RESUMEN

This study investigated the effects of cyclic adenosine monophosphate modulating during cumulus-oocyte complexes (COCs) pre-maturation and the role of melatonin on in vitro maturation (IVM) of bovine COCs. In experiment one, COCs were pre-matured for 8 h in control medium or with 3-isobutyl-1-methylxanthine (IBMX) and forskolin, IBMX and C-type natriuretic peptide, c-type natriuretic peptide and forskolin or IBMX, forskolin and c-type natriuretic peptide. Then, meiotic progression was evaluated. In experiment two, COCs were pre-matured, followed by IVM in control medium alone or with 10-6, 10-7 or 10-8 M melatonin. After IVM, chromatin configuration, transzonal projections (TZPs), reactive oxygen species, mitochondrial distribution, ultrastructure and mRNA expression for antioxidant enzymes were evaluated. In experiment 1, COCs pre-matured with both C-type natriuretic peptide and forskolin or C-type natriuretic peptide, forskolin and IBMX had lower meiotic resumption rate when compared to control. Considering that IBMX had not an additional effect to potentiate inhibition of meiotic resumption, a combination of C-type natriuretic peptide and forskolin was chosen. In experiment 2, COCs matured with 10-8 M melatonin had greater rates of meiotic resumption when compared to the other treatments (P < 0.05). The COCs matured with 10-7 or 10-8 M melatonin had greater mitochondrial activity (P < 0.05), while those matured with 10-6 or 10-8 M of melatonin had greater levels of TZPs. Ultrastructure of oocyte and cumulus cells after IVM with melatonin was relatively well preserved. COCs matured with 10-8 M melatonin increased mRNA expression for superoxide dismutase (SOD) and catalase (CAT) (P < 0.05), when compared to non-cultured and pre-matured COCs, respectively. In conclusion, bovine COC pre-maturation with C-type natriuretic peptide and forskolin, followed by IVM with 10-8 M melatonin improves meiotic resumption rates, TZPs, mitochondrial distribution and mRNA expression for SOD and CAT.

Asunto(s)

Melatonina , Animales , Bovinos , Femenino , Melatonina/farmacología , Melatonina/metabolismo , 1-Metil-3-Isobutilxantina/farmacología , Técnicas de Maduración In Vitro de los Oocitos/veterinaria , Péptido Natriurético Tipo-C/farmacología , Colforsina/farmacología , Colforsina/metabolismo , Oocitos/fisiología , AMP Cíclico/metabolismo , ARN Mensajero/metabolismo , Superóxido Dismutasa/metabolismo , Adenosina Monofosfato/metabolismo , Adenosina Monofosfato/farmacología , Células del Cúmulo

RESUMEN

This study aimed to investigate the effects of different concentrations of N-acetylcysteine (NAC) on the growth, antrum formation, viability, and ultrastructure of bovine secondary follicles cultured in vitro for 18 days. To this end, the follicles were cultured in TCM-199+ medium alone or supplemented with 1.0, 5.0, or 25.0 mM NAC. Follicular growth, antrum formation, viability (calcein-AM and ethidium homodimer-1) and ultrastructure were evaluated at the end of culture period. The results showed that 1.0 mM NAC increased the percentage of growing follicles and the fluorescence intensity for calcein-AM when compared to other treatments (p < 0.05). On the other hand, follicles cultured with 25.0 mM NAC had higher fluorescence intensity for ethidium homodimer-1, which is a sign of degeneration. Ultrastructural analysis showed that oocytes from follicles cultured in control medium alone or with 1 mM NAC had intact zonae pellucidae in close association with oolemmae, but the ooplasm showed mitochondria with a reduced number of cristae. On the other hand, oocytes from follicles cultured with 5 or 25 mM NAC had extremely vacuolated cytoplasm and no recognizable organelles. In conclusion, 1 mM NAC increases cytoplasmic calcein staining and the growth rate in bovine secondary follicles cultured in vitro, but the presence of 5 or 25 mM NAC causes damage in cellular membranes and organelles, as well as reducing the percentages of growing follicles.

RESUMEN

The effect of L-165041 (PPARδ-agonist) on decreasing apoptosis and intracellular lipid content was assessed in fresh and vitrified-warmed in vitro -produced bovine embryos. It was hypothesised that the addition of L-165041 to the culture medium enhances development and cryopreservation. Oocytes were allocated to one of two treatments: control-standard culture medium, or L-165041 added to the medium on day1 with no media change. Ultrastructure, cleavage, and blastocyst rates were evaluated in fresh, and in post-vitrification cultured embryos by optical and electronic microscopy. A subset of fresh embryos were fixed for TUNEL assay and for Sudan-Black-B histochemical staining. Vitrified-warmed embryos were assessed using MALDI-MS technique. Cleavage and blastocyst rates (control 49.4±5.2, L-165041 51.8±4.3) were not influenced by L-165041. The proportion of inner cell mass cells (ICM) was higher in fresh embryos, and the rate of total and ICM apoptosis was lower in L-165041. In warmed-embryos, total and ICM apoptosis was lower in L-165041. The overall hatching rate was higher in L-165041 (66.62±2.83% vs 53.19±2.90%). There was less lipid accumulation in fresh L-165041-embryos. In conclusion, the use of L-165041 is recommended to improve the viability of in vitro -derived bovine embryos.

