Tripartite motif-containing protein 11 promotes hepatocellular carcinogenesis through ubiquitin-proteasome-mediated degradation of pleckstrin homology domain leucine-rich repeats protein phosphatase 1.

BACKGROUND AND AIMS: HCC is one of the main types of primary liver cancer, with high morbidity and mortality and poor treatment effect. Tripartite motif-containing protein 11 (TRIM11) has been shown to promote tumor formation in lung cancer, breast cancer, gastric cancer, and so on. However, the specific function and mechanism of TRIM11 in HCC remain open for study. APPROACH AND RESULTS: Through clinical analysis, we found that the expression of TRIM11 was up-regulated in HCC tissues and was associated with high tumor node metastasis (TNM) stages, advanced histological grade, and poor patient survival. Then, by gain- and loss-of-function investigations, we demonstrated that TRIM11 promoted cell proliferation, migration, and invasion in vitro and tumor growth in vivo. Mechanistically, RNA sequencing and mass spectrometry analysis showed that TRIM11 interacted with pleckstrin homology domain leucine-rich repeats protein phosphatase 1 (PHLPP1) and promoted K48-linked ubiquitination degradation of PHLPP1 and thus promoted activation of the protein kinase B (AKT) signaling pathway. Moreover, overexpression of PHLPP1 blocked the promotional effect of TRIM11 on HCC function. CONCLUSIONS: Our study confirmed that TRIM11 plays an oncogenic role in HCC through the PHLPP1/AKT signaling pathway, suggesting that targeting TRIM11 may be a promising target for the treatment of HCC.

Vitamin B12-Induced Autophagy Alleviates High Glucose-Mediated Apoptosis of Islet ß Cells.

High glucose levels can lead to the apoptosis of islet ß cells, while autophagy can provide cytoprotection and promote autophagic cell death. Vitamin B12, a water-soluble B vitamin, has been shown to regulate insulin secretion and increase insulin sensitivity. However, the precise mechanism of action remains unclear. In this study, we investigated the influence of vitamin B12 on high glucose-induced apoptosis and autophagy in RIN-m5F cells to elucidate how vitamin B12 modulates insulin release. Our results demonstrate that exposure to 45 mM glucose led to a significant increase in the apoptosis rate of RIN-m5F cells. The treatment with vitamin B12 reduced the apoptosis rate and increased the number of autophagosomes. Moreover, vitamin B12 increased the ratio of microtubule-associated protein 1 light chain 3 beta to microtubule-associated protein 1 light chain 3 alpha (LC3-II/LC3-I), while decreasing the amount of sequestosome 1 (p62) and inhibiting the phosphorylation of p70 ribosomal protein S6 kinase (p70S6K) under both normal- and high-glucose conditions. The additional experiments revealed that vitamin B12 inhibited high glucose-induced apoptosis. Notably, this protective effect was attenuated when the autophagy inhibitor 3-methyladenine was introduced. Our findings suggest that vitamin B12 protects islet ß cells against apoptosis induced by high glucose levels, possibly by inducing autophagy.

Heliothis virescens ascovirus 3h blocks the cell cycle of Spodoptera exigua fat body cells at G2 /M phase by downregulating cyclin B1 and cyclin-dependent kinase 1.

Ascoviruses are double-stranded DNA viruses that are pathogenic to noctuid larvae. In vitro infection causes the cells to fail to replicate and proliferate normally. However, the molecular mechanisms are unclear. In this study, the transmission electron microscopy data of infected-Spodoptera exigua (Hübner) fat body cells (SeFB, IOZCAS-SpexII-A cells) showed that virions were internalized in phagocytic vesicles, but not in the nucleus. FACS of cell-cycle progression was performed in SeFB cells infected with Heliothis virescens ascovirus 3h (HvAV-3h). The cell cycle phase distributions of the SeFB cells were G1 = 29.52 ± 1.10%, S = 30.33 ± 1.19%, and G2 /M = 40.06 ± 0.75%. The cell culture doubling time was approximately 24 h. The G1 , S, and G2 /M phases were each approximately 8 h. The unsynchronized or synchronized cells were arrested at G2 /M phase after infection with HvAV-3h. Our data also showed that cells with more than 4N DNA content appeared in the HvAV-3h-treated group. While the mRNA levels of cyclin B1 , cyclin H, and cyclin-dependent kinase 1 (CDK1) were downregulated after HvAV-3h infection, the mRNA expression levels of cyclin A, cyclin D, and cyclin B2 were not significantly changed. Western blotting results showed that the expression of cyclin B1 and CDK1 in infected SeFB cells within 24 h postinfection (hpi), and HvAV-3h infection inhibited the expression of cyclin B1 and CDK1 at 12-24 hpi. Overall, these data implied that HvAV-3h infection leads to an accumulation of cells in the G2 /M phases by downregulating the expression of cyclin B1 and CDK1.

