Bioinformatics and LC-QTOF-MS based discovery of pharmacodynamic and Q-markers of Pitongshu against functional dyspepsia.

ETHNOPHARMACOLOGICAL RELEVANCE: Pitongshu (PTS) is a clinically effective empirical formula for the treatment of FD. The efficacy and safety of PTS have been demonstrated in randomized, controlled, double-blind trials, but there is a lack of understanding of the systematic evaluation of the efficacy of PTS and its material basis. OBJECTIVE: To investigate the efficacy of PTS in Functional dyspepsia (FD) mice and possible Q-markers. METHOD: In this study, we used "irregular feeding + chronic unpredictable chronic stimulation" to establish a mice model of FD with hepatogastric disharmony. The efficacy of PTS was assessed from hair condition, behavioral, pain, gastrointestinal function, and serum 5-HT, GAS, MTL levels in mice by instillation of different doses of PTS. In addition, the composition of drugs in blood was analyzed by LC-QTOF-MS and potential Q-markers were selected by combining network pharmacology, molecular docking and actual content. RESULT: Our study showed that different doses of PTS increased pain threshold and writhing latency, decreased the number of writhings, increased gastric emptying rate and small intestinal propulsion rate, decreased total acidity of gastric contents and gastric acid secretion, and increased serum levels of 5-HT, GAS, and MTL in mice to different degrees. Enrichment analysis showed that PTS may be anti-FD through multiple pathways such as Serotonergic synapse, thyroid hormone signaling pathway, cholinergic synapse, and dopaminergic synapse. In addition, potential active ingredient substances were explored by LC-QTOF-MS combined with bioinformatics. Combined with the actual contentselected six constituents, hesperidin, neohesperidin, naringin, paeoniflorin, magnolol and honokiol, possible as Q-markers. CONCLUSION: PTS may exert its anti-FD effects through multi-component, multi-target and multi-pathway". Constituents, hesperidin, neohesperidin, naringin, paeoniflorin, magnolol and honokiol may be the Q-markers of its anti-FD effects.

Molecular mechanism of the effect of Gegen Qinlian decoction on COVID-19 comorbid with diabetes mellitus based on network pharmacology and molecular docking: A review.

To explore the potential mechanism of Gegen Qinlian decoction (GGQL) in the treatment of COVID-19 comorbid with diabetes mellitus (DM) through network pharmacology and molecular docking, and to provide theoretical guidance for clinical transformation research. Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform was used to screen the active compounds and targets of GGQL, the targets of COVID-19 comorbid with DM were searched based on Genecards database. Protein-protein interaction network was constructed using String data platform for the intersection of compounds and disease targets, the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis of the intersection targets was performed using DAVID database. Cytoscape software was used to construct the "compound target-pathway (C-T-P)" of GGQL in the treatment of COVID-19 comorbid with DM, the molecular docking platform was used to complete the simulated docking of key compounds and targets. We obtained 141 compounds from GGQL, revealed 127 bioactive compounds and 283 potential targets of GGQL. Quercetin, kaempferol and formononetin in GGQL play a role by modulating the targets (including AR, GSK3B, DPP4, F2, and NOS3). GGQL might affect diverse signaling pathways related to the pathogenesis of coronavirus disease - COVID-19, AGE-RAGE signaling pathway in diabetic complications, IL-17 signaling pathway, human cytomegalovirus infection and Th17 cell differentiation. Meanwhile, molecular docking showed that the selected GGQL core active components had strong binding activity with the key targets. This study revealed that GGQL play a role in the treatment of COVID-19 comorbid with DM through multi-component, multi-target and multi-pathway mode of action, which provided good theoretical basis for further verification research.

[Origin identification of Poria cocos based on hyperspectral imaging technology].

