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1.
J Ethnopharmacol ; 289: 115019, 2022 May 10.
Article in English | MEDLINE | ID: mdl-35074453

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE: Hydrocotyle umbellata L. is a medicinal herb for the treatment of some health problems including hypertension, according to traditional medicine. Even so, its vascular effects and the pharmacological action mechanisms have not been analyzed. AIM OF THE STUDY: This experiment aimed to analyze the effects of hydroalcoholic extract of Hydrocotyle umbellata L. (HEHU) on isolated vessels and verify the interaction of hibalactone (chemical marker) against Cav1.2 channels using molecular docking. MATERIALS AND METHODS: Vascular reactivity experiments were performed using rat aortas with (E+) or without endothelium (E-) in an isolated organ bath. Computational molecular docking approaches were used to show the direct effect on L-type Ca2+ Channels. RESULTS: HEHU (0-560 µg/mL) induced relaxation of the pre-contracted arteries in a concentration-dependent manner. The maximum effect was higher in E+ (76.8 ± 4.1%) as compared to E- (47.3 ± 5.5%). Pre-treatment of E+ arteries with L-NAME or ODQ reduced the relaxation to similar level of E- arteries. The treatment of arteries with MDL-12,330 A, diclofenac, propranolol and atropine did not change the relaxation induced by HEHU. The contraction caused by internal Ca2+ release induced by caffeine was reduced after HEHU treatment. Moreover, the HEHU also impaired the contraction induced by Ca2+ influx stimulated with phenylephrine or high KCl. The docking study demonstrated the effectiveness of hibalactone in blocking the Cav1.2 channel. CONCLUSIONS: These findings show that HEHU induces vascular relaxation which is potentiated (but not dependent) by endothelial cells. Blocking of Ca2+ influx seems to be the main mechanism for the vascular effects of HEHU.


Subject(s)
Calcium Channel Blockers/pharmacology , Calcium Channels, L-Type/drug effects , Centella/chemistry , Plant Extracts/pharmacology , Animals , Aorta/drug effects , Aorta/metabolism , Calcium/metabolism , Calcium Channel Blockers/isolation & purification , Male , Molecular Docking Simulation , Rats , Rats, Wistar , Vasodilation/drug effects , Vasodilator Agents/isolation & purification , Vasodilator Agents/pharmacology
2.
Biomed Pharmacother ; 144: 112231, 2021 Dec.
Article in English | MEDLINE | ID: mdl-34610498

ABSTRACT

The hypotensive and antihypertensive activities of the aqueous extract (AE) and butanolic fraction (ButF) isolated from Cecropia glaziovii Sneth have been demonstrated in previous studies in animal models. This study aimed to evaluate the molecular mechanism of action responsible for the vasodilatory effect of procyanidins, flavanols, and flavonoids found in C. glaziovii in endothelial cell culture. For this purpose, we analyzed the effect of procyanidin B2 and B3 compounds, catechin, epicatechin, orientin, isoorientin, and isovitexin in the mobilization of Ca2+ in rat endothelial cell cultures. Parallel associations with different antagonists were examined by considering the following in vivo hypotensive mechanisms: blockage of L-type calcium channels, action on ß-2 adrenergic receptors, and vasodilation via the nitric oxide pathway. All measurements of calcium mobilization were carried out by using the fluorescence measurement methodology in a Flexstation M3 spectrophotometer. The results indicate that some of the compounds have mixed actions, acting through different calcium mobilization pathways. The mobilization induced by such compounds significantly decreased when they were incubated with their corresponding antagonists. Taken together, our data suggest that the beneficial effects seen with the popular use of Cecropia glaziovii Sneth in pathological conditions, such as systemic arterial hypertension, seem to be related to the plant's hypotensive effect, very probably promoted by the actions of flavonols, flavonoids, and procyanidins, by different pathways of calcium mobilization.


Subject(s)
Calcium Channel Blockers/pharmacology , Calcium Channels, L-Type/drug effects , Cecropia Plant , Endothelial Cells/drug effects , Flavonoids/pharmacology , Flavonols/pharmacology , Lung/blood supply , Phytochemicals/pharmacology , Plant Extracts/pharmacology , Proanthocyanidins/pharmacology , Vasodilator Agents/pharmacology , Animals , Calcium Channel Blockers/isolation & purification , Calcium Channels, L-Type/metabolism , Calcium Signaling/drug effects , Cecropia Plant/chemistry , Cells, Cultured , Endothelial Cells/metabolism , Flavonoids/isolation & purification , Flavonols/isolation & purification , Male , Phytochemicals/isolation & purification , Phytotherapy , Plant Extracts/isolation & purification , Proanthocyanidins/isolation & purification , Rats, Wistar , Vasodilator Agents/isolation & purification
3.
Bioorg Chem ; 107: 104523, 2021 02.
Article in English | MEDLINE | ID: mdl-33339668

ABSTRACT

Chuanxiongdiolides R4-R6 (1-3), three novel phthalide dimers featuring two classes of unreported monomeric units (ligustilide/senkyunolide A and ligustilide/neocnidilide) with an unprecedented linkage style (3a,7'/7a,7'a), were isolated from the aerial parts of Ligusticum chuanxiong, together with three pairs of enantiomeric phthalide dimers [(-)/(+)-4a/4b, 5a/5b, and 6a/6b]. The bioassays revealed that compounds 1, 3, 4, 5, and 6 showed significant vasodilation effects, and the mechanism may be attributed to Cav1.2 activation blockade. Based on the established compounds library, the structure activity relationship of the phthalides was proposed. Our findings afford possible leads for developing new vasodilator against cardiovascular and cerebrovascular diseases such as hypertension and ischemic stroke.


