Development of a virus-based affinity ultrafiltration method for screening virus-surface-protein-targeted compounds from complex matrixes: Herbal medicines as a case study.

Herbal medicines (HMs) are one of the main sources for the development of lead antiviral compounds. However, due to the complex composition of HMs, the screening of active compounds within these is inefficient and requires a significant time investment. We report a novel and efficient virus-based screening method for antiviral active compounds in HMs. This method involves the centrifugal ultrafiltration of viruses, known as the virus-based affinity ultrafiltration method (VAUM). This method is suitable to identify virus specific active compounds from complex matrices such as HMs. The effectiveness of the VAUM was evaluated using influenza A virus (IAV) H1N1. Using this method, four compounds that bind to the surface protein of H1N1 were identified from dried fruits of Terminalia chebula (TC). Through competitive inhibition assays, the influenza surface protein, neuraminidase (NA), was identified as the target protein of these four TC-derived compounds. Three compounds were identified by high performance liquid chromatography (HPLC) and liquid chromatography/mass spectrometry (LC/MS), and their anti-H1N1 activities were verified by examining the cytopathic effect (CPE) and by performing a virus yield reduction assay. Further mechanistic studies demonstrated that these three compounds directly bind to NA and inhibit its activity. In summary, we describe here a VAUM that we designed, one that can be used to accurately screen antiviral active compounds in HMs and also help improve the efficiency of screening antiviral drugs found in natural products.

Integrated serum pharmacochemistry and investigation of the anti-influenza A virus pneumonia effect of Qingjin Huatan decoction.

ETHNOPHARMACOLOGICAL RELEVANCE: Qingjin Huatan Decoction (QJHTT) consists of 11 herbal medicines: Scutellaria baicalensis Georgi, Gardenia jasminoides J. Ellis, Platycodon grandiflorus (Jacq.) A. DC., Ophiopogon japonicus (Thunb.) Ker Gawl., Morus alba L., Fritillaria thunbergii Miq., Anemarrhena asphodeloides Bunge, Trichosanthes kirilowii Maxim., Citrus reticulata Blanco, Poria cocos (Schw.) Wolf, and Glycyrrhiza uralensis Fisch. As a traditional Chinese medicinal formula, QJHTT has been used for more than 400 years in China. It has shown promising results in treating influenza A virus (IAV) pneumonia. AIM OF THE STUDY: To elusive the specific pharmacological constituents and mechanisms underlying its anti-IAV pneumonia effects. MATERIALS AND METHODS: The components in QJHTT were analyzed through the use of a serum pharmacology-based ultra high-performance liquid chromatography Q- Exactive Orbitrap mass spectrometry (UHPLC-Q Exactive Orbitrap-MS) method. Simultaneously, the dynamic changes in IAV-infected mouse lung viral load, lung index, and expression of lung inflammation factors were monitored by qRT-PCR. RESULTS: We successfully identified 152 chemical components within QJHTT, along with 59 absorbed chemical prototype constituents found in the serum of mice treated with QJHTT. 43.45% of these chemical components and 43.10% of the prototype constituents were derived from the monarch drugs, namely Huangqin and Zhizi, aligning perfectly with traditional Chinese medicine theory. Notably, our analysis led to the discovery of 14 compounds within QJHTT for the first time, three of which were absorbed into the bloodstream. Simultaneously, we observed that QJHTT not only reduced the viral load but also modulated the expression of inflammation factors in the lung tissue including TNF-α, IL-1ß, IL-4, IL-6, IFN-γ, and IL17A. A time-effect analysis further revealed that QJHTT intervention effectively suppressed the peak of inflammatory responses, demonstrating a robust anti-IAV pneumonia effect. CONCLUSIONS: We comprehensively analyzed the pharmacological material basis of QJHTT by a highly sensitive and high-resolution UHPLC-Q Exactive Orbitrap-MS method, and demonstrated its efficacy in combating IAV pneumonia by reducing lung viral load and inflammatory factors. This study has significant importance for elucidating the pharmacological basis and pharmacological mechanism of QJHTT in combating IAV pneumonia.

