Human Colorectal Adenocarcinoma HT-29 Cells Research Articles

Microbial synthesis of zinc oxide nanoparticles and their potential biological application as an antimicrobial and anticancer agent

Zinc oxide nanoparticles (ZnO-NPs) are a type of metal oxide nanomaterial, recognized as a valuable and adaptable inorganic compound owing to its distinctive physical and chemical properties. Nanosized ZnO particles exhibit substantial antibacterial properties attributable to their diminutive size, which can activate various bactericidal mechanisms within the bacterial cell, including interactions with the bacterial surface or core, the generation of reactive oxygen species (ROS), the release of Zn2+, and potential endocytosis by cells. ZnO NPs nanoparticles were extracellularly produced using pigment extracts from the PP6 strain. Agar well screening indicated that PP6 secondary metabolites possess antibacterial properties. UV–Vis spectroscopy was used to analyze the external growth of nanoparticles. Scanning Electron Microcopy (SEM), Fourier Transform Infrared Spectroscopy (FTIR), and X-ray diffraction (XRD) techniques were employed to analyze the shape, stability, crystalline structure, and coating of strain PP6 ZnO NPs. The PP6 ZnO NPs demonstrated an antibiofilm impact on the bacterial pathogens tested, which was dependent on the dosage. Elevated levels of lipid peroxidation (LPO) and reduced antioxidant activity are indicative of apoptosis in cancer cells. The synthesized ZnO NPs nanoparticles were assessed for their anticancer properties by performing the MTT assay on HT-29 human colorectal adenocarcinoma cells. ZnO NPs nanoparticles exposed to HT-29 cells the viability was reduced significantly in proportion to the concentration of nanoparticles. Additional comprehensive study will be needed to fully understand their mechanism.

Probiotic and Postbiotic Potentials of Enterococcus faecalis EF-2001: A Safety Assessment.

Probiotics, which are live microorganisms that, when given in sufficient quantities, promote the host's health, have drawn a lot of interest for their ability to enhance gut health. Enterococcus faecalis, a member of the human gut microbiota, has shown promise as a probiotic candidate due to its functional attributes. However, safety concerns associated with certain strains warrant comprehensive evaluation before therapeutic application. In this study, E. faecalis EF-2001, originally isolated from fecal samples of a healthy human infant, was subjected to a multi-faceted assessment for its safety and probiotic potential. In silico analysis, CAZyme, biosynthetic, and stress-responsive proteins were identified. The genome lacked biogenic amine genes but contained some essential amino acid and vitamin synthetic genes, and carbohydrate-related enzymes essential for probiotic properties. The negligible difference of 0.03% between the 1st and 25th generations indicates that the genetic information of the E. faecalis EF-2001 genome remained stable. The live E. faecalis EF-2001 (E. faecalis EF-2001L) demonstrated low or no virulence potential, minimal D-Lactate production, and susceptibility to most antibiotics except some aminoglycosides. No bile salt deconjugation or biogenic amine production was observed in an in vitro assay. Hemolytic activity assessment showed a β-hemolytic pattern, indicating no red blood cell lysis. Furthermore, the EF-2001L did not produce gelatinase and tolerated simulated gastric and intestinal fluids in an in vitro study. Similarly, heat-killed E. faecalis EF-2001 (E. faecalis EF-2001HK) exhibits tolerance in both acid and base conditions in vitro. Further, no cytotoxicity of postbiotic EF-2001HK was observed in human colorectal adenocarcinoma HT-29 cells. These potential properties suggest that probiotic and postbiotic E. faecalis EF-2001 could be considered safe and retain metabolic activity suitable for human consumption.

Phytochemical Profile of Cymbopogon citratus (DC.) Stapf Lemongrass Essential Oil from Northeastern Thailand and Its Antioxidant and Antimicrobial Attributes and Cytotoxic Effects on HT-29 Human Colorectal Adenocarcinoma Cells.

Colorectal cancer is the third most prevalent cancer in Thailand, prompting the search for alternative or preventive treatments using natural constituents. In this study, the authors employed hydrodistillation to extract Cymbopogon citratus (DC.) Stapf (lemongrass) essential oil (LEO) from plants in northeastern Thailand and assessed its chemical profile, antioxidant, antimicrobial, and anticancer properties. The LEO displayed potent antioxidant activities in DPPH and hydroxyl scavenging assays with IC50 values of 2.58 ± 0.08 and 4.05 ± 0.12 mg/mL, respectively, and demonstrated antimicrobial activities against Escherichia coli, Cutibacterium acnes, Streptococcus agalactiae, and Staphylococcus aureus at 8-10 µg/mL. At 48 h, the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay showed the LEO exhibiting low cell viability (3%) at concentrations of 200-400 µg/mL, with an IC50 value of 82.46 ± 1.73 µg/mL, while in the clonogenic assay it exhibited a lower IC50 value of 23.11 ± 1.80 µg/mL. The GC-MS analysis identified citral (79.24%) consisting of 44.52% geranial and 34.72% neral, and β-myrcene (5.56%). The addition of LEO significantly influenced apoptotic genes (Bcl-2, Bax, p21, and Caspase-3) and proteins, as indicated by real-time polymerase chain reaction (RT-PCR) and Western blot studies. Results suggested that LEO initiated apoptosis through intrinsic pathways and demonstrated potential as a chemopreventive, antimicrobial, and antioxidant agent with substantial health advantages.

Sialic acid detection and theranostic activity of phenylboronic acid-based fluorescent probe in human colorectal adenocarcinoma HT-29 cells

A new probe, 4-(((3′,6′-bis(diethylamino)-3-oxospiro[isoindoline-1,9′-xanthen]-2-yl)imino)methyl)phenyl)boronic acid (R4B) was prepared by facile condensation of 4-formylphenylboronic acid and rhodamine B hydrazide. R4B was characterized by spectroscopic methods and single crystal X-ray diffraction. The sensor R4B solution turned pink and emitted orange fluorescence only in the presence of sialic acid but remained colorless and non-fluorescent otherwise. The sugar recognition performance was investigated via UV–vis and fluorescence spectroscopic studies. Our results revealed that R4B has good affinity and selectivity for sialic acid over common monosaccharides, with a detection limit as low as 10−7 M. Furthermore, R4B selectively inhibited growth of human colorectal adenocarcinoma HT-29 (IC50 <20 µM) without significant cytotoxicity to normal human colon fibroblasts CCD-18Co. Treatment with R4B suppressed HT-29 colony formation via mitochondrial apoptosis in a time-dependent manner. Cellular imaging studies also revealed the ability of R4B as a fluorescence dye to detect intracellular sialic acid and showed mitochondria-tracking ability in HT-29 cells. In summary, R4B is a potential theranostic for the detection of intracellular sialic acid during the early incubation period, followed by induction of cancer apoptotic cell death at a later treatment point.

Dentatacid A: An Unprecedented 2, 3-Seco-arbor-2, 3-dioic Triterpenoid from the Invasive Plant Euphorbia dentata, with Cytotoxicity Effect on Colon Cancer.

Euphorbia dentata Michx. is an invasive plant species in China, known for its toxicity and potential to reduce crop yields, posing numerous threats. To gain a deeper understanding of this invasive plant, phytochemical methods were employed to isolate 13 terpenoids (1-11, 19, 20) and 7 sterols (12-18) from the ethanol extract of E. dentata, identifying one new compound and 19 known compounds. Within spectroscopic methods such as NMR, HR-ESI-MS, and ECD, the structures and absolute configurations of these compounds were established. Among them, dentatacid A (11) possesses an unprecedented 2, 3-seco-arbor-2, 3-dioic skeleton within the potential biosynthetic pathway proposed. Dentatacid A also exhibited excellent anti-proliferative activity against the HT-29 (human colorectal adenocarcinoma) cell line, with an IC50 value of 2.64 ± 0.78 μM, which was further confirmed through network pharmacology and molecular docking. This study significantly expands the chemical diversity of E. dentata and offers new insights into the resource utilization and management of this invasive plant from the perspective of natural product discovery.

Inducing Immunogenic Cancer Cell Death through Oxygen‐Economized Photodynamic Therapy with Nitric Oxide‐Releasing Photosensitizers

AbstractPhotodynamic therapy (PDT) utilizes reactive oxygen species (ROS) for eradication of cancer cells. Its effectiveness is governed by the oxygen content, which is scarce in the hypoxic tumor microenvironment. We report herein two zinc(II) phthalocyanines substituted with two or four nitric oxide (NO)‐releasing moieties, namely ZnPc‐2NO and ZnPc‐4NO, which can suppress the mitochondrial respiration, thereby sparing more intracellular oxygen for PDT. Using HT29 human colorectal adenocarcinoma cells and A549 human lung carcinoma cells, we have demonstrated that both conjugates release NO upon interaction with the intracellular glutathione, which can reduce the cellular oxygen consumption rate and adenosine triphosphate generation and alter the mitochondrial membrane potential. They can also relieve the hypoxic status of cancer cells and decrease the expression of hypoxia‐inducible factor protein HIF‐1α. Upon light irradiation, both conjugates can generate ROS and induce cytotoxicity even under a hypoxic condition, overcoming the oxygen‐dependent nature of PDT. Interestingly, the photodynamic action of ZnPc‐2NO elicits the release of damage‐associated molecular patterns, inducing the maturation of dendritic cells and triggering an antitumor immune response. The immunogenic cell death caused by this oxygen‐economized PDT has been demonstrated through a series of in vitro and in vivo experiments.

Inducing Immunogenic Cancer Cell Death through Oxygen-Economized Photodynamic Therapy with Nitric Oxide-Releasing Photosensitizers.

Photodynamic therapy (PDT) utilizes reactive oxygen species (ROS) for eradication of cancer cells. Its effectiveness is governed by the oxygen content, which is scarce in the hypoxic tumor microenvironment. We report herein two zinc(II) phthalocyanines substituted with two or four nitric oxide (NO)-releasing moieties, namely ZnPc-2NO and ZnPc-4NO, which can suppress the mitochondrial respiration, thereby sparing more intracellular oxygen for PDT. Using HT29 human colorectal adenocarcinoma cells and A549 human lung carcinoma cells, we have demonstrated that both conjugates release NO upon interaction with the intracellular glutathione, which can reduce the cellular oxygen consumption rate and adenosine triphosphate generation and alter the mitochondrial membrane potential. They can also relieve the hypoxic status of cancer cells and decrease the expression of hypoxia-inducible factor protein HIF-1α. Upon light irradiation, both conjugates can generate ROS and induce cytotoxicity even under a hypoxic condition, overcoming the oxygen-dependent nature of PDT. Interestingly, the photodynamic action of ZnPc-2NO elicits the release of damage-associated molecular patterns, inducing the maturation of dendritic cells and triggering an antitumor immune response. The immunogenic cell death caused by this oxygen-economized PDT has been demonstrated through a series of in vitro and in vivo experiments.

Novel tetraaza macrocyclic Schiff base complexes of bivalent zinc: microwave-assisted green synthesis, spectroscopic characterization, density functional theory calculations, molecular docking studies, in vitro antimicrobial and anticancer activities.

In the present manuscript, novel macrocyclic Schiff base complexes [Zn(N4MacL1)Cl2-Zn(N4MacL3)Cl2] were synthesized by the reaction of ZnCl2 and macrocyclic ligands (N4MacL1-N4MacL3) derived from diketone and diamines under microwave irradiation method and conventional method. The structures of the obtained complexes were identified by various spectrometric methods such as Fourier transformation infra-red (FT-IR), nuclear magnetic resonance (NMR), high-resolution mass spectrometry (HR-MS), powder X-ray diffraction, molar conductivity, and UV-vis. The structures of the synthesized compounds were optimized by using the def2-TZV/J and def2-SVP/J Coulomb fitting basis sets at B3LYP level in density functional theory (DFT) calculations. The macrocyclic Schiff base complexes exhibited higher activities against Gram-positive bacteria (Staphylococcus aureus and Bacillus cereus), Gram-negative bacteria (Escherichia coli and Xanthomonas campestris), and fungal strains (Fusarium oxysporum and Candida albicans) in comparison to macrocyclic Schiff base ligands. Furthermore, the newly synthesized macrocyclic compounds were assessed for their anticancer activity against three cell lines: A549 (human alveolar adenocarcinoma epithelial cell line), HT-29 (human colorectal adenocarcinoma cell line), and MCF-7 (human breast adenocarcinoma cell line) using the MTT assay. The obtained results showed that the macrocyclic complex [Zn(N4MacL3)Cl2] displayed the highest cytotoxic activity (2.23 ± 0.25µM, 6.53 ± 0.28µM, and 7.40 ± 0.45µM for A549, HT-29, and MCF-7 cancer cell lines, respectively). Additionally, molecular docking investigations were conducted to elucidate potential molecular interactions between the synthesized macrocyclic compounds and target proteins. The results revealed a consistent agreement between the docking calculations and the experimental data.

Aminocarbyne-Alkyne Coupling in Diruthenium Complexes: Exploring the Anticancer Potential of the Resulting Vinyliminium Complexes and Comparison with Diiron Homologues.

New diruthenium complexes based on the scaffold Ru2Cp2(CO)2 (Cp = η5-C5H5) and containing a bridging vinyliminium ligand, [2a-d]CF3SO3, were synthesized through regioselective coupling of alkynes with an aminocarbyne precursor (85-90% yields). The reaction involving phenylacetylene proceeded with the formation of a diruthenacyclobutene byproduct, [4]CF3SO3 (10% yield). Complexes [2a-d]+ undergo partial alkyne extrusion in contact with alumina or CDCl3. All products were characterized by elemental analysis, infrared and multinuclear NMR spectroscopy, and single crystal X-ray diffraction in two cases. Complexes [2a-d]+ revealed an outstanding stability in DMEM cell culture medium at 37 °C (<1% degradation over 72 h). These complexes exhibited cytotoxicity in human colon colorectal adenocarcinoma HT-29 cells in the low micromolar range, with lower IC50 values than those obtained with the homologous diiron complexes previously reported. Evaluation of ROS (reactive oxygen species) production and O2 consumption rate (OCR) highlighted the higher potential of Ru2 complexes, compared to the Fe2 counterparts, to impact mitochondrial activity, with the heterometallic Ru2-ferrocenyl complex [2d]+ showing the best performance.

Synthesis of selenophene-containing flavonols and 2-styrylchromones: Evaluation of their activities compared with selenophene-containing chalcones as potential anticancer agents.

Previously, we documented the synthesis and assessed the biological effects of chalcones containing selenium against HT-29 human colorectal adenocarcinoma cells, demonstrating their significant potential. As research on selenium-containing flavonoids remains limited, this article outlines our design and synthesis of three selenium-based flavonols and three 2-styrylchromones. We conducted evaluations of these compounds to determine their impact on human lung cancer cells (A549, H1975, CL1-0, and CL1-5) and their influence on normal lung fibroblast MRC5 cells. Additionally, we included selenium-based chalcones in our testing for comparative purposes. Our findings highlight that the simplest compound, designated as compound 1, exhibited the most promising performance among the tested molecules.

Miniature wireless LED-device for photodynamic-induced cell pyroptosis

The inability of visible light to penetrate far through biological tissue limits its use for phototherapy and photodiagnosis of deep-tissue sites of disease. This is unfortunate because many visible dyes are excellent photosensitizers and photocatalysts that can induce a wide range of photochemical processes, including photogeneration of reactive oxygen species. One potential solution is to bring the light source closer to the site of disease by using a miniature implantable LED. With this goal in mind, we fabricated a wireless LED-based device (volume of 23 mm3) that is powered by RF energy and emits light with a wavelength of 573 nm. It has the capacity to excite the green absorbing dye Rose Bengal, which is an efficient type II photosensitizer. The wireless transfer of RF power is effective even when the device is buried in chicken breast and located 6 cm from the transmitting antenna. The combination of a wireless device as light source and Rose Bengal as photosensitizer was found to induce cell death of cultured HT-29 human colorectal adenocarcinoma cells. Time-dependent generation of protruding bubbles was observed in the photoactivated cells suggesting cell death by light-induced pyroptosis and supporting evidence was gained by cell staining with the fluorescence probes Annexin-V FITC and Propidium Iodide. The results reveal a future path towards a wireless implanted LED-based device that can trigger photodynamic immunogenic cell death in deep-seated cancerous tissue.

Synthesis of reduced graphene oxide using methanolic extract of Sargassum tenerrimum and its antiproliferative activity against human colorectal adenocarcinoma HT-29 cell lines

Synthesis of reduced graphene oxide using methanolic extract of Sargassum tenerrimum and its antiproliferative activity against human colorectal adenocarcinoma HT-29 cell lines

Rheum khorasanicum. Hydroalcoholic root extract induces cell death in human colorectal adenocarcinoma: An in vitro and in silico study

Colorectal cancer (CRC) is the second greatest cause of cancer-related death in the world and chemotherapy, as an important part of CRC treatment, has some drawbacks, including systemic toxicity. Therefore, it is crucial to discover new and more effective CRC treatment plans. Rheum khorasanicum (R.khorasanicum) is a medicinal plant with high flavonoids, stilbenes, and anthraquinone contents, so it can be a potential source of antioxidants and can be used for therapeutic purposes and trigger apoptosis in cancer cells. In this study, we investigated the effects of hydroalcoholic root extract of R.khorasanicum treatment on inducing mitochondrial apoptosis of HT-29 and Caco-2 human colorectal adenocarcinoma cells. Firstly, the total phenolic and flavonoid content was determined. Then, the cytotoxic effects of R.khorasanicum on cells of three different types, including HT-29 and Caco-2 colon cancer cells as well as normal 3T3 cells were assessed using the MTT assay. To investigate the characteristics of cellular death, flow cytometry, and western blotting were performed. The results of this study indicated considerable phenolic (356.4±9.4 GAE/gDW) and flavonoid (934.55±17.1 QE/gDW) contents in R.khorasanicum. MTT assay's finding indicated that 100, 60, and 30μg/mL concentrations of R.khorasanicum reduce cell viability in HT-29 and Caco-2 cell lines significantly (P<0.05). It has been also revealed that R.khorasanicum extract induces apoptosis rather than necrosis in these cell lines. Moreover, Bcl-2 expression was significantly reduced in both HT-29 and Caco-2 cell lines, while Bax and cleaved caspase-3 expression soared considerably in the groups under R.khorasanicum treatment (P<0.05). In conclusion, our findings have suggested that high phenol and flavonoid contents of R.khorasanicum root extract possibly play an important role in cell cytotoxicity and apoptosis induction in HT-29 and Caco-2 colon cancer cells.

Phytochemical profile by LC-MS/MS analysis and evaluation of antioxidant, antidiabetic, anti-Alzheimer, and anticancer activity of Onobrychis argyrea leaf extracts

IntroductionStudies on different species of Onobrychis, a member of the Fabaceae family, have revealed a number of biological activities with potential applications in alternative medicine. The phytochemical content of the plant Onobrychis argyrea subsp. argyrea and some of the important biological activities linked to its metabolism are quite limited. Therefore, this study was the first to determine the content of secondary metabolites in this species and to elucidate important biological activities related to metabolism. MethodsLC-MS/MS was used for the quantitative analysis of bioactive compounds in the aerial part of O. argyrea extracts. The antioxidant properties of the extracts were assessed based on their ability to scavenge DPPH radicals and reduce iron ions. The anti-diabetic effect of the extracts was tested by the inhibition of α-amylase and α-glycosidase enzymes, and the anti-Alzheimer's ability was tested by the inhibition of the enzymes AChE and BChE. The cytotoxic effects of the extracts were tested on four different cancer lines, namely A-549, HT-29, MCF-7 and MDA-MB 453 using the XTT assay. The mechanism of the anti-cancer effect was determined by means of flow cytometry analysis on HT-29 cells. ResultsTwenty-nine phytochemicals were identified via the LC-MS/MS analysis in the plant extracts. The most abundant phytochemicals in the extracts were quinic acid, isoquercitrin, epicatechin, and routine, respectively. Antioxidant analysis showed that among all extracts, methanol extract (ME) was most effective in scavenging DPPH (IC50: 23.93 ± 0.96 µg/mL) and in reducing iron ions (53.24 ± 2.14 mgTEAC/g). Enzyme activity studies demonstrated that ME is a potent inhibitor of α-glycosidase (IC50: 15.06±0.64 µg/mL), AChE (IC50: 24.75±0.76 µg/mL), and BChE (IC50: 3.36±0.14 µg/mL). XTT assay results disclosed the strongest anti-proliferative activity of ME against the human colorectal adenocarcinoma HT-29 cell line with an IC50 value of 33.35±0.92 µg/mL. Flow cytometry analysis revealed that ME caused mitochondrial membrane damage in HT-29 cells (**p<0.01) and subsequently induced apoptosis (**p<0.01) by activating the caspase pathway (**p<0.001). ConclusionO. argyrea, rich in bioactive compounds, showed remarkable antioxidant, antidiabetic, Alzheimer's, and anticancer properties in vitro, demonstrating that this plant contains secondary metabolites that may be potential drug candidates for the treatment of metabolic diseases.

