RESUMO
Caffeine exerts a biphasic effect on zebrafish behavior. High doses of caffeine have been associated with increased stress and anxiety, whereas low doses have been found to enhance performance on tasks requiring focus and attention. However, the sex-specific nature of these biphasic effects on behavior and physiology remains unclear. This study assessed the behavioral responses and hormone levels in male and female zebrafish after acute exposure to caffeine ranging from 0.3 to 600â¯mg/L. The results showed no significant difference in caffeine intake between males and females after acute exposure at each concentration. Caffeine-induced behavioral and physiological responses indicated a threshold dosage existed between 30 and 300â¯mg/L. Female fish displayed increased anxiety-like behavioral phenotypes, i.e., latency to upper and freezing, whereas males exhibited more erratic movement following acute exposure to a high-dose treatment. In addition, females exhibited a significant increase in whole-body cortisol levels, while males experienced a testosterone elevation at 300â¯mg/L of caffeine acute exposure. There was a significant decrease in the duration of erratic movements in males treated with the androgen receptor antagonist flutamide compared to the control group. The transcriptome analysis uncovered 511 and 592 up-regulated and 761 and 922 down-regulated differential expression genes in males and females, respectively, compared to the control. The Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) pathway analysis revealed that caffeine has the potential to impact various pathways in zebrafish, including phototransduction and steroid hormone biosynthesis. Our findings demonstrate that testosterone and cortisol play a combined role in regulating stress responses in both behavior and physiology. Furthermore, our study highlights the significance of encompassing both male and female zebrafish as a model system.
Assuntos
Ansiedade , Comportamento Animal , Cafeína , Hidrocortisona , Caracteres Sexuais , Testosterona , Peixe-Zebra , Animais , Cafeína/farmacologia , Cafeína/administração & dosagem , Feminino , Masculino , Ansiedade/induzido quimicamente , Ansiedade/metabolismo , Hidrocortisona/metabolismo , Hidrocortisona/sangue , Testosterona/metabolismo , Testosterona/sangue , Comportamento Animal/efeitos dos fármacos , Estimulantes do Sistema Nervoso Central/farmacologia , Estimulantes do Sistema Nervoso Central/administração & dosagem , Flutamida/farmacologia , Fatores Sexuais , Relação Dose-Resposta a DrogaRESUMO
In brief: Atrazine, like oestrogen, disorganises laminin formation and reduces the number of germ cells and Sertoli cells in the developing testes of the tammar wallaby. This study suggests that interfering with the balance of androgen and oestrogen affects the integrity of laminin structure and testis differentiation. Abstract: The herbicide atrazine was banned in Europe in 2003 due to its endocrine disrupting activity but remains widely used. The integrity of the laminin structure in fetal testis cords requires oestrogen signalling but overexposure to xenoestrogens in the adult can cause testicular dysgenesis. However, whether xenoestrogens affect laminin formation in developing testes has not been investigated. Here we examined the effects of atrazine in the marsupial tammar wallaby during early development and compare it with the effects of the anti-androgen flutamide, oestrogen, and the oestrogen degrader fulvestrant. The tammar, like all marsupials, gives birth to altricial young, allowing direct treatment of the developing young during the male programming window (day 20-40 post partum (pp)). Male pouch young were treated orally with atrazine (5 mg/kg), flutamide (10 mg/kg), 17ß-oestradiol (2.5 mg/kg) and fulvestrant (1 mg/kg) daily from day 20 to 40 pp. Distribution of laminin, vimentin, SOX9 and DDX4, cell proliferation and mRNA expression of SRY, SOX9, AMH, and SF1 were examined in testes at day 50 post partum after the treatment. Direct exposure to atrazine, flutamide, 17ß-oestradiol, and fulvestrant all disorganised laminin but had no effect on vimentin distribution in testes. Atrazine reduced the number of germ cells and Sertoli cells when examined at day 40-50 pp and day 20 to 40 pp, respectively. Both flutamide and fulvestrant reduced the number of germ cells and Sertoli cells. Atrazine also downregulated SRY expression and impaired SOX9 nuclear translocation. Our results demonstrate that atrazine can compromise normal testicular differentiation during the critical male programming window.
