HT-29 Cells Research Articles

Fermentative Characteristics and Metabolic Profiles of Japanese Apricot Juice Fermented with Lactobacillus acidophilus and Torulaspora delbrueckii.

Functional fermented fruit juices produced using a combination of non-Saccharomyces yeast and lactic acid bacteria (LAB) are relatively unexplored. The effects of three inoculation protocols, single inoculation with Lactobacillus acidophilus (La), single inoculation with Torulaspora delbrueckii (Td), and co-culture of both La + Td, on the physicochemical, microbiological, sensory properties, and metabolic profile of fermented JA juices after 24 h at 30 °C were investigated. Uninoculated (UI) Japanese apricot (JA) juice was used as a control. The results show significant increases in the color intensity of the fermented-JA juices, whereas an enhancement of total phenolic contents is observed in the fermented JA-juices acquired through the use of La for both single and co-culture inoculations. The colony counts of LAB and yeast in the inoculated JA juices increased by approximately 2.0 and 1.7 log CFU/mL at 24 h, respectively. The antibacterial activity of JA juices against four pathogenic bacteria was detected. All JA juices exhibited antimicrobial activity against the tested pathogenic strains, with strong antibacterial properties of La-fermented juice being recorded against Bacillus cereus at the lowest MIC of 124 µL/mL. Additionally, La + Td-fermented and UI-JA juices demonstrated comparable anticancer activity against HT-29 cells with IC50 values of 823.37 and 754.87 µg/mL, respectively. Furthermore, a total of 995 compounds was identified as differential fermentation metabolites through non-targeted metabolome analysis across different fermentation groups. These findings illustrate the significant potential of using JA juice for La and Td fermentation to develop functional juices.

Evaluation of Targeted Alpha Therapy Using [211At]FAPI1 in Triple-Negative Breast Cancer Xenograft Models.

Triple-negative breast cancer (TNBC) presents limited therapeutic options and is associated with poor prognosis. Early detection and the development of novel therapeutic agents are therefore imperative. Fibroblast activation protein (FAP) is a membrane protein expressed on cancer-associated fibroblasts (CAFs) that plays an essential role in TNBC proliferation, migration, and invasion. Consequently, it is hypothesized that the Astatine (211At)-labeled FAP inhibitor (FAPI) selectively exerts anti-tumor effects through alpha-particle emission. In this study, we aimed to assess its theranostic capabilities by integrating [18F]FAPI-74 PET imaging with targeted alpha therapy using [211At]FAPI1 in TNBC models. Mice xenografts were established by transplanting MDA-MB-231 and HT1080 cells (control). As a parallel diagnostic method, [18F]FAPI-74 was administered for PET imaging to validate FAP expression. A single dose of [211At]FAPI1 (1.04 ± 0.10 MBq) was administered to evaluate the therapeutic efficacy. [18F]FAPI-74 exhibited high accumulation in MDA-MB-231 xenografts, and FAP expression was pathologically confirmed via immunostaining. The group that received [211At]FAPI1 (n = 11) demonstrated a significantly enhanced anti-tumor effect compared with the control group (n = 7) (p = 0.002). In conclusion, [18F]FAPI-74 PET imaging was successfully used to diagnose FAP expression, and as [211At]FAPI1 showed promising therapeutic efficacy in TNBC models, it is expected to be a viable therapeutic option.

Potential probiotic Lactiplantibacillus plantarum strains alleviate TNF-α by regulating ADAM17 protein and ameliorate gut integrity through tight junction protein expression in in vitro model

