A549. A total of 651 proteins was identified in the EV samples using at least two peptides. These proteins were highly enriched in exosome markers. We found 11 universal, eight tissue-specific, and 29 line-specific markers, the levels of which were increased in EVs compared to the whole lysates. The EV proteins were involved in the EGFR, Rap1, integrin, and microRNA signaling associated with metastasis and cancer progression. An EV protein-based assay could be developed as a liquid biopsy tool.

Arachidonic acid induces ER stress and apoptosis in HT-29 human colon cancer cells

ABSTRACT Polyunsaturated fatty acids (PUFAs) have important functions in biological systems. The beneficial effects of dietary PUFAs against inflammatory diseases, cardiovascular diseases, and metabolic disorders have been shown. Studies using cancer cells have presented the anti-tumorigenic effects of docosahexaenoic acid (DHA), an n-3 PUFA, while arachidonic acid (AA), an n-6 PUFA, has been shown to elicit both pro- and anti-tumorigenic effects. In the current study, the anti-tumorigenic effects of AA were evaluated in HT-29 human colon cancer cells. Upon adding AA in the media, more than 90% of HT-29 cells died, while the MCF7 cells showed good proliferation. AA inhibited the expression of SREBP-1 and its target genes that encode enzymes involved in fatty acid synthesis. As HT-29 cells contained lower basal levels of fatty acid synthase, a target gene of SREBP-1, than that in MCF7 cells, the inhibitory effects of AA on the fatty acid synthase levels in HT-29 cells were much stronger than those in MCF-7 cells. When oleic acid (OA), a monounsaturated fatty acid that can be synthesized endogenously, was added along with AA, the HT-29 cells were able to proliferate. These results suggested that HT-29 cells could not synthesize enough fatty acids for cell division in the presence of AA because of the suppression of lipogenesis. HT-29 cells may incorporate more AA into their membrane phospholipids to proliferate, which resulted in ER stress, thereby inducing apoptosis. AA could be used as an anti-tumorigenic agent against cancer cells in which the basal fatty acid synthase levels are low.

A Natural Flavone Tricin from Grains Can Alleviate Tumor Growth and Lung Metastasis in Colorectal Tumor Mice.

Tricin, a flavone isolated from rice bran, has been shown to be chemopreventive in a colorectal cancer (CRC) mouse model. This study aimed to illustrate the inhibitory activities of tricin in colon cancer cells and in a metastatic CRC mouse model. BALB/c mice injected with mouse Colon26-Luc cells into the rectum wall were treated with tricin (37.5 mg/kg) daily for 18 days. Orthotopic colon tumor growth and metastasis to lungs were assessed by in vivo bioluminescence imaging. Results showed that tricin suppressed Colon-Luc cells motility and downregulated phosphorylated Akt, Erk1/2 and NF-κB expressions of human colon cancer HT-29 cells. While tricin treatment suppressed tumor growth and lung metastasis as well as altered the populations of myeloid-derived suppressor cells and regulatory T cells in spleens. In summary, the tumor microenvironment modulatory and anti-metastatic effects of tricin in colon cancer mouse model were shown for the first time, suggesting the potential development of tricin-containing food supplements for CRC patients.

A Scaffold-Free 3-D Co-Culture Mimics the Major Features of the Reverse Warburg Effect In Vitro.

Most tumors consume large amounts of glucose. Concepts to explain the mechanisms that mediate the achievement of this metabolic need have proposed a switch of the tumor mass to aerobic glycolysis. Depending on whether primarily tumor or stroma cells undergo such a commutation, the terms ‘Warburg effect’ or ‘reverse Warburg effect’ were coined to describe the underlying biological phenomena. However, current in vitro systems relying on 2-D culture, single cell-type spheroids, or basal-membrane extract (BME/Matrigel)-containing 3-D structures do not thoroughly reflect these processes. Here, we aimed to establish a BME/Matrigel-free 3-D microarray cancer model to recapitulate the metabolic interplay between cancer and stromal cells that allows mechanistic analyses and drug testing. Human HT-29 colon cancer and CCD-1137Sk fibroblast cells were used in mono- and co-cultures as 2-D monolayers, spheroids, and in a cell-chip format. Metabolic patterns were studied with immunofluorescence and confocal microscopy. In chip-based co-cultures, HT-29 cells showed facilitated 3-D growth and increased levels of hexokinase-2, TP53-induced glycolysis and apoptosis regulator (TIGAR), lactate dehydrogenase, and: translocase of outer mitochondrial membrane 20 (TOMM20), when compared with HT-29 mono-cultures. Fibroblasts co-cultured with HT-29 cells expressed higher levels of mono-carboxylate transporter 4, hexokinase-2, microtubule-associated proteins 1A/1B light chain 3, and ubiquitin-binding protein p62 than in fibroblast mono-cultures, in both 2-D cultures and chips. Tetramethylrhodamin-methylester (TMRM) live-cell imaging of chip co-cultures revealed a higher mitochondrial potential in cancer cells than in fibroblasts. The findings demonstrate a crosstalk between cancer cells and fibroblasts that affects cellular growth and metabolism. Chip-based 3-D co-cultures of cancer cells and fibroblasts mimicked features of the reverse Warburg effect.