Assuntos
Atrazina , Diferenciação Celular , Herbicidas , Laminina , Testículo , Masculino , Animais , Testículo/efeitos dos fármacos , Testículo/metabolismo , Testículo/citologia , Atrazina/farmacologia , Laminina/metabolismo , Diferenciação Celular/efeitos dos fármacos , Herbicidas/farmacologia , Macropodidae/metabolismo , Células de Sertoli/efeitos dos fármacos , Células de Sertoli/metabolismo , Células de Sertoli/citologia , Estrogênios/farmacologia , Estrogênios/metabolismo , Disruptores Endócrinos/farmacologia , Contagem de Células , Antagonistas de Androgênios/farmacologia , Flutamida/farmacologiaRESUMO
Flutamide is frequently used in the management of prostate cancer, hirsutism, and acne. It is a non-steroidal anti-androgenic drug and causes hepatotoxicity. The current study's objective is to evaluate sophorin's hepatoprotective effectiveness against flutamide-induced hepatotoxicity in Wistar rats. Sophorin is a citrus flavonoid glycoside, also known as rutin, which is a low molecular weight polyphenolic compound with natural antioxidant properties and reported to have promising hepatoprotective efficacy. In this study, sophorin was used at a dose of 100 mg/kg body weight in purified water via oral route for 4 week daily whereas, flutamide was used at a dose of 100 mg kg/b.wt for 4 weeks daily in 0.5% carboxy methyl cellulose (CMC) through the oral route for the induction of hepatotoxicity. Flutamide administration leads to enhanced reactive oxygen species (ROS) generation, an imbalance in redox homeostasis and peroxidation of lipid resulted in reduced natural antioxidant level in liver tissue. Our result demonstrated that sophorin significantly abrogate flutamide induced lipid peroxidation, protein carbonyl (PC), and also significantly increasesed in enzymatic activity/level of tissue natural antioxidant such as reduced glutathione(GSH), glutathione reductase(GR), catalase, and superoxide dismutase(SOD). Additionally, sophorin reduced the activity of cytochrome P450 3A1 in liver tissue which was elevated due to flutamide treatment. Furthermore, sophorin treatment significantly decreased the pro-inflammatory cytokines (TNF-α and IL-6) level. Immunohistochemical analysis for the expression of inflammatory proteins (iNOS and COX-2) in hepatic tissue was decreased after sophorin treatment against flutamide-induced hepatotoxicity. Moreover, sophorin suppressed the infiltration of mast cells in liver tissue which further showed anti-inflammatory potential of sophorin. Our histological investigation further demonstrated sophorin's hepatoprotective function by restoring the typical histology of the liver. Based on the aforementioned information, we are able to come to the conclusion that sophorin supplementation might benefit wistar rats with flutamide-induced hepatic damage by reducing oxidative stress and hepatocellular inflammation.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas , Flutamida , Fígado , Ratos Wistar , Animais , Flutamida/farmacologia , Ratos , Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Masculino , Fígado/efeitos dos fármacos , Fígado/patologia , Estresse Oxidativo/efeitos dos fármacos , Antioxidantes/farmacologia , Peroxidação de Lipídeos/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Antagonistas de Androgênios/farmacologiaRESUMO
RATIONALE: Numerous epidemiological studies have reported a link between low testosterone levels and an increased risk of cerebrovascular disease in men. However, there is ongoing controversy surrounding testosterone replacement therapy due to potential side effects. PBMT has been demonstrated to improve cerebrovascular function and promote testosterone synthesis in peripheral tissues. Despite this, the molecular mechanisms that could connect PBMT with testosterone and vascular function in the brain of photothrombosis (PT)-induced stroke rats remain largely unknown. METHODS: We measured behavioral performance, cerebral blood flow (CBF), vascular permeability, and the expression of vascular-associated and apoptotic proteins in PT-induced stroke rats treated with flutamide and seven consecutive days of PBM treatment (350 mW, 808 nM, 2 min/day). To gain further insights into the mechanism of PBM on testosterone synthesis, we used testosterone synthesis inhibitors to study their effects on bEND.3 cells. RESULTS: We showed that PT stroke caused a decrease in cerebrovascular testosterone concentration, which was significantly increased by 7-day PBMT (808 nm, 350 mW/cm2 , 42 J/cm2 ). Furthermore, PBMT significantly increased cerebral blood flow (CBF) and the expression of vascular-associated proteins, while inhibiting vascular permeability and reducing endothelial cell apoptosis. This ultimately mitigated behavioral deficits in PT stroke rats. Notably, treatment with the androgen receptor antagonist flutamide reversed the beneficial effects of PBMT. Cellular experiments confirmed that PBMT inhibited cell apoptosis and increased vascular-associated protein expression in brain endothelial cell line (bEnd.3) subjected to oxygen-glucose deprivation (OGD). However, these effects were inhibited by flutamide. Moreover, mechanistic studies revealed that PBMT-induced testosterone synthesis in bEnd.3 cells was partly mediated by 17ß-hydroxysteroid dehydrogenase 5 (17ß-HSD5). CONCLUSIONS: Our study provides evidence that PBMT attenuates cerebrovascular injury and behavioral deficits associated with testosterone/AR following ischemic stroke. Our findings suggest that PBMT may be a promising alternative approach for managing cerebrovascular diseases.
Assuntos
Terapia com Luz de Baixa Intensidade , Acidente Vascular Cerebral , Humanos , Masculino , Ratos , Camundongos , Animais , Testosterona/metabolismo , Androgênios/metabolismo , Receptores Androgênicos/metabolismo , Células Endoteliais/metabolismo , Flutamida/farmacologia , Flutamida/uso terapêutico , Flutamida/metabolismo , Acidente Vascular Cerebral/terapiaRESUMO
PURPOSE: Polycystic ovarian syndrome (PCOS) is an endocrine-metabolic condition affecting 5-10% of reproductive-aged women and characterized by hyperandrogenism, insulin resistance (IR), and hyperinsulinemia. CFTR is known to be regulated by steroid hormones, and our previous study has demonstrated an essential role of CFTR in ß-cell function. This study aims to investigate the contribution of androgen and CFTR to hypersecretion of insulin in PCOS and the underlying mechanism. METHODS: We established a rat PCOS model by subcutaneously implanting silicon tubing containing Dihydrotestosterone (DHT). Glucose tolerance test with insulin levels was performed at 9 weeks after implantation. A rat ß-cell line RINm5F, a mouse ß-cell line ß-TC-6, and mouse islets were treated with DHT, and with or without the androgen antagonist flutamide for CFTR and insulin secretion-related functional assays or mRNA/protein expression measurement. The effect of CFTR inhibitors on DHT-promoted membrane depolarization, glucose-stimulated intracellular Ca2+ oscillation and insulin secretion were examined by membrane potential imaging, calcium imaging and ELISA, respectively. RESULTS: The DHT-induced PCOS model showed increased body weight, impaired glucose tolerance, and higher blood glucose and insulin levels after glucose stimulation. CFTR was upregulated in islets of PCOS model and DHT-treated cells, which was reversed by flutamide. The androgen receptor (AR) could bind to the CFTR promoter region, which was enhanced by DHT. Furthermore, DHT-induced membrane depolarization, enhanced glucose-stimulated Ca2+ oscillations and insulin secretion, which could be abolished by CFTR inhibitors. CONCLUSIONS: Excessive androgen enhances glucose-stimulating insulin secretion through upregulation of CFTR, which may contribute to hyperinsulinemia in PCOS.
Assuntos
Hiperinsulinismo , Resistência à Insulina , Síndrome do Ovário Policístico , Camundongos , Feminino , Ratos , Humanos , Animais , Adulto , Síndrome do Ovário Policístico/metabolismo , Androgênios/farmacologia , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Flutamida/farmacologia , Regulação para Cima , Resistência à Insulina/fisiologia , Insulina , Di-Hidrotestosterona/farmacologia , Glucose/farmacologiaRESUMO
Ganoderma lucidum polysaccharide is the most widely used complementary therapy in cancer. The present study aims to investigate the possible interaction between Ganoderma lucidum polysaccharide and Docetaxel (a chemotherapy drug) and the first-line medication for prostate cancer treatment (Flutamide) and sensitizing the cells to these treatments. The cytotoxic effects of Ganoderma lucidum polysaccharide in combination with Docetaxel and Flutamide on prostate cancer cells were investigated by the MTT test, Hoechst staining, and flow cytometry. In addition, the expression of genes related to apoptosis, angiogenesis, Epithelial-Mesenchymal Transition pathway (EMT), and prostate cancer biomarkers by Real-Time PCR was investigated. The results demonstrated that IC50 values for Ganoderma lucidum polysaccharide (30 µM and 20 µM), Docetaxel (10 µM and 5 µM), and Flutamide (20 µM and 12 µM) with MTT were confirmed by flow cytometry in a dose and time-dependent manner. Regarding the high efficacy of Ganoderma lucidum polysaccharide in combination with Flutamide and Docetaxel, 10 µM and 5 µM Flutamide were used instead of 20 µM and 12 µM and 5 µM and 2 µM Docetaxel was used instead of 10 µM and 5 µM in PC3 and LNCap, respectively. Moreover, for the first time, it was shown that Ganoderma lucidum polysaccharide alone and in combination with Docetaxel and Flutamide significantly augmented apoptosis, reduced cell migration and colonization, and downregulated expression of KLK2 and EMT pathway genes in both PC3 and LNCap cell line (P < 0.01). Ganoderma lucidum polysaccharide synergistically increased the effect of Docetaxel and Flutamide and increased the sensitivity of the prostate cancer cell lines to these drugs. Therefore, it may provide a new therapeutic strategy against prostate cancer.
Assuntos
Neoplasias da Próstata , Reishi , Masculino , Humanos , Docetaxel/farmacologia , Docetaxel/uso terapêutico , Próstata/metabolismo , Flutamida/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/metabolismo , Linhagem Celular Tumoral , Polissacarídeos/farmacologia , Polissacarídeos/uso terapêuticoRESUMO
In this study, we examined the metabolic adaptations of a chemoresistant prostate cancer cell line in comparison to a sensitive cell line. We utilized prostate cancer LNCaP cells and subjected them to a stepwise increase in the antiandrogen 2-hydroxy-flutamide (FLU) concentration to generate a FLU-resistant cell line (LN-FLU). These LN-FLU cells displayed characteristics of cancer stem cells, exhibited drug resistance, and showed a significantly reduced expression of Cyclin D1, along with the overexpression of p16, pointing to a proliferation arrest. In comparing the cancer stem-like LN-FLU cells to the LNCaP cells, we observed a decrease in the expression of CTP-choline cytidylyl transferase α (CCTα), as well as a decline in choline kinase, suggesting altogether a downregulation of the phosphatidylcholine biosynthetic pathway. In addition, we found decreased levels of the protein methyl transferase PRMT2 and the upregulation of the histone deacetylase Sirtuin1 (Sirt1). Analysis of the human prostate cancer samples revealed similar results in a population with high expressions of the stem cell markers Oct4 and ABCB1A1. Our findings suggest that the adaptation of prostate cancer cells to antiandrogens could induce reprogramming into stem cells that survive in a low phosphocholine metabolism and cell cycle arrest and display drug resistance.