Asunto(s)

PPAR delta , Vitrificación , Animales , Blastocisto , Bovinos , Criopreservación/métodos , Criopreservación/veterinaria , Medios de Cultivo , Técnicas de Cultivo de Embriones/métodos , Técnicas de Cultivo de Embriones/veterinaria , Desarrollo Embrionario , Lípidos/farmacología , Fenoxiacetatos

RESUMEN

This study evaluated the potential of Cimicifuga racemosa (L.) Nutt extract (CIMI) to reduce the deleterious effects of doxorubicin (DOXO) in oocytes, follicles and stromal cells in mice ovaries cultured in vitro. In experiment 1, mice ovaries were cultured in DMEM+ alone or supplemented with 5, 50 or 500 ng/mL CIMI, while in experiment 2, mice ovaries were cultured in DMEM+ alone or supplemented with 5 ng/mL CIMI (better concentration), 0.3 µg/mL DOXO or both. Thereafter, the ovaries were processed for histological (morphology, growth, activation, extracellular matrix configuration and stromal cell density), immunohistochemical (caspase-3) analyses. Follicle viability was evaluated by fluorescence microscopy (ethidium homodimer-1 and calcein) while real-time PCR was performed to analyses the levels of (mRNA for SOD, CAT and nuclear factor erythroid 2-related factor 2 (NRF2) analyses. The results showed that DOXO reduces the percentage of normal follicles and the density of stromal cells in cultured ovaries, but these harmful effects were blocked by CIMI. The DOXO reduced the percentage of primordial follicles, while the presence of CIMI alone did not influence percentage of primordial follicles. A higher staining for caspase-3 was seen in ovaries cultured in control medium alone or with DOXO when compared with those cultured with CIMI alone or both CIMI and DOXO. In addition, follicles from ovaries cultured with both CIMI and DOXO were stained by calcein, while those follicles cultured with only DOXO were stained with ethidium homodimer-1. Furthermore, ovaries cultured with CIMI or both CIMI and DOXO had higher levels of mRNA for SOD and CAT, respectively, than those cultured with only DOXO. In conclusion, the extract of CIMI protects the ovaries against deleterious effects of DOXO on follicular survival and ovarian stromal cells.

RESUMEN

BACKGROUND: In this article, a series of 20 new thiosemicarbazone derivatives containing indole were synthesized and evaluated for their anti-inflammatory potential. METHODS: The compounds were obtained through a synthetic route of only two steps, with yields that varied between 33.6 and 90.4%, and characterized by spectroscopic and spectrometric techniques. RESULTS: An initial screening through the lymphoproliferation assay revealed that compounds LT76, LT81, and LT87 were able to inhibit lymphocyte proliferation, with CC50 of 0.56 ± 0.036, 0.9 ± 0.01 and 0.5 ± 0.07 µM, respectively, better results than indomethacin (CC50 > 12 µM). In addition, these compounds were able to suppress the in-vitro production of TNF-α and NO, in addition to stimulating the production of IL-4. Reinforcing in-vitro assays, the compounds were able to inhibit COX-2 similar to Celecoxib showing greater selectivity for this isoform (LT81 SI: 23.06 versus Celecoxib SI: 11.88). Animal studies showed that compounds LT76 (64.8% inhibition after 6 h), LT81 (89% inhibition after 6 h) and LT87 (100% inhibition after 4 h) were able to suppress edema in mice after inoculation carrageenan with greater potency than indomethacin, and immunohistochemistry revealed that the groups treated with LT76, LT81 and LT87 reduced the expression of COX-2, similar or better results when compared to indomethacin. Complementarily, in-silico studies have shown that these compounds have a good pharmacokinetic profile, for respecting the parameters of Lipinski and Veber, showing their good bioavailability. CONCLUSIONS: These results demonstrate the potency of thiosemicarbazone derivatives containing indole and confirm their importance as scaffolds of molecules with notorious anti-inflammatory activity.

Asunto(s)

Antiinflamatorios no Esteroideos/farmacología , Ciclooxigenasa 1/metabolismo , Inhibidores de la Ciclooxigenasa 2/farmacología , Ciclooxigenasa 2/metabolismo , Tiosemicarbazonas/farmacología , Animales , Carragenina/farmacología , Celecoxib/farmacología , Proliferación Celular/efectos de los fármacos , Edema/tratamiento farmacológico , Edema/metabolismo , Indoles/farmacología , Indometacina/farmacología , Linfocitos/efectos de los fármacos , Linfocitos/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C