BackgroundLactiplantibacillus species are extensively studied for their ability to regulate host immune responses and functional therapeutic potentials. Nevertheless, there is a lack of understanding on the mechanisms of interactions with the hosts during immunoregulatory activities.MethodsTwo Lactiplantibacillus plantarum strains MKMB01 and MKMB02 were tested for probiotic potential following Indian Council of Medical Research (ICMR) guidelines. Human colorectal adenocarcinoma cells such as HT-29, caco-2, and human monocytic cell THP-1 were also used to study the potential of MKMB01 and MKMB02 in regulating the host immune response when challenged with enteric pathogen Salmonella enterica typhimurium. Cells were pre-treated with MKMB01 and MKMB02 for 4 h and then stimulated with Salmonella. qRT-PCR and ELISA were used to analyze the genes and protein expression. Confocal microscopy and field emission scanning electron microscopy (FESEM) were used to visualize the effects. An Agilent Seahorse XF analyzer was used to determine real-time mitochondrial functioning.ResultsBoth probiotic strains could defend against Salmonella by maintaining gut integrity via expressing tight junction proteins (TJPs), MUC-2, and toll-like receptors (TLRs) negative regulators such as single Ig IL-1-related receptor (SIGIRR), toll-interacting protein (Tollip), interleukin-1 receptor-associated kinase (IRAK)-M, A20, and anti-inflammatory transforming growth factor-β and interleukin-10. Both strains also downregulated the expression of pro-inflammatory cytokines/chemokines interleukin-1β, monocyte chemoattractant protein (MCP)-1, tumor necrosis factor-alpha (TNF-α), interleukin 6, and nitric oxide (NO). Moreover, TNF-α sheddase protein, a disintegrin and metalloproteinase domain 17 (ADAM17), and its regulator iRhom2 were downregulated by both strains. Moreover, the bacteria also ameliorated Salmonella-induced mitochondrial dysfunction by restoring bioenergetic profiles, such as non-mitochondrial respiration, spare respiratory capacity (SRC), basal respiration, adenosine triphosphate (ATP) production, and maximal respiration.ConclusionsMKMB01 and MKMB02 can reduce pathogen-induced gut-associated disorders and therefore should be further explored for their probiotic potential.

KEFIRAN in vitro biological activity on enterocytes and mesodermal origin cell lines: Focus on adenocarcinoma cells HT29

Kefiran, the exopolysaccharide found in the fermented milk named “kefir” has been receiving attention over the last years due to its interesting biological and functional properties. It is a non-digestible branched glucogalactan with a molecular weight higher than 1 106 Da. Kefiran was isolated from kefir grains and its biological activity was assessed on different cell lines. Proliferation (trypan blue exclusion assay), mitochondrial activity (MTT assay), viability (Alive-Vulnerable-Dead model, AVD) and cell death were studied. It was found that proliferation was not inhibited in mesodermal cell line MC3 (osteoblast cell line) or in not-invasive colonocytes (Caco-2/TC7 cell line); on the contrary, the high invasive HT29 cell line and the UMR106 osteosarcoma cell line were sensitive to kefiran 0.5 mg/mL, and a significant diminishment on cell proliferation was found by trypan blue dye exclusion assay. Kefiran maintained mitochondrial activity while increased cell death processes in a dose-dependent way in HT29 cell line. When viability was assessed by AVD model, it was found that kefiran increased vulnerability status of neoplasic cell line HT29. In summary, in the present work, we shown that kefiran has a different effect on tumor and non-tumor cell lines. Our results contribute to the understanding of kefiran effect on the empirical antitumoral effect attributed to kefir consumption.

Design, Synthesis, and Evaluation of Doxifluridine Derivatives as Nitroreductase-Responsive Anticancer Prodrugs.

Antimetabolite antitumor drugs interfere with nucleic acid and DNA synthesis, causing cancer cell death. However, they also affect rapidly dividing normal cells and cause serious side effects. Doxifluridine (5'-deoxy-5-fluorouridine [5'-DFUR]), a 5-fluorouracil (5-FU) prodrug converted to 5-FU by thymidine phosphorylase (TP), exerts antitumor effects. Since TP is distributed in tumor and normal tissues, 5'-DFUR features side effects. Here we designed a series of novel 5'-DFUR derivatives based on high nitroreductase (NTR) levels in the hypoxic microenvironment of tumor tissues by introducing nitro-containing moieties into the 5'-DFUR structure. These derivatives exert their antitumor effects by producing 5-FU under the dual action of TP and NTR in the tumor microenvironment. The derivatives were synthesized and their stability, release, and cytotoxicity evaluated in vitro and antitumor activity evaluated in vivo. Compound 2c, featuring nitrofuran fragments, was stable in phosphate-buffered saline and plasma at different pH values and reduced rapidly in the presence of NTR. The in vitro cytotoxicity evaluation indicated that compound 2c showed excellent selectivity in the MCF-7 and HT29 cell lines. Moreover, it exhibited antitumor effects comparable to those of 5'-DFUR in vivo without significant toxic side effects. These results suggest that compound 2c is a promising antitumor prodrug.