To realize the non-destructive and rapid origin discrimination of Poria cocos in batches, this study established the P. cocos origin recognition model based on hyperspectral imaging combined with machine learning. P. cocos samples from Anhui, Fujian, Guangxi, Hubei, Hunan, Henan and Yunnan were used as the research objects. Hyperspectral data were collected in the visible and near infrared band(V-band, 410-990 nm) and shortwave infrared band(S-band, 950-2 500 nm). The original spectral data were divided into S-band, V-band and full-band. With the original data(RD) of different bands, multiplicative scatter correction(MSC), standard normal variation(SNV), S-G smoothing(SGS), first derivative(FD), second derivative(SD) and other pretreatments were carried out. Then the data were classified according to three different types of producing areas: province, county and batch. The origin identification model was established by partial least squares discriminant analysis(PLS-DA) and linear support vector machine(LinearSVC). Finally, confusion matrix was employed to evaluate the optimal model, with F1 score as the evaluation standard. The results revealed that the origin identification model established by FD combined with LinearSVC had the highest prediction accuracy in full-band range classified by province, V-band range by county and full-band range by batch, which were 99.28%, 98.55% and 97.45%, respectively, and the overall F1 scores of these three models were 99.16%, 98.59% and 97.58%, respectively, indicating excellent performance of these models. Therefore, hyperspectral imaging combined with LinearSVC can realize the non-destructive, accurate and rapid identification of P. cocos from different producing areas in batches, which is conducive to the directional research and production of P. cocos.

[Origin identification of Gardeniae Fructus based on hyperspectral imaging technology].

In order to realize rapid and non-destructive identification of the origin of Gardeniae Fructus, a technical method based on hyperspectral imaging technology was established in this study. Spectral information of Gardeniae Fructus samples from eight production origins was acquired from visible NIR(410-990 nm, VNIR) and short wavelength NIR(950-2 500 nm, SWIR) bands based on hyperspectral imaging techniques. The average spectral reflectance within the region of interest was extracted and calculated using the ENVI 5.3 software, resulting in 1 600 sample data. The visible short wavelength infrared band(fused bands) spectral data covering the range 410-2 500 nm were obtained after combining the spectral data of VNIR and SWIR. Data were de-noised by five common preprocessing methods, including multivariate scatter correction, Savitzky-Golay smoothing, standard normal variate, first derivative(FD), and second derivative from VNIR, SWIR, and fused bands(VNIR+SWIR). Partial least squares discriminant analysis, linear support vector classification(LinearSVC), and random forest were used to establish the model for origin identification of Gardeniae Fructus. The results indicated that the identification model of Gardeniae Fructus origin established after FD pretreatment of the spectral data in the fused bands could yield good results. According to the confusion matrix evaluation results, the model prediction set using LinearSVC reached 100% accuracy, so the optimum identification model of Gardeniae Fructus origin was determined as fusion bands-FD-LinearSVC. Therefore, the hyperspectral imaging technology can achieve rapid, nondestructive, and accurate identification of Gardeniae Fructus samples of different origins, which provides a technical reference for the differential detection of Gardeniae Fructus and other Chinese medicines.

Underlying mechanisms of acupuncture therapy on polycystic ovary syndrome: Evidences from animal and clinical studies.

Polycystic ovary syndrome (PCOS) is a common endocrine and metabolic disorder among women of reproductive age. Current standard treatment includes lifestyle change, oral pharmacological agents, and surgical modalities. However, the efficacy of current therapies is less than satisfactory. Clinical evidence has shown that acupuncture is effective for regulating hormone levels, promoting ovulation, and attenuating insulin resistance in patients with PCOS. Acupuncture may affect the production of ß-endorphin, which may lead to gonadotropin-releasing hormone secretion and then affect ovulation, menstrual cycle, and fertility. The mechanism of acupuncture for patients with PCOS has not been comprehensively reviewed so far. Better understanding of the mechanisms of acupuncture would help popularize the use of acupuncture therapy for patients with PCOS. In this narrative review, we aimed to overview the potential mechanisms and evidence-based data of acupuncture on PCOS, and analyze the most frequently used acupoints based on animal and clinical studies. The results of this study will contribute to a better understanding of the current situation in this field.

Biochemical and structural characterization of the cyanophage-encoded phosphate-binding protein: implications for enhanced phosphate uptake of infected cyanobacteria.

To acquire phosphorus, cyanobacteria use the typical bacterial ABC-type phosphate transporter, which is composed of a periplasmic high-affinity phosphate-binding protein PstS and a channel formed by two transmembrane proteins PstC and PstA. A putative pstS gene was identified in the genomes of cyanophages that infect the unicellular marine cyanobacteria Prochlorococcus and Synechococcus. However, it has not been determined whether the cyanophage PstS protein is functional during infection to enhance the phosphate uptake rate of host cells. Here we showed that the cyanophage P-SSM2 PstS protein was abundant in the infected Prochlorococcus NATL2A cells and the host phosphate uptake rate was enhanced after infection. This is consistent with our biochemical and structural analyses showing that the phage PstS protein is indeed a high-affinity phosphate-binding protein. We further modelled the complex structure of phage PstS with host PstCA and revealed three putative interfaces that may facilitate the formation of a chimeric ABC transporter. Our results provide insights into the molecular mechanism by which cyanophages enhance the phosphate uptake rate of cyanobacteria. Phosphate acquisition by infected bacteria can increase the phosphorus contents of released cellular debris and virus particles, which together constitute a significant proportion of the marine dissolved organic phosphorus pool.

