José Maria Soares Júnior

The effects of topical isoflavones on postmenopausal skin: Double-blind and randomized clinical trial of efficacy

OBJECTIVEnThe aim of this study was to evaluate the effects of estrogen and isoflavones on postmenopausal skin morphological parameters.nnnSTUDY DESIGNnA randomized, double-blind, estrogen-controlled trial was performed on postmenopausal women treated in the Gynecology Department of the Federal University of São Paulo. This study was designed to analyze the effects of topical administration of estradiol and isoflavones on facial skin for 24 weeks. The participants were divided into two groups: G1-17-betaestradiol 0.01% (n=18) and G2-isoflavones 40% (genistein 4%, n=18). Skin biopsies were performed on each patient before and after the treatment. The skin samples were processed for histological analysis, stained with haematoxylin and eosin, and examined using light microscopy.nnnRESULTSnAfter 24 weeks of treatment, the estradiol group had a significant increase in skin parameters analyzed compared to the isoflavone group and to the baseline measurements: epidermal thickness (a 75% increase in the estrogen group and 20% in the isoflavone group), number of dermal papillae (a rise of 125% with estrogen, no significant gain with isoflavones), fibroblasts (a 123% accretion with estradiol, no significant gain with isoflavones), and vessels (a 77% increase with estrogen and 36% with isoflavones).nnnCONCLUSIONnOur data suggest that estrogens may have a stronger effect on histomorphometrical parameters than isoflavones.

Evaluation of the implantation in pinealectomized and/or submitted to the constant illumination rats

En este trabajo evaluamos los aspectos morfologicos y cuantitativos de los sitios de implantaciones en uteros de ratas pinealectomizadas y/o sometidas a iluminacion constante. Los grupos experimentales fueron: I xad ratas normales mantenidas en ciclo claro / oscuro de 12/12 horas, durante dos meses, cruzadas y sacrificadass en el 60 dia de prenez (control); II xad ratas sham-pinealectomizadas mantenidas en ciclo claro/oscuro de 12/12 horas, durante dos meses, cruzadas y sacrificadas en el 60 dia de prenez (control); III xad ratas pinealectomizadas mantenidas en ciclo claro/oscuro de 12/12 horas, durante dos meses, cruzadas y sacrificadas en el 60 dia de prenez; IV xad ratas normales mantenidas en constante iluminacion durante dos meses acasaladas y sacrificadas en el 60 dia de prenez; V xad ratas sham-pinealectomiazadas mantenidas en constante iluminacion durante dos meses, cruzadas y sacrificadas en el 60 dia de prenez; VI xad ratas pinealectomizadas, mantenidas en constante iluminacion durante dos meses, cruzadas y sacrificadas en el 60 dia de prenez. El analisis estadistico de las medias de los sitios de implantaciones mostro que hubo diferencia significativa, donde los grupos III, IV, V y VI diferian de los grupos I y II, los cuales presentaron las mayores medias. Los grupos III, IV, V y VI no son diferentes entre si. El analisis morfologico de los sitios en los grupos controles revelo la presencia de pequenas lagunas conteniendo sangre, ademas de celulas trofoblasticas y algunos citrofoblastos poliploides. En los grupos III, IV, V y VI se evidencio lagunas muy desarrolladas en los sitios, con trofoblastos, citrofoblastos con alto grado de poliploidia y sinciciotrofoblastos. Concluimos que la pinealectomia y/o constante iluminacion llevan a una reduccion del numero de blastocistos implantados en ratas, ademas estimula el desarrollo de los sitios de implantacion, indicando asi; que la melatonina puede tener importante funcion en la viabilidad de implantacion del blastocisto

Rat pinealectomy: a modified direct visual approach

PURPOSEnTo report a new, direct visual approach for rat pinealectomy.nnnMETHODSnEighty adult female rats (Rattus norvegicus albinus EPM-1 strain) were weighted and anesthetized intraperitoneally with 15 mg/kg xylazine and 30 mg/kg ketamine. The animal was fastened to a dissection table, an incision was made in the skin and the subcutaneous tissue, bringing the lambda into view. The skullcap was opened with a dental drill, bringing the cerebral hemispheres and the superior sagittal sinus into view. The pineal gland, located under the venous sinus, was removed in a single piece using tweezers. Next, the bone fragment was returned to its place and the surgical layers were sutured.nnnRESULTSnThis new technique is easy to be done, avoids bleedings and removes only the pineal gland without damage to the remaining encephalon. In addition it makes possible the achievement of a sham surgery, allowing the pineal gland to remain intact.nnnCONCLUSIONnThe proposed technique intends to facilitate studies aiming to better understanding the complexity and importance of the pineal gland on reproductive and other body systems.