RESUMEN

Parkinson's disease (PD) induced by environmental toxins involves a multifactorial cascade of harmful factors, thus motivating the search for therapeutic agents able to act on the greatest number of molecular targets. This study evaluated the efficacy of 50 mg/kg purified anacardic acids (AAs), isolated from cashew nut shell liquid, on multiple steps of oxidative stress and inflammation induced by rotenone in the substantia nigra (SN) and striatum. Adult mice were divided into four groups: Control, rotenone, AAs + rotenone, and AAs alone. Lipoperoxidation, nitric oxide (NO) levels, and reduced glutathione (GSH)/oxidized gluthatione (GSSG) ratio were evaluated. NF-kB-p65, pro-IL-1ß, cleaved IL-1ß, metalloproteinase-9, Tissue Inhibitory Factor-1 (TIMP-1), tyrosine hydroxylase (TH), and glial fibrillary acidic protein (GFAP) levels were assessed by Western blot. In silico studies were also made using the SwissADME web tool. Rotenone increased lipoperoxidation and NO production and reduced TH levels and GSH/GSSG ratio in both SN and striatum. It also enhanced NF-kB-p65, pro, and cleaved IL-1ß, MMP-9, GFAP levels compared to control and AAs groups. The AAs alone reduced pro-IL-1ß in the striatum while they augmented TIMP1 and reduced MMP-9 amounts in both regions. AAs reversed rotenone-induced effects on lipoperoxidation, NO production, and GSH/GSSG ratio, as well as increased TH and attenuated pro-IL-1ß and MMP-9 levels in both regions, NF-kB-p65 in the SN and GFAP in the striatum. Altogether, the in vivo and in silico analysis reinforced multiple and defined molecular targets of AAs, identifying that they are promising neuroprotective drug candidates for PD, acting against oxidative and inflammatory conditions induced by rotenone.

Asunto(s)

Ácidos Anacárdicos/farmacología , Fármacos Neuroprotectores/farmacología , Enfermedad de Parkinson Secundaria/tratamiento farmacológico , Enfermedad de Parkinson/tratamiento farmacológico , Plaguicidas/toxicidad , Ácidos Anacárdicos/química , Ácidos Anacárdicos/aislamiento & purificación , Animales , Simulación por Computador , Cuerpo Estriado/efectos de los fármacos , Cuerpo Estriado/metabolismo , Cuerpo Estriado/patología , Proteína Ácida Fibrilar de la Glía/genética , Glutatión/metabolismo , Disulfuro de Glutatión/metabolismo , Humanos , Interleucina-1beta/genética , Peroxidación de Lípido/efectos de los fármacos , Metaloproteinasa 9 de la Matriz/genética , Ratones , Óxido Nítrico/metabolismo , Estrés Oxidativo/efectos de los fármacos , Enfermedad de Parkinson/etiología , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/patología , Enfermedad de Parkinson Secundaria/inducido químicamente , Enfermedad de Parkinson Secundaria/genética , Enfermedad de Parkinson Secundaria/patología , Inhibidor Tisular de Metaloproteinasa-1/genética , Factor de Transcripción ReIA/genética , Tirosina 3-Monooxigenasa/genética

RESUMEN

Zika virus (ZIKV) is a flavivirus that has recently been associated with an increased incidence of neonatal microcephaly and other neurological disorders. The virus is primarily transmitted by mosquito bite, although other routes of infection have been implicated in some cases. The Aedes aegypti mosquito is considered to be the main vector to humans worldwide; however, there is evidence that other mosquito species, including Culex quinquefasciatus, transmit the virus. To test the potential of Cx. quinquefasciatus to transmit ZIKV, we experimentally compared the vector competence of laboratory-reared Ae. aegypti and Cx. quinquefasciatus. Interestingly, we were able to detect the presence of ZIKV in the midgut, salivary glands and saliva of artificially fed Cx. quinquefasciatus. In addition, we collected ZIKV-infected Cx. quinquefasciatus from urban areas with high microcephaly incidence in Recife, Brazil. Corroborating our experimental data from artificially fed mosquitoes, ZIKV was isolated from field-caught Cx. quinquefasciatus, and its genome was partially sequenced. Collectively, these findings indicate that there may be a wider range of ZIKV vectors than anticipated.

Asunto(s)

Culex/virología , Mosquitos Vectores/virología , Replicación Viral , Virus Zika/fisiología , Aedes/virología , Animales , Brasil/epidemiología , Genoma Viral , Humanos , Microcefalia/epidemiología , Mosquitos Vectores/fisiología , Saliva/virología , Glándulas Salivales/virología , Análisis de Secuencia de ADN , Virus Zika/genética , Virus Zika/aislamiento & purificación , Infección por el Virus Zika/epidemiología , Infección por el Virus Zika/transmisión , Infección por el Virus Zika/virología

RESUMEN

This study evaluates the effect of different concentrations (0, 10, 50 and 100ng/mL) of bone morphogenetic protein-2 (BMP-2) on primordial and secondary follicle development. It also investigates the effects of FSH and BMP-2 on the growth, morphology, ultrastructure and expression of mRNA for GDF9, NLRP5 and NPM2 genes in secondary follicles cultured for 18 days. The presence of BMP-2 at all tested concentrations increased the development of primordial follicles in vitro, but the highest concentration of BMP-2 (100 ng/mL) reduced the percentage of normal follicles when compared with tissues cultured with 10 ng/mL BMP-2. During culture of secondary follicles, in contrast to higher concentrations (50 or 100 ng/mL), 10 ng/mL BMP-2 kept the morphology of follicles during initial stages of in vitro culture. This concentration of BMP-2 also benefits maintenance of the ultrastructure of 18-day cultured follicles. The presence of both BMP-2 and FSH in culture medium resulted in a significant (P<0.05) increase in follicular diameter after 18 days of culture. However, both FSH and BMP-2 reduced follicular mRNA expression of GDF9 and NLRP5 when compared to follicles cultured in media containing only FSH. In combination with FSH, BMP-2 reduced the mRNA levels of NPM2, when compared to follicles cultured in control medium. It is concluded from these data that 10 ng/mL BMP-2 promotes the growth of primordial in vitro and it helps to maintain the ultrastructure of secondary follicles, while FSH is more important for better expression of follicular markers like GDF9 and NLRP5.

