Parishin treatment alleviates cardiac aging in naturally aged mice.

Background: Cardiac aging progressively decreases physiological function and drives chronic/degenerative aging-related heart diseases. Therefore, it is crucial to postpone the aging process of heart and create products that combat aging. Aims & methods: The objective of this study is to examine the effects of parishin, a phenolic glucoside isolated from traditional Chinese medicine Gastrodia elata, on anti-aging and its underlying mechanism. To assess the senescent biomarkers, cardiac function, cardiac weight/body weight ratio, cardiac transcriptomic changes, and cardiac histopathological features, heart tissue samples were obtained from young mice (12 weeks), aged mice (19 months) treated with parishin, and aged mice that were not treated. Results: Parishin treatment improved cardiac function, ameliorated aging-induced cardiac injury, hypertrophy, and fibrosis, decreased cardiac senescence biomarkers p16Ink4a, p21Cip1, and IL-6, and increased the "longevity factor" SIRT1 expression in heart tissue. Furthermore, the transcriptomic analysis demonstrated that parishin treatment alleviated the cardiac aging-related Gja1 downregulation and Cyp2e1, Ccna2, Cdca3, and Fgf12 upregulation in the heart tissues. The correlation analysis suggested a strong connection between the anti-aging effect of parishin and its regulation of gut microbiota and metabolism in the aged intestine. Conclusion: The present study demonstrates the protective role and underlying mechanism of parishin against cardiac aging in naturally aged mice.

Exploring the therapeutic mechanism of Banxia Xiexin Decoction in mild cognitive impairment and diabetes mellitus: a network pharmacology approach.

The incidence of mild cognitive impairment (MCI) and diabetes mellitus (DM) is increasing year by year. Clinical findings show that Banxia Xiexin Decoction (BXD) can be combined to treat MCI and DM. However, the principle and mechanism of BXD in treating MCI and DM remain unclear. In this study, to explore the common mechanism of BXD in treating MCI and DM by using the method of network pharmacology. Traditional Chinese Medicine Systems Pharmacology Database (TCMSP) was used to screen the main active components of BXD, as well as to predict and screen its potential targets. Using Online Mendelian Inheritance in Man (OMIM), Therapeutic Target Database (TTD), DisGeNET, GeneCards to select the target proteins of two diseases, and intersecting the drug target and the disease target to obtain the common target of drug diseases, which is imported into cytoscape software to draw the network diagram of "drug components-target diseases" and the interaction network diagram between the common target proteins. According to the Database for Annotation, Visualization and Integrated Discovery (DAVID) database, we analyzed the common targets using two methods, gene ontology Kyoto Encyclopedia of Genes and Genomes (KEGG) biological pathway enrichment analysis and Gene Ontology (GO) function enrichment analysis, as well as studied the interaction mechanism of the two diseases, with the results validated using molecular docking. A total of 267 main active components of BXD were screened, together with the two diseases shared 233 common targets. The top five key targets identified by the topological analysis were TP53, AKT1, STAT3, TNF, and MAPK3. Go enrichment results indicated that it was primarily related to response to drug, extracellular space, enzyme binding, RNA polymerase II transcription factor activity, ligand-activated sequence-specific DNA binding. t KEGG enrichment pathway analysis identified 20 significant pathways, the majority of which are AGE-RAGE signaling pathways in diabetic complications, lipid and atherosclerosis, fluid shear stress and atherosclerosis, IL-17 signaling pathway, TNF signaling pathway, and so on. The results of molecular docking revealed that the key components of BXD, baicalein, licochalcone a, quercetin, and naringenin, had strong binding ability with core targets TP53, AKT1, STAT3, TNF, MAPK3. BXD can treat MCI and DM by multi-targets and multi-channels,and plays a role of "homotherapy for heteropathy" mainly through response to drug, positive regulation of gene expression, extracellular space and enzyme binding and other ways.

Effectiveness and safety analysis of SanHanHuaShi granules for the treatment of coronavirus disease 2019: Study protocol and statistical analysis plan for a randomized, parallel-controlled, open-label clinical trial.