Perfil de glicosaminoglicanos sulfatados no útero de camundongas durante o ciclo estral

OBJECTIVE: Identification and quantitation of sulphated glycosaminoglycans (GAGs) in the uterus of female mice during the estrous cycle. METHODS: Four groups (n = 10 each) of virgin, 100-day old female mice were assembled according to the estrous cycle phase: proestrus, estrus, metaestrus and diestrus. Samples of the median portion of uterine horns were processed for light microscopy examination (H/E and Alcian blue + PAS). The GAGs were extracted and characterized by agarose gel electrophoresis. Data were analyzed by the unpaired Students t-test. RESULTS: At light microscopy GAGs appear in all layers of the uterus, especially in the endometrium, between collagen fibers, in the basal membrane and around fibroblasts. Biochemical analyses disclosed presence of dermatan sulphate (DS), chondroitin sulphate (CS and heparan sulphate (HS) during all estral cycle phases. There was no clear electrophoretic separation between DS and CS, thus these two GAGs were considered together (DS+CS) (proestrus = 0.854 ± 0.192; estrus = 1.073 ± 0.254; metaestrus = 1.003 ± 0.255; diestrus = 0.632 ± 0.443 µg/mg). HS was as follows: proestrus = 0.092 ± 0.097; estrus = 0.180 ± 0.141; metaestrus = 0.091 ± 0.046; diestrus = 0.233 ± 0.147 µg/mg. The uterine content of DS+CS peaked at estrus (estrogenic action) and that of HS at diestrus (progestagen action). CONCLUSION: Due to a constant turnover process, there are definite alterations in the uterine profile of GAGs content during the estrous cycle in mice, which may be modulated by female sex hormones.

Efeitos da associação estro-androgênica em mulheres na pós-menopausa

OBJETIVO: avaliar os efeitos da associacao de estrogenios e androgenios sobre a qualidade de vida e a sexualidade de mulheres durante o climaterio. METODOS: foram incluidas 96 pacientes com sintomas vasomotores e disfuncao sexual na pos-menopausa. As mulheres foram aleatoriamente divididas em tres grupos de tratamento com 32 pacientes cada: placebo, estrogenios conjugados equinos (0,625 mg por dia) e associacao de estrogenios conjugados equinos (0,625 mg por dia) e de metiltestosterona (2,5 mg por dia). O tratamento foi realizado por tres meses. A avaliacao da qualidade de vida e da sexualidade foi realizada por entrevista usando o Questionario de Saude da Mulher (QSM) e o Questionario Simplificado sobre Sexualidade, antes e apos o tratamento. Alguns parâmetros do risco cardiovascular, o eco endometrial e a toxidade hepatica foram avaliados. Para analise dos dados aplicou-se o teste ANOVA, seguido pelo teste post hoc de Fisher e teste de Shapiro-Wilk. RESULTADOS: houve melhora nos parâmetros do QSM nos grupos que receberam estrogenioterapia isolada e associada ao androgenio em comparacao ao grupo placebo. Entretanto, nao houve diferenca entre os tres grupos em relacao as questoes referentes aos sintomas somaticos. A associacao estro-androgenica foi superior ao estrogenio isolado nas questoes relacionadas com funcao sexual (media da pontuacao: 64 vs 67; p<0,05) e humor deprimido (media da pontuacao: 75 vs 80; p<0,05). No grupo que recebeu a terapia estro-androgenica, houve reducao do colesterol total (212±42 e 194±43, antes e apos o tratamento, respectivamente) e do HDL-colesterol (56±16 e 48±14, antes e apos o tratamento, respectivamente), bem como discreto aumento do eco endometrial (4,7±2,3 e 5,5±2,3 antes e apos o tratamento, respectivamente). Nao foi constatada alteracao significante das enzimas hepaticas entre os grupos, durante o periodo de estudo. CONCLUSOES: a associacao estro-androgenica resultou em melhora na qualidade de vida e nos disturbios sexuais. Este efeito foi superior a estrogenioterapia isolada e ao placebo. O efeito do tratamento estro-androgenico foi mais marcante nos itens de humor deprimido e de funcao sexual no questionario QSM.

