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2.
Nutrients ; 13(8)2021 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-34444651

RESUMO

Cow's milk allergy is a common food allergy in infants, and is associated with an increased risk of developing other allergic diseases. Dietary selenium (Se), one of the essential micronutrients for humans and animals, is an important bioelement which can influence both innate and adaptive immune responses. However, the effects of Se on food allergy are still largely unknown. In the current study it was investigated whether dietary Se supplementation can inhibit whey-induced food allergy in an animal research model. Three-week-old female C3H/HeOuJ mice were intragastrically sensitized with whey protein and cholera toxin and randomly assigned to receive a control, low, medium or high Se diet. Acute allergic symptoms, allergen specific immunoglobulin (Ig) E levels and mast cell degranulation were determined upon whey challenge. Body temperature was significantly higher in mice that received the medium Se diet 60 min after the oral challenge with whey compared to the positive control group, which is indicative of impaired anaphylaxis. This was accompanied by reductions in antigen-specific immunoglobulins and reduced levels of mouse mast cell protease-1 (mMCP-1). This study demonstrates that oral Se supplementation may modulate allergic responses to whey by decreasing specific antibody responses and mMCP-1 release.


Assuntos
Dieta , Hipersensibilidade a Leite/dietoterapia , Selenometionina/administração & dosagem , Proteínas do Soro do Leite/imunologia , Anafilaxia/dietoterapia , Anafilaxia/imunologia , Ração Animal , Animais , Biomarcadores/sangue , Degranulação Celular , Células Cultivadas , Quimases/sangue , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Dermatite Alérgica de Contato/dietoterapia , Dermatite Alérgica de Contato/imunologia , Modelos Animais de Doenças , Feminino , Imunoglobulina E/sangue , Mastócitos/imunologia , Mastócitos/metabolismo , Camundongos Endogâmicos C3H , Hipersensibilidade a Leite/sangue , Hipersensibilidade a Leite/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo
3.
Comput Biol Med ; 136: 104671, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34332348

RESUMO

Antiviral culinary plants are potential bioresources for preventive nutraceuticals and/or antiviral drugs in COVID-19. Structure-based virtual screening was undertaken to screen 173 compounds previously reported from Vernonia amygdalina and Occinum gratissimum for direct interaction with the active site of the 3-Chymotrypsin-Like Protease (3CLpro) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Based on docking scores and comparison with reference inhibitors, a hit-list of 10 top phytocompounds was defined, which also had strong interactions with the catalytic centre of 3CLpro from three related strains of coronavirus (SARS-CoV, MERS-CoV, HKU4). Among these, six compounds (neoandrographolide, vernolide, isorhamnetin, chicoric acid, luteolin, and myricetin) exhibited the highest binding tendencies to the equilibrated conformers of SARS-CoV-2 3CLpro in an in-depth docking analysis to 5 different representative conformations from the cluster analysis of the molecular dynamics simulation (MDS) trajectories of the protein. In silico drug-likeness analyses revealed two drug-like terpenoids viz: neoandrographolide and vernolide as promising inhibitors of SARS-CoV-2 3CLpro. These structures were accommodated within the substrate-binding pocket; and interacted with the catalytic dyad (Cys145 and His41), the oxyanion loop (residues 138-145), and the S1/S2 sub-sites of the enzyme active site through the formation of an array of hydrogen bonds and hydrophobic interactions. Molecular dynamics simulation and binding free energy calculation revealed that the terpenoid-enzyme complexes exhibit strong interactions and structural stability. Therefore, these compounds may stabilize the conformation of the flexible oxyanion loop; and thereby interfere with the tetrahedral oxyanion intermediate formation during the proteolytic activity of the enzyme.


Assuntos
Antivirais/farmacologia , Proteases 3C de Coronavírus/antagonistas & inibidores , Ocimum/química , Compostos Fitoquímicos/farmacologia , SARS-CoV-2/efeitos dos fármacos , Vernonia , COVID-19 , Quimases , Humanos , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Inibidores de Proteases , Vernonia/química
4.
J Gen Virol ; 102(7)2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34319869

