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Journal of Food Biochemistry Oct 2022Coronavirus disease 2019 (COVID-19) is caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Several vaccines against SARS-CoV-2 have been...
Coronavirus disease 2019 (COVID-19) is caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Several vaccines against SARS-CoV-2 have been approved; however, variants of concern (VOCs) can evade vaccine protection. Therefore, developing small compound drugs that directly block the interaction between the viral spike glycoprotein and ACE2 is urgently needed to provide a complementary or alternative treatment for COVID-19 patients. We developed a viral infection assay to screen a library of approximately 126 small molecules and showed that peimine inhibits VOCs viral infections. In addition, a fluorescence resonance energy transfer (FRET) assay showed that peimine suppresses the interaction of spike and ACE2. Molecular docking analysis revealed that peimine exhibits a higher binding affinity for variant spike proteins and is able to form hydrogen bonds with N501Y in the spike protein. These results suggest that peimine, a compound isolated from Fritillaria, may be a potent inhibitor of SARS-CoV-2 variant infection. PRACTICAL APPLICATIONS: In this study, we identified a naturally derived compound of peimine, a major bioactive alkaloid extracted from Fritillaria, that could inhibit SARS-CoV-2 variants of concern (VOCs) viral infection in 293T/ACE2 and Calu-3 lung cells. In addition, peimine blocks viral entry through interruption of spike and ACE2 interaction. Moreover, molecular docking analysis demonstrates that peimine has a higher binding affinity on N501Y in the spike protein. Furthermore, we found that Fritillaria significantly inhibits SARS-CoV-2 viral infection. These results suggested that peimine and Fritillaria could be a potential functional drug and food for COVID-19 patients.
Topics: Angiotensin-Converting Enzyme 2; Binding Sites; COVID-19 Vaccines; Cevanes; Glycoproteins; Humans; Molecular Docking Simulation; Peptidyl-Dipeptidase A; Protein Binding; SARS-CoV-2; Spike Glycoprotein, Coronavirus; Viral Proteins; Virus Internalization; COVID-19 Drug Treatment
PubMed: 35894128
DOI: 10.1111/jfbc.14354 -
Toxins Jul 2022Jervine, protoveratrine A (proA), and protoveratrine B (proB) are alkaloids that are presented in some remedies obtained from , such as . This paper reports on a...
Jervine, protoveratrine A (proA), and protoveratrine B (proB) are alkaloids that are presented in some remedies obtained from , such as . This paper reports on a single-center pilot cardiotoxic mechanism study of jervine, proA, and proB in case series. The molecular aspects were studied via molecular dynamic simulation, molecular docking with cardiac sodium channel Na1.5, and machine learning-based structure-activity relationship modeling. HPLC-MS/MS method in combination with clinical events were used to analyze alkaloid cardiotoxicity in patients. Jervine demonstrates the highest docking score (-10.8 kcal/mol), logP value (4.188), and p value (9.64) compared with proA and proB. Also, this compound is characterized by the lowest calculated IC. In general, all three analyzed alkaloids show the affinity to Na1.5 that highly likely results in cardiotoxic action. The clinical data of seven cases of intoxication by confirms the results of molecular modeling. Patients exhibited nausea, muscle weakness, bradycardia, and arterial hypotension. The association between alkaloid concentrations in blood and urine and severity of patient condition is described. These experiments, while primary, confirmed that jervine, proA, and proB contribute to cardiotoxicity by Na1.5 inhibition.
Topics: Alkaloids; Cardiotoxicity; Humans; Molecular Docking Simulation; Pilot Projects; Tandem Mass Spectrometry; Veratrum; Veratrum Alkaloids
PubMed: 35878228
DOI: 10.3390/toxins14070490 -
American Journal of Physiology. Cell... Sep 2022Isolated smooth muscle cells (SMCs) from mouse bronchus were studied using the whole cell patch-clamp technique at ∼21°C. Stepping from -100 mV to -20 mV evoked...