Efeitos das isoflavonas em altas doses sobre o útero da rata

OBJECTIVE: To evaluate the effects of high-dose isoflavones on the uterus of castrated adult rats. METHODS: Adult, ovariectomized virgin rats (n = 40) were treated by gavage during 30 consecutive days with vehicle (propylene glycol, group GCtrl) or different doses of genistein: 42 (group GES42), 125 (GES125), or 250 (GES250) µg/g body weight per day. Animals were killed, weighed, vaginal and uterine samples were taken for cytologic evaluation, and serum levels of 17 β-estradiol and progesterone were determined. The middle third of the uterine horns was dissected, fixed in 10% formaldehyde and processed for paraffin inclusion; 5-µm thick sections were obtained and stained with HE for further histological study under light microscopy. The endometrial morphology and area, number and area of glands, and number of eosinophils in the lamina propria were analyzed. ANOVA and the Tukey-Kramer test were used for statistical analyses. RESULTS: Uterine weight, endometrial glandular area, and number of glands and eosinophils were all higher in GES250 > G125 than in the other groups (GES250 > GES125 > GES42 = GCtrl; p < 0.05). Morphological data showed signs of endometrial proliferation upon treatment with genistein, especially in animals in GES125 and GES250 compared to other groups. In all animals in GES250, signs of uterine squamous metaplasia were observed. CONCLUSION: A short treatment period with high daily doses of isoflavones can promote endometrial squamous metaplasia in ovariectomized rats.

Efeitos da isoflavona e dos estrogênios conjugados eqüinos sobre a qualidade de vida na pós-menopausa

PuRPOse: to analyze the isoflavone and estrogen effects on the postmenopausal quality of life. MethOds: this is a randomized and double-blind study with 79 postmenopausal patients, 12 months of amenorrhea, 40 years old or more and body mass index (BMI) above 30 kg/m2. The participants were randomly divided into two treatment groups: GECP received orally two capsules, every 12 hours, one contained 0.625 mg conjugated equine estrogen and another placebo (n=33); GECS received two capsules of 150 mg extract of soy, with 60 mg isoflavone (n=32). Both treatments were administered for six months. The Quality Menopause Specific Questionnaire of Life was applied before and after one, three and six months of treatment. The parameters of gynecological cancer risk were evaluated. ANOVA and the Tukey test were used for data analysis. Results: there was a reduction in the values of the vasomotor parameters after six months of treatment, 1.6±0.8 and 2.4±1.6, compared to before therapy, 4.0±2.2 and 4.2±2.3 in GECP and GECS, respectively. The psychological aspects showed reduction in values after six months of therapy, 2.5±1.2 and 2.9±1.4, compared to before treatment, 3.6±1.6 and 4.1±1.9 in GECP and GECS, respectively. Similar results were obtained on the physical aspects and in the sexual symptoms. COnClusiOns: isoflavones may positively act on life quality of postmenopausal women. This effect was similar to conjugated equine estrogen.

Estudo morfológico e molecular da mama de ratas castradas tratadas com isoflavona ou estrogênios

