Mechanisms of cuproptosis and its relevance to distinct diseases.

Copper is a trace element required by the organism, but once the level of copper exceeds the threshold, it becomes toxic and even causes death. The underlying mechanisms of copper-induced death are inconclusive, with different studies showing different opinions on the mechanism of copper-induced death. Multiple investigations have shown that copper induces oxidative stress, endoplasmic reticulum stress, nucleolar stress, and proteasome inhibition, all of which can result in cell death. The latest research elucidates a copper-dependent death and denominates it as cuproptosis. Cuproptosis takes place through the combination of copper and lipoylated proteins of the tricarboxylic acid cycle, triggering agglomeration of lipoylated proteins and loss of iron-sulfur cluster proteins, leading to proteotoxic stress and ultimately death. Given the toxicity and necessity of copper, abnormal levels of copper lead to diseases such as neurological diseases and cancer. The development of cancer has a high demand for copper, neurological diseases involve the change of copper contents and the binding of copper to proteins. There is a close relationship between these two kinds of diseases and copper. Here, we summarize the mechanisms of copper-related death, and the association between copper and diseases, to better figure out the influence of copper in cell death and diseases, thus advancing the clinical remedy of these diseases.

TM9SF1 promotes bladder cancer cell growth and infiltration.

BACKGROUND: Bladder cancer (BC) is the most common urological tumor. It has a high recurrence rate, displays tutor heterogeneity, and resists chemotherapy. Furthermore, the long-term survival rate of BC patients has remained unchanged for decades, which seriously affects the quality of patient survival. To improve the survival rate and prognosis of BC patients, it is necessary to explore the molecular mechanisms of BC development and progression and identify targets for treatment and intervention. Transmembrane 9 superfamily member 1 (TM9SF1), also known as MP70 and HMP70, is a member of a family of nine transmembrane superfamily proteins, which was first identified in 1997. TM9SF1 can be expressed in BC, but its biological function and mechanism in BC are not clear. AIM: To investigate the biological function and mechanism of TM9SF1 in BC. METHODS: Cells at 60%-80% confluence were transfected with lentiviral vectors for 48-72 h to achieve stable TM9SF1 overexpression or silencing in three BC cell lines (5637, T24, and UM-UC-3). The effect of TM9SF1 on the biological behavior of BC cells was then investigated through CCK8, wound-healing assay, transwell assay, and flow cytometry. RESULTS: Overexpression of TM9SF1 increased the in vitro proliferation, migration, and invasion of BC cells by promoting the entry of BC cells into the G2/M phase. Silencing of TM9SF1 inhibited in vitro proliferation, migration, and invasion of BC cells and blocked BC cells in the G1 phase. CONCLUSION: TM9SF1 may be an oncogene in BC.

Application of near-infrared spectroscopy for hay evaluation at different degrees of sample preparation.

OBJECTIVE: A study was conducted to quantify the performance differences of the nearinfrared spectroscopy (NIRS) calibration models developed with different degrees of hay sample preparations. METHODS: A total of 227 imported alfalfa (Medicago sativa L.) and another 360 imported timothy (Phleum pratense L.) hay samples were used to develop calibration models for nutrient value parameters such as moisture, neutral detergent fiber, acid detergent fiber, crude protein, and in vitro dry matter digestibility. Spectral data of hay samples prepared by milling into 1-mm particle size or unground were separately regressed against the wet chemistry results of the abovementioned parameters. RESULTS: The performance of the developed NIRS calibration models was evaluated based on R2, standard error, and ratio percentage deviation (RPD). The models developed with ground hay were more robust and accurate than those with unground hay based on calibration model performance indexes such as R2 (coefficient of determination), standard error, and RPD. Although the R2 of calibration models was mainly greater than 0.90 across the feed value indexes, the R2 of cross-validations was much lower. The R2 of cross-validation varies depending on feed value indexes, which ranged from 0.61 to 0.81 in alfalfa, and from 0.62 to 0.95 in timothy. Estimation of feed values in imported hay can be achievable by the calibrated NIRS. However, the NIRS calibration models must be improved by including a broader range of imported hay samples in the modeling. CONCLUSION: Although the analysis accuracy of NIRS was substantially higher when calibration models were developed with ground samples, less sample preparation will be more advantageous for achieving rapid delivery of hay sample analysis results. Therefore, further research warrants investigating the level of sample preparations compromising analysis accuracy by NIRS.