Assuntos
Flutamida , Neoplasias da Próstata , Masculino , Humanos , Flutamida/farmacologia , Regulação para Baixo , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Antagonistas de Androgênios/farmacologia , Linhagem Celular Tumoral , Transferases/metabolismoRESUMO
Because a wide range of environmental contaminants are known to cause endocrine disorders in humans and animals, in vivo tests are needed to identify such endocrine disrupting chemicals (EDCs) and to assess their biological effects. Despite the lack of a standardized guideline, the avian embryo has been shown to be a promising model system which responds sensitively to EDCs. After previous studies on the effects of estrogenic, antiestrogenic and androgenic substances, the present work focuses on the effects of in ovo exposure to p,p'-DDE, flutamide and cyproterone acetate (CPA) as antiandrogenic model compounds regarding gonadal sex differentiation and embryonic development of the domestic fowl (Gallus gallus domesticus). The substances were injected into the yolk of fertilized eggs on embryonic day one. On embryonic day 19 sex genotype and phenotype were determined, followed by gross morphological and histological examination of the gonads. Treatment with flutamide (0.5, 5, 50 µg/g egg), p,p'-DDE (0.5, 5, 50 µg/g egg) or CPA (0.2, 2, 20 µg/g egg) did not affect male or female gonad development, assessed by gonad surface area and cortex thickness in both sexes and by the percentage of seminiferous tubules in males as endpoints. This leads to the conclusion that antiandrogens do not affect sexual differentiation during embryonic development of G. gallus domesticus, reflecting that gonads are not target organs for androgens in birds. In ovo exposure to 2 and 20 µg CPA/g egg, however, resulted in significantly smaller embryos as displayed by shortened lengths of skull, ulna and tarsometatarsus. Although gonadal endpoints were not affected by antiandrogens, the embryo of G. gallus domesticus is shown to be a suitable test system for the identification of substance-related mortality and developmental delays.
Assuntos
Antagonistas de Androgênios , Flutamida , Animais , Humanos , Masculino , Feminino , Antagonistas de Androgênios/efeitos adversos , Flutamida/farmacologia , Acetato de Ciproterona/efeitos adversos , Galinhas , Diclorodifenil Dicloroetileno/farmacologia , Diferenciação Sexual , Aves Domésticas , Androgênios/efeitos adversos , Desenvolvimento EmbrionárioRESUMO
In males, skeletal muscle function may be altered by shifts in either circulating testosterone or estrogen. We examined the effect of acute (2-week) exposures to 17α-ethinyl estradiol (EE2), an estrogen receptor (ER) agonist, or flutamide, an androgen receptor (AR) antagonist, on the contractile function of individual skeletal muscle fibers from slow-contracting soleus and fast-contracting extensor digitorum longus muscles from adult male mice. Single fiber specific tension (force divided by cross-sectional area) was decreased with flutamide treatment in all myosin heavy chain (MHC) fiber types examined (I, IIA, and IIB); similar effects were observed with EE2 treatment but only in the fastest-contracting MHC IIB fibers. The decreases in maximally Ca2+-activated specific tension were primarily a result of fewer strongly bound myosin-actin cross-bridges, with flutamide treatment also showing lower myofilament lattice stiffness. Myosin-actin cross-bridge kinetics were slower in MHC IIA fibers in flutamide-treated mice, but faster in EE2-treated mice, indicating that contractile velocity may be affected differently in this fiber type, which is commonly expressed in human skeletal muscle. Importantly, these effects were observed in the absence of outcomes previously used to evaluate ER agonists or AR antagonists in rodents including weight of reproductive organs or mammary gland morphology. Our findings indicate that substantial shifts in skeletal muscle function occur in male mice following acute exposures to low doses of a pharmacological ER agonist and an AR antagonist. These results suggest that countermeasures to maintain physical function may be needed early in situations that induce similar ER agonist and AR antagonist conditions.