Fabrication of a bio-based polymer adsorbent and its application for extraction and determination of glycosides from Huangqi Liuyi decoction.

Huangqi Liuyi Decoction, a famous classical Chinese prescription, shows significant curative effect on diabetes and its complications, in which calycosin-7-glucoside, liquiritin and glycyrrhizic acid are the main components that playing these mentioned pharmacological activity, under the synergistic action of various other ingredients in the decoction. However, there are significant differences in the content of active compounds in Chinese medicinal materials, which mainly due to origin, picking seasons, and processing methods. Hence, the accurate content of the glycosides is the prerequisite for ensuring the pharmacological efficacy. Aiming at establishing an efficient extraction and determination method for accurate quantitative analysis of calycosin-7-glucoside, liquiritin and glycyrrhizic acid in Huangqi Liuyi Decoction, an on line solid-phase extraction-high-performance liquid chromatography method was developed, using a homemade bio-based monolithic adsorbent. The bio-based adsorbent was prepared in a stainless steel tube, using bio-monomers of methyleugenol and S-allyl-L-cysteine, which effectively reduced the dependence of the polymer field on non-renewable fossil resources and reduced carbon emissions. Furthermore, the prepared adsorbent owned abundant chemical groups, which can produce interactions of hydrogen bond, dipole-dipole, π-π and hydrophobic force with the target glycosides, thus improving the specific recognition ability of the adsorbent. The experiments were carried out on an LC-3000 HPLC instrument with a six-way valve. Methodology validation indicates that the recovery is in the range of 97.0%-103.4% with the RSD in the range of 1.6%-4.0%, due to the specific selectivity of the bio-based monolithic adsorbent for these three glycosides, and good matrix-removal ability for Huangqi Liuyi decoction. The limit of detection is 0.17, 0.50 and 0.33 µg/mL for calycosin-7-glucoside, liquiritin and glycyrrhizic acid, respectively, and the limit of quantitation is 0.50, 1.50 and 1.00 µg/mL, respectively, with the linear range of 2-200 µg/mL for calycosin-7-glucoside, and 5-500 µg/mL for liquiritin and glycyrrhizic acid. The present work provided a simple and efficient method for the extraction and determination of glycosides in complex medicinal plants.

A common thalamic hub for general and defensive arousal control.

The expression of defensive responses to alerting sensory cues requires both general arousal and a specific arousal state associated with defensive emotions. However, it remains unclear whether these two forms of arousal can be regulated by common brain regions. We discovered that the medial sector of the auditory thalamus (ATm) in mice is a thalamic hub controlling both general and defensive arousal. The spontaneous activity of VGluT2-expressing ATm (ATmVGluT2+) neurons was correlated with and causally contributed to wakefulness. In sleeping mice, sustained ATmVGluT2+ population responses were predictive of sensory-induced arousal, the likelihood of which was markedly decreased by inhibiting ATmVGluT2+ neurons or multiple downstream pathways. In awake mice, ATmVGluT2+ activation led to heightened arousal accompanied by excessive anxiety and avoidance behavior. Notably, blocking their neurotransmission abolished alerting stimuli-induced defensive behaviors. These findings may shed light on the comorbidity of sleep disturbances and abnormal sensory sensitivity in specific brain disorders.