RESUMO

Rapid repurposing of existing drugs as new therapeutics for COVID-19 has been an important strategy in the management of disease severity during the ongoing SARS-CoV-2 pandemic. Here, we used high-throughput docking to screen 6000 compounds within the DrugBank library for their potential to bind and inhibit the SARS-CoV-2 3 CL main protease, a chymotrypsin-like enzyme that is essential for viral replication. For 19 candidate hits, parallel in vitro fluorescence-based protease-inhibition assays and Vero-CCL81 cell-based SARS-CoV-2 replication-inhibition assays were performed. One hit, diclazuril (an investigational anti-protozoal compound), was validated as a SARS-CoV-2 3 CL main protease inhibitor in vitro (IC50 value of 29 µM) and modestly inhibited SARS-CoV-2 replication in Vero-CCL81 cells. Another hit, lenvatinib (approved for use in humans as an anti-cancer treatment), could not be validated as a SARS-CoV-2 3 CL main protease inhibitor in vitro, but serendipitously exhibited a striking functional synergy with the approved nucleoside analogue remdesivir to inhibit SARS-CoV-2 replication, albeit this was specific to Vero-CCL81 cells. Lenvatinib is a broadly-acting host receptor tyrosine kinase (RTK) inhibitor, but the synergistic effect with remdesivir was not observed with other approved RTK inhibitors (such as pazopanib or sunitinib), suggesting that the mechanism-of-action is independent of host RTKs. Furthermore, time-of-addition studies revealed that lenvatinib/remdesivir synergy probably targets SARS-CoV-2 replication subsequent to host-cell entry. Our work shows that combining computational and cellular screening is a means to identify existing drugs with repurposing potential as antiviral compounds. Future studies could be aimed at understanding and optimizing the lenvatinib/remdesivir synergistic mechanism as a therapeutic option.


Assuntos
Monofosfato de Adenosina/análogos & derivados , Alanina/análogos & derivados , COVID-19/tratamento farmacológico , COVID-19/virologia , Quimases/antagonistas & inibidores , Compostos de Fenilureia/farmacologia , Quinolinas/farmacologia , SARS-CoV-2/efeitos dos fármacos , Monofosfato de Adenosina/farmacologia , Alanina/farmacologia , Animais , Antivirais/farmacologia , COVID-19/enzimologia , Células Cultivadas , Avaliação Pré-Clínica de Medicamentos , Humanos , Simulação de Acoplamento Molecular , Inibidores de Proteínas Quinases/farmacologia , SARS-CoV-2/isolamento & purificação , SARS-CoV-2/patogenicidade
5.
Int J Mol Sci ; 22(10)2021 May 16.
Artigo em Inglês | MEDLINE | ID: mdl-34065716

RESUMO

Chronic respiratory diseases are often characterized by impaired epithelial function and remodeling. Mast cells (MCs) are known to home into the epithelium in respiratory diseases, but the MC-epithelial interactions remain less understood. Therefore, this study aimed to investigate the effect of MC proteases on bronchial epithelial morphology and function. Bronchial epithelial cells were stimulated with MC tryptase and/or chymase. Morphology and epithelial function were performed using cell tracking analysis and holographic live-cell imaging. Samples were also analyzed for motility-associated gene expression. Immunocytochemistry was performed to compare cytoskeletal arrangement. Stimulated cells showed strong alterations on gene, protein and functional levels in several parameters important for maintaining epithelial function. The most significant increases were found in cell motility, cellular speed and cell elongation compared to non-stimulated cells. Also, cell morphology was significantly altered in chymase treated compared to non-stimulated cells. In the current study, we show that MC proteases can induce cell migration and morphological and proliferative alterations in epithelial cells. Thus, our data imply that MC release of proteases may play a critical role in airway epithelial remodeling and disruption of epithelial function.


Assuntos
Brônquios/citologia , Brônquios/metabolismo , Quimases/metabolismo , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Mastócitos/enzimologia , Triptases/metabolismo , Divisão Celular , Linhagem Celular , Movimento Celular , Proliferação de Células , Citoesqueleto/metabolismo , Holografia , Humanos , Imageamento Tridimensional , Imuno-Histoquímica , Análise Serial de Tecidos
6.
PLoS One ; 16(5): e0252624, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34048501