Entecavir versus tenofovir for prevention of hepatitis B virus-associated hepatocellular carcinoma after curative resection: study protocol for a randomized, open-label trial.

BACKGROUND: Entecavir and tenofovir disoproxil fumarate (TDF) are standard first-line treatments to prevent viral reactivation and hepatocellular carcinoma (HCC) in individuals chronically infected with the hepatitis B virus (HBV), but the long-term efficacy of the two drugs remains controversial. Also unclear is whether the drugs are effective at preventing viral reactivation or HCC recurrence after hepatectomy to treat HBV-associated HCC. This trial will compare recurrence-free survival, overall survival, viral indicators and adverse events in the long term between patients with HBV-associated HCC who receive entecavir or TDF after curative resection. METHODS: This study is a randomized, open-label trial. A total of 240 participants will be randomized 1:1 into groups receiving TDF or entecavir monotherapy. The two groups will be compared in terms of recurrence-free and overall survival at 1, 3, and 5 years after surgery; adverse events; virological response; rate of alanine transaminase normalization; and seroreactivity at 24 and 48 weeks after surgery. DISCUSSION: This study will compare long-term survival between patients with HBV-associated HCC who receive TDF or entecavir monotherapy. Numerous outcomes related to prognosis will be analyzed and compared in this study. TRIAL REGISTRATION: ClinicalTrials.gov NCT02650271. Registered on January 7, 2016.

Assessment of growing condition variables on alfalfa productivity.

This study was conducted to assess the impact of growing condition variables on alfalfa (Medicago sativa L.) productivity. A total of 197 alfalfa yield results were acquired from the alfalfa field trials conducted by the South Korean National Agricultural Cooperative Federation or Rural Development Administration between 1983 and 2008. The corresponding climate and soil data were collected from the database of the Korean Meteorological Administration. Twenty-three growing condition variables were developed as explaining variables for alfalfa forage biomass production. Among them, twelve variables were chosen based on the significance of the partial-correlation coefficients or potential agricultural values. The selected partial correlation coefficients between the variables and alfalfa forage biomass ranged from -0.021 to 0.696. The influence of the selected twelve variables on yearly alfalfa production was summarized into three dominant factors through factor analysis. Along with the accumulated temperature variables, the loading scores of the daily mean temperature higher than 25°C were over 0.88 in factor 1. The sunshine duration at temperature between 0°C-25°C was 0.939 in factor 2. Precipitation days were 0.82, which was the greatest in factor 3. Stepwise regression applied with the three dominant factors resulted in the coefficients of factors 1, 2, and 3 for 0.633, 0.485, and 0.115, respectively, and the R-square of the model was 0.602. The environmental conditions limiting alfalfa growth, such as daily temperature higher than 25°C or daily mean temperature affected annual alfalfa production most substantially among the growing condition variables. Therefore, future cultivar selection should consider the capability of alfalfa to be tolerant to extreme summer weather along with biomass production potential.

Insights into the metabolite profiles of Rubus chingii Hu at different developmental stages of fruit.

The fruits of Rubus chingii Hu have high medicinal and nutritional values. However, the metabolite profiles of R. chingii, especially the alterations during different development stages of fruit, have not been comprehensively analyzed, hindering the effective utilization of the unique species. In this study, we comprehensively analyzed the metabolites of R. chingii fruit at four developmental stages using systematic untargeted and targeted liquid chromatography-mass spectrometry metabolomics analysis and identified 682 metabolites. Significant changes were observed in metabolite accumulation and composition in fruits during the different developmental stages. The contents of the index components, kaempferol-3-O-rutinoside and ellagic acid, were the highest in immature fruit. The analysis identified 64 differentially expressed flavonoids and 39 differentially expressed phenolic acids; the accumulation of most of these differentially expressed metabolites decreased with the developmental stages of fruit from immaturity to maturity. These results confirmed that the developmental stages of fruit are a critical factor in determining its secondary metabolite compositions. This study elucidated the metabolic profile of R. chingii fruit at different stages of development to understand the dynamic changes in metabolites.

