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1.
Biomed Chromatogr ; 35(9): e5140, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33830528

RESUMO

Owing to the complexity of the composition of herbal and dietary supplements, it is a challenging problem to efficiently screen and identify active or toxic compounds. Psoralea corylifolia L. (PCL) was selected as the subbject to establish a methodology for rapid screening and identification of hepatotoxic compounds. High-content imaging, ultra-performance liquid chromatography and high-resolution mass spectrometry were used in this study to detect the hepatotoxicity and identify unknown compounds in PCL samples. Then, putative toxic compounds which are highly related to hepatotoxicity were screened by spectrum-toxicity correlation analysis, and the toxicity intensity verified by high-content imaging. The maximum nontoxic dose of processed samples with good detoxification effect reduced more than 9 times compared with unprocessed raw medicinal materials. Spectrum-toxicity correlation analysis showed that bavachinin A, bavachin, isobavachalcone and neobavaisoflavone had high correlation with the hepatotoxicity of PCL, and psoralen and isopsoralen had low correlation with hepatotoxicity. This study verified the hepatotoxicity of these six putative compound monomers, proving the results of spectrum-toxicity correlation analysis. Based on the correlation analysis of high-resolution mass spectrometry of detection compounds and high-content imaging of hepatocyte toxicity data, the potential toxic compound of herbal and dietary supplement products can be quickly and accurately screened.


Assuntos
Suplementos Nutricionais/toxicidade , Medicamentos de Ervas Chinesas/toxicidade , Hepatócitos/efeitos dos fármacos , Psoralea/química , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Ficusina/toxicidade , Flavonoides/toxicidade , Humanos , Isoflavonas/toxicidade , Espectrometria de Massas/métodos , Imagem Molecular/métodos
2.
Hum Exp Toxicol ; 40(6): 1012-1021, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33317360

RESUMO

Fructus Psoraleae (FP), widely used in traditional medicine, is increasingly reported to cause serious hepatotoxicity in recent years. However, the main toxic constituents responsible for hepatotoxicity and the underlying mechanisms are poorly understood. In the present study, psoralen, a main and quality-control constituent of FP, was intragastrically administered to Sprague-Dawley rats at a dose of 60 mg/kg for 1, 3 and 7 days. Blood and selected tissue samples were collected and analyzed for biochemistry and histopathology to evaluate hepatotoxicity. The results showed that psoralen could induce hepatotoxicity by enhanced liver-to-body weight ratio and alterations of serum alanine aminotransferase (ALT), aspartate aminotransferase (AST) and total cholesterol after administration for 3 days. In addition, histopathological examinations also indicated the hepatotoxicity induced by psoralen. Furthermore, the mRNA and protein levels of hepatic bile acid transporters were significantly changed, in which MRP4, ABCG5 and ABCG8 were repressed, while the protein level of NTCP tended to increase in the rat liver. Taken together, psoralen caused liver injury possibly through affecting bile acid transporters, leading to the disorder of bile acid transport and accumulation in hepatocytes.


Assuntos
Bile/metabolismo , Proteínas de Transporte/efeitos dos fármacos , Proteínas de Transporte/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Reagentes para Ligações Cruzadas/farmacocinética , Ficusina/metabolismo , Ficusina/toxicidade , Animais , Reagentes para Ligações Cruzadas/metabolismo , Reagentes para Ligações Cruzadas/farmacologia , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Ratos , Ratos Sprague-Dawley
3.
Dermatitis ; 32(3): 140-143, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33273237

RESUMO

Phytophotodermatitis is a cutaneous reaction that occurs after exposure to plant-derived furocoumarins and ultraviolet A light. Psoralen is the most common phototoxic furocoumarin and is present in varying levels within many different plant species. This article focuses on the diagnosis and management of psoralen-induced phytophotodermatitis along with other clinical applications.


Assuntos
Dermatite Fototóxica/etiologia , Ficusina/toxicidade , Furocumarinas/toxicidade , Transtornos de Fotossensibilidade/etiologia , Raios Ultravioleta/efeitos adversos , Humanos , Plantas Tóxicas
4.
Leg Med (Tokyo) ; 47: 101740, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32634765

