Nanopore analysis of salvianolic acids in herbal medicines.

Natural herbs, which contain pharmacologically active compounds, have been used historically as medicines. Conventionally, the analysis of chemical components in herbal medicines requires time-consuming sample separation and state-of-the-art analytical instruments. Nanopore, a versatile single molecule sensor, might be suitable to identify bioactive compounds in natural herbs. Here, a phenylboronic acid appended Mycobacterium smegmatis porin A (MspA) nanopore is used as a sensor for herbal medicines. A variety of bioactive compounds based on salvianolic acids, including caffeic acid, protocatechuic acid, protocatechualdehyde, salvianic acid A, rosmarinic acid, lithospermic acid, salvianolic acid A and salvianolic acid B are identified. Using a custom machine learning algorithm, analyte identification is performed with an accuracy of 99.0%. This sensing principle is further used with natural herbs such as Salvia miltiorrhiza, Rosemary and Prunella vulgaris. No complex sample separation or purification is required and the sensing device is highly portable.

DNA-induced assembly of biocatalytic nanocompartments for sensitive and selective aptasensing of aflatoxin B1.

Enzyme cascade with high specificity and catalytic efficiency has significant applications for developing efficient bioanalysis methods. In this work, a sensitive and selective aptasensor was constructed based on the DNA-induced assembly of biocatalytic nanocompartments. Different from the conventional co-immobilization in one pot, the cascade enzymes of glucose oxidase (GOX) and horseradish peroxidase (HRP) were separately encapsulated in ZIF-90 nanoparticles. After conjugating complementary DNA or aptermer on enzyme@ZIF-90, DNA hybridization drove enzyme@ZIF-90 connected into clusters or linked on other DNA modified biocatalytic nanocompartment (such as invertase loaded Fe3O4@SiO2). Owing to the shortened distance between enzymes, the catalytic efficiency of connected clusters was significantly enhanced. However, the specifically interaction between the substrate molecule and aptermer sequence would lead to the disassembly of DNA duplexes, resulting in the gradual "switching-off" of cascade reactions. With aflatoxin B1 (AFB1) as the model substrate, the compartmentalized three-enzyme nanoreactors showed good analytical performance in the linear range from 0.01 ng mL-1 to 50 ng mL-1 with a low detection limit (3.3 pg mL-1). In addition, the proposed aptasensor was applied to detect AFB1 in corn oil and wheat powder samples with total recoveries ranging from 94 % to 109 %. As a result, this DNA-induced strategy for enzyme cascade nanoreactors opens new avenues for stimuli-responsive applications in biosensing.

Unambiguous discrimination of all 20 proteinogenic amino acids and their modifications by nanopore.

Natural proteins are composed of 20 proteinogenic amino acids and their post-translational modifications (PTMs). However, due to the lack of a suitable nanopore sensor that can simultaneously discriminate between all 20 amino acids and their PTMs, direct sequencing of protein with nanopores has not yet been realized. Here, we present an engineered hetero-octameric Mycobacterium smegmatis porin A (MspA) nanopore containing a sole Ni2+ modification. It enables full discrimination of all 20 proteinogenic amino acids and 4 representative modified amino acids, Nω,N'ω-dimethyl-arginine (Me-R), O-acetyl-threonine (Ac-T), N4-(ß-N-acetyl-D-glucosaminyl)-asparagine (GlcNAc-N) and O-phosphoserine (P-S). Assisted by machine learning, an accuracy of 98.6% was achieved. Amino acid supplement tablets and peptidase-digested amino acids from peptides were also analyzed using this strategy. This capacity for simultaneous discrimination of all 20 proteinogenic amino acids and their PTMs suggests the potential to achieve protein sequencing using this nanopore-based strategy.

Effects of lumbar-pelvic training combined with electroacupuncture on chronic nonspecific low back pain.

