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1.
Molecules ; 26(15)2021 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-34361789

RESUMO

Isobavachalcone (IBC) is an active substance from the medicinal plant Psoralea corylifolia. This prenylated chalcone was reported to possess antioxidative, anti-inflammatory, antibacterial, and anticancer activities. Multidrug resistance (MDR) associated with the over-expression of the transporters of vast substrate specificity such as ABCB1 (P-glycoprotein) belongs to the main causes of cancer chemotherapy failure. The cytotoxic, MDR reversing, and ABCB1-inhibiting potency of isobavachalcone was studied in two cellular models: human colorectal adenocarcinoma HT29 cell line and its resistant counterpart HT29/Dx in which doxorubicin resistance was induced by prolonged drug treatment, and the variant of MDCK cells transfected with the human gene encoding ABCB1. Because MDR modulators are frequently membrane-active substances, the interaction of isobavachalcone with model phosphatidylcholine bilayers was studied by means of differential scanning calorimetry. Molecular modeling was employed to characterize the process of membrane permeation by isobavachalcone. IBC interacted with ABCB1 transporter, being a substrate and/or competitive inhibitor of ABCB1. Moreover, IBC intercalated into model membranes, significantly affecting the parameters of their main phospholipid phase transition. It was concluded that isobavachalcone interfered both with the lipid phase of cellular membrane and with ABCB1 transporter, and for this reason, its activity in MDR cancer cells was presumptively beneficial.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Chalconas/farmacologia , Doxorrubicina/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Psoralea/química , Subfamília B de Transportador de Cassetes de Ligação de ATP/antagonistas & inibidores , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Animais , Antibióticos Antineoplásicos/farmacologia , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/isolamento & purificação , Ligação Competitiva , Linhagem Celular Tumoral , Chalconas/química , Chalconas/isolamento & purificação , Cães , Combinação de Medicamentos , Resistencia a Medicamentos Antineoplásicos/genética , Expressão Gênica , Células HT29 , Humanos , Concentração Inibidora 50 , Bicamadas Lipídicas/química , Bicamadas Lipídicas/metabolismo , Células Madin Darby de Rim Canino , Membranas Artificiais , Modelos Moleculares , Fosfatidilcolinas/química , Fosfatidilcolinas/metabolismo , Extratos Vegetais/química , Plantas Medicinais , Ligação Proteica , Transgenes , Verapamil/farmacologia
2.
Chem Biol Interact ; 346: 109570, 2021 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-34217686

RESUMO

Verapamil is reported to prevent scar formation. However, whether verapamil is involved in the ureteral stricture scar and the underlying mechanism need further investigation. Fibroblasts were isolated from ureteral scar tissues. TGF-ß1 stimulation was used to induce fibrosis of fibroblasts. Inhibition of CaMK II was achieved by shRNA transfection. CCK-8 was performed to evaluate cell viability. qRT-PCR was applied to determine the level of mRNA while western blotting was used to determine the level of proteins. Immunofluorescence was used to detect the level of vimentin, collagen I and collagen III. Primary fibroblasts was successfully isolated from ureteral scar tissues. TGF-ß1 stimulation was capable to induce collagen production and fibrosis in primary fibroblasts while inhibition of CaMK II attenuate collagen production. Overexpression of wild type CaMK II lead to further increase of collagen production upon TGF-ß1 stimulation while the mutated CaMK II did not exert this promotion. Treatment of verapamil inhibits TGF-ß1 induced collagen production via inhibiting CaMK II. In present study, we revealed a vital role of Verapamil and CaMK II in the formation of ureteral scar. Verapamil inhibited TGF-ß1 induced collagen fiber formation by regulating CaMK II. Our finding might provide new insight into mechanism of prevention and treatment of ureteral scar.


Assuntos
Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/metabolismo , Verapamil/farmacologia , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/antagonistas & inibidores , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Células Cultivadas , Cicatriz/tratamento farmacológico , Cicatriz/metabolismo , Cicatriz/patologia , Colágeno Tipo I/metabolismo , Colágeno Tipo III/metabolismo , Fibroblastos/citologia , Fibroblastos/metabolismo , Humanos , Mutagênese , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Fator de Crescimento Transformador beta1/farmacologia , Regulação para Cima/efeitos dos fármacos , Verapamil/uso terapêutico , Vimentina/metabolismo
3.
Chronobiol Int ; 38(6): 839-850, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33829951

RESUMO

Verapamil is the first-line preventive medication for cluster headache, an excruciating disorder with strong circadian features. Whereas second- and third-line preventives include known circadian modulators, such as melatonin, corticosteroids, and lithium, the circadian effects of verapamil are poorly understood. Here, we characterize the circadian features of verapamil using both in vitro and in vivo models. In Per2::LucSV reporter fibroblasts, treatment with verapamil (0.03-10 µM) showed a dose-dependent period shortening of the reporter rhythm which reached a nadir at 1 µM, and altered core clock gene expression at 10 µM. Mouse wheel-running activity with verapamil (1 mg/mL added to the drinking water) also resulted in significant period shortening and activity reduction in both male and female free-running wild-type C57BL6/J mice. The temporal patterns of activity reduction, however, differ between the two sexes. Importantly, piezo sleep recording revealed sexual dimorphism in the effects of verapamil on sleep timing and bout duration, with more pronounced adverse effects in female mice. We also found altered circadian clock gene expression in the cerebellum, hypothalamus, and trigeminal ganglion of verapamil-treated mice. Verapamil did not affect reporter rhythms in ex vivo suprachiasmatic nucleus (SCN) slices from Per2:Luc reporter mice, perhaps due to the exceptionally tight coupling in the SCN. Thus, verapamil affects both peripheral (trigeminal ganglion) and central (hypothalamus and cerebellum) nervous system structures involved in cluster headache pathophysiology, possibly with network effects instead of isolated SCN effects. These studies suggest that verapamil is a circadian modulator in laboratory models at both molecular and behavioral levels, and sex is an important biological variable for cluster headache medications. These observations highlight the circadian system as a potential convergent target for cluster headache medications with different primary mechanisms of action.