Background: Coronavirus disease 2019 (COVID-19) was declared a global pandemic in March 2020 by the World Health Organization (WHO). As of July 2, 2022, COVID-19 has caused more than 545 million infections and 6.3 million deaths worldwide, posing a significant threat to human health. Currently, there is still a lack of effective prevention and control strategies for the variation and transmission of SARS-CoV-2. Traditional Chinese medicine (TCM), which has a unique theoretical system, has treated various conditions for thousands of years. Importantly, recent studies have revealed that TCM contributed significantly to COVID-19. SanHanHuaShi (SHHS) granules, a Chinese herbal medicine, which has been included in Protocol for the Diagnosis and Treatment of Novel Coronavirus Disease 2019 (6th to 9th editions) issued by the National Health Commission of China and used to prevent and treat COVID-19 disease. A previous retrospective cohort study showed that SHHS could significantly reduce the severity of mild and moderate COVID-19. However, there is an absence of high-quality randomized controlled clinical studies to confirm the clinical effectiveness of SHHS. Therefore, a clinical study protocol and a statistical analysis plan were designed to investigate the efficacy and safety of SHHS for the prevention and treatment of COVID-19. This study will increase the integrity and data transparency of the clinical research process, which is of great significance for improving the practical application of SHHS granules in the future. Methods and analysis: The study was designed as a 7-day, randomized, parallel controlled, open-label, noninferiority clinical trial of positive drugs. A total of 240 patients with mild and moderate COVID-19 will be enrolled and randomly assigned to receive SanHanHuaShi granules or LianHuaQingWen granules treatment in a 1:1 ratio. Disease classification, vital signs, SARS-CoV-2 nucleic acid testing, symptoms, medications, adverse events, and safety evaluations will be recorded at each visit. The primary outcome will be the clinical symptom recovery rate. Secondary outcomes will include the recovery time of clinical symptoms, negative conversion time of SARS-CoV-2 nucleic acid test negative conversion rate, hospitalization time, antipyretic time, rate of conversion to severe patients, and time and rate of single symptom recovery. Adverse incidents and safety assessments will be documented. All data will be analyzed using a predetermined statistical analysis plan, including our method for imputation of missing data, primary and secondary outcome analyses, and safety outcomes. Discussion: The results of this study will provide robust evidence to confirm the effectiveness and safety of SHHS in the treatment of COVID-19. Clinical Trial Registration: http://www.chictr.org.cn. Trial number: ChiCTR2200058080. Registered on 29 March 2022.

Role of gut microbiota-derived branched-chain amino acids in the pathogenesis of Parkinson's disease: An animal study.

Neuroinflammation caused by the disorder of gut microbiota and its metabolites is associated with the pathogenesis of Parkinson's disease (PD). Thus, it is necessary to identify certain molecules derived from gut microbiota to verify whether they could become intervention targets for the treatment of PD. The branched-chain amino acids (BCAAs), as a common dietary supplement, could modulate brain function. Herein, we investigated the longitudinal shifts of microbial community in mice treated with rotenone for 0, 3 and 4 weeks by 16S rRNA gene sequencing to identify the microbial markers at different PD stages. Serum BCAAs were determined by gas chromatography-mass spectrometry. Then, rotenone-induced mice were given a high BCAA diet to evaluate the motor and non-motor functions, dopaminergic neuron loss, and inflammation levels. Using a PD mouse model, we discovered that during PD progression, the alterations of gut microbiota compositions led to the peripheral decrease of BCAAs. Based on the serum lipopolysaccharide binding protein concentrations and the levels of pro-inflammatory factors (including tumor necrosis factor-α, interleukin [IL]-1ß, and IL-6) in the colon and substantia nigra, we found that the high BCAA diet could attenuate the inflammatory levels in PD mice, and reverse motor and non-motor dysfunctions and dopaminergic neuron impairment. Together, our results emphasize the dynamic changes of gut microbiota and BCAA metabolism and propose a novel strategy for PD therapy: a high BCAA diet intervention could improve PD progression by regulating the levels of inflammation.

An affinity interaction guided two-dimensional separation system for the screening of neuraminidase inhibitors from Reynoutria japonica Houtt. roots.

Discovering bioactive compounds from medicinal herbs is crucial for drug discovery. Ultrafiltration is often used in the screening of bioactive compounds from natural herbs because of its simple and rapid operations. However, the ultrafiltration results are often disturbed by the undissolved compounds and the non-target compounds, which reduces the accuracy of the results. Herein, an affinity interaction guided two-dimensional (2D) separation system was developed. Discovery of the potential neuraminidase (NA) inhibitors from the dried roots of Reynoutria japonica Houtt. (RRJ) was used as an example. Only the small molecules showing affinity interaction with NA could be screened by the affinity interaction guided 2D separation system. Firstly, the NA and crude extract were incubated to form a sample solution (containing NA-inhibitor complexes, NA, and three types of small molecules with different polarities) by affinity interaction. Then the sample solution was separated and detected by the 2D separation system. This aimed to reduce the interference of the undissolved compounds and non-target compounds, and pick out the NA-inhibitor complexes (NA-Is). The collected NA-Is were denatured to release small molecular inhibitors (Is) for LC-MS/MS analysis. Compared with the ultrafiltration, more obvious peak area differences were observed in the results, and four potential NA inhibitors were successfully identified. In all, we provided a simple strategy with better performance in the screening of natural bioactive compounds.

Effects of two types nitrogen sources on humification processes and phosphorus dynamics during the aerobic composting of spent mushroom substrate.

Aerobic composting is increasingly regarded as a promising technology for the recycling of spent mushroom substrate (SMS), and an applicable nitrogen source is necessary to improve the process. This study is the first to investigate the effects of protein-like N source (chicken manure, CM) and high-N source (urea, UR) on humification process and P dynamics during SMS composting. The effect of different N sources on microbial succession was also studied. Results showed that CM addition achieved a longer thermophilic phase (16 d vs 9 d), greater germination indices (131.6% vs 106.3%), and higher total phosphorus content (13.1 g/kg vs 6.56 g/kg) in the end products, as compared to UR. The addition of CM showed beneficial effects on humification and stabilization, including decreased weight loss and fluctuations in the level of functional groups. The P produced in the compost was interconverted and leached in the P pool. In this case, the P detected in the compost was in the form of orthophosphate and MgNH4PO4⋅6H2O crystal as inorganic P and orthophosphate monoester as organic P. The most abundant microorganisms at the phylum level mainly include Firmicutes, Actinobacteria, and Proteobacteria, accounting for more than 88% of the total microorganisms. The addition of CM to SMS compost resulted in higher organic matter degradation rates. This work clarified the role of various N sources in SMS composting and presented an appropriate waste management method beneficial to bioresource technology and sustainable development of the edible fungi business.

The Protective Effect of Ethyl Acetate and n-Butanol Fractions of Wine-Steamed Ligustri Lucidi Fructus on Diabetic Nephropathy in Rats.

Ligustri Lucidi Fructus (LLF), the dry and ripe fruit of Ligustrum lucidum W. T. Aiton (Oleaceae), is a traditional Chinese medicine for nourishing the liver and kidney in clinics for thousands of years. Wine-steamed Ligustri Lucidi Fructus (WLL) can alleviate coolness and smoothness of LLF and enhance the function of nourishing the liver and kidney, so ancient and modern medicine usually used it in clinics. First of all, we prepared the extracts of different polar fractions of WLL to explore the effective fractions and potential mechanisms of WLL in the treatment of diabetic nephropathy (DN). Then, HPLC method was used to determine the contents of 12 active components in WLL and its different polar components. Finally, the potential relationship between 12 active components and physicochemical parameters of DN rats was explored. The pharmacological experiments showed that WLL, ethyl acetate (EtOAc), and n-butanol (n-BuOH) extracts not only significantly alleviated the clinical symptoms and kidney damage of DN rats but also had obvious anti-inflammatory and antioxidant effects. In addition, the results of HPLC analysis showed that the 12 active components of WLL mainly existed in the extracts of EtOAc and n-BuOH. The Pearson correlation analysis showed 12 active components and physicochemical parameters had different degrees of correlation. In conclusion, we proved that the extracts of EtOAc and n-BuOH were the effective fractions of WLL in treating DN in rats, and they could regulate the levels of inflammatory cytokines and decrease oxidation stress, which provides a basis for further research on the mechanism of WLL in treating DN and provides a pharmacological and chemical foundation for the development of new anti-DN drugs.

Music Therapy Alleviates Motor Dysfunction in Rats With Focal Cerebral Ischemia-Reperfusion Injury by Regulating BDNF Expression.

Background/Aim: Music-based therapy plays a role in central nervous system diseases. We aimed to explore the effect of different doses and durations of music therapy on motor function recovery after stroke and the underlying molecular mechanisms. Methods: Adult male Sprague-Dawley rats were subjected to middle cerebral artery occlusion (MCAO) for 1 h, which was followed by reperfusion. In experiment 1, the rats that survived 1 week after MCAO surgery were randomly allocated into four groups (n = 10 per group): MCAO group, 1 h music group (Mozart K.448 music therapy 1 h per day for 2 weeks), 12 h music group (Mozart K.448 music therapy 12 h/day for 2 weeks), and accelerated music group (reversely accelerated music therapy 12 h for 2 weeks, AM group). In experiment 2, the survived rats were randomly divied into three groups: MCAO group, 12 h music group (music therapy 12 h/day for 3 weeks), and 12 h music-R group (music therapy 12 h/day for 2 weeks and rest for 1 week). Three neuroscores were evaluated daily, starting on the first day after surgery until the end of the experiment. The rats were killed 3 weeks after MCAO surgery in experiment 1 or 4 weeks after surgery in experiment 2. Nissl staining of infart core, peri-infarct zone, and motor cortex was performed to assess neuronal survival and regeneration. Western blot and immunofluorescence were used to detect the expression and distribution of brain-derived neurotrophic factor (BDNF) and glial fibrillary acidic protein (GFAP) in ipsilateral hemispheres. Results: In the experiment of different music therapy doses, the motor function in the 12-h music group but not in the 1-h music group and AM group was significantly improved compared with that of the MCAO group. The BDNF protein level of the ipsilateral hemisphere motor cortex in the 12-h music group and the 1-h music group was higher than that of the MCAO group. The neurons and Nissl bodies were more in the 12-h music group than in the MCAO group. Immunofluorescence assay showed that a 12 h music therapy induces BDNF and GFAP accumulation at the damage boundary. In the experiment of different music therapy durations, 3 weeks music therapy (12 h music group) induced more longer cell synapses and more clearer cell-to-cell connections than 2 weeks music intervention (12 h music-R group). Moreover, the GFAP morphology in the 12-h music group was more similar to mature activated astrocytes than that in the 12-h music-R group. Conclusions: Music therapy may improve poststroke motor function and promote neuronal repair in the long term. The mechanism may be through stimulating BDNF and GFAP secretion in the injured motor cortex.

Analysis of chemical variations between raw and wine-processed Ligustri Lucidi Fructus by ultra-high-performance liquid chromatography-Q-Exactive Orbitrap/MS combined with multivariate statistical analysis approach.

Ligustri Lucidi Fructus (LLF) is the dried and mature fruit of Ligubtrum lucidum Ait., which has the effect of nourishing the liver and kidney, brightening the eyes and promoting the growth of black hair. Wine-processed LLF is commonly used in traditional Chinese medicine; however, the processing mechanisms are still unclear. Herein, a system data acquisition and mining strategy was designed to investigate the chemical profile differences between the raw and wine-processed LLF, based on high-performance liquid chromatography-Orbitrap high resolution mass spectrometry coupled with multivariate statistical analysis including principal component analysis and partial least square analysis. Afterwars, a total of 55 components were found to be the main contributors to the significant difference between raw and wine-processed LLF by comparison with chromatographic behaviors, intact precursor ions, and characteristic MS fragmentation patterns. In addition, 10 main constituents of raw and wine-processed LLF were simultaneously determined by UHPLC-MS/MS for analyzing the content variations. Some structural transformation mechanisms during wine processing were deduced from the results. The results may provide a scientific foundation for deeply elucidating the wine-processing mechanism of LLF.

Pharmacokinetic comparison of nine bioactive components in rat plasma following oral administration of raw and wine-processed Ligustri Lucidi Fructus by ultra-high-performance liquid chromatography coupled with triple quadrupole mass spectrometry.

An accurate and sensitive ultra-high-performance liquid chromatography coupled with triple quadrupole mass spectrometry method was established and validated for the determination of nine bioactive compounds of Ligustri Lucidi Fructus in rat plasma. Separation was performed on Halo® C18 column with a mobile phase of acetonitrile and 0.1% formic acid in water. The eluate was detected by multiple reaction monitoring scanning operating in the negative ionization mode. This assay method was validated for selectivity, linearity, intra- and interday precision, accuracy, recovery, matrix effect, and stability, and all methodological parameters fulfilled the Food and Drug Administration criteria for bioanalytical validation. The established method was successfully applied to a comparative pharmacokinetic study of raw and wine-processed Ligustri Lucidi Fructus in rats for the first time. It was found that the AUC0-24 and Cmax value of salidroside, hydroxytyrosol, and nuezhenidic acid were increased significantly after processing, while the AUC0-24 and Cmax value of oleoside 11-methyl ester, 1'''-O-ß-d-glucosylformoside, specnuezhenide, G13, oleonuezhenide, and oleanolic acid were decreased, which suggested that processing affects the absorption and bioavailability of Ligustri Lucidi Fructus. The results might be valuable for the clinical reasonable application and understanding the processing mechanism of Ligustri Lucidi Fructus.

Research progress on nanotechnology for delivery of active ingredients from traditional Chinese medicines.

There is growing acceptance of traditional Chinese medicines (TCMs) as potential sources of clinical agents based on the demonstrated efficacies of numerous bioactive compounds first identified in TCM extracts, such as paclitaxel, camptothecin, and artemisinin. However, there are several challenges to achieving the full clinical potential of many TCMs, particularly the generally high hydrophobicity and low bioavailability. Recently, however, numerous studies have attempted to circumvent the limited in vivo activity and systemic toxicity of TCM ingredients by incorporation into nanoparticle-based delivery systems. Many of these formulations demonstrate improved bioavailability, enhanced tissue targeting, and greater in vivo stability compared to the native compound. This review summarizes nanoformulations of the most promising and extensively studied TCM compounds to provide a reference for further research. Combining these natural compounds with nanotechnology-based delivery systems may further improve the clinical utility of these agents, in turn leading to more intensive research on traditional medicinal compounds.

Vancomycin-resistant Enterococcus faecium pneumonia in a uremic patient on hemodialysis: a case report and review of the literature.

BACKGROUND: Even though enterococci can cause serious infections in multiple sites, they are a rare cause of pneumonia. We reported a uremic patient with vancomycin-resistant E. faecium (VRE-fm) pneumonia, possibly related to epileptic seizures. CASE PRESENTATION: A 57-year old man with uremia on hemodialysis was admitted to the hospital with complaint of recurrent epileptic seizures, followed by a two-week history of recurrent fever and cough with purulent sputum. Chest CT demonstrated multiple exudation of both lungs. He was diagnosed as community acquired pneumonia. Despite antibiotic combination therapy, abnormal chest shadows aggravated. Sputum and blood cultures were initially negative, but later blood culture grew VRE-fm. We suspected aspiration of gastrointestinal content induced by epilepsy as the most likely mechanism. The patient was successfully treated with a four-week course of linezolid according to the antibiotic susceptibility testing. CONCLUSIONS: Physicians should consider multi-drug resistant organisms such as VRE in uremic patients with pneumonia that fails to resolve with broad-spectrum antibiotics, especially in the cases with aspiration induced by epilepsy, immunocompromised conditions, and repeated or prolonged hospitalizations.

Short Communication: Fructose-Enhanced Antibacterial Activity of Self-Assembled Nano-Peptide Amphiphiles for Treating Antibiotic-Resistant Bacteria.