Isolated smooth muscle cells (SMCs) from mouse bronchus were studied using the whole cell patch-clamp technique at ∼21°C. Stepping from -100 mV to -20 mV evoked inward currents of mean amplitude -275 pA. These inactivated (tau = 1.1 ms) and were abolished when external Na was substituted with -Methyl-d-glucamine. In current-voltage protocols, current peaked at -10 mV and reversed between +20 and +30 mV. The Vs of activation and inactivation were -25 and -86 mV, respectively. The current was highly sensitive to tetrodotoxin (IC = 1.5 nM) and the Na1.7 subtype-selective blocker, PF-05089771 (IC = 8.6 nM), consistent with Na1.7 as the underlying pore-forming α subunit. Two Na1.7-selective antibodies caused membrane-delineated staining of isolated SMC, as did a nonselective pan-Na antibody. RT-PCR, performed on groups of ∼15 isolated SMCs, revealed transcripts for Na1.7 in 7/8 samples. Veratridine (30 µM), a nonselective Na channel activator, reduced peak current evoked by depolarization but induced a sustained current of 40 pA. Both effects were reversed by tetrodotoxin (100 nM). In tension experiments, veratridine (10 µM) induced contractions that were entirely blocked by atropine (1 µM). However, in the presence of atropine, veratridine was able to modulate the pattern of activity induced by a combination of U-46619 (a thromboxane A2 mimetic) and PGE (prostaglandin E), by eliminating bursts in favor of sustained phasic contractions. These effects were readily reversed to control-like activity by tetrodotoxin (100 nM). In conclusion, mouse bronchial SMCs functionally express Na1.7 channels that are capable of modulating contractile activity, at least under experimental conditions.
Topics: Animals; Atropine Derivatives; Bronchi; Mice; Myocytes, Smooth Muscle; Sodium; Tetrodotoxin; Veratridine
PubMed: 35876287
DOI: 10.1152/ajpcell.00011.2022 -
Computational and Mathematical Methods... 2022and record that Radix Veratri root is and the root of . According to () , Radix Veratri is a Liliaceae plant Veratrum taliense. Another literature pointed out that...
BACKGROUND
and record that Radix Veratri root is and the root of . According to () , Radix Veratri is a Liliaceae plant Veratrum taliense. Another literature pointed out that the aliases of Veratrum taliense and Veratrum angustifolia are both Radix Veratri, and their effects are basically the same. The main active ingredient of Veratrum is veratramine, of which veratramine and Jervine are higher in content, reaching 24.60% and 21.28% of the total alkaloids, respectively. Veratrum alkaloids are both toxic and effective ingredients. In addition to its good clinical efficacy, attention should also be paid to its pharmacokinetic characteristics in vivo. It is particularly important to study the pharmacokinetic characteristics of veratramine and Jervine in vivo.
OBJECTIVE
The goal of this study was to develop a simple and effective method for measuring veratramine and Jervine in rat plasma at the same time. This method was used to study the pharmacokinetic characteristics of veratramine and Jervine in the alcohol extract of Radix Veratri in rats, to provide a reasonable basis for the clinical use of Radix Veratri.
METHODS
Eighteen SD rats were randomly assigned into three groups, half male and half female, and were given 0.04 g/kg, 0.08g/kg, and 0.16 g/kg Radix Veratri alcohol extract, respectively. Blood samples were collected at different time points and were analyzed by LC-MS/MS after protein precipitation. Bullatine was set as the internal standard; the plasma samples were extracted with ethyl acetate. After the sample was processed, acetonitrile-10 mM ammonium acetate, whose pH was adjusted to 8.8 with ammonia water, was taken as the mobile phase. Veratramine quantitative ion pair was 410.1⟶295.1/, Jervine quantitative ion pair was 426.2⟶114.1/, and Bullatine B (IS) quantitative ion pair was 438.2⟶420.1/. In the positive ion mode, the multireaction monitoring (MRM) mode was used to determine the blood concentration of veratramine and Jervine. DAS 3.3.0 was used to calculate the relevant pharmacokinetic parameters.
RESULTS
Veratramine had a good linear relationship in the concentration range of 0.0745~18.2 ng/mL, and that of Jervine was 1.11~108 ng/mL. The correlation coefficient of three consecutive batches of the standard curve was greater than 0.995. Veratramine's lower quantification limit was 0.745 ng/mL, Jervine's was 1.11 ng/mL, and precision and accuracy were both less than 15%. The accuracy of veratramine was between 88.96% and 101.85%, and the accuracy of Jervine was between 92.96% and 104.50%. This method was adopted for the pharmacokinetic study of alcohol extracts of Radix Veratri. The results showed that only of veratramine female rats did not show linear kinetic characteristics in the dose range of Radix Veratri alcohol extract from 0.04 g/kg to 0.16 g/kg. For AUC and of veratramine and Jervine, it could not determine whether the Radix Veratri alcohol extract showed linear kinetic characteristics within the dosage range of 0.04 g/kg~0.16 g/kg. Veratramine and Jervine showed obvious gender differences in the absorption and elimination stages. The absorption rate of veratramine and Jervine by male mice was about 10 times higher than that of female mice, and the elimination rate of male mice is about 20 times lower than that of female mice. It was suggested that the clinical application of the steroidal alkaloids veratramine and Jervine in Radix Veratri required rational use of drugs based on gender.
CONCLUSION
An LC-MS/MS analysis method suitable for the pharmacokinetic study of veratramine and Jervine in Radix Veratri in SD rats was established to provide a basis for in vivo pharmacokinetic studies. The pharmacokinetic characteristics of veratramine and Jervine in the alcohol extract of Radix Veratri were significantly different in female and male rats. During the clinical use of Radix Veratri, it should pay close attention to the obvious gender differences that may occur after the medication.
Topics: Alkaloids; Animals; Chromatography, Liquid; Female; Humans; Male; Mice; Plant Extracts; Rats; Rats, Sprague-Dawley; Tandem Mass Spectrometry; Veratrum; Veratrum Alkaloids
PubMed: 35785141
DOI: 10.1155/2022/8289548 -
Ecotoxicology and Environmental Safety Jul 2022Fine particulate matter (PM2.5) exposure can cause lung injury and a large number of respiratory diseases. Sipeimine is a steroidal alkaloid isolated from Fritillaria...
Fine particulate matter (PM2.5) exposure can cause lung injury and a large number of respiratory diseases. Sipeimine is a steroidal alkaloid isolated from Fritillaria roylei which has been associated with anti-inflammatory, antitussive and antiasthmatic properties. In this study, we explored the potential effects of sipeimine against PM2.5-induced lung injury in Sprague Dawley rats. Sipeimine alleviated lung injury caused by PM2.5 and decreased pulmonary edema, inflammation and the levels of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in the bronchoalveolar lavage fluid. In addition, sipeimine upregulated the glutathione (GSH) expression and downregulated the expression of 4-hydroxynonenal (4-HNE), tissue iron and malondialdehyde (MDA). The downregulation of proteins involved in ferroptosis, including nuclear factor E2-related factor 2 (Nrf2), glutathione peroxidase 4 (GPX4), heme oxygenase-1 (HO-1) and solute carrier family 7 member 11 (SLC7A11) was reversed by sipeimine. The administration of RSL3, a potent ferroptosis-triggering agent, blocked the effects of sipeimine. Using network pharmacology, we found that the effects of sipeimine were presumably mediated through the phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signaling pathway. A PI3K inhibitor (LY294002) blocked the PI3K/Akt signaling pathway and reversed the effects of sipeimine. Overall, this study suggested that the protective effect of sipeimine against PM2.5-induced lung injury was mainly mediated through the PI3K/Akt pathway, ultimately leading to a reduction in ferroptosis.
Topics: Animals; Cevanes; Ferroptosis; Lung Injury; NF-E2-Related Factor 2; Network Pharmacology; Particulate Matter; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley
PubMed: 35567927
DOI: 10.1016/j.ecoenv.2022.113615 -
Frontiers in Plant Science 2022Pimacao is a traditional Chinese folk medicine and is the main component of the famous Chinese herbal remedy "Yunnan Baiyao" for its significant analgesic activity in...
Pimacao is a traditional Chinese folk medicine and is the main component of the famous Chinese herbal remedy "Yunnan Baiyao" for its significant analgesic activity in the treatment of wounds. Due to increases in consumption, its wild population is now difficult to find, and adulterant from the same genus has occurred. However, this is challenging to distinguish the species of in Pimacao using dried roots and rhizomes or medicinal powder. ITS2 sequences and steroidal alkaloids by the non-targeted and pseudo-targeted metabolomics methods were taken advantage of establishing an effective identification method. Based on the ITS2 sequence, metabolite profiling of steroidal alkaloids and morphological characteristics, the classification of two distinct subspecies in has been reinforced. In addition, the new subspecies subsp. was collected in China for the first time. The ITS2 sequence could be used in the identification of , , , and , but is insufficient for intraspecific identification. Simultaneously, 147 variables were labeled by non-targeted analysis accomplished utilizing an ultra-high-performance liquid chromatography electrospray ionization orbitrap tandem mass spectrometry (UPLC-ESI-QE-Orbitrap-MS) system consisting of an Orbitrap QE HF-X. Followed by a pseudo-targeted analysis method developed for the Qtrap 6500-plus mass spectrometry system coupled with an ESI source, 29 labeled steroidal alkaloids detected by the MRM mode could distinguish between four species. Notably, 25 labeled steroidal alkaloids could distinguish between three closely related species. These have the potential to be used as markers for identification. Furthermore, there were several variables with statistical differences between two subspecies of and populations of , , and .