PURPOSE: to analyze the effects of isoflavones and estrogens on the morphology, morphometry and VEGF expression of the adult female rat mammary gland. METHODS: Forty-five adult female rats were oophorectomized; 28 days after surgery they were divided into 3 groups of 15 animals each: CON - control (treated with propylenoglycol); ISO - isoflavones (100 mg/kg) and CEE - conjugated equine estrogens (50 µg/Kg). Drugs or vehicle were given orally once a day for 60 days. After this, the animals were killed and the first pair of inguinal mammary glands was immediately removed; part of the material was processed for routine histological study and the remaining tissue was frozen for further analyses of the expression of VEGF mRNA by means of the RT-PCR technique. RESULTS: We observed that mammary ducts were atrophic in the control (CON) and isoflavone-treated (ISO) groups. In these groups the mammary glands were composed of a large concentration of adipose tissue with some ducts and rare alveolar structures. In the CEE group the ducts were well developed with many buds and alveolar structures. The number of mammary gland alveoli was higher in CEE than in the other groups (CON = 1.4 ± 2.1; ISO = 1.6 ± 3.8; CEE = 12.3 ± 7.1 alveoli/mm2; p<0.05%); also, the cell volume was higher (CON = 14.9 ± 4.9; ISO = 11.4 ± 6.9; CEE = 27.4 ± 9.7 µm3, p< 0.05%). The same was observed with regard to the number of blood vessels (CON = 16.4 ± 1.5; ISO = 18.4 ± 2.1; CEE = 37.1 ± 4.1 vessels/mm2, p< 0.05). The expression of VEGF in the CEE group was higher than in the other groups, which did not significantly differ from each other in this respect. CONCLUSION: Our data did not show any proliferation effect in the mammary tissue of adult oophorectomized rats treated with isoflavones (100 mg/kg) during 60 days.

Efeitos dos fitoestrogênios sobre alguns parâmetros clínicos e laboratoriais no climatério

Purpose: to evaluate the estrogenic effects of soy phytoestrogens in postmenopausal patients. Method: we selected postmenopausal patients from the Climacteric Service of the Gynecology Department of the Federal University of Sao Paulo - Escola Paulista de Medicina. The patients were divided into two groups: GI (n=40) received daily 100 mg isoflavone (3 x 33.3 mg/day); and GCtr (n=40) received daily placebo. We included in this study the Kuppermann menopausal index, physical examination and the biochemical determination of total and fractions of cholesterol, triglycerides, estradiol, FSH e LH. We also performed sonographic examination to measure endometrial thickness. To compare baseline and after treatment values of both groups, the nonparametric Kruskal-Wallis test was used for the Kuppermann index, while the t test was performed for other data. Results: evaluation of the Kupperman Menopausal Index, 80% patients of GI showed a decrease in the total score, while the improvement of symptoms in GCtr was 12,5%. The analysis of total cholesterol levels showed that 35 patients of GI had levels above 200 mg/dL, and only 13 of GCtr presented similar levels. The body mass index reduction occurred in GI and the control group showed no changes. We also verified that echo endometrial thickness and FSH levels were similar to baseline values. However the values of estrogen increased after isoflavone treatment. Conclusion: we concluded that 100 mg/day isoflavone may be an alternative treatment for attenuating the climacteric symptoms in 80% of isoflavone users and that it reduced total cholesterol.

Morfologia das células intersticiais de ovários policísticos de ratas: um estudo experimental

PURPOSES: To evaluate the histomorphometry of ovarian interstitial cells, as well as the blood sex steroid concentrations of female rats with polycystic ovaries induced by continuous light. METHODS: Twenty female rats were divided into two groups: Control Group - in the estrous phase (CtrlG), and a group of rats with polycystic ovaries induced by continuous illumination (POG). CtrlG animals were maintained on a light period from 07:00 a.m. to 07:00 p.m., and POG animals with continuous illumination (400 Lux) for 60 days. After this period all animals were anesthetized and blood was collected for the determination of serum estradiol (E2), progesterone (P4), and testosterone (T), followed by removal of the ovaries that were fixed in 10% formalin and processed for paraffin embedding. Five-µm histological sections were stained with hematoxylin and eosin and used for histomorphometric analysis. Morphological analyses, cyst count, determination of concentration and of the nuclear volume of interstitial cells were performed with the aid of a light microscope adapted to a high resolution camera (AxioCam), whose images were transmitted to and analyzed by the computer using AxioVision Rel 4.8 software (Carl Zeiss). Data were analyzed statistically by the Students t-test (p CtrlG=73.2±6.5, p CtrlG=80.6±3.9, p POG=4.2±1.5, p CtrlG=63.6±16.5, p CtrlG=6.9±3.2, p<0.05) compared to CtrlG animals. CONCLUSION: The interstitial cells of the rat polycystic ovary probably originate from ovarian cysts due to the degeneration of granulosa cells and differentiation of the internal theca cells. The elevations of serum testosterone and estradiol were probably due to the significant increase in cell activity and in the area occupied by interstitial cells.