Phytoconstituents, Antioxidant Activity and Cytotoxicity of Puya chilensis Mol. Extracts in Colon Cell Lines.

Puya chilensis Mol. is a plant of the Bromeliaceae family, which has been traditionally used for medicinal applications in various digestive disorders. In this study, the phytoconstituents of six extracts of stems and flowers of P. chilensis were evaluated: phenols, flavonoids and total anthraquinones, as well as their antioxidant capacity and cytotoxicity in colon cancer cell lines HT-29. The data demonstrate that the ethyl acetate extract of P. chilensis flowers is cytotoxic in HT-29 cell lines (IC50 = 41.70 µg/mL) without causing toxic effects on healthy colon cells (IC50 > 100 µg/mL); also, this extract concentrated the highest amount of phenols (4.63 μg GAE/g d.e.), flavonoids (31.5 μg QE/g d.e.) and anthraquinones (12.60 μg EE/g d.e.) among all the extracts tested, which also correlated with its highlighted antioxidant capacity (DPPH∙IC50 = 4.15 mg/mL and FRAP 26.52 mM TEAC) over the other extracts. About thirty-five compounds were identified in this extract-the fatty acid esters present have been shown to have therapeutic effects on several types of cancer and could explain its antiproliferative activity.

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.

Anti-cancer Activity Evaluation of Naphthalenonic and Chromanonic Spiroisoxazoline Derivatives as Selective COX-2 Inhibitors on HT29 Cell Lines

Background: Cyclooxygenase-2 (COX-2) is induced in response to proinflammatory conditions, and it is not only a key enzyme in the inflammatory process, but also seems to be highly expressed in various types of cancer cells. On the other hand, it is well documented that chemical compounds with spiro scaffolds in their structure could be effective chemical agents against cancer types. Objective: In this study, the cytotoxicity effects of spiroisoxazoline derivatives containing naphthalinone and chromanone spiro-bridge were investigated. Methods: The cytotoxicity effects of compounds 7a-7h were evaluated by performing the MTT assay on the HT-29 (colorectal cancer), MCF-7 (breast cancer), and HEK-293 (normal kidney) cell lines. After that, a compound with high yield and remarkable cytotoxic activity was selected to analyze the cell cycle and apoptosis mechanism. Results: The most effective cytotoxic activity was observed on HT-29 and MCF-7 cell lines of compounds 7b (IC50 value: 1.07±0.28 µM) and 7f (IC50 value: 11.92±1.07 µM). None of the compounds had a toxic effect on normal HEK-293 cells, except for compound 7g with an IC50 value of 21.30±16.14 µM, whose effect was much lower than that of cisplatin and doxorubicin, known as anti-cancer agents. Subsequently, compound 7e with significant yield and cytotoxic activity was investigated to evaluate cell cycle and apoptosis. The result showed that compound 7e induced significant G0/G1 cell cycle arrest and apoptosis in HT-29 cells Conclusion: The selective COX-2 inhibitor compounds with spiroisoxazoline core structure could be suitable scaffolds for cytotoxic effects.

Onion Peel Extract Prevents Intestinal Inflammation via AMK-Activated Protein Kinase Activation in Caco-2/HT-29 Cells.

Obesogenic diets cause intestinal inflammation and dysfunction. Polyphenols have shown a positive impact on reducing inflammation in in vitro studies. However, their bioactivity may not be the same in the in vivo system due to structural alteration by the gastrointestinal digestive process. The purpose of this study was to investigate the anti-inflammatory effect of onion peel and its major bioactive compound, quercetin, in the intestine and further examine the impact of intestinal digestion on this effect. Onion peel extract (OPE) and quercetin (Q) were digested using gastrointestinal digestive enzymes in vitro and then treated into lipopolysaccharide (LPS)-stimulated Caco-2/HT-29 cells. Genes and proteins related to tight junction, inflammation, and epithelial integrity were measured. OPE and digested OPE (DOPE) had a higher protective effect on LPS-induced tight junction and inflammatory genes and paracellular permeability than Q and digested Q (DQ). DOPE was more effective than OPE, while digestion did not change the activity of Q. The anti-inflammatory effect of OPE and Q with or without digestion was achieved by inhibiting nuclear factor kappa B through AMP-activated protein kinase-activated silent mating-type information regulation 2 homolog 1. It was the first to find that a crude extract, after undergoing gastrointestinal digestion, demonstrated a notably superior anti-inflammatory effect in the cell study, suggesting the consumption of onion peels could potentially yield similar benefits in the human intestine. This discovery underscores the potential of onion peel polyphenols in combating intestinal inflammation, making them a compelling area of research for future therapeutic applications using food byproducts.