Subject(s)
Benzofurans/pharmacology , Heterocyclic Compounds, Bridged-Ring/pharmacology , Ligusticum/chemistry , Vasodilator Agents/pharmacology , Animals , Benzofurans/chemistry , Benzofurans/isolation & purification , Benzofurans/metabolism , Calcium Channel Blockers/chemistry , Calcium Channel Blockers/isolation & purification , Calcium Channel Blockers/metabolism , Calcium Channel Blockers/pharmacology , Calcium Channels, L-Type/metabolism , HEK293 Cells , Heterocyclic Compounds, Bridged-Ring/chemical synthesis , Heterocyclic Compounds, Bridged-Ring/isolation & purification , Heterocyclic Compounds, Bridged-Ring/metabolism , Humans , Molecular Docking Simulation , Molecular Structure , Plant Components, Aerial/chemistry , Protein Binding , Rabbits , Rats, Sprague-Dawley , Stereoisomerism , Structure-Activity Relationship , Vasodilator Agents/chemistry , Vasodilator Agents/isolation & purification , Vasodilator Agents/metabolism
4.
Neurochem Int ; 143: 104899, 2021 02.
Article in English | MEDLINE | ID: mdl-33181239

ABSTRACT

Silexan®, a proprietary essential oil manufactured by steam distillation from Lavandula angustifolia flowers showed pronounced anxiolytic effects in patients with subthreshold anxiety disorders and was also efficacious in patients with Generalized Anxiety disorder (GAD). Moreover, evidences for antidepressant-like properties of Silexan® have been observed in anxious patients suffering from comorbid depressive symptoms and in patients with mixed anxiety-depression disorder (ICD-10 F41.2). In accordance with the clinical data Silexan® is active in several behavioral models in rodents at rather low concentrations indicating potent anxiolytic and antidepressive properties. As possible mechanism of action a moderate inhibition of voltage dependent calcium channels (VDCC) has been found showing some similarities to the anxiolytic drug pregabalin. However, while pregabalin mainly inhibits P/Q-type channels by binding to a modulatory subunit, Silexan® moderately inhibits mainly T-type and N-type channels and to some extent P/Q-type channels. Unlike pregabalin Silexan® is free of hypnotic or sedative side effects and seems to be devoid of any abuse potential. With respect to its specific antidepressant like properties Silexan® improves several aspects of neuroplasticity which seems to be the common final pathway of all antidepressant drugs. As a potential mechanism of its effects on neuroplasticity an activation of the transcription factor CREB via activation of intracellular signaling kinases like PKA and MAPK has been found. Since the concentrations of Silexan® needed to inhibit VDCC function and to improve neuroplasticity are quite similar, the effects of Silexan® on PKA or MAPK could constitute a common intracellular signaling cascade leading to VDCC modulation as well as CREB activation and improved neuroplasticity.


Subject(s)
Anti-Anxiety Agents/pharmacology , Antidepressive Agents/pharmacology , Calcium Channel Blockers/pharmacology , Lavandula , Oils, Volatile/pharmacology , Plant Oils/pharmacology , Animals , Anti-Anxiety Agents/isolation & purification , Anti-Anxiety Agents/therapeutic use , Antidepressive Agents/isolation & purification , Antidepressive Agents/therapeutic use , Anxiety/drug therapy , Calcium Channel Blockers/isolation & purification , Calcium Channel Blockers/therapeutic use , Calcium Channels/physiology , Depression/drug therapy , Flowers , Humans , Oils, Volatile/isolation & purification , Oils, Volatile/therapeutic use , Plant Oils/isolation & purification , Plant Oils/therapeutic use
5.
J Ethnopharmacol ; 266: 113399, 2021 Feb 10.
Article in English | MEDLINE | ID: mdl-32961278

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE: Tagetes lucida Cav. commonly known as "yauhtli" or "pericón" is used in Mexican traditional medicine for the treatment of anxiety, depressant diseases, pain, hypertension, among others. AIM: To evaluate the antihypertensive and vasorelaxant modes of action of a crude ethanolic extract from T. lucida aerial parts and to isolate the bioactive compounds. MATERIALS AND METHODS: Ethanolic extract was tested in an in vivo assay in SHR rats by intragastric administration at 10 and 100 mg/kg dosages, to measure and to compare hemodynamic parameters like diastolic and systolic blood pressure and heart rate. Also, extract (3.03-1000 µg/ml), fractions (3.03-1000 µg/ml) and pure isolated compounds (1.75-550 µM) were evaluated on isolated aortic rings contracted with noradrenaline (0.1 µM) to determine their vasorelaxant effect and extract-mode of action. RESULTS: Ethanolic extract of T. lucida lowered systolic and diastolic blood pressure on SHR rats without heart rate modification (P > 0.05). Moreover, the extract showed concentration-dependent relaxant effect in a partially endothelium-dependent manner (P < 0.05), through NO/cGMP system activation and calcium channel blockade. 6,7,8-trimethoxycoumarin (1), 6,7-dimethoxycoumarin (2), and 7-methoxycoumarin (3) from T. lucida are the main bioactive compounds of the extract and showed significant vasorelaxant activity. CONCLUSIONS: Results provide evidence and endorsed the antihypertensive properties attributed to T. lucida in traditional medicine, which is produced by vasorelaxant effect mainly through multitarget NO/cGMP system activation and calcium channel blockade. Coumarin derivatives 1, 2 and 3 are the responsible of the vasorelaxant activity.


