Angiotensin-converting enzyme 2 improves liver fibrosis in mice by regulating autophagy of hepatic stellate cells.

BACKGROUND: Liver fibrosis is the common pathological process associated with the occurrence and development of various chronic liver diseases. At present, there is still a lack of effective prevention and treatment methods in clinical practice. Hepatic stellate cell (HSC) plays a key role in liver fibrogenesis. In recent years, the study of liver fibrosis targeting HSC autophagy has become a hot spot in this research field. Angiotensin-converting enzyme 2 (ACE2) is a key negative regulator of renin-angiotensin system, and its specific molecular mechanism on autophagy and liver fibrosis needs to be further explored. AIM: To investigate the effect of ACE2 on hepatic fibrosis in mice by regulating HSC autophagy through the Adenosine monophosphate activates protein kinases (AMPK)/mammalian target of rapamycin (mTOR) pathway. METHODS: Overexpression of ACE2 in a mouse liver fibrosis model was induced by injection of liver-specific recombinant adeno-associated virus ACE2 vector (rAAV2/8-ACE2). The degree of liver fibrosis was assessed by histopathological staining and the biomarkers in mouse serum were measured by Luminex multifactor analysis. The number of apoptotic HSCs was assessed by terminal deoxynucleoitidyl transferase-mediated dUTP nick-end labeling (TUNEL) and immunofluorescence staining. Transmission electron microscopy was used to identify the changes in the number of HSC autophagosomes. The effect of ACE2 overexpression on autophagy-related proteins was evaluated by multicolor immunofluorescence staining. The expression of autophagy-related indicators and AMPK pathway-related proteins was measured by western blotting. RESULTS: A mouse model of liver fibrosis was successfully established after 8 wk of intraperitoneal injection of carbon tetrachloride (CCl4). rAAV2/8-ACE2 administration reduced collagen deposition and alleviated the degree of liver fibrosis in mice. The serum levels of platelet-derived growth factor, angiopoietin-2, vascular endothelial growth factor and angiotensin II were decreased, while the levels of interleukin (IL)-10 and angiotensin- (1-7) were increased in the rAAV2/8-ACE2 group. In addition, the expression of alpha-smooth muscle actin, fibronectin, and CD31 was down-regulated in the rAAV2/8-ACE2 group. TUNEL and immunofluorescence staining showed that rAAV2/8-ACE2 injection increased HSC apoptosis. Moreover, rAAV2/8-ACE2 injection notably decreased the number of autophagosomes and the expression of autophagy-related proteins (LC3I, LC3II, Beclin-1), and affected the expression of AMPK pathway-related proteins (AMPK, p-AMPK, p-mTOR). CONCLUSION: ACE2 overexpression can inhibit HSC activation and promote cell apoptosis by regulating HSC autophagy through the AMPK/mTOR pathway, thereby alleviating liver fibrosis and hepatic sinusoidal remodeling.

Combining transcriptomic analysis and network pharmacology to explore the mechanism by which Shaofu Zhuyu decoction improves diabetes mellitus erectile dysfunction.

BACKGROUND: Erectile dysfunction is common among the complications of diabetes mellitus. Shaofu Zhuyu decoction (SFZYD) is commonly used to treat diabetic mellitus erectile dysfunction (DMED). However, its main active components and specific mechanism are still unknown. PURPOSE: To confirm the activity of SFZYD in improving DMED, explore the main active components of SFZYD, and clarify the underlying mechanism. METHODS: A diabetic rat model was induced with streptozotocin (STZ). After intragastric administration, erectile function was assessed by the maximum intracavernous pressure (ICPmax)/mean arterial pressure (MAP). Corpus cavernosum fibrosis was evaluated by Masson staining, and ELISA methods were used to determine the serum levels of IL-6, TNF-α, IL-10, IL-4 and IL-1ß to evaluate inflammation. Then, the main active components of SFZYD were identified by UPLC‒MS/MS. Finally, the target and biological mechanism of SFZYD in improving DMED were predicted by combined network pharmacology and transcriptomics, which was also validated by molecular docking and cellular thermal shift assay (CETSA) experiments. RESULTS: SFZYD significantly improved erectile dysfunction and inhibited inflammatory responses and local tissue fibrosis in diabetic rats. A total of 1846 active components were identified by UPLC‒MS/MS, and isorhamnetin was the main active component. The transcriptomic results were used to identify differentially expressed genes among the control, DM and SFZYD groups, and 1264 differentially expressed genes were obtained from the intersection. The network pharmacology results showed that SFZYD acts on core targets such as AKT1, ALB, HSP90AA1 and ESR1 through core components such as isorhamnetin, quercetin and chrysophanic acid. Further combined analysis revealed that multiple targets, such as CYP1B1, DPP4, NOS2 and LCN2, as well as the regulation of the PI3K-AKT signaling pathway, may be important mechanisms by which SFZYD improves DMED. Molecular docking verification showed that isorhamnetin, the key component of SFZYD, has good binding ability with several core targets, and its binding ability with CYP1B1 was the strongest. The CETSA results showed that isorhamnetin binds to CYP1B1 in CCECs. CONCLUSION: SFZYD improves DMED, inhibits the inflammatory response and alleviates local tissue fibrosis. The combined application of transcriptomic, network pharmacology, molecular docking and CETSA approaches was helpful for revealing the mechanism by which SFZYD improves DMED, which may be related to the regulation of CYP1B1 and the PI3K-Akt signaling pathway.

Detection of Soybean-Derived Components in Dairy Products Using Proofreading Enzyme-Mediated Probe Cleavage Coupled with Ladder-Shape Melting Temperature Isothermal Amplification (Proofman-LMTIA).

Food adulteration is a serious problem all over the world. Establishing an accurate, sensitive and fast detection method is an important part of identifying food adulteration. Herein, a sequence-specific ladder-shape melting temperature isothermal amplification (LMTIA) assay was reported to detect soybean-derived components using proofreading enzyme-mediated probe cleavage (named Proofman), which could realize real-time and visual detection without uncapping. The results showed that, under the optimal temperature of 57 °C, the established Proofman-LMTIA method for the detection of soybean-derived components in dairy products was sensitive to 1 pg/µL, with strong specificity, and could distinguish soybean genes from those of beef, mutton, sunflower, corn, walnut, etc. The established Proofman-LMTIA detection method was applied to the detection of actual samples of cow milk and goat milk. The results showed that the method was accurate, stable and reliable, and the detection results were not affected by a complex matrix without false positives or false negatives. It was proved that the method could be used for the detection and identification of soybean-derived components in actual dairy products samples.

A Sterically Tuned Directing Auxiliary Promotes Catalytic 1,2-Carbofluorination of Alkenyl Carbonyl Compounds.

The site-selective palladium-catalyzed three-component coupling of unactivated alkenyl carbonyl compounds, aryl- or alkenylboronic acids, and N-fluorobenzenesulfonimide is described herein. Tuning of the steric environment on the bidentate directing auxiliary enhances regioselectivity and facilitates challenging C(sp3 )-F reductive elimination from a PdIV intermediate to afford 1,2-carbofluorination products in moderate to good yields.

Effects of acceleration on skill level in skiers / Efeitos da aceleração sobre o nível de habilidade em esquiadores / Efectos de la aceleración en el nivel de habilidad de los esquiadores

ABSTRACT Introduction: In a "U" shaped field, the skier must take off using snow slides in a determined territory, using the distribution of his body mass over his feet to control the direction, completing a variety of complex movements inherent to aerial sports. Objective: Study the effect of acceleration on skiers' skill levels. Methods: Peak moment angle and range of motion were measured in 12 U-shaped cross-country snow ski athletes with an average training period of over 5 years. The isokinetic muscle strength test parameters were analyzed with the characteristics and abilities inherent to the sport. The corresponding range of motion and the angle of acceleration were determined. The results were confronted with a literature search and discussed afterward. results: The peak torque and relative peak torque of the flexor and extensor muscle groups on both sides of the athletes' hip and knee joints reduced with increasing angular velocity; the maximum peak torque of the extensors was significantly greater than that of the flexors at the same velocity (P﹤0.05). The current scientific literature predicted this inversely proportional relationship, but the angles obtained had not been explored. Conclusion: Athletes should pay attention to improving cognitive ability and active adjustment ability to complete their technical movements, promoting the development of controlled acceleration required by snow sports. Level of evidence II; Therapeutic studies - investigation of treatment outcomes.

RESUMO Introdução: Em campo em forma de "U", o esquiador deve decolar utilizando deslizamentos de neve em um território determinado, utilizando a distribuição de sua massa corporal sobre os pés para controlar a direção, completando uma variedade de movimentos complexos inerentes aos esportes aéreos. Objetivo: Estudar o efeito da aceleração sobre o nível de habilidade dos esquiadores. Métodos: Aferiu-se o ângulo do momento de pico e a amplitude de movimento em 12 atletas de esqui na neve em campos em forma de "U" com um período médio de treinamento de mais de 5 anos. Foram analisados os parâmetros do teste de força muscular isocinético combinados com as características e habilidades inerentes ao esporte. Determinou-se a gama correspondente do movimento e o ângulo de aceleração. Os resultados foram confrontados com uma pesquisa bibliográfica e discutidos posteriormente. Resultados: O torque de pico e o torque de pico relativo dos grupos de músculos flexores e extensores em ambos os lados das articulações do quadril e joelho dos atletas reduziram com o aumento da velocidade angular; o torque de pico máximo dos extensores foi significativamente maior do que o dos flexores na mesma velocidade (P﹤0.05). Essa relação inversamente proporcional foi prevista pela literatura científica vigente, porém os ângulos obtidos não haviam sido explorados. Conclusão: Os atletas devem atentar-se em melhorar a capacidade cognitiva e a capacidade de ajuste ativa para completar seus movimentos técnicos promovendo o desenvolvimento da aceleração controlada requerida pelos esportes na neve. Nível de evidência II; Estudos terapêuticos - investigação dos resultados do tratamento.

RESUMEN Introducción: En un campo en forma de "U", el esquiador debe despegar utilizando deslizamientos de nieve en un territorio determinado, utilizando la distribución de su masa corporal sobre los pies para controlar la dirección, completando una variedad de movimientos complejos inherentes a los deportes aéreos. Objetivo: Estudiar el efecto de la aceleración en el nivel de habilidad de los esquiadores. Métodos: Se midieron el ángulo de momento máximo y la amplitud de movimiento en 12 atletas de esquí de fondo en forma de U con un período de entrenamiento medio de más de 5 años. Se analizaron los parámetros de la prueba de fuerza muscular isocinética combinados con las características y habilidades propias del deporte. Se determinó la correspondiente amplitud de movimiento y el ángulo de aceleración. Los resultados se confrontaron con una búsqueda bibliográfica y se debatieron posteriormente. Resultados: El par máximo y el par máximo relativo de los grupos musculares flexores y extensores de ambos lados de las articulaciones de la cadera y la rodilla de los atletas se redujo con el aumento de la velocidad angular; el par máximo de los extensores fue significativamente mayor que el de los flexores a la misma velocidad (P﹤0,05). Esta relación inversamente proporcional fue predicha por la literatura científica actual, pero los ángulos obtenidos no habían sido explorados. Conclusión: Los atletas deben prestar atención a la mejora de la capacidad cognitiva y la capacidad de ajuste activo para completar sus movimientos técnicos promoviendo el desarrollo de la aceleración controlada que requieren los deportes de nieve. Nivel de evidencia II; Estudios terapéuticos - investigación de los resultados del tratamiento.

Regio- and Stereoselective 1,2-Oxyhalogenation of Non-Conjugated Alkynes via Directed Nucleopalladation: Catalytic Access to Tetrasubstituted Alkenes.

A catalytic 1,2-oxyhalogenation method that converts non-conjugated internal alkynes into tetrasubstituted alkenes with high regio- and stereoselectivity is described. Mechanistically, the reaction involves a PdII /PdIV catalytic cycle that begins with a directed oxypalladation step. The origin of regioselectivity is the preference for formation of a six-membered palladacycle intermediate, which is facilitated by an N,N-bidentate 2-(pyridin-2-yl)isopropyl (PIP) amide directing group. Selectivity for C(alkenyl)-X versus -N (X=halide) reductive elimination from the PdIV center depends on the identity of the halide anion; bromide and iodide engage in C(alkenyl)-X formation, while intramolecular C(alkenyl)-N reductive elimination occurs with chloride to furnish a lactam product. DFT calculations shed light on the origins of this phenomenon.

Activated platelet membrane nanovesicles recruit neutrophils to exert the antitumor efficiency.

Platelets play a crucial role in the recruitment of neutrophils, mediated by P-selectin, CCL5, and ICAM-2. In this study, we prepared platelet membrane nanovesicles from activated platelets. Whether activated platelet membrane nanovesicles can recruit neutrophils has not been reported, nor has their role in antitumor immunity. The results of SDS-PAGE showed that the platelet membrane nanovesicles retained almost all the proteins of platelets. Western blotting showed that both the activated platelets and the platelet membrane nanovesicles expressed P-selectin, ICAM-2, and CCL5. In vivo results of a mouse model of breast cancer-transplanted tumor showed that tumor volume reduced significantly, Ki-67-positive tumor cells decreased, and TUNEL-positive tumor cells increased in tumors after treatment with activated platelet membrane nanovesicles (aPNs). After treatment with aPNs, not only the number of neutrophils, CD8+, CD4+ T cells, and B cells increased, but also IL-12, TNF-α, and IFN-γ levels elevated significantly in tumor tissues.

Development of a Ladder-Shape Melting Temperature Isothermal Amplification (LMTIA) Assay for the Identification of Cassava Component in Sweet Potato Starch Noodles.

Food authenticity has become increasingly important as a result of food adulteration. To identify the authenticity of sweet potato starch noodles, the ladder-shape melting temperature isothermal amplification (LMTIA) method of determining cassava (Manihot esculenta Crantz) DNA in sweet potato starch noodles was used. A set of primers targeted at the internal transcription spacer (ITS) of cassava was designed, genomic DNA was extracted, the LMTIA reaction temperature was optimized, and the specificity of the primer was verified with the genomic DNAs of cassava, sweet potato (Ipomoea batatas L.), Solanum tuberosum L., Zea mays L., Vigna radiate L., Triticum aestivum L., and Glycine max (L.) Merr. The sensitivity with the serially diluted genomic DNA of cassava and the suitability for the DNA extracted from sweet potato starch adulterated with cassava starch were tested. The LMTIA assay for identifying the cassava component in sweet potato starch noodles was established. At the optimal temperature of 52 °C, the primers could specifically distinguish a 0.01% (w/w) cassava component added to sweet potato starch. Additionally, the LMTIA method was applied to the cassava DNA detection of 31 sweet potato starch noodle samples purchased from retail markets in China. Of these, 14 samples were positive. The LMTIA assay could be a reliable method for the rapid detection of cassava components in sweet potato starch noodles, to protect the rights of consumers and to regulate the sale market order of starch noodles.

PdII -Catalyzed C(alkenyl)-H Activation Facilitated by a Transient Directing Group.

Palladium(II)-catalyzed C(alkenyl)-H alkenylation enabled by a transient directing group (TDG) strategy is described. The dual catalytic process takes advantage of reversible condensation between an alkenyl aldehyde substrate and an amino acid TDG to facilitate coordination of the metal catalyst and subsequent C(alkenyl)-H activation by a tailored carboxylate base. The resulting palladacycle then engages an acceptor alkene, furnishing a 1,3-diene with high regio- and E/Z-selectivity. The reaction enables the synthesis of enantioenriched atropoisomeric 2-aryl-substituted 1,3-dienes, which have seldom been examined in previous literature. Catalytically relevant alkenyl palladacycles were synthesized and characterized by X-ray crystallography, and the energy profiles of the C(alkenyl)-H activation step and the stereoinduction model were elucidated by density functional theory (DFT) calculations.

A high-quality genome assembly of Jasminum sambac provides insight into floral trait formation and Oleaceae genome evolution.

As one of the most economically significant Oleaceae family members, Jasminum sambac is renowned for its distinct sweet, heady fragrance. Using Illumina reads, Nanopore long reads, and HiC-sequencing, we efficiently assembled and annotated the J. sambac genome. The high-quality genome assembly consisted of a total of 507 Mb sequence (contig N50 = 17.6 Mb) with 13 pseudomolecules. A total of 21,143 protein-coding genes and 303 Mb repeat sequences were predicted. An ancient whole-genome triplication event at the base of Oleaceae (~66 million years ago [Ma], Late Cretaceous) was identified and this may have contributed to the diversification of the Oleaceae ancestor and its divergence from the Lamiales. Stress-related (e.g., WRKY) and flowering-related (e.g., MADS-box) genes were located in the triplicated regions, suggesting that the polyploidy event might have contributed adaptive potential. Genes related to terpenoid biosynthesis, for example, FTA and TPS, were observed to be duplicated to a great extent in the J. sambac genome, perhaps explaining the strong fragrance of the flowers. Copy number changes in distinct phylogenetic clades of the MADS-box family were observed in J. sambac genome, for example, AGL6- and Mα- were lost and SOC- expanded, features that might underlie the long flowering period of J. sambac. The structural genes implicated in anthocyanin biosynthesis were depleted and this may explain the absence of vivid colours in jasmine. Collectively, assembling the J. sambac genome provides new insights into the genome evolution of the Oleaceae family and provides mechanistic insights into floral properties.

Ladder-shape melting temperature isothermal amplification of nucleic acids.

A novel method, termed ladder-shape melting temperature isothermal amplification (LMTIA), was developed in this study. As a proof of concept, one pair of primers or two pairs of nested primers and a thermostable DNA polymerase were employed to amplify the internal transcribed spacer of Oryza sativa with the ladder-shape melting temperature curve. Our results demonstrated that the LMTIA assay with nested primers was 50-fold more sensitive than the LAMP assay with the same level of specificity. The LMTIA method has the potential to be used for the prevention and control of emerging epidemics caused by different types of pathogens.

Clinical characteristics of small bowel tumors diagnosed by double-balloon endoscopy: Experience from a Chinese tertiary hospital.

BACKGROUND/AIMS: To determine the characteristics of small bowel tumors (SBTs) in patients underwent double balloon endoscopy (DBE) and to compare the clinical value of DBE with other diagnostic tools. MATERIALS AND METHODS: A retrospective study was conducted in patients underwent DBE procedures from March 2008 to April 2017.The demographic, clinical and pathological characteristics of patients with SBTs were recorded, while the diagnosis of SBTs was achieved either by DBE biopsy or surgical specimens. RESULTS: One thousand one hundred and two patients (761 males, range 3-85 years) were enrolled in this study, with 1140 procedures completed in total. 99/1102 patients (9.0%) had SBTs, including benign polyps (20, 20.2%), gastrointestinal stromal tumors (GISTs) (24, 24.2%), lymphomas (13, 13.1%), adenocarcinoma (39, 39.4%), and neuroendocrine tumors (3, 3.0%). The most common clinical symptom for benign polyps was obscure gastrointestinal bleeding (OGIB) (75.0%). But among patients with malignant SBTs, the main indication for DBE was chronic abdominal pain (43.8%), followed by OGIB (36.3%), vomit (10.0%), abnormal images (6.3%) and diarrhea (3.8%) (P<0.001). Moreover, SBTs were primarily located in the jejunum alone (40/99, 40.4%). DBE had better sensitivity (89.2%), specificity (95.2%), positive predictive value (PPV) (90.0%), and negative predictive value (NPV) (94.8%) than other tools for suspected SBTs. CONCLUSION: Small bowel tumor is mainly located in jejunum and with OGIB and abdominal pain as major complaints. DBE is a reliable method for the diagnosis of SBTs compared with other diagnostic tools.

TRIM29 facilitates the epithelial-to-mesenchymal transition and the progression of colorectal cancer via the activation of the Wnt/ß-catenin signaling pathway.

BACKGROUND: Tripartite Motif 29 (TRIM29) has been newly identified as being implicated in cancer progression. However, the biological role and molecular mechanism of TRIM29 in the invasion and metastasis of colorectal cancer (CRC) remain to be determined. METHODS: The expression levels of TRIM29 and ß-catenin in CRC patient specimens were detected by immunohistochemistry. Recombinant lentivirus vectors containing the TRIM29 gene and its small hairpin interfering RNAs were constructed and transduced into CRC cells. Wound-healing and Transwell assays were performed to evaluate the migration and invasion abilities of CRC cells in vitro. Hepatic metastasis models in nude mice were established to validate the function of TRIM29 in vivo. Moreover, the expressions of epithelial-to-mesenchymal transition (EMT)-associated proteins were detected by qRT-PCR and Western blotting in CRC cells. Finally, Western blotting, qRT-PCR, luciferase reporter assays, and immunofluorescence assays were used to explore the molecular mechanisms of TRIM29 in CRC progression. RESULTS: Increased TRIM29 expression positively correlated with lymph node metastasis and ß-catenin expression in patient CRC tissues. Overexpression of TRIM29 promoted invasion and metastasis of CRC cells in vitro and in vivo by regulating EMT, whereas the knockdown of TRIM29 had the opposite effect. Further mechanistic studies suggest that TRIM29 can activate the Wnt/ß-catenin signaling pathway via up-regulating CD44 expression in colorectal cancer. CONCLUSIONS: TRIM29 induces EMT through activating the Wnt/ß-catenin signaling pathway via up-regulating CD44 expression, thus promoting invasion and metastasis of CRC.