This observational study was conducted to investigate the effect of lumbar-pelvic training (LP) combined with electroacupuncture (EA) in the treatment of chronic nonspecific low back pain. One hundred and twenty patients diagnosed with chronic nonspecific low back pain were evenly randomized to receive the following 4 treatments for 2 weeks: LP combined with EA (Group A), EA (Group B), LP (Group C) or no intervention (Group D). The LP was a self-developed training program containing 5 movements and was conducted three times a week to build up the strength of abdomen muscle groups. Four acupoints along the foot-taiyang bladder meridian and the governor vessel were chosen for EA five times a week based on the theory of Traditional Chinese Medicine. The Visual Analog Scale and Oswestry Disability Index were measured before and after treatment to assess the reduction of pain intensity and functional disability, respectively. Following the treatments, Visual Analog Scale and Oswestry Disability Index scores in all 3 intervention groups were significantly lower than those in the Group D without intervention (P < .01). Among the intervention groups, Group A's scores were lower than those of Group B or Group C (P < .01). The overall efficacy of Group A was 93.33%, which was higher than that of Group B (76.67%) and Group C (70.00%) (P < .01). In conclusion, this study suggest that our self-developed lumbar-pelvic training combined with electroacupuncture is effective for chronic nonspecific low back pain in terms of pain and disability reduction.

Selenium Protects Follicular Granulosa Cells from Apoptosis Induced by Mercury Through Inhibition of ATF6/CHOP Pathway in Laying Hens.

The purpose of this research was to explore the effect of selenium on mercury-mediated apoptosis of follicular granulosa cells in laying hens. Moreover, the ATF6/CHOP pathway was investigated to explore the mechanism in this progress. Hg, Se, and 4-phenyl butyric acid were used alone or in combination to treat the cells. Our results showed that the nuclear in cells became condensate after Hg exposure, while Se addition significantly alleviated this change. Hg exposure significantly induced the apoptosis and the reduction of mitochondrial membrane potential in cells (P < 0.05). Nevertheless, co-treatment of Se significantly inhibited these effects (P < 0.05). Additionally, Hg exposure dramatically elevated the gene expressions of Bax/Bcl-2 (P < 0.05), caspase-3 (P < 0.05), caspase-9 (P < 0.05), protein kinase RNA-like endoplasmic reticulum kinase (P < 0.05), activating transcription factor 6 (P < 0.05), C/EBP homologous protein (CHOP; P < 0.05), inositol-requiring enzyme 1α (P < 0.05), tumor necrosis factor-associated factor 2 (P < 0.05), activating transcription factor 6 (ATF6; P < 0.05), and apoptosis signal-regulating kinase 1 (P < 0.05) in cells, whereas Se addition avoided these changes. The exposure to Hg considerably boosted the expression of ATF6 and CHOP protein (P < 0.05), while Se addition significantly alleviated the above-mentioned enhancements (P < 0.05). In summary, Hg exposure induced apoptosis, which was considerably reduced alleviated by Se addition, which was linked to the ATF6/CHOP pathway in follicular granulosa cells in laying hens.

Effects of Glutamine on Rumen Digestive Enzymes and the Barrier Function of the Ruminal Epithelium in Hu Lambs Fed a High-Concentrate Finishing Diet.

The present experiment aimed to research the effects of glutamine (Gln) on the digestive and barrier function of the ruminal epithelium in Hu lambs fed a high-concentrate finishing diet containing some soybean meal and cottonseed meal. Thirty healthy 3-month-old male Hu lambs were randomly divided into three treatments. Lambs were fed a high-concentrate diet and supplemented with 0, 0.5, and 1% Gln on diet for 60 days. The experimental results show that the Gln treatment group had lower pepsin and cellulase enzyme activity, propionate acid concentration, and IL-6, TNF-α, claudin-1, and ZO-1 mRNA expression in the ruminal epithelium (p < 0.05); as well as increases in lipase enzyme activity, the ratio of propionic acid to acetic acid, the IL-10 content in the plasma, and the mRNA expression of IL-2 and IL-10 in the ruminal epithelium, in contrast to the CON (control group) treatment (p < 0.05). Taken together, the findings of this present study support the addition of Gln to improve digestive enzyme activity, the ruminal epithelium's barrier, and fermentation and immune function by supplying energy to the mononuclear cells, improving the ruminal epithelium's morphology and integrity, and mediating the mRNA expression of tight junction proteins (TJs) and cytokines.

Protective effect of nano-selenium on mercury-induced prehierarchical follicular atresia in laying hens.

This study investigated the effect of nano-selenium (nano-Se) in protecting laying hens from mercury (Hg)-induced prehierarchical follicular atresia. Furthermore, the endoplasmic reticulum stress (ERS) was explored to reveal the molecular mechanism. In vivo, 720 Hyline-Brown laying hens were treated with Hg and nano-Se alone or in combination. In vitro, the prehierarchical follicles were treated with Hg, nano-Se and 4-phenyl butyric acid (4-PBA) alone or in combination (Control, 25 µM Hg group, 10 µM nano-Se group, 20 µM nano-Se group, 25 µM Hg + 10 µM nano-Se group, 25 µM Hg + 20 µM nano-Se group, 25 µM Hg + 4-PBA group, and 25 µM Hg + 20 µM nano-Se + 4-PBA group). The GCs were treated with Hg and nano-Se alone or in combination (Control, 15 µM Hg group, 6 µM nano-Se group, 12 µM nano-Se group, 15 µM Hg + 6 µM nano-Se group, 15 µM Hg + 12 µM nano-Se group). The results revealed that dietary Hg significantly reduced laying performance (P < 0.05) and egg quality (P < 0.05), whereas nano-Se addition prevented these reductions (P < 0.05). Hg exposure significantly induced the accumulation of Hg in PHFs (P < 0.05), prehierarchical follicular atresia (P < 0.05) and apoptosis in PHFs, whereas nano-Se addition significantly prevented these effects (P < 0.05). The levels of sex hormones (P < 0.05) were significantly decreased after Hg exposure in vivo and in vitro, while nano-Se addition prevented the reductions. Furthermore, the RNA-Seq results showed that the key factors of the ERS presented differential expression, including C/EBP homologous protein, protein kinase RNA-like endoplasmic reticulum kinase (PERK) and activating transcription factor 6 (ATF6) in GCs. Hg exposure significantly increased the key gene expression of endoplasmic reticulum stress in GCs, whereas nano-Se addition prevented the induction of expression of these genes. In addition, the protein levels of PERK, inositol requiring protein 1α (IRE1α) and ATF6 were significantly increased, whereas nano-Se addition prevented the enhancements of protein expression in GCs. In conclusion, this study shows that Hg exposure can reduce induce prehierarchical follicular atresia, whereas nano-Se can prevent these effects. Our results also elucidate a key role of ERS in these protective effects of nano-Se in laying hens.

Effects of Dietary Supplementation with Glutamine on the Immunity and Intestinal Barrier Gene Expression in Broiler Chickens Infected with Salmonella Enteritidis.

The effects of glutamine (Gln) on immunity and intestinal barrier gene expression levels in broilers challenged with Salmonella Enteritidis were evaluated. A total of 400 1-day-old broilers were randomly assigned to four groups, 10 repetition treatments per group with 10 broiler chickens for a 21-day feeding trial. The groups were the normal control group (CON, no infected group, fed with a basal diet); the S. Enteritidis-infected control group (SCC, infected with 2.0 × 104 CFU/mL of S. Enteritidis, fed a basal diet); and the Gln 1 and 2 groups, who were challenged with S. Enteritidis and fed a basal diet plus Gln at 0.5% and 1.0%, respectively. The results show that S. Enteritidis had adverse effects on the average daily feed intake, average daily gain, and the feed conversion ratio of infected broilers compared with those of CON broilers on d 7 (p < 0.05); decreased serum immunoglobulin A (IgA), immunoglobulin M (IgM), and immunoglobulin G (IgG) concentrations, and intestinal mucosa Bcl-2 mRNA expression levels (p < 0.05); increased the Lysozyme (LZM, only serum), NO, inducible NO synthase (iNOS) (except at 4 d), and total nitric oxide synthase (TNOS) (except at 4 d) activities in serum and the intestinal mucosa; and increased intestinal mucosa polymeric immunoglobulin receptor (pIgR) (except at 21 d), Avian beta-defensin 5 (AvBD5), AvBD14, Bax, and Bak mRNA expression levels during the experimental period (p < 0.05). Supplementation with Gln improved growth performance; increased serum IgA, IgG, and IgM concentrations and intestinal mucosa Bcl-2 mRNA expression levels (p < 0.05); decreased the LZM (only serum), NO, iNOS (except at 4 d), and TNOS (except at 4 d) activities in serum and the intestinal mucosa; and decreased intestinal mucosa pIgR (except at 21 d), AvBD5, AvBD14, Bax, and Bak mRNA expression levels during the experimental period (p < 0.05). These results suggest that Gln might lessen the inflammatory reaction of the small intestine and enlarge the small bowel mucosa immune and barrier function in broiler chickens challenged with S. Enteritidis.

S2Snet: deep learning for low molecular weight RNA identification with nanopore.

Ribonucleic acid (RNA) is a pivotal nucleic acid that plays a crucial role in regulating many biological activities. Recently, one study utilized a machine learning algorithm to automatically classify RNA structural events generated by a Mycobacterium smegmatis porin A nanopore trap. Although it can achieve desirable classification results, compared with deep learning (DL) methods, this classic machine learning requires domain knowledge to manually extract features, which is sophisticated, labor-intensive and time-consuming. Meanwhile, the generated original RNA structural events are not strictly equal in length, which is incompatible with the input requirements of DL models. To alleviate this issue, we propose a sequence-to-sequence (S2S) module that transforms the unequal length sequence (UELS) to the equal length sequence. Furthermore, to automatically extract features from the RNA structural events, we propose a sequence-to-sequence neural network based on DL. In addition, we add an attention mechanism to capture vital information for classification, such as dwell time and blockage amplitude. Through quantitative and qualitative analysis, the experimental results have achieved about a 2% performance increase (accuracy) compared to the previous method. The proposed method can also be applied to other nanopore platforms, such as the famous Oxford nanopore. It is worth noting that the proposed method is not only aimed at pursuing state-of-the-art performance but also provides an overall idea to process nanopore data with UELS.

Role of Endoplasmic Reticulum Stress in Nano-Selenium Alleviating Prehierarchical Follicular Atresia Induced by Mercury in Laying Hens.

This study investigated that the effect of nano-selenium (nano-Se) addition preventing prehierarchical follicular atresia induced by mercury (Hg) exposure in laying hens. Furthermore, endoplasmic reticulum (ER) stress pathway was explored to reveal the protective mechanism of nano-Se in vitro. The results revealed that Hg could significantly reduce laying performance (P < 0.05) and egg quality (P < 0.05), whereas nano-Se addition partially reversed the reductions. Besides, Hg significantly induced the deposition of Hg in prehierarchical follicles (P < 0.05) and prehierarchical follicular atresia (P < 0.05), whereas nano-Se addition could alleviate these toxicities in vitro. In addition, Hg exposure could significantly reduce cell viability (P < 0.05) and induce pyknotic nucleus in prehierarchical granulosa cells, while nano-Se addition reversed these effects. The levels of follicle-stimulating hormone (P < 0.05), luteinizing hormone (P < 0.05), progesterone (P < 0.05), and estradiol (P < 0.05) were significantly decreased after Hg exposure in vitro. However, nano-Se addition reversed the decreases of sex hormone levels. Furthermore, Hg exposure significantly increased the gene expressions of CHOP (P < 0.05), PERK (P < 0.05), ATF4 (P < 0.05), ATF6 (P < 0.05), ASK1 (P < 0.05), IRE1α (P < 0.05), TRAF2 (P < 0.05), caspase-9 (P < 0.05), caspase-3 (P < 0.05), and Bax/Bcl-2 (P < 0.05), whereas nano-Se addition reversed these increases of gene expressions in vitro. In summary, this study provides that Hg can induce prehierarchical follicular atresia, whereas nano-Se addition can ameliorate it, and elucidates an important role of ER stress in nano-Se alleviating prehierarchical follicular atresia induced by Hg in laying hens.

Distinctive quality control method for solid-state fermented Isaria cicadae from strain Ic-17-7 and application in a rat model of type 2 diabetes.