Subject(s)
Antihypertensive Agents/pharmacology , Plant Extracts/pharmacology , Tagetes/chemistry , Vasodilator Agents/pharmacology , Animals , Antihypertensive Agents/administration & dosage , Antihypertensive Agents/isolation & purification , Blood Pressure/drug effects , Calcium Channel Blockers/administration & dosage , Calcium Channel Blockers/isolation & purification , Calcium Channel Blockers/pharmacology , Cyclic GMP/metabolism , Dose-Response Relationship, Drug , Hypertension/drug therapy , Male , Medicine, Traditional , Nitric Oxide/metabolism , Plant Components, Aerial , Plant Extracts/administration & dosage , Rats , Rats, Inbred SHR , Rats, Wistar , Vasodilator Agents/administration & dosage , Vasodilator Agents/isolation & purification
6.
Article in English | MEDLINE | ID: mdl-32901591

ABSTRACT

Aims The aim of the study was to investigate the effect of aqueous aerial part extract of Mentha pulegium L. (Pennyrile) (MPAE) on arterial pressure parameters in rats. BACKGROUND: Mentha pulegium is a medicinal plant used to treat hypertension in the Moroccon population. METHODS: In the current study, MPAE was prepared and its antihypertensive activity was pharmacologically investigated. L-NAME-hypertensive and normotensive rats received MPAE (180 and 300 mg/kg) orally for six hours for acute experiment and during seven days for the sub-chronic treatment. Thereafter, systolic, diastolic, mean arterial blood pressure and heart rate were evaluated. In the in vitro experiment, isolated denuded and intact thoracic aortic rings were suspended in a tissue bath system and the tension changes were recorded. RESULTS: A fall in blood pressure was observed in L-NAME-induced hypertensive treated with MPAE. The extract also produced a dose-dependent relaxation of aorta pre-contracted with NE and KCl. The study showed that the vasorelaxant ability of MPAE seems to be exerted through the blockage of extracellular Ca2+ entry. CONCLUSION: The results demonstrate that the extract of pennyrile exhibits antihypertensive activity. In addition, the effect may be, at least in part, due to the dilation of blood vessels via blockage of Ca2+ channels.


Subject(s)
Antihypertensive Agents/pharmacology , Aorta, Thoracic/drug effects , Arterial Pressure/drug effects , Calcium Channel Blockers/pharmacology , Hypertension/drug therapy , Mentha pulegium , Plant Extracts/pharmacology , Vasodilation/drug effects , Vasodilator Agents/pharmacology , Animals , Antihypertensive Agents/isolation & purification , Aorta, Thoracic/physiopathology , Calcium Channel Blockers/isolation & purification , Calcium Signaling/drug effects , Disease Models, Animal , Hypertension/chemically induced , Hypertension/physiopathology , Male , Mentha pulegium/chemistry , NG-Nitroarginine Methyl Ester , Plant Extracts/isolation & purification , Rats, Wistar , Vasodilator Agents/isolation & purification
7.
Toxins (Basel) ; 12(6)2020 06 15.
Article in English | MEDLINE | ID: mdl-32549266

ABSTRACT

The biological activity of Rhinella icterica parotoid secretion (RIPS) and some of its chromatographic fractions (RI18, RI19, RI23, and RI24) was evaluated in the current study. Mass spectrometry of these fractions indicated the presence of sarmentogenin, argentinogenin, (5ß,12ß)-12,14-dihydroxy-11-oxobufa-3,20,22-trienolide, marinobufagin, bufogenin B, 11α,19-dihydroxy-telocinobufagin, bufotalin, monohydroxylbufotalin, 19-oxo-cinobufagin, 3α,12ß,25,26-tetrahydroxy-7-oxo-5ß-cholestane-26-O-sulfate, and cinobufagin-3-hemisuberate that were identified as alkaloid and steroid compounds, in addition to marinoic acid and N-methyl-5-hydroxy-tryptamine. In chick brain slices, all fractions caused a slight decrease in cell viability, as also seen with the highest concentration of RIPS tested. In chick biventer cervicis neuromuscular preparations, RIPS and all four fractions significantly inhibited junctional acetylcholinesterase (AChE) activity. In this preparation, only fraction RI23 completely mimicked the pharmacological profile of RIPS, which included a transient facilitation in the amplitude of muscle twitches followed by progressive and complete neuromuscular blockade. Mass spectrometric analysis showed that RI23 consisted predominantly of bufogenins, a class of steroidal compounds known for their cardiotonic activity mediated by a digoxin- or ouabain-like action and the blockade of voltage-dependent L-type calcium channels. These findings indicate that the pharmacological activities of RI23 (and RIPS) are probably mediated by: (1) inhibition of AChE activity that increases the junctional content of Ach; (2) inhibition of neuronal Na+/K+-ATPase, leading to facilitation followed by neuromuscular blockade; and (3) blockade of voltage-dependent Ca2+ channels, leading to stabilization of the motor endplate membrane.


Subject(s)
Bufanolides/toxicity , Bufonidae , Neurotoxins/toxicity , Parotid Gland/chemistry , Animals , Brain/drug effects , Brain/metabolism , Brain/pathology , Bufanolides/isolation & purification , Calcium Channel Blockers/isolation & purification , Calcium Channel Blockers/toxicity , Cell Survival/drug effects , Chickens , Cholinesterase Inhibitors/isolation & purification , Cholinesterase Inhibitors/toxicity , Dose-Response Relationship, Drug , Neuromuscular Junction/drug effects , Neuromuscular Junction/metabolism , Neurotoxins/isolation & purification , Secretory Pathway , Sodium-Potassium-Exchanging ATPase/antagonists & inhibitors , Sodium-Potassium-Exchanging ATPase/metabolism
8.
J Ethnopharmacol ; 253: 112643, 2020 May 10.
Article in English | MEDLINE | ID: mdl-32035218