Age-related heterogeneity revealed by disruption of white matter structural networks in patients with first-episode untreated major depressive disorder: WM Network In OA-MDD.

The clinical treatment and prognosis of major depressive disorder (MDD) are limited by the high degree of disease heterogeneity. It is unclear whether there is a potential network mechanism for age-related heterogeneity. We aimed to uncover the heterogeneity of the white matter (WM) network at different ages of onset and its correlation with different symptom characteristics. 85 first-episode MDD patients and 84 corresponding healthy controls (HCs) were recruited and underwent diffusion tensor imaging scans. Structural network characteristics were analyzed using graph theory methods. We observed an accelerated age-related decline of the WM network in MDD patients compared with HCs. Distinct symptom-related networks were identified in three MDD groups with different onset-age. For early-onset MDD (18-29 years; EOD), higher guilt and loss of interest were correlated with the insula, and inferior parietal lobe which in default mode network and salience network. For mid-term-onset MDD (30-44 years; MOD), higher somatic symptoms were correlated with thalamus which in cortico-striatal-thalamic-cortical circuit. For later-onset MDD (45-60 years; LOD), poor sleep symptoms were correlated with the caudate in the basal ganglia, which suggests the cingulate operculum network in the control of sleep. These results supported a circuit-based heterogeneity associated with the age of onset in MDD. Understanding this circuit-based heterogeneity might help to develop a new target for clinical treatment strategies.

Secondary Amine Pendant ß-Peptide Polymers Displaying Potent Antibacterial Activity and Promising Therapeutic Potential in Treating MRSA-Induced Wound Infections and Keratitis.

Interest in developing antibacterial polymers as synthetic mimics of host defense peptides (HPDs) has accelerated in recent years to combat antibiotic-resistant bacterial infections. Positively charged moieties are critical in defining the antibacterial activity and eukaryotic toxicity of HDP mimics. Most examples have utilized primary amines or guanidines as the source of positively charged moieties, inspired by the lysine and arginine residues in HDPs. Here, we explore the impact of amine group variation (primary, secondary, or tertiary amine) on the antibacterial performance of HDP-mimicking ß-peptide polymers. Our studies show that a secondary ammonium is superior to either a primary ammonium or a tertiary ammonium as the cationic moiety in antibacterial ß-peptide polymers. The optimal polymer, a homopolymer bearing secondary amino groups, displays potent antibacterial activity and the highest selectivity (low hemolysis and cytotoxicity). The optimal polymer displays potent activity against antibiotic-resistant bacteria and high therapeutic efficacy in treating MRSA-induced wound infections and keratitis as well as low acute dermal toxicity and low corneal epithelial cytotoxicity. This work suggests that secondary amines may be broadly useful in the design of antibacterial polymers.

Effects of Macroporous Resin Extract of Dendrobium officinale Leaves in Rats with Hyperuricemia Induced by Fructose and Potassium Oxonate.

AIMS AND OBJECTIVES: Fructose, as a ubiquitous monosaccharide, can promote ATP consumption and elevate circulating Uric Acid (UA) levels. Our previous studies have confirmed that the macroporous resin extract of Dendrobium officinale leaves (DoMRE) could reduce the UA level of rats with hyperuricemia induced by a high-purine diet. This study aimed to investigate whether DoMRE had a UA-lowering effect on rats with hyperuricemia caused by fructose combined with potassium oxonate, so as to further clarify the UA-lowering effect of DoMRE, and to explore the UAlowering effect of DoMRE on both UA production and excretion. MATERIALS AND METHODS: Rats with hyperuricemia induced by fructose and potassium oxonate were administered with DoMRE and vehicle control, respectively, to compare the effects of the drugs. At the end of the experiment, the Serum Uric Acid (SUA) and Creatinine (Cr) levels were measured using an automatic biochemical analyzer, the activities of xanthine oxidase (XOD) were measured using an assay kit, and the protein expressions of Urate Transporter 1 (URAT1), glucose transporter 9 (GLUT9), and ATP-Binding Cassette Superfamily G member 2 (ABCG2) were assessed using immune-histochemical and western blot analyses. Hematoxylin and eosin staining was used to assess the histological changes in the kidney, liver, and intestine. RESULTS: Fructose and potassium induced hyperuricemia in rats. Meanwhile, the activities of XOD were markedly augmented, the expression of URAT1 and GLUT9 was promoted, and the expression of ABCG2 was reduced, which were conducive to the elevation of UA. However, exposure to DoMRE reversed these fructose- and potassium oxonate-induced negative alternations in rats. The activities of XOD were recovered to the normal level, reducing UA formation; the expressions of URAT1, ABCG2, and GLUT9 returned to the normal level, resulting in an increase in renal urate excretion. CONCLUSION: DoMRE reduces UA levels in rats with hyperuricemia induced by fructose combined with potassium oxonate by inhibiting XOD activity and regulating the expression of ABCG2, URAT1, and GLUT9. DoMRE is a potential therapeutic agent for treating hyperuricemia through inhibiting UA formation and promoting UA excretion.