Assuntos
Relógios Circadianos , Cefaleia Histamínica , Animais , Ritmo Circadiano , Cefaleia Histamínica/tratamento farmacológico , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Circadianas Period/genética , Proteínas Circadianas Period/metabolismo , Sono , Núcleo Supraquiasmático/metabolismo , Verapamil/farmacologia
4.
J Nutr Biochem ; 94: 108749, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33910062

RESUMO

Overexpression of drug efflux transporters is commonly associated with multidrug-resistance in cancer therapy. Here for the first time, we investigated the ability of diindolylmethane (DIM), a dietary bioactive rich in cruciferous vegetables, in enhancing the efficacy of Centchroman (CC) by modulating the drug efflux transporters in human breast cancer cells. CC is a selective estrogen receptor modulator, having promising therapeutic efficacy against breast cancer. The combination of DIM and CC synergistically inhibited cell proliferation and induced apoptosis in breast cancer cells. This novel combination has also hindered the stemness of human breast cancer cells. Molecular docking analysis revealed that DIM had shown a strong binding affinity with the substrate-binding sites of ABCB1 (P-gp) and ABCC1 (MRP1) drug-efflux transporters. DIM has increased the intracellular accumulation of Hoechst and Calcein, the substrates of P-gp and MRP1, respectively, in breast cancer cells. Further, DIM stimulates P-gp ATPase activity, which indicates that DIM binds at the substrate-binding domain of P-gp, and thereby inhibits its efflux activity. Intriguingly, DIM enhanced the intracellular concentration of CC by inhibiting the P-gp and MRP1 expression as well as activity. The intracellular retaining of CC has increased its efficacy against breast cancer. Overall, DIM, a dietary bioactive, enhances the anticancer efficiency of CC through modulation of drug efflux ABC-transporters in breast cancer cells. Therefore, DIM-based nutraceuticals and functional foods can be developed as adjuvant therapy against human breast cancer.


Assuntos
Antineoplásicos/farmacologia , Centocromano/farmacologia , Indóis/farmacologia , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Antineoplásicos/metabolismo , Sítios de Ligação , Transporte Biológico/efeitos dos fármacos , Linhagem Celular Tumoral , Centocromano/metabolismo , Antagonistas de Estrogênios/metabolismo , Antagonistas de Estrogênios/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Paclitaxel/química , Paclitaxel/farmacologia , Ligação Proteica , Verapamil/química , Verapamil/farmacologia
5.
Oxid Med Cell Longev ; 2021: 6667074, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33927797

RESUMO

Sepsis-induced myocardial dysfunction considerably increases mortality risk in patients with sepsis. Previous studies from our group have shown that sepsis alters the expression of structural proteins in cardiac cells, resulting in cardiomyocyte degeneration and impaired communication between cardiac cells. Caveolin-3 (CAV3) is a structural protein present in caveolae, located in the membrane of cardiac muscle cells, which regulates physiological processes such as calcium homeostasis. In sepsis, there is a disruption of calcium homeostasis, which increases the concentration of intracellular calcium, which can lead to the activation of potent cellular enzymes/proteases which cause severe cellular injury and death. The purpose of the present study was to test the hypotheses that sepsis induces CAV3 overexpression in the heart, and the regulation of L-type calcium channels directly relates to the regulation of CAV3 expression. Severe sepsis increases the expression of CAV3 in the heart, as immunostaining in our study showed CAV3 presence in the cardiomyocyte membrane and cytoplasm, in comparison with our control groups (without sepsis) that showed CAV3 presence predominantly in the plasma membrane. The administration of verapamil, an L-type calcium channel inhibitor, resulted in a decrease in mortality rates of septic mice. This effect was accompanied by a reduction in the expression of CAV3 and attenuation of cardiac lesions in septic mice treated with verapamil. Our results indicate that CAV3 has a vital role in cardiac dysfunction development in sepsis and that the regulation of L-type calcium channels may be related to its expression.


Assuntos
Caveolina 3/metabolismo , Coração/efeitos dos fármacos , Sepse/tratamento farmacológico , Verapamil/uso terapêutico , Animais , Canais de Cálcio Tipo L , Humanos , Masculino , Camundongos , Sepse/mortalidade , Sepse/patologia , Análise de Sobrevida , Verapamil/farmacologia
6.
Phytomedicine ; 85: 153528, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33735724