ABSTRACT

ETHNOPHARMACOLOGICAL IMPORTANCE: Achillea millefolium L. (Asteraceae) is used for the treatment of respiratory diseases, diabetes, and hypertension. AIM: to explore its tracheal relaxant properties and clarify its functional mechanism of action on smooth muscle cells, which allow us to propose it as a potential anti-asthmatic drug. MATERIAL AND METHODS: organic and hydro-alcoholic extracts from A. millefolium were obtained by macerations, then their relaxing effect on ex vivo isolated rat trachea rings was determined. Most active extract (hexanic extract, EHAm) was studied to determine its functional mechanism of action using synergic, antagonist and inhibitor agents related with the contraction/relaxation process of the smooth muscle. Also, EHAm was subjected to bio-guided fractionation by open-column chromatography (on silica gel) using cyclohexane-EtOAc (80:20) in an isocratic way to isolate main bioactive compounds. RESULTS: organic and hydro-alcoholic extracts showed relaxant effect in a concentration-response dependent manner, being EHAm the most active. The functional mechanism of action indicates that EHAm induced a non-competitive antagonism to the muscarinic receptors ; in addition, the NO/cGMP pathway is involved in the relaxation process of the tracheal smooth muscle. However, the most important mechanism of action showed by EHAm was related with the calcium channel blockade influx into the smooth muscle cells. On the other hand, epimeric sesquiterpene lactones leucodin (1) and achillin (2) were isolated and purified, which are responsible for the observed smooth muscle relaxant activity of the extract. CONCLUSION: hexanic extract of A. millefollium induced a significant relaxant effect on tracheal rat rings by calcium channel blockade and NO release.


Subject(s)
Achillea/chemistry , Calcium Channel Blockers/pharmacology , Muscle Relaxation/drug effects , Plant Extracts/pharmacology , Trachea/drug effects , Animals , Anti-Asthmatic Agents/administration & dosage , Anti-Asthmatic Agents/isolation & purification , Anti-Asthmatic Agents/pharmacology , Calcium Channel Blockers/administration & dosage , Calcium Channel Blockers/isolation & purification , Dose-Response Relationship, Drug , Male , Muscle, Smooth/drug effects , Muscle, Smooth/metabolism , Myocytes, Smooth Muscle/drug effects , Myocytes, Smooth Muscle/metabolism , Plant Extracts/administration & dosage , Rats , Rats, Wistar , Trachea/metabolism
9.
J Ethnopharmacol ; 248: 112321, 2020 Feb 10.
Article in English | MEDLINE | ID: mdl-31655146

ABSTRACT

ETHNOPHARMACOLOGY RELEVANCE: In traditional Mexican medicine, Echeveria gibbiflora DC has been used as a vaginal post-coital rinse to prevent pregnancy. The aqueous crude extract (OBACE) induces sperm immobilization/agglutination and a hypotonic-like effect, likely attributed to the high concentration of calcium bis-(hydrogen-1-malate) hexahydrate [Ca2+ (C4H5O5)2•6H2O]. Likewise, OBACE impedes the increase of [Ca2+]i during capacitation. AIM OF THE STUDY: Evaluate the effect of OBACE on sperm energy metabolism and the underlying mechanism of action on sperm-specific channel. MATERIAL AND METHODS: In vitro, we quantified the mouse sperm immobilization effect and the antifertility potential of OBACE. The energetic metabolism status was also evaluated by assessing the ATP levels, general mitochondrial activity, mitochondrial membrane potential, and enzymatic activity of three key enzymes of energy metabolism. Furthermore, the effect of the ion efflux of Cl- and K+, as well as the pHi, were investigated in order to elucidate which channel is suitable to perform an in silico study. RESULTS: Total and progressive motility notably decreased, as did fertility rates. ATP levels, mitochondrial activity and membrane potential were reduced. Furthermore, the activities of the three enzymes decreased. Neither Cl- or K+ channels activities were affected at low concentrations of OBACE; nevertheless, pHi did not alkalinize. Finally, an in silico analysis was performed between the Catsper channel and calcium bis-(hydrogen-1-malate) hexahydrate, which showed a possible blockade of this sperm cation channel. CONCLUSION: The results were useful to elucidate the effect of OBACE and to propose it as a future male contraceptive.


Subject(s)
Calcium Channel Blockers/pharmacology , Calcium Channels/drug effects , Calcium Signaling/drug effects , Contraceptive Agents, Male/pharmacology , Crassulaceae , Energy Metabolism/drug effects , Plant Extracts/pharmacology , Spermatozoa/drug effects , Animals , Binding Sites , Calcium Channel Blockers/chemistry , Calcium Channel Blockers/isolation & purification , Calcium Channels/chemistry , Calcium Channels/metabolism , Contraceptive Agents, Male/chemistry , Contraceptive Agents, Male/isolation & purification , Crassulaceae/chemistry , Fertility/drug effects , Hydrogen-Ion Concentration , Male , Mice , Mitochondria/drug effects , Mitochondria/metabolism , Molecular Docking Simulation , Plant Extracts/isolation & purification , Protein Conformation , Sperm Motility/drug effects , Spermatozoa/metabolism , Structure-Activity Relationship
10.
Int J Oncol ; 55(5): 1090-1096, 2019 Nov.
Article in English | MEDLINE | ID: mdl-31545410

ABSTRACT

Uveal malignant melanoma (UMM), the most common primary adult intraocular tumor with a marked metastatic potential, is genetically unique and has unfortunately had few treatment breakthroughs. In this study, we subjected a UMM cell line to high­throughput library screening with 1,018 FDA­approved compounds to identify potential UMM­selective cytotoxic agents. Amlodipine, a dihydropyridine calcium channel blocker (CCB), ranked no. 2 and no. 8 of the most cytotoxic compounds. Thus, we further characterized the differential effects of calcium blockade on UMM and cutaneous malignant melanoma (CMM) lines in vitro using growth inhibition, cell cycle progression, apoptosis and senescence assays. Amlodipine had a significantly higher growth inhibitory potency in UMM (IC50=13.1 µM) than CMM (IC50=15.9 µM, P<0.05) lines. In 3D spherical cell culture, amlodipine treatment significantly impaired tissue volume growth in two UMM lines, but exerted no significant effects among all 3 CMM lines tested. Treatment with 10 and 20 µM amlodipine induced a significant impairment of cell cycle progression and the apoptosis of UMM lines, implicating both of these processes as mediators of the observed growth inhibition in UMM compared to CMM. On the whole, the findings of this study suggest that calcium channel blockade is a potentially effective strategy for selective uveal melanoma targeting.