Asunto(s)

Proteína Morfogenética Ósea 2/metabolismo , Folículo Ovárico/fisiología , Animales , Proteína Morfogenética Ósea 2/farmacología , Bovinos , Células Cultivadas , Femenino , Hormona Folículo Estimulante/metabolismo , Hormona Folículo Estimulante/farmacología , Regulación de la Expresión Génica , Factor 9 de Diferenciación de Crecimiento/biosíntesis , Técnicas In Vitro , Nucleoplasminas/biosíntesis , Oocitos , Folículo Ovárico/efectos de los fármacos , ARN Mensajero/análisis

RESUMEN

This study aims to investigate the effects of jacalin and follicle-stimulating hormone (FSH) on activation and survival of goat primordial follicles, as well as on gene expression in cultured ovarian tissue. Ovarian fragments were cultured for 6 days in minimum essential medium (MEM) supplemented with jacalin (10, 25, 50 or 100 µg/ml - Experiment 1) or in MEM supplemented with jacalin (50 µg/ml), FSH (50 ng/ml) or both (Experiment 2). Non-cultured and cultured tissues were processed for histological and ultrastructural analysis. Cultured tissues from Experiment 2 were also stored to evaluate the expression of BMP-15, KL (Kit ligand), c-kit, GDF-9 and proliferating cell nuclear antigen (PCNA) by real-time polymerase chain reaction (PCR). The results of Experiment 1 showed that, compared with tissue that was cultured in control medium, the presence of 50 µg/ml of jacalin increased both the percentages of developing follicles and viability. In Experiment 2, after 6 days, higher percentages of normal follicles were observed in tissue cultured in presence of FSH, jacalin or both, but no synergistic interaction between FSH and jacalin was observed. These substances had no significant effect on the levels of mRNA for BMP-15 and KL, but FSH increased significantly the levels of mRNA for PCNA and c-kit. On the other hand, jacalin reduced the levels of mRNA for GDF-9. In conclusion, jacalin and FSH are able to improve primordial follicle activation and survival after 6 days of culture. Furthermore, presence of FSH increases the expression of mRNA for PCNA and c-kit, but jacalin resulted in lower GDF-9 mRNA expression.

Asunto(s)

Hormona Folículo Estimulante/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Folículo Ovárico/efectos de los fármacos , Lectinas de Plantas/farmacología , Animales , Proteína Morfogenética Ósea 15/genética , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Femenino , Cabras , Factor 9 de Diferenciación de Crecimiento/genética , Folículo Ovárico/citología , Folículo Ovárico/fisiología , Antígeno Nuclear de Célula en Proliferación/genética , Factor de Células Madre/genética , Técnicas de Cultivo de Tejidos

RESUMEN

The aim of the present study was to analyze the effect of a combination of metformin hydrochloride and melatonin on oxidative stress together with a biochemical and histopathological analysis of the livers of Wistar rats induced with PCOS. The results indicated that a combination of the drugs was more effective in the reduction of plasmatic levels of liver enzyme alanine aminotransferase, nitric oxide and total glutathione, and decreased the inflammatory response and histopathological damage, producing results that were significantly similar to animals from the control group. A mixture of the drugs produced more effective results against liver toxicity caused by PCOS, encouraging the normalization of biochemical parameters. During pregnancy, there was reduced oxidative stress compared to monotherapeutic use of these drugs. Interestingly, the combination of the drugs caused a physiological reaction similar to responses identified in healthy rats without induction of the PCOS control group. However, the clinical and physiological effectiveness of the combination should be further explored, especially with respect to the possible side effects on offspring.

Asunto(s)

Antioxidantes/administración & dosificación , Hígado/efectos de los fármacos , Melatonina/administración & dosificación , Metformina/administración & dosificación , Estrés Oxidativo/efectos de los fármacos , Síndrome del Ovario Poliquístico/tratamiento farmacológico , Animales , Quimioterapia Combinada , Femenino , Hígado/metabolismo , Hígado/patología , Estrés Oxidativo/fisiología , Síndrome del Ovario Poliquístico/metabolismo , Síndrome del Ovario Poliquístico/patología , Embarazo , Ratas , Ratas Wistar , Resultado del Tratamiento

