Here, we investigated the possible anti-cancer properties of bromelain in Kras mutant human colorectal carcinoma cell lines and a mouse model harboring a Kras mutation. Cell growth and proliferation were significantly reduced in the Kras mutant colorectal carcinoma cell lines following treatment with 50 μg/mL bromelain as assessed by crystal violet staining and a proliferation assay. To identify the molecules responsible for this action, the expression levels of genes involved in signaling pathways and miRNAs were analyzed by real-time PCR. Among the genes tested, down-regulation of ACSL-4 and up-regulation of miRNAs targeting ASCL-4 were observed in Caco2 cells. Compared to the Kras wild-type colorectal carcinoma cell lines, Kras mutant colorectal carcinoma cell lines exhibited a remarkably up-regulated expression of ACSL-4, which is responsible for ferroptosis sensitivity. Moreover, the knockdown of ACSL-4 by a specific shRNA inhibited erastin-induced ferroptosis in Kras mutant DLD-1 cells as assessed by propidium iodide staining and lipid reactive oxygen species measurement. Our findings indicate that bromelain effectively exerts cytotoxic effects in Kras mutant colorectal cancer cells compared to in Kras wild-type colorectal cancer cells. Differential expression of ACSL-4 is responsible for the differential action of bromelain in regulating ferroptotic cell death.

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New approaches to veterinary drug screening based on liquid chromatography-mass spectrometry (LC-MS/MS) and time-of-flight mass spectrometry (ToF/MS) are rapid and have high selectivity and sensitivity. In this study, we developed a multiresidue method for screening over 100 veterinary drug residues using ion trap (IT)-ToF/MS. The screened compounds comprised major drug classes used in veterinary practice, representing the following: amphenicols, anthelmintics, benzimidazoles, β-lactams, coccidiostats, ionophores, macrolides, non-steroidal anti-inflammatory drugs, quinolones, sulfonamides, tetracyclines, and tranquilizers. The method was developed based on chromatographic retention time, specific accurate mass, isotope distribution, and fragment data. Each compound was validated at three levels, and the mass accuracy, accuracy, and repeatability were calculated. All parameters showed acceptable values and conformed to the Commission Decision 2002/657/EC criteria. This screening method can simultaneously analyze over 100 veterinary drugs in meat, milk, eggs, and fish in a single analytical run.

ObjectiveGlucosamine is widely used to improve the symptoms and to delay the structural progression of osteoarthritis. However, its efficacy in osteoarthritis has been controversial and its underlying mechanism of action remains unclear. The aim of this study was to investigate the effects of glucosamine and the underlying mechanisms in human chondrocytes.MethodsChondrocytes from normal human articular cartilage were treated with glucosamine (10–100 mM). Subsequently, cell death was analyzed by Annexin V staining and FACS and mitochondrial function was studied by measuring the mitopotential. Peroxisomal function was analyzed by BODIPY staining, and gene expression of PMP70 and acyl-CoA oxidase 1, by real-time PCR. Total lipids were analyzed by gas chromatography/mass spectrometry. Autophagy activation was determined by western blotting of beclin and light chain 3B. Autophagosome formation was analyzed by introduction of green fluorescent protein (GFP) LC3, and pexophagy was determined by introduction of mRFP-EGFP-SKL plasmids.ResultsTreatment of chondrocytes with glucosamine exerts exposure time-dependent dual effects on apoptosis/autophagy. Short time exposure of glucosamine to chondrocytes activated autophagy, pexophagy, and peroxidation. On the other hand, long time exposure of glucosamine had opposite effects, namely accumulation of very long chain fatty acids and peroxisomal dysfunction.ConclusionWe highlight the dual role of glucosamine in apoptosis/autophagy in human chondrocytes depending on exposure time. Although further research is required to fully understand the dual effects of glucosamine, dosage and duration of glucosamine treatment are clear contributing factors towards the line of beneficial reward-to-risk action.