RESUMO

Hepatotoxicity from paracetamol/acetaminophen has occasionally been reported at lower than expected doses. As herbal preparations may interact with pharmaceutical drugs the following in vitro study was undertaken to determine whether the toxic effects of paracetamol on liver cell growth in culture would be exacerbated by the addition of psoralen, a furanocoumarin compound that is present in Psoralea corylifolia, a common Chinese herb. The following study utilising a liver carcinoma cell line (HepG2) showed that Psoralea corylifolia was significantly toxic from 0.3 mg/ml to 5 mg/ml (p < 0.05), whereas paracetamol was not toxic below 50 mM (p = 0.0026). Interactions between previously non-toxic levels of 0.1 mg/ml of Psoralea corylifolia and increasing concentrations of paracetamol (0-50 mM), however, were observed, with a significant increase in toxicity compared to paracetamol alone (30% cell death vs. 72% cell death with Psoralea corylifolia). A significant synergistic interaction was observed at 40 mM paracetamol with 0.1 mg/ml of Psoralea (p = 0.038). This study has, therefore, shown significantly increased hepatotoxicity in cell cultures exposed to paracetamol when herbal compounds containing furanocoumarins were added. Fulminant acute liver failure occurring after the ingestion of low doses of paracetamol may not, therefore, always be due to an occult idiosyncratic response to paracetamol, but instead possibly to the combined effects of paracetamol and herbal preparations. Given the widespread use of both paracetamol and herbal preparations this possibility should be considered in cases of unexplained hepatic necrosis and liver failure that present for medicolegal investigation.


Assuntos
Acetaminofen/toxicidade , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Ficusina/toxicidade , Fígado/patologia , Morte Celular/efeitos dos fármacos , Doença Hepática Induzida por Substâncias e Drogas/patologia , Sinergismo Farmacológico , Ficusina/isolamento & purificação , Células Hep G2 , Humanos , Falência Hepática/induzido quimicamente , Necrose/induzido quimicamente , Psoralea/química
5.
Toxicol Mech Methods ; 30(1): 39-47, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31364909

RESUMO

Psoralen has potential hepatotoxicity and has a certain promoting effect on the clinical liver injury of Psoralea corylifolia L (Fructus Psoraleae). This study investigated the underlying mechanisms of psoralen-induced hepatotoxicity in vitro. HepG2 cells were treated with psoralen for 6, 12, 24, or 48 h, and an endoplasmic reticulum (ER) stress-specific inhibitor, 4-PBA, was employed to investigate the mechanism of psoralen on ER stress and unfolded protein response (UPR). Cell viability was tested by MTT assay, ATP assay, and cell death by LDH. The apoptosis was reflected by the flow cytometry, caspase-8, and caspase-3 activates. The expression of ER stress-related markers was determined by RT-PCR and western blot. We found that psoralen significantly decreased cell viability, increased activities of caspase-8 and caspase-3, and upregulated expression of CHOP and BAX in a time- and dose-dependent manner. Moreover, psoralen significantly increased the expression and transcription levels of ER stress-related markers, including Grp78, PERK, eIF2α, ATF4, and ATF6, while IRE1α was not significantly affected. And 4-PBA could effectively inhibit psoralen-induced cell death and apoptosis along with the inhibition of ER stress responses. These results suggested that psoralen causes liver injury due to the induction of the ER stress-mediated apoptosis via PERK-eIF2α-ATF4-CHOP and ATF6-CHOP related pathways.


Assuntos
Fator 6 Ativador da Transcrição/metabolismo , Apoptose/efeitos dos fármacos , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Ficusina/toxicidade , Fígado/efeitos dos fármacos , eIF-2 Quinase/metabolismo , Fator 4 Ativador da Transcrição/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/enzimologia , Doença Hepática Induzida por Substâncias e Drogas/patologia , Fator de Iniciação 2 em Eucariotos/metabolismo , Células Hep G2 , Humanos , Fígado/enzimologia , Fígado/patologia , Transdução de Sinais , Fator de Transcrição CHOP/metabolismo , Resposta a Proteínas não Dobradas/efeitos dos fármacos
6.
Exp Eye Res ; 187: 107767, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31437439

RESUMO

Limbal Stem Cell Deficiency (LSCD) is a painful and debilitating disease that results from damage or loss of the Corneal Epithelial Stem Cells (CESCs). Therapies have been developed to treat LSCD by utilizing epithelial stem cell transplants. However, effective repair and recovery depends on many factors, such as the source and concentration of donor stem cells, and the proper conditions to support these transplanted cells. We do not yet fully understand how CESCs heal wounds or how transplanted CESCs are able to restore transparency in LSCD patients. A major hurdle has been the lack of vertebrate models to study CESCs. Here we utilized a short treatment with Psoralen AMT (a DNA cross-linker), immediately followed by UV treatment (PUV treatment), to establish a novel frog model that recapitulates the characteristics of cornea stem cell deficiency, such as pigment cell invasion from the periphery, corneal opacity, and neovascularization. These PUV treated whole corneas do not regain transparency. Moreover, PUV treatment leads to appearance of the Tcf7l2 labeled subset of apical skin cells in the cornea region. PUV treatment also results in increased cell death, immediately following treatment, with pyknosis as a primary mechanism. Furthermore, we show that PUV treatment causes depletion of p63 expressing basal epithelial cells, and can stimulate mitosis in the remaining cells in the cornea region. To study the response of CESCs, we created localized PUV damage by focusing the UV radiation on one half of the cornea. These cases initially develop localized stem cell deficiency characteristics on the treated side. The localized PUV treatment is also capable of stimulating some mitosis in the untreated (control) half of those corneas. Unlike the whole treated corneas, the treated half is ultimately able to recover and corneal transparency is restored. Our study provides insight into the response of cornea cells following stem cell depletion, and establishes Xenopus as a suitable model for studying CESCs, stem cell deficiency, and other cornea diseases. This model will also be valuable for understanding the nature of transplanted CESCs, which will lead to progress in the development of therapeutics for LSCD.