Qingjin Huatan decoction protects mice against influenza a virus pneumonia via the chemokine signaling pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Qingjin Huatan Decoction (QJHTT) consists of 11 herbal medicines: Scutellaria baicalensis Georgi, Gardenia jasminoides J.Ellis, Platycodon grandiflorus (Jacq.) A.DC., Ophiopogon japonicus (Thunb.) Ker Gawl., Morus alba L., Fritillaria thunbergii Miq., Anemarrhena asphodeloides Bunge, Trichosanthes kirilowii Maxim., Citrus reticulata Blanco, Poria cocos (Schw.) Wolf, and Glycyrrhiza uralensis Fisch. As a traditional compound Chinese medicinal formula, QJHTT has been used for more than 400 years in China. Historically, it was used to treat respiratory diseases and had shown beneficial clinical results for diseases related to lung inflammation. AIM OF THE STUDY: To investigate the therapeutic effect of QJHTT on influenza A virus (IAV) pneumonia in mice and explore its possible mechanism of action. MATERIALS AND METHODS: The components in QJHTT were analyzed by UPLC-Q-TOF-MS and some antiviral active components reported in the literature were determined and quantified by HPLC. The protective effects of QJHTT were investigated using lethal and sublethal doses (2 LD50 or 0.8 LD50 viral suspension, separately) of H1N1-infected mice. Mortality and lung lesions in H1N1-infected mice were used to evaluate the efficacy of QJHTT. The potential mechanism of QJHTT in the treatment of viral pneumonia was determined at the gene level by RNA sequencing and validated by qRT-PCR. Following this, the changes in protein levels of JAK2/STAT3 were analyzed since it is a key downstream target of the chemokine signaling pathways. Preliminary elucidation of the mechanism of QJHTT to protect mice against IAV pneumonia through this pathway was conducted. RESULTS: In this study, 12 antiviral active constituents including baicalin, geniposide, and mangiferin were identified from QJHTT. In vivo treatment of QJHTT reduced the virus titers of lung tissue significantly and improved the survival rate, lung index, and pulmonary histopathological changes; additionally, a reduction in the serum levels of TNF-α, IL-1ß, IL-6, and IFN-γ inflammatory factors in H1N1-infected mice was observed. RNA-seq analysis and qRT-PCR showed that QJHTT primarily reversed the activities CCL2, CCL7, CCR1, and other chemokines and their reception-related genes, suggesting that QJHTT may produce disease-resistant pneumonia by inhibiting the downstream JAK2/STAT3 pathway. Western blot analysis confirmed that QJHTT effectively reduced the protein levels of JAK2, STAT3, and related phosphorylated products in the lung tissue of H1N1-infected mice. CONCLUSIONS: Our results indicated that QJHTT alleviated IAV pneumonia in mice by regulating related chemokines and their receptor-related genes in lung tissue, thereby inhibiting JAK2/STAT3 pathway. This could pave way for the design of novel therapeutic strategies to treat viral pneumonia.

Biosurfactant promoted enzymatic saccharification of alkali­pretreated reed straw.

The decrease of cellulase activity and unproductive adsorption of lignin are important obstructive factors for inefficient enzymatic hydrolysis. This paper applied five different kinds of biosurfactants including rhamnolipid, sophorolipid, chitin, tea saponin, and sodium lignosulfonate in the enzymatic hydrolysis process of alkali-pretreated reed straw (RS) to enhance the saccharification efficiency. When 8 g/L sophorolipid is added, the efficiency of enzymatic hydrolysis is 91.68 %, which is 30.65 % higher than that without using any biosurfactant. The efficiency of enzymatic hydrolysis can be further increased to 99.56 % when 7.5 g/L sophorolipid and 1.5 g/L tea saponin are added together. This is because the sophorolipid, rhamnolipid, and chitin can synergistically hamper the enzymatic inactivation during enzymatic hydrolysis, while tea saponin and sodium lignosulfonate can inhibit the non-productive adsorption of lignin. This work proposed a very effective method to improve the efficiency of enzymatic hydrolysis and reduce the dosage of the enzyme by adding biosurfactants.

Hesperetin ameliorates ischemia/hypoxia-induced myocardium injury via inhibition of oxidative stress, apoptosis, and regulation of Ca2+ homeostasis.

Ischemia/hypoxia (I/H)-induced myocardial injury has a large burden worldwide. Hesperetin (HSP) has a cardioprotective effect, but the molecular mechanism underlying this is not clearly established. Here, we focused on the protective mechanisms of HSP against I/H-induced myocardium injury. H9c2 cardiomyocytes were challenged with CoCl2 for 22 h to imitate hypoxia after treatment groups received HSP for 4 h. The viability of H9c2 cardiomyocytes was evaluated, and cardiac function indices, reactive oxygen species, apoptosis, mitochondrial membrane potential (MMP), and intracellular Ca2+ concentration ([Ca2+ ]i ) were measured. L-type Ca2+ current (ICa-L ), myocardial contraction, and Ca2+ transients in isolated ventricular myocytes were also recorded. We found that HSP significantly increased the cell viability, and MMP while significantly decreasing cardiac impairment, oxidative stress, apoptosis, and [Ca2+ ]i caused by CoCl2 . Furthermore, HSP markedly attenuated ICa-L , myocardial contraction, and Ca2+ transients in a concentration-dependent manner. Our findings suggest a protective mechanism of HSP on I/H-induced myocardium injury by restoring oxidative balance, inhibiting apoptosis, improving mitochondrial function, and reducing Ca2+ influx via L-type Ca2+ channels (LTCCs). These data provide a new direction for HSP applied research as a LTCC inhibitor against I/H-induced myocardium injury.

Garlic essential oil in water nanoemulsion prepared by high-power ultrasound: Properties, stability and its antibacterial mechanism against MRSA isolated from pork.

Food-borne methicillin-resistance Staphylococcus aureus (MRSA) has caused significant health threats and economic loss in livestock and poultry products. Garlic essential oil (GEO) is an effective antibacterial agent but presents strong instability and hydrophobicity. In this study, GEO in water nanoemulsion (GEON) with good stability was produced by emulsification technique of high-power ultrasound. Its antibacterial activity and underlying mechanism against MRSA isolated from retailed pork were investigated. Results showed that ultrasonic treatment significantly reduced the particle size of GENO from 820.3 to 215.0 nm as time increased from 0 to 10 min. Comparatively, GEON of 10 min ultrasound was more stable than other GEONs (0, 1, 5 min) during 30 d storage. It also displayed good thermal stability and relatively good ion stability (NaCl, MgCl2, and glucose). Antibacterial analysis showed that GEON (10 min) exhibited the best anti-MRSA activity among all GEONs, and the minimum inhibitory concentration of GEO in this nanoemulsion was 0.125 % (1.25 mg/mL). Treatment of GEON (10 min) significantly suppressed the cell proliferation of MRSA, which was mainly achieved by damaging the cell membrane as evidenced by membrane depolarization and considerable leakage of intracellular nucleic acids and protein. Laser scanning confocal microscope and scanning electron microscopy showed that treatment of GEON (10 min) significantly altered the membrane integrity and severely damaged the cellular membrane and structure. The present work illustrated that GEON produced by ultrasonic emulsification is a promising alternative to inhibit the contamination and spread of MRSA in livestock and poultry products.

CaCO3 Crystals with Unique Morphologies Controlled by the Hydrogen-Bonded Supramolecular Assemblies of Ureido-Pyrimidinone-Amino Acid Derivatives.

Biomineral materials such as nacre of shells exhibit high mechanical strength and toughness on account of their unique "brick-mortar" multilayer structure. 2-Ureido-4[1H]-pyrimidinone (UPy) derivatives with different types of end groups, due to the self-complementary quadruple hydrogen bonds and abundant Ca2+ binding sites, can easily self-assemble into supramolecular aggregates and act as templates and skeleton in the process of inducing mineral crystallization. In this work, UPy derivatives were used as templates to induce the mineralization and growth of CaCO3 through a CO2 diffusion method. The morphology of CaCO3 crystals was modulated and analyzed by adjusting the synthesizing parameters including Ca2+ concentration, pH, and end groups. The results showed that, by the regulatory role of the mineralization template, it was easier to realize the multilayer crystal structure at a lower concentration of Ca2+ (less than 0.01 mol L-1). Under alkaline regulation, the quadruple hydrogen bonds would be destroyed, and the template's regulation effect on the morphology of CaCO3 crystals would be weakened. Moreover, by comparing different types of end groups, it was proven that the UPy derivatives with carboxylic acid groups (-COOH) played a crucial role in the process of CaCO3 crystallization with unique morphologies.