This work was aimed to establish a quality control method for evaluating the effects on glucose and lipids of the fruiting body of Isaria cicadae Miquel from strain Ic-17-7 (Ic-17-7fb) using a rat model of type 2 diabetes (T2DM). Random amplified polymorphic DNA, sequence-characterized amplified region, and high-performance liquid chromatography (HPLC) were used for the quality control of Ic-17-7fb. The pharmacological effects on streptozocin (STZ)-induced high fat diet (HFD)-fed Albino Wistar rats were evaluated. The rats underwent the following treatments: control, metformin, Ic-17-7fb (0.166 and 0.5 g·kg-1) or without treatment. The fasting blood glucose (FBG), blood glucose, total cholesterol (TC), triglyceride (TG), high-density lipoprotein (HDL-c), and low-density lipoprotein (LDL-c) were measured. Ic-17-7fb amplified a single specific band by S11-2-F3 and S11-2-R3 primers. An HPLC-based quality and quantity method was established for industrial application. The contents of adenosine and N6-(2-hydroxyethyl) adenosine (HEA) of the cultivated Ic-17-7fb were analyzed. All of the validation lots of cultured Ic-17-7fb passed the quantity control of the training set (0.90 mg·g-1 of adenosine and 0.89 mg·g-1 of HEA). After two weeks of administration, the average FBG was 4.89 ± 0.42 (control), 26.10 ± 5.77 (model), 23.63 ± 6.15 (metformin), 17.96 ± 9.36 (Ic-17-7fb for 0.166 g·kg-1), and 19.69 ± 8.71 mmol·L-1 (Ic-17-7fb for 0.5 g·kg-1). The FBG of Ic-17-7fb (0.166 g·kg-1) treatment significantly reduced by 31.19%, compared with the model after two weeks of administration (P < 0.01). Metformin, Ic-17-7fb (0.166 g·kg -1), and Ic-17-7fb (0.5 g·kg-1) reduced TC, TG, HDL-c, and LDL-c compared with the T2DM model treatment at the 6th week of treatment (P < 0.05). This study established the first quality standard for Ic-17-7fb, which can be effectively applied in the treatment of T2DM. The reliable quality control method and pharmacological effect will broaden its application space.

Kaempferol-3-O-Glucuronide Ameliorates Non-Alcoholic Steatohepatitis in High-Cholesterol-Diet-Induced Larval Zebrafish and HepG2 Cell Models via Regulating Oxidation Stress.

NAFLD (non-alcoholic fatty liver disease) is one of the most prominent liver diseases in the world. As a metabolic-related disease, the development of NAFLD is closely associated with various degrees of lipid accumulation, oxidation, inflammation, and fibrosis. Ilex chinensis Sims is a form of traditional Chinese medicine which is used to treat bronchitis, burns, pneumonia, ulceration, and chilblains. Kaempferol-3-O-glucuronide (K3O) is a natural chemical present in Ilex chinensis Sims. This study was designed to investigate the antioxidative, fat metabolism-regulating, and anti-inflammatory potential of K3O. A high-cholesterol diet (HCD) was used to establish steatosis in larval zebrafish, whereby 1mM free fatty acid (FFA) was used to induce lipid accumulation in HepG2 cells, while H2O2 was used to induce oxidative stress in HepG2. The results of this experiment showed that K3O reduced lipid accumulation and the level of reactive oxygen species (ROS) both in vivo (K3O, 40 µM) and in vitro (K3O, 20 µM). Additionally, K3O (40 µM) reduced neutrophil aggregation in vivo. K3O (20 µM) also decreased the level of malondialdehyde (MDA) and significantly increased the level of glutathione peroxidase (GSH-px) in both the HCD-induced larval zebrafish model and H2O2-exposed HepG2 cells. In the mechanism study, keap1, nrf2, tnf-α, and il-6 mRNA were all significantly reversed by K3O (20 µM) in zebrafish. Changes in Keap1 and Nrf2 mRNA expression were also detected in H2O2-exposed HepG2 cells after they were treated with K3O (20 µM). In conclusion, K3O exhibited a reduction in oxidative stress and lipid peroxidation, and this may be related to the Nrf2/Keap1 pathway in the NAFLD larval zebrafish model.

Protective effect of selenomethionine on T-2 toxin-induced liver injury in New Zealand rabbits.