Subject(s)
Calcium Channel Blockers/pharmacology , Calcium Channels/chemistry , Calcium/metabolism , Melanoma/drug therapy , Uveal Neoplasms/drug therapy , Apoptosis , Calcium Channel Blockers/isolation & purification , Cell Cycle , Cell Movement , Cell Proliferation , High-Throughput Screening Assays , Humans , Melanoma/metabolism , Melanoma/pathology , Tumor Cells, Cultured , Uveal Neoplasms/metabolism , Uveal Neoplasms/pathology
11.
Elife ; 82019 08 12.
Article in English | MEDLINE | ID: mdl-31403402

ABSTRACT

Inhibiting high-voltage-activated calcium channels (HVACCs; CaV1/CaV2) is therapeutic for myriad cardiovascular and neurological diseases. For particular applications, genetically-encoded HVACC blockers may enable channel inhibition with greater tissue-specificity and versatility than is achievable with small molecules. Here, we engineered a genetically-encoded HVACC inhibitor by first isolating an immunized llama nanobody (nb.F3) that binds auxiliary HVACC CaVß subunits. Nb.F3 by itself is functionally inert, providing a convenient vehicle to target active moieties to CaVß-associated channels. Nb.F3 fused to the catalytic HECT domain of Nedd4L (CaV-aßlator), an E3 ubiquitin ligase, ablated currents from diverse HVACCs reconstituted in HEK293 cells, and from endogenous CaV1/CaV2 channels in mammalian cardiomyocytes, dorsal root ganglion neurons, and pancreatic ß cells. In cardiomyocytes, CaV-aßlator redistributed CaV1.2 channels from dyads to Rab-7-positive late endosomes. This work introduces CaV-aßlator as a potent genetically-encoded HVACC inhibitor, and describes a general approach that can be broadly adapted to generate versatile modulators for macro-molecular membrane protein complexes.


Subject(s)
Biological Products/pharmacology , Calcium Channel Blockers/pharmacology , Calcium Channels/metabolism , Single-Domain Antibodies/pharmacology , Animals , Biological Products/isolation & purification , Calcium Channel Blockers/isolation & purification , Camelids, New World , HEK293 Cells , Humans , Protein Binding
12.
Toxins (Basel) ; 11(9)2019 08 27.
Article in English | MEDLINE | ID: mdl-31461913

ABSTRACT

Pamphobeteus verdolaga is a recently described Theraphosidae spider from the Andean region of Colombia. Previous reports partially characterized its venom profile. In this study, we conducted a detailed analysis that includes reversed-phase high-performance liquid chromatography (rp-HPLC), calcium influx assays, tandem mass spectrometry analysis (tMS/MS), and venom-gland transcriptome. rp-HPLC fractions of P. verdolaga venom showed activity on CaV2.2, CaV3.2, and NaV1.7 ion channels. Active fractions contained several peptides with molecular masses ranging from 3399.4 to 3839.6 Da. The tMS/MS analysis of active fraction displaying the strongest activity to inhibit calcium channels showed sequence fragments similar to one of the translated transcripts detected in the venom-gland transcriptome. The putative peptide of this translated transcript corresponded to a toxin, here named ω-theraphositoxin-Pv3a, a potential ion channel modulator toxin that is, in addition, very similar to other theraphositoxins affecting calcium channels (i.e., ω-theraphotoxin-Asp1a). Additionally, using this holistic approach, we found that P. verdolaga venom is an important source of disulfide-rich proteins expressing at least eight superfamilies.


Subject(s)
Calcium Channel Blockers/pharmacology , Disulfides/pharmacology , Peptides/pharmacology , Spider Venoms/chemistry , Spiders , Transcriptome/genetics , Amino Acid Sequence , Animals , Calcium Channel Blockers/isolation & purification , Calcium Channels/metabolism , Cell Line, Tumor , Disulfides/isolation & purification , Female , Humans , Molecular Sequence Annotation , Peptides/genetics , Peptides/isolation & purification , Sequence Alignment , Spider Venoms/genetics , Spiders/genetics
13.
Phytochem Anal ; 30(4): 456-463, 2019 Jul.
Article in English | MEDLINE | ID: mdl-30859657

ABSTRACT

INTRODUCTION: Calcium channel blockers (CCBs) are currently the most commonly used drugs for the treatment of hypertension. Moutan Cortex (MC), a traditional Chinese herb, has been found to have an anti-hypertensive effect. However, its potential mechanisms in the regulation of intracellular calcium concentration ([Ca2+ ]i ) remain poorly understood. OBJECTIVE: The main objective of this work was to identify the potential calcium antagonists from MC and study their molecular mechanisms. METHODS: Ultra-high performance liquid chromatography-quadrupole-time-of-fight-mass spectrometry (UHPLC-QTOF-MS) analysis combined with a dual-luciferase reporter assay was utilised to systematically screen the calcium antagonistic active ingredients in the methanol extract of MC. Additionally, the molecular mechanism of these compounds was further studied using live-cell imaging analysis with the calcium ion (Ca2+ ) probe dye fluo-4/AM to monitor changes in [Ca2+ ]i . RESULTS: Three monoterpenoids (paeoniflorin, benzoylpaeoniflorin and mudanpioside C), one phenolic acid (paeonol) and one gallotannin (1,2,3,4,6-O-pentagalloylglucose) were screened out as potential calcium antagonists in MC. Among them, the calcium antagonistic activity of benzoylpaeoniflorin, mudanpioside C and 1,2,3,4,6-O-pentagalloylglucose is first reported. Additionally, paeoniflorin, benzoylpaeoniflorin, mudanpioside C and paeonol can effectively block voltage-operated Ca2+ channels (VOCCs) to exert calcium antagonism, while 1,2,3,4,6-O-pentagalloylglucose plays a role in blocking inositol 1,4,5-trisphosphate receptors (IP3Rs). CONCLUSION: This work indicated that the anti-hypertensive efficacy of MC acted through multiple components selectively antagonising multiple cell signalling pathways to regulate [Ca2+ ]i . Furthermore, they could be considered as a reference standard for controlling the quality of Chinese medicinal materials.