Cardioprotective effects of alantolactone on isoproterenol-induced cardiac injury and cobalt chloride-induced cardiomyocyte injury.

OBJECTIVES: Alantolactone (AL) is a compound extracted from the roots of Inula Racemosa that has shown beneficial effects in cardiovascular disease. However, the cardioprotective mechanism of AL against hypoxic/ischemic (H/I) injury is still unclear. This research aimed to determine AL's ability to protect the heart against isoproterenol (ISO)-induced MI injury in vivo and cobalt chloride (CoCl2) induced H/I injury in vitro. METHODS: Electrocardiography (ECG), lactate dehydrogenase (LDH), creatine kinase (CK), and cardiac troponin I (cTnI) assays in addition to histological analysis of the myocardium were used to investigate the effects of AL in vivo. Influences of AL on L-type Ca2+ current (ICa-L) in isolated rat myocytes were observed by the patch-clamp technique. Furthermore, cell viability, apoptosis, oxidative stress injury, mitochondrial membrane potential, and intracellular Ca2+ concentration were examined in vitro. RESULTS: The results indicated that AL treatment ameliorated the morphological and ECG changes associated with MI, and decreased levels of LDH, CK, and cTnI. Furthermore, pretreatment with AL elevated antioxidant enzyme activity and suppressed ROS production. AL prevented H/I-induced apoptosis, mitochondria damage, and calcium overload while reducing ICa-L in a concentration and time dependent fashion. The 50% inhibiting concentration (IC50) and maximal inhibitory effect (Emax) of AL were 17.29 µmol/L and 57.73 ± 1.05%, respectively. CONCLUSION: AL attenuated MI-related injury by reducing oxidative stress, apoptosis, calcium overload, and mitochondria damage. These cardioprotective effects may be related to the direct inhibition of ICa-L.

Extraction and determination of tussilagone from Farfarae Flos with online solid-phase extraction-high-performance liquid chromatography using a homemade monolithic cartridge doped with porous organic cage material.

A solid-phase extraction cartridge was fabricated using diallyl isophthalate as the monomer with the addition of porous organic cage material via in situ free-radical polymerization in a stainless-steel column. The resulting monolithic adsorbent exhibited a relatively uniform porous structure, a high specific surface area of 113.98 m2 /g, and multiple functional chemical groups according to the characterization results. An online solid-phase extraction-high-performance liquid chromatography procedure was fabricated to extract and determine tussilagone from Farfarae Flos. The results show that the complex sample matrices can be removed in the solid-phase extraction procedure. Simultaneously, tussilagone can remain, which can be subsequently switched to an octadecylsilane bonded analytical column. The methodological validation showed that the correlation coefficient was 0.9999 with a linear range of 0.6-200.0 µg/mL, the limit of detection was 0.2 µg/mL, the limit of quantification was 0.6 µg/mL, accuracy was 100.3-100.6%, and relative standard deviation of precision was ≤1.9%. The present monolithic cartridge exhibits good reusability of not more than 100 times. The real sample of Farfarae Flos was determined with a tussilagone content of 0.74 mg/g.

Progress in Traditional Chinese Medicine Against Respiratory Viruses: A Review.

Respiratory viruses, such as severe acute respiratory syndrome coronavirus (SARS-CoV)-1, SARS-CoV-2, influenza A viruses, and respiratory syncytial virus, pose a serious threat to society. Based on the guiding principles of "holism" and "syndrome differentiation and treatment", traditional Chinese medicine (TCM) has unique advantages in the treatment of respiratory virus diseases owing to the synergistic effect of multiple components and targets, which prevents drug resistance from arising. According to TCM theory, there are two main strategies in antiviral treatments, namely "dispelling evil" and "fu zheng". Dispelling evil corresponds to the direct inhibition of virus growth and fu zheng corresponds to immune regulation, inflammation control, and tissue protection in the host. In this review, current progress in using TCMs against respiratory viruses is summarized according to modern biological theories. The prospects for developing TCMs against respiratory viruses is discussed to provide a reference for the research and development of innovative TCMs with multiple components, multiple targets, and low toxicity.