BACKGROUND: T-2 toxin is a mycotoxin produced by Fusarium species that is highly toxic to animals. Recent studies have indicated that Selenomethionine (SeMet) have protective effect against mycotoxins-induced toxicity. The aim of the present study was to investigate the protective effect of SeMet on T-2-toxin-induced liver injury in rabbit and explore its molecular mechanism. Fifty rabbits (30 d, 0.5 ± 0.1 kg) were randomly divided into 5 groups: control group, T-2 toxin group, low, medium and high dose SeMet treatment group. The SeMet-treated group was orally pretreated with SeMet (containing selenium 0.2 mg/kg, 0.4 mg/kg and 0.6 mg/kg) for 21 days. On the 17th day, T-2 toxin group and SeMet-treated group were orally administered with T-2 toxin (0.4 mg/kg body weight) for 5 consecutive days. RESULTS: The results showed that low-dose SeMet significantly improved T-2 toxin-induced liver injury. We found that low-dose SeMet can reduce the level of oxidative stress and the number of hepatocyte apoptosis. Moreover, the levels of Bax, caspase-3 and caspase-9 were significantly reduced and the levels of Bcl-2 were increased. CONCLUSIONS: Therefore, we confirmed that low-dose SeMet may protect rabbit hepatocytes from T-2 toxin by inhibiting the mitochondrial-caspase apoptosis pathway.

Protective effect of selenomethionine on intestinal injury induced by T- 2 toxin.

T-2 toxin is the most toxic as a type A trichothecenes, which could contaminate grains, especially in wheat and corn. It can cause immune suppression, neurotoxicity, the apoptosis of cells and even induce tumorigenesis. Recent studies have indicated that selenium (Se) have protective effect against mycotoxins-induced toxicity. The present studies was designed to investigate the protective role of Selenomethionine (SeMet) on T-2 toxin-induced toxicity in rabbit's jejunum. 50 New Zealand rabbits were divided into five group (Control group, T-2 group, low-dose Se + T-2 group, medium-dose + T-2 group and high-dose Se + T-2 group). New Zealand rabbits were orally administered with SeMet (0.2, 0.4 and 0.6 mg/kg, Adding diet) for 21 days. On 17th days, each group began to take 0.4 mg/kg of T-2 toxin orally every day for 5 days. We found that rabbit exposed to T-2 toxin could increase the levels of ROS, and decrease activities of antioxidant enzymes and the expression of Occludin and ZO-1. In addition, T-2 toxin could trigger jejunal inflammatory response and enhance the expression of IL-1ß, IL-6 and TNF-α. After SeMet pretreatment, our results indicated that Se attenuated the T-2 toxin-induced oxidative stress, decreasing the level of ROS, MDA and enhancing the activity of SOD and GSH-Px. Moreover, SeMet can alleviate jejunal inflammatory response, and protect the integrity of the intestinal barrier through up-regulating the expression of ZO-1 and Occludin. In the present research, supplementation of 0.2 mg/kg SeMet in the diet could effectively alleviate the T-2 toxin poisoning in rabbits.

Potential therapeutic effects of Cordyceps cicadae and Paecilomyces cicadae on adenine-induced chronic renal failure in rats and their phytochemical analysis.

BACKGROUND: Natural Cordyceps cicadae (C. cicadae) has been utilized extensively in traditional Chinese medicine to treat chronic renal diseases, heart palpitations, infantile convulsions, and dizziness. However, given its slow growth and immoderate exploitation, C. cicadae resources have been severely depleted. By contrast, Paecilomyces cicadae (P. cicadae), as the anamorph stage of C. cicadae, is easy to cultivate, and this kind of cultivated P. cicadae has good and controllable quality. PURPOSE: This study aimed to compare the therapeutic effects of C. cicadae and P. cicadae on adenine-induced chronic renal failure (CRF) rats. In accordance with the aforementioned studies, our work subsequently analyzed the intrinsic relationships between the efficacy and pharmacodynamic substances of C. cicadae and P. cicadae to conclude whether or not P. cicadae could be used as an alternative to C. cicadae in treating CRF. METHODS: Rats were administered with C. cicadae (1.0 g/kg) or P. cicadae (1.0 g/kg) by gavage for 4 weeks. Furthermore, we applied Fourier transform infrared spectroscopy, gas chromatography-mass spectrometry, liquid chromatography-tandem mass spectrometry, and ultraviolet spectrophotometry to comprehensively detect and analyze the chemical constituent differences from ten batches each of C. cicadae and P. cicadae. RESULTS: This study revealed that both C. cicadae and P. cicadae exerted obvious therapeutic effects on CRF and were more consistent with their chemical compositions. CONCLUSION: P. cicadae can be used as an alternative to C. cicadae for treating CRF to cater to market demands.