A deep learning approach for the detection and counting of colon cancer cells (HT-29 cells) bunches and impurities.

HT-29 has an epithelial appearance as a human colorectal cancer cell line. Early detection of colorectal cancer can enhance survival rates. This study aims to detect and count HT-29 cells using a deep-learning approach (ResNet-50). The cell lines were procured from Procell Life Science & Technology Co., Ltd. (Wuhan, China). Further, the dataset is self-prepared in lab experiments, cell culture, and collected 566 images. These images contain two classes; the HT-29 human colorectal adenocarcinoma cells (blue shapes in bunches) and impurities (tinny circular grey shapes). These images are annotated with the help of an image labeller as impurity and cancer cells. Then afterwards, the images are trained, validated, and tested against the deep learning approach ResNet50. Finally, in each image, the number of impurity and cancer cells are counted to find the accuracy of the proposed model. Accuracy and computational expense are used to gauge the network's performance. Each model is tested ten times with a non-overlapping train and random test splits. The effect of data pre-processing is also examined and shown in several tasks. The results show an accuracy of 95.5% during training and 95.3% in validation for detecting and counting HT-29 cells. HT-29 cell detection and counting using deep learning is novel due to the scarcity of research in this area, the application of deep learning, and potential performance improvements over traditional methods. By addressing a gap in the literature, employing a unique dataset, and using custom model architecture, this approach contributes to advancing colon cancer understanding and diagnosis techniques.

Enterococcus Induces MHC-II Expression in the Colonic Epithelium to Aggravate Graft-Versus-Host Disease

Intestinal domination by Enterococcus, a genus of Gram-positive facultative bacteria, has been associated with an increased risk of GVHD mortality in allo-HCT patients and in preclinical models. However, the immunological mechanisms underlying this association remain unclear. Several preclinical studies have demonstrated that MHC-II expression by intestinal epithelial cells (IECs) is involved in allo-antigen presentation and GVHD and can be modulated by the intestinal microbiota. MHC-II genes are constitutively expressed by professional antigen-presenting cells (APCs) but can be induced in non-professional APCs by IFN-γ. Here we reported that Enterococcus could potentially aggravate GVHD through upregulation of MHC-II expression by IECs in an IFN-γ-dependent manner. Using an MHC-disparate mouse model of GVHD (C57BL/6J into BALB/cJ), we profiled the fecal microbiome of GVHD mice by 16S rRNA sequencing and the intestinal epithelium by flow cytometry on day 7 post-transplant. We found that endogenous Enterococcus relative abundance was positively correlated with MHC-II expression by colonic IECs (CD45- CD31- EpCAM+) during GVHD ( Fig. 1a; p=0.0004; 2 replicates; n=19). To determine whether this phenotype occurs in the absence of systemic inflammation, we utilized gnotobiotic mice treated with either E. faecalis, B. producta (a bacteria genus previously associated with improved GVHD outcomes)or PBS via oral gavage. Monocolonization with E. faecalis was sufficient to upregulate MHC-II expression by colonic IECs in non-transplanted (steady-state) animals compared to PBS-treated germfree controls ( Fig. 1b; p=0.0027; 3 replicates; n=8-12/group). On the contrary, monocolonization by B. producta did not induce MHC-II expression by colonic IECs compared to germfree controls ( p=0.96). We also observed that MHC-II upregulation was promoted by specific species within the genus Enterococcus, such that compared to PBS-treated germfree controls, only E. faecalis ( p=0.0007; 3 replicates; n=8-12/group) was able to induce MHC-II expression by colonic IECs, but not E. durans ( p=0.06) , E. gallinarium ( p=0.98) and E. hirae ( p=0.99). Using a Villin-Cre IFN- γ R fl/flmouse model with genetic ablation of the IFN-γ receptor specifically in epithelial cells, we investigated whether MHC-II expression induced by Enterococcus was dependent on IFN-γ signaling. We colonized Villin-Cre pos and Villin-Cre neg littermate controls with E. faecalis via oral gavage after antibiotics decontamination (ampicillin/enrofloxacin) and profiled the intestinal epithelium. We observed that Villin-Cre posmice exhibited a significantly lower MHC-II expression in the small intestine compared to Villin-Cre negmice on day 3 post- Enterococcus colonization ( p=0.012; 2 replicates; n=7/group), suggesting that IFN-γ signaling is necessary to mediate Enterococcus interaction with the intestinal epithelium. Finally, to explore a potential intervention to reduce intestinal inflammation upon Enterococcus domination, we treated GVHD mice with either butyrate-producing B. producta or E. faecalis. Colonization with B. producta reduced fecal Enterococcus burden on day 7 post-transplant ( p=0.007; 2 replicates; n=8/group) and improved survival in GVHD mice, compared to the E. faecalis-colonized group ( p=0.002; 2 replicates; n=18-19/group). We and others have previously shown that butyrate administration can alleviate GVHD through its effects on epithelial homeostasis. To determine whether butyrate could regulate allo-antigen presentation by the intestinal epithelium, we employed the human colorectal adenocarcinoma HT-29 cell culture. We pre-treated the cells with IFNγ (100U/ml) to induce MHC-II expression, then added butyrate (5mM) to the culture for 24 hours. Butyrate treatment was sufficient to suppress MHC-II expression by HT-29 cells, compared to DMSO-treated controls ( p=0.006; 3 replicates; n=6/group). Altogether, our results suggest that one mechanism by which Enterococcus can aggravate GVHD involves the upregulation of MHC-II expression by colonic IECs, and treatment with butyrate might serve as an intervention to prevent intestinal inflammation. Further identification of the upstream mediators involved will provide additional mechanistic insight into the interaction between Enterococcus and the intestinal epithelium at steady state and during GVHD.