RESUMO

BACKGROUND: P-glycoprotein (P-gp) over-expression plays a vital role in not only systemic drug bioavailability but also cancer multi-drug resistance (MDR). Develop functional inhibitors of P-gp can conquer both problems. PURPOSE AND STUDY DESIGN: The aim of the present study was to research the P-gp modulating effects and MDR reversing ability of a novel flavonoid from Fissistigma cupreonitens, the underlying inhibitory mechanisms were further elucidated as well. METHODS: Calcein-AM, rhodamine 123, and doxorubicin were fluorescent substrates for the evaluation of P-gp inhibitory function and detailed drug binding modes. Docking simulation was performed to reveal the in silico molecular bonding. ATPase assay and MDR1 shift assay were adopted to reveal the ATP consumption and conformational change of P-gp. The MDR reversing effects were demonstrated through cytotoxicity, cell cycle, and apoptosis analyses. RESULTS: 5­hydroxy­7,8­dimethoxyflavanone inhibited the efflux of rhodamine 123 and doxorubicin in a competitive manner, and increased the intracellular fluorescence of calcein at a concentration as low as 2.5 µg/ml. 5­hydroxy­7,8­dimethoxyflavanone slightly changed P-gp's conformation and only stimulated ATPase at very high concentration (100 µg/ml). The docking results showed that 5­hydroxy­7,8­dimethoxyflavanone and verapamil exhibited similar binding affinity to P-gp. The MDR reversing effects were prominent in the vincristine group, the reversal folds were 23.01 and 13.03 when combined with 10 µg/ml 5­hydroxy­7,8­dimethoxyflavanone in the P-gp over-expressing cell line (ABCB1/Flp-In™-293) and MDR cancer cell line (KB/VIN), respectively. CONCLUSION: The present study demonstrated that 5­hydroxy­7,8­dimethoxyflavanone was a novel effective flavonoid in the P-gp efflux inhibition and in vitro cancer MDR reversion.


Assuntos
Annonaceae/química , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Flavonoides/farmacologia , Subfamília B de Transportador de Cassetes de Ligação de ATP/antagonistas & inibidores , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Doxorrubicina/metabolismo , Fluoresceínas/metabolismo , Humanos , Simulação de Acoplamento Molecular , Compostos Fitoquímicos/farmacologia , Rodamina 123/metabolismo , Verapamil/farmacologia
7.
Int J Mol Sci ; 22(4)2021 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-33671517

RESUMO

Recently, it has been suggested that progesterone affects the contractile activity of pregnant myometrium via nongenomic pathways; therefore, we aimed to clarify whether progesterone causes and/or inhibits pregnant myometrial contractions via nongenomic pathways. Our in vitro experiments using myometrial strips obtained from rats at 20 days of gestation revealed that progesterone caused myometrial contractions in a concentration- and time-dependent manner at concentrations up to 5 × 10-7 M; however, this effect decreased at concentrations higher than 5 × 10-5 M. Similarly, progesterone enhanced oxytocin-induced contractions up to 5 × 10-7 M and inhibited contractions at concentrations higher than 5 × 10-5 M. Conversely, progesterone did not enhance high-KCl-induced contractions but inhibited contractions in a concentration- and time-dependent manner at concentrations higher than 5 × 10-7 M. We also found that RU486 did not affect progesterone-induced contractions or the progesterone-induced inhibition of high-KCl-induced contractions; however, progesterone-induced contractions were blocked by calcium-free phosphate saline solution, verapamil, and nifedipine. In addition, FPL64176, an activator of L-type voltage-dependent calcium channels, enhanced high-KCl-induced contractions and rescued the decrease in high-KCl-induced contractions caused by progesterone. Together, these results suggest that progesterone exerts conflicting nongenomic effects on the contractions of pregnant myometrium via putative L-type voltage-dependent calcium channels.


Assuntos
Miométrio/fisiologia , Progesterona/fisiologia , Contração Uterina/fisiologia , Animais , Agonistas dos Canais de Cálcio/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Canais de Cálcio Tipo L/metabolismo , Feminino , Antagonistas de Hormônios/farmacologia , Mifepristona/farmacologia , Miométrio/efeitos dos fármacos , Nifedipino/farmacologia , Técnicas de Cultura de Órgãos , Ocitocina/farmacologia , Cloreto de Potássio/farmacologia , Gravidez , Progesterona/farmacologia , Pirróis/farmacologia , Ratos Wistar , Contração Uterina/efeitos dos fármacos , Verapamil/farmacologia
8.
Gastroenterology ; 160(6): 2072-2088.e6, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33581123

RESUMO

BACKGROUND & AIMS: In upper airway cells, T helper 2 cytokines that signal through interleukin-4 (IL-4) receptor-α have been shown to stimulate eotaxin-3 secretion via a nongastric proton pump (ngH+,K+ATPase). To seek novel targets for eosinophilic esophagitis (EoE) treatments, we evaluated ngH+,K+ATPase expression in EoE squamous cells, and explored molecular pathways involved in eotaxin-3 secretion by IL-4 receptor-α signaling. METHODS: ngH+,K+ATPase expression in EoE cells was evaluated by quantitative real-time polymerase chain reaction and Western blotting. IL-4-stimulated eotaxin-3 secretion was measured by enzyme-linked immunosorbent assay after treatment with omeprazole, SCH 28080 (potassium-competitive acid blocker), ethylene glycol-bis(ß-aminoethyl)-N,N,N',N'-tetraacetoxymethyl ester (calcium chelator), 2-aminoethoxydiphenyl borate (inhibitor of endoplasmic reticulum calcium release), verapamil, and diltiazem (L-type calcium channel inhibitors). Intracellular calcium transients were measured by Fluo-4 fluorescence. Key experiments were confirmed in EoE primary cells and in RNA sequencing datasets from mucosal biopsies of patients with EoE and controls. RESULTS: EoE cells expressed ngH+,K+ATPase messenger RNA and protein. Omeprazole and SCH 28080 decreased IL-4-stimulated eotaxin-3 secretion. IL-4 increased intracellular calcium transients, and IL-4-stimulated eotaxin-3 secretion was blocked by ethylene glycol-bis(ß-aminoethyl)-N,N,N',N'-tetraacetoxymethyl ester, 2-aminoethoxydiphenyl borate, verapamil, and diltiazem. The combination of omeprazole and verapamil suppressed IL-4-stimulated eotaxin-3 secretion more than either agent alone. EoE biopsies expressed higher ngH+,K+ATPase and exhibited more calcium signaling than controls. CONCLUSIONS: EoE cells express a nongastric proton pump that mediates T helper 2 cytokine-stimulated eotaxin-3 secretion. IL-4 induces calcium release from the endoplasmic reticulum and calcium entry via L-type calcium channels, increasing intracellular calcium that contributes to eotaxin-3 secretion by EoE cells. L-type calcium channel inhibitors block T helper 2 cytokine-stimulated eotaxin-3 secretion, suggesting a potential role for these agents in EoE treatment.