PubMed: 35481147
DOI: 10.3389/fpls.2022.831562 -
International Journal of Molecular... May 2022Clear cell renal cell carcinoma (ccRCC) is the most common and aggressive subtype of kidney cancer, with high mortality rates worldwide. The sonic hedgehog (SHH)...
Clear cell renal cell carcinoma (ccRCC) is the most common and aggressive subtype of kidney cancer, with high mortality rates worldwide. The sonic hedgehog (SHH) molecular cascade is altered in various malignancies in tumorigenesis, and several SHH pathway inhibitors have been considered as potential anticancer drugs. The aim of the present study was to determine the expression profile of SHH signaling components and their target genes in ccRCC. Additionally, the present study examined the effects of SHH pathway inhibitory drugs (RU‑SKI43, cyclopamine and GLI‑antagonist 61) on cell viability, cell cycle progression, expression levels of SHH target genes and migration ability in 786‑O, ACHN and HK2 cells. The study also included paired tumor and normal samples from 62 patients with ccRCC. The mRNA levels in clinical samples and cell lines were measured via reverse transcription‑quantitative PCR. Cell viability was examined using a sulforhodamine B assay. Flow cytometry was used to investigate cell cycle progression and the migratory rate of cells was assessed using a wound healing assay. High mRNA levels of , smoothened (), glioma‑associated zinc finger protein (), BCL2 apoptosis regulator (), MYC proto‑oncogene (), vascular endothelial growth factor A () and cyclin D1 () were observed in the tumor tissues, especially in early ccRCC, according to the TNM stage or World Health Organization/International Society of Urological Pathology (ISUP) grade. High expression levels of , as well as low mRNA expression, were associated with short overall survival, and increased expression was an independent prognostic factor of a poor outcome in patients with advanced ISUP grade (Cox hazard ratio test). Cyclopamine treatment was found to arrest 786‑O cells in the G/M phase and decreased the expression levels of , , and . RU‑SKI43 inhibited cell migration and decreased the expression levels of , and in ACHN cells. Overall, the results of the present study suggested that SHH signaling may be involved in the early development of ccRCC, and the expression levels of and may serve as prognostic factors of this disease. Cyclopamine and RU‑SKI43 appear to be potential anti‑renal cell carcinoma drugs; however, this hypothesis requires verification by further studies.
Topics: Carcinoma, Renal Cell; Hedgehog Proteins; Humans; Kidney Neoplasms; Vascular Endothelial Growth Factor A; Veratrum Alkaloids; Zinc Finger Protein GLI1
PubMed: 35266008
DOI: 10.3892/ijmm.2022.5114 -
International Journal of Molecular... Feb 2022The cardiac sodium ion channel (Na1.5) is a protein with four domains (DI-DIV), each with six transmembrane segments. Its opening and subsequent inactivation results in...
The cardiac sodium ion channel (Na1.5) is a protein with four domains (DI-DIV), each with six transmembrane segments. Its opening and subsequent inactivation results in the brief rapid influx of Na ions resulting in the depolarization of cardiomyocytes. The neurotoxin veratridine (VTD) inhibits Na1.5 inactivation resulting in longer channel opening times, and potentially fatal action potential prolongation. VTD is predicted to bind at the channel pore, but alternative binding sites have not been ruled out. To determine the binding site of VTD on Na1.5, we perform docking calculations and high-throughput electrophysiology experiments in the present study. The docking calculations identified two distinct binding regions. The first site was in the pore, close to the binding site of Na1.4 and Na1.5 blocking drugs in experimental structures. The second site was at the "mouth" of the pore at the cytosolic side, partly solvent-exposed. Mutations at this site (L409, E417, and I1466) had large effects on VTD binding, while residues deeper in the pore had no effect, consistent with VTD binding at the mouth site. Overall, our results suggest a VTD binding site close to the cytoplasmic mouth of the channel pore. Binding at this alternative site might indicate an allosteric inactivation mechanism for VTD at Na1.5.
Topics: Binding Sites; Cell Line; HEK293 Cells; Humans; Ion Channel Gating; Mouth; NAV1.5 Voltage-Gated Sodium Channel; Neurotoxins; Sodium; Veratridine
PubMed: 35216338
DOI: 10.3390/ijms23042225 -
Microbiology Spectrum Feb 2022The limited number of available effective agents necessitates the development of new antifungals. We report that jervine, a jerveratrum-type steroidal alkaloid isolated...