Genoprotective potential of Macaranga species phytochemical compounds on HT-29 human colorectal adenocarcinoma cell line

BackgroundThe species of genus Macaranga are widely found in Malaysian secondary forests and has been used as an alternative for treating varieties of illness. Studies have shown that the medicinal properties of this genus contain anti-inflammatory, antioxidant, and anti-cancer effects. This study aimed to determine the cytotoxicity of six isolated phytochemicals from Macaranga heynei (M. heynei), Macaranga lowii and Shorea leprosula on HT-29 human colorectal adenocarcinoma cell lines.ResultsOne out of six isolated phytochemical compounds, identified as “Laevifolin A”, showed a cytotoxicity with an IC50 value of 21.2 µM following 48 h treatment as detected using Sulforhodamine B (SRB) assay. Additionally, no induction of apoptosis and oxidative stress were observed on Laevifolin A treated HT-29 cells as determined using Annexin V-FITC/PI assay and dihydroethidine (HE) staining, respectively. Additionally, no damage to the DNA were observed as measured using the Alkaline Comet assay. Further investigation on menadione-induced oxidative DNA damage showed the genoprotective potential of Laevifolin A on HT-29 cells.ConclusionsIn conclusion, this study indicated that only one compound (Laevifolin A) that extracted from M. heynei has the cytotoxicity potential to be developed as an anticancer agent in human colorectal adenocarcinoma. However, besides exhibiting cytotoxic effect, the compound also exhibits genoprotective capability that warrant further investigation.

Increased Production of Interleukin-10 and Decreased Production of Tumor Necrosis Factor-α by Lacticaseibacillus rhamnosus PL60

Lacticaseibacillus rhamnosus PL60, originally isolated from healthy baby feces, has beneficial activities including resistance to acid and bile acids, immune-enhancing activity, inhibition of various intestinal pathogens, anti-oxidation activity, adherence to the intestines, susceptibility to antimicrobials, an anti-obesity effect, and an anti-fatty liver effect. In the present study, the binding activity of L. rhamnosus PL60 to the HT-29 human colorectal adenocarcinoma cell line and mouse intestines was established. L. rhamnosus PL60 increased interleukin-10 production and decreased tumor necrosis factor-α production in a dose-dependent manner in HT-29 cells treated with lipopolysaccharide. Similar results were observed in an in vivo experiment with BALB/c mice fed with L. rhamnosus PL60. Taken together, results from previous studies and the present study establish that L. rhamnosus PL60 is a probiotic candidate for inflammation-related diseases.