BACKGROUND: In recent years, numerous bacteria have become resistant to conventional antibiotics. Fortunately, an increasing body of research indicates that through the addition of specific metabolites (like sugars), the antibacterial activity of certain drugs can be enhanced. A new type of self-assembled nano-peptide amphiphile (SANPA) was designed in this study to treat antibiotic-resistant bacterial infections and to reduce the use of antibiotics. METHODS: Here, SANPAs were self-assembled into nanorod structures with a diameter of ca. 10.5 nm at concentrations greater than the critical micelle concentration (CMC) of 44.67 µM. Both Gram-positive and Gram-negative bacteria were treated with SANPAs with fructose supplementation. RESULTS: After a 30-min fructose pre-incubation, SANPAs reduced bacteria growth relative to non-fructose treatments at all concentrations. Cytotoxicity assays indicated that the presence of fructose seemed to slightly ameliorate the cytotoxic effect of the treatment on model human fetal osteoblasts (or bone-forming cells) and human dermal fibroblasts. CONCLUSION: We demonstrated here that SANPAs-like nanomaterials have a promising potential to treat antibiotic-resistant bacteria, especially when added to fructose, potentially limiting their associated infections.

Analysis of variations in the contents of steroidal saponins in Fructus Tribuli during stir-frying treatment.

Just as natural saponins transform into aglycones, secondary glycosides and their derivatives using biotransformation technology, steroidal saponins may also undergo similar transformation after stir-frying. The purpose of this study was to elucidate the variations and the reasons for these variations in the contents of steroidal saponins in Fructus Tribuli (FT) during a stir-frying treatment. Stir-fried FT was processed in different time-temperature conditions. An UHPLC-MS/MS method was established and fully validated for quantitative analysis. In addition, the simulation processing products of tribuluside A, terrestroside B, terrestrosin K, terrestrosin D and 25R-tribulosin were determined by qualitative analysis using UHPLC-Q-TOF-MS. The established UHPLC-MS/MS method provides a rapid, flexible, and reliable method for the quality assessment of FT. The present study revealed that furostanol saponins with a C22-OH group could transform into corresponding furostanol saponins with a C-20-C-22 double bond (FSDB) via dehydroxylation. Additionally, FSDB could be successively converted into its secondary glycosides via a deglycosylation reaction. The transformation of spirostanol saponins into corresponding aglycones via deglycosylation led to a decrease in spirostanol saponins and an increase in aglycones. The results of this research provided scientific evidence of variation and structural transformation among steroidal saponins. These findings might be helpful for elucidating the processing mechanism of FT.

Carbon sources derived from maize cobs enhanced nitrogen removal in saline constructed wetland microcosms treating mariculture effluents under greenhouse condition.

This study investigated an alternative carbon source derived from maize cobs (MCs) to enhance nitrogen removal in saline constructed wetlands (SCWs). The main objectives were to select the proper pretreatment method of MCs for rapid carbon release; and to investigate the effects of maize cob pieces (i.e. MCP) and three addition levels of maize cob lixiviums (i.e. L-MCL, M-MCL and H-MCL) on nitrogen purification performance and microbial characteristics of SCWs. Results showed NaOH pretreatment enhanced carbon release of MCs in seawater (from 7.5 ± 0.4 mgCOD g-1 to 16.4 ± 0.2 mgCOD g-1). The 80-d trial showed SCWs with M-MCL addition performed well on nitrogen removal: NO3-N, 88.8 ± 11.6%; NO2-N, 91.1 ± 3.5%; TAN, 96.5 ± 1.6%; TIN, 89.8 ± 10.4%; with 2 mg L-1 effluent COD. Denitrification parameters confirmed MCL to be a high quality carbon source: denitrification potential (PDN) = 0.16 gN gCOD-1; heterotrophy anoxic yield coefficient (YH) = 0.54 gCOD gCOD-1. The MCP and H-MCL treatments improved substrate dehydrogenase activity, indicating a higher microbial activity in these SCWs. Sequencing analysis revealed that, regardless of addition manners, carbon sources from MCs changed the rhizosphere microbial community. At genus level, Anaerophaga (10.1%), Granulosicoccus (8.2%) and Sulfurimonas (6.6%) dominated in SCWs under MCP treatment. Increased MCL addition levels improved the relative abundance of Vibrio, Malonomonas and Caldithrix, suggesting the enhancement of denitrification. Relative high proportions of Desulfotignum and Desulfovibrio, and Sulfurimonas were observed in MCP and H-MCL SCWs, implying that sulfate reduction occurred in SCWs with excess carbon sources.