Distribution of elements extracted from symptom patterns and characteristics of polysomnograph of common symptom patterns of insomnia with Traditional Chinese Medicine.

OBJECTIVE: To analyze the distribution and combined regulation of elements of symptom patterns in the diagnosis of insomnia with Traditional Chinese Medicine (TCM). METHODS: The samples were collected from the patients, diagnosed with insomnia, of Henan Province Hospital of TCM between June 2011 and September 2013. The symptom patterns in insomnia were extracted. Next, symptom differentiation, characteristics of polysomnography (PSG), distribution and combined regulation of these symptom patterns were conducted by tests. RESULTS: In total, 286 eligible patients were recruited. The main locations of the disease symptom elements were the brain and heart, and the main characteristics of the disease symptom elements were phlegm-heat, Yin-deficiency and Qi-stagnation. The elements from two or three symptom patterns were commonly manifested in patients with insomnia, especially from three symptom patterns. We also found that all TCM symptom patterns had an effect on polysomnographic indicators in PSG tests. CONCLUSION: The elements of symptom patterns in insomnia were identified as mainly fire-heat and phlegm-heat. The most common patterns of excess were pathogenic fire derived from stagnation of liver- Qi, and mental disturbance due to phlegm-heat, while the most common patterns of deficiency in both the heart and the spleen. There are many differences in PSG indicators of different syndrome patterns of insomnia.

Fuzheng Huayu Recipe Ameliorates Liver Fibrosis by Restoring Balance between Epithelial-to-Mesenchymal Transition and Mesenchymal-to-Epithelial Transition in Hepatic Stellate Cells.

Activation of hepatic stellate cells (HSCs) depending on epithelial-to-mesenchymal transition (EMT) reflects the key event of liver fibrosis. Contrastively, mesenchymal-to-epithelial transition (MET) of HSCs facilitates the fibrosis resolution. Here we investigated the effect of Fuzheng Huayu (FZHY) recipe, a Chinese herbal decoction made of Radix Salviae Miltiorrhizae, Semen Persicae, Cordyceps sinensis, Pollen Pini, and Gynostemma pentaphyllum, on liver fibrosis concerning the balance of EMT and MET in HSCs. In contrast to the increased TGF-ß 1/BMP-7 ratio in activated HSCs, FZHY administration induced significant upregulation of BMP-7 and downregulation of TGF-ß 1 at both transcription and translation levels. Restoration of TGF-ß 1/BMP-7 ratio inhibited the expression of p38 MAPK and phosphorylated p38 MAPK, resulting in the reversal of epithelial-to-mesenchymal transition (EMT) to mesenchymal-to-epithelial transition (MET) as characterized by the abolishment of EMT markers (α-SMA and desmin) and reoccurrence of MET marker (E-cadherin). In vivo treatment of FZHY recipe also demonstrated the statistical reduction of activated HSCs with EMT phenotype, which attenuated the carbon tetrachloride- (CCl4-) induced liver fibrosis in a dose-dependent manner. These findings may highlight a novel antifibrotic role of FZHY recipe on the basis of rebalancing EMT and MET in HSCs.

[Virtual screening of small molecular HIV-1 entry inhibitor NC-2 targeting gp120 and its action mechanism].

OBJECTIVE: To screen the HIV-1 entry inhibitors targeting HIV-1 gp120 from the IBS natural product database by virtual screening based on the binding mode of the neutralizing antibody VRC01 with HIV-1 gp120 and investigate the anti-viral activities of the inhibitors and their action mechanisms. METHODS: The binding interaction of the candidate molecules binding gp120 and changes of the binding free energy were analyzed by MM-PBSA calculation. The anti-HIV-1 activities of the tested compounds were detected by HIV-1 pseudotyped virus, laboratory-adapted HIV-1 and a cell-cell fusion assay. The cytotoxicity of the studied molecules was examined by XTT colorimetric assay. The mechanisms of the anti-viral activities of the candidate molecules were analyzed using enzyme-linked immunosorbent assay. RESULTS: A total of 19 molecules with distinct reduction of the binding free energy after binding with gp120 were screened from 40000 molecules. Among them, NC-2 showed anti-HIV-1 activities against HIV-1 pseudotyped virus and laboratory-adapted HIV-1, and was capable of blocking HIV-1 envelope-mediated cell-cell fusion. The IC50 of NC-2 for inhibiting HIV-1IIIB and pseudotyped HIV-1JRFL infection were 1.95∓0.44 µmol/L and 10.58∓0.13 µmol/L, respectively. The results of ELISA suggested that NC-2 could inhibit the binding of HIV-1 gp120 to CD4 without blocking the formation of gp41 six-helix bundle in vitro. CONCLUSION: This computer-based virtual screening method can be used to screen HIV-1 entry inhibitors targeting gp120. Using this virtual screening approach combined with anti-viral activity screening, we obtained a potent HIV-1 entry inhibitor NC-2 with novel structure.