A comprehensive review on phytochemistry, bioactivities, toxicity studies, and clinical studies on Ficus carica Linn. leaves.

The leaves of Ficus carica Linn. (FC) have been widely used for medicine purposes since ancient times, and its decoction is consumed as tea. Many scientific papers have been published in the literature and the researchers across the world are still exploring the health benefits of FC leaves. In this review, we have collected the literature published since 2010 in the databases: Pubmed, Scopus, Web of Science, SciFinder, Google Scholar, Baidu Scholar and local classic herbal literature. The summary of the chemical constituents in FC leaves, biological activities, toxicity studies, and clinical studies carried out on FC leaves is provided in this review. In addition, the molecular mechanisms of the active constituents in FC leaves are also comprehended. FC leaves are reported to 126 constituents out of which the polyphenolic compounds are predominant. Many scientific studies have proven the antidiabetic, antioxidant, anti-inflammatory, anticancer, anticholinesterase, antimicrobial, hepatoprotective, and renoprotective activities. Many studies have carried out to provide the insights on molecular pathways involved in the biological activities of FC leaves. The toxicity studies have suggested that FC leaves exhibit toxicity only at very high doses. We believe this review serve as a comprehensive resource for those who are interested to understand the scientific evidence that support the medicinal values of FC leaves and also the research gaps to further improve the commercial value and health benefits of FC leaves.

An L-type lectin receptor-like kinase promotes starch accumulation during rice pollen maturation.

Starch accumulation is key for the maturity of rice pollen grains; however, the regulatory mechanism underlying this process remains unknown. Here, we have isolated a male-sterile rice mutant, abnormal pollen 1 (ap1), which produces nonviable pollen grains with defective starch accumulation. Functional analysis revealed that AP1 encodes an active L-type lectin receptor-like kinase (L-LecRLK). AP1 is localized to the plasma membrane and its transcript is highly accumulated in pollen during the starch synthesis phase. RNA-seq and phosphoproteomic analysis revealed that the expression/phosphorylation levels of numerous genes/proteins involved in starch and sucrose metabolism pathway were significantly altered in the mutant pollen, including a known rice UDP-glucose pyrophosphorylase (OsUGP2). We further found that AP1 physically interacts with OsUGP2 to elevate its enzymatic activity, likely through targeted phosphorylation. These findings revealed a novel role of L-LecRLK in controlling pollen maturity via modulating sucrose and starch metabolism.

Multiple modes of action of myricetin in influenza A virus infection.

Influenza A virus remains a major threat to public health worldwide after its first pandemic. Scientists keep searching novel anti-influenza drugs, of which natural products present to be an important source. Myricetin, a natural flavonol compound, which exists in many edible plants, which has a wide range of biological activities, but its anti-influenza A virus activity is ambiguous. This study aims to evaluate the anti-influenza activity of myricetin and elucidate its underlying mechanism. Our results demonstrated that myricetin could significantly inhibit influenza A virus replication, reduce viral polymerase activity via selective inhibition of viral PB2 subunit, and the production of inflammatory cytokines by inhibiting TLR3 signaling pathway. The binding affinity analysis and the result of molecular docking revealed that myricetin interacted with the PB2 cap-binding pocket of influenza A virus. The above results suggested myricetin could exhibit anti-influenza virus activity with low cytotoxicity as well, and myricetin had low toxicity in BALB/c mice in vivo. Results from this study highlighted myricetin could be considered as a promising anti-influenza virus agent with dual inhibition profile. Furthermore, the compound with similar structure would provide a new option for the development of novel inhibitors against influenza A virus.

Effects of Auricular Point Acupressure on Pain Relief: A Systematic Review.