Designing a pH-responsive drug delivery system for the release of black-carrot anthocyanins loaded in halloysite nanotubes for cancer treatment

Anthocyanins extracted from black-carrot were encapsulated into halloysite nanotubes (HNT) as a potential carrier for the first time. Anthocyanins were loaded into HNT under vacuum conditions. TEM images indicated the successful entrance of anthocyanins into the lumen of HNT. Anthocyanins loading efficiency was determined to be 4% using thermogravimetric analysis (TGA). XRD patterns showed that all reflections shifted to higher values for HNT loaded with anthocyanins compared to pure HNT which indicates reduced d-value of the crystalline forms. In vitro release of anthocyanins was studied under gastrointestinal conditions using the pH-differential method. The drug release from HNT was modeled and based on R2 values, the Korsmeyer-Peppas model was introduced as the best model predicting the release behavior. Cytotoxic effects of the samples against MCF-7 human breast cancer and HT-29 colon cancer cell lines were evaluated using SRB assay. The cell viability was reduced by 2 folds against anthocyanins-loaded HNT compared to pure anthocyanins in both cell lines. Anthocyanins-loaded HNT is suggested as a promising pH-responsive drug delivery system for the treatment of cancer.

Anticancer Effects of Extracts from Three Different Chokeberry Species

Colon cancer risk appears to be lowered by consumption of a diet rich in fruits and vegetables. Chokeberries are rich in phytochemicals that may act as potent anticancer agents. Phytochemicals that are particularly abundant in chokeberries include anthocyanins and phenolic acids. In this study, we compared the growth inhibitory activity of three chokeberry extracts in HT-29 human colon cancer cells. The three extracts tested were derived from Aronia arbutifolia (red), Aronia prunifolia (purple), and Aronia melanocarpa (black). Cells were incubated with either red, purple, or black chokeberry extracts and cell viability was quantified using the thiazolyl blue tetrazolium bromide (MTT) assay. The black chokeberry extract had the greatest effect in reducing cell proliferation. The extracts were also characterized for total phenols (Folin-Ciocalteu assay), total antioxidant activity (oxygen radical absorbance capacity assay), and levels of bioactive phenolic acids (high-performance liquid chromatography). The growth inhibitory activities of the extracts correlated well with total phenolic content, antioxidant activity, and levels of caffeic and chlorogenic acids. The black chokeberry extract had the greatest level of total phenols, antioxidant activity, and individual phenolic acids. This research suggests that the phenolic profile of foods such as chokeberries can help determine their cancer cell growth inhibitory activity.

Development, characterization and optimization of polymeric mucoadhesive microcapsules containing anticancer agent using response surface method: in vitro and in vivo study

The present research work covenants the preparation, characterisation and optimisation of mucoadhesive microcapsules containing paclitaxel through ionic gelation method using 32 statistical factorial designs. The effect of mixing proportion of primary polymer sodium alginate to copolymer (X1) and speed of magnetic stirrer (X2) on the microcapsules size (Y1), efficiency of paclitaxel encapsulation (Y2), and percentage yield (Y3) was optimised. The morphology of microcapsules was characterised and evaluated by in vitro and in vivo tests to study the swelling characteristics, mucoadhesion and drug release characteristics, followed by MTT assay on human HT-29 colon cancer cell lines. The size of prepared microcapsules was within the range of 361 ± 4.50 to 931 ± 22.41; encapsulation efficiency (%) was within the range of 42.72 ± 0.43 to 98.12 ± 0.43 %. The in vitro paclitaxel released over 24 hours were in a range of 82.15 ± 3.43 % to 96.75 ± 2.41 %. The controlled release pattern of paclitaxel was observed from the in vitro drug release study of microcapsules. The prepared microcapsules that showed better mucoadhesion were in the range of 73.66 ± 1.42 to 97.85 ± 1.08 % for a period of 6 h. The in vivo pharmacokinetic study conducted in rats resulted in high Tmax, the area under the curve and mean residence time for microcapsules as compared to that of the marketed formulation. It could be concluded that the microcapsules containing povidone polymer showed superior results.

