Increased ENT2 Expression and Its Association With Altered Purine Metabolism in Different Stages of Colorectal Cancer Cell Lines

Background Colorectal cancer (CRC) is one of the most prevalent malignant cancers worldwide. Although the purine metabolism pathway is known to be vital for cancer cells survival mechanism, not much is known on ENT2 role in CRC development and its association with purine metabolites. Hence this study is aimed to determine the level of hypoxanthine phosphoribosyl transferase (HPRT), hypoxanthine, uric acid (UA), and the activity of xanthine oxidase (XO) and relate the findings with the ENT2 expression level in different CRC stages. Methods and results Normal colon cell line; CCD-841CoN and a panel of human CRC cell lines; SW480, HCT15 and HCT116, representing different CRC stages; Dukes’ B, C, and D respectively, have been used to measure HPRT, hypoxanthine/xanthine, UA levels and the activity of XO by biochemical assays. The level of ENT2 gene expression was also performed by qRT-PCR. The levels of HPRT, hypoxanthine were significantly higher (P< 0.05), while XO and UA were lower (P< 0.05) in all CRC stages as compared to the normal colon cells. Furthermore, ENT2 expression was found to be increased in all CRC stages. Despite having the highest level of HPRT and hypoxanthine, ENT2 level is lower in Dukes' D when compared to Dukes' B and C. Conclusion The rate of salvage pathway is increased in CRC development as indicated by the elevated levels of HPRT and hypoxanthine in different CRC stages. Increase ENT2 expression implies its importance in assisting hypoxanthine uptake. This step is vital in order to increase DNA synthesis via hypoxanthine recycling. Thus, ENT2 may be a potential marker in therapeutic development.

Nigella Sativa’s Protection Against 7,12 Dimethylbenz [A] Anthracene -Induced Colon Carcinogenesis in Rats

Colon cancer is the third most common cause of death from cancer worldwide. Recently, natural products have been widely used as an alternative therapy for colon cancer. Previous studies have reported that Nigella sativa has chemopreventive activity in vitro and in vivo.This study aimed to evaluate the effect of Nigella sativa seed (NSS) on rat-colon cell after initiation of 7,12-dimethylbenz [a] anthracene. Rats were divided into five groups, 12 rats in each group: Group I was given 7,12dimetilbenz [a] anthracene (DMBA) orally 20 mg/kgBW twice a week for five weeks, group V is the solvent control group was given corn oil. The other three groups were given DMBA + NSS, at the dosage of 250 mg/kgBW, 500 mg/kgBW and 750 mg/kgBW. NSS extract was dissolved in corn oil and administered daily per oral during the next two weeks before and during the initiation of DMBA. After 16 weeks, all rats were sacrificed. H&E staining showed that necrosis activity was lower in treated groups compared to DMBA group. AgNOR staining showed mAgNOR was significantly decrease following the increasing dose of NSS (250 mg/kgBW, 500 mg/kgBW and 750 mg/kgBW) were subsequently 1.62 ± 0.086, 1.60 ± 0.101 and 1.39 ± 0.049 (p<0.05). The results showed that NNS reduce the damage of colon cells and inhibit colon cell proliferation in DMBA induced rats.

Potential Antioxidant and Anticancer Activities of Secondary Metabolites of Nostoc linckia Cultivated under Zn and Cu Stress Conditions

The objective of the present study is to determine the antioxidant and anticancer activities of Nostoc linckia extracts cultivated under heavy metal stress conditions (0.44, 0.88, and 1.76 mg/L for zinc and 0.158, 0.316, 0.632 mg/L for copper). Phycobiliprotein, phenolic compounds, flavonoids, and tannins were measured. Active ingredients of extracts were evaluated by GC-mass spectroscopy. The obtained results revealed that higher zinc and copper concentrations showed growth inhibition while 0.22 mg/L (Zn) and 0.079 mg/L (Cu) enhanced growth, reaching its maximum on the 25th day. Increases in catalase, lipids peroxidation, and antioxidants, as well as tannins and flavonoids, have been induced by integration of 0.88 mg/L (Zn) and 0.316 mg/L (Cu). Elevation of Zn concentration induced augmentation of antioxidant activity of crude extract (DPPH or ABTS), with superior activity at 0.44 mg/L zinc concentration (81.22%). The anticancer activity of Nostoc linckia extract (0.44 mg/L Zn) tested against four cancer cell lines: A549, Hela, HCT 116, and MCF-7. The extract at 500 µg/mL appeared the lowest cell viability of tested cell lines. The promising extract (0.44 mg/L Zn) recorded the lowest cell viability of 25.57% in cervical cell line, 29.74% in breast cell line, 33.10% in lung cell line and 34.53% in the colon cell line. The antioxidant active extract showed significant stability against pH with attributed increase in antioxidant activity in the range between 8–12. The extract can be used effectively as a natural antioxidant and anticancer after progressive testing.