Assuntos
Actinas , Antagonistas de Receptores de Andrógenos , Adulto , Humanos , Masculino , Animais , Camundongos , Flutamida/farmacologia , Músculo Esquelético , EstrogêniosRESUMO
Background: Evidence supports an important link between mitochondrial DNA (mtDNA) variation and adverse drug reactions such as idiosyncratic drug-induced liver injury (iDILI). Here, we describe the generation of HepG2-derived transmitochondrial cybrids, to investigate the impact of mtDNA variation on mitochondrial (dys)function and susceptibility to iDILI. This study created 10 cybrid cell lines, each containing distinct mitochondrial genotypes of haplogroup H or haplogroup J backgrounds. Methods: HepG2 cells were depleted of mtDNA to make rho zero cells, before the introduction of known mitochondrial genotypes using platelets from healthy volunteers (n=10), thus generating 10 transmitochondrial cybrid cell lines. The mitochondrial function of each was assessed at basal state and following treatment with compounds associated with iDILI; flutamide, 2-hydroxyflutamide, and tolcapone, and their less toxic counterparts bicalutamide and entacapone utilizing ATP assays and extracellular flux analysis. Results: Whilst only slight variations in basal mitochondrial function were observed between haplogroups H and J, haplogroup-specific responses were observed to the mitotoxic drugs. Haplogroup J showed increased susceptibility to inhibition by flutamide, 2-hydroxyflutamide, and tolcapone, via effects on selected mitochondrial complexes (I and II), and an uncoupling of the respiratory chain. Conclusions: This study demonstrates that HepG2 transmitochondrial cybrids can be created to contain the mitochondrial genotype of any individual of interest. This provides a practical and reproducible system to investigate the cellular consequences of variation in the mitochondrial genome, against a constant nuclear background. Additionally, the results show that inter-individual variation in mitochondrial haplogroup may be a factor in determining sensitivity to mitochondrial toxicants. Funding: This work was supported by the Centre for Drug Safety Science supported by the Medical Research Council, United Kingdom (Grant Number G0700654); and GlaxoSmithKline as part of an MRC-CASE studentship (grant number MR/L006758/1).
Assuntos
Doença Hepática Induzida por Substâncias e Drogas , Flutamida , Humanos , Flutamida/metabolismo , Flutamida/farmacologia , Tolcapona/metabolismo , Tolcapona/farmacologia , Haplótipos , Mitocôndrias/metabolismo , DNA Mitocondrial/genética , DNA Mitocondrial/metabolismo , Genótipo , Doença Hepática Induzida por Substâncias e Drogas/genética , Doença Hepática Induzida por Substâncias e Drogas/metabolismoRESUMO
Flutamide is an antagonist of testosterone, an essential hormone in male reproduction. However, the use of flutamide as a contraceptive agent for nonsurgical castration in veterinary practice remains challenging due to its poor bioavailability. Here, the flutamide-loaded nanostructure lipid carrier (FLT-NLC) was synthesized, and its biological effects were demonstrated by an in vitro blood-testis barrier model. The flutamide was incorporated into the nanostructure lipid carrier by a homogenization method resulting in a high encapsulation efficiency (99.7 ± 0.04%). The FLT-NLC was negatively charged (-27.90 ± 0.10 mV), with a nano size (182.13 ± 0.47 nm) and narrow dispersity index (0.17 ± 0.01). An in vitro release study demonstrated a slower release profile of FLT-NLC when compared with flutamide solution (FLT). The FLT-NLC at doses up to 50 µM showed no significant cytotoxic effects against mouse Sertoli cells (TM4) or mouse fibroblast cells (NIH/3T3) (p > 0.05). An in vitro blood-testis barrier with FLT-NLC demonstrated remarkable lower transepithelial electrical resistance when compared with those lacking FLT-NLC (p < 0.01). Moreover, FLT-NLC significantly decreased the mRNA expression of blood-testis barrier proteins, CLDN11 and OCLN. In conclusion, we successfully synthesized FLT-NLC and confirmed its potential antifertility effects on in vitro blood-testis barrier, thus indicating its possible application as nonsurgical contraception for male animals.
Assuntos
Flutamida , Nanoestruturas , Camundongos , Masculino , Animais , Flutamida/farmacologia , Portadores de Fármacos/química , Anticoncepcionais , Barreira Hematotesticular , Nanoestruturas/química , Lipídeos/químicaRESUMO
Although aryl hydrocarbon receptors (AhRs) are related to the metabolic pathway of xenobiotics, recent studies have revealed that this receptor is also associated with the life cycle of viruses and inflammatory reactions. For example, flutamide, used to treat prostate cancer, inhibits hepatitis C virus proliferation by acting as an AhR antagonist, and methylated-pelargonidin, an AhR agonist, suppresses pro-inflammatory cytokine production. To discover a novel class of AhR ligands, we screened 1000 compounds derived from fungal metabolites using a reporter assay and identified methylsulochrin as a partial agonist of the aryl hydrocarbon receptor. Methylsulochrin was found to inhibit the production of hepatitis C virus (HCV) in Huh-7.5.1 cells. Methylsulochrin also suppressed the production of interleukin-6 in RAW264.7 cells. Furthermore, a preliminary structure-activity relationship study using sulochrin derivatives was performed. Our findings suggest the use of methylsulochrin derivatives as anti-HCV compounds with anti-inflammatory activity.