Assuntos
Quimiocina CCL26/metabolismo , Esofagite Eosinofílica/metabolismo , Esofagite Eosinofílica/patologia , Células Epiteliais/metabolismo , ATPase Trocadora de Hidrogênio-Potássio/genética , ATPase Trocadora de Hidrogênio-Potássio/metabolismo , Transporte Biológico/efeitos dos fármacos , Compostos de Boro/farmacologia , Cálcio/metabolismo , Bloqueadores dos Canais de Cálcio/farmacologia , Linhagem Celular , Diltiazem/farmacologia , Ácido Egtázico/análogos & derivados , Ácido Egtázico/farmacologia , Mucosa Esofágica/metabolismo , Mucosa Esofágica/patologia , Famotidina/farmacologia , Feminino , Antagonistas dos Receptores H2 da Histamina/farmacologia , Humanos , Subunidade alfa de Receptor de Interleucina-4/metabolismo , Masculino , Omeprazol/farmacologia , Cultura Primária de Células , Inibidores da Bomba de Prótons/farmacologia , Bombas de Próton/efeitos dos fármacos , Bombas de Próton/metabolismo , RNA Mensageiro/metabolismo , Ranitidina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Células Th2/metabolismo , Verapamil/farmacologia
9.
Hum Mol Genet ; 30(3-4): 294-304, 2021 04 26.
Artigo em Inglês | MEDLINE | ID: mdl-33577681

RESUMO

Approaches toward new therapeutics using disease genomics, such as genome-wide association study (GWAS), are anticipated. Here, we developed Trans-Phar [integration of transcriptome-wide association study (TWAS) and pharmacological database], achieving in silico screening of compounds from a large-scale pharmacological database (L1000 Connectivity Map), which have inverse expression profiles compared with tissue-specific genetically regulated gene expression. Firstly we confirmed the statistical robustness by the application of the null GWAS data and enrichment in the true-positive drug-disease relationships by the application of UK-Biobank GWAS summary statistics in broad disease categories, then we applied the GWAS summary statistics of large-scale European meta-analysis (17 traits; naverage = 201 849) and the hospitalized COVID-19 (n = 900 687), which has urgent need for drug development. We detected potential therapeutic compounds as well as anisomycin in schizophrenia (false discovery rate (FDR)-q = 0.056) and verapamil in hospitalized COVID-19 (FDR-q = 0.068) as top-associated compounds. This approach could be effective in disease genomics-driven drug development.


Assuntos
COVID-19/tratamento farmacológico , Desenvolvimento de Medicamentos/métodos , Regulação da Expressão Gênica/efeitos dos fármacos , Estudo de Associação Genômica Ampla/métodos , Estudo de Associação Genômica Ampla/estatística & dados numéricos , Esquizofrenia/tratamento farmacológico , Transcriptoma/genética , Anisomicina/farmacologia , Bases de Dados Genéticas , Bases de Dados de Produtos Farmacêuticos , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/genética , Genômica/métodos , Humanos , Preparações Farmacêuticas , Software , Verapamil/farmacologia
10.
Ann Surg Oncol ; 28(9): 5400-5411, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33566246

RESUMO

BACKGROUND: The membrane transporters activated in cancer stem cells (CSCs) are the target of novel cancer therapies for gastric cancer. The present study investigated ion channel expression profiles in gastric CSCs (GCSCs). METHODS: Cells strongly expressing CD44 were separated from MKN74 cells, a human gastric cancer cell line, by fluorescence-activated cell sorting (FACS), and GCSCs were identified based on tumorsphere formation. Gene expression profiles in GCSCs were examined by a microarray analysis. RESULTS: Among MKN74 cells, CD44 messenger RNA levels were higher in CSCs than in non-CSCs. These CSCs also exhibited resistance to cisplatin. The microarray analysis revealed that the expression of several genes related to voltage-gated Ca2+ channels (VGCCs), including CACNA2D1 and CACNB4, was upregulated. The cytotoxicities of the CACNA2D1 inhibitor amlodipine and the CACNB4 inhibitor verapamil were greater at lower concentrations in CSCs than in non-CSCs, and markedly reduced tumorsphere numbers. Tumor volumes were significantly smaller in a xenograft nude mouse model treated with amlodipine or verapamil in combination with cisplatin than in that treated with cisplatin alone. CONCLUSIONS: The present results indicate that VGCCs play a role in maintaining CSCs, and demonstrated the potential of their specific inhibitors, amlodipine and verapamil, as targeted therapeutic agents against gastric cancer.