Cytotoxicity of norstictic acid derivatives, a depsidone from Ramalina anceps Nyl.

Structural modifications in lichen phenolic compounds have been one of the tools to potentiate their biological activity. In the present work, seven alkyl derivatives of norstictic acid were prepared and evaluated against eight cell lines. Norstictic acid was isolated from the lichen Ramalina anceps and the alkyl derivatives were obtained through reactions with alcohols. Cytotoxicity was evaluated against the 786-0 (kidney carcinoma), MCF7 (breast carcinoma), HT-29 (colon carcinoma), PC-03 (prostate carcinoma), HEP2 (laryngeal carcinoma), B16-F10 (murine melanoma), UACC-62 (human melanoma), and NIH/3T3 (mouse embryonic fibroblast) cell lines using the sulforhodamine B assay. Norstictic acid exhibited poor activity, while the 8'-O-n-butyl-norstictic acid and 8'-O-sec-butyl-norstictic acid derivatives showed potential activity (GI50 values of 6.37-45.0 μM and 6.8-52.40 μM, respectively) and high selectivity (selectivity index (SI) values of 13.88-98.11 and SI 11.30-87.40, respectively) against all tumor cells. The 8'-O-n-hexyl-norstictic acid showed good activity (5.96-9.53 μM) and moderate selectivity (SI 9.2-5.76) against MCF7, HT-29, PC-03, and HEP2 cells, while 8'-O-isopropyl-norstictic acid demonstrated high activity and selectivity against PC-03 cells (GI50 1.28 μM and SI 33.8), and was highly active but moderately selective against UACC, HEP2, and B16-F10 cells (GI50 6.2, 7.78, and 9.65 μM; SI 7.0, 5.5, and 4.5, respectively). Additionally, 8'-O-n-pentyl- and 8'-O-tert-butyl-norstictic acids were active and selective against PC-03 cells (GI50 8.77 and 7.60 μM; SI 6.53 and 5.0, respectively). Chemometric analysis revealed a clear relationship between all compounds and their biological activities. The insertion of a four-carbon alkyl chain (n-butyl and sec-butyl) produced potentially active compounds on all tested tumor cells.

Antioxidant and Anti-Inflammatory Effects of Nettle Polyphenolic Extract: Impact on Human Colon Cells and Cytotoxicity Against Colorectal Adenocarcinoma.

Urtica dioica L. is one of the most widely utilized medicinal plants commonly applied in the form of tea, juice, and dietary supplements. This study aimed to assess the effect of the U. dioica ethanol-water extract (UdE) and polyphenolic fraction isolated from the extract (UdF) on normal human colon epithelial cells and to evaluate their protective activity against induced oxidative stress. The cytotoxic potential against human colorectal adenocarcinoma (HT29) and the anti-inflammatory effects were also investigated. UPLC-MS-DAD analysis revealed that both extracts were abundant in caffeic acid derivatives, specifically chlorogenic and caffeoylmalic acids, and therefore, they showed significant protective and ROS scavenging effects in normal human colon epithelial cells. Moreover, they had no negative impact on cell viability and morphology in normal cells and the extracts, particularly UdF, moderately suppressed adenocarcinoma cells. Furthermore, UdF significantly decreased IL-1β levels in HT29 cells. Our research indicates that U. dioica may provide significant health advantages because of its antioxidant and anti-inflammatory effects.

Potential probiotic Lactiplantibacillus plantarum DS1800 extends lifespan and enhances stress resistance in Caenorhabditis elegans model.