[Electroacupuncture postpones growth of tumors by activating SIRT1 expression to reduce inflammatory reaction and facilitate cellular autophagy in mice with transformed colorectal carcinomas].

OBJECTIVE: To observe the effect of electroacupuncture (EA) on tumor number, body conditions, inflammatory factors and expression levels of silent information regulator 1 (sirtuin 1, SIRT1) and autophagy-related proteins Beclin-1, P62, and LC3 in colorectal tissues in inflammatory-transformed colorectal cancer mice, so as to explore its underlying mechanisms in resisting tumor growth. METHODS: A total of 100 C57BL/6 male mice were randomly divided into normal control, model, EA, EA + SIRT1 inhibitor (EA+inhibitor) and agonist resveratrol (agonist) groups, with 20 mice in each group. EA (2 Hz, 1 mA) was applied to "Zusanli"(ST36)and "Fenglong"(ST40) for 20 min every time, 3 times a week for 11 weeks. Mice of the EA +inhibitor group received intraperitoneal injection of SIRT1 inhibitor EX527 (5 mg/kg) at the same time of EA treatment, and those of the agonist group received gavage of resveratrol (200 mg/kg, an agonist of SIRT1), 3 times a week for 11 weeks. The body mass was measured weekly. The disease activity index (DAI), colorectal length and tumor number in each group were recorded. The histopathological changes of colorectal tissues were observed by H.E. staining; the contents of interleukin 6 (IL-6), IL-10, IL-17, in the colorectal tissues were detected by enzyme-linked immunosorbent assay, and the expression levels of SITR1, Beclin-1, P62, and LC3 in colorectal tissues were detected by Western blot and real-time fluorescence quantitative PCR, respectively. RESULTS: Compared with the normal control group, the body weight, length of colorectum, the contents of IL-10, and the expression levels of SIRT1,Beclin-1 and LC3 mRNAs and proteins were significantly decreased (P<0.001), whereas the DAI score, the number of tumors, the contents of IL-6 and IL-17, and the expression levels of P62 mRNA and protein were significantly increased (P<0.000 1, P<0.001) in the model group. In comparison with the model group, the body weight, the length of colorectum, the contents of IL-10, and the expression levels of SIRT1,Beclin-1 and LC3 mRNAs and proteins were significantly increased (P<0.01,P<0.05,P<0.001), while the DAI scores, the numbers of tumors, the contents of IL-6 and IL-17, and the expression levels of P62 mRNA and protein were obviously decreased in the EA and agonist groups (P<0.01,P<0.05, P<0.001). No significant changes were found in all the above-mentioned indexes in the EA+inhibitor group in comparison with the model group (P>0.05). CONCLUSION: EA can reduce the number of tumors and inflammation reaction in colorectal tissue and improve the body condition in mice with colorectal cancer, which may be related to its functions in activating the expression of intestinal SIRT1, and then facilitating cellular autophagy.

Effect of oxaliplatin plus 5-fluorouracil or capecitabine on circulating and imaging biomarkers in patients with metastatic colorectal cancer: a prospective biomarker study.

BACKGROUND: Patients with metastatic colorectal cancer are treated with cytotoxic chemotherapy supplemented by molecularly targeted therapies. There is a critical need to define biomarkers that can optimise the use of these therapies to maximise efficacy and avoid unnecessary toxicity. However, it is important to first define the changes in potential biomarkers following cytotoxic chemotherapy alone. This study reports the impact of standard cytotoxic chemotherapy across a range of circulating and imaging biomarkers. METHODS: A single-centre, prospective, biomarker-driven study. Eligible patients included those diagnosed with colorectal cancer with liver metastases that were planned to receive first line oxaliplatin plus 5-fluorouracil or capecitabine. Patients underwent paired blood sampling and magnetic resonance imaging (MRI), and biomarkers were associated with progression-free survival (PFS) and overall survival (OS). RESULTS: Twenty patients were recruited to the study. Data showed that chemotherapy significantly reduced the number of circulating tumour cells as well as the circulating concentrations of Ang1, Ang2, VEGF-A, VEGF-C and VEGF-D from pre-treatment to cycle 2 day 2. The changes in circulating concentrations were not associated with PFS or OS. On average, the MRI perfusion/permeability parameter, Ktrans, increased in response to cytotoxic chemotherapy from pre-treatment to cycle 2 day 2 and this increase was associated with worse OS (HR 1.099, 95%CI 1.01-1.20, p = 0.025). CONCLUSIONS: In patients diagnosed with colorectal cancer with liver metastases, treatment with standard chemotherapy changes cell- and protein-based biomarkers, although these changes are not associated with survival outcomes. In contrast, the imaging biomarker, Ktrans, offers promise to direct molecularly targeted therapies such as anti-angiogenic agents.

4-Hydroxyisoleucine relieves inflammation through iRhom2-dependent pathway in co-cultured macrophages and adipocytes with LPS stimulation.

BACKGROUND: 4-Hydroxyisoleucine (4-HIL) is an active ingredient extracted from Trigonella foenum-graecum L., a Chinese traditional herbal medicine, which exerts the efficacy of anti-obesity and anti-diabetes. We previously reported that 4-HIL potentiates anti-inflammatory and anti-insulin resistance effects through down-regulation of TNF-α and TNF-α converting enzyme (TACE) in 3 T3-L1 adipocytes and HepG2 cells. In the present study, we further investigate the effects and mechanisms of 4-HIL on obesity-induced inflammation in RAW264.7 macrophages and 3 T3-L1 adipocytes co-culture system. METHODS: RAW264.7 macrophages and 3 T3-L1 adipocytes were co-cultured to mimic the microenvironment of adipose tissue. siRNA-iRhom2 transfection was performed to knockdown iRhom2 expression in RAW264.2 macrophages. The mRNA and protein expression of iRhom2 and TACE were measured by real-time quantitative PCR (RT-qPCR) and western blotting. The production of tumor necrosis factor-α (TNF-α), monocyte chemotactic protein-1 (MCP-1), IL-6 and IL-10 were evaluated by ELISA. The ratio of M2/M1 was detected by flow cytometry. RESULTS: 4-HIL significantly repressed the mRNA and protein levels of iRhom2 and TACE in RAW264.7 macrophages after LPS stimulated. Meanwhile, the levels of pro-inflammatory cytokines, including TNF-α, MCP-1, and IL-6, were substantially suppressed by 4-HIL in the co-culture system. Moreover, the level of anti-inflammatory cytokine IL-10 was increased significantly by 4-HIL in the co-culture system after LPS stimulation. Additionally, the ratio of M2/M1 was also increased by 4-HIL in the co-culture system after LPS stimulation. Finally, these effects of 4-HIL were largely enhanced by siRNA-iRhom2 transfection. CONCLUSION: Taken together, our results indicated that obesity-induced inflammation was potently relieved by 4-HIL, most likely through the iRhom2-dependent pathway.

Fenugreek attenuates obesity-induced inflammation and improves insulin resistance through downregulation of iRhom2/TACE.