Assuntos
Córnea/fisiologia , Doenças da Córnea/fisiopatologia , Epitélio Corneano/patologia , Células-Tronco/patologia , Cicatrização/fisiologia , Animais , Proliferação de Células , Reagentes para Ligações Cruzadas/toxicidade , Modelos Animais de Doenças , Epitélio Corneano/efeitos dos fármacos , Ficusina/toxicidade , Técnica Indireta de Fluorescência para Anticorpo , Homeostase/fisiologia , Marcação In Situ das Extremidades Cortadas , Fenótipo , Regeneração/fisiologia , Células-Tronco/efeitos dos fármacos , Raios Ultravioleta , Xenopus laevis
7.
Basic Clin Pharmacol Toxicol ; 125(6): 527-535, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31271704

RESUMO

BACKGROUND: The main bioactive components of Fructus psoraleae, such as psoralen and isopsoralen, are known to be hepatotoxic. However, its underlying mechanism is to be elucidated. METHODS: To address this, SD rats were randomly divided into control group, 60 mg/kg psoralen group and 60 mg/kg isopsoralen group. Blood was collected to detect serum biochemical indices. RNA was extracted from liver samples, and then, cDNA gene expression profiles were analysed. RESULTS: Psoralen administration significantly up-regulated serum AST (aspartate aminotransferase) while addition of isopsoralen increased serum ALT (alanine aminotransferase), AST, TBA (total bile acid) and TG (total triglyceride) levels. A total of 172 differentially expressed genes (DEGs) were acquired between psoralen group and control group while 884 DEGs were screened between isopsoralen group and control group. Chemical Carcinogenesis and Metabolism of Xenobiotics by Cytochrome P450 were the two most significantly enriched pathways as revealed by DEGs. Liver was the most impacted organ, and endoplasmic reticulum was the most impacted organelle in subcellular level. Finally, some kinds of cancers and cytochrome p450 oxidoreductase deficiency were predicted. Taken together, psoralen and isopsoralen might cause hepatotoxicity mainly through cytochrome P450 metabolism of xenobiotics. Furthermore, Cyp1a1, Cyp1a2, Gstm1 and Akr7a3 worked as key genes in hepatotoxicity. Moreover, endoplasmic reticulum was the main target subcellular structure in hepatotoxicity induced by psoralen and isopsoralen.


Assuntos
Doença Hepática Induzida por Substâncias e Drogas/etiologia , Ficusina/toxicidade , Furocumarinas/toxicidade , Fígado/efeitos dos fármacos , Animais , Doença Hepática Induzida por Substâncias e Drogas/genética , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Perfilação da Expressão Gênica , Fígado/metabolismo , RNA Mensageiro , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley
8.
Food Chem Toxicol ; 125: 133-140, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30597224

RESUMO

Fructus Psoraleae (FP) causes cholestatic liver injury; however, its main toxic constituents that are responsible for causing hepatotoxicity remained undetermined in previous studies. In the present study, psoralen and isopsoralen, the two main constituents of FP, were administered orally to rats (80 and 40 mg/kg, respectively) and mice (320 and 160 mg/kg, respectively) for 28 days, followed by biochemical and histopathological examinations to evaluate their hepatotoxicity. The results showed that psoralen and isopsoralen could induce the toxic reactions of liver and other organs in rats, while mice were not sensitive to these two compounds. Furthermore, the corresponding results indicated that administration of psoralen and isopsoralen repressed the expression of CYP7A1, BSEP, MRP2 and SULT2A1 and increased the expression of FXR and MRP3 in the rat liver. In summary, the toxic reactions of psoralen and isopsoralen are different in different species. In this study, multiple organ toxicity, such as cholestatic liver injury, occurs in rats, but not in mice. Psoralen and isopsoralen are the two main toxic constituents of FP. In addition, psoralen and isopsoralen cause liver injury, possibly through inhibiting bile acid excretion in the liver, leading to the accumulation of toxin in hepatocytes.