Probiotics are live microorganisms that provide health benefits when administered in appropriate amounts by improving or restoring the balance of intestinal microbiota. Various functional probiotic products have been developed due to the growing interest in the health-promoting and anti-aging effects of enhancing the gut microbiome. Lactiplantibacillus plantarum species are known for their potential to extend lifespan. However, this activity is strain or isolation source specific, necessitating the identification of individual strain functionalities. This study used the C. elegans model to screen probiotics for life-extension effects and analyze their functions. The 43 lactic-acid bacteria strains isolated from fermented foods, breast milk, and human feces were subjected to longevity assays, and L. plantarum DS1800 was selected to demonstrate the most effective lifespan extension. The average lifespan of Caenorhabditis elegans fed DS1800 increased by 17.36% compared with those fed Escherichia coli OP50. Further analysis of the expression of key genes related to longevity revealed the high expression of the skinhead-1 (skn-1), antibacterial, and heat stress resistance genes via the p38 MAPK pathway. These expression patterns suggest that DS1800 extends the lifespan of C. elegans by enhancing its stress resistance and protecting it against pathogens. Additionally, DS1800 exhibited excellent intestinal adhesion, with 7.56% adhesion to HT-29 cells. Therefore, L. plantarum DS1800 is effective in extending the lifespan of C. elegans and can be used as a functional probiotic.

Scutellarein ameliorates dextran sulfate sodium-induced ulcerative colitis by inhibiting colonic epithelial cell proinflammation and barrier disruption.

Scutellarein (Scu) is a natural occurring flavonoid found in multiple traditional Chinese medicines such as Oroxylum indicum (L.) Kurz and Scutellaria baicalensis, with various pharmacological activities including anti-inflammation, anti-oxidation and myocardial protection. Here, we investigated the therapeutic efficacy of Scu on ulcerative colitis (UC) and the underlying mechanism. Efficacy of Scu on UC was evaluated in dextran sulfate sodium (DSS) induced colitis mouse model. Inflammation in colonic tissues was assessed by myeloperoxidase activity assay and RT-qPCR. Barrier proteins expression was examined using immunostaining and Western blot. IL-1β-treated HT-29 cells was used for mechanical investigation. Gavage of Scu significantly decreased the DAI score, improved colon shortening, ameliorated the pathological score in DSS-treated mice with better efficacy than the positive drug, 5-aminosalicylic acid. Scu also inhibited the expression levels of cytokines (Il-1β, Tnf-α, Il-1α, Il-6, and Cxcl1) as well as barrier proteins (E-cadherin, Occludin, and ZO-1) in colon tissues of DSS mice. In intestinal epithelial HT-29 cells, Scu attenuated the IL-1β-downregulated expression levels of E-cadherin, occludin, and ZO-1, while reduced IL-1β-upregulated IL-6 and IL-8 mRNA levels. Moreover, Scu inhibited the phosphorylation and nuclear translocation of NF-κB and suppression of NF-κB phosphorylation abolished IL-1β-disrupted epithelial barrier integrity and IL-1β-upregulated proinflammatory mediators expression in HT-29 cells. These data demonstrate that Scu is an efficacious therapeutic agent to treat UC. Inhibition of inflammatory responses and maintenance of epithelial barrier integrity through NF-κB signaling pathway underlines Scu therapeutic effect on UC.

Myrciaria jaboticaba Fruit Peel: Bioactive Composition as Determined by Distinct Harvest Seasons and In Vitro Anti-Cancer Activity.

Jaboticaba (Myrciaria jaboticaba) is a recognizable and unique crop from Brazil. The fruit's byproducts are currently being studied, given their bioactive composition and promising anti-cancer potential. It is not evident, however, if different harvesting seasons can modify the chemical profile and antioxidant capacity of jaboticaba fruit fractions. Furthermore, as there is limited data for jaboticaba's anti-proliferative effects, additional assessments are required to improve the robustness of these findings. Therefore, this study aimed to determine the composition of the peel of jaboticaba collected in two periods (May-off-season, sample 1-and August-October-peak season, sample 2) and test the peel's richest anthocyanin sample against colorectal cancer (CRC) cell lines. To accomplish this, proximate, spectrophotometric, and chromatographic analyses were performed in two freeze-dried samples; and anti-proliferative and/or colony-forming assays were carried out in Caco-2, HT29, and HT29-MTX cells. As a result, sample 2 showed the highest levels of polyphenols overall, including flavonoids and anthocyanins. This sample displayed significative higher contents of cyanidin-3-O-glucoside (48%) and delphinidin-3-O-glucoside (105%), in addition to a superior antioxidant capacity (23% higher). Sample 1 showed higher amounts of total protein, gallic acid (20% higher), and specific carotenoids. An aqueous extract from sample 2 was tested against CRC, showing anti-proliferative effects for Caco-2 cells at 1 and 2 mg/mL concentrations, with IC50 values of 1.2-1.3 mg/mL. Additionally, the extract was able to inhibit cell colony formation when tested at both low and high concentrations. In conclusion, jaboticaba collected in the main season stands out regarding its polyphenol composition and holds potential against cancer cell growth.