Study on steroidal saponins in crude and stir-fried Fructus Tribuli by ultra-high-performance liquid chromatography-mass spectrometry coupled with multivariate statistical analysis.

Fructus Tribuli is a traditional Chinese medicine used clinically for many years. Crude Fructus Tribuli and stir-fried Fructus Tribuli are recorded in the Pharmacopoeia of the People's Republic of China. However, the differences between steroidal saponins in crude Fructus Tribuli and stir-fried Fructus Tribuli have not been compared. In this study, ultra-high-performance liquid chromatography quadrupole time-of-flight mass spectrometry along with multivariate statistical analysis was developed to discriminate the chemical profiles and identify the steroidal saponins of crude Fructus Tribuli and stir-fried Fructus Tribuli. Additionally, an ultra-high-performance liquid chromatography triple-quadrupole mass spectrometer was used for the simultaneous quantification of nine major steroidal saponins to analyze the variations between crude Fructus Tribuli and stir-fried Fructus Tribuli. Finally, a total of 30 steroidal saponins whose structures or contents changed significantly after processing were found and identified. The mechanism of structural transformations deduced indicated that during the stir-frying of Fructus Tribuli, C-22 hydroxy furostanol saponins were converted to the corresponding furostanol saponins containing C-20-C-22 double bonds by dehydroxylation and deglycosylation reactions that occurred in the spirostanol saponins causing the generation of steroidal sapogenins. This study was successfully applied to the global analysis of crude Fructus Tribuli and stir-fried Fructus Tribuli. The results of this research will be beneficial to explore the processing mechanism of Fructus Tribuli.

Curcumin Inhibits Joint Contracture through PTEN Demethylation and Targeting PI3K/Akt/mTOR Pathway in Myofibroblasts from Human Joint Capsule.

Joint contracture is increasingly regarded as a clinical problem that leads to irreversible dysfunction of the joint. It is a pathophysiological process following joint injury, which is marked by the activation of myofibroblasts. There is currently no effective treatment for the prevention of joint contracture. Curcumin is a polyphenol pigment extracted from turmeric, which possesses anti-inflammatory, antioxidative, and antitumor properties. In the present study, we demonstrated that curcumin exerts a protective effect against joint contracture via the inhibition of myofibroblast proliferation and migration in a time- and concentration-dependent manner. Moreover, we indicated that phosphatase and tension homolog (PTEN) was downregulated in myofibroblasts in vitro and in the contracture capsule tissues of patients in vivo. Additionally, western blot analysis revealed a negative correlation between the expression levels of PTEN and the fibrosis marker protein alpha smooth muscle cell actin. Methylation-specific PCR results suggested that curcumin was able to demethylate PTEN in a similar manner to the demethylation agent 5-azacytidine, increasing PTEN expression and further inhibiting phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin signaling. In conclusion, our data illustrate part of the mechanism of curcumin inhibition in joint contracture. These results support the hypothesis that curcumin may potentially be used as a novel candidate for the treatment of joint contracture.

An aptamer assay for aflatoxin B1 detection using Mg2+ mediated free zone capillary electrophoresis coupled with laser induced fluorescence.

We described an aptamer based and Mg2+ mediated free zone capillary electrophoresis-laser induced fluorescence (CE-LIF) assay for aflatoxin B1 (AFB1) detection. This CE-LIF assay applied an anti-AFB1 aptamer with a single fluorescein (FAM) label at 5' end and a short complementary DNA (cDNA). In the absence of AFB1, the cDNA hybridized with the aptamer probe and formed a duplex DNA. The use of running buffer containing MgCl2 allowed good isolation of the duplex DNA from the single stranded DNA in CE. We found introducing a biotin label on the cDNA further improved the isolation. When AFB1 existed in sample solution, the aptamer probe bound with AFB1, dissociating from the duplex DNA. Thus, the duplex DNA peak decreased, while the aptamer probe peak increased during CE-LIF analysis. We achieved detection of AFB1 by measuring the aptamer probe peak. The length of cDNA, the ratio of aptamer to cDNA, and the concentration of MgCl2 in sample buffer and separation buffer had great effect on the aptamer based CE-LIF assay. Under optimized conditions, the detection limit of AFB1 was 0.2 nM, and the dynamic range was from 0.2 nM to 500 nM. Limit of quantitation was 0.5 nM. This CE-LIF assay enabled detection of AFB1 spiked in diluted human serum, diluted human urine, and corn flour samples. This assay exhibits potential for wide application as it integrates the rapidity, high sensitivity, low sample consumption of CE-LIF analysis and the strengths of aptamer.