RESUMO

Ruminants have a very complex digestive system adapted for the digestion of cellulose rich food. Gene duplications have been central in the process of adapting their digestive system for this complex food source. One of the new loci involved in food digestion is the lysozyme c locus where cows have ten active such genes compared to a single gene in humans and where four of the bovine copies are expressed in the abomasum, the real stomach. The second locus that has become part of the ruminant digestive system is the chymase locus. The chymase locus encodes several of the major hematopoietic granule proteases. In ruminants, genes within the chymase locus have duplicated and some of them are expressed in the duodenum and are therefore called duodenases. To obtain information on their specificities and functions we produced six recombinant proteolytically active duodenases (three from cows, two from sheep and one from pigs). Two of the sheep duodenases were found to be highly specific tryptases and one of the bovine duodenases was a highly specific asp-ase. The remaining two bovine duodenases were dual enzymes with potent tryptase and chymase activities. In contrast, the pig enzyme was a chymase with no tryptase or asp-ase activity. These results point to a remarkable flexibility in both the primary and extended specificities within a single chromosomal locus that most likely has originated from one or a few genes by several rounds of local gene duplications. Interestingly, using the consensus cleavage site for the bovine asp-ase to screen the entire bovine proteome, it revealed Mucin-5B as one of the potential targets. Using the same strategy for one of the sheep tryptases, this enzyme was found to have potential cleavage sites in two chemokine receptors, CCR3 and 7, suggesting a role for this enzyme to suppress intestinal inflammation.


Assuntos
Duodeno/enzimologia , Serina Endopeptidases/metabolismo , Sequência de Aminoácidos , Animais , Bovinos , Quimases/classificação , Quimases/genética , Biblioteca de Peptídeos , Filogenia , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Serina Endopeptidases/genética , Ovinos , Especificidade por Substrato , Suínos
7.
Biochem Biophys Res Commun ; 551: 127-132, 2021 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-33725574

RESUMO

Mast cell-deficient mice are helpful for understanding the roles of mast cells in vivo. To date, a dozen mouse models for mast cell deficiency have been reported. However, mice with a specific depletion of all populations of mast cells have not been reported. We generated knock-in mice, termed Mcpt5/Cma1DTR mice, expressing human diphtheria toxin A (DT) receptor under the endogenous promoter of Mcpt5 (also known as Cma1), which encodes mouse mast cell protease-5. Flow cytometry and histological analysis showed that intraperitoneal injection of DT induced almost complete depletion of mast cells in heterozygote Mcpt5/Cma1DTR/+ mice. The deletion rates of mast cells in peritoneal cavity, mesentery, abdominal skin, ear skin, and glandular stomach were 99.9%, 100%, 98.7%, 97.7%, and 100%, respectively. Passive cutaneous anaphylaxis reaction also revealed mast cell deficiency in ear skin after DT treatment. Other than mast cells, a small percentage of marginal zone B cells in Mcpt5/Cma1DTR/+ mice were killed by DT treatment. In conclusion, the Mcpt5/Cma1DTR/+ mouse model is valuable for achieving conditional depletion of all populations of mast cells without inducing a marked reduction in other cells.


Assuntos
Separação Celular/métodos , Quimases/genética , Mastócitos/citologia , Modelos Animais , Animais , Células do Tecido Conjuntivo/citologia , Feminino , Humanos , Injeções Intraperitoneais , Camundongos , Membrana Mucosa/citologia , Regiões Promotoras Genéticas/genética
8.
Biomed J ; 44(1): 74-85, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33736953

RESUMO

The newly emerged SARS-CoV-2 strains from the coronavirus (CoV) family is causing one of the most disruptive pandemics of the past century. Developing antiviral drugs is a challenge for the scientific community and pharmaceutical industry. Given the health emergency, repurposing of existing antiviral, antiinflammatory or antimalarial drugs is an attractive option for controlling SARS-CoV-2 with drugs. However, phytochemicals selected based on ethnomedicinal information as well as in vitro antiviral studies could be promising as well. Here, we summarise the phytochemicals with reported anti-CoV activity, and further analyzed them computationally to accelerate validation for drug development against SARS-CoV-2. This systematic review started from the most potent phytocompounds (IC50 in µM) against SARS-CoV, followed by a cluster analysis to locate the most suitable lead(s). The advanced molecular docking used the crystallography structure of SARS-CoV-2-cysteine-like protease (SARS-CoV-2-3CLpro) as a target. In total, seventy-eight phytochemicals with anti-CoV activity against different strains in cellular assays, were selected for this computational study, and compared with two existing repurposed FDA-approved drugs: lopinavir and ritonavir. This review brings insights in the potential application of phytochemicals and their derivatives, which could guide researchers to develop safe drugs against SARS-CoV-2.