Phosphorylation of nucleolin is indispensable to upregulate miR-21 and inhibit apoptosis in cardiomyocytes.

Nucleolin is a multifunctional phosphoprotein and is involved in protecting from myocardial ischemia/reperfusion (I/R) injury. The function of nucleolin is regulated by posttranslational modifications, including phosphorylation and glycosylation. To study whether phosphorylation of nucleolin (P-nucleolin) was involved in the protection from myocardial I/R injury. We investigated the expression pattern of P-nucleolin (Thr-76 and 84) in hearts subjected to I/R injury, or rat cardiac myoblast cells (H9C2) subjected to hydrogen peroxide (H 2 O 2 ). The results showed that the expression of P-nucleolin and the ratio of P-nucleolin/nucleolin were significantly increased both in vivo and in vitro. Mutant nucleolin was obtained by site directed mutagenesis in vitro: threonine at 76 and 84 was replaced by alanine, and we found that the protective effect of nucleolin on apoptosis induced by oxidative stress was dependent on its phosphorylation at 76 and 84 in H9C2 cells. Furthermore, the cardio-protective roles of P-nucleolin (Thr-76 and 84) in H9C2 cardiomyocytes, were attributable to the upregulation of microRNA (miR)-21. Further analysis found that P-nucleolin (Thr-76 and 84) could bind to miR-21, and P-nucleolin colocalized with argonaute 2 (Ago2) in cytoplasm and could interact with Ago2 in a RNA-independent manner under cell oxidative stress. The current study revealed that P-nucleolin (Thr-76 and 84) increased in I/R injury myocardium, P-nucleolin was indispensable to upregulate miR-21 and inhibited apoptosis induced by H 2 O 2 in H9C2 cardiomyocytes. These findings provided new insight into the molecular mechanisms of nucleolin in myocardial I/R injury and oxidative stress cells.

Gymnemic acid I triggers mechanistic target of rapamycin-mediated ß cells cytoprotection through the promotion of autophagy under high glucose stress.

Gymnemic acid I (GA I) is a bioactive component of Gymnema sylvestre. It is an Indian traditional medicinal herb which has antidiabetic effect. However, the molecular mechanism is remaining to be elucidated. Here, we showed that high glucose promoted the rate of apoptosis, GA I decreased the apoptosis under the high glucose stress. Our further study explored that GA I increased the number of autophagosome and the ratio of light chain 3-I (LC3-I)/LC3-II in MIN-6 cells under the normal or high glucose stress by the methods of western blot analysis and immunofluorescence. It induced autophagy flux and inhibited the phosphorylation of mammalian target of rapamycin (mTOR) and ribosomal protein S6 kinase ß-1 (p70 S6K/S6K1), which is a substrate of mTOR. GA I decreased the rate of apoptosis and the activity of caspase-3 under the high glucose stress. The inhibition of apoptosis and caspase-3 activity by GA I were increased after treating with autophagy inhibitor in mouse islet ß cells MIN-6. Our data suggested that GA I-induced autophagy protected MIN-6 cells from apoptosis under high glucose stress via inhibition the phosphorylation activity of mTOR.

Rapid and visual detection of Lawsonia intracellularis with an improved recombinase polymerase amplification assay combined with a lateral flow dipstick.