Assuntos
Anlodipino , Neoplasias Gástricas , Anlodipino/farmacologia , Linhagem Celular Tumoral , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Humanos , Receptores de Hialuronatos/metabolismo , Células-Tronco Neoplásicas/metabolismo , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/genética , Verapamil/farmacologia
11.
Virulence ; 12(1): 231-243, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-33410730

RESUMO

The emergence of resistance requires alternative methods to treat Candida albicans infections. We evaluated efficacy of the efflux pump inhibitor (EPI) verapamil (VER) with fluconazole (FLC) against FLC-resistant (CaR) and -susceptible C. albicans (CaS). The susceptibility of both strains to VER and FLC was determined, as well as the synergism of VER with FLC. Experiments were performed in vitro for planktonic cultures and biofilms and in vivo using Galleria mellonella. Larval survival and fungal recovery were evaluated after treatment with VER and FLC. Data were analyzed by analysis of variance and Kaplan-Meier tests. The combination of VER with FLC at sub-lethal concentrations reduced fungal growth. VER inhibited the efflux of rhodamine 123 and showed synergism with FLC against CaR. For biofilms, FLC and VER alone reduced fungal viability. The combination of VER with FLC at sub-lethal concentrations also reduced biofilm viability. In the in vivo assays, VER and FLC used alone or in combination increased the survival of larvae infected with CaR. Reduction of fungal recovery was observed only for larvae infected with CaR and treated with VER with FLC. VER reverted the FLC-resistance of C. albicans. Based on the results obtained, VER reverted the FLC-resistance of C. albicans and showed synergism with FLC against CaR. VER also increased the survival of G. mellonella infected with CaR and reduced the fungal recovery.


Assuntos
Antifúngicos/farmacologia , Candida albicans/efeitos dos fármacos , Fluconazol/farmacologia , Larva/efeitos dos fármacos , Proteínas de Membrana Transportadoras/metabolismo , Verapamil/farmacologia , Animais , Biofilmes/efeitos dos fármacos , Transporte Biológico , Candidíase/tratamento farmacológico , Candidíase/microbiologia , Sinergismo Farmacológico , Larva/microbiologia , Testes de Sensibilidade Microbiana , Mariposas/efeitos dos fármacos , Mariposas/microbiologia
12.
Tissue Eng Regen Med ; 18(1): 179-186, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33515165

RESUMO

BACKGROUND: Verapamil is used in the treatment of hypertension, angina pectoris, cardiac arrhythmia, hypertrophic scars, and keloids to block transmembrane calcium ion flux. Verapamil has antioxidant activity, which enhances the production of nitric oxide (NO). NO promotes the proliferation of fibroblasts, keratinocytes, endothelial cells, and epithelial cells during wound healing. In this study, we investigated the effect of verapamil and its antioxidant properties on the enhancement of acute wound healing via NO. METHODS: A full-thickness wound healing model was created on the rat dorsal with a silicone ring. The wound closure rate was estimated every 2 days for 14 days. A histological study was performed to evaluate wound healing. Immunofluorescence staining was analyzed for angiogenesis. The expressions of collagen type I (COL I), collagen type III (COL III), and vascular endothelial growth factor (VEGF) were assessed by Western blot. Real-time polymerase chain reaction (qRT-PCR) was performed to examine the expression of endothelial NO synthase and inducible NO synthase, which are related to antioxidant activity in the process of wound healing. RESULTS: The wound closure rate was faster in the verapamil group compared to the control and silicone groups. Histologic analysis revealed capillaries and stratum basale in the verapamil group. Immunofluorescence staining was shown vessel formation in the verapamil group. Western blot and qRT-PCR analysis revealed high expression levels of COL I, VEGF, eNOS, and FGF in the verapamil. CONCLUSION: Verapamil's antioxidant activity enhances NO production in acute wound healing. We suggest that verapamil can be used to promote acute wound healing.


Assuntos
Antioxidantes , Óxido Nítrico , Animais , Antioxidantes/farmacologia , Células Endoteliais , Ratos , Fator A de Crescimento do Endotélio Vascular , Verapamil/farmacologia , Cicatrização
13.
Biomolecules ; 11(2)2021 01 21.
Artigo em Inglês | MEDLINE | ID: mdl-33494474

RESUMO

Recent evidence suggests that the reason Extra Virgin Olive Oil (EVOO) lowers blood pressure and reduces the risk of developing hypertension is partly due to minor components of EVOO, such as phenols. However, little is still known about the mechanism(s) through which EVOO phenols mediate anti-hypertensive effects. The aim of the present study was to investigate the mechanisms of action of EVOO phenols on mesenteric resistance arteries. A pressure myograph was used to test the effect of EVOO phenols on isolated mesenteric arteries in the presence of specific inhibitors of: 1) BKca channels (Paxillin, 10-5 M); 2) L-type calcium channels (Verapamil, 10-5 M); 3) Ryanodine receptor, RyR (Ryanodine, 10-5 M); 4) inositol 1,4,5-triphosphate receptor, IP3R, (2-Aminoethyl diphenylborinate, 2-APB, 3 × 10-3 M); 5) phospholipase C, PLC, (U73122, 10-5 M), and 6) GPCR-Gαi signaling, (Pertussis Toxin, 10-5 M). EVOO phenols induced vasodilation of mesenteric arteries in a dose-dependent manner, and this effect was reduced by pre-incubation with Paxillin, Verapamil, Ryanodine, 2-APB, U73122, and Pertussis Toxin. Our data suggest that EVOO phenol-mediated vasodilation requires activation of BKca channels potentially through a local increase of subcellular calcium microdomains, a pivotal mechanism on the base of artery vasodilation. These findings provide novel mechanistic insights for understanding the vasodilatory properties of EVOO phenols on resistance arteries.