JPH203, a newly developed anti-cancer drug, shows a preincubation inhibitory effect on L-type amino acid transporter 1 function

JPH203 is a novel anti-cancer drug targeting L-type amino acid transporter 1 (LAT1), which plays a primary role in the uptake of essential amino acids in tumor cells. Although a co-incubation inhibitory effect of JPH203 has been shown in a conventional uptake assay, its preincubation inhibitory effects have remained undetermined. Therefore, we aimed to characterize the preincubation inhibitory effects of JPH203 on LAT1 function using leucine uptake assays in LAT1-positive human colon cancer HT-29 cells. Preincubation of the cells with JPH203 (0.3 μM for 120 min) decreased the activity level to 30% of that in dimethylsulfoxide-treated cells. Similarly, in time-dependency analysis, preincubation of HT-29 cells with 10 μM JPH203 for 30, 60, and 120 min decreased the leucine uptake activity (42%, 32%, and 28% of that in control cells, respectively). Furthermore, the IC50 value of the combination of preincubation and co-incubation effects was lower than that of co-incubation inhibition alone (34.2 ± 3.6 nM vs. 99.2 ± 11.0 nM). In conclusion, we revealed that JPH203 has the capability to inhibit LAT1 function through preincubation effects. Moreover, preincubation synergistically enhances the co-incubation inhibitory effects. These findings provide a novel insight into the anti-cancer effects of JPH203 in cancer therapy.

Negative impact of high body mass index on cetuximab-mediated cellular cytotoxicity against human colon carcinoma cells

This study assessed the relationship between the ability of Natural Killer (NK) cells to activate antibody-dependent cellular cytotoxicity against human HT29 colorectal cancer cells exposed to cetuximab and the body mass index of the human subjects from whom the NK cells had been obtained. NK cells obtained from 73 human donors were co-incubated with HT-29 human colon cancer cells in the presence or absence of cetuximab. Antibody-dependent cellular cytotoxicity was assessed by measuring LDH release. A significant negative correlation was observed between body mass index and cetuximab-induced antibody-dependent cellular cytotoxicity. NK cells obtained from subjects who were overweight or with obesity were less efficient in killing cetuximab-treated HT29 cells than those derived from normal weight donors. Our results suggest that the success of cetuximab-containing regimens might be impaired in overweight and obese patients with colorectal cancer.

The heparan sulfate proteoglycan syndecan‐1 regulates colon cancer stem cell function via a focal adhesion kinase—Wnt signaling axis

In colon cancer, downregulation of the transmembrane heparan sulfate proteoglycan syndecan-1 (Sdc-1) is associated with increased invasiveness, metastasis, and dedifferentiation. As Sdc-1 modulates signaling pathways relevant to stem cell function, we tested the hypothesis that it may regulate a tumor-initiating cell phenotype. Sdc-1 small-interfering RNA knockdown in the human colon cancer cell lines Caco2 and HT-29 resulted in an increased side population (SP), enhanced aldehyde dehydrogenase 1 activity, and higher expression of CD133, LGR5, EPCAM, NANOG, SRY (sex-determining region Y)-box 2, KLF2, and TCF4/TCF7L2. Sdc-1 knockdown enhanced sphere formation, cell viability, Matrigel invasiveness, and epithelial-to-mesenchymal transition-related gene expression. Sdc-1-depleted HT-29 xenograft growth was increased compared to controls. Decreased Sdc-1 expression was associated with an increased activation of β1-integrins, focal adhesion kinase (FAK), and wingless-type (Wnt) signaling. Pharmacological FAK and Wnt inhibition blocked the enhanced stem cell phenotype and invasive growth. Sequential flow cytometric SP enrichment substantially enhanced the stem cell phenotype of Sdc-1-depleted cells, which showed increased resistance to doxorubicin chemotherapy and irradiation. In conclusion, Sdc-1 depletion cooperatively enhances activation of integrins and FAK, which then generates signals for increased invasiveness and cancer stem cell properties. Our findings may provide a novel concept to target a stemness-associated signaling axis as a therapeutic strategy to reduce metastatic spread and cancer recurrence. DATABASES: The GEO accession number of the Affymetrix transcriptomic screening is GSE58751.