AIMS: We previously reported that fenugreek-derived 4-hydroxyisoleucine ameliorates insulin resistance via regulation of TNF-α converting enzyme (TACE) expression. In the present study, we further investigate the effects and mechanisms of fenugreek on obesity-induced inflammation and insulin signaling in the high-fat diet (HFD)-challenged obese mice. MAIN METHODS: After 12 weeks of HFD intervention, mice were treated with the low or high dosages of fenugreek. Serum levels of glucose, insulin, lipid profile, inflammation cytokines, and adipokines were detected. Macrophage infiltration and adipose tissue morphology were observed. Western blot was conducted to investigate the expressions of inactive rhomboid 2 (iRhom2) and TACE as well as other signaling pathways in subcutaneous adipose tissue. KEY FINDINGS: We showed that fenugreek significantly suppressed body weight gain and fat accumulation in HFD-challenged obese mice. Meanwhile, fasting glucose, insulin, and HOMA-IR in fenugreek-treated mice were remarkably decreased, which were properly explained by fenugreek-induced activation of the insulin receptor signaling pathway. Moreover, the anti-inflammatory properties of fenugreek were shown by the decrease of systemic and local expressions of pro-inflammatory cytokines as well as reduced macrophage infiltration into adipose tissue. Additionally, fenugreek markedly deactivated NF-κB and JNK pathways. Finally, we demonstrated that fenugreek strikingly repressed the transcriptions and expressions of iRhom2 and TACE. SIGNIFICANCE: Fenugreek shows an encouraging and promising property in ameliorating insulin resistance and suppressing inflammation in obesity, which might be realized by fenugreek-mediated inhibition of iRhom2/TACE axis-facilitated TNF-α release from adipocytes.

Soporific Effect of Modified Suanzaoren Decoction and Its Effects on the Expression of CCK-8 and Orexin-A.

Suanzaoren decoction (SZRT), a classic Chinese herbal prescription, has been used as a treatment for insomnia for more than a thousand years. However, recent studies have found no significant effects of SZRT as a treatment for insomnia caused by gastric discomfort. Herein, we studied the effects of modified Suanzaoren decoction (MSZRD) on gastrointestinal disorder-related insomnia. The main constituents of MSZRD were spinosin (2.21 mg/g) and 6-feruloylspinosin (0.78 mg/g). A pentobarbital-induced animal model of insomnia showed that MSZRD shortened sleep latency and prolonged sleep time of the male Institute of Cancer Research (ICR) mice treated for 7 days with oral MSZRD. Sprague-Dawley male rats were treated daily with oral MSZRD or placebo for 11 days and then deprived of sleep for the last 4 days to establish a model of insomnia. Of note, MSZRD-treated animals had significantly improved body weight, organ index scores, and fecal moisture relative to placebo-treated animals, as well as reduced temperature. Sleep-deprived rats exhibited more exploratory behaviors in an open-field anxiety test; however, this effect was significantly reduced in MSZRD-treated animals. We found that MSZRD treatment decreased gastric acid pH, decreased the production of gastrin, pepsin, and Orexin-A, and increased the expression of MTL and CCK-8. Importantly, serum GABA concentration was increased by treatment with MSZRD, as reflected by a decreased Glu/GABA ratio. Treated animals had increased the expression of GAD1, GABARA1, and CCKBR but decreased the expression of Orexin R1. In summary, these results suggest that MSZRD has soporific and gastroprotective effects that may be mediated by differential expression of CCK-8 and Orexin-A.

Physicochemical characterization of polysaccharide from the leaf of Dendrobium officinale and effect on LPS induced damage in GES-1 cell.

The polysaccharide was first successfully isolated from the leaf of Dendrobium officinale by hot water extraction and alcohol precipitation and further purified using DEAE-52 and Sephadex G-100 chromatography. The structure of LDOP-1 was characterized by HPLC, GPC, and FT-IR and NMR spectroscopy, and its protective effect on LPS-induced GES-1 cell injury was analyzed. Results showed that LDOP was a homogeneous polysaccharide with average molecular weight of 91.8 kDa and consisted of Man, Gla, Glc, Glc acid, and Ara at a molar ratio of 2.0:1.3:1.6:1.7:0.7. LDOP had two types of residues, including 1,6-linked α-d-Glup and 1,4-linked α-d-Manp. Activity studies indicated that LDOP-1 can significantly suppress the release of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and IL-6 from LPS-induced GES-1 cell injury, decreased the protein expressions of TLR4, phospho-NF-κB, ASC, NLRP3, cleaved-IL-1ß, IL-6, and Bax, increased the protein expression of Bcl-2, and downregulated the ratios of cleaved caspase-1 to pro-caspase-1, phospho-IκBα to IκBα, and phospho-NF-кB to NF-κB. These findings strongly suggested that LDOP can prevent LPS-induced GES-1 cell injury by inhibiting the release of inflammatory cytokines regulated via the TLR4/NF-κB signal pathways.

Triptolide-induced hepatotoxicity can be alleviated when combined with Panax notoginseng saponins and Catapol.