Assuntos
COVID-19/tratamento farmacológico , Quimases/antagonistas & inibidores , Compostos Fitoquímicos/uso terapêutico , SARS-CoV-2/efeitos dos fármacos , Quimases/química , Reposicionamento de Medicamentos , Humanos , Simulação de Acoplamento Molecular , Compostos Fitoquímicos/química , SARS-CoV-2/enzimologia
9.
Clin Exp Hypertens ; 43(5): 392-401, 2021 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-33687310

RESUMO

Background: Chymase generates angiotensin II (ANG II) independently of angiotensin-converting enzyme in tissues and it contributes to vascular remodeling and development of hypertension, however the exact mechanism of its action is unclear. Methods: Hence, the effects of chymase inhibition were examined in anesthetized spontaneously hypertensive rats (SHR) in two stages of the disease development, ie. pre-hypertensive (SHR7) and with established hypertension (SHR16). Chymostatin, a commercial chymase inhibitor, was infused intravenously alone or in subsequent groups co-infused with captopril. Results: Mean blood pressure (MBP), total renal blood flow (RBF) and ANG II content (plasma and tissues) were measured. In SHR16 chymase blockade significantly decreased MBP (-6%) and plasma (-38%), kidney (-71%) and heart (-52%) ANG II levels. In SHR7 chymostatin did not influence MBP or RBF, but significantly decreased heart ANG II level. Conclusion: Jointly, functional studies and ANG II determinations support the evidence that in SHR chymase can raise plasma ANG II and contribute to blood pressure elevation. We propose that addition of chymase blockade to ACE inhibition could be a promising approach in the treatment of hypertensive patients resistant to therapy with ACE-inhibitors alone.


Assuntos
Angiotensina II/sangue , Pressão Sanguínea/fisiologia , Quimases/metabolismo , Hemodinâmica , Inibidores da Enzima Conversora de Angiotensina/farmacologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Quimases/antagonistas & inibidores , Taxa de Filtração Glomerular/efeitos dos fármacos , Hemodinâmica/efeitos dos fármacos , Hipertensão/fisiopatologia , Ílio/irrigação sanguínea , Ílio/efeitos dos fármacos , Rim/irrigação sanguínea , Rim/efeitos dos fármacos , Rim/fisiopatologia , Masculino , Oligopeptídeos , Perfusão , Potássio/metabolismo , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Fluxo Sanguíneo Regional/efeitos dos fármacos , Sódio/metabolismo
10.
Cells ; 10(2)2021 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-33546258

RESUMO

The mast cell granule metalloprotease CPA3 is proposed to have important tissue homeostatic functions. However, the basal CPA3 mRNA and protein expression among mast cell populations has remained poorly investigated. Using a novel histology-based methodology that yields quantitative data on mRNA and protein expression at a single-cell level, the present study maps CPA3 mRNA and protein throughout the MCT and MCTC populations in healthy skin, gut and lung tissues. MCTC cells had both a higher frequency of CPA3 protein-containing cells and a higher protein-staining intensity than the MCT population. Among the tissues, skin MCs had highest CPA3 protein intensity. The expression pattern at the mRNA level was reversed. Lung mast cells had the highest mean CPA3 mRNA staining. Intriguingly, the large alveolar MCT population, that lack CPA3 protein, had uniquely high CPA3 mRNA intensity. A broader multi-tissue RNA analysis confirmed the uniquely high CPA3 mRNA quantities in the lung and corroborated the dissociation between chymase and CPA3 at the mRNA level. Taken together, our novel data suggest a hitherto underestimated contribution of mucosal-like MCT to baseline CPA3 mRNA production. The functional consequence of this high constitutive expression now reveals an important area for further research.


Assuntos
Carboxipeptidases/metabolismo , Quimases/metabolismo , Mastócitos/metabolismo , RNA Mensageiro/metabolismo , Triptases/metabolismo , Humanos
11.
Yakugaku Zasshi ; 141(2): 215-233, 2021.
Artigo em Japonês | MEDLINE | ID: mdl-33518643

RESUMO

Studies on functional molecules starting from syntheses of cysteine-containing peptides and protein are described. Starting from evaluation of a cysteine specific side-reaction, a specific reaction for disulfide-bond formation was developed. The reaction made it possible to independently construct a disulfide bridge without effecting the existing disulfide bonds, which resulted in a unique approach for the synthesis of human insulin by site-specific disulfide bond formation. In a series of studies on sulfur-containing amino acids, another cysteine related un-natural amino acid, α-methyl cysteine, was used for the total syntheses of natural products containing a unique thiazorine/thiazole ring system. Chloroimidazolidium coupling reagent developed by us was effective for the successive couplings of the α-methyl cysteine residues. Based on these synthetic studies, design and evaluation of protease inhibitors were then studied, since a stereo-specific synthesis of the key structure is crucial to make the inhibitor an effective functional molecule in the interactions with its target protease. As the target proteases, ß-site amyloid precursor protein cleaving enzyme 1 (BACE1) and chymotrypsin-like protease of severe acute respiratory syndrome (SARS 3CL protease) were selected: the former is a crucial enzyme for amyloid ß production and the latter is an essential enzyme for the re-construction of SARS corona virus in host cells. Structure optimization procedure of the respective inhibitors are described based on X-ray crystal structure analyses of the inhibitor-protease complex.