Five previously undescribed compounds from Ajuga lupulina Maxim. and their in vitro activities.

A new biflavone (philonotisflavone-3'''-methyl ether), three diterpenes (lupulin G, lupulin H, lupulin I), a new ecdysteroid (ajugasterone E), and four known compounds were isolated from the whole plant of Ajuga lupulina Maxim. Their structures were determined by spectroscopic analysis, including MS, NMR and ECD spectral data. Compounds 1-3 has DPPH radical scavenging ability, and compound 1 has stronger activity than vitamin C. Compounds 2, 3, 7 and 8 have potential cytotoxic activity against Hela, with IC50 values less than 20.0 µM. Abietane diterpenes 2, 3, 7 and 8 are also found to have NO inhibitory effects with IC50 values less than 40.0 µM.

Beef Steer Performance on Irrigated Monoculture Legume Pastures Compared with Grass- and Concentrate-Fed Steers.

Fall- or spring-born steers grazed monoculture irrigated birdsfoot trefoil (BFT; Lotus corniculatus L.) or cicer milkvetch (CMV; Astragalus cicer L.) pastures for approximately 12 weeks for 3 years and were compared with steers on concentrate diets. In the 3rd year, an irrigated meadow bromegrass (MBG; Bromus biebersteinii Roem. and Schult.) pasture treatment was added for further comparison. Steer average daily gain (ADG) was 1.31, 0.94, 0.83 and 0.69 kg d−1 on concentrate, 'Norcen' BFT, 'Oberhaunstadter' BFT, and 'Monarch' CMV diets, respectively; ADG on grass pastures was 0.43 kg d−1. The ADG on the concentrate diet was greater than ADG on legume or grass pastures, ADG was greater on BFT than CMV in every year (p < 0.03), and ADG on BFT was greater than ADG on grass (p < 0.03). The rate constant of gas production of an in vitro rumen fermentation demonstrated a slower rate of microbial digestion for CMV than for BFT. The elevated ADG on BFT pastures may be due to greater non-fiber carbohydrate (NFC) concentration and reduced neutral detergent fiber (NDF) concentration combined with condensed tannins that protect proteins in the rumen but do not impede protein digestion in the abomasum and intestines.

Four sesquiterpenoids from the vegetable raw material Schisandra chinensis (Turcz.) Baill.: leaves and stems.

Four sesquiterpenoids A-D (1-4) were isolated from the ethanol extracts of the leaves and stems from Schisandra chinensis. Their structures and absolute configurations were elucidated by a combination of NMR, MS and ECD. Compounds 1-4 (10 µM) exhibited moderate hepatoprotective activities against APAP-induced LO2 cell damage with increasing cell viability by 18%, 17%, 16%, and 19% compared to the model group (bicyclol, 26%) at 10 µM, respectively. All the compounds displayed no cytotoxic activity against five human cell lines, which also suggested the safety of leaves and stems of S. chinensis as an edible vegetable in a degree.

PARK14 (D331Y) PLA2G6 Causes Early-Onset Degeneration of Substantia Nigra Dopaminergic Neurons by Inducing Mitochondrial Dysfunction, ER Stress, Mitophagy Impairment and Transcriptional Dysregulation in a Knockin Mouse Model.