Cytotoxic and Optically Active Pyrisulfoxins From the Endophytic Streptomyces albolongus EA12432.

R-Pyrisulfoxin C (1), S-pyrisulfoxin D [(+)-2], R-pyrisulfoxin D [(–)-2], pyrisulfoxin E (13), S-pyrisulfoxin F [(+)-14], and R-pyrisulfoxin F [(–)-14], six new caerulomycin derivatives with a 2,2′-bipyridine skeleton, were obtained from the cultures of the endophytic Streptomyces albolongus EA12432 with Aconitum carmichaeli (Ranunculaceae). Additionally, the racemic pyrisulfoxins A [(±)-3] and B [(±)-4] were further purified as optically pure compounds and identified the configurations for the first time. The racemic pyrisulfoxin D [(±)-2] displayed significant cytotoxicity against a series of cancer cell lines with IC50 values ranging from 0.92 to 9.71 μM. Compounds 7, 8, and (±)-3 showed cytotoxicity against the HCT-116, HT-29, BXPC-3, P6C, and MCF-7 cell lines. Notably, compounds 7 and 8 have a strong inhibition both on the proliferation of human colon cancer cells HCT-116 and HT-29 with IC50 values ranging from 0.048 to 0.2 μM (doxorubicin, 0.21 and 0.16 μM), and compound 1 showed a selective inhibition on the proliferation of the gastric carcinoma cell lines, N87, with an IC50 value of 8.09 μM. Optically pure compounds R(–)-14 and S(+)-14 showed weak cytotoxicity against HCT-116 and MCF-7 cell lines with the IC50 values of 14.7 μM and 10.4 μM, respectively. Interestingly, compounds 1 and (±)-2 didn't show cytotoxic activity against two human normal cell lines, HEK-293F and L02, with IC50 values >100 μM.

The roles of SHANK1 in the development of colon cancer.

SH3 and multiple ankyrin repeat domains protein 1 (SHANK1) belongs to a family of postsynaptic scaffolding proteins. In this study, we found that SHANK1 was abnormally high expressed in colon cancer tissues compared to normal tissues. Colon cancer patients with low SHANK1 expression had better prognosis. Furthermore, the expression of SHANK1 was knocked down in human colon cancer cell lines HCT116 and HT29 and the role of SHANK1 was investigated in colon cancer tumorigenesis. Our results showed that the knockdown of SHANK1 inhibited the survival and proliferation of both cells. The migration of these two cell lines was significantly reduced and the apoptosis was induced compared with control cells. The Bax/Bcl-2 ratio in both cell lines that SHANK1 was knocked down was increased, which is a signal that the mitochondrial apoptotic pathway was triggered. In addition, we observed that knockdown of SHANK1 reduced the expression of phosphorylated forms of AKT and mTOR. These data suggested that loss of SHANK1 inhibited viability and induced apoptosis of HCT116 and HT29 cells through the AKT/mTOR signaling pathway. Our data revealed that SHANK1 played important roles in the growth of colon cancer cells and may be used as a novel strategy for colon cancer therapy. SIGNIFICANCE OF THE STUDY: Herein, we reported that SHANK1 was abnormally high expressed in colon cancer tissues and associated with worse prognosis of patients. In addition, knockdown of SHANK1 inhibited viability and induced apoptosis in colon cancer cell lines through AKT/mTOR signaling pathways. These data suggest that SHANK1 may be a new oncogene in colon cancer. This study reveals the role of SHANK1 in addition to neuronal development and cognitive development. And it provides a new potential target for the prediction and treatment of colon cancer.

Preparation and characterization of PLGA-PEG-PLGA polymeric nanoparticles for co-delivery of 5-Fluorouracil and Chrysin