Assuntos
Aminoácidos/química , Peptídeos/síntese química , Secretases da Proteína Precursora do Amiloide/química , Ácido Aspártico Endopeptidases/química , Produtos Biológicos/síntese química , Produtos Biológicos/química , Quimases/química , Cristalografia por Raios X , Cisteína , Dissulfetos/química , Insulina/síntese química , Peptídeos/química , Inibidores de Proteases/síntese química , Inibidores de Proteases/química , Vírus da SARS , Enxofre/química , Tiazóis/química
12.
Am J Physiol Lung Cell Mol Physiol ; 320(3): L422-L429, 2021 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33404363

RESUMO

The unique clinical features of COVID-19 disease present a formidable challenge in the understanding of its pathogenesis. Within a very short time, our knowledge regarding basic physiological pathways that participate in SARS-CoV-2 invasion and subsequent organ damage have been dramatically expanded. In particular, we now better understand the complexity of the renin-angiotensin-aldosterone system (RAAS) and the important role of angiotensin converting enzyme (ACE)-2 in viral binding. Furthermore, the critical role of its major product, angiotensin (Ang)-(1-7), in maintaining microcirculatory balance and in the control of activated proinflammatory and procoagulant pathways, generated in this disease, have been largely clarified. The kallikrein-bradykinin (BK) system and chymase are intensively interwoven with RAAS through many pathways with complex reciprocal interactions. Yet, so far, very little attention has been paid to a possible role of these physiological pathways in the pathogenesis of COVID-19 disease, even though BK and chymase exert many physiological changes characteristic to this disorder. Herein, we outline the current knowledge regarding the reciprocal interactions of RAAS, BK, and chymase that are probably turned-on in COVID-19 disease and participate in its clinical features. Interventions affecting these systems, such as the inhibition of chymase or blocking BKB1R/BKB2R, might be explored as potential novel therapeutic strategies in this devastating disorder.


Assuntos
COVID-19/patologia , Quimases/metabolismo , Cininas/metabolismo , Sistema Renina-Angiotensina , SARS-CoV-2/isolamento & purificação , COVID-19/metabolismo , COVID-19/virologia , Humanos
13.
J Pharmacol Exp Ther ; 376(2): 213-221, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33154104

RESUMO

Mouse mast cell protease 4 (mMCP-4), the murine functional analog to the human chymase, is a serine protease synthesized and stored in mast cell secretory granules. Our previous studies reported physiologic and pathologic roles for mMCP-4 in the maturation and synthesis of the vasoactive peptide endothelin-1 (ET-1) from its precursor, big ET-1. The aim of this study was to investigate the impact of mast cell degranulation or stabilization on mMCP-4-dependent pressor responses after the administration of big ET-1 or angiotensin I (Ang I). In anesthetized mice, mast cell degranulation induced by compound 48/80 (C48/80) or stabilization by cromolyn enhanced or repressed, respectively, the dose-dependent vasopressor responses to big ET-1 in wild-type (WT) mice but not in mMCP-4 knockout mice in a chymase inhibitor (TY-51469)-sensitive fashion. In addition, mMCP-4-dependent hydrolysis of the fluorogenic substrate Suc-Leu-Leu-Val-Tyr-7-amino-4-methylcoumarin was depleted or enhanced in peritoneal mast cells isolated from mice pretreated with C48/80 or cromolyn, respectively. Furthermore, C48/80 or cromolyn markedly increased or abolished, respectively, ET-1 (1-31) conversion from exogenous big ET-1 in WT mice peritoneal fluid-isolated mast cells, in vitro. Finally, the vasopressor responses to Ang I were unaffected by mast cell activation or stabilization, whereas those induced by the angiotensin-converting enzyme-resistant Ang I analog, [Pro11, D-Ala12] Ang I, were potentiated by C48/80. Altogether, the present study shows that mast cell activation enhances the mMCP-4-dependent vasoactive properties of big ET-1 but not Ang I in the mouse model. SIGNIFICANCE STATEMENT: The current work demonstrates a significant role for mast cell stability in the cardiovascular pharmacology of big endothelin-1 but not angiotensin I in the murine systemic circulation.