OBJECTIVES: This systematic review aimed to summarize and provide an update on clinical studies investigating the effects of auricular point acupressure (APA) on pain relief, in addition to the APA methods of delivery and operation. DESIGN: A systematic review. DATA SOURCES: A systematic review on literatures published on five English (PubMed, Web of Science, Embase, EBSCO, and Cochrane databases) and four major Chinese databases (China National Knowledge Infrastructure, Wan Fang Data, Chinese Scientific Journals Database [VIP], and SinoMed) was conducted. METHOD: We screened nine electronic databases from the time of their respective establishment until December 20, 2019. Randomized controlled trials and studies that defined an APA intervention measure and evaluated pain intensity were considered. We individually categorized and analyzed 46 studies considering the following: (1) acute or chronic pain and (2) whether the outcomes positively or negatively support the effectiveness of APA on pain intensity. We also summarized the methods of delivery used (including the acupoint selection, stimulator selection, method of taping seeds on the ears, frequency of replacing seeds, suitability of acupressure intensity, acupressure frequency, and pressing time) and APA operator. RESULTS: Regardless of pain intensity, APA effectively treated most acute pain when combined with other interventions. Although it was used alone to treat low back pain and dysmenorrhea, other chronic diseases typically underwent a combination of APA with other interventions. The 43 positive studies revealed that acute pain required shorter APA intervention periods than chronic pain. Corresponding acupoints and nervous system acupoints were chosen. Vaccaria seeds, the single-ear method (including the alternate-ear method), and daily seed replacement were commonly adopted. Deqi was considered an effective signal for appropriate acupressure intensity. Additionally, the patients could effectively apply acupressure. CONCLUSION: This systematic review revealed important trends in APA treatments, which could be essential in determining treatment efficacy.

Indole primes defence signalling and increases herbivore resistance in tea plants.

Upon herbivore attack, plants emit herbivore-induced plant volatiles (HIPVs). HIPVs can prime defences and resistance of intact plants. However, how HIPVs are decoded and translated into functional defence responses is not well understood, especially in long-lived woody plants. Here, we investigated the impact of the aromatic HIPV indole on defence-related early signalling, phytohormone accumulation, secondary metabolite biosynthesis and herbivore resistance in tea plants. We find that tea plants infested with tea geometrid caterpillars release indole at concentrations >450 ng*hr-1 . Exposure to corresponding doses of synthetic indole primes the expression of early defence genes involved in calcium (Ca2+ ) signalling, MPK signalling and jasmonate biosynthesis. Indole exposure also primes the production of jasmonates and defence-related secondary metabolites. These changes are associated with higher herbivore resistance of indole-exposed tea plants. Chemical inhibition of Ca2+ and jasmonate signalling provides evidence that both are required for indole-mediated defence priming and herbivore resistance. Our systematic assessment of the impact of indole on defence signalling and deployment shows that indole acts by boosting Ca2+ signalling, resulting in enhanced jasmonate-dependent defence and resistance in a woody plant. Our work extends the molecular basis of HIPV-induced defence priming from annual plants to an economically important tree species.

Mechanisms Underlying the Cardioprotection of YangXinDingJi Capsule against Myocardial Ischemia in Rats.