Assuntos
Colestase Intra-Hepática/induzido quimicamente , Ficusina/toxicidade , Furocumarinas/toxicidade , Hepatócitos/efeitos dos fármacos , Extratos Vegetais/química , Animais , Fabaceae , Feminino , Camundongos Endogâmicos ICR , RNA Mensageiro/metabolismo , Ratos Wistar
9.
Chem Res Toxicol ; 31(9): 852-860, 2018 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-30132663

RESUMO

Psoralen is the main active component of Psoralea corylifolia and is used as a marker to assess its quality. The effects of psoralen on animals have been well characterized. However, the molecular pathway of its toxicity is not fully understood. In this study, the toxic effects of psoralen administration (60 mg/kg) for 7 days in Sprague-Dawley rats were observed. Serum biochemistry and liver histopathology were further investigated. Proton nuclear magnetic resonance was applied to characterize the metabolic profile of liver toxicity induced by psoralen and to find changed metabolites in rat serum and liver. It was revealed that visceral coefficients and serum biochemistry indexes were significantly changed in rats with psoralen-induced liver injury. Furthermore, the histopathological examination exhibited that the liver might be the target organ for psoralen. Metabolic analysis of both serum and liver samples further proved the liver was the target of toxicity of psoralen. Multivariate analysis identified 7 metabolites in serum samples and 15 in liver samples as potential biomarkers in liver injury induced by psoralen. In addition, our results suggest that psoralen can cause a disturbance in amino acid metabolism, especially valine, leucine, and isoleucine biosynthesis in both serum and liver samples. In conclusion, we combined the results of toxicity and metabolomics induced by psoralen and provide useful information about the safety and potential risks of psoralen and Psoralea corylifolia.


Assuntos
Doença Hepática Induzida por Substâncias e Drogas/sangue , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Ficusina/toxicidade , Fígado/efeitos dos fármacos , Espectroscopia de Ressonância Magnética/métodos , Metabolômica , Animais , Biomarcadores/sangue , Biomarcadores/metabolismo , Fígado/lesões , Fígado/metabolismo , Ratos , Ratos Sprague-Dawley
10.
Bioorg Med Chem ; 21(17): 5047-53, 2013 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-23886808

RESUMO

New tetracyclic benzofurocoumarin (benzopsoralen) analogues were synthesized and their inhibitory effect on the growth of tumor cell lines was evaluated. The human tumor cell lines used were MDA MB231 (breast adenocarcinoma), HeLa (cervix adenocarcinoma) and TCC-SUP (bladder transitional cell carcinoma). The in vitro antitumor activity of the new benzopsoralens was discussed in terms of structure-activity relationship. Molecular docking studies with human-CYP2A6 enzymes were also carried out with the synthesized compounds in order to evaluate the potential of these compounds to interact with the heme group of the enzymes. The results have demonstrated that the linear compounds have the most pronounced activity against tumor cell lines and this might be related to the better accessibility that these compounds have to the active site in relation to the angular ones that have shown in the majority of the cases multiple binding poses in the active site of CYP2A6.


Assuntos
Antineoplásicos/síntese química , Ficusina/química , Antineoplásicos/química , Antineoplásicos/toxicidade , Hidrocarboneto de Aril Hidroxilases/química , Hidrocarboneto de Aril Hidroxilases/metabolismo , Sítios de Ligação , Domínio Catalítico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Citocromo P-450 CYP2A6 , Ensaios de Seleção de Medicamentos Antitumorais , Ficusina/síntese química , Ficusina/toxicidade , Células HeLa , Humanos , Simulação de Acoplamento Molecular , Relação Estrutura-Atividade
11.
Carcinogenesis ; 34(10): 2218-30, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23715498

RESUMO

Interstrand cross-links (ICLs) are very severe lesions as they are absolute blocks of replication and transcription. This property of interstrand cross-linking agents has been exploited clinically for the treatment of cancers and other diseases. ICLs are repaired in human cells by specialized DNA repair pathways including components of the nucleotide excision repair pathway, double-strand break repair pathway and the Fanconi anemia pathway. In this report, we identify the role of RECQL5, a member of the RecQ family of helicases, in the repair of ICLs. Using laser-directed confocal microscopy, we demonstrate that RECQL5 is recruited to ICLs formed by trioxalen (a psoralen-derived compound) and ultraviolet irradiation A. Using single-cell gel electrophoresis and proliferation assays, we identify the role of RECQL5 in the repair of ICL lesions. The domain of RECQL5 that recruits to the site of ICL was mapped to the KIX region between amino acids 500 and 650. Inhibition of transcription and of topoisomerases did not affect recruitment, which was inhibited by DNA-intercalating agents, suggesting that the DNA structure itself may be responsible for the recruitment of RECQL5 to the sites of ICLs.