Cupric-ion-promoted fabrication of oxygen-replenishing nanotherapeutics for synergistic chemo and photodynamic therapy against tumor hypoxia.

Mixing a glutathione (GSH)-responsive carboxy zinc(II) phthalocyanine (ZnPc*) and CuSO4·5H2O in water with or without the presence of the anticancer drug SN38 resulted in the formation of self-assembled nanotherapeutics labeled as ZnPc*/Cu/SN38@NP and ZnPc*/Cu@NP, respectively. The Cu2+ ions not only promoted the self-assembly of the carboxy phthalocyanine through metal complexation, but also catalyzed the transformation of H2O2 to oxygen via a catalase-like reaction, rendering an oxygen-replenishing property to the nanosystems. Both nanosystems exhibited high stability in aqueous media, but the nanoparticles disassembled gradually in an acidic or GSH-enriched environment and inside human colorectal adenocarcinoma HT29 cells, releasing the encapsulated therapeutic components. The disassembly process together with the activation by the intracellular GSH led to relaxation of the intrinsic quenching of the nanophotosensitizers and restoration of the photoactivities of ZnPc*. Under a hypoxic condition, ZnPc*/Cu/SN38@NP could attenuate the intracellular hypoxia level and maintain the photodynamic activity due to its Cu2+-promoted oxygen-replenishing ability. The photodynamic effect of ZnPc* and the anticancer effect of SN38 worked cooperatively, causing substantial apoptotic cell death. The dual therapeutic actions could also effectively inhibit the tumor growth in HT29 tumor-bearing nude mice without initiating notable adverse effects to the mice. STATEMENT OF SIGNIFICANCE: The oxygen-dependent nature of photodynamic therapy generally reduces its efficacy against tumor hypoxia, which is a common characteristic of advanced solid tumors and usually leads to resistance toward various anticancer therapies. We report herein a facile approach to assemble a glutathione-responsive carboxy phthalocyanine-based photosensitizer and an anticancer drug in aqueous media, in which Cu(II) ions were used to promote the self-assembly through metal complexation and catalyze the conversion of H2O2 to oxygen through a catalase-like reaction, making the resulting nanoparticles possessing an oxygen-replenishing property that could promote the photodynamic effect against hypoxic cancer cells and tumors. The use of Cu(II) ions to achieve the aforementioned dual functions in the fabrication of advanced nano-photosensitizing systems has not been reported.

Design, Synthesis, Characterization, and Evaluation of the Anti-HT-29 Colorectal Cell Line Activity of Novel 8-Oxyquinolinate-Platinum(II)-Loaded Nanostructured Lipid Carriers Targeted with Riboflavin.

Colorectal cancer is occasionally called colon or rectal cancer, depending on where cancer begins to form, and is the second leading cause of cancer death among both men and women. The platinum-based [PtCl(8-O-quinolinate)(dmso)] (8-QO-Pt) compound has demonstrated encouraging anticancer activity. Three different systems of 8-QO-Pt-encapsulated nanostructured lipid carriers (NLCs) with riboflavin (RFV) were investigated. NLCs of myristyl myristate were synthesized by ultrasonication in the presence of RFV. RFV-decorated nanoparticles displayed a spherical shape and a narrow size dispersion in the range of 144-175 nm mean particle diameter. The 8-QO-Pt-loaded formulations of NLC/RFV with more than 70% encapsulation efficiency showed sustained in vitro release for 24 h. Cytotoxicity, cell uptake, and apoptosis were evaluated in the HT-29 human colorectal adenocarcinoma cell line. The results revealed that 8-QO-Pt-loaded formulations of NLC/RFV showed higher cytotoxicity than the free 8-QO-Pt compound at 5.0 µM. All three systems exhibited different levels of cellular internalization. Moreover, the hemotoxicity assay showed the safety profile of the formulations (less than 3.7%). Taken together, RFV-targeted NLC systems for drug delivery have been investigated for the first time in our study and the results are promising for the future of chemotherapy in colon cancer treatment.