MORINGA OLEIFERA LEAVES AQUEOUS EXTRACT INDUCE APOPTOSIS IN HT29 CELL LINE

Moringa oleifera (MO) tree has been recognized for the nutritional and medicinal values, attributing anti-inflammatory, antioxidants, and antidiabetic activities of the alcoholic and/or water extracts to its various parts. Recent research has focused on therapeutic anti-cancerous effects of the water extract of MO leaves (MOE), commonly consumed as hot beverage. In-vitro cytotoxicity on esophageal, lung, and breast cancer cells were reported by MOE. The aim in this study is to investigate the anti-cancerous potential of MOE on the colon carcinoma HT29 cells. MO water extract (MOE) was prepared at concentrations (0.1%-0.5%). MOE anti-proliferative effect on HT29-cells was assessed by measuring cellular viability using trypan blue assay. MOE cytotoxicity was further examined by monitoring the change in levels of ROS (NBT assay), ATP (Luciferase assay), and LDH release (cytotoxicity assay) and on depolarization of mitochondrial membrane potential (Mito PTTMJC-1). Protective effect of N-Acetyl cysteine and catalase were assayed. Cell cycle analysis and Annexin-V/PI staining were also performed to detect cell death. Our result showed that MOE induced a dose and time-dependent decrease in viability; increase in ROS and LDH release; dissipation of mitochondrial membrane potential with concomitant decrease in ATP level. In addition, we obtained an increase in preG0 and late apoptotic events. Pretreatment with the antioxidant N-acetyl cysteine suppressed completely LDH release and restored partially and significantly cell viability while decreasing ROS level. Catalase was not protective. MOE also induced cell death with related colon cancer cell lines: Caco2, SW837, SW948, and HCT116. The induced apoptosis in HT29 cells and related CCC identified MOE with anti-cancerous therapeutic potential

EFFECT OF SORAFENIB ON HUMAN COLON CANCER CELLS

ABSTRACT: Sorafenib, a kinase inhibitor, is among the approved drugs for the treatment of radioactive iodine resistant thyroid carcinoma, primary kidney and liver cancers. Reported targets of Sorafenib include VEGFR, Raf family, and PDGFR belonging to the general class of tyrosine kinases. Blocking growth signals in kidney and breast cancers underlie one of the mechanisms of Sorafenib antitumor effects’ leading to cell death. We hereby examine the effect of Sorafenib on human colon carcinoma cell-line HCT116. We also investigate the possible role of p53 in mediating this effect using mutant HCT116 p53-/- cells. Cultured wild and mutant cells are treated with Sorafenib (0-75µM) for 24 hr. This is followed by assessing the viability of cells using MTT and trypan blue exclusion assays. We also examined if Sorafenib mode of action is mediated by ROS. Levels of ROS were determined in the presence and absence of antioxidants using the colorimetric NBT assay. Our preliminary results show a concentration dependent decrease in viability (trypan blue) with an estimated EC50 of 10 and 25 µM for HCT116 and HCT116 p53-/- respectively. Compared to trypan blue, MTT results were similar in case of HCT116 p53-/- but were significantly different with HCT116. Furthermore we obtained a significant increase in level of ROS of: 37.11% and 31.30% for HCT116 and HCT116p53-/- respectively. However, 2 hours pre-incubation of cells with antioxidants, Trolox, N-acetylcysteine (NAC), and catalase, prior to Sorafenib treatment, exerted no different effect. No restoration of viability or decrease in generated ROS level was noted except when pre-incubated with NAC. Our preliminary findings show that Sorafenib action is independent of ROS level and p53 expression and further investigations on the mechanism(s) of Sorafenib action are ongoing

The Appearance of a Leptin Effect on Glucose Absorption in Caco2 Cells Depends on Their Differentiation Level

Backdround/Aims: The aim of this work was to study the effect and mechanism of action of leptin added apically, on glucose absorption, using Caco-2 cells as a model. Methods: Cells were grown on inserts and treated with leptin, at different time points after confluence. Radiolabelled glucose was added to the upper chamber and samples from the lower chamber were collected and assayed for radioactivity. Results: Glucose absorption increased with an increase in the level of differentiation and was associated with an increase in the protein expression level of glucose transporters. Leptin reduced glucose absorption only by day 16 after confluence, the time at which apical leptin receptors started appearing. This inhibitory effect became higher the longer the post confluence period, and was prominent on day 23. The hormone effect was found to be mediated via a decrease in the number of glucose transporters (SGLT1 and GLUT2) and a decrease in the activity of the Na+/K+ ATPase which was assayed by measuring the amount of inorganic phosphate liberated in presence and absence of enzyme activators. Conclusion: It was concluded that by day 23 post confluence, Caco-2 cells are differentiated and are appropriate to use as a model for intestinal transport studies