Assuntos
Antivirais , Receptores de Hidrocarboneto Arílico , Masculino , Humanos , Receptores de Hidrocarboneto Arílico/agonistas , Receptores de Hidrocarboneto Arílico/metabolismo , Antivirais/farmacologia , Flutamida/farmacologia , Anti-Inflamatórios/farmacologia , LigantesRESUMO
AIM: this study aims to explore the effect of androgen receptor (AR) blockade by flutamide on some renal pathologic changes such as inflammation, apoptosis, and fibrosis in male rats. MAIN METHODS: Firstly, we investigated the potential effect of AR blockade on renal inflammatory intermediates including IL-1ß, IL-6, TNF-α, NF-Òß proteins, and the renal gene expression of NF-Òß. Besides inflammation, we also assessed the apoptosis pathways including the caspases 3 & 9, mTOR, pAKT proteins, and BAX gene expression. Besides inflammation and apoptosis pathways, we also investigated the effect of androgen blockade on renal fibrosis intermediates including vimentin, TGFß-1, α-SMA, MMP-9, collagen type-III, collagen type-IV, and the renal expression of the col1A1 gene. Besides previous pathological pathways, we assessed the expression of chloride channel protein-5 (ClC-5), as an important regulator of many renal pathological changes. Finally, we assessed the impact of previous pathological changes on renal function at biochemical and pathological levels. KEY FINDINGS: We found that AR blockade by flutamide was associated with the down-regulation of renal inflammation, apoptosis, and fibrosis markers. It was associated with expression down-regulation of IL-1ß & IL-6, TNF-α, NF-Òß, caspases 3 & 9, mTOR, MMP-9, collagens, TGFß-1, and α-SMA. Away from down-regulation, we also found that AR blockade has upregulated ClC-5 and pAKT proteins. SIGNIFICANCE: AR is a major player in androgens-induced nephrotoxicity. AR blockade downregulates renal fibrosis, inflammation, and apoptosis pathways. It may be helpful as a strategy for alleviation of renal side effects associated with some drugs. However; this needs further investigations.
Assuntos
Flutamida , Nefropatias , Ratos , Masculino , Animais , Flutamida/farmacologia , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Interleucina-6/farmacologia , NF-kappa B/metabolismo , Androgênios/farmacologia , Fibrose , Apoptose , Serina-Treonina Quinases TOR , Inflamação/tratamento farmacológico , CaspasesRESUMO
With the objective to develop a potential 99mTc radiopharmaceutical for imaging the androgen receptor (AR) in prostate cancer, four ligands bearing the same pharmacophore derived from the AR antagonist flutamide were prepared, labeled with 99mTc, and their structures corroborated via comparison with the corresponding stable rhenium analogs. All complexes were obtained with high radiochemical purity. Three of the complexes were highly stable, and, due to their favorable physicochemical properties, were further evaluated using AR-positive and AR-negative cells in culture. All complexes exhibited considerable uptake in AR-positive cells, which could be blocked by an excess of flutamide. The efflux from the cells was moderate. They also showed significantly lower uptakes in AR-negative cells, indicating interactions with the AR receptor. However, the binding affinities were considerably reduced by the coordination to 99mTc, and the complex that exhibited the best biological behavior did not show sufficient specificity towards AR-positive cells.
Assuntos
Flutamida , Receptores Androgênicos , Masculino , Humanos , Flutamida/farmacologia , Diagnóstico por Imagem , Compostos Radiofarmacêuticos/química , Tecnécio/química , Compostos de Organotecnécio/químicaRESUMO
ABSTRACT: Chronic pain is a significant health problem associated with disability and reduced quality of life. Current management of chronic pain is inadequate with only modest effects of pharmacological interventions. Thus, there is a need for the generation of analgesics for treating chronic pain. Although preclinical and clinical studies demonstrate the analgesic effects of testosterone, clinical use of testosterone is limited by adverse androgenic effects. Selective androgen receptor modulators (SARMs) activate androgen receptors and overcome treatment limitations by minimizing androgenic side effects. Thus, we tested whether daily soluble SARMs or a SARM-loaded microparticle formulation alleviated muscle hyperalgesia in a mouse-model of widespread pain (male and female C57BL/6J mice). We tested whether the analgesic effects of the SARM-loaded microparticle formulation was mediated through androgen receptors by blocking androgen receptors with flutamide pellets. In vitro and in vivo release kinetics were determined for SARM-loaded microparticles. Safety and toxicity of SARM treatment was determined using serum cardiac and liver toxicity panels, heart histology, and conditioned place preference testing. Subcutaneous daily SARM administration, and 2 injections, 1 week apart, of SARM-loaded microparticles alleviated muscle hyperalgesia in both sexes and was prevented with flutamide treatment. Sustained release of SARM, from the microparticle formulation, was observed both in vitro and in vivo for 4 weeks. Selective androgen receptor modulator treatment produced no cardiac or liver toxicity and did not produce rewarding behaviors. These studies demonstrate that SARM-loaded microparticles, which release drug for a sustained period, alleviate muscle pain, are safe, and may serve as a potential therapeutic for chronic muscle pain.
