Long-term treatment of familial male-limited precocious puberty (testotoxicosis) with cyproterone acetate or ketoconazole

Background Familial male-limited precocious puberty (FMPP) or testotoxicosis is a rare gonadotrophin-independent form of sexual precocity caused by constitutively activating mutations of the LH receptor. Several clinical therapeutic approaches have been reported for this disorder, but with a paucity of long-term outcome data. Objective To evaluate the long-term treatment of testotoxicosis with cyproterone acetate or ketoconazole. Design A multicentric retrospective clinical study. Patients Ten boys from eight unrelated Brazilian families who carried known LH-receptor activating mutations were treated with 70 mg/m(2) cyproterone acetate (n = 5) or 10 mg/kg ketoconazole (n = 5) for a mean period of 5 and 8 years, respectively. Measurements Chronological and bone ages, bone age/chronological age ratio, target height (TH) range, adult height, basal and GnRH-stimulated gonadotrophin levels and basal testosterone levels were assessed. Results Growth velocity decreased significantly during treatment with cyproterone acetate or ketoconazole when compared to pretreatment value in each group (P < 0.05). Bone age/chronological age ratio decreased significantly after cyproterone acetate or ketoconazole therapy. Basal testosterone levels were significantly lower in patients undergoing ketoconazole compared to cyproterone acetate treatment [0.6 +/- 0.3 nmol/l (42 +/- 21 ng/dl) vs. 5.6 +/- 4.0 nmol/l (392 +/- 280 ng/dl); P < 0.05], as expected. Secondary gonadotrophin-dependent precocious puberty occurred at a similar frequency (40%) in both groups. Five patients have attained adult height and two patients have already reached 90% of their adult height. Two of them achieved their TH range and one patient, for whom TH was not available, had an adult height of 0.3 SDS. Four boys (two in each group) did not attain their TH range. Conclusion Long-term treatment with cyproterone acetate or ketoconazole resulted in similar outcomes without important side-effects in boys with testotoxicosis. However, both therapies showed limited efficacy in attaining normal adult height.

Effect of peritoneal fluid from patients with minimal/mild endometriosis on progesterone release by human granulosa-lutein cells obtained from infertile patients without endometriosis: A pilot study

Objective: To evaluate the effect of peritoneal fluid (PF) from women without and with minimal/mild endometriosis on progesterone (P) release by cultured human granulosa-lutein cells obtained from infertile patients without endometriosis submitted to ovarian hyperstimulation for in vitro fertilization (IVF). Study design: A pilot study was performed. Human granulosa-lutein cells, obtained from 11 infertile patients without endometriosis (tubal or male factors of infertility) submitted to ovarian hyperstimulation for IVF, were cultured without PF (basal production) and with increasing volumes of steroid-extracted PF samples from 11 patients with endometriosis and 11 patients without endometriosis. Progesterone (P) levels in the media after 72 h culture were measured by chemoluminescence assay. The non-parametric Mann-Whitney-test was used for statistical analysis. Results: PF from patients without endometriosis stimulated P release in a dose-dependent manner up to the dose of 100 mu l/ml (10% concentration) when compared with basal production (without adding PF). P release was similar in cultures stimulated with PF from patients with or without endometriosis at 1% (10 mu l/ml) and 5% (50 ml/ml) concentrations. At 10% concentration, there was a non-statistically significant reduction in progesterone release by granulosa cells stimulated with PF from patients with endometriosis. PF from patients with endometriosis significantly reduced P release at 30% concentration (300 mu l/ml). Conclusions: PF stimulates P release by human granulosa-lutein cells in a dose-dependent manner. However, higher concentrations of PF from patients with minimal/mild endometriosis reduce P release, suggesting it contains factors that may compromise ovarian steroidogenesis. (c) 2007 Elsevier Ireland Ltd. All rights reserved.

Influence of aging, tidal volume, and respiratory rate on Doppler echocardiographic indices of diastolic function in normal male subjects: A quantitative evaluation

Background: Although the influence of respiration on ventricular filling, as evaluated by Doppler technique, and the evaluation of diastolic velocities of mitral valve annulus (MVA), as measured by Doppler tissue imaging (DTI), can provide valuable information for the study of left ventricular (LV) diastolic function, