Integrating Transcriptome and Chemical Analyses to Provide Insights into Biosynthesis of Terpenoids and Flavonoids in the Medicinal Industrial Crop Andrographis paniculate and Its Antiviral Medicinal Parts.

Andrographis paniculata is a medicinal plant traditionally used to produce diterpene lactones and flavonoids, which possess various biological activities. Widely distributed in China, India, and other Southeast Asia countries, A. paniculata has become an important economic crop, significantly treating SARS-CoV-2, and is being cultivated on a large scale in southern China. The biosynthesis of active ingredients in A. paniculata are regulated and controlled by genes, but their specific roles are still not fully understood. To further explore the growth regulation factors and utilization of its medicinal parts of this industrial crop, chemical and transcriptome analyses were conducted on the roots, stems, and leaves of A. paniculata to identify the biosynthesis pathways and related candidate genes of the active ingredients. The chemical analysis revealed that the main components of A. paniculata were diterpene lactones and flavonoids, which displayed potential ability to treat SARS-CoV-2 through molecular docking. Moreover, the transcriptome sequencing annotated a total of 40,850 unigenes, including 7962 differentially expressed genes. Among these, 120 genes were involved in diterpene lactone biosynthesis and 60 genes were involved in flavonoid biosynthesis. The expression of diterpene lactone-related genes was the highest in leaves and the lowest in roots, consistent with our content determination results. It is speculated that these highly expressed genes in leaves may be involved in the biosynthesis pathway of diterpenes. Furthermore, two class Ⅰ terpene synthases in A. paniculata transcriptome were also annotated, providing reference for the downstream pathway of the diterpene lactone biosynthesis. With their excellent market value, our experiments will promote the study of the biosynthetic genes for active ingredients in A. paniculata and provide insights for subsequent in vitro biosynthesis.

Curcumin improves the egg quality, antioxidant activity, and intestinal microbiota of quails during the late laying period.

This study aimed to investigate the effects of dietary curcumin supplementation on laying performance, egg quality, egg metabolites, lipid metabolism, antioxidant activity, and intestinal microbial composition of quails in the late laying period. A total of 960 late-laying quails (240-day-old) were randomly divided into 4 groups of 6 replicates each (n = 40/replicate). The experimental diets of the 4 groups consisted of basal diets supplemented with 0, 50, 100, and 200 mg/kg curcumin, respectively. The feeding experiment lasted for 8 wk. The results showed that 200 mg/kg curcumin supplementation decreased mortality and increased eggshell thickness and strength compared with the 0 mg/kg curcumin supplementation during wk 5 to 8. In addition, dietary supplementation of curcumin promoted lipid metabolism, enhanced antioxidant activity, and modified intestinal microbiota structure. In conclusion, dietary supplemented with 200 mg/kg curcumin significantly improved the egg quality of quails in the late laying period, primarily by improving lipid metabolism and selectively regulating the intestinal microbial community.

Preparative separation of iridoid glucosides and crocins from Gardeniae Fructus using sequential macroporous resin column chromatography and evaluation of their anti-inflammatory and antioxidant activities.

Iridoid glycosides (geniposide (GP), genipin-1-gentiobioside (GB), etc.) and crocins (crocin Ⅰ (CR1), crocin Ⅱ(CR2), etc.) are two main bioactive components in Gardeniae Fructus (GF), which is a famous traditional Chinese medicine. Iridoid glycosides exhibit many activities and are used to manufacture gardenia blue pigment for the food industry. Crocins are rare natural water-soluble carotenoids that are often used as food colorants. A sequential macroporous resin column chromatography technology composed of HC-500B and HC-900B resins was developed to selectively separate iridoid glucosides and crocins from GF. The adsorption of GP on HC-900B resin was an exothermic process. The adsorption of CR1 on HC-500B resin was an endothermic process. The two kinds of components were completely separated by a sequential resin column. GB and GP were mainly found in product 1 (P1) with purities of 11.38% and 46.83%, respectively, while CR1 and CR2 were mainly found in product 2 (P2) with purities of 12.32% and 1.40%, respectively. The recovery yields of all the compounds were more than 80%. The above results showed that sequential resin column chromatography technology achieved high selectivity and recovery yields. GF extract, P1 and P2 could significantly inhibit the secretion of nitric oxide (NO), tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6) in lipopolysaccharide (LPS)-induced RAW264.7 cells, indicating that iridoid glycosides and crocins provide a greater contribution to the anti-inflammatory activity of GF. At the same time, compared to the GF extract and P1, P2 exhibited stronger scavenging activities against 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radicals, indicating that crocins may provide a significant contribution to the antioxidant activity of GF.

Dual-Mechanism Glycolipidpeptide with High Antimicrobial Activity, Immunomodulatory Activity, and Potential Application for Combined Antibacterial Therapy.

Bacterial drug resistance is becoming increasingly serious, and it is urgent to develop effective antibacterial drugs. Antimicrobial peptides (AMPs), as potential candidates against bacteria, have a broad prospect for development. Herein, a series of AMPs with biological characteristics (net positive charge, amphiphilicity, and α-helix), an AXA motif recognized by membrane bound serine protease type I signal peptidases (SPase I), an FLPII motif to reduce hemolysis, and a monosaccharide motif to improve the stability and activity were designed and synthesized, and among which, the glycolipidpeptide GLP6 (glycosylated LP6 lipopeptide) had excellent antibacterial and immunomodulatory activity, good stability and biocompatibility, and excellent biofilm eradication and membrane penetrating activity. The positively charged spherical aggregates formed by self-assembly of GLP6 could encapsulate tetracycline (TC) to form GLP6@TC with a sustained-release effect, which could enhance the sensitivity of bacteria to the antibiotic and realize combined sterilization. The results of acute peritonitis and bacterial keratitis showed that GLP6@TC had a good combined antibacterial effect and the ability to inhibit interleukin-2 (IL-2), which could significantly reduce the inflammatory response while treating bacterial infection, and it had great potential for application. The results of computer molecular docking showed the AXA motif could effectively bind to SPase I, which was consistent with the results of biological experiments. In general, the study could provide a perspective for the design of AMPs and combined antibacterial therapy.

The Bridge Between Ischemic Stroke and Gut Microbes: Short-Chain Fatty Acids.

Short-chain fatty acids (SCFAs) are monocarboxylates produced by the gut microbiota (GM) and result from the interaction between diet and GM. An increasing number of studies about the microbiota-gut-brain axis (MGBA) indicated that SCFAs may be a crucial mediator in the MGBA, but their roles have not been fully clarified. In addition, there are few studies directly exploring the role of SCFAs as a potential regulator of microbial targeted interventions in ischemic stroke, especially for clinical studies. This review summarizes the recent studies concerning the relationship between ischemic stroke and GM and outlines the role of SCFAs as a bridge between them. The potential mechanisms by which SCFAs affect ischemic stroke are described. Finally, the beneficial effects of SFCAs-mediated therapeutic measures such as diet, dietary supplements (e.g., probiotics and prebiotics), fecal microbiota transplantation, and drugs on ischemic brain injury are also discussed.

Analysis on the Effect of Wushu Project Propagation in Nonmaterial Cultural Field of Environmental Protection Based on Artificial Intelligence Analysis Technology.

Since the beginning of the new century, the party and the government have always attached great importance to youth sports work and cared about the healthy growth of youth. The CPC Central Committee and the State Council have issued the opinions on strengthening youth sports and strengthening youth physique, which has initially formed a social consensus on promoting the healthy growth of young students. With the process of school education reform and the promulgation and implementation of the Ministry of Education's opinions on implementing and ensuring the time of daily sports activities for primary and middle school students, especially the official launch of the "national sunshine sports for hundreds of millions of students" in 2021, the school sports work in China has entered a new historical stage. Under this premise, the status and importance of Wushu in the intangible cultural field of schools have been gradually improved. As an important part of school sports, Wushu in the field of intangible culture plays an important role in strengthening students' physique, improving students' health, and promoting students' intellectual development. It is also one of the basic ways to realize school sports. How to better play its role has attracted the attention of many experts and scholars. This study focuses on the current situation of Wushu projects in the field of intangible culture in Weixian Middle School of Xingtai City, trying to understand its advantages and existing problems through the current situation, and put forward targeted solutions, so as to enable Wushu projects in the field of intangible culture to play their educational and fitness functions in school sports and provide help for the comprehensive development of middle school students.

Integrating transcriptome and chemical analyses to reveal the anti-Alzheimer's disease components in Verbena officinalis Linn.

Verbena officinalis Linn. is a kind of traditional Chinese medicine, which has a long history of application and shows good effects on neuroprotection. Therefore, we consider that V. officinalis may be a potential drug for treating Alzheimer's disease (AD). First, ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) pointed out that the main chemical components in V. officinalis were iridoid glycosides, phenylethanoid glycosides, and flavonoids. These compounds were used for molecular docking and the results showed that these compounds had good anti-AD activity. To explore the biosynthetic pathway of anti-AD components in V. officinalis, UPLC and ultraviolet (UV) spectrophotometry were used for contents determination and the result was leaf > stem > root. At the same time, 92,867 unigenes were annotated in V. officinalis transcriptome; 206, 229, 115 related unigenes were, respectively, annotated in iridoid glycoside, phenylethanoid glycoside, and flavonoid pathway, of which 61, 73, and 35 were differential expression genes. The components had relatively high expression in leaves, which was consistent with the quantitative results. In addition, the tissue distribution particularity of verbenalin may be related to the branching of pathways. Meanwhile transcription factors VoWRKY6 and VoWRKY7 may be involved in the regulation of iridoid glycoside biosynthesis. Further, VoWRKY3, VoWRKY9, and VoWRKY12 may be related to flavonoid biosynthesis. The above research is helpful to explore the biosynthetic pathway of anti-AD components and the regulation mechanism of active components and to further explore the anti-AD effect of V. officinalis.

Compounds with NO Inhibitory Effect from the Rattan Stems of Sinomenium acutum, a Kind of Chinese Folk Medicine for Treating Rheumatoid Arthritis.

Three new alkaloids (1-3), one new diphenyl ether (4) and fifteen known alkaloids (5-19) were isolated from the rattan stems of Sinomenium acutum. Comprehensive analyses of HR-ESI-MS, 1D (1 H and 13 C), 2D-NMR (1 H-1 H COSY, HSQC, HMBC, NOESY), circular dichroism (CD), UV and IR revealed the structures and absolute configurations of these new compounds. The structures of other compounds were determined by comparison of their 1 H and 13 C-NMR data with previous literature reports. By measuring the amount of NO produced, the anti-inflammatory properties of the isolated compounds were studied. The results showed that compounds 4 and 5 had strong NO inhibitory activity.

Full-length transcriptome and metabolite analysis reveal reticuline epimerase-independent pathways for benzylisoquinoline alkaloids biosynthesis in Sinomenium acutum.

Benzylisoquinoline alkaloids (BIAs) are a large family of plant natural products with important pharmaceutical applications. Sinomenium acutum is a medicinal plant from the Menispermaceae family and has been used to treat rheumatoid arthritis for hundreds of years. Sinomenium acutum contains more than 50 BIAs, and sinomenine is a representative BIA from this plant. Sinomenine was found to have preventive and curative effects on opioid dependence. Despite the broad applications of S. acutum, investigation on the biosynthetic pathways of BIAs from S. acutum is limited. In this study, we comprehensively analyzed the transcriptome data and BIAs in the root, stem, leaf, and seed of S. acutum. Metabolic analysis showed a noticeable difference in BIA contents in different tissues. Based on the study of the full-length transcriptome, differentially expressed genes, and weighted gene co-expression network, we proposed the biosynthetic pathways for a few BIAs from S. acutum, such as sinomenine, magnoflorine, and tetrahydropalmatine, and screened candidate genes involved in these biosynthesis processes. Notably, the reticuline epimerase (REPI/STORR), which converts (S)-reticuline to (R)-reticuline and plays an essential role in morphine and codeine biosynthesis, was not found in the transcriptome data of S. acutum. Our results shed light on the biogenesis of the BIAs in S. acutum and may pave the way for the future development of this important medicinal plant.

Taurochenodeoxycholic acid inhibits the proliferation and invasion of gastric cancer and induces its apoptosis.

Taurochenodeoxycholic acid (TCDCA) is the principal ingredient of Compound Shougong Powder. Despite traditional Chinese medicine (TCM) research demonstrates that Compound Shougong Powder can restrict tumor growth, whether TCDCA exerts a role in suppressing cancer as the major ingredient of Compound Shougong Powder remains unknown. This study aims to clarify the regulatory mechanism of TCDCA on gastric cancer. Gastric cancer cells SGC-7901 were cultured to investigate the effects of TCDCA on proliferation and apoptosis. Furthermore, a subcutaneously implanted tumor model was established using SGC-7901 cells in BALB/C nude mice and tumor volume was measured under low and high dose treatment of TCDCA. Cell proliferation, apoptosis, and invasion were subjected to 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide (MTT) assay, flow cytometry, and transwell assay. Differentially expressed genes were screened by transcriptome sequencing. Nude mouse tumorigenicity assay was initiated to identify the effect of TCDCA on both tumor volume and weight, and the expression of candidate genes screened by transcriptome sequencing was determined by real-time fluorescence quantification (qPCR) and Western blot. The experiments revealed that TCDCA could significantly inhibit the proliferation and invasion of gastric cancer cells and induce apoptosis of these cells. Meanwhile, test findings via in vivo indicated that TCDCA severely diminished the volume and weight of tumors. This study first demonstrated that TCDCA inhibited the proliferation and invasion of gastric cancer and induced apoptosis, which is expected to serve as an experimental basis for the application of TCM in tumor therapeutic options. PRACTICAL APPLICATIONS: Through this study, the inhibitory effect of Taurochenodeoxycholic acid on gastric cancer can be clarified, which provides a new research basis for the application of traditional Chinese medicine (TCM) and TCM monomer in cancer. In addition, this study can further promote the research and application of Chinese traditional medicine, which has important application value and economic benefits.

Global identification and determination of the major constituents in Kai-Xin-San by ultra-performance liquid chromatography-quadrupole-Orbitrap mass spectrometry and gas chromatography-mass spectrometry.

Kai-Xin-San (KXS) is a traditional Chinese medicine (TCM) formula containing four herbal medicines: Ginseng Radix Rhizoma, Polygalae Radix, Poria and Acori Tatarinowii Rhizoma. A large number of pharmacological studies in vitro and in vivo have shown that KXS is characterized by anti-depression, anti-Alzheimer's disease, anti-oxidation and other activities. However, the pharmacodynamic substance basis studies of KXS are hitherto quite limited. Here, KXS was identified and determined by ultra-performance liquid chromatography-quadrupole-Orbitrap mass spectrometry (UPLC-Q-Orbitrap MS) and gas chromatography-mass spectrometry (GC-MS). Firstly, the data-dependent acquisition mode (DDA) of UPLC-Q-Orbitrap MS combined with the inclusion list were used to collected the chemical composition. The chemical constituents of KXS were identified by local database on compound discoverer™ 3.1 software and Xcalibur 4.1 software. With the use of this approach, a total of 211 compounds were identified from KXS. Wherein 60 compounds were from Ginseng Radix Rhizoma, 40 compounds were from Poria, and 111 compounds were from Polygala Radix, respectively. Secondly, 105 volatile constituents were identified by GC-MS analysis, which were mainly derived from Acori Tatarinowii Rhizoma. Besides, an adjusted parallel reaction monitoring method was established and validated to quantify the seventeen major compounds in different herbal medicines of KXS, which were chosen as the benchmarked substances to evaluate the quality of KXS. In conclusion, this study provided a generally applicable strategy for global metabolite identification of the complicated components and determination of multi-component content in traditional Chinese medicines.

Phytochemical Constituents and Biological Activities of Plants from the Genus Cissampelos.

Cissampelos is a significant genus comprising of approximately 21 species of the medicinal plants (Menispermaceae). The plants of this genus are used in traditional medicine for the treatment of various ailments such as asthma, arthritis, dysentery, hyperglycemia, cardiopathy, hypertension and other related problems. These plants are rich in bioactive dibenzylisoquinoline and aborphine as well as small amounts of other ingredients. In recent years, the chemical constituents and pharmacological activities of Cissampelos genus have been paid more and more attention due to their diversity. Herein, we compile the chemical constituents and biological activities on this genus, and summarize the 13 C-NMR data of the main bioactive ingredients. All information comes from scientific databases such as Google Scholar, PubMed, Sci-Finder, ScienceDirect, Web of Science and CNKI. It provides valuable data for the future research and development of Cissampelos genus.

Separation of three chromones from Saposhnikovia divaricata using macroporous resins followed by preparative high-performance liquid chromatography.

Prim-O-glucosylcimifugin, cimifugin, and 5-O-methylvisamminoside are three major chromone derivatives of Saposhnikovia divaricata that have many pharmacological activities, such as anti-inflammatory and antitumor activities. In the present work, an effective method for the simultaneous separation of prim-O-glucosylcimifugin, cimifugin, and 5-O-methylvisamminoside with high purities was established using HPD-300 resin coupled with preparative high-performance liquid chromatography. The adsorption kinetics curves of the three compounds on the HPD-300 resin were studied and found to fit well according to the pseudo-second-order equation. The adsorption isotherm results indicated that the adsorption process of the three compounds was exothermic. After a one-run treatment with the resin, the contents of prim-O-glucosylcimifugin, cimifugin, and 5-O-methylvisamminoside increased from 0.29, 0.06, and 0.37% to 13.07, 2.83, and 16.91% with recovery yields of 76.38, 78.25, and 76.73%, respectively. Finally, the purities of the three compounds were found to reach more than 95% after further separation using preparative high-performance liquid chromatography. The method developed in this study was effective and could simultaneously separate three chromones from Saposhnikovia divaricate. The experimental results also showed that the HPD-300 resin is suitable for the separation of chromone derivatives.

Ten New Dammarane-Type Saponins with Hypolipidemia Activity from a Functional Herbal Tea-Gynostemma pentaphyllum.

Gynostemma pentaphyllum (thumb.) Makino is a functional herbal tea commonly used in Asian countries and regions to reduce blood lipid levels. G. pentaphyllum saponin is the main component, but there are still a large number of components with lipid-lowering activity that have not been found. In this study, 10 novel dammarane-type saponins, (1-10) and a known one (11) were isolated from G. pentaphyllum. Ten new compounds were identified and named as yunnangypenosides A-J (1-10), and another known one (11) was also obtained. Their chemical structures were determined by MS, NMR spectroscopic analyses. Moreover, the cytotoxicities on human HepG-2 hepatocellular carcinoma cells of these isolates were evaluated, and the results showed that compounds 1-11 had no obvious cytotoxicity. Finally, all these compounds were evaluated for their lipid-lowering effect by means of the oil red O staining method. Ten compounds could significantly reduce lipid levels except of 2, especially 8 exhibite the strongest hypolipidemia activity.

Clinical evidence-guided network pharmacology analysis reveals a critical contribution of ß1-adrenoreceptor upregulation to bradycardia alleviation by Shenxian-Shengmai.

BACKGROUND: Shenxian-Shengmai (SXSM) Oral Liquid is a CFDA-approved patent Chinese Herbal medicine, which has been clinically used for the treatment of bradycardia. However, its active components and action mechanism remain to be established. The present study aimed to evaluate the efficacy of SXSM on bradycardia and to identify the possible active components and their pharmacological targets for this action. METHODS: A literature-based meta-analysis was performed to evaluate the clinical efficacy of SXSM on bradycardia, which was confirmed by a rat ex vivo cardiac model. Network pharmacology analysis was then conducted to reveal the potential targets of SXSM active components and their anti-arrhythmia mechanisms. Finally, the identified drug-target interaction was confirmed by immunofluorescence assay in cardiomyocyte. RESULTS: Meta-analysis of the available clinical study data shows that Shenxian-Shengmai Oral Liquid has a favorable effect for bradycardia. In an ex vivo bradycardia model of rat heart, SXSM restored heart rate by affecting Heart rate variability (HRV) which is associated with autonomic nervous system activity. A drug-target-pathway network analysis connecting SXSM components with arrhythmia suggested that a prominent anti-arrhythmia mechanisms of SXSM was via ß1-adrenergic signaling pathway, which was subsequently validated by immunofluorescence assay showing that SXSM indeed increased the expression of ADRB1 in cultured cardiomyocytes. CONCLUSION: By combining approaches of clinical evidence mining, experimental model confirmation, network pharmacology analyses and molecular mechanistic validation, we show that SXSM is an effective treatment for bradycardia and it involves multiple component interacting via multiple pathways, among which is the critical ß1-adrenergic receptor upregulation. Our integrative approach could be applied to other multi-component traditional Chinese medicine investigation where ample clinical data are accumulated but advanced mechanistic studies are lacking.

The effect of the inhalation of and topical exposure to zinc oxide nanoparticles on airway inflammation in mice.