Assuntos
Angiotensina I/farmacologia , Pressão Sanguínea , Degranulação Celular , Endotelina-1/farmacologia , Mastócitos/fisiologia , Serina Endopeptidases/metabolismo , Animais , Células Cultivadas , Quimases/antagonistas & inibidores , Cromolina Sódica/farmacologia , Inibidores Enzimáticos/farmacologia , Masculino , Estabilizadores de Mastócitos/farmacologia , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Peritônio/citologia , Serina Endopeptidases/genética , Sulfonamidas/farmacologia , Tiofenos/farmacologia
14.
J Biomol Struct Dyn ; 39(4): 1203-1212, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32036760

RESUMO

A recent research has identified chymase, a mast cell-specific protease as an exclusive novel therapeutic target to prevent Japanese encephalitis virus (JEV) induced encephalitis. Interestingly, JEV activates mast cell specific chymase during its penetration through blood brain barrier (BBB) which eventually guide to viral encephalitis. Hence, in this study, natural chemical entities (NCE) from multiple databases (MPD3, TIPDB and MTDP) were virtually screened for their binding affinity as chymase inhibitors, a promising negotiator for prolong survival against JEV tempted encephalitis. Merged computational programs, Maestro software, QikProp, ProTox and Gromacs were applied to screen the NCEs against target receptor (PDB: 4KP0). Three hits (C00008437, C00014417 and 8141903) were identified after employing a series of sieves such as High Throughput Virtual Screening (HTVS), Standard precision (SP) and Xtra precision (XP) molecular docking simulations followed by desired pharmacokinetic-toxicity profile predictions and molecular dynamics (MD) examinations. Maestro simulations resulted in best three binding energy scores as -11.992 kcal/mol (first ranked; C00008437), -11.673 kcal/mol (second ranked; C00014417) and -11.456 kcal/mol (third ranked; 8141903), respectively. The top three hits revealed an ideal range of pharmacokinetic and toxicity descriptors values. In addition, MD simulations enabled us to confirm top hits higher selectivity toward chymase receptor. In conclusion, this might potentially represent remarkable novel classes with an effective chymase mediated treatment to combat JEV induced encephalitis, which need to justify with further detail studies.


Assuntos
Produtos Biológicos , Encefalite Japonesa , Quimases , Encefalite Japonesa/tratamento farmacológico , Humanos , Mastócitos , Simulação de Acoplamento Molecular
15.
J Biomol Struct Dyn ; 39(7): 2607-2616, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-32238094

RESUMO

Recently, the world has witnessed outbreak of a novel Coronavirus (SARS-CoV-2), the virus which initially emerged in Wuhan, China has now made its way to a large part of the world, resulting in a public emergency of international concern. The functional importance of Chymotrypsin-like protease (3CLpro) in viral replication and maturation turns it into an attractive target for the development of effective antiviral drugs against SARS and other coronaviruses. At present, there is no standard drug regime nor any vaccine available against the infection. The rapid development and identification of efficient interventions against SARS-CoV-2 remains a major challenge. Based on the available knowledge of closely related coronavirus and their safety profiles, repurposing of existing antiviral drugs and screening of available databases is considered a near term strategic and economic way to contain the SARS-CoV-2 pandemic. Herein, we applied computational drug design methods to identify Chymotrypsin-like protease inhibitors from FDA approved antiviral drugs and our in-house database of natural and drug-like compounds of synthetic origin. As a result three FDA approved drugs (Remdesivir, Saquinavir and Darunavir) and two natural compounds (. flavone and coumarine derivatives) were identified as promising hits. Further, MD simulation and binding free energy calculations were performed to evaluate the dynamic behavior, stability of protein-ligand contact, and binding affinity of the hit compounds. Our results indicate that the identified compounds can inhibit the function of Chymotrypsin-like protease (3CLpro) of Coronavirus. Considering the severity of the spread of coronavirus, the current study is in-line with the concept of finding the new inhibitors against the vital pathway of the corona virus to expedite the process of drug discovery.Communicated by Ramaswamy H. Sarma.