RESUMEN

The aim of this study was to examine the effect of maternal exposure to Panax ginseng extract (GE) on the prenatal dexamethasone (DEXA)-induced increase in testosterone production by isolated Leydig cells in adult rats. Pregnant rats were treated with (i) GE (200 mg/kg) or vehicle on days 10-21; (ii) DEXA (100 µg/kg) or vehicle on days 14-21; or (iii) a combination of GE plus DEXA at the same doses and with the same regimen. Testosterone production was induced either by the activator of protein kinase A (dbcAMP) or substrates of steroidogenesis [22(R)-hydroxycholesterol (22(R)-OH-C)] and pregnenolone. The capacity of rat Leydig cells exposed to DEXA to synthesize testosterone induced by dbcAMP, 22(R)-OH-C or pregnenolone was increased in comparison with the control group. Combined exposure to DEXA + GE prevented the effect of DEXA on the responsiveness of Leydig cells to all inductors of testosterone synthesis, whereas GE alone did not modify the response to inductors. No modifications in testosterone production were observed under basal conditions. StAR immunoexpression in Leydig cells was not modified by prenatal exposure to DEXA, GE or DEXA + GE. P450scc and glucocorticoid receptor immunoexpression was higher in offspring exposed to DEXA in comparison with the control group. This increased expression was prevented by combined treatment with DEXA + GE. The present findings demonstrate that GE is capable of reversing the effect of DEXA on testosterone synthesis by rat Leydig cells.

Asunto(s)

Dexametasona/farmacología , Células Intersticiales del Testículo/metabolismo , Panax/química , Extractos Vegetales/farmacología , Efectos Tardíos de la Exposición Prenatal/metabolismo , Testosterona/biosíntesis , Factores de Edad , Animales , Peso Corporal/efectos de los fármacos , Enzima de Desdoblamiento de la Cadena Lateral del Colesterol/metabolismo , Interacciones Farmacológicas , Femenino , Glucocorticoides/farmacología , Células Intersticiales del Testículo/ultraestructura , Masculino , Microscopía Electrónica de Transmisión , Tamaño de los Órganos/efectos de los fármacos , Fosfoproteínas/metabolismo , Embarazo , Ratas , Ratas Wistar , Testículo/citología , Testículo/efectos de los fármacos , Testículo/metabolismo , Testosterona/sangre

RESUMEN

Microorganisms with immunomodulating effects beneficially affect the host organism by improving the microbial equilibrium and balancing the immune system. Zymomonas mobilis is reported to have antagonistic properties against yeast and other pathogenic microorganisms in humans and animals. This study assessed the effects of Z. mobilis UFPEDA 202 (10(9)CFU/mL) cultures on polymicrobial sepsis induced by cecal ligation and puncture (CLP). The survival of animals subjected to lethal sepsis was evaluated after pre-treatment, post-treatment or a combination of both. 6h after the induction of sepsis, neutrophil migration, the number of bacteria, myeloperoxidase, TNF-α, MCP-1, and IL-10 were performed in the peritoneal lavage of animals. Histopathological changes in the spleen of animals were evaluated by light microscopy, and apoptosis of splenocytes was analyzed by transmission electron microscopy. The results showed that the combination of prophylactic and therapeutic treatment with Z. mobilis increased the survival of animals by 50% at 96 h after the induction of sepsis. There was a reduction in the levels of TNF-α and myeloperoxidase (MPO) in lung tissue. There was also a reduction in the number of viable bacteria in peritoneal fluid. However, increases in neutrophil migration and IL-10 levels were observed. The observed levels of MCP-1 remained similar to the control. Histopathology analysis showed a decrease in acute lung injury. The group pre-treated with the Z. mobilis culture demonstrated a marked decrease in the number of apoptotic cells in the spleen (24%). This study demonstrates that Z. mobilis cultures increased the survival of animals with severe sepsis. This survival was mediated by improvement of neutrophil migration, enhanced activity against pathogenic enteric bacteria and reduced lung injury.

Asunto(s)

Lesión Pulmonar Aguda/prevención & control , Sepsis/prevención & control , Zymomonas , Lesión Pulmonar Aguda/inmunología , Lesión Pulmonar Aguda/microbiología , Lesión Pulmonar Aguda/patología , Animales , Apoptosis , Carga Bacteriana , Quimiocina CCL2/inmunología , Citocinas/inmunología , Interleucina-10/inmunología , Recuento de Leucocitos , Masculino , Ratones , Cavidad Peritoneal/citología , Cavidad Peritoneal/microbiología , Peroxidasa/inmunología , Sepsis/inmunología , Sepsis/microbiología , Sepsis/patología , Bazo/citología , Factor de Necrosis Tumoral alfa/inmunología

RESUMEN

This study evaluated the levels of bone morphogenetic protein receptors BMPRIB and BMPRII mRNA in goat follicles and the effects of bone morphogenetic protein-15 (BMP-15) on the in vitro development of cultured preantral follicles. Real-time polymerase chain reaction (PCR) was used to analyze the levels of BMPRIB and BMPRII mRNA in caprine preantral follicles and in small and large antral follicles. Preantral follicles (≥150 µm) were also isolated from goat ovaries and cultured for 18 days in α-MEM(+) supplemented with or without BMP-15 (10, 50, or 100 ng/ml). At the end of culture, some follicles were fixed for ultrastructural evaluation. Real-time PCR showed a reduction in BMPRII mRNA levels from the primary to secondary follicles. Higher levels of BMPRIB mRNA were observed in granulosa/theca cells from large antral follicles compared with small antral follicles. Moreover, BMPRII mRNA was expressed to a greater extent in cumulus-oocyte complexes from large antral follicles than in their respective granulosa/theca cells. In culture, 50 ng/ml BMP-15 positively influenced antral cavity formation and follicle growth after 18 days and also maintained follicular integrity. Thus, BMPRIB and BMPRII mRNAs are present in all follicular categories. BMP-15 (50 ng/ml) stimulates growth, antrum formation and the ultrastructural integrity of isolated caprine preantral follicles after 18 days of culture.