Zinc oxide nanoparticles (ZnONPs) are widely used in the manufacturing of many commercial products. Workers exposed to ZnO particles may develop metal fume fever. Our previous study suggested that the oropharyngeal aspiration of ZnONPs could cause eosinophilic airway inflammation and increase T helper 2 (Th2) cytokine expression in the absence of allergens in mice. ZnO has been used topically as a sunscreen and a therapeutic agent for dermatological conditions. To understand whether inhalation and topically applied ZnONPs might cause or exert an adjuvant effect on the development of allergic airway inflammation in mice, C57BL/6 J mice were exposed to filtered air or 2.5 mg/m3 ZnONPs via whole-body inhalation for 5 h a day over 5 days, and BALB/c mice were topically exposed to ZnONPs using modified mouse models of atopic dermatitis (AD) and asthma. Ovalbumin (OVA) solution was used as an allergen in the topical exposure experiments. A significantly increased eosinophil count and mixed Th1/Th2 cytokine expression were detected in the bronchoalveolar lavage fluid (BALF) after ZnONP inhalation. However, only mild eosinophilia and low Th2 cytokine expression were detected in the BALF after oropharyngeal OVA aspiration in the high-dose ZnONP topical treatment group. These results suggest that ZnONP inhalation might play a role in the development of allergic airway inflammation in mice. However, topically applied ZnONPs only play a limited role in the development of allergic airway inflammation in mice.

Purification of spinosin from Ziziphi Spinosae Semen using macroporous resins followed by preparative high-performance liquid chromatography.

As a well-known traditional Chinese medicine, Ziziphi Spinosae Semen has been used for treating anxiety and insomnia for a long time. Spinosin, the main active C-glycoside flavonoid in Ziziphi Spinosae Semen, has attracted much attention because of its many pharmacological activities including strong hypnotic effects, anxiolytic-like effects, and so on. In the present work, high-purity spinosin was separated from Ziziphi Spinosae Semen using the HPD-300 resin followed by preparative high-performance liquid chromatography. The adsorption kinetics curve of spinosin on the HPD-300 resin was studied and fitted well by the pseudo-second-order equation. The adsorption isotherms were also constructed and low temperature favored the adsorption reaction. The separation parameters were optimized using dynamic adsorption and desorption tests. After a one-run treatment with HPD-300 resin, the concentration of spinosin increased 11.8-fold from 0.99 to 11.7% with a recovery yield of 80.4%. Furthermore, the purity of spinosin could surpass above 98% after separation by preparative high-performance liquid chromatography and recrystallization with a recovery yield of 72.6%. The developed method was effective and suitable for the large-scale preparation of spinosin. Moreover, it was confirmed that HPD-300 resin could enable good selection for the enrichment of flavonoids from different plants.

A Strategy for Selecting "Q-Markers" of Chinese Medical Preparation via Components Transfer Process Analysis with Application to the Quality Control of Shengmai Injection.

Chinese medical preparation has complicated chemical constituents. Consequently, the proper quality control methods for these Chinese medical preparations have been great challenges to the traditional Chinese medicine modernization and internationalization. What components should be chosen for quality control is a big challenge in the development of traditional Chinese medicine. A new concept of "Quality Marker" was proposed by Liu et al. to solve this problem and established a new research paradigm for traditional Chinese medicine quality study. Several strategies were proposed by the researchers in traditional Chinese medicine, here, we used Shengmai injection as an example to discuss a strategy for selecting "Quality Markers" of Chinese medical preparation by the components transfer process analysis in the Shengmai injection manufacturing process. Firstly, a total of 87 compounds were identified or partially characterized in shengmai injection. Secondly, referenced to the quality control method in China pharmacopeia and considered the biomarkers in the original medicines and representative components in the manufacturing process, four ginsenosides in Panax ginseng (Hongshen), two compounds in Schisandra chinensis (Wuweizi), and a sugar from Ophiopogon japonicas (Maidong) were quantified. As a result, these seven representative compounds exhibited an acceptable transitivity throughout the Shengmai injection manufacturing process. Finally, combined with the active ingredients, components transfer process analysis, and comprehensive evaluation by "Spider-web" analysis, six compounds were selected as the quality markers for the quality control of Shengmai injection. Through this strategy of optimization for quality markers of Shengmai injection, we found that these six compounds could represent the main bioactive substances and be easily detected in the whole process of production. Furthermore, the quality control method was developed for quality assessment and control of these six quality markers in the Shengmai injection. The total content range of the selected quality markers in the 10 batches of the Shengmai injection is 13.844-22.557 mg/mL.