Assuntos
Microdomínios da Membrana/química , Artérias Mesentéricas/efeitos dos fármacos , Azeite de Oliva/química , Canais de Potássio/química , Fosfolipases Tipo C/metabolismo , Animais , Pressão Sanguínea/efeitos dos fármacos , Compostos de Boro/farmacologia , Canais de Cálcio/química , Estrenos/farmacologia , Receptores de Inositol 1,4,5-Trifosfato/química , Masculino , Paxilina/farmacologia , Toxina Pertussis/farmacologia , Fenol/química , Fenóis/farmacologia , Pirrolidinonas/farmacologia , Ratos , Ratos Sprague-Dawley , Receptores Acoplados a Proteínas G/metabolismo , Rianodina/farmacologia , Canal de Liberação de Cálcio do Receptor de Rianodina/química , Vasodilatação/efeitos dos fármacos , Verapamil/farmacologia
14.
Rhinology ; 59(2): 205-211, 2021 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-33459729

RESUMO

BACKGROUND: P-glycoprotein (P-gp) is a membrane efflux pump which is overexpressed in Chronic Rhinosinusitis with Nasal Polyps (CRSwNP) and promotes Type 2 inflammation. Glucocorticoids (GC) are substrates of P-gp suggesting that overexpression may additionally contribute to GC resistance in CRSwNP. This study aims to determine whether P-gp inhibition using verapamil enhances mometasone retention and efficacy in nasal polyp explants. METHODOLOGY: IRB approved study in which organotypic polyp explants were exposed to mometasone (4.15 µg/mL) and verapa- mil (125 µg/mL) as mono and combination therapy. The effect of verapamil on mometasone tissue retention over time was deter- mined using HPLC. The effect of verapamil on mometasone anti-inflammatory function was determined using ELISA for secreted IL-5. Groups were compared using Kruskal-Wallis test. RESULTS: P-gp expression strongly and significantly inversely correlated with mometasone retention 1hr after exposure, with a ne- arly 6-fold reduction in tissue retention between the lowest and highest P-gp expressing polyp explants. P-gp inhibition reversed this effect and significantly improved mometasone retention at 1hr relative to mometasone alone. The combination of mome- tasone and verapamil significantly reduced IL-5 secretion relative to vehicle control and outperformed either treatment alone. CONCLUSIONS: Our study confirms that P-gp contributes to mometasone resistance. This P-gp mediated resistance was successfully reversed by addition of the P-gp inhibitor verapamil. Verapamil further significantly enhanced the anti-inflammatory effect of mometasone when given as a combination therapy.


Assuntos
Pólipos Nasais , Rinite , Sinusite , Subfamília B de Transportador de Cassetes de Ligação de ATP , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Doença Crônica , Humanos , Furoato de Mometasona/farmacologia , Pólipos Nasais/tratamento farmacológico , Sinusite/tratamento farmacológico , Verapamil/farmacologia
15.
Medicine (Baltimore) ; 100(2): e24032, 2021 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-33466149

RESUMO

RATIONALE: Long QT syndrome (LQTS) is an inheritable disease characterized by prolonged QT interval on the electrocardiogram. The pathogenesis of LQTS is related to mutations in LQTS-susceptible genes encoding cardiac ion channel proteins or subunits. PATIENT CONCERNS: Here, we reported a 37-year-old female Uygur patient with palpitation and loss of consciousness. DIAGNOSES: At the time of admission, a 12-lead electrocardiogram showed a QTc interval of 514 ms. Genetic analysis revealed KCNQ1 G219E and TRPM4 T160M mutations. INTERVENTIONS: Although beta-blockers remain the mainstay in treating LQTS, the patient underwent implantation of an automatic cardioverter defibrillator due to life-threatening arrhythmias. OUTCOMES: To explore the effect of the calcium ion antagonist verapamil on ion channels, we generated human induced pluripotent stem cell cardiomyocytes (hiPSC-CMs) from the peripheral blood mononuclear cells of the patient. The changes of action potential duration in response to verapamil were observed. LESSONS: Our results showed that patient-derived hiPSC-CMs could recapitulate the electrophysiological features of LQTS and display pharmaceutical responses to verapamil.


Assuntos
Canal de Potássio KCNQ1/genética , Síndrome do QT Longo/genética , Canais de Cátion TRPM/genética , Adulto , Bloqueadores dos Canais de Cálcio/farmacologia , Desfibriladores Implantáveis , Feminino , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Síndrome do QT Longo/cirurgia , Verapamil/farmacologia
16.
Otolaryngol Head Neck Surg ; 164(1): 104-109, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32633618

RESUMO

OBJECTIVE: Papaverine is a topical vasodilator commonly used during microvascular surgery to inhibit undesired vasoconstriction. A previous national shortage of papaverine prompted evaluation of an alternative, effective vasodilator. This study aims to assess the experience of a solution of verapamil and nitroglycerin (VG) as a potential alternative pharmacologic vasodilator. STUDY DESIGN: Retrospective case series. SETTING: Two tertiary academic medical centers. SUBJECTS AND METHODS: Among 298 patients, 306 consecutive free tissue transfers performed between 2014 and 2017 for head and neck defect reconstruction utilized a VG solution. Patient and flap characteristics, intraoperative patient and flap complications, and postoperative complications were reviewed. Diameter of the cervical recipient artery was measured intraoperatively before and after topical application of the VG solution in a subset of 43 patients (44 flaps). RESULTS: Flaps included fibula, radial forearm, subscapular system, and anterolateral thigh. In total, 3 (0.98%) flaps failed with varied etiology unrelated to the VG solution (venous thrombosis, arterial anastomosis thrombosis, physical damage to the perforator). Specific to topical application of the VG solution, the mean recipient artery diameter increased from 2.1 to 3.1 mm, a 48% increase (P < .01). There were no intraoperative cardiac events or complications attributable to the VG solution. CONCLUSION: We describe the use of a VG solution for pharmacologic vasodilation during microvascular free tissue transfer. Its use was associated with an acceptable incidence of adverse events, none of which were directly attributable to the VG solution. Apparent and sustained vasodilation was demonstrated. The VG solution represents a safe and efficacious alternative to papaverine in microvascular surgery.