Asunto(s)

Proteína Morfogenética Ósea 15/farmacología , Receptores de Proteínas Morfogenéticas Óseas de Tipo II/genética , Receptores de Proteínas Morfogenéticas Óseas de Tipo 1/genética , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Folículo Ovárico/crecimiento & desarrollo , Folículo Ovárico/metabolismo , Animales , Receptores de Proteínas Morfogenéticas Óseas de Tipo 1/metabolismo , Receptores de Proteínas Morfogenéticas Óseas de Tipo II/metabolismo , Proliferación Celular/efectos de los fármacos , Femenino , Cabras/genética , Humanos , Meiosis/efectos de los fármacos , Microscopía Fluorescente , Oocitos/citología , Oocitos/efectos de los fármacos , Oocitos/metabolismo , Folículo Ovárico/citología , Folículo Ovárico/ultraestructura , ARN Mensajero/genética , ARN Mensajero/metabolismo , Técnicas de Cultivo de Tejidos , Supervivencia Tisular/efectos de los fármacos

RESUMEN

Insulin-degrading enzyme (IDE) has been shown to enhance the binding of androgen and glucocorticoid receptors to DNA in the nuclear compartment. Glucocorticoids cause hyperglycaemia, peripheral resistance to insulin and compensatory hyperinsulinaemia. The aim of the present study was to investigate the effect of dexamethasone (D), testosterone (T) and dexamethasone plus testosterone (D + T) on the regulation of IDE and on the remodelling of rat ventral prostate after castration (C). Castration led to a marked reduction in prostate weight (PW). Body weight was significantly decreased in the castrated animals treated with dexamethasone, and the relative PW was 2.6-fold (±0.2) higher in the D group, 2.8-fold (±0.3) higher in the T group and 6.6-fold (±0.6) higher in the D + T group in comparison with the castrated rats. Ultrastructural alterations in the ventral prostate in response to androgen deprivation were restored after testosterone and dexamethasone plus testosterone treatments and partially restored with dexamethasone alone. The nuclear IDE protein level indicated a 4.3-fold (±0.4) increase in castrated rats treated with D + T when compared with castration alone. Whole-cell IDE protein levels increased approximately 1.5-fold (±0.1), 1.5-fold (±0.1) and 2.9-fold (±0.2) in the D, T and D + T groups, respectively, when compared with castration alone. In conclusion, the present study reports that dexamethasone-induced hyperinsulinaemic condition plus exogenous testosterone treatment leads to synergistic effects of insulin and testosterone in the prostatic growth and in the amount of IDE in the nucleus and whole epithelial cell.

Asunto(s)

Castración , Dexametasona/farmacología , Insulisina/metabolismo , Próstata/metabolismo , Próstata/patología , Testosterona/farmacología , Andrógenos/farmacología , Animales , Peso Corporal/efectos de los fármacos , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Núcleo Celular/patología , Dexametasona/efectos adversos , Glucocorticoides/efectos adversos , Glucocorticoides/farmacología , Hiperinsulinismo/inducido químicamente , Hiperinsulinismo/metabolismo , Insulisina/efectos de los fármacos , Masculino , Modelos Animales , Próstata/efectos de los fármacos , Ratas , Ratas Wistar

RESUMEN

BACKGROUND: The present study was designed to examine the effect of chronic treatment with rosiglitazone - thiazolidinedione used in the treatment of type 2 diabetes mellitus for its insulin sensitizing effects - on the Leydig cell steroidogenic capacity and expression of the steroidogenic acute regulatory protein (StAR) and cholesterol side-chain cleavage enzyme (P450scc) in normal adult rats. METHODS: Twelve adult male Wistar rats were treated with rosiglitazone (5 mg/kg) administered by gavage for 15 days. Twelve control animals were treated with the vehicle. The ability of rosiglitazone to directly affect the production of testosterone by Leydig cells ex vivo was evaluated using isolated Leydig cells from rosiglitazone-treated rats. Testosterone production was induced either by activators of the cAMP/PKA pathway (hCG and dbcAMP) or substrates of steroidogenesis [22(R)-hydroxy-cholesterol (22(R)-OH-C), which is a substrate for the P450scc enzyme, and pregnenolone, which is the product of the P450scc-catalyzed step]. Testosterone in plasma and in incubation medium was measured by radioimmunoassay. The StAR and P450scc expression was detected by immunocytochemistry. RESULTS: The levels of total circulating testosterone were not altered by rosiglitazone treatment. A decrease in basal or induced testosterone production occurred in the Leydig cells of rosiglitazone-treated rats. The ultrastructural and immunocytochemical analysis of Leydig cells from rosiglitazone-treated rats revealed cells with characteristics of increased activity as well as increased StAR and P450scc expression, which are key proteins in androgen biosynthesis. However, a number of rosiglitazone-treated cells exhibited significant mitochondrial damage. CONCLUSION: The results revealed that the Leydig cells from rosiglitazone-treated rats showed significant reduction in testosterone production under basal, hCG/dbcAMP- or 22 (R)-OH-C/pregnenolone-induced conditions, although increased labeling of StAR and P450scc was detected in these cells by immunocytochemistry. The ultrastructural study suggested that the lower levels of testosterone produced by these cells could be due to mitochondrial damage induced by rosiglitazone.