Subject(s)
Calcium Channel Blockers/isolation & purification , Calcium/metabolism , Drugs, Chinese Herbal/chemistry , Hypertension/drug therapy , Monoterpenes/chemistry , Paeonia/chemistry , Phytochemicals/chemistry , Plant Extracts/chemistry , Animals , Calcium Channel Blockers/chemistry , Cell Line , Chromatography, High Pressure Liquid , Genes, Reporter , Humans , Inositol 1,4,5-Trisphosphate Receptors/metabolism , Medicine, Chinese Traditional , Monoterpenes/isolation & purification , Monoterpenes/pharmacology , Phytochemicals/isolation & purification , Phytochemicals/pharmacology , Plant Extracts/isolation & purification , Plant Extracts/pharmacology , Rats , Tandem Mass Spectrometry
14.
Org Lett ; 21(4): 1197-1201, 2019 02 15.
Article in English | MEDLINE | ID: mdl-30702896

ABSTRACT

Two novel sesquiterpenoids, curcumanes A (1) and B (2), possessing unprecedented skeletons with a dicyclo[3.2.1]octane and a dicyclo[3.3.1]nonane moiety, respectively, were isolated from Curcuma longa. Both of them had remarkable vasorelaxant activity on rat aorta via blocking extracellular Ca2+ influx through VDCCs and ROCCs. The activity of 1 was endothelium-independent, while that of 2 was endothelium-dependent. Compound 2 also prolonged APTT and TT to inhibit blood coagulation.


Subject(s)
Aorta, Thoracic/drug effects , Calcium Channel Blockers/pharmacology , Calcium Channels/metabolism , Curcuma/chemistry , Polycyclic Sesquiterpenes/pharmacology , Receptors, Calcium-Sensing/antagonists & inhibitors , Vasodilator Agents/pharmacology , Animals , Calcium Channel Blockers/chemistry , Calcium Channel Blockers/isolation & purification , Polycyclic Sesquiterpenes/chemistry , Polycyclic Sesquiterpenes/isolation & purification , Rats , Receptors, Calcium-Sensing/metabolism , Vasodilator Agents/chemistry , Vasodilator Agents/isolation & purification
15.
J Asian Nat Prod Res ; 21(1): 17-24, 2019 Jan.
Article in English | MEDLINE | ID: mdl-29373928

ABSTRACT

Three new Lycopodium alkaloids (1-3), together with 15 known alkaloids, were isolated from club moss Lycopodium japonicum. Their structures were determined by extensive spectroscopic analysis, including 1D and 2D NMR spectra. Compound 1 has an unusual ß-oriented methyl group substituted at C-15 and an α-hydroxy cyclopentenone moiety. All new alkaloids were evaluated for the inhibition of T-type calcium channel.


Subject(s)
Alkaloids/isolation & purification , Lycopodium/chemistry , Alkaloids/chemistry , Alkaloids/pharmacology , Calcium Channel Blockers/isolation & purification , Calcium Channels, T-Type/drug effects , HEK293 Cells , Humans , Magnetic Resonance Spectroscopy
16.
Pak J Pharm Sci ; 32(6(Supplementary)): 2893-2898, 2019 Nov.
Article in English | MEDLINE | ID: mdl-32024630

ABSTRACT

Hypertension is persistent elevation in blood pressure for 3-4 weeks. Estimated global prevalence of hypertension suggested that by the Year 2025 (29%) of adult worldwide are suffering from hypertension (1.56 billion). Hypertension complications are hemorrhage, atherosclerosis, renal artery stenosis, angina pectoris end organ damage, cardiomyopathy, myocardial infarction and retinopathy. Along with other drug class Calcium channel blocker are also used for the treatment of hypertension. In this study the possible action of the n-hexane leaves fraction of the Androsace foliosa on isolated rabbit aorta was examined. Antihypertensive activity was examined in the existence of standard agonist like phenylephrine and antagonist like Verapamil. Phenylephrine (PE 1µM) high K+ was used to steady the tissue materials. Additionally to observe the calcium channel blocking effect the tissues were treated with n-hexane segment of A. foliosa leaves. Aortic tissues were treated 4-5intervals with Ca+2- free preparation earlier to control calcium reaction curve (CRCs). Verapamil is utilized as standard calcium channel inhibitory mediator and is used as an antagonist. The Af. n-hexane leaves fraction completely inhibited the precontractions induced by Phenylephrine (1µM) and K+ (80 mM) precontractions, with EC50 standards of 1.0mM (0.3-1.0mg/mL) and 4.90mM (1-3mg/mL), respectively. Androsace foliosa n-hexane leaves fraction was tested for calcium channel inhibitory effect on isolated rabbit aorta. A. foliosa n- hexane leaves segment at the dosage of 1mg/mL block the calcium channel approximately (35±5%). Consequence indicates that A. foliosa n-hexane leaves segment block calcium channel in the similar manner as compared to the standard calcium channel blocker drug (verapamil).