Assuntos
Reagentes para Ligações Cruzadas/toxicidade , Dano ao DNA/efeitos dos fármacos , Reparo do DNA/fisiologia , Ficusina/toxicidade , RecQ Helicases/metabolismo , Linhagem Celular , DNA Topoisomerases/metabolismo , Exodesoxirribonucleases/metabolismo , Humanos , Cinética , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , RecQ Helicases/química , Inibidores da Topoisomerase/farmacologia , Transcrição Genética , Helicase da Síndrome de Werner
12.
Methods Mol Biol ; 817: 165-81, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22147573

RESUMO

The single cell gel electrophoresis (SCGE) assay, more commonly known as the comet assay, due to the "comet-like" appearance of the cells, was originally developed as a technique to measure the presence of DNA single-strand breaks. The assay is performed on single cells embedded in agar and placed in an electrical field at alkaline pH, so that fragments of negatively charged single-stranded DNA move through the gel toward the positively charged anode. Undamaged DNA moves relatively slowly, forming the head of the comet, while DNA fragmented due to the presence of single-strand breaks, moves more quickly giving the appearance of the tail. The extent of DNA migration is a measure of the DNA damage present. Since it was first developed, the comet assay has been adapted for measuring other types of DNA damage. The neutral comet assay has been employed for DNA double-strand breaks, while techniques using DNA repair enzymes to cleave specific adducts, UvrABC for ultraviolet radiation induced adducts, for example, have also been described. Here, we describe a modified version of the comet assay for the measurement of interstrand crosslinks (ICLs). Interstrand crosslinking agents include the chemotherapeutic agents mitomycin C and cis-platin, psoralen plus UVA light (PUVA) used to treat hyperproliferative skin disorders and diepoxybutane, a metabolite of 1,3-butadiene used in industrial processes and an environmental pollutant. ICLs are a potent and cytotoxic form of DNA damage as they prevent DNA strand separation, thereby preventing DNA replication. Their removal requires several different DNA repair processes including translesion synthesis and homologous recombination. As ICLs prevent separation of the DNA strands, their presence results in less DNA migration in the comet assay. To successfully measure ICLs, it is necessary to incorporate a step that induces single-strand breaks (using a defined dose of ionizing radiation) that allows the crosslinked DNA to migrate.


Assuntos
Ensaio Cometa/métodos , Dano ao DNA/efeitos dos fármacos , DNA/genética , Mutagênicos/toxicidade , Animais , Linhagem Celular , Reagentes para Ligações Cruzadas/toxicidade , DNA/análise , Dano ao DNA/efeitos da radiação , Compostos de Epóxi/toxicidade , Ficusina/toxicidade , Raios gama , Humanos , Mitomicina/toxicidade , Raios Ultravioleta
13.
Pharm Biol ; 49(4): 422-7, 2011 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21391886

RESUMO

CONTEXT: Psoralens are naturally occurring furanocumarins used in photochemotherapy of several skin diseases. They are obtained from dried ripe fruits of Psoralea corylifolia Linn. (Fabaceae). However, little research has been done to study the melanogenic activity of P. corylifolia seeds and their active ingredients on the pigment cells, the melanophores taking account of their cholinergic activity. OBJECTIVE: The present work was carried out to determine the effects of lyophilized seed extracts of P. corylifolia, along with pure psoralen on the isolated scale melanophores of Channa punctatus Bloch. (Channidae), which are a disguised type of smooth muscle cells and offer excellent in vitro opportunities for studying the effects of drugs. MATERIALS AND METHODS: Effects of lyophilized extracts of P. corylifolia and pure psoralen were studied on the isolated scale melanophores of C. punctatus as per the modified method of Bhattacharya et al. (1976) . RESULTS: The lyophilized extract of P. corylifolia and its active ingredient psoralen caused significant melanin dispersal responses leading to darkening of the fish scale melanophores, which were completely antagonized by atropine and hyoscine. These melanin dispersal effects were also found to be markedly potentiated by neostigmine, an anticholinesterase agent. DISCUSSION: In the present study, the lyophilized extract of P. corylifolia seeds and standard psoralen in different dose ranges induced powerful melanin dispersal effects of the previously adrenaline-aggregated isolated scale melanophores of C. punctatus. Comparatively, psoralen caused a more sustained and powerful melanin dispersal within the isolated fish melanophores and interestingly the concentrations required to achieve maximal dispersion of melanophore were 10 times less than that of lyophilized seed extract of P. corylifolia. The physiologically significant dose-related melanin dispersion effects of lyophilized P. corylifolia seeds and synthetic psoralen per se were found to be completely abolished by atropine and hyoscine, which are specific cholino-muscarinic receptor blockers. These data strongly indicate that in the fish C. punctatus, the dispersion of melanin granules within the scale melanophores is mediated by choline receptors of muscarinic nature. CONCLUSION: It appears that the melanin dispersal effects of the extracts of P. corylifolia and pure psoralen leading to skin darkening are mediated by cholino-muscarinic- or cholino-psoralen-like receptors having similar properties.