The limited number of available effective agents necessitates the development of new antifungals. We report that jervine, a jerveratrum-type steroidal alkaloid isolated from Veratrum californicum, has antifungal activity. Phenotypic comparisons of cell wall mutants, K1 killer toxin susceptibility testing, and quantification of cell wall components revealed that β-1,6-glucan biosynthesis was significantly inhibited by jervine. Temperature-sensitive mutants defective in essential genes involved in β-1,6-glucan biosynthesis, including , , , , and , were hypersensitive to jervine. In contrast, point mutations in or its paralog produced jervine resistance, suggesting that jervine targets Kre6 and Skn1. Jervine exhibited broad-spectrum antifungal activity and was effective against human-pathogenic fungi, including Candida parapsilosis and Candida krusei. It was also effective against phytopathogenic fungi, including Botrytis cinerea and Puccinia recondita. Jervine exerted a synergistic effect with fluconazole. Therefore, jervine, a jerveratrum-type steroidal alkaloid used in pharmaceutical products, represents a new class of antifungals active against mycoses and plant-pathogenic fungi. Non-Candida albicans species (NCAC) are on the rise as a cause of mycosis. Many antifungal drugs are less effective against NCAC, limiting the available therapeutic agents. Here, we report that jervine, a jerveratrum-type steroidal alkaloid, is effective against NCAC and phytopathogenic fungi. Jervine acts on Kre6 and Skn1, which are involved in β-1,6-glucan biosynthesis. The skeleton of jerveratrum-type steroidal alkaloids has been well studied, and more recently, their anticancer properties have been investigated. Therefore, jerveratrum-type alkaloids could potentially be applied as treatments for fungal infections and cancer.
Topics: Alkaloids; Antifungal Agents; Candida; Cell Wall; Fungi; Humans; Mycoses; Plant Extracts; Veratrum; beta-Glucans
PubMed: 35019680
DOI: 10.1128/spectrum.00873-21 -
British Journal of Pharmacology Mar 2022Corpus cavernosum smooth muscle (CCSM) exhibits phasic contractions that are coordinated by ion channels. Mouse models are commonly used to study erectile dysfunction,...
BACKGROUND AND PURPOSE
Corpus cavernosum smooth muscle (CCSM) exhibits phasic contractions that are coordinated by ion channels. Mouse models are commonly used to study erectile dysfunction, but there are few published electrophysiological studies of mouse CCSM. We describe the voltage-dependent sodium (Na ) currents in mouse CCSM and investigate their function.
EXPERIMENTAL APPROACH
We used electrophysiological, pharmacological and immunocytochemical methods to study the Na currents in isolated CCSM cells from C57BL/6 mice. Tension measurements were carried out using crural sections of the corpus cavernosum in whole tissue.
KEY RESULTS
Fast, voltage-dependent, sodium currents in mouse CCSM were induced by depolarising steps. Steady-state activation and inactivation curves revealed a window current between -60 and -30 mV. Two populations of Na currents, 'TTX-sensitive' and 'TTX-insensitive', were identified. TTX-sensitive currents showed 48% block with the Na channel subtype-specific blockers ICA-121431 (Na 1.1-1.3), PF-05089771 (Na 1.7) and 4,9-anhydro-TTX (Na 1.6). TTX-insensitive currents were resistant to blockade by A803467, specific for Na 1.8 channels. Immunocytochemistry confirmed expression of Na 1.5 and Na 1.4 in freshly dispersed CCSM cells. Veratridine, a Na channel activator, reduced time-dependent inactivation of Na currents and increased duration of evoked action potentials. Veratridine induced phasic contractions in CCSM strips, reversible with TTX and nifedipine but not KB-R7943.
CONCLUSION AND IMPLICATIONS
There are fast, voltage-dependent, sodium currents in mouse CCSM. Stimulation of these currents increased contractility of CCSM in vitro, suggesting an involvement in detumescence and potentially providing a clinically relevant target in erectile dysfunction. Further work will be necessary to define its role.
Topics: Animals; Erectile Dysfunction; Humans; Male; Mice; Mice, Inbred C57BL; Myocytes, Smooth Muscle; Sodium; Sodium Channel Blockers; Tetrodotoxin; Veratridine
PubMed: 34767251
DOI: 10.1111/bph.15728