Subject(s)
Antihypertensive Agents/pharmacology , Aorta, Thoracic/drug effects , Calcium Channel Blockers/pharmacology , Hypertension/drug therapy , Plant Extracts/pharmacology , Primulaceae/chemistry , Animals , Antihypertensive Agents/isolation & purification , Aorta, Thoracic/metabolism , Aorta, Thoracic/physiopathology , Blood Pressure/drug effects , Calcium Channel Blockers/isolation & purification , Calcium Channels/metabolism , Female , Hexanes/chemistry , In Vitro Techniques , Plant Extracts/isolation & purification , Plant Leaves/chemistry , Rabbits , Vasodilation/drug effects
17.
Neuropharmacology ; 138: 232-244, 2018 08.
Article in English | MEDLINE | ID: mdl-29913186

ABSTRACT

Since Cav3.2 T-type Ca2+ channels (T-channels) expressed in the primary afferents and CNS contribute to intractable pain, we explored T-channel-blocking components in distinct herbal extracts using a whole-cell patch-clamp technique in HEK293 cells stably expressing Cav3.2 or Cav3.1, and purified and identified sophoraflavanone G (SG) as an active compound from SOPHORAE RADIX (SR). Interestingly, hop-derived SG analogues, (2S)-6-prenylnaringenin (6-PNG) and (2S)-8-PNG, but not naringenin, also blocked T-channels; IC50 (µM) of SG, (2S)-6-PNG and (2S)-8-PNG was 0.68-0.75 for Cav3.2 and 0.99-1.41 for Cav3.1. (2S)-6-PNG and (2S)-8-PNG, but not SG, exhibited reversible inhibition. The racemic (2R/S)-6-PNG as well as (2S)-6-PNG potently blocked Cav3.2, but exhibited minor effect on high-voltage-activated Ca2+ channels and voltage-gated Na+ channels in differentiated NG108-15 cells. In mice, the mechanical allodynia following intraplantar (i.pl.) administration of an H2S donor was abolished by oral or i.p. SR extract and by i.pl. SG, (2S)-6-PNG or (2S)-8-PNG, but not naringenin. Intraperitoneal (2R/S)-6-PNG strongly suppressed visceral pain and spinal ERK phosphorylation following intracolonic administration of an H2S donor in mice. (2R/S)-6-PNG, administered i.pl. or i.p., suppressed the neuropathic allodynia induced by partial sciatic nerve ligation or oxaliplatin, an anti-cancer agent, in mice. (2R/S)-6-PNG had little or no effect on open-field behavior, motor performance or cardiovascular function in mice, and on the contractility of isolated rat aorta. (2R/S)-6-PNG, but not SG, was detectable in the brain after their i.p. administration in mice. Our data suggest that 6-PNG, a hop component, blocks T-channels, and alleviates neuropathic and visceral pain with little side effects.


Subject(s)
Analgesics, Non-Narcotic/pharmacology , Calcium Channel Blockers/pharmacology , Flavonoids/pharmacology , Neuralgia/drug therapy , Visceral Pain/drug therapy , Analgesics, Non-Narcotic/chemistry , Analgesics, Non-Narcotic/isolation & purification , Animals , Calcium Channel Blockers/chemistry , Calcium Channel Blockers/isolation & purification , Calcium Channels, T-Type/genetics , Calcium Channels, T-Type/metabolism , Disease Models, Animal , Flavonoids/chemistry , Flavonoids/isolation & purification , HEK293 Cells , Humans , Humulus , Hyperalgesia/drug therapy , Hyperalgesia/metabolism , Male , Mice, Inbred C57BL , Mice, Transgenic , Neuralgia/metabolism , Phytochemicals/chemistry , Phytochemicals/isolation & purification , Phytochemicals/pharmacology , Plant Extracts/chemistry , Plant Extracts/pharmacology , Random Allocation , Rats, Wistar , Visceral Pain/metabolism
18.
Pak J Pharm Sci ; 31(1): 95-102, 2018 Jan.
Article in English | MEDLINE | ID: mdl-29348090

ABSTRACT

In vivo and in vitro research study was conducted on Cyperus rotundus to evaluate the sound mechanistic background in the treatment of gastrointestinal, bronchial and vascular disorders as well as in pain, emesis, pyrexia and bacterial infections. Results showed that crude extract of Cyperus rotundus (Cr.Cr) exhibited the dose-dependent spasmolytic effect in rabbit jejunum by inhibiting the spontaneous and K+ (80 mM)-induced contractions. Pretreatment of tissue with Cr. Cr caused the rightward shift of calcium concentration response curves, similar to verapamil. Cr. Cr also caused the relaxation of K+(80 mM)- and carbachol (1 µM)-induced contractions of trachea preparations, similar to that of verapamil. Moreover, Cr. Cr also relaxed the contraction induced by the K+ (80 mM) and phenylephrine (1 µM) of aorta preparations. Data show that C. rotundus possess the spasmolytic, bronchodilator and vasodilator activities possibly through calcium channels blockade; validating its folkloric use in diarrhea, dyspepsia, bronchitis, asthma and hypertension in addition to antibacterial, antiemetic, antipyretic and analgesic activities.


Subject(s)
Bronchodilator Agents/pharmacology , Calcium Channel Blockers/pharmacology , Cyperus/chemistry , Medicine, Traditional/methods , Plant Extracts/pharmacology , Vasodilator Agents/pharmacology , Animals , Bronchodilator Agents/isolation & purification , Calcium Channel Blockers/isolation & purification , Chickens , Female , In Vitro Techniques , Male , Mice , Plant Extracts/isolation & purification , Rabbits , Vasodilator Agents/isolation & purification
19.
Pharm Biol ; 56(1): 559-566, 2018 Dec.
Article in English | MEDLINE | ID: mdl-31070538