Assuntos
Ficusina/farmacologia , Melanóforos/efeitos dos fármacos , Fármacos Fotossensibilizantes/farmacologia , Extratos Vegetais/farmacologia , Óleos Vegetais/farmacologia , Animais , Atropina/farmacologia , Peixes-Gato , Inibidores da Colinesterase/farmacologia , Relação Dose-Resposta a Droga , Fabaceae , Ficusina/toxicidade , Antagonistas Muscarínicos/farmacologia , Neostigmina/farmacologia , Fármacos Fotossensibilizantes/toxicidade , Extratos Vegetais/análise , Extratos Vegetais/toxicidade , Óleos Vegetais/toxicidade , Escopolamina/farmacologia , Sementes
14.
Xenobiotica ; 41(3): 198-211, 2011 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-21070145

RESUMO

1. PAP-1 (5-(4-phenoxybutoxy)psoralen), a potent small-molecule blocker of the voltage-gated potassium Kv1.3 channel, is currently in preclinical development for psoriasis. This study was undertaken to identify the major phase I metabolites of PAP-1 in Sprague-Dawley (SD) rats. 2. Five phase I metabolites, that is 5-(oxybutyric-acid)psoralen (M1), 5-[4-(4-hydroxybutoxy)]psoralen (M2), 5-[4-(4-hydroxyphenoxy)butoxy]psoralen (M3), 5-[4-(3-hydroxyphenoxy)butoxy]psoralen (M4), and 8-hydroxyl-5-(4-phenoxybutoxy)psoralen (M5), were isolated from the bile of rats and identified by mass spectrometry and NMR spectroscopy. The last four metabolites are new compounds. 3. Incubation of PAP-1 with SD rat liver microsomes rendered the same five major metabolites in a nicotinamide adenine dinucleotide phosphate (NADPH)-dependent manner suggesting that cytochrome P450 (CYP) enzymes are involved in PAP-1 metabolism. Inhibitors of rat CYP1A1/2 (alpha-naphthoflavone) and CYP3A (ketoconazole) but not CYP2D6 (quinidine), CYP2E (diethyldithiocarbamate), or CYP2C9 (sulphaphenazole) blocked the metabolism of PAP-1 in rat microsomes. 4. Of the five metabolites M3, M4, and M5 were found to inhibit Kv1.3 currents with nanomolar IC50s, while M1 and M2 were inactive. Our results identified the Kv1.3-inactive M1 as the major phase I metabolite, and suggest that hydroxylation and O-dealkylation are the major pathways of PAP-1 metabolism. 5. We further conducted a 6-month repeat-dose toxicity study with PAP-1 at 50 mg/kg in both male and female Lewis rats and did not observe any toxic effects.


Assuntos
Ficusina/toxicidade , Canal de Potássio Kv1.3/metabolismo , Desentoxicação Metabólica Fase I , Bloqueadores dos Canais de Potássio/toxicidade , Animais , Bile/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Relação Dose-Resposta a Droga , Fezes/química , Ficusina/química , Ficusina/metabolismo , Masculino , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/metabolismo , Proteínas Associadas a Pancreatite , Bloqueadores dos Canais de Potássio/química , Bloqueadores dos Canais de Potássio/metabolismo , Ratos , Ratos Sprague-Dawley , Testes de Toxicidade Crônica
15.
Nucleic Acids Res ; 37(19): 6378-88, 2009 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-19726585

RESUMO

Correction of a defective gene is a promising approach for both basic research and clinical gene therapy. However, the absence of site-specific targeting and the low efficiency of homologous recombination in human cells present barriers to successful gene targeting. In an effort to overcome these barriers, we utilized triplex-forming oligonucleotides (TFOs) conjugated to a DNA interstrand crosslinking (ICL) agent, psoralen (pTFO-ICLs), to improve the gene targeting efficiency at a specific site in DNA. Gene targeting events were monitored by the correction of a deletion on a recipient plasmid with the homologous sequence from a donor plasmid in human cells. The mechanism underlying this event is stimulation of homologous recombination by the pTFO-ICL. We found that pTFO-ICLs are efficient in inducing targeted gene conversion (GC) events in human cells. The deletion size in the recipient plasmid influenced both the recombination frequency and spectrum of recombinants; i.e. plasmids with smaller deletions had a higher frequency and proportion of GC events. The polarity of the pTFO-ICL also had a prominent effect on recombination. Our results suggest that pTFO-ICL induced intermolecular recombination provides an efficient method for targeted gene correction in mammalian cells.