BACKGROUND: Lawsonia intracellularis (L. intracellularis) is the etiologic agent of porcine proliferative enteropathy (PPE), which is reported in many swine breeding countries all over the world, and has caused enormous economic losses in intensive pig production systems. Therefore, the aim of this study was to develop a simple and rapid method for on-site detection of Lawsonia intracellularis (L. intracellularis). As the isothermal recombinase polymerase amplification (RPA) can be performed at a constant temperature and its product is directly observed on a lateral-flow dipstick (LFD) with naked eyes without electrophoresis, the RPA-LFD assay should be useful for field diagnosis of L. intracellularis as well as its detection from clinical samples. RESULTS: The established RPA-LFD assay could be finished in 30 min at a wide temperature range of 25 to 40 °C, and the amplicons could be visualized by naked eyes. The developed RPA-LFD assay was specific to dnaA gene of L. intracellularis, and did not detect nucleic acids extracted from other common gastrointestinal pathogens. The minimum detection of this RPA-LFD method was 400 L. intracellularis per reaction, which was as sensitive as conventional PCR. Further, the RPA-LFD assay was performed with 150 clinical fecal samples and the detection results were compared with conventional PCR. Results showed that the coincidence rate of RPA-LFD and conventional PCR was 100%. CONCLUSIONS: The combined RPA with LFD assay provides a simple, rapid, specific and sensitive alternative for field diagnosis of L. intracellularis infection.

Mogroside V inhibits LPS-induced COX-2 expression/ROS production and overexpression of HO-1 by blocking phosphorylation of AKT1 in RAW264.7 cells.

Momordica grosvenori is a valuable edible plant with medicinal purposes, and it is widely used in medicated diets and traditional Chinese medicine in Asia. Mogroside V (MV), the main bioactive component from M. grosvenori, is commonly used as a natural sweetener. M. grosvenori extracts have been reported to exert potent anti-inflammatory property, however the underlying molecular mechanism still remains unknown. In the present study, the biological effect of MV in inflammation was investigated in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. The ELISA and western blot analysis results showed that MV significantly inhibited LPS-induced prostaglandin E2 (PGE2) production and cyclooxygenase-2 (COX-2) expression. MV markedly decreased the phosphorylation of IκB-α, increased IκB-α, and reduced nuclear p-65 and C/EBPδ. Furthermore, MV attenuated LPS-induced phosphorylation of MAPKs and AKT1, and only the phosphorylation status of AKT1 was found to be consistent with the expression trend of COX-2. Moreover, MV reduced ROS level and restored overexpressed HO-1 and AP-1 to basal level, which can be markedly reversed by AKT1 inhibitor LY294002. These results revealed that AKT1 plays a key role in LPS-induced COX-2 expression, and acts as a mediator to keep the redox balance in LPS-stimulated RAW264.7 cells. MV exerts anti-inflammatory property by blocking AKT1-mediated NF-κB and C/EBPδ activation, ROS generation and AP-1/ HO-1 expression. Therefore, the present study indicated that MV has a significant chemopreventive effect on the inflammatory lesions and suggested that AKT1 is a potential specific target of MV for relieving inflammation.

Delphinidin-induced autophagy protects pancreatic ß cells against apoptosis resulting from high-glucose stress via AMPK signaling pathway.

Hyperglycemia, a diagnostic characteristic of diabetes mellitus, is detrimental to pancreatic ß cells. Delphinidin, a member of the anthocyanin family, inhibits glucose absorption, increases glucagon-like peptide-1 (GLP-1) secretion, and improves insulin secretion in diabetes. However, whether delphinidin plays a protective role in pancreatic ß-cell mass and function is not clear. In this study, delphinidin was found to decrease the high-glucose-induced apoptosis of RIN-m5F pancreatic ß cells. In addition, delphinidin induced autophagy in RIN-m5F cells under the normal and high-glucose conditions, while 3-methyladenine (3-MA) inhibition of autophagy significantly diminished the protective role of delphinidin against high-glucose-induced apoptosis of pancreatic ß cells. Delphinidin also decreased the level of cleaved caspase 3 and increased the phosphorylation level of AMP-activated protein kinase α (AMPKα) Thr172. Compound C, an AMPK inhibitor, was found to decrease the ratio of LC3-II/LC3-I, and the apoptotic rate of high-glucose-injured cells was increased after treatment with delphinidin, indicating that delphinidin attenuated the negative effects of high-glucose stress to cells. In conclusion, our data demonstrate that delphinidin protects pancreatic ß cells against high-glucose-induced injury by autophagy regulation via the AMPK signaling pathway. These findings might shed light on the underlying mechanisms of diabetes and help improve the prevention and therapy of this common disease.

Nucleolin protects against doxorubicin-induced cardiotoxicity via upregulating microRNA-21.