High-Throughput Screening and Identification of Potent Broad-Spectrum Inhibitors of Coronaviruses.

Coronaviruses (CoVs) act as cross-species viruses and have the potential to spread rapidly into new host species and cause epidemic diseases. Despite the severe public health threat of severe acute respiratory syndrome coronavirus and Middle East respiratory syndrome CoV (MERS-CoV), there are currently no drugs available for their treatment; therefore, broad-spectrum inhibitors of emerging and endemic CoVs are urgently needed. To search for effective inhibitory agents, we performed high-throughput screening (HTS) of a 2,000-compound library of approved drugs and pharmacologically active compounds using the established genetically engineered human CoV OC43 (HCoV-OC43) strain expressing Renilla luciferase (rOC43-ns2Del-Rluc) and validated the inhibitors using multiple genetically distinct CoVs in vitro We screened 56 hits from the HTS data and validated 36 compounds in vitro using wild-type HCoV-OC43. Furthermore, we identified seven compounds (lycorine, emetine, monensin sodium, mycophenolate mofetil, mycophenolic acid, phenazopyridine, and pyrvinium pamoate) as broad-spectrum inhibitors according to their strong inhibition of replication by four CoVs in vitro at low-micromolar concentrations. Additionally, we found that emetine blocked MERS-CoV entry according to pseudovirus entry assays and that lycorine protected BALB/c mice against HCoV-OC43-induced lethality by decreasing viral load in the central nervous system. This represents the first demonstration of in vivo real-time bioluminescence imaging to monitor the effect of lycorine on the spread and distribution of HCoV-OC43 in a mouse model. These results offer critical information supporting the development of an effective therapeutic strategy against CoV infection.IMPORTANCE Currently, there is no approved therapy to treat coronavirus infection; therefore, broad-spectrum inhibitors of emerging and endemic CoVs are needed. Based on our high-throughput screening assay using a compound library, we identified seven compounds with broad-spectrum efficacy against the replication of four CoVs in vitro Additionally, one compound (lycorine) was found to protect BALB/c mice against HCoV-OC43-induced lethality by decreasing viral load in the central nervous system. This inhibitor might offer promising therapeutic possibilities for combatting novel CoV infections in the future.

Investigation of ovatodiolide, a macrocyclic diterpenoid, as a potential inhibitor of oral cancer stem-like cells properties via the inhibition of the JAK2/STAT3/JARID1B signal circuit.

BACKGROUND: The cancer stem cells (CSCs) have been shown to play key roles in the oral cancer initiation, distant metastasis, the development of chemoresistance and recurrence after treatment. Therefore, the inhibition of oral CSCs has been the target for therapeutic development. PURPOSE: In this study, we investigated the anti-CSCs potential of Ovatodiolide (Ova), a diterpenoid isolate of Anisomeles indica, in vitro and in vivo. METHODS: Oral CSCs were treated with Ova, and the expression of pluripotency factors Oct4, Sox-2, and Nanog were evaluated by western blot. Effect of Ova on self-renewal capacity and clonogenicity were assessed with the sphere formation and clonogenic assay in CSCs model derived from oral cancer cell. The effect of Ova was also investigated in a mouse xenograft model obtained by injecting nude mice with oral CSCs cells. RESULTS: We demonstrated that Ova significantly and dose-dependently suppressed oral cancer cell viability and colony formation; Ova markedly inhibited the ALDH1 activities and reduced the CD44high/ALDHrich cell sub-population. Additionally, Ova suppressed orosphere formation by down-regulating CD133, Klf4, Oct4A, Nanog and JARID1B expression. Furthermore, Ova-mediated anti-cancer effects were associated with the dose-dependent reduction in the expression levels of STAT3, p-STAT3, pJAK2, pAKT and pERK1/2 protein. Moreover, Ova synergistically enhanced the anticancer effect of cisplatin against the SAS, FaDu, HSC-3 and TW2.6 orospheres. Ova significantly attenuated the tumor-initiating potential of orosphere in mouse xegnograft model. CONCLUSION: These results demonstrate that Ova effectively suppressed oral tumorigenesis and stemness properties via JAK2/STAT3 signaling. Ova may be considered for future clinical usage.