Assuntos
COVID-19 , SARS-CoV-2 , Quimases , Humanos , Simulação de Acoplamento Molecular , Pandemias , Inibidores de Proteases/farmacologia
16.
J Biomol Struct Dyn ; 39(9): 3396-3408, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32367767

RESUMO

The novel coronavirus disease 2019 (COVID-19) caused by SARS-COV-2 has raised myriad of global concerns. There is currently no FDA approved antiviral strategy to alleviate the disease burden. The conserved 3-chymotrypsin-like protease (3CLpro), which controls coronavirus replication is a promising drug target for combating the coronavirus infection. This study screens some African plants derived alkaloids and terpenoids as potential inhibitors of coronavirus 3CLpro using in silico approach. Bioactive alkaloids (62) and terpenoids (100) of plants native to Africa were docked to the 3CLpro of the novel SARS-CoV-2. The top twenty alkaloids and terpenoids with high binding affinities to the SARS-CoV-2 3CLpro were further docked to the 3CLpro of SARS-CoV and MERS-CoV. The docking scores were compared with 3CLpro-referenced inhibitors (Lopinavir and Ritonavir). The top docked compounds were further subjected to ADEM/Tox and Lipinski filtering analyses for drug-likeness prediction analysis. This ligand-protein interaction study revealed that more than half of the top twenty alkaloids and terpenoids interacted favourably with the coronaviruses 3CLpro, and had binding affinities that surpassed that of lopinavir and ritonavir. Also, a highly defined hit-list of seven compounds (10-Hydroxyusambarensine, Cryptoquindoline, 6-Oxoisoiguesterin, 22-Hydroxyhopan-3-one, Cryptospirolepine, Isoiguesterin and 20-Epibryonolic acid) were identified. Furthermore, four non-toxic, druggable plant derived alkaloids (10-Hydroxyusambarensine, and Cryptoquindoline) and terpenoids (6-Oxoisoiguesterin and 22-Hydroxyhopan-3-one), that bind to the receptor-binding site and catalytic dyad of SARS-CoV-2 3CLpro were identified from the predictive ADME/tox and Lipinski filter analysis. However, further experimental analyses are required for developing these possible leads into natural anti-COVID-19 therapeutic agents for combating the pandemic.Communicated by Ramaswamy H. Sarma.


Assuntos
Alcaloides , COVID-19 , Plantas Medicinais , Alcaloides/farmacologia , Quimases , Simulação por Computador , Humanos , Inibidores de Proteases , SARS-CoV-2 , Terpenos
17.
J Med Virol ; 93(6): 3330-3337, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-32857465

RESUMO

We conducted a prospective, observational study to assess the serum chymase level, a mast cell derived protease, as a predictor of dengue severity. NS1-positive non-severe dengue patients of age ≥14 years with duration of fever ≤4 days were included in the study. At the time of admission, the serum sample was taken for chymase estimation. Patients were followed up to four days after they became afebrile to find out the final diagnosis. Total of 338 non-severe dengue patients were recruited (mean age: 29.15 years; male: 66%). On follow-up, 26 patients (7.8%) developed severe dengue. Only chymase level (adjusted odds ratio [aOR]: 1.787; 95% confidence interval [CI]: 1.309-2.440) and platelet count at admission (aOR: 0.981; 95% CI: 0.968-0.993) were able to predict the severity after adjustment for all variables. But, for prediction of severe dengue, the area under receiver's operating curve of chymase was 0.835 (95% CI: 0.765-0.905), which was significantly higher than that of the platelet count at admission (0.760, 95% CI: 0.650-0.870) (p < .001). Patients who developed severe dengue in due course of illness had significantly higher serum chymase level at admission as compared with the rest of the patients. Similar findings were noted across all age-groups. At an optimum cut-off value of 1.35 ng/ml, chymase had a positive likelihood ratio (LR) of 3.5 and a negative LR of 0.15, for predicting severe dengue. This study demonstrated the potential ability of serum chymase levels at admission, as a biomarker for prediction of severe dengue in due course of illness.


Assuntos
Quimases/sangue , Dengue Grave/diagnóstico , Dengue Grave/epidemiologia , Índice de Gravidade de Doença , Adolescente , Adulto , Biomarcadores/sangue , Quimases/genética , Vírus da Dengue , Feminino , Febre , Hospitalização , Humanos , Masculino , Razão de Chances , Estudos Prospectivos , Dengue Grave/sangue , Centros de Atenção Terciária/estatística & dados numéricos , Adulto Jovem
19.
J Ethnopharmacol ; 270: 113610, 2021 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-33246121