Assuntos
Reagentes para Ligações Cruzadas/toxicidade , Ficusina/toxicidade , Conversão Gênica , DNA/metabolismo , Deleção de Genes , Células HeLa , Humanos , Oligonucleotídeos/metabolismo , Reação em Cadeia da Polimerase , Recombinação Genética
16.
Bioorg Med Chem Lett ; 19(10): 2874-6, 2009 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-19359172

RESUMO

Following our results with benzopsoralens as potent photochemotherapeutic agents, we report the antiproliferative evaluation of nitrogenated isoster upon and without UVA irradiation. The evaluated pyridazinopsoralen showed a higher photochemotherapeutic activity with respect to the well-known drug, 8-MOP, and a significant cytotoxicity, also in the dark. This result enlarges the interest in this tetracyclic psoralen derivative skeleton in the search of new anticancer agents.


Assuntos
Antineoplásicos/química , Ficusina/química , Fármacos Fotossensibilizantes/química , Piridazinas/química , Animais , Antineoplásicos/síntese química , Antineoplásicos/toxicidade , Linhagem Celular , Ficusina/síntese química , Ficusina/toxicidade , Cobaias , Células HeLa , Humanos , Metoxaleno/toxicidade , Fármacos Fotossensibilizantes/síntese química , Piridazinas/síntese química , Piridazinas/toxicidade , Raios Ultravioleta
17.
J Pharm Pharm Sci ; 12(3): 378-87, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-20067712

RESUMO

PURPOSE: To discuss the contribution of psoralen and bergapten metabolites on psoralens toxicity. METHODS: Computational chemistry prediction of metabolic reactions and toxicophoric groups based on the expert systems Derek and Meteor. RESULTS: a total of 15 metabolites were suggested for both psoralen and bergapten based on phase 1 and 2 biotransformations until the 3rd generation. Five toxicophoric substructures were shared among psoralen, bergapten and their corresponding metabolites; one toxicophoric marker (resorcinol) was only identified in bergapten and its biotransformation products. CONCLUSION: Although the toxic effects of psoralens are well known and documented, there is little information concerning the role of their metabolites in this process. We believe this work add to the knowledge of which molecular substructures are relevant to the process of metabolism and toxicity induction, thus guiding the search and development of more effective and less toxic drugs to treat vitiligo.


Assuntos
Sistemas Especialistas , Ficusina/metabolismo , Ficusina/toxicidade , Metoxaleno/análogos & derivados , Fármacos Fotossensibilizantes/metabolismo , Fármacos Fotossensibilizantes/toxicidade , 5-Metoxipsoraleno , Animais , Biotransformação , Biologia Computacional , Simulação por Computador , Descoberta de Drogas/métodos , Humanos , Metoxaleno/metabolismo , Metoxaleno/toxicidade , Vitiligo/tratamento farmacológico
18.
Mol Pharmacol ; 75(3): 599-607, 2009 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-19064630

RESUMO

Psoralen plus UVA light (PUVA) is commonly used to treat psoriasis, a common skin disorder associated with rapid proliferation of cells. PUVA exerts its antiproliferative activity through formation of DNA monoadducts and interstrand cross-links (ICLs). However, this treatment may lead to skin malignancies as a direct result of inducing carcinogenic DNA damage. Inactivation of the p53 tumor suppressor gene is an important event in the development of skin cancer. p53 is rapidly phosphorylated and stabilized in response to DNA damage, and the induction of apoptosis by p53 is an important mechanism by which p53 exerts its tumor-suppressive activity. To better understand the mechanism by which PUVA treatment induces p53, we exposed human skin fibroblasts with PUVA under conditions that differentially produce monoadducts and ICLs and found that psoralen-induced ICLs induced phosphorylation of the Ser-15 site of p53 and apoptosis much more effectively than psoralen-induced monoadducts. The induction of p53 phosphorylation by psoralen ICLs did not require factors believed to be involved in the repair of psoralen ICLs [xeroderma pigmentosum (XP)-A, XP-C, XP-F, Cockayne's syndrome-B, Fanconi anemia] but did require the ataxia-telangiectasia and Rad3-related but not the ataxia-telangiectasia mutated kinase. Psoralen-induced ICLs blocked transcription and replication more efficiently than monoadducts, and induction of p53 and apoptosis correlated with doses causing interference with transcription rather than DNA replication. Our finding that cells underwent apoptosis preferentially during S-phase suggests that the combined blockade of transcription and DNA replication by psoralen ICLs during S-phase elicits a strong apoptotic response.