Nucleolin is a multifunctional protein and participates in many important biological processes. Our previous study found that nucleolin protects the heart against myocardial ischemia-reperfusion injury. In this study, we aimed to investigate the role of nucleolin in doxorubicin (DOX)-induced cardiotoxicity. The expression pattern of nucleolin in hearts subjected to DOX injury was investigated, and we found that administration of DOX induced nucleolin expression significantly in vivo and in vitro. Gene transfection and RNA interference approaches were used in cardiomyocytes to investigate the function of nucleolin. Nucleolin overexpression protects cardiomyocytes against DOX-induced injury. Nucleolin-ablated cardiomyocytes become susceptible to the injury induced by DOX. The hearts of cardiac-myocyte-specific nucleolin transgenic mice are more resistant to DOX injury. Furthermore, nucleolin upregulates microRNA(miRNA)-21 expression in vivo and in vitro, and the miRNA-21 inhibitor negates the protective effect of nucleolin against injury induced by DOX. These results have demonstrated that nucleolin is involved in the regulation of DOX-induced cardiac injury and dysfunction via the regulation of miRNA-21 expression, and may be a novel therapeutic target for DOX-induced cardiotoxicity.

Organosulfur compounds induce cytoprotective autophagy against apoptosis by inhibiting mTOR phosphorylation activity in macrophages.

Organosulfur compounds (OSCs) are the bioactive components of garlic. Some OSCs have apoptotic or autophagy-inducing effects. Autophagy plays roles in both cytoprotection and apoptosis-related cell death, and the interaction between autophagy and apoptosis is important in the modulation of immune responses. The mechanism of an OSC-mediated effect via the interaction of autophagy and apoptosis is unknown. In this study, the effects of five OSC compounds on autophagy in the macrophage cell line RAW264.7 and primary macrophages were investigated. We found that S-allylcysteine (SAC), diallyl disulde (DADS) and diallyl tetrasulfide (DTS) treatment increased the number of autophagosomes of RAW264.7 cells, inhibited the phosphorylation of ribosomal protein S6 kinase beta-1 (p70S6K/S6K1) which is a substrate of mammalian target of rapamycin (mTOR), and significantly enhanced autophagy flux. The induction of autophagy by SAC, DADS and DTS was inhibited by stably knocking down the expression of autophagy-related gene 5 (ATG5) with short hairpin RNA (shRNA). Further experiments confirmed that SAC, DADS and DTS also induced apoptosis in RAW264.7 cells. The induction of apoptosis and Caspase 3 activity by SAC, DADS and DTS were increased by stably knocking down of ATG5 expression with shRNA in RAW264.7 cells or treating with 5 mM 3-MA in primary macrophages. Our results suggest that SAC, DADS and DTS induce both autophagy and apoptosis. The autophagy induction protects macrophages from apoptosis by inhibiting mTOR phosphorylation activity to maintain the mass of immune cells.

MiR-21 Protected Cardiomyocytes against Doxorubicin-Induced Apoptosis by Targeting BTG2.

Doxorubicin (DOX) is an anthracycline drug with a wide spectrum of antineoplastic activities. However, it causes cardiac cytotoxicity, and this limits its clinical applications. MicroRNA-21 (miR-21) plays a vital role in regulating cell proliferation and apoptosis. While miR-21 is preferentially expressed in adult cardiomyocytes and involved in cardiac development and heart disease, little is known regarding its biological functions in responding to DOX-induced cardiac cytotoxicity. In this study, the effects of DOX on mouse cardiac function and the expression of miR-21 were examined in both mouse heart tissues and rat H9C2 cardiomyocytes. The results showed that the cardiac functions were more aggravated in chronic DOX injury mice compared with acute DOX-injury mice; DOX treatment significantly increased miR-21 expression in both mouse heart tissue and H9C2 cells. Over-expression of miR-21 attenuated DOX-induced apoptosis in cardiamyocytes whereas knocking down its expression increased DOX-induced apoptosis. These gain- and loss- of function experiments showed that B cell translocation gene 2 (BTG2) was a target of miR-21. The expression of BTG2 was significantly decreased both in myocardium and H9C2 cells treated with DOX. The present study has revealed that miR-21 protects mouse myocardium and H9C2 cells against DOX-induced cardiotoxicity probably by targeting BTG2.

Dietary supplementation with quercetin: an ideal approach for improving meat quality and oxidative stability of broiler chickens.