PARK14 patients with homozygous (D331Y) PLA2G6 mutation display motor deficits of pure early-onset Parkinson's disease (PD). The aim of this study is to investigate the pathogenic mechanism of mutant (D331Y) PLA2G6-induced PD. We generated knockin (KI) mouse model of PARK14 harboring homozygous (D331Y) PLA2G6 mutation. Then, we investigated neuropathological and neurological phenotypes of PLA2G6D331Y/D331Y KI mice and molecular pathogenic mechanisms of (D331Y) PLA2G6-induced degeneration of substantia nigra (SN) dopaminergic neurons. Six-or nine-month-old PLA2G6D331Y/D331Y KI mice displayed early-onset cell death of SNpc dopaminergic neurons. Lewy body pathology was found in the SN of PLA2G6D331Y/D331Y mice. Six-or nine-month-old PLA2G6D331Y/D331Y KI mice exhibited early-onset parkinsonism phenotypes. Disrupted cristae of mitochondria were found in SNpc dopaminergic neurons of PLA2G6D331Y/D331Y mice. PLA2G6D331Y/D331Y mice displayed mitochondrial dysfunction and upregulated ROS production, which may lead to activation of apoptotic cascade. Upregulated protein levels of Grp78, IRE1, PERK, and CHOP, which are involved in activation of ER stress, were found in the SN of PLA2G6D331Y/D331Y mice. Protein expression of mitophagic proteins, including parkin and BNIP3, was downregulated in the SN of PLA2G6D331Y/D331Y mice, suggesting that (D331Y) PLA2G6 mutation causes mitophagy dysfunction. In the SN of PLA2G6D331Y/D331Y mice, mRNA levels of eight genes that are involved in neuroprotection/neurogenesis were decreased, while mRNA levels of two genes that promote apoptotic death were increased. Our results suggest that PARK14 (D331Y) PLA2G6 mutation causes degeneration of SNpc dopaminergic neurons by causing mitochondrial dysfunction, elevated ER stress, mitophagy impairment, and transcriptional abnormality.

Optimisation of extraction conditions for total saponins from Cynanchum wallichii using response surface methodology and its anti-tumour effects.

Cynanchum wallichii Wight, is a traditional Chinese medicine herb, which is rich in saponins and has varieties of pharmacological activities. In this study, a standardized C. wallichii extract was established and the anti-tumor activity of the total saponins was evaluated by MTT assay. The extraction conditions of the standardized extract was optimized using response surface methodology. The experimental value was in good agreement (the yield 4.28%) with predicted values. The total saponins of the extract showed significant anti-tumor activity against three human tumor cell lines (A549, HepG2 and MCF-7), especially for MCF-7 (IC50. 67.63 µg/mL) cells in vitro.

Effect of piperine on the bioavailability and pharmacokinetics of rosmarinic acid in rat plasma using UPLC-MS/MS.

1. The purpose of the present study was to investigate the effect of piperine (PP) on the pharmacokinetics of rosmarinic acid (RA) in rat plasma and to determine whether PP could enhance the oral bioavailability of RA via inhibition of its glucuronidation. 2. The pharmacokinetic profiles of RA between oral administration of RA (50 mg/kg) alone and in combination with different oral dose PP (20, 40, 60, and 80 mg/kg) to rats were investigated via a validated UPLC/MS/MS method. 3. The AUC and Cmax of RA were significantly increased in combination with different dose PP dose dependently, especially in the presence of 60 and 80 mg/kg PP (p < 0.01). The relative bioavailability of RA in the presence of 20, 40, 60, and 80 mg/kg PP was 1.24-, 1.32-, 2.02-, and 2.26-folds higher, respectively, compared with the control group given RA alone. Compared with RA, the pharmacokinetic modulations of RA glucuronide were even more apparent, and the glucuronidation of RA was remarkedly inhibited. 4. This study demonstrated that PP significantly improved the in vivo bioavailability of RA partly attributing to the inhibition of gut and hepatic metabolism enzymes of RA.

(R1441C) LRRK2 induces the degeneration of SN dopaminergic neurons and alters the expression of genes regulating neuronal survival in a transgenic mouse model.

Mutation of leucine-rich repeat kinase 2 (LRRK2) is the most common genetic cause of both familial and sporadic Parkinson's disease (PD) cases. Several mutations in LRRK2 gene were reported in PD patients. R1441 is the second most frequent site of LRRK2 mutation. We generated (R1441C) LRRK2 transgenic mice that displayed motor deficits at the age of 16 months. Compared with wild-type mice, 16-month-old (R1441C) LRRK2 mice exhibited a significant reduction in the number of substantia nigra (SN) dopaminergic neurons. To elucidate molecular pathogenic pathways involved in (R1441C) LRRK2-induced death of SN dopaminergic neurons, we performed microarray analysis to visualize altered mRNA expressions in the SN of (R1441C) LRRK2 mouse. In the SN of (R1441C) LRRK2 transgenic mouse, the mRNA expression of three genes that promote cell death was upregulated, while the mRNA expression of seven genes that contribute to neurogenesis/neuroprotection was significantly downregulated. Our results suggest that altered expression of these genes involved in regulating neuronal survival may contribute to the pathogenesis of (R1441C) LRRK2-induced PD.