BACKGROUND: YangXinDingJi (YXDJ) capsule is one of traditional Chinese medicines (TCMs) derived from Zhigancao decoction, which is usually used for the treatment of cardiovascular disease in China. Aim of the Study. Cardiovascular events are one of the leading causes of death worldwide. Myocardial ischemia (MI) severely reduces myocyte longevity and function. The YangXinDingJi (YXDJ) capsule has been used in the treatment of clinical cardiac disease in China. Nevertheless, the underlying cellular mechanisms for the benefits to heart function resulting from the use of this capsule are still unclear. The aim of this study was to evaluate the protective effects of the YXDJ on isoprenaline-induced MI in rats and to clarify its underlying myocardial protective mechanisms based on L-type calcium channels and myocardial contractility. MATERIALS AND METHODS: Rats were randomly divided into five groups with ten rats in each group: (1) control; (2) ISO-induced model; (3) high-dose YXDJ (2.8 g/kg/day intraperitoneally for five days), (4) low-dose YXDJ (1.4 g/kg/day for five days); and (5) verapamil (n = 10 in each group). Isoproterenol (ISO) was injected subcutaneously for two consecutive days to induce the rat model of MI. Heart and biochemical parameters were obtained. The patch-clamp technique was used to observe the regulatory effects of YXDJ on the L-type calcium current (ICa-L) in isolated cardiomyocytes. An IonOptix MyoCam detection system was used to observe the contractility of YXDJ on isolated cardiomyocytes. RESULTS: YXDJ caused a significant improvement in pathological heart morphology and alleviated oxidative stress and inflammatory responses. Exposure to YXDJ caused a decrease in blockade of ICa-L in a concentration-dependent manner. CONCLUSIONS: The results indicate that YXDJ significantly inhibited inflammatory cytokine expressions, oxidative stress, and L-type Ca2+ channels, and decreased contractility in isolated rat cardiomyocytes. These findings may be relevant to the cardioprotective efficacy of YXDJ.

Positive detection of SARS-CoV-2 combined HSV1 and HHV6B virus nucleic acid in tear and conjunctival secretions of a non-conjunctivitis COVID-19 patient with obstruction of common lacrimal duct.

BACKGROUND: The current outbreak of COVID-19 has spread rapidly all over the world. Respiratory droplets and contaction with infected patients are the two major transmission routes. However, the value of tear virus nucleic acid is still not clear. We dynamic detected the SARS-CoV-2 in eye sample of one COVID-19 patient with obstruction of common lacrimal ducts. METHODS: Besides the routine examination, nasopharyngeal and eye swab were continuously measured by polymerase chain reaction assay and next-generation sequencing (NGS). Gene detection was performed for drug use guidance, and flow cytometry was performed to analyse the lymphocyte subsets. RESULTS: Nasopharyngeal swabs were positive for 22 days, but eye swabs were still continuously positive for 2 weeks after nasopharyngeal swabs turned negative. The low level of lymphocyte and the high level IL-6 lasted for almost 4 weeks, then became near normal. Next-generation sequencing (NGS) confirmed the existing of SARS-CoV-2, HSV1 and HHV6B virus nucleic acid. The gene detection for drug use guidance showed the genetic locus ABCB1 (3435T>C) rs1045642 belonged to type CC and it mean the efficiency of lopinavir-ritonavir would be significantly decreased. The flow cytometry of lymphocyte subsets showed PD-1+  CD95+ cells was accounting for 94.8% in CD3+  CD8+ T subset and for 94.8% in CD3+  TCRγδ+ T subset. CONCLUSIONS: As obstruction of common lacrimal duct, positively detection in one eye for 2 weeks more after nasopharyngeal swab became negative. More eye swabs should be collected from COVID-19 patients, especially from those immunocompromised, those with eye symptoms and those had a history of ocular diseases.

Discovery of a Natural Product That Binds to the Mycobacterium tuberculosis Protein Rv1466 Using Native Mass Spectrometry.

Elucidation of the mechanism of action of compounds with cellular bioactivity is important for progressing compounds into future drug development. In recent years, phenotype-based drug discovery has become the dominant approach to drug discovery over target-based drug discovery, which relies on the knowledge of a specific drug target of a disease. Still, when targeting an infectious disease via a high throughput phenotypic assay it is highly advantageous to identifying the compound's cellular activity. A fraction derived from the plant Polyalthia sp. showed activity against Mycobacterium tuberculosis at 62.5 µge/µL. A known compound, altholactone, was identified from this fraction that showed activity towards M. tuberculosis at an minimum inhibitory concentration (MIC) of 64 µM. Retrospective analysis of a target-based screen against a TB proteome panel using native mass spectrometry established that the active fraction was bound to the mycobacterial protein Rv1466 with an estimated pseudo-Kd of 42.0 ± 6.1 µM. Our findings established Rv1466 as the potential molecular target of altholactone, which is responsible for the observed in vivo toxicity towards M. tuberculosis.