Assessing the Biocompatibility of Multi-Anchored Glycoconjugate Functionalized Iron Oxide Nanoparticles in a Normal Human Colon Cell Line CCD-18Co

We have previously demonstrated that iron oxide nanoparticles with dopamine-anchored heterobifunctional polyethylene oxide (PEO) polymer, namely PEO-IONPs, and bio-functionalized with sialic-acid specific glycoconjugate moiety (Neu5Ac(α2-3)Gal(β1-4)-Glcβ-sp), namely GM3-IONPs, can be effectively used as antibacterial agents against target Escherichia coli. In this study, we evaluated the biocompatibility of PEO-IONPs and GM3-IONPs in a normal human colon cell line CCD-18Co via measuring cell proliferation, membrane integrity, and intracellular adenosine triphosphate (ATP), glutathione GSH, dihydrorhodamine (DHR) 123, and caspase 3/7 levels. PEO-IONPs caused a significant decrease in cell viability at concentrations above 100 μg/mL whereas GM3-IONPs did not cause a significant decrease in cell viability even at the highest dose of 500 μg/mL. The ATP synthase activity of CCD-18Co was significantly diminished in the presence of PEO-IONPs but not GM3-IONPs. PEO-IONPs also compromised the membrane integrity of CCD-18Co. In contrast, cells exposed to GM3-IONPs showed significantly different cell morphology, but with no apparent membrane damage. The interaction of PEO-IONPs or GM3-IONPs with CCD-18Co resulted in a substantial decrease in the intracellular GSH levels in a time- and concentration-dependent manner. Conversely, levels of DHR-123 increased with IONP concentrations. Levels of caspase 3/7 proteins were found to be significantly elevated in cells exposed to PEO-IONPs. Based on the results, we assume GM3-IONPs to be biocompatible with CCD-18Co and could be further evaluated for selective killing of pathogens in vivo.

Antiproliferative Efficacy of N-(3-chloro-4-fluorophenyl)-6,7-dimethoxyquinazolin-4-amine, DW-8, in Colon Cancer Cells Is Mediated by Intrinsic Apoptosis

A novel series of 4-anilinoquinazoline analogues, DW (1–10), were evaluated for anticancer efficacy in human breast cancer (BT-20) and human colorectal cancer (CRC) cell lines (HCT116, HT29, and SW620). The compound, DW-8, had the highest anticancer efficacy and selectivity in the colorectal cancer cell lines, HCT116, HT29, and SW620, with IC50 values of 8.50 ± 2.53 µM, 5.80 ± 0.92 µM, and 6.15 ± 0.37 µM, respectively, compared to the non-cancerous colon cell line, CRL1459, with an IC50 of 14.05 ± 0.37 µM. The selectivity index of DW-8 was >2-fold in colon cancer cells incubated with vehicle. We further determined the mechanisms of cell death induced by DW-8 in SW620 CRC cancer cells. DW-8 (10 and 30 µM) induced apoptosis by (1) producing cell cycle arrest at the G2 phase; (2) activating the intrinsic apoptotic pathway, as indicated by the activation of caspase-9 and the executioner caspases-3 and 7; (3) nuclear fragmentation and (4) increasing the levels of reactive oxygen species (ROS). Overall, our results suggest that DW-8 may represent a suitable lead for developing novel compounds to treat CRC.