Simultaneous Determination of 10 Active Components in Hulisan Tablets by a High-Performance Liquid Chromatographic-Diode Array Detector.

Hulisan tablets (HLST), a famous classic traditional Chinese prescriptions consisting of four medicinal herbs, have been applied for treating fractures, rheumatoid arthritis, mechanical phlebitis and traumatic bleeding extensively. In this study, a simple and reliable method using high-performance liquid chromatography with diode array detector (HPLC-DAD) was established for the simultaneous determination of 10 active compounds in HLST. The chromatographic analysis was performed on a Symmetry(®)-C18 column (4.6 × 250 mm, i.d., 5 µm; Waters, Wexford, Leinster, Ireland) at 30°C with a gradient elution of 0.1 mol/L ammonium acetate (containing 0.5 mL glacial acetic acid per 1,000 mL) and acetonitrile at a flow rate of 1.0 mL/min. The detection UV wavelengths were set at 232, 254 and 280 nm. The method was validated by linearity, precision, stability and recovery. Calibration curves for the 10 compounds showed good linear regressions (R(2) > 0.9992). The limits of detection and quantification fell in the ranges of 0.03-0.36 and 0.11-1.01 µg/mL, respectively. The results of the recovery test were 97.19-102.04% with a RSD value of 0.65-2.47%. The developed method was subsequently applied to evaluate five batches of HLST and testified to be suitable for the quality control.

Lipophilic constituents from two processed products and three different parts of Changium smyrnioides Wolff.

This article reports the lipophilic chemical composition of different processed products (Changii Radix, Changii Radix Alba) and parts (root bark, leaf and fruit) of Changium smyrnioides Wolff.. The lipophilic constituents were extracted with petroleum ether in Soxhlet apparatus, subsequently identified and determined by gas chromatography-mass spectroscopy (GC-MS). Yield of lipophilic constituents from Changii Radix (3.65%) was about three times more than Changii Radix Alba's (1.07%), which indicated processing by boiling in water had an impact on the content of lipophilic constituents. Moreover, the major compounds in different processed products and parts were found to be fatty acids and sesquiterpenes. The results are a contribution for the lipophilic chemical composition and can serve as a reference for product development of Changium smyrnioides Wolff..

Evaluation of alfalfa inter-seeding effect on bahiagrass baleage fermentation and lactating Holstein performance.

BACKGROUND: Previous research indicates that bahiagrass may be successfully conserved as baleage, but nutritive value is typically low for lactating dairy cows. The purpose of this study was to determine the effect of adding modest amounts of alfalfa forage (22%), achieved by inter-seeding alfalfa into an existing bahiagrass pasture, on baleage nutritive value and lactation performance of Holstein cows. Forage treatments employed were monoculture bahiagrass baleage (MBB; negative control), bahiagrass-alfalfa mixture baleage (BAB) and conventional corn silage (CCS; positive control). Thirty six mid lactation Holstein cows [34.8 ± 5.8 kg 3.5% fat-corrected milk and 112 ± 19 d in milk (DIM)] were stratified according to milk yield and DIM and assigned randomly to 1 of 3 forage treatments. Cows were trained to Calan feeding gates and were offered a common CCS-based TMR in a 10-d covariance period followed by a 42-d treatment feeding period. RESULTS: The BAB contained more protein and less NDF than MBB (12.6 vs 10.3% CP and 71.8 vs 76.6% NDF). Diet DMI was similar for MBB and BAB (19.5 vs 21.6 kg/hd/d), but cows consumed more of the CCS diet (25.5 kg/hd/d) than either baleage-based diet. Cows offered BAB tended to produce more milk than cows offered MBB based TMR (28.4 vs 26.1 kg/hd/d), but both baleage diets generated less milk than CCS-based diets (33.1 kg/hd/d). Milk composition was similar across diets except for milk protein concentrations which were higher for CCS than either MBB or BAB diets; however, milk urea nitrogen (MUN) was lowest for cows fed CCS diets. Cow BW gain was higher for BAB than MBB implying that a portion of the higher energy contributed by the alfalfa was being used to replenish weight on these mid lactation cows. CONCLUSIONS: Data from this study indicate that alfalfa inter-seeded in bahiagrass sod that produces BAB with as little as 22% alfalfa may improve nutritive value compared to monoculture bahiagrass baleage and marginally improve lactation performance of Holstein cows. However, the CCS diet was vastly superior to either MBB or BAB-based diets for milk production.