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.

Thiazepine-based Hybrids as Promising Anti-colon Cancer Agents: Design, Synthesis, Computational and In Vitro Screening.

Novel thiazepine-based hybrids (9a-d) were designed and synthesized to create lead molecules with exceptional anti-colon cancer efficacy. Analytical methods, including IR, NMR, and HR-MS, characterized the synthesized compounds. The in vitro colorectal study was carried out to compare the biological activity of newly developed compounds with the computational data. The tested compounds induced cytotoxicity in HT-29 cells for both 24h and 48h in a dose-dependent manner. However, compound 9a induced cytotoxicity at much higher concentrations compared to the rest of the compounds. 9b and 9c caused 50% cell death (compared to the untreated cells) at a dose of ~ 50µM and 40 µM in case of 24-hour exposure, respectively. On the contrary, for 48h exposure, both 9b and 9c induced 50% cell death concerning untreated cells at a dose of around ~20 µM, whereas 9d exhibited 50% cell death at 5 µM in the case of 48 h exposure. In silico ADMET was also carried out to understand the pharmacokinetics and safety profiles of the drug candidates. We found some of the critical targets of these compounds, which eventually will be integral to exploring the mechanistic actions of these compounds in colon cancer.

Co-delivery of Doxorubicin and VEGF siRNA using carcinoembryonic antigen-targeted chitosan nanoparticles for colorectal cancer

Developing a targeted nanocarrier-based therapeutic approach is highly effective in delivering therapeutics to specific cancer cells. Combining two or more therapeutic approaches with different mechanisms of action can effectively maintain their unique benefits while considerably improving cancer treatment. Doxorubicin (DOX) binds to DNA by intercalation and triggers a cascade of biochemical processes in tumor cells leading to death. Anti-VEGF medicines may sensitize cancer cells to chemotherapeutics by limiting blood supply and increasing penetration by disrupting tumor vasculature. In this study, DOX and vascular endothelial growth factor siRNA (VEGF siRNA) loaded chitosan nanoparticles (ChNPs) were prepared by using the ionic gelation method. The DOX-siRNA-ChNPs surface was then functionalized with carcinoembryonic antigen (CEA) targeted monoclonal antibody (DOX-siRNA-ChNPs-CMAb), thereby delivering therapeutic moieties into the cancer milieu. The cytotoxicity and cellular uptake of the developed nanocarrier were tested in-vitro on HT-29, which revealed that DOX-siRNA-ChNPs-CMAb possessed the highest cytotoxicity when compared with other prepared formulations and free DOX. The cellular uptake using fluorescence microscopy displayed a significantly higher uptake of DOX-siRNA-ChNPs-CMAb (targeted) than free DOX and DOX-siRNA-ChNPs (non-targeted). Apoptosis analysis in flow cytometry using annexin-V/PI staining demonstrated a substantially higher apoptosis rate of HT-29 cells using DOX-siRNA-ChNPs-CMAb than DOX-siRNA-ChNPs. Moreover, DOX-siRNA-ChNPs-CMAb were studied in CRC induced animal models, showing that DOX-siRNA-ChNPs-CMAb specifically targets CRC cells without showing organ toxicity. Thus, the novel approach of simultaneous delivery of DOX and siRNA using CMAb conjugated ChNPs demonstrated significant targeting in both in-vitro and in-vivo with minimal organ toxicity.