Assuntos
Dor Crônica , Receptores Androgênicos , Camundongos , Animais , Masculino , Feminino , Flutamida/farmacologia , Flutamida/uso terapêutico , Mialgia/induzido quimicamente , Mialgia/tratamento farmacológico , Hiperalgesia/tratamento farmacológico , Qualidade de Vida , Camundongos Endogâmicos C57BL , Músculos , Testosterona , Androgênios/farmacologia , Androgênios/uso terapêuticoRESUMO
BACKGROUND: The aim of this study was to evaluate modifications in proteoglycan morphology and composition in the prostatic stroma of 18-month-old gerbils after surgical castration, in association or not with an androgenic blockade. METHODS: The animals (n = 5) were sorted into groups subjected or not to antiandrogen treatment (flutamide 10 mg/kg/day) administered for the total postsurgery period and euthanized at 7- or 30-day postcastration; the control group consisted of intact animals. Tissue analysis included immunohistochemical assessment (perlecan and chondroitin sulfate) and proteoglycan morphology was analyzed by transmission electron microscopy. RESULTS: Chondroitin sulfate frequency was increased 7 days postcastration with an androgenic blockade. The presence of these carbohydrates was rare after 30 days of androgenic blockade treatment. There was a significant increase in the amount of perlecan in the prostate stroma from groups subjected to castration plus flutamide for 7 or 30 days. Ultrastructural analysis showed that the incidence of areas occupied by proteoglycans and basement membrane was altered by treatment. In addition, androgenic blockade results in changes in the amount, thickness, and morphology of these structures. At 30 days postcastration, with or without flutamide treatment, larger proteoglycans were common. CONCLUSIONS: In this study, in particular, the decrease in chondroitin sulfate after the longer period might be understood as a prostatic response to androgenic deprivation, while the high frequency and permanence of perlecan led to the assumption that its modulation could be androgen-independent. Length and form alterations in proteoglycans as well as associations among them and with the basement membrane were dynamic events in the prostate microenvironment.
Assuntos
Flutamida , Próstata , Masculino , Animais , Flutamida/farmacologia , Gerbillinae , Androgênios/farmacologia , Sulfatos de Condroitina/farmacologia , OrquiectomiaRESUMO
BACKGROUND: Reduced androgen action during early fetal development has been suggested as the origin of reproductive disorders comprised within the testicular dysgenesis syndrome (TDS). This hypothesis has been supported by studies in rats demonstrating that normal male development and adult reproductive function depend on sufficient androgen exposure during a sensitive fetal period, called the masculinization programming window (MPW). The main aim of this study was therefore to examine the effects of manipulating androgen production during different timepoints during early human fetal testis development to identify the existence and timing of a possible window of androgen sensitivity resembling the MPW in rats. METHODS: The effects of experimentally reduced androgen exposure during different periods of human fetal testis development and function were examined using an established and validated human ex vivo tissue culture model. The androgen production was reduced by treatment with ketoconazole and validated by treatment with flutamide which blocks the androgen receptor. Testicular hormone production ex vivo was measured by liquid chromatography-tandem mass spectrometry or ELISA assays, and selected protein markers were assessed by immunohistochemistry. RESULTS: Ketoconazole reduced androgen production in testes from gestational weeks (GW) 7-21, which were subsequently divided into four age groups: GW 7-10, 10-12, 12-16 and 16-21. Additionally, reduced secretion of testicular hormones INSL3, AMH and Inhibin B was observed, but only in the age groups GW 7-10 and 10-12, while a decrease in the total density of germ cells and OCT4+ gonocytes was found in the GW 7-10 age group. Flutamide treatment in specimens aged GW 7-12 did not alter androgen production, but the secretion of INSL3, AMH and Inhibin B was reduced, and a reduced number of pre-spermatogonia was observed. CONCLUSIONS: This study showed that reduced androgen action during early development affects the function and density of several cell types in the human fetal testis, with similar effects observed after ketoconazole and flutamide treatment. The effects were only observed within the GW 7-14 period-thereby indicating the presence of a window of androgen sensitivity in the human fetal testis.
Assuntos
Hormônios Testiculares , Testículo , Humanos , Masculino , Androgênios/farmacologia , Androgênios/metabolismo , Flutamida/farmacologia , Flutamida/metabolismo , Cetoconazol/metabolismo , Cetoconazol/farmacologia , Receptores Androgênicos/metabolismo , Hormônios Testiculares/metabolismo , Hormônios Testiculares/farmacologia , Testosterona/farmacologiaRESUMO
In this study, molecular dynamics simulation is applied to investigate drug transport in both pure state and conjugated with neutral gold nanoparticle (AuNP) as a drug carrier inside dipalmitoylphosphatidylcholine (DPPC) membrane. Flutamide (Flu) as a hydrophobic and Glutathione (GSH) as a hydrophilic anticancer drug are selected as the case studies. Dynamics of each drug including adhesion on and penetration into the cell membrane are investigated. Pure and conjugated form of drugs inside the water and near the membrane are studied. Simulation results show that the interaction between drug molecules and DPPC changes after drug conjugating with AuNP. GSH, as a hydrophilic drug, intends to remain above the membrane bilayer and after conjugating with AuNP diffuses inside DPPC. However, hydrophobic Flu molecule likes to diffuse inside DPPC, but after conjugating with AuNP, its diffusion inside the lipid bilayer decreases, and its retention time at the surface of DPPC increases. Presence of Flu-NP at the surface of DPPC could enhance its impact on blocking dihydrotestosterone binding at androgen receptors resulting in tumor cell growth arrest. In addition, the tendency of GSH-NP for diffusion to the DPPC is a positive factor for the successful transport of heavy metals such as AuNP without rapid clearance through either the hepatobiliary pathway or the renal system. In conclusion, such MD simulation results may solve problems in nanomedicine translation and turn into a bridge toward maximizing targeting and minimizing nanotoxicity of metal NPs.