Evaluation of nutritive value and in vitro rumen fermentation gas accumulation of de-oiled algal residues.

BACKGROUND: Algae are widely recognized for their high oil content and for exponentially accumulating biomass with particular potential to provide single cell protein for human consumption or animal feed. It is believed that along with biodiesel from algae, the high protein de-oiled algal residue may become an alternative feed supplement option in the future. This study was conducted to investigate de-oiled algal residue obtained from the common Chlorella species, Thalassiosira weissflogii, Selenarstrum capricornutum, Scenedesmus sp., and Scenedesmus dimorphus for assessment as potential feed supplements for ruminants by comparing with soybean (Glycine max) meal and alfalfa (Medicago sativa) hay. RESULTS: With the exception of T. weissflogii, algal residue had higher concentrations of Cu, Zn, and Mn and lower concentration of Ca, Mg, and K than soybean meal and alfalfa hay. The algal residue CP (crude protein) concentrations ranged from 140 to 445 g/kg DM and varied among the de-oiled residues. In vitro rumen fermentation gas accumulation curves indicated that algal biomass degradation potential was less than that of soybean meal or alfalfa hay by up to 41.7%. The gas production curve, interpreted with a dual pool logistic model, confirmed that the fraction sizes for fast fermenting and slow fermenting of de-oiled algal residues were smaller than those in soybean meal and alfalfa hay, and the fermenting rate of the fractions was also low. CONCLUSIONS: Inferior in vitro rumen gas accumulation from the five de-oiled algal residues suggests that these algal byproducts are less degradable in the rumen.

Xanthone-rich dichloromethane fraction of Securidaca inappendiculata, the possible antirheumatic material base with anti-inflammatory, analgesic, and immunodepressive effects.

CONTEXT: Securidaca inappendiculata Hassk. is an traditional Chinese medicine curing rheumatoid arthritis, but there is a lack of reports on material base research. OBJECTIVE: To find the active fraction of S. inappendiculata contributing the most to antirheumatic activity. MATERIALS AND METHODS: Prior to assays in vivo, mice were treated with different fractions from S. inappendiculata for 5 d at doses relative to 10, 5, and 2.5 g/kg of crude drug. Hot plate test and carrageenan-induced paw edema test were used to investigate analgesic and anti-inflammatory activities. PGE2 levels in inflammatory paws were determined by a colorimetric method. Carbon clearance test in vivo and lymphocyte transformation test in vitro were employed to assess the immune regulation activity. HPLC was used to explore the main compounds in the active fraction. RESULTS: All the fractions, especially the dichloromethane fraction (SID), alleviated inflammation. High dose of SID (112 mg/kg) inhibited paw swelling by 63.1%, and decreased PGE2 level to 38 ng/mL. The ethyl acetate fraction (SIE) and SID suppressed the carbon clearance rate (K = 0.044, 0.038 for high dose) efficiently. All fractions hindered the transformation and proliferation of lymphocyte, and prolonged the reaction time of rats in the hot plate test. The concentrations of two typical xanthones: 2-hydroxyl-1,7-dimethoxyl-xanthone and 1,7-dihydroxyl-xanthone in SID were 0.93% and 1.19%, respectively, by HPLC analysis. CONCLUSION: SID exhibited significant anti-inflammatory, analgesic, and immunodepressive effects in vivo and vitro, and deemed as the main material base for the antirheumatic activity.

Solvent dependent triphenylamine based D-(pi-A)n type dye molecules and optical properties.

D-(pi-A)n type dyes of triphenylamine derivatives were synthesized and their absorption and luminescence in different solvents were examined to investigate solvent dependent properties observed for their emissions in solvents with different dielectric constants. The emission wavelengths showed a dramatic blue shift with increasing solvent polarity. The results of molecular orbital calculations by computer simulation, based on Material Studio suite of programs, were found to reasonably account for the spectral properties. Relative levels of HOMO and LUMO were measured and calculated and all derivatives exhibited strong solid fluorescence with distinctively different FWHMs.