ETHNOPHARMACOLOGICAL RELEVANCE: The hepatotoxicity of Tripterygium wilfordii Hook. f. (TW), due to the presence of triptolide (TP), limits its therapeutic potential. Based on the traditional Chinese medicine theory, the theory of "Yi lei xiang zhi" was proposed that Chinese herbs with different efficacy can restrict each other to achieve the least adverse reactions. AIM OF THE STUDY: To observe the effects of Catapol (CAT) and Panax notoginseng saponins (PNS), active ingredients in Rehmannia glutinosa (RG) and Panax notoginseng (PN) respectively, on reducing TP-induced hepatotoxicity, and further to explore the mechanisms. MATERIALS AND METHODS: The human hepatic cell line L-02 was cultured and treated with CAT, PNS or Combinations, and then treated with TP. The cytotoxic assay, the levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactate dehydrogenase (LDH), apoptosis, mitochondrial membrane potential and the expressions of NF-E2-related factor 1 (Nrf1) and its downstream targets were detected. Rats were treated with TP, TP + CAT, TP + PNS, or the combinations for 4 weeks. The levels of ALT, AST and LDH in serum, apoptosis of liver cells, mitochondria injury and the protein expressions of Caspase 3 and Nrf1 were investigated. RESULTS: CAT, PNS or CAT+PNS pre-treatment inhibited TP-induced toxicity in L-02 cells, distinctly decreased the apoptosis, alleviated the reduction of mitochondrial membrane potential, and modulated the expressions of Nrf1 and its downstream target, the mitochondrial transcription factor A (TFAM) and cytochrome C (Cyt-C). CAT, PNS or CAT+PNS inhibited the TP-induced hepatotoxicity in SD rats by reducing the mitochondria injury, decreasing the cells apoptosis and increasing the Nrf1 protein expression. Noticeably, TP + PNS + CAT combinations exhibited more effective than any single ingredient alone. CONCLUSION: PNS and CAT were able to effectively attenuate TP-induced hepatotoxicity. The efficiency benefits from their modulating Nrf1 and its downstream genes TFAM and Cyt-C, and further influencing mitochondrial functions and cells apoptosis. The combination is more effective than single ingredient alone.

Self-protection against triptolide-induced toxicity in human hepatic cells via Nrf2-ARE-NQO1 pathway.

OBJECTIVE: To find the signaling pathway of triptolide (TP)-induced liver injury and to reveal whether NF-E2-related factor 2 (Nrf2) plays an important role in cellular self-protection. METHODS: The L-02 and HepG2 cells were cultured and treated with various concentrations of TP. The cell viability was observed, and the cell medium was collected for detecting the aspartate aminotransferase (ALT), alanine aminotransferase (AST), lactate dehydrogenase (LDH), superoxide dismutase (SOD) and L-glutathione production (GSH) levels. Nrf2 and its downstream target NAD(P)H: quinine oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1) expression, the nuclear translocation of Nrf2, and the binding ability of Nrf2 and antioxidant response element (ARE) were also identified. Meanwhile, shRNA was used to silence Nrf2 in L-02 cells to find out whether Nrf2 plays a protective role. RESULTS: The viability of the L-02 and HepG2 cells treated with TP decreased in a doseand time-dependent manner, and TP (20-80 µg/mL) markedly induced the release of ALT, AST and LDH (P<0.05 or P<0.01), reduced the levels of SOD and GSH (P<0.01), and increased the intracellular reactive oxygen species. Meanwhile, TP augmented the Nrf2 expression in L-02 and HepG2 cells (P<0.05 or P<0.01), induced Nrf2 nuclear translocation, increased the Nrf2 ARE binding activity, and increased HO-1 and NQO1 expressions. Nrf2 knockdown revealed a more severe toxic effect of TP (P<0.05 or P<0.01). CONCLUSIONS: Human hepatic cells treated with TP induced oxidative stress, and led to cytotoxicity. Self-protection against TP-induced toxicity in human hepatic cells might be via Nrf2-ARE-NQO1 transcriptional pathway.

Inhibition mechanism of Qingluo Tongbi Granule () on osteoclast differentiation induced by synovial fibroblast and monocytes co-culture in adjuvant-induced arthritic rats.