Despite significant advances in cancer therapy, chemotherapeutic agents are still the main types of drugs used to treat cancer patients. 5-Fluorouracil (5-FU) is the first-line treatment in several types of human cancers, however, nonspecific function, low plasma half-life, and high doses toxicity are the important barrier to achieve efficient response in cancer patients. The use of polymeric nanoparticles (NPs) for tumor targeted delivery of 5-FU in combination with other potent anticancer agent is considered an important strategy to enhance the therapeutic efficacy of 5-FU. In this study, we proposed to use PLGA-PEG-PLGA NPs to co-encapsulate 5-FU and Chrysin, a natural compound known to enhance the therapeutic efficacy of chemotherapy. NPs were prepared by double emulsion method and characterized for size and drug encapsulation efficacy. The cell growth inhibitory effect of prepared NPs was assessed by MTT assay in HT29 human colon cancer cell line. The analysis of NPs by dynamic light scattering showed that the developed NPs have average size of 40 nm. The encapsulation efficiency of NPs was 81.3% and 97.5% for 5-FU and Chrysin, respectively. Furthermore, the NPs showed a remarkable uptake in HT29 cells. NPs loaded with both 5-FU and Chrysin (5-FU@Chrysin loaded NPs) were found to have significantly higher growth inhibitory effects compared with NPs loaded with each drug alone in HT29 cell line. The synergistic anticancer effects of 5-FU and Chrysin loaded in NPs were confirmed with the combination index (CI) being 0.35. CI for combination therapy with free 5-FU and Chrysin was found to be 0.73, indicating weaker synergistic anticancer effects of these two drugs in free forms as compared with 5-FU@Chrysin loaded NPs. These finding indicates that co-delivery of 5-FU and Chrysin with PLGA-PEG-PLGA copolymer can be used to improve the therapeutic and functional delivery efficacy of 5-FU and Chrysin in cancer.

Effect of Hydrogen Sulfide‐Releasing Compounds on Proliferation of Human Colon Cancer HT29 Cells

Both natural and synthetic compounds that release hydrogen sulfide (H2S) such as diallyl trisulfide and naproxen‐H2S hybrids growth and proliferation of human colon HT29 cells (Lai et al. J. Cell. Mol. Med 19: 474, 2014; Kodela et al. Drug Des. Dev. Ther. 9:4873, 2015). Aims of the present study were: (a) to investigate the pharmacological action of L‐cysteine (substrate for the biosynthesis of H2S) and NaHS (H2S‐releasing compound) on proliferation of HT29 cells at different time‐points, and (b) to study the role of enzymes (cystathionine β‐synthase, CBS and cystathionine γ‐lyase, CSE) and K+‐ATP channels on the response elicited by H2S‐producing compounds in HT29 cells. The colon adenoma HT29 cells were treated with different concentrations of L‐cysteine (1 μM – 1 mM) and NaHS (1 μM – 1 mM) for 24 and 48 hours, in the absence and presence of inhibitors of CBS and CSE, amino‐oxyacetic acid (AOA, 1 mM), an inhibitor of K+‐ATP channels, glibenclamide (GLB, 10 μM). After incubation, the anti‐proliferative response on cells caused by various agents was determined by MTT Assay. At equimolar concentrations (10 μM and 1 mM), both L‐cysteine and NaHS caused concentration‐dependent inhibitions of cell proliferation after incubation for 24 h, but increased cell growth at these same concentrations after 48 h of incubation. For instance, at 10 μM, both L‐cysteine and NaHS increased cell proliferation by 37% and 51% after 48 h of incubation. Pretreatment of cells with AOA (1 mM) had no significant (P>0.05) effect on the inhibitory response caused by L‐cysteine and NaHS after 24 h incubation. Interestingly, AOA blocked and even reversed the increase in proliferation caused by L‐cysteine (10 μM and 1 mM) and NaHS (10 μM and 1 mM) after 48 h incubation. After 24 h, both L‐cysteine and NaHS caused additional inhibitory effects on cell proliferation in HT 29 cells pretreated with GLB (10 μM). GLB blocked and even reversed the increase in proliferation caused by L‐cysteine and NaHS in HT29 cells after 48 h incubation. We conclude that H2S‐releasing compounds can elicit a time‐dependent inhibitory or excitatory action on the proliferation of HT29 cells. The inhibitory/excitatory effects caused by these compounds depends, at least in part, on endogenous biosynthesis of new gas and the activity of K+‐ATP channels in HT29 cells.Support or Funding InformationThis research is supported by Tittle III under the Award Number P031B090216;Title III, Part B, Historically Black Graduate Institutions (HBGI) (CFDA No. 84.031B).