Exploring the impact of sEH inhibition on intestinal cell differentiation and Colon Cancer: Insights from TPPU treatment

Inhibition of soluble epoxide hydrolase (sEH) appears to be promising for the treatment of many diseases. Studies have focused on the beneficial effects of epoxyeicosatrienoic acids (EETs), which are sEH substrates. However, our recent studies have shown that the sEH activity is crucial for the proper intestinal cell differentiation. In this recent study, we investigated the impact of TPPU, an inhibitor of sEH, on the colon cancer cell lines Caco2 and HT-29. We analysed the changes in the expression of the cytoskeletal protein ezrin and the phosphorylated protein kinase p38 (p-p38). Our results showed a decrease in ezrin expression in differentiated cells and an increase in p-p38 expression after TPPU treatment. Immunocytochemical staining revealed a higher staining intensity of p-p38 in the nuclei of HT-29 cells following TPPU treatment. Immunohistochemical staining was performed on human samples of normal colon tissue, grade 2 tumours, and embryonal/foetal tissues. The staining intensity of ezrin in tumours was reduced in the surface area compared to the crypts. Additionally, we observed the translocation of p-p38 expression from the cytoplasm to the nucleus during differentiation. The tumour samples exhibited higher levels of p-p38 in the cytoplasm, similar to normal undifferentiated tissue. To observe the disruption of the cytoskeleton after TPPU treatment, confocal microscopy was used. It was found that β-actin associated with ezrin forms clusters under the plasma membranes. All of these results are significant because sEH inhibitors are being tested in clinical trials, but they could cause an unexpected adverse effects.

Bioactivities of Quinic Acids from Vitex rotundifolia Obtained by Supercritical Fluid Extraction.

Acyl-quinic acids (AQAs), present in various plants with many health benefits, are regarded as therapeutic agents in the prevention and treatment of chronic and cardiovascular diseases. The molecular network-guided identification of ten AQA compounds, two new (5 and 7) and eight known compounds, were isolated from V. rotundifolia L. f. by using a newly applied extraction method. Their structures were determined through spectroscopic means, reaction mixtures, and modified Mosher and PGME techniques. These compounds were assessed for their anti-inflammatory and antioxidant capabilities. Notably, compounds 1, 3, 4, 6, 8, and 9 exhibited notable DPPH radical scavenging activity. In LPS-induced HT-29 cells, compounds 2-7 significantly inhibited IL-8 production. Furthermore, compounds 3-5 and 7 markedly suppressed NO production, while compounds 1-10 effectively inhibited IL-6 production in LPS-induced RAW264.7 cells. Western blot analyses revealed that compounds 3-5, and 7 reduced iNOS and COX-2 expression, and compounds 2-5, 7, and 8 also diminished the expression levels of p38 MAPK phosphorylation. Docking studies demonstrated the active compounds' binding affinity with the IL-8, iNOS, COX-2, and p38 MAPK proteins through interactions with essential amino acids within the binding pockets of complexes. The findings suggest that compounds 1, 3, 4, 6, 8, and 9, and compounds 3-5, and 7, hold promise as potential therapeutic agents for treating antioxidative and inflammatory diseases, respectively.

A polysaccharide with a triple helix structure from Agaricus bisporus: Characterization and anti-colon cancer activity

In this study, A polysaccharide WAAP-2 (121 kDa) with a triple-helical structure was isolated and purified from Agaricus bisporus for the first time. The physicochemical properties, structural characteristics and anti-colon cancer activity were preliminarily investigated. The primary structure indicated that WAAP-2 was composed of mannose, glucose and galactose and determined the position of the linkage between monosaccharide residues. The advanced structure revealed that WAAP-2 has a triple helix and tangled chain conformation. In the anti-colon cancer activity investigation, WAAP-2 exerted an apoptosis-inducing effect by causing HT-29 cell cycle arrest in S phase. WAAP-2 promoted HT-29 cell apoptosis by up-regulating the expression of Caspase-3 and Bax proteins while down-regulating the expression of Bcl-2 protein. Besides, WAAP-2 could inhibit the migration and invasion of colorectal cancer cells by inducing E-cadherin expression and inhibiting Vimentin expression to affect epithelial mesenchymal transition. This paper is of importance for the application of WAAP-2, a triple-helical structural polysaccharide from Agaricus bisporus, to low-toxicity anti-colon cancer drugs.