This study aimed to improve the eating quality of yellow-feathered broiler chicks by feeding them corn-soybean meal diets supplemented with 250, 500, and 1,000 mg/kg quercetin. we examined the impact of varying doses of dietary quercetin on the sensory quality of chicken breast meat as well as on the antioxidant enzymes, antioxidant-related signaling molecules, structure and thermal stability of myofibrillar protein (MPs), and microstructure of myogenic fibers in the meat during 24 h of postslaughter aging. Additionally, we investigated the potential correlations among antioxidant capacity, MP structure, and meat quality parameters. The results indicated that dietary supplementations with 500 and 1,000 mg/kg quercetin improved the physicochemical properties and eating quality of yellow-feathered broiler chicken breast meat during 12 to 24 h postslaughter. Additionally, quercetin improved the postslaughter oxidative stress status and reduced protein and lipid oxidation levels. It also increased hydrogen bonding interactions and α-helix content during 6 to 12 h postslaughter and decreased ß-sheet content during 12 to 24 h postslaughter in chicken breast MP. This resulted in improved postslaughter MP structure and thermal stability. The correlation results indicated that the enhancement of antioxidant capacity and MP structure enhanced the physicochemical and edible qualities of yellow-feathered broiler chicken breast meat. In conclusion, the current findings suggest that dietary supplementation with quercetin is an ideal approach for improving the eating quality of chicken meat, thereby broadening our understanding of theoretical and technological applications for improving the quality of chicken.

Probiotic Characteristics and Anti-Inflammatory Effects of Limosilactobacillus fermentum 664 Isolated from Chinese Fermented Pickles.

Limosilactobacillus fermentum (L. fermentum) is widely used in industrial food fermentations, and its probiotic and health-promoting roles attracted much attention in the past decades. In this work, the probiotic potential of L. fermentum 664 isolated from Chinese fermented pickles was assessed. In addition, the anti-inflammatory properties and mechanisms were investigated using lipopolysaccharide (LPS)-stimulated RAW264.7 cells. Results indicated that L. fermentum 664 demonstrated excellent acid and bile salt tolerance, adhesion capability, antimicrobial activity, and safety profile. L. fermentum 664 downregulated the release of inflammatory mediators, including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1ß (IL-1ß), and cyclooxygenase-2 (COX-2) stimulated with LPS. Moreover, L fermentum 664 inhibited the nuclear translocation of the nuclear factor κB (NF-κB) and the activation of mitogen-activated protein kinases (MAPKs) induced by LPS. This action was associated with a reduction in reactive oxygen species (ROS) levels and an enhanced expression of heme oxygenase-1 (HO-1) protein. Additionally, whole genome sequencing indicated that L. fermentum 664 contained genes that encode proteins with antioxidant and anti-inflammatory functions, including Cytochrome bd ubiquinol oxidase subunit I (CydA), Cytochrome bd ubiquinol oxidase subunit II (CydB), and NAD(P)H dehydrogenase quinone 1 (NQO1). In conclusion, our study suggested that L. fermentum 664 has the potential to become a probiotic and might be a promising strategy for the prevention of inflammation.

A Novel Method in Identifying Pyroptosis and Apoptosis Based on the Double Resonator Piezoelectric Cytometry Technology.

In this study, a double resonator piezoelectric cytometry (DRPC) technology based on quartz crystal microbalance (QCM) was first employed to identify HeLa cell pyroptosis and apoptosis by monitoring cells' mechanical properties in a real-time and non-invasive manner. AT and BT cut quartz crystals with the same frequency and surface conditions were used concurrently to quantify the cells-exerted surface stress (ΔS). It is the first time that cells-exerted surface stress (ΔS) and cell viscoelasticity have been monitored simultaneously during pyroptosis and apoptosis. The results showed that HeLa pyroptotic cells exerted a tensile stress on quartz crystal along with an increase in the elastic modulus (G'), viscous modulus (G″), and a decrease of the loss tangent (G″/G'), whereas apoptotic cells exerted increasing compressive stress on quartz crystal along with a decrease in G', G″ and an increase in G″/G'. Furthermore, engineered GSDMD-/--DEVD- HeLa cells were used to investigate drug-induced disturbance and testify the mechanical responses during the processes of pyroptosis and non-pyroptosis. These findings demonstrated that the DRPC technology can serve as a precise cytomechanical sensor capable of identifying pyroptosis and apoptosis, providing a novel method in cell death detection and paving the road for pyroptosis and apoptosis related drug evaluation and screening.

Expression and correlation of COX-2 and NUCB1 in colorectal adenocarcinoma.