Asunto(s)

Células Intersticiales del Testículo/efectos de los fármacos , Esteroides/biosíntesis , Tiazolidinedionas/farmacología , Animales , Peso Corporal/efectos de los fármacos , Células Cultivadas , Enzima de Desdoblamiento de la Cadena Lateral del Colesterol/metabolismo , Evaluación Preclínica de Medicamentos , Hipoglucemiantes/efectos adversos , Hipoglucemiantes/farmacología , Células Intersticiales del Testículo/metabolismo , Células Intersticiales del Testículo/ultraestructura , Masculino , Tamaño de los Órganos/efectos de los fármacos , Fosfoproteínas/metabolismo , Ratas , Ratas Wistar , Rosiglitazona , Vesículas Seminales/citología , Vesículas Seminales/efectos de los fármacos , Testículo/citología , Testículo/efectos de los fármacos , Testosterona/sangre , Tiazolidinedionas/efectos adversos , Factores de Tiempo

RESUMEN

RATIONALE: The mechanism of action of diethylcarbamazine (DEC), an antifilarial drug effective against tropical pulmonary eosinophilia, remains controversial. DEC effects on microfilariae depend on inducible NO synthase (iNOS). In eosinophilic pulmonary inflammation, its therapeutic mechanism has not been established. We previously described the rapid up-regulation of bone marrow eosinophilopoiesis in ovalbumin (OVA)-sensitized mice by airway allergen challenge, and further evidenced the down-regulation of eosinophilopoiesis by iNOS- and CD95L-dependent mechanisms. OBJECTIVES: We investigated whether: (1) DEC can prevent the effects of airway challenge of sensitized mice on lungs and bone marrow, and (2) its effectiveness depends on iNOS/CD95L. METHODS: OVA-sensitized BALB/c mice were intranasally challenged for 3 consecutive days, with DEC administered over a 12-, 3-, or 2-day period, ending at the day of the last challenge. We evaluated: (1) airway resistance, cytokine (IFN-gamma, IL-4, IL-5, and eotaxin) production, and pulmonary eosinophil accumulation; and (2) bone marrow eosinophil numbers in vivo and eosinophil differentiation ex vivo. MEASUREMENTS AND MAIN RESULTS: DEC effectively prevented the effects of subsequent challenges on: (1) airway resistance, Th1/Th2 cytokine production, and pulmonary eosinophil accumulation; and (2) eosinophilopoiesis in vivo and ex vivo. Recovery from unprotected challenges included full responses to DEC during renewed challenges. DEC directly suppressed IL-5-dependent eosinophilopoiesis in naive bone marrow. DEC was ineffective in CD95L-deficient gld mice and in mice lacking iNOS activity because of gene targeting or pharmacological blockade. CONCLUSIONS: DEC has a strong impact on pulmonary eosinophilic inflammation in allergic mice, as well as on the underlying hemopoietic response, suppressing the eosinophil lineage by an iNOS/CD95L-dependent mechanism.

Asunto(s)

Enfermedades de la Médula Ósea/tratamiento farmacológico , Dietilcarbamazina/farmacología , Eosinofilia/tratamiento farmacológico , Proteína Ligando Fas/fisiología , Filaricidas/farmacología , Óxido Nítrico Sintasa de Tipo II/fisiología , Eosinofilia Pulmonar/tratamiento farmacológico , Resistencia de las Vías Respiratorias/efectos de los fármacos , Animales , Broncoconstrictores/farmacología , Eosinófilos/efectos de los fármacos , Hematopoyesis/efectos de los fármacos , Interferón gamma/biosíntesis , Interleucina-4/biosíntesis , Interleucina-5/biosíntesis , Recuento de Linfocitos , Cloruro de Metacolina/farmacología , Ratones , Ratones Endogámicos BALB C

RESUMEN

The objective of the present study was to evaluate, by light and transmission electron microscopy, the efficacy of a single intratesticular injection of a novel zinc-based solution, as a contraceptive for male dogs. Fifteen mongrel dogs were assigned to three groups (five dogs/group). Group 1, the control group, which consisted of animals ranging from 8 mo to 4 yr, was injected with saline solution. Group 2, which consisted of animals ranging from 8 mo to 1 yr old and Group 3, animals ranging from 2 to 4 yr old, were injected with a zinc-based solution (0.2-1.0mL; volume based on testicular width). There were no histopathological changes detected in testes from control dogs. Histological examination of treated groups revealed degeneration, vacuolation, fewer germ cells, formation of multinucleated giant cells, and a lack of elongated spermatids in atrophic seminiferous tubules. Leydig cells had varying degrees of lipid degeneration and necrosis. The majority of seminiferous tubules in all zinc-treated dogs were lined only by Sertoli cells, which were vacuolated. Ultrastructure of testis of treated groups had degenerate Sertoli and Leydig cells, characterized by numerous mitochondria with the lack of a matrix and agglomeration of lysosomal bodies. The cytoplasm of elongated spermatids was characterized by tubules of hyperplastic and hypertrophic smooth endoplasmic reticulum and numerous Golgi apparati. Round spermatids in Golgi phase had lysis of acrosomal vesicles. The degree of histological changes suggested irreversibility. In conclusion, intratesticular injection of a zinc-based solution effectively impaired spermatogenesis.