Assuntos
Neoplasias de Cabeça e Pescoço/cirurgia , Microcirurgia , Nitroglicerina/farmacologia , Retalhos Cirúrgicos/irrigação sanguínea , Procedimentos Cirúrgicos Vasculares , Vasodilatação/efeitos dos fármacos , Verapamil/farmacologia , Administração Tópica , Rejeição de Enxerto , Humanos , Nitroglicerina/administração & dosagem , Papaverina/farmacologia , Estudos Retrospectivos , Soluções , Verapamil/administração & dosagem
17.
Biomed Chromatogr ; 35(2): e4976, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32852057

RESUMO

Multidrug resistance remains a huge challenge in the chemotherapy of cancer and numerous studies have reported that P-glycoprotein is the most common mechanism of multidrug resistance. Verapamil has been shown to be able to reverse development of multidrug resistance mediated by P-glycoprotein. However, the mechanism of action for verapamil in reversing multidrug resistance at the metabolic level has been rarely reported. In this research, we report the reversal effect of verapamil on multidrug resistance and its mechanisms of action using metabolomics. The results show that the P-glycoprotein-mediated chemotherapy drug resistance was significantly reversed by verapamil in resistant SW620/Ad300 cells. In-depth studies demonstrated that verapamil at reversal concentration had no effect on the P-glycoprotein expression level, but increased intramolecular accumulation of paclitaxel in SW620/Ad300 cells. Metabolomics revealed that the multidrug resistance of SW620/Ad300 cells was related to changes in glycerophospholipid metabolism, sphingolipid metabolism and citric acid cycle, and verapamil could antagonize the multidrug resistance by reversing the above-mentioned glycerophospholipid metabolism and sphingolipid metabolism. This research shows the multidrug resistance reversal mechanism of verapamil at the metabolic level, which helps in understanding the exact multidrug resistance mechanism of verapamil and might be potentially useful to find new multidrug resistance reversal agents. The combination of verapamil (VRP) and paclitaxel (PTX) yielded synergistic effects. VRP had no effect on the expression of P-gp, but increased intramolecular accumulation of PTX. VRP antagonized the MDR by regulating glycerophospholipid metabolism and sphingolipid metabolism.


Assuntos
Neoplasias Colorretais/metabolismo , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Metaboloma/efeitos dos fármacos , Verapamil/farmacologia , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Antineoplásicos/farmacocinética , Antineoplásicos/farmacologia , Biomarcadores/análise , Biomarcadores/metabolismo , Linhagem Celular Tumoral , Cromatografia Líquida de Alta Pressão , Sinergismo Farmacológico , Humanos , Metabolômica/métodos , Paclitaxel/farmacocinética , Paclitaxel/farmacologia , Espectrometria de Massas em Tandem , Verapamil/farmacocinética
18.
FEBS J ; 288(1): 127-141, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32338825

RESUMO

Most neurodegenerative disorders are associated with aggregation and accumulation of misfolded proteins. One of these proteins, tau, is involved in a number of pathologies including Alzheimer's disease and frontotemporal dementia. Aggregation and phosphorylation of tau have been shown to be a trigger for abnormal signal transduction and disruption of cellular homeostasis. Here, we have studied the effect of extracellular tau at different stages of aggregation in cortical co-cultures of neurons and astrocytes, to understand how this process affects tau pathogenicity. We found that the species formed after prolonged in vitro aggregation of tau (longer than 1 day) are able to stimulate reactive oxygen species (ROS) production through the activation of NADPH oxidase without decreasing the level of the endogenous antioxidant glutathione. The same late insoluble aggregates of tau induced calcium signals in neurons and a gradual increase in the ionic current of artificial membranes. Both tau-induced calcium signals and ROS production in NADPH oxidase were reduced in the presence of the inhibitor of voltage-gated calcium channels (VGCC) nifedipine. This suggests that insoluble aggregates of tau incorporate into the membrane and modify ionic currents, changing plasma membrane potential and activating VGCCs, which induces a calcium influx that triggers ROS production in NADPH oxidase. The combination of all these effects likely leads to toxicity, as only the same insoluble tau aggregates which demonstrated membrane-active properties produced neuronal cell death.