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Acmella oleracea (L.) R. K. Jansen (Asteraceae), known as jambú in Brazil, is used in traditional medicine as analgesic and for inflammatory conditions, characterized by the presence of N-alkylamides, mainly spilanthol. This bioactive compound is responsible for the above-described pharmacological properties, including sialagogue and anesthetic. AIM OF THE STUDY: This study aimed to characterize the anti-inflammatory effects of A. oleracea leaves (AOEE-L) and flowers (AOEE-F) extracts, including an isolated alkylamide (spilanthol), using in vitro and in vivo models. The mechanism underlying this effect was also investigated. MATERIALS AND METHODS: Extracts were analyzed by HPLC-ESI-MS/MS in order to characterize the N-alkylamides content. AOEE-L, AOEE-F (25-100 µg/mL) and spilanthol (50-200 µM) were tested in vitro on VSMC after stimulation with hyperglycemic medium (25 mM glucose). Their effects over nitric oxide (NO) generation, chymase inhibition and expression, catalase (CAT), superoxide anion (SOD) radical activity were evaluated. After an acute administration of extracts (10-100 mg/mL) and spilanthol (6.2 mg/mL), the anti-inflammatory effects were evaluated by applying the formalin test in rats. Blood was collected to measure serum aminotransferases activities, NO activity, creatinine and urea. RESULTS: A number of distinct N-alkylamides were detected and quantified in AOEE-L and AOEE-F. Spilanthol was identified in both extracts and selected for experimental tests. Hyperglycemic stimulation in VSMC promoted the expression of inflammatory parameters, including chymase, NO, CAT and SOD activity and chymase expression, all of them attenuated by the presence of the extracts and spilanthol. The administration of extracts or spilanthol significantly inhibited edema formation, NO production and cell tissue infiltration in the formalin test, without causing kidney and liver toxicity. CONCLUSION: Taken together, these results provide evidence for the anti-inflammatory activity of leaves and flowers extracts of jambú associated distinctly with their chemical profile. The effects appear to be associated with the inhibition of chymase activity, suppression of the proinflammatory cytokine NO and antioxidant activities.


Assuntos
Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Asteraceae/química , Quimases/antagonistas & inibidores , Extratos Vegetais/farmacologia , Alcamidas Poli-Insaturadas/farmacologia , Animais , Anti-Inflamatórios/uso terapêutico , Antioxidantes/química , Antioxidantes/uso terapêutico , Brasil , Linhagem Celular , Quimases/metabolismo , Edema/induzido quimicamente , Edema/tratamento farmacológico , Edema/patologia , Etanol/química , Flores/química , Formaldeído/toxicidade , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Inflamação/patologia , Masculino , Medicina Tradicional , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Óxido Nítrico/antagonistas & inibidores , Óxido Nítrico/metabolismo , Extratos Vegetais/química , Extratos Vegetais/uso terapêutico , Folhas de Planta/química , Alcamidas Poli-Insaturadas/uso terapêutico , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo
20.
Curr Opin Allergy Clin Immunol ; 21(1): 71-78, 2021 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-33369571

RESUMO

PURPOSE OF REVIEW: Mast cells have previously been thought to function solely as effector cells in asthma but more recent studies have indicated that mast cells may play a more central role in propagating and regulating lower airway inflammation in asthma. RECENT FINDINGS: Initial studies have found increased numbers of mast cell progenitors (MCPs) in the peripheral blood of patients with asthma and these cells could contribute to the increased number of progenitors identified in the airways of patients with asthma. There are unique subpopulations of mast cells within the asthmatic airway, which are characterized by their physical location and distinguished by their expression profile of mast cell proteases. Intraepithelial mast cells are tightly associated with type-2 (T2) inflammation but additional studies have suggested a role for anti-mast cell therapies as a treatment for T2-low asthma. Mast cells have recently been shown to closely communicate with the airway epithelium and airway smooth muscle to regulate lower airway inflammation and airway hyperresponsiveness. SUMMARY: Recent studies have better illuminated the central role of mast cells in regulating lower airway inflammation and airway hyperresponsiveness.


Assuntos
Asma/imunologia , Diferenciação Celular/imunologia , Mastócitos/imunologia , Mucosa Respiratória/patologia , Asma/sangue , Asma/patologia , Carboxipeptidases A/metabolismo , Quimases/metabolismo , Humanos , Inflamação/sangue , Inflamação/imunologia , Inflamação/patologia , Mastócitos/metabolismo , Mucosa Respiratória/citologia , Mucosa Respiratória/imunologia , Transdução de Sinais/imunologia , Triptases/metabolismo
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