Asunto(s)

Perros/fisiología , Gluconatos/farmacología , Bloqueadores de Espermatogénesis/farmacología , Testículo/efectos de los fármacos , Zinc/farmacología , Animales , Anticoncepción/métodos , Gluconatos/administración & dosificación , Inyecciones , Masculino , Microscopía Electrónica de Transmisión , Bloqueadores de Espermatogénesis/administración & dosificación , Espermatozoides/efectos de los fármacos , Espermatozoides/crecimiento & desarrollo , Espermatozoides/ultraestructura , Testículo/patología , Testículo/ultraestructura , Zinc/administración & dosificación

RESUMEN

PURPOSE: To demonstrate the effects of the use of sunflower seed oil on the treatment of skin wounds. METHODS: Eighteen male Saint Inês lambs were divided in 3 groups according to the pos-operative (7, 14 and 21 days). After antisepsis and local anestesia, two 4cm2 wounds on each side of the thoracic region, close to the scapule were surgically produced. The experimental wounds were treated with sunflower seed oil, with high concentration of linoleic acid (LA), and the control ones with sterilized Vaseline. Biopsies of the pos-operative wounds tissue were performed on the 7th, 14th, 21st days and histologically evaluated. RESULTS: Topic application of sunflower seed oil accelerated healing process at the 7th and 21st days, reducing wound area and increasing wound contraction. Granulation tissue increased faster on treated wounds. The epidermis of the treated wounds was completely recovered when compared to control wounds. CONCLUSION: The topic use of sunflower seed oil accelerated the healing process, and it can be used as an alternative therapy on second intention wound healing.

Asunto(s)

Animales , Masculino , Cicatrización de Heridas , Helianthus , Aceites de Plantas/administración & dosificación , Aceites de Plantas/uso terapéutico , Semillas , Administración Tópica , Ovinos

RESUMEN

Ultrathin sections of L3 of Wuchereria bancrofti embedded in hydrophilic resin were incubated with antisera pools from individuals (1) asymptomatic microfilaremic with different microfilaria (mf) densities (1-100, 101-500, and >1,000 mf/ml); (2) chronic with hydrocele or lymphedema; and (3) with no evidence of microfilaremia or clinical filariasis but residing in an endemic area. The groups of microfilaremic subjects studied presented differences relative to the intensity of labeling, with the density of gold particles per square micrometer proportional to microfilaremia. Incubation of ultrathin sections of W. bancrofti L3 larvae in the presence of antisera from patients exhibiting chronic obstructive lymphatic pathology of hydrocele and from individuals with clear clinical evidence of lymphedema exhibited a strong reaction in the same tissues. Except for the endemic normal group, all groups studied showed reactivity against epitopes in all tissues of infective larvae of W. bancrofti. The cuticle presented an intense labeling, suggesting a possible target structure for immune response.

Asunto(s)

Antígenos Helmínticos/análisis , Filariasis/inmunología , Sueros Inmunes/inmunología , Wuchereria bancrofti/inmunología , Adolescente , Adulto , Animales , Anticuerpos Antihelmínticos/inmunología , Antígenos Helmínticos/inmunología , Culex , Femenino , Humanos , Inmunohistoquímica , Insectos Vectores , Larva/inmunología , Masculino , Persona de Mediana Edad , Parasitemia/inmunología , Wuchereria bancrofti/ultraestructura

RESUMEN

The bli-1 gene of Caenorhabditis elegans has previously been described as a mutation which disrupts the structure of the adult-stage cuticle, causing the formation of fluid-filled blisters. We investigated the blistering phenotype exhibited of n361 allele through immunocytochemical and freeze-fracture techniques. In the course of the blistering process several fine changes occurred, including a high-electron-density granulous material filling the intermediate layer, alterations in strut structure, and finally the total disappearance of the fibrous and basal layers. A polyclonal antibody against a synthetic 18-amino-acid peptide of the 3A3-collagen sequence labeled all the cuticular regions of the N(2) strain of the nematode C. elegans except for the intermediate layer. Similarly, no reaction was observed in the intermediate layer of the mutant strain DR 847 bli-1 (n361), which was filled by the granulous and electron-dense material. Replicas of quick-frozen, freeze-fracture, deep-etched, and rotatory-shadowed adult forms of the mutant DR 847 bli-1 (n361) of C. elegans revealed an increased number of the filamentous structures filling the intermediate layer in older nematodes, and also a gradual destruction and disappearance of the fibrous and basal layers. Based on these results, we postulated that the blistering phenotype is due to an altered function of bli-1 gene, which is probably enzymatic.

Asunto(s)

Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/genética , Animales , Vesícula/genética , Vesícula/patología , Colágeno/metabolismo , Técnica de Fractura por Congelación , Inmunohistoquímica , Microscopía Electrónica