Assuntos
Ataxia Telangiectasia , Proteínas de Ciclo Celular/fisiologia , Reagentes para Ligações Cruzadas/toxicidade , Dano ao DNA/efeitos dos fármacos , Ficusina/toxicidade , Proteína Supressora de Tumor p53/biossíntese , Ataxia Telangiectasia/induzido quimicamente , Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia , Linhagem Celular Transformada , Dano ao DNA/efeitos da radiação , Fibroblastos/efeitos dos fármacos , Fibroblastos/fisiologia , Humanos , Proteína Supressora de Tumor p53/genética , Raios Ultravioleta/efeitos adversos
19.
PLoS Genet ; 4(9): e1000189, 2008 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-18787700

RESUMO

DNA interstrand crosslinks (ICLs) are among the most toxic types of damage to a cell. For this reason, many ICL-inducing agents are effective therapeutic agents. For example, cisplatin and nitrogen mustards are used for treating cancer and psoralen plus UVA (PUVA) is useful for treating psoriasis. However, repair mechanisms for ICLs in the human genome are not clearly defined. Previously, we have shown that MSH2, the common subunit of the human MutSalpha and MutSbeta mismatch recognition complexes, plays a role in the error-free repair of psoralen ICLs. We hypothesized that MLH1, the common subunit of human MutL complexes, is also involved in the cellular response to psoralen ICLs. Surprisingly, we instead found that MLH1-deficient human cells are more resistant to psoralen ICLs, in contrast to the sensitivity to these lesions displayed by MSH2-deficient cells. Apoptosis was not as efficiently induced by psoralen ICLs in MLH1-deficient cells as in MLH1-proficient cells as determined by caspase-3/7 activity and binding of annexin V. Strikingly, CHK2 phosphorylation was undetectable in MLH1-deficient cells, and phosphorylation of CHK1 was reduced after PUVA treatment, indicating that MLH1 is involved in signaling psoralen ICL-induced checkpoint activation. Psoralen ICLs can result in mutations near the crosslinked sites; however, MLH1 function was not required for the mutagenic repair of these lesions, and so its signaling function appears to have a role in maintaining genomic stability following exposure to ICL-induced DNA damage. Distinguishing the genetic status of MMR-deficient tumors as MSH2-deficient or MLH1-deficient is thus potentially important in predicting the efficacy of treatment with psoralen and perhaps with other ICL-inducing agents.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Dano ao DNA , Reparo do DNA/fisiologia , Genoma Humano , Proteína 2 Homóloga a MutS/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Linhagem Celular Tumoral , Reagentes para Ligações Cruzadas/toxicidade , DNA/metabolismo , Ficusina/toxicidade , Genes cdc/fisiologia , Humanos , Proteína 1 Homóloga a MutL , Mutação , Proteínas Nucleares/genética , Transdução de Sinais/genética
20.
Curr Genet ; 53(6): 361-71, 2008 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-18437386

RESUMO

In order to extend the understanding of the genetical and biochemical basis of photo-activated psoralen-induced DNA repair in the yeast Saccharomyces cerevisiae we have identified and cloned 10 pso mutants. Here, we describe the phenotypic characterization and molecular cloning of the pso10-1 mutant which is highly sensitive to photoactivated psoralens, UV(254) (nm) radiation and the alkylating agent methylmethane sulphonate. The pso10-1 mutant allele also confers a block in the mutagenic response to photoactivated psoralens and UV(254) (nm) radiation, and homoallelic diploids do not sporulate. Molecular cloning using a yeast genomic library, sequence analysis and genetic complementation experiments proved pso10-1 to be a mutant allele of gene MMS21 that encodes a SUMO ligase involved in the sumoylation of several DNA repair proteins. The ORF of pso10-1 contains a single nucleotide C-->T transition at position 758, which leads to a change in amino acid sequence from serine to phenylalanine [S253F]. Pso10-1p defines a leaky mutant phenotype of the essential MMS21 gene, and as member of the Smc5-Smc6 complex, still has some essential functions that allow survival of the mutant. DNA repair via translesion synthesis is severely impaired as the pso10-1 mutant allele confers severely blocked induced forward and reverse mutagenesis and shows epistatic interaction with a rev3Delta mutant allele. By identifying the allelism of PSO10 and MMS21 we demonstrate the need of a fully functional Smc5-Smc6 complex for a WT-like adequate repair of photoactivated psoralen-induced DNA damage in yeast.


Assuntos
Alelos , Dano ao DNA , Reparo do DNA , Ficusina/toxicidade , Proteína SUMO-1/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Sequência de Aminoácidos , Sequência de Bases , Ciclo Celular , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Clonagem Molecular , Reparo do DNA/efeitos dos fármacos , Reparo do DNA/genética , Genes Fúngicos , Dados de Sequência Molecular , Mutação , Proteína SUMO-1/metabolismo , Saccharomyces cerevisiae/enzimologia , Proteínas de Saccharomyces cerevisiae/metabolismo
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