Assuntos
Ouro , Nanopartículas Metálicas , 1,2-Dipalmitoilfosfatidilcolina/química , Membrana Celular/química , Portadores de Fármacos , Flutamida/análise , Flutamida/farmacologia , Glutationa/análise , Ouro/química , Bicamadas Lipídicas/química , Nanopartículas Metálicas/química , Simulação de Dinâmica MolecularRESUMO
Melatonin (MEL) is involved in homeostasis of the epididymis lumen environment. Dihydrotestosterone (DHT) partakes in the development of gonads and organs in male animals. However, whether MEL secretion, the expression of its receptors, MT1 and MT2, and sheep epididymal epithelial cell apoptosis is regulated by DHT remains unclear. In this study, we used immunohistochemical staining to detect the distribution patterns of DHT synthetases [5α-reductase (5α-red)] and its androgen receptor (AR) in sheep epididymides. 5α-red1, 5α-red2 and AR were positively expressed in sperm, epididymal epithelial cells, and the smooth muscle cells of the caput, corpus and cauda regions of the epididymis. DHT concentration and the expression levels of 5α-red and AR in the caput, corpus and cauda regions were measured by enzyme-linked immunosorbent assay, liquid chromatography-mass spectrometry, real-time quantitative polymerase chain reaction and western blot analysis. DHT concentration in the caput was significantly higher than those in corpus and cauda, probably because of the high expression of 5α-red2 in the caput and secretion and transport of DHT by the testicles. DHT inhibited MEL secretion, the expression of its membrane receptors and MEL synthetases in cultured sheep epididymal epithelial cells in vitro. In addition, the Bax/Bcl-2 ratio, ACT CASP3 and caspase-3 mRNA expression were also decreased. The decreasing effect was partially reversed after flutamide treatment. Therefore, DHT regulates sheep epididymal function by influencing MEL expression and apoptosis-related factors. This study provides basic data for further research on the reproductive physiology of male animals.
Assuntos
Epididimo , Melatonina , Animais , Apoptose , Caspase 3/metabolismo , Di-Hidrotestosterona/metabolismo , Di-Hidrotestosterona/farmacologia , Epididimo/metabolismo , Flutamida/metabolismo , Flutamida/farmacologia , Ligases/metabolismo , Ligases/farmacologia , Masculino , Melatonina/metabolismo , Melatonina/farmacologia , RNA Mensageiro/metabolismo , Receptores Androgênicos/genética , Receptores de Melatonina/metabolismo , Sêmen/química , Ovinos , Proteína X Associada a bcl-2/metabolismoRESUMO
RESEARCH QUESTION: Polycystic ovary syndrome (PCOS) alters oocyte developmental competence and so the treatment of PCOS patients with assisted reproductive techniques is a major challenge for an infertility specialist. Does ovulation induction therapy with clomiphene citrate, metformin and flutamide affect abnormal PCOS follicle gene expression, its quality, and nuclear and cytoplasmic maturation in the oocyte matured in vitro? DESIGN: Fifty NMRI mice (7-8 weeks old) were randomly divided into five groups, including non-PCOS, PCOS and PCOS groups treated with clomiphene citrate (18 mg/kg body weight for 2 days), metformin (50 mg/100 g body weight for 30 days) and flutamide (10 mg/kg body weight injection for 15 days). The oocytes were collected for quantitative real-time polymerase chain reaction analysis (the evaluation of genes associated with oocyte maturation), transmission electron microscopy (the evaluation of cytoplasmic maturation) and in-vitro maturation (IVM) and IVF outcomes. RESULTS: The dysregulated expression of some genes of insulin-like growth factor (IGF) families involved in oocyte competence and steroid metabolism (e.g. Cyp19a1 and Cyp11a1), transforming growth factor beta (TGF-ß) family (e.g. Tgfb1, Gdf9, Bmp15, Inhbb and Bmpr1a), and oestrogen receptor signalling (e.g. Ncoa3, Pgr) was, to some extent, restored in the PCOS follicles following their with clomiphene citrate, metformin and flutamide. The improved cytoplasmic maturation was observed particularly in the PCOS mice treated with clomiphene citrate. The better IVM-IVF outcomes were also observed in the clomiphene citrate and metformin groups compared with the PCOS group. CONCLUSIONS: This study opens up a new perspective on knowing the effect of ovulation induction therapy on not only nuclear maturation but also ultrastructural characteristics, IVM-IVF outcomes and PCOS oocyte developmental competence.