Assuntos
Astrócitos/efeitos dos fármacos , Canais de Cálcio/genética , Regulação da Expressão Gênica/efeitos dos fármacos , NADPH Oxidases/genética , Neurônios/efeitos dos fármacos , Proteínas tau/farmacologia , Peptídeos beta-Amiloides/agonistas , Peptídeos beta-Amiloides/genética , Peptídeos beta-Amiloides/metabolismo , Animais , Animais Recém-Nascidos , Astrócitos/citologia , Astrócitos/metabolismo , Cálcio/metabolismo , Canais de Cálcio/metabolismo , Morte Celular/efeitos dos fármacos , Técnicas de Cocultura , Humanos , NADPH Oxidases/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Nifedipino/farmacologia , Oxirredução , Cultura Primária de Células , Agregados Proteicos , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/agonistas , Espécies Reativas de Oxigênio/metabolismo , Tapsigargina/farmacologia , Verapamil/farmacologia , Proteínas tau/genética , Proteínas tau/metabolismo
19.
Lancet Neurol ; 20(1): 29-37, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33245858

RESUMO

BACKGROUND: Prednisone is commonly used for initial short-term therapy of episodic cluster headaches before preventive medication such as verapamil becomes effective, but this strategy has not been tested in large randomised trials. We aimed to access the safety and efficacy of this treatment approach. METHODS: This study was a multicentre, randomised, double-blind, placebo-controlled trial done in ten specialised headache centres in Germany. Patients with episodic cluster headaches who were aged between 18 and 65 years and within a current pain episode for not more than 30 days, received 100 mg oral prednisone for 5 days followed by tapering of 20 mg every 3 days, or matching placebo (17 days total exposure). All patients received oral verapamil for long-term prevention, starting with 40 mg three times daily and increasing to 120 mg three times daily by day 19; patients then continued with verapamil 120 mg throughout the study. Randomisation was computer-generated at a 1:1 ratio by use of an interactive web-response system, with stratification according to age, sex, and participating site. Participants, investigators, and those assessing outcomes were unaware of treatment allocation. The primary endpoint was the mean number of attacks within the first week of treatment with prednisone compared with placebo. An attack was defined as a unilateral headache with moderate-to-severe intensity of at least five on a numerical rating scale. All efficacy and safety analyses were done in the modified intention-to-treat (mITT) population, which consisted of all patients who had been randomly assigned to a trial group and received at least one dose of prednisone or placebo. The study was stopped early due to slow recruitment and expired funding. The study was registered with EudraCT (2011-006204-13) and with the German Clinical Trials Register (DRKS00004716). FINDINGS: Between April 5, 2013, and Jan 11, 2018, 118 patients were enrolled in the study. Two patients dropped out immediately and 116 patients were randomly assigned (57 patients to prednisone and 59 patients to placebo); 109 patients were included in the mITT analysis (53 patients assigned to prednisone and 56 patients assigned to placebo). Participants in the prednisone group had a mean of 7·1 (SD 6·5) attacks within the first week compared with 9·5 (6·0) attacks in the placebo group (difference -2·4 attacks, 95% CI -4·8 to -0·03; p=0·002). Two serious adverse events occurred, both in the placebo group (inguinal hernia and severe deterioration of cluster headache). A total of 270 adverse events were observed: in the prednisone group, 37 (71%) of 52 patients reported 135 adverse events (most common were headache, palpitations, dizziness, and nausea) and in the placebo group, 39 (71%) of 55 patients had 135 adverse events (most common were nausea, dizziness, and headache). INTERPRETATION: Oral prednisone was an effective short-term preventive therapy in our population of patients with episodic cluster headache. Our findings support the use of prednisone as a first-line treatment in parallel to the up-titration of verapamil, although the efficacy of prednisone alongside other long-term prevention requires additional investigation. FUNDING: German Federal Ministry for Education and Research.


Assuntos
Bloqueadores dos Canais de Cálcio/farmacologia , Cefaleia Histamínica/prevenção & controle , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Glucocorticoides/farmacologia , Avaliação de Resultados em Cuidados de Saúde , Prednisona/farmacologia , Verapamil/farmacologia , Adulto , Bloqueadores dos Canais de Cálcio/administração & dosagem , Método Duplo-Cego , Quimioterapia Combinada , Feminino , Glucocorticoides/administração & dosagem , Glucocorticoides/efeitos adversos , Humanos , Masculino , Pessoa de Meia-Idade , Prednisona/administração & dosagem , Prednisona/efeitos adversos , Verapamil/administração & dosagem
20.
An Acad Bras Cienc ; 92(4): e20200703, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33331390

RESUMO

Candida is a human fungal pathogen that causes a wide range of diseases. Candida albicans is the main etiologic agent in these diseases; however, infections can be caused by non-albicans Candida species. Virulence factors such as biofilm production, which protect the fungus from host immunity and anti-fungal drugs, are important for the infection. Therefore, available antifungal drugs for candidiasis treatment are limited and the investigation of new and effective drugs is needed. Verapamil is a calcium channel blocker with an inhibitory effect on hyphae development, adhesion, and colonization of C. albicans. In this study, we investigated the effect of verapamil on cell viability and its antifungal and anti-biofilm activity in non-albicans Candida species. Verapamil was not toxic to keratinocyte cells; moreover, C. krusei, C. parapsilosis, and C. glabrata were susceptible to verapamil with a minimal inhibitory concentration (MIC) of 1250 µM; in addition, this drug displayed fungistatic effect at the evaluated concentrations. After treatment with verapamil, reduced viability, biomass, and mitochondrial activity were observed in biofilms of the non-albicans Candida species C. krusei, C. glabrata, and C. parapsilosis. These findings highlight the importance of the study of verapamil as an alternative treatment for infections caused by non-albicans Candida species.


Assuntos
Antifúngicos , Candida , Antifúngicos/farmacologia , Biofilmes , Bloqueadores dos Canais de Cálcio/farmacologia , Candida albicans , Humanos , Testes de Sensibilidade Microbiana , Verapamil/farmacologia
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