OBJECTIVE: To study the mechanism underlying the inhibitory effect of Qingluo Tongbi Granule (, QTG) on osteoclast differentiation in rheumatoid arthritis in rats. METHODS: Fibroblast and monocyte co-culture were used to induce osteoclast differentiation in adjuvant-induced arthritic (AIA) rats. Serum containing QTG was prepared and added to the osteoclasts, and activation of the tumor necrosis factor receptor-associated factor 6/mitogen-activated protein kinase/nuclear factor of activated T cells, cytoplasmic1 (TRAF6/MAPK/NFATc1) pathways was examined. RESULTS: The induced osteoclasts were multinucleated and stained positive for tartrate-resistant acid phosphatase (TRAP) staining. Serum containing QTG at 14.4, 7.2 or 3.6 g/kg inhibited the activation of TRAF6, extracellular regulated protein kinase (ERK)1/2, c-Jun N-terminal kinase (JNK) and p38 and decreased the percentage of cells with nuclear NFATc1 in a dose-dependent manner, the high and middle doses exhibited clear inhibitory activity (P<0.01 and P<0.05, respectively). After the addition of MAPK inhibitors, the NFATc1 expression showed no significant difference compared with the control group (P>0.05). CONCLUSIONS: Serum containing QTG could generally inhibit the TRAF6/MAPK pathways and possibly inhibit the NFATc1 pathway. In addition, QTG may regulate other signaling pathways that are related to osteoclast differentiation and maturation.

Structures of yeast Apa2 reveal catalytic insights into a canonical AP4A phosphorylase of the histidine triad superfamily.

The homeostasis of intracellular diadenosine 5',5″'-P(1),P(4)-tetraphosphate (Ap4A) in the yeast Saccharomyces cerevisiae is maintained by two 60% sequence-identical paralogs of Ap4A phosphorylases (Apa1 and Apa2). Enzymatic assays show that, compared to Apa1, Apa2 has a relatively higher phosphorylase activity towards Ap3A (5',5″'-P(1),P(3)-tetraphosphate), Ap4A, and Ap5A (5',5″'-P(1),P(5)-tetraphosphate), and Ap4A is the favorable substrate for both enzymes. To decipher the catalytic insights, we determined the crystal structures of Apa2 in the apo-, AMP-, and Ap4A-complexed forms at 2.30, 2.80, and 2.70Å resolution, respectively. Apa2 is an α/ß protein with a core domain of a twisted eight-stranded antiparallel ß-sheet flanked by several α-helices, similar to the galactose-1-phosphate uridylyltransferase (GalT) members of the histidine triad (HIT) superfamily. However, a unique auxiliary domain enables an individual Apa2 monomer to possess an intact substrate-binding cleft, which is distinct from previously reported dimeric GalT proteins. This cleft is perfectly complementary to the favorable substrate Ap4A, the AMP and ATP moieties of which are perpendicular to each other, leaving the α-phosphate group exposed at the sharp turn against the catalytic residue His161. Structural comparisons combined with site-directed mutagenesis and activity assays enable us to define the key residues for catalysis. Furthermore, multiple-sequence alignment reveals that Apa2 and homologs represent canonical Ap4A phosphorylases, which could be grouped as a unique branch in the GalT family.

[Chuzhen therapy for sub-health: a randomized controlled study].

OBJECTIVE: To observe the effect on intervention of sub-health with pestle needle (Chuzhen). METHOD: Randomized controlled trail was adopted for this research. One hundred and fifty-three cases were randomly divided into two groups of a Chuzhen group (79 cases) and a massage group (74 cases). Acupoint of Bazhen (Baihui Bazhen, Shendao Bazhen, Zhiyang Bazhen, Mingmen Bazhen, Yaoyangguan Bazhen), Hechelu on the head, the neck and the lumbar area were adopted in Chuzhen group. While regular whole-body massage was applied in the massage group. The human sub-health score, the cornell medical index (CMI) and thermal texture maps system (TTM) technology of the two groups before and after the intervention were observed. RESULTS: 1) After treatment, sub-health condition score, the CMI score, the M-R score and the TTM index were all increased in both groups (all P<0.01) 2) Comparison of D-value of the two groups before and after the intervention: the level of the sub-health score, the total score of CMI, and the index of sleep, pressure, Governor Vessel, Hukou (first web), blood lipid, viscosity of blood, microcirculation of TTM index of the Chuzhen group changed more obvious (all P<0.01), but there was no statistic significances in the M-R score and blood sugar of the TTM (both P>0.05). 3) The sub-health condition score in Chuzhen group was higher than that in the massage group (P<0.01). CONCLUSION: Chuzhen therapy has definite effect on intervention of sub-health, which is better than regular general massage.