2,3,4',5-Tetrahydroxystilbene-2-O-ß-d-glucoside inhibits platelet-derived growth factor-induced proliferation of vascular smooth muscle cells by regulating the cell cycle.

1. 2,3,4',5-Tetrahydroxystilbene-2-O-ß-d-glucoside (TSG) has been shown to have an anti-atherosclerotic effect. Vascular smooth muscle cell (VSMC) proliferation contributes to the pathobiology of atherosclerosis. The aim of the present study was to investigate the effects of TSG on platelet-derived growth factor (PDGF)-BB-induced VSMC proliferation and to explore the molecular mechanisms underlying the effects. 2. Cultured rat VSMC were pretreated with TSG (l-50 µmol/L) for 1 h, followed by exposure to PDGF-BB (10 ng/mL) for 24 h, after which cell proliferation and cell cycle stages were examined. The expression of protein cell cycle regulators, including retinoblastoma (Rb), cyclin D1/E, cyclin-dependent kinase (CDK) 2/4, CDK inhibitors p21 and p27 and proliferative cell nuclear antigen (PCNA), was examined. Activation of extracellular signal-regulated kinase (ERK) 1/2 was evaluated to elucidate the possible upstream mechanism by which TSG affects cell cycle regulators. 3. The results showed that TSG dose-dependently inhibited PDGF-BB-induced VSMC proliferation, possibly by blocking the progression of the cell cycle from the G(1) to S phase. In addition, TSG significantly inhibited PDGF-BB-induced phosphorylation of Rb and the expression of cyclin D1, CDK4, cyclin E, CDK2 and PCNA. In addition, TSG suppressed PDGF-BB-induced downregulation of p27 and upregulation of p21, as well as PDGF-BB-induced activation of ERK1/2. 4. Together, the findings of the present study provide the first evidence that TSG can inhibit PDGF-BB-stimulated VSMC proliferation via cell cycle arrest in association with modulation of the expression of cell cycle regulators, which may be mediated, at least in part, by suppression of ERK1/2 activation.

Comparison of ginsenosides Rg1 and Rb1 for their effects on improving scopolamine-induced learning and memory impairment in mice.

Rg1 and Rb1 are two major active compounds of ginseng that facilitate learning and memory. The present study aimed to compare the nootropic effects of Rg1 and Rb1 in a scopolamine induced dementia mice model. After 6 and 12 mg/kg of Rg1 and Rb1 intraperitoneal administration to mice for 7 days, their effects were assessed using the step-down passive avoidance (SD) and the Morris water maze (MWM) tests, the acetylcholinesterase (AChE) activity, acetylcholine (ACh) content and serotonin (5-HT) level in the hippocampus were analysed after SD and MWM tests. The results showed that Rg1 and Rb1 ameliorated cognition-deficiency in mice with dementia. Rg1 showed stronger effects than Rb1 on escape acquisition in MWM. Both Rg1 and Rb1 increased ACh levels in the hippocampus, but Rg1 inhibited AChE activity while Rb1 had no effect on AChE activity. Both Rg1 and Rb1 inhibited the decrease of 5-HT induced by scopolamine, but Rb1 was more active than the same dose of Rg1. These results demonstrate that multiple administrations of Rg1 and Rb1 are effective in improving memory deficiency induced by scopolamine. Rg1 appears to be more potent than Rb1 in improving acquisition impairment, and the two ginsenosides may act through different mechanisms.