ABSTRACT

CONTEXT: Aquilariae Lignum Resinatum (ALR), the dry rhizome of Aquilaria agallocha R. (Thymelaeaeeae), has been widely used to treat emesis, stomachache and gastrointestinal dysfunction. OBJECTIVE: This study evaluates the effects of ALR methanol extract on gastrointestinal motility (GIM) and possible mechanisms of the action involved. MATERIALS AND METHODS: In vivo, the study evaluated the effects of ALR (200-800 mg/kg) on gastric emptying and small intestinal motility in normal and neostigmine-induced adult KM mice. The in vitro effects of ALR (0.2-1.6 mg/mL) on GIM were performed on isolated jejunum of Wistar rats, pretreated with acetylcholine (ACh), KCl, CaCl2, and pre-incubation with l-NAME (a selective inhibitor of the nitric oxide synthase). RESULTS: In vivo, ALR (800 mg/kg) decreased gastric emptying (70.82 ± 9.81%, p < 0.01, compared with neostigmine group 91.40 ± 7.81%), small intestinal transit (42.82 ± 3.82%, p < 0.01, compared with neostigmine group 85.53 ± 5.57%). In vitro, ALR concentration dependently decreased the contractions induced by ACh (10-5 M) and KCl (60 mM) with respective EC50 values of 0.35 and 0.32 mg/mL. The Ca2+ concentration-response curves were shifted by ALR to the right, similar to that caused by verapamil (the positive). The spasmolytic activity of ALR was inhibited by pre-incubation with l-NAME. DISCUSSION AND CONCLUSIONS: ALR played a spasmolytic role in GIM, which is probably mediated through inhibition of muscarinic receptors, blockade of Ca2+ influx and NO release. This is the first study presenting a comprehensive description of the effects of ALR on GIM.


Subject(s)
Calcium Channel Blockers/pharmacology , Gastrointestinal Motility/drug effects , Muscarinic Antagonists/pharmacology , Nitric Oxide/antagonists & inhibitors , Parasympatholytics/pharmacology , Plant Extracts/pharmacology , Thymelaeaceae , Animals , Calcium Channel Blockers/isolation & purification , Calcium Channels/physiology , Dose-Response Relationship, Drug , Gastric Emptying/drug effects , Gastric Emptying/physiology , Gastrointestinal Motility/physiology , Mice , Muscarinic Antagonists/isolation & purification , Nitric Oxide/physiology , Organ Culture Techniques , Parasympatholytics/isolation & purification , Plant Extracts/isolation & purification , Random Allocation , Rats , Rats, Wistar , Receptors, Muscarinic/physiology
20.
J Ethnopharmacol ; 220: 188-196, 2018 Jun 28.
Article in English | MEDLINE | ID: mdl-28965754

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE: The aerial parts of Pogostemon cablin (Blanco) Benth. for the treatment of cardiodynia have been documented in Mingyi Bielu of late Han Dynasty, in addition to that the Ca2+ antagonized activities of P. cablin and its critically pharmacological ingredient patchouli alcohol (PA) were reported previously. AIM OF THE STUDY: To investigate the relaxant effects of PA on rat isolated thoracic aortas and further explore its potential mechanisms of actions. MATERIALS AND METHODS: The aortas with endothelium and without endothelium were prepared and suspended in the organ bath for isometric tension recordings. The responses to accumulative concentrations of PA in endothelium-intact (E + ) aortas with basal tension and in different treated aortas pre-contracted with potassium chloride (KCl) or phenylephrine (PHE) were observed; the effects of L-NAME and indomethacin in aortas with intact endothelium, and of L-NAME, propranolol, tetraethtylamine (TEA), 4-aminopyridine (4-AP), barium chloride (BaCl2), glyburide in aortas with endothelial denudation on PA-produced vasorelaxation were assessed; the influences of PA on extracellular Ca2+ influx and intracellular Ca2+ release were measured in Ca2+-free medium. Finally, the abilities of PA to inhibit KCl- and PHE-induced contractions were compared to that of verapamil in E- aortas. RESULTS: PA produced vasorelaxant effects in KCl- and PHE-precontracted E + aortas in a concentration-dependent manner, which had no statistically different from that in KCl- and PHE-precontracted E- aortas. PA (10 µM) significantly reduced KCl-induced contractions while PHE-induced contractions were significantly reduced by 100 µM of PA instead of 10 µM and 30 µM in aortas with endothelium. Pre-incubation of E + aortas with L-NAME as well as indomethacin and of E- aortas with L-NAME, propranolol, TEA, 4-AP, BaCl2 and glyburide had no obvious effects on vasorelaxation of PA. In endothelium-removed aortas around Ca2+-free solution, PA remarkably lowered the contractions stimulated with Ca2+ and PHE, and application of ruthenium red and heparin further enhanced the abilities of PA to reduce PHE-caused contractions. In aortas without endothelium, 100 µM of PA markedly attenuated KCl-induced contractions but not affect PHE-induced contractions. Verapamil at the equal dose markedly attenuated KCl- and PHE-induced contractions, and the inhibitory effects on KCl-induced contractions were more forceful than that on PHE-induced contractions. In combined usage, the inhibitory effects on the contractions elicited by KCl were evidently weaker than that of verapamil alone and indifferently stronger than that of PA alone, and the inhibitory effects on the contractions elicited by PHE were evidently weaker than that of single verapamil but stronger than that of single PA. CONCLUSION: PA may act as a Ca2+ antagonist to exert an intensively vasorelaxant effects through endothelium-independent pathway, and its mechanisms underlying the vasoactivities seem to be associated with the blockade of extracellular Ca2+ influx through VDCCs and ROCCs in vascular smooth muscle cells (VSMCs) membrane and intracellular Ca2+ releases through IP3R- and RYR-mediated Ca2+ channels in sarcolemma.


Subject(s)
Calcium Channel Blockers/pharmacology , Pogostemon/chemistry , Sesquiterpenes/pharmacology , Vasodilation/drug effects , Animals , Aorta, Thoracic/drug effects , Aorta, Thoracic/metabolism , Calcium/metabolism , Calcium Channel Blockers/isolation & purification , Calcium Channels/drug effects , Calcium Channels/metabolism , Endothelium, Vascular/drug effects , Endothelium, Vascular/metabolism , Male , Myocytes, Smooth Muscle/drug effects , Myocytes, Smooth Muscle/metabolism , Plant Components, Aerial , Rats , Rats, Sprague-Dawley
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