Aptamer-Structure Switch Coupled with Horseradish Peroxidase Labeling on a Microplate for the Sensitive Detection of Small Molecules.

Detection of small molecules with good sensitivity, high throughput, simplicity, and generality using aptamers is desired but still remains challenging. We described an aptamer-structure-switch assay coupled with horseradish peroxidase (HRP) labeling on microplates for sensitive absorbance and chemiluminescence detection of small molecules. This assay relies on competition for affinity binding to a limited HRP-labeled aptamer between small-molecule targets and immobilized short DNA strands complementary to the aptamer (cDNA) on a microplate. In the absence of targets, the HRP-labeled aptamer hybridizes with the cDNA on the microplate, and HRP catalyzes substrate into product, generating absorbance or chemiluminescence signals. The binding of small-molecule targets to aptamers causes displacement of HRP-labeled aptamers from the cDNA and signal decrease. In chemiluminescence-analysis mode, the assay achieved detection of aflatoxin B1 (AFB1), ochratoxin A (OTA), and adenosine triphosphate (ATP) with detection limits of 10 pM, 20 pM, and 20 nM, respectively. This assay does not require enzyme-labeled small molecules or the conjugation of small molecules on solid phase. HRP, as an enzyme label, here allows for easily obtainable and highly active signal amplification. This microplate assay is rapid and promising for high-throughput analysis. It shows potential for wide applications in the detection of small molecules.

Effects of pyrroloquinoline quinone supplementation on growth performance and small intestine characteristics in weaned pigs.

This study was conducted to explore the effect of graded levels of pyrroloquinoline quinone disodium (PQQ·Na2) on the performance and intestinal development of weaned pigs. A total of 216 pigs weaned at 28 d were assigned in a randomized complete block design to 6 diets containing 0, 1.5, 3.0, 4.5, 6.0, or 7.5 mg/kg PQQ·Na2 for 28 d. Performance, diarrhea incidence, intestinal morphology, redox status, cytokines, and the expression of tight junction proteins were determined. Pigs had increased ADG (linear, P < 0.01), G:F (quadratic, P < 0.01), and lower diarrhea incidence (P < 0.01) with the increase of PQQ·Na2 supplementation. Villus height increased (quadratic, P < 0.01) in all segments of the small intestine, and crypt depth in the duodenum and jejunum was decreased (linear, P < 0.05) in pigs with the increase of PQQ·Na2 supplementation. Pigs fed PQQ·Na2-supplemented diets had higher (P < 0.05) activities of antioxidant enzymes including total superoxide dismutase in duodenum, jejunum, and ileum; glutathione peroxidase (GSH-Px) in jejunum and ileum; catalase (CAT) in duodenum and ileum; and lower (P < 0.05) malondialdehyde concentrations in the intestinal mucosa of all segments. In the intestinal mucosa, cytokines including interleukin (IL)-1ß, IL-2, and interferon-γ were significantly decreased (P < 0.05) in pigs fed PQQ·Na2-supplemented diets. The protein expression of zonula occluden protein-1 (ZO-1) and occludin in the jejunum was significantly increased (P < 0.05) in pigs fed diets containing PQQ·Na2. In conclusion, these results have indicated that dietary PQQ·Na2 supplementation improves growth performance and gut health in weaned pigs. Moreover, pigs fed diet with as low as 1.5-mg/kg PQQ·Na2 have better performance compared with pigs fed no PQQ·Na2-supplemented diet; pigs fed diet with 4.5-mg/kg PQQ·Na2 have highest G:F among treatments during the whole period.