Mannose Inhibits the Pentose Phosphate Pathway in Colorectal Cancer and Enhances Sensitivity to 5-Fluorouracil Therapy

Colorectal cancer (CRC) is one of the leading cancers and causes of death in patients. 5-fluorouracil (5-FU) is the therapy of choice for CRC, but it exhibits high toxicity and drug resistance. Tumorigenesis is characterized by a deregulated metabolism, which promotes cancer cell growth and survival. The pentose phosphate pathway (PPP) is required for the synthesis of ribonucleotides and the regulation of reactive oxygen species and is upregulated in CRC. Mannose was recently reported to halt tumor growth and impair the PPP. Mannose inhibitory effects on tumor growth are inversely related to the levels of phosphomannose isomerase (PMI). An in silico analysis showed low PMI levels in human CRC tissues. We, therefore, investigated the effect of mannose alone or in combination with 5-FU in human CRC cell lines with different p53 and 5-FU resistance statuses. Mannose resulted in a dose-dependent inhibition of cell growth and synergized with 5-FU treatment in all tested cancer cell lines. Mannose alone or in combination with 5-FU reduced the total dehydrogenase activity of key PPP enzymes, enhanced oxidative stress, and induced DNA damage in CRC cells. Importantly, single mannose or combination treatments with 5-FU were well tolerated and reduced tumor volumes in a mouse xenograft model. In summary, mannose alone or in combination with 5-FU may represent a novel therapeutic strategy in CRC

Fragrance chemicals lyral and lilial decrease viability of HaCat cells’ by increasing free radical production and lowering intracellular ATP level : protection by antioxidants

We investigate in this study the biochemical effects on cells in culture of 2 commonly used fragrance chemicals: lyral and lilial. Whereas both chemicals exerted a significant effect on primary keratinocyte (s), HaCat cells, no effect was obtained with any of HepG2, Hek293, Caco2, NIH3T3, and MCF7 cells. Lyral and lilial: a) decreased the viability of HaCat cells with a 50% cell death at 100nM and 60nM respectively; b) decreased significantly in a dose dependant manner the intracellular ATP level following 12-hrs of treatment; c) inhibited complexes I and II of electron transport chain in liver sub-mitochondrial particles; d) increased reactive oxygen species generation that was reversed by N-acetyl cysteine and trolox and the natural antioxidant lipoic acid, without influencing the level of free and/or oxidized glutathione. Lipoic acid protected HaCat cells against the decrease in viability induced by either compound. Dehydrogenation of lyral and lilial produce α-β-unsaturated aldehydes, that reacts with lipoic acid requiring proteins resulting in their inhibition. We propose lyral and lilial as toxic to mitochondria that have a direct effect on electron transport chain, increase ROS production, derange mitochondrial membrane potential, and decrease cellular ATP level, leading thus to cell death. ⺠Fragrant chemicals Lyral /lilial decreased viability of HaCat cells. ⺠Lyral / lilial- decreased ATP level and increased: ROS level and LDH release in HaCat cells. ⺠Lyral /lilial inhibited liver sub-mitochondrial particles complexes I and II activities of respiratory chain. ⺠Antioxidants restored control ROS level; lipoic acid protected the viability of lilial and lyral treated Hacat cells. ⺠Lyral and lilila are mitochondriotoxin with direct and indirect effect on HaCat cells

Clinical, biochemical and genetic profile of WD patients in the S family.

a<p>Screening refers to genetic screening of the ATP7B gene exons' by PCR followed by sequencing; <b>^b</b>Few years later developed liver cirrhosis and portal hypertension;</p><p><b>Normal ranges of</b>: Serum ceruloplasmin: 0.15–0.60 g/; Serum Cu: 70–150 µg/dL; 24h urine Cu: 15–50 µg/24h; Bilirubin T/D: 0.2–1.2/0–0.5; ^blevel determined after 7 years of treatment; ^cUrinary Cu level while S41was on penicillamine; * c.2298_2299insC is referred to as 2299insC. DNA Mutation numbering is based on cDNA numbering where nucleotide +1 as the A of the ATG translation initiation codon, in the reference sequence # NM_000053 with the initiation codon aa codon +1.</p><p>Clinical, biochemical and genetic profile of WD patients in the S family.</p