Prognostic Analysis of Duodenal Gastrointestinal Stromal Tumors.

AIM: This study aims to analyze factors possibly related to the prognosis of duodenal gastrointestinal stromal tumors (DGISTs). METHODS: We collected and retrospectively analyzed clinical and pathological data of 62 patients with primary DGISTs. All the patients were hospitalized and received complete surgical resection at Shanghai Ruijin Hospital from September 2003 to April 2015. We followed up the patients to determine survival outcomes. We also analyzed the effect of clinical and pathological factors on disease-free survival (DFS) and overall survival (OS) of the patients. RESULTS: Kaplan-Meier univariate survival analysis demonstrated that tumor size, mitotic index, Ki-67 index, and pathological risk were correlated with the DFS and OS of the patients (DFS P = 0.039, 0.001, <0.001, and 0.005, resp.; OS P = 0.027, 0.007, <0.001, and 0.012, resp.). Cox multivariate regression analysis revealed that Ki-67 index was an independent prognostic factor affecting DFS and OS (P = 0.007 and 0.028, resp.). Moreover, Kaplan-Meier survival analysis showed that imatinib treatment for patients with recurrence was correlated with prolonged OS (P = 0.002). CONCLUSION: Prognosis for DGIST treated by R0 resection is favorable. High level of Ki-67 can be an independent risk factor of DGIST prognosis. Adjuvant imatinib therapy for patients with tumor recurrence could probably lead to prolonged survival.

Removal of caffeine from green tea by microwave-enhanced vacuum ice water extraction.

In order to selectively remove caffeine from green tea, a microwave-enhanced vacuum ice water extraction (MVIE) method was proposed. The effects of MVIE variables including extraction time, microwave power, and solvent to solid radio on the removal yield of caffeine and the loss of total phenolics (TP) from green tea were investigated. The optimized conditions were as follows: solvent (mL) to solid (g) ratio was 10:1, microwave extraction time was 6 min, microwave power was 350 W and 2.5 h of vacuum ice water extraction. The removal yield of caffeine by MVIE was 87.6%, which was significantly higher than that by hot water extraction, indicating a significant improvement of removal efficiency. Moreover, the loss of TP of green tea in the proposed method was much lower than that in the hot water extraction. After decaffeination by MVIE, the removal yield of TP tea was 36.2%, and the content of TP in green tea was still higher than 170 mg g(-1). Therefore, the proposed microwave-enhanced vacuum ice water extraction was selective, more efficient for the removal of caffeine. The main phenolic compounds of green tea were also determined, and the results indicated that the contents of several catechins were almost not changed in MVIE. This study suggests that MVIE is a new and good alternative for the removal of caffeine from green tea, with a great potential for industrial application.

Construction and characterization of a fusion ß-1,3-1,4-glucanase to improve hydrolytic activity and thermostability.

A new fusion gene (Bgl-licMB), encoding ß-1,3-1,4-glucanase both from Bacillus amyloliquefaciens (Bgl) and Clostridium thermocellum (licMB), was constructed via end-to-end fusion and expressed in Escherichia coli to improve hydrolytic activity and thermostability of ß-1,3-1,4-glucanase. The results of enzymatic properties showed that the catalytic efficiency (K(cat)/K(m)) of the fusion enzyme for oat ß-glucan was 2.7 and 20-fold higher than that of the parental Bgl and licMB, respectively, and that the fusion enzyme can retain more than 50% of activity following incubation at 80°C for 30 min, whereas the residual activities of Bgl and licMB were both less than 30%. These properties make this particular ß-1,3-1,4-glucanase a good candidate for application in brewing and animal-feed industries.