Objective: To investigate the expression and correlation of COX-2 and NUCB1 in colorectal adenocarcinoma and adjacent tissues. Methods: The expression of COX-2 and NUCB1 and their effects on prognosis were predicted using bioinformatics. Immunohistochemistry was used to identify the expression of two molecules in 56 cases of colorectal adenocarcinoma and the surrounding tissues. The expression of two molecules and their association with clinicopathological variables were examined using the chi-square test. The association between COX-2 and NUCB1 was investigated using the Spearman correlation test. Results: The STRING database revealed that COX-2 and NUCB1 were strongly linked. According to the UALCAN and HPA database, COX-2 was upregulated while NUCB1 was downregulated in colorectal adenocarcinoma, both at the protein and gene levels. The OS times for COX-2 and NUCB1 high expression, however, exhibited the same patterns. The rate of positive COX-2 immunohistochemical staining in cancer tissues was 69.64% (39/56), which was significantly higher than the rate in healthy tissues 28.57% (16/56). NUCB1 was expressed positively in cancer tissues at a rate of 64.29% (36/56) compared to just 19.64% (11/56) in neighboring tissues. The positive expression levels of COX-2 and NUCB1 were both closely related to clinical stage, differentiation degree, and lymphatic metastases (P < 0.05). In colorectal cancer, COX-2 and NUCB1 expression were significantly correlated (rs = 0.6312, P < 0.001). Conclusion: Both COX-2 and NUCB1 are overexpressed and significantly associated in colorectal adenocarcinoma.

Separation of m-cresol and p-cresol by NaZSM-5 with different Si/Al ratios.

m-Cresol and p-cresol are widely used in medicine and pesticides as important chemical intermediates. They are generally produced as a mixture in industry and are difficult to separate due to the similarities in both chemical structures and physical properties. The adsorption behaviours of m-cresol and p-cresol on zeolites (NaZSM-5 and HZSM-5) with different Si/Al ratios have been compared by static experiments. Selectivity on NaZSM-5(Si/Al = 80) could be greater than 6.0. Adsorption kinetics and isotherms were investigated in detail. The kinetic data was correlated by PFO, PSO, and ID models, the NRMSE of which were 14.03%, 9.41%, and 21.11%, respectively. In the meanwhile, according to the NRMSE of Langmuir (6.01%), Freundlich (57.80%), D-R (1.1%), and Temkin (0.56%) isotherms, adsorption on NaZSM-5(Si/Al = 80) was mainly a monolayer and chemical adsorption process. It was endothermic for m-cresol and exothermic for p-cresol. The Gibbs free energy, entropy, and enthalpy were calculated accordingly. The adsorption of cresol isomers on NaZSM-5(Si/Al = 80) were both spontaneous, and it was exothermic (ΔH = -37.11kJ/mol) for p-cresol while endothermic (ΔH = 52.30kJ/mol) for m-cresol. Additionally, ΔS were respectively -0.05 and 0.20 kJ·mol-1·K-1for p-cresol and m-cresol, which were both close to zero. The adsorption was mainly driven by enthalpy. The result of breakthrough further demonstrated m-cresol and p-cresol could be separated effectively by NaZSM-5(Si/Al = 80). Additionally, the selectivity increased from 7.53 to 14.72 after four cycles regeneration with 9.95% and 53.96% decreases in the adsorption amounts for m-cresol and p-cresol, respectively. In conclusion, NaZSM-5(Si/Al = 80) could be a feasible adsorbent for the separation of m-cresol and p-cresol.

Carbon Quantum Dots from Roasted Coffee Beans: Their Degree and Mechanism of Cytotoxicity and Their Rapid Removal Using a Pulsed Electric Field.

Carbon quantum dots (CQDs) from heat-treated foods show toxicity, but the mechanisms of toxicity and removal of CQDs have not been elucidated. In this study, CQDs were purified from roasted coffee beans through a process of concentration, dialysis and lyophilization. The physical properties of CQDs, the degree and mechanism of toxicity and the removal method were studied. Our results showed that the size of CQDs roasted for 5 min, 10 min and 20 min were about 5.69 ± 1.10 nm, 2.44 ± 1.08 nm and 1.58 ± 0.48 nm, respectively. The rate of apoptosis increased with increasing roasting time and concentration of CQDs. The longer the roasting time of coffee beans, the greater the toxicity of CQDs. However, the caspase inhibitor Z-VAD-FMK was not able to inhibit CQDs-induced apoptosis. Moreover, CQDs affected the pH value of lysosomes, causing the accumulation of RIPK1 and RIPK3 in lysosomes. Treatment of coffee beans with a pulsed electric field (PEF) significantly reduced the yield of CQDs. This indicates that CQDs induced lysosomal-dependent cell death and increased the rate of cell death through necroptosis. PEF is an effective way to remove CQDs from roasted coffee beans.