Cytotoxic and pharmacokinetic studies of Indian seaweed polysaccharides for formulating raindrop synbiotic candy

Gut microbiome evidenced as the assembling mode of action facilitates the relationship of environmental factors (such as diet and lifestyle) with colorectal cancer. The cytotoxic and anticancer studies of the enzymatically extracted polysaccharides from selected Indian seaweeds (such as S. wightii, E. compressa, and A. spicifera) on Raw 264.7 macrophage and HT-29 human colon cancer cell line were investigated. E. compressa showed nitric oxide production up to a concentration of 6.99 ± 0.05 μM. The polysaccharide extract of seaweed (PES), A. spicifera (100 μg/ml) had shown the highest in-vitro cytotoxicity effect on HT-29 cells up to 52.13 ± 1.4%. Absorption, distribution, metabolism and excretion (ADME) predictions were performed for exploring the possibility of anti-cancer drug development. The formulated synbiotic candy exhibited post storage survivability of probiotic species L. plantarum NCIM 2083 up to 107 CFU/ml until three weeks and it could be an aesthetic functional food for treating colon cancer.

Anticancer Activities of Meroterpenoids Isolated from the Brown Alga Cystoseira usneoides against the Human Colon Cancer Cells HT-29.

Colorectal cancer (CRC) is one of the most common types of cancers and a leading cause of cancer death worldwide. The current treatment for CRC mainly involves surgery, radiotherapy, and chemotherapy. However, due to the side effects and the emergence of drug resistance, the search for new anticancer agents, pharmacologically safe and effective, is needed. In the present study, we have investigated the anticancer effects of eight algal meroterpenoids (AMTs, 1-8) isolated from the brown seaweed Cystoseira usneoides and their underlying mechanisms of action using HT-29, a highly metastatic human colon cancer cell line. All the tested meroterpenoids inhibited the growth of HT-29 malignant cells and were less toxic towards non-cancer colon cells, with the AMTs 1 and 5 exhibiting selectivity indexes of 5.26 and 5.23, respectively. Treatment of HT-29 cells with the AMTs 1, 2, 3, 4, 5, and 7 induced cell cycle arrest in G2/M phase and, in some instances, apoptosis (compounds 2, 3, and 5). Compounds 1-8 also exhibited significant inhibitory effects on the migration and/or invasion of colon cancer cells. Mechanistic analysis demonstrated that the AMTs 1, 2, 5, 6, 7, and 8 reduced phosphorylation levels of extracellular signal-regulated kinase (ERK) and the AMTs 2, 3, 4, 5, 7, and 8 decreased phosphorylation of c-JUN N-terminal kinase (JNK). Moreover, the AMTs 1, 2, 3, 4, 7, and 8 inhibited phosphorylation levels of protein kinase B (AKT) in colon carcinoma cells. These results provide new insights into the mechanisms and functions of the meroterpenoids of C. usneoides, which exhibit an anticancer effect on HT-29 colon cancer cells by inducing cell cycle arrest and apoptosis via the downregulation of ERK/JNK/AKT signaling pathways.

High Levels of Apoptosis Are Induced in the Human Colon Cancer HT-29 Cell Line by Co-Administration of Sulforaphane and a Peptide Nucleic Acid Targeting miR-15b-5p.

Sulforaphane (SFN) is one of most important dietary constituents of broccoli (Brassica oleracea) and other cruciferous vegetables, which have been reported to exhibit health benefits, including prevention and therapy of cancer, such as colorectal carcinoma (CRC). The objective of this study was to determine whether the anticancer effect of SFN on colon cancer HT-29 cell line could be improved by the combined treatment with molecules inhibiting microRNAs (miRNAs) involved in CRC. As miRNA inhibiting molecules we focused on peptide-nucleic acids (PNAs). As miRNA to be targeted, miR-15b-5p was selected on the basis of several information present in the literature and confirming that miR-15b-5p is overexpressed in colon cancer patients, and that its targeting decreases cell migration and metastasis in colorectal cancer. In this article, we described for the first time the efficacy of targeting miR-15b-5p by using a PNA against miR-15b-5p (R8-PNA-a15b), functionalized with an octoarginine peptide (R8) for maximizing cellular uptake. The miR-15b-5p downregulation in the colon cancer HT-29 cell line was associated with inhibition of in vitro cell growth and activation of the proapoptotic pathway, demonstrated by a sharp increase of late apoptotic cells in HT-29-treated cell populations. A second conclusion of this study is that the R8-PNA-a15b might be proposed in "combo-therapy" associated with SFN. To our knowledge, no report is available in the literature on a combination between SFN and miRNA-targeting molecules. Our data demonstrate that this combined treatment leads to a very high proportion of apoptotic HT-29 cells (over 85%), a value higher than the sum of the values of apoptotic cells obtained after singularly administered regents (either SFN or R8-PNA-a15b).