Recently, intermittent fasting, also known as time-restricted eating (TRE), has become a popular diet trend. Compared to animal studies, there have been few studies and inconclusive findings investigating the effects of TRE in humans. In this study, we examined the effects of 8 h TRE on body weight and cardiometabolic risk factors in young adults who were mainly active at night. A total of 33 young adults completed the 8 h TRE for 4 weeks. Body composition was measured by bioelectrical impedance analysis at baseline and every 2 weeks, and blood samples were collected at baseline and week 4. Daily dietary records were logged throughout the intervention period. Participants experienced significant changes in body weight (−1.0 ± 1.4 kg), body mass index (−0.4 ± 0.5 kg/m2), and body fat (−0.4 ± 1.9%) after 4 weeks of TRE. When participants were divided into weight loss/gain groups based on their weight change in week 4, fat mass reduction was significantly higher in the weight loss group than in the weight gain group. Regarding cardiometabolic risk factors, levels of fasting insulin and insulin resistance improved in the weight loss group after intervention, but not in the weight gain group. All subjects showed late-shifted sleeping patterns, but no significant differences in sleep duration, sleep quality, or psychological measures between the two groups. When meal frequency and energy proportion were evaluated, the average meal frequency was 2.8 ± 0.5 and energy proportions of breakfast, lunch, dinner, and snacks were 4.5, 39.2, 37.6, and 18.5%, respectively; there were no significant differences between the two groups. However, the saturated fat intake at dinner was lower in the weight loss group (3.1 ± 3.2%, 6.0 ± 2.5% respectively). In conclusion, 8 h TRE can be applied as a lifestyle strategy to manage body weight and cardiometabolic risk factors among young adults with late chronotypes.

Extracellular vesicles are cell-derived membranous vesicles that are secreted into biofluids. Emerging evidence suggests that EVs play an essential role in the pathogenesis of many diseases by transferring proteins, genetic material, and small signaling molecules between cells. Among these molecules, microRNAs (miRNAs), a type of small noncoding RNA, are one of the most important signals and are involved in various biological processes. Lung cancer is one of the leading causes of cancer-related deaths worldwide. Early diagnosis of lung cancer may help to reduce mortality and increase the 5 years survival rate and thereby reduce the associated socioeconomic burden. In the past, EV-miRNAs have been recognized as biomarkers of several cancers to assist in diagnosis or prognosis. In this review, we discuss recent findings and clinical practice for EV-miRNAs of lung cancer in several biofluids, including blood, bronchoalveolar lavage fluid (BALF), and pleural lavage.

Carbon monoxide dehydrogenase (CO-DH) is an enzyme catalysing the oxidation of CO to carbon dioxide in Mycobacterium sp. strain JC1 DSM 3803. Cloning of the genes encoding CO-DH from the bacterium and sequencing of overlapping clones revealed the presence of duplicated sets of genes for three subunits of the enzyme, cutB1C1A1 and cutB2C2A2, in operons, and a cluster of genes encoding proteins that may be involved in CO metabolism, including a possible transcriptional regulator. Phylogenetic analysis based on the amino acid sequences of large subunits of CO-DH suggested that the CO-DHs of Mycobacterium sp. JC1 and other mycobacteria are distinct from those of other types of bacteria. The growth phenotype of mutant strains lacking cutA genes and of a corresponding complemented strain showed that both of the duplicated sets of CO-DH genes were functional in this bacterium. Transcriptional fusions of the cutB genes with lacZ revealed that the cutBCA operons were expressed regardless of the presence of CO and were further inducible by CO. Primer extension analysis indicated two promoters, one expressed in the absence of CO and the other induced in the presence of CO. This is believed to be the first report to show the presence of multiple copies of CO-DH genes with identical sequences and in close proximity in carboxydobacteria, and to present the genetic evidence for the function of the genes in mycobacteria.

RT-PCR, nucleotide sequencing, and phylogenetic analysis were performed for genotyping and molecular characterization of noroviruses isolated from Korean groundwater. Among 160 samples collected from 80 sites between 2008 and 2010, 14 samples (8.7 %) from 12 sites were positive for noroviruses (NoVs). The percentages of NoV-positive samples in 2008, 2009, and 2010 were 22.2, 3.2, and 0 %, respectively, representing a yearly decrease. GII-positive samples (n = 9, 5.6 %) outnumbered GI-positive samples (n = 5, 3.1 %). The genotypes of the GI NoVs were GI.2, GI.5, and GI.6, and the genotypes of the GII NoVs were all GII.4. One sample, HM623465, was very similar to CUK-3 and CBNU2 and two GII.4 sequences isolated from the stool of Korean gastroenteritis patients. A BLASTN search revealed several nucleotide sequences highly similar to those of NoVs isolated in this study. The original isolation sources for these similar NoVs were mostly stool (n = 731, 80.0 %) and groundwater (n = 135, 14.8 %), and all the countries from which they were isolated were almost in Asia (96.0 %); specifically, China (n = 192, 21.0 %), Japan (n = 383, 41.9 %), Korea (n = 296, 32.4 %), and other Asian countries (n = 6, 0.7 %). These results suggest that Korean groundwater might be contaminated with NoVs from the stool of infected patients and that these NoVs in turn cause new cases of gastroenteritis through a typical fecal-oral route with region-specific circulation. Therefore, it is important to properly treat sewage, which may include waterborne viruses and manage point sources in groundwater for national health and sanitation. In addition, continuous molecular surveillance remains important for understanding circulating NoVs.