Vitamin B6 Inhibits High Glucose-Induced Islet ß Cell Apoptosis by Upregulating Autophagy.

Vitamin B6 may alleviate diabetes by regulating insulin secretion and increasing insulin sensitivity, but its mechanism remains to be explored. In this study, vitamin B6-mediated autophagy and high glucose-induced apoptosis were tested to investigate the mechanism by which vitamin B6 regulates insulin release. The results showed that 20 mM glucose increased the apoptosis rate from 10.39% to 22.44%. Vitamin B6 reduced the apoptosis rate of RIN-m5F cells from 22.44% to 11.31%. Our data also showed that the vitamin B6 content in processed eggs was decreased and that the hydrothermal process did not affect the bioactivity of vitamin B6. Vitamin B6 increased the number of autophagosomes and the ratio of autophagosome marker protein microtubule associated protein 1 light chain 3 beta to microtubule associated protein 1 light chain 3 alpha (LC3-II/LC3-I). It also decreased the amount of sequetosome 1 (SQSTM1/p62) and inhibited the phosphorylation of p70 ribosomal protein S6 kinase (p70S6K) under normal and high glucose stress. Another study showed that vitamin B6 inhibited the apoptosis rate, whereas the autophagy inhibitor 3-methyladenine (3-MA) blocked the protective effect of vitamin B6 against apoptosis induced by high glucose. The hydrothermal process decreased the vitamin B6 content in eggs but had no effect on the cytoprotective function of vitamin B6 in RIN-m5f cells. In conclusion, we demonstrated that vitamin B6-mediated autophagy protected RIN-m5f cells from high glucose-induced apoptosis might via the mTOR-dependent pathway. Our data also suggest that low temperatures and short-term hydrothermal processes are beneficial for dietary eggs.

Lactate: The Mediator of Metabolism and Immunosuppression.

The Warburg effect, one of the hallmarks of tumors, produces large amounts of lactate and generates an acidic tumor microenvironment via using glucose for glycolysis. As a metabolite, lactate not only serves as a substrate to provide energy for supporting cell growth and development but also acts as an important signal molecule to affect the biochemical functions of intracellular proteins and regulate the biological functions of different kinds of cells. Notably, histone lysine lactylation (Kla) is identified as a novel post-modification and carcinogenic signal, which provides the promising and potential therapeutic targets for tumors. Therefore, the metabolism and functional mechanism of lactate are becoming one of the hot fields in tumor research. Here, we review the production of lactate and its regulation on immunosuppressive cells, as well as the important role of Kla in hepatocellular carcinoma. Lactate and Kla supplement the knowledge gap in oncology and pave the way for exploring the mechanism of oncogenesis and therapeutic targets. Research is still needed in this field.

Tumor microenvironment-associated lactate metabolism regulates the prognosis and precise checkpoint immunotherapy outcomes of patients with lung adenocarcinoma.

BACKGROUND: Despite the wide clinical application of checkpoint inhibitor immunotherapy in lung adenocarcinoma, its limited benefit to patients remains puzzling to researchers. One of the mechanisms of immunotherapy resistance may be the dysregulation of lactate metabolism in the immunosuppressive tumor microenvironment (TME), which can inhibit dendritic cell maturation and prevent T-cell invasion into tumors. However, the key genes related to lactate metabolism and their influence on the immunotherapeutic effects in lung adenocarcinoma have not yet been investigated in depth. METHODS: In this study, we first surveyed the dysregulated expression of genes related to lactate metabolism in lung adenocarcinoma and then characterized their biological functions. Using machine learning methods, we constructed a lactate-associated gene signature in The Cancer Genome Atlas cohort and validated its effectiveness in predicting the prognosis and immunotherapy outcomes of patients in the Gene Expression Omnibus cohorts. RESULTS: A 7-gene signature based on the metabolomics related to lactate metabolism was found to be associated with multiple important clinical features of cancer and was an independent prognostic factor. CONCLUSIONS: These results suggest that rather than being simply a metabolic byproduct of glycolysis, lactate in the TME can affect immunotherapy outcomes. Therefore, the mechanism underlying this effect of lactate is worthy of further study.