ORAI3 contributes to hypoxia-inducible factor 1/2α-sensitive colon cell migration

BackgroundHypoxia is a pivotal initiator of tumor angiogenesis and growth through the stabilization of hypoxia-inducible factors (HIFs). This study set out to examine the involvement of HIF-1α and HIF-2α in colon cancer and ascertained whether ORAI3 was involved in the pathway.Materials and methodsPatients and murine models as well as human colorectal adenocarcinoma tumor (CW2) cells were included to examine the levels of ORAI1/3 and HIF-1/2α levels. Calcium imaging was utilized to ascertain the activity of calcium channel. Scratch assay was used to assess the migration capacity of the cells.ResultsTumors from murine colon cancer xenograft models and patients with colon cancer displayed high ORAI1/3 and HIF-1/2α levels. Hypoxia treatment, mimicking the tumor microenvironment in vitro, increased ORAI1/3 and HIF-1/2α expression as well as store-operated Ca2+ entry (SOCE). Of note is that HIF-1/2α silencing decreased SOCE, and HIF-1/2α overexpression facilitated SOCE. Furthermore, ORAI3 rather than ORAI1 expression was inhibited by HIF-1/2α silencing while increased by ML228. Luciferase assay also confirmed that ORAI3 was elevated in the presence of ML228, indicating the linkage between HIF-1/2α and ORAI3. Additionally, colony-forming potential and cell migration capacity were decreased in siHIF-1α and siHIF-2α as well as siORAI3 cells, and the facilitating effect of ML228 on cell migration and colony-forming potential was also decreased in siORAI3 CW-2 cells, which points out the importance of ORAI3 in HIF1/2α pathway.ConclusionOur findings allow to conclude that both HIF-1α and HIF-2α facilitate ORAI3 expression, hence enhancing colon cancer progression.

Targeting the hallmarks of cancer: the effects of silibinin on proliferation, cell death, angiogenesis, and migration in colorectal cancer

Background Silibinin, as a chemopreventive agent, has shown anti-cancer efficacy against different types of cancers. In the present study, we investigated the anti-cancer activities of silibinin on CT26 mouse colon cell line. Methods CT26 cells were treated with different concentrations of silibinin. To examine the cytotoxic effect of silibinin on proliferation, apoptosis, autophagy, angiogenesis, and migration, MTT, colony-forming assay, Annexin V/PI flow cytometry, RT-qPCR, and Scratch assay were used. Results Silibinin was found to significantly reduce CT26 cells survival. Furthermore, silibinin strongly induced apoptosis and autophagy by up-regulating the expression of Bax, Caspase-3, Atg5, Atg7 and BECN1 and down-regulating Bcl-2. Silibinin considerably down-regulated the expression of COX-2, HIF-1α, VEGF, Ang-2, and Ang-4 as well as the expression of MMP-2, MMP-9, CCR-2 and CXCR-4. Conclusions The present study revealed that silibinin shows anticancer activities by targeting proliferation, cell survival, angiogenesis, and migration of CT26 cells.

6,7,4′-Trihydroxyflavanone Protects against Dextran Sulfate Sodium-Induced Colitis by Regulating the Activity of T Cells and Colon Cells In Vivo

Colitis is a multifactorial disorder that mostly occurs in the gastrointestinal tract. Despite improvements in mucosal inflammation research, little is known regarding the small bioactive molecules that are beneficial for regulating T cells and colon cell activity. 6,7,4′-trihydroxyflavanone (THF) is a flavanone that possesses anti-osteoclastogenesis activity and exerts protective effects against methamphetamine-induced immunotoxicity. Whether THF mitigates intestinal inflammation by regulating T cells and colon cell activity remains unknown. In the present study, Jurkat and HT-29 cells were used for in vitro experiments, and dextran sulfate sodium (DSS)-induced colitis model in mice was used for in vivo experiment. We observed that THF did not have a negative effect on the viability of Jurkat and HT-29 cells. Quantitative PCR and Western blot analysis revealed that THF regulates the activity of Jurkat cells and HT-29 cells via the NFκB and MAPK pathways under stimulated conditions. In the DSS-induced colitis model, oral administration of THF attenuated the manifestations of DSS-induced colitis, including a reduction in body weight, shrinkage of the colon, and enhanced expression of pro-inflammatory cytokines in the colon and mesenteric lymph nodes. These data suggest that THF alleviates DSS-induced colitis by modulating the activity of T cells and colon cells in vivo.

Oxidative stress induced by tBHP in human normal colon cells by label free Raman spectroscopy and imaging. The protective role of natural antioxidants in the form of β-carotene

The present study aimed to investigate the protective effect of β-carotene on the oxidative stress injury of human normal colon cell line CCD-18Co triggered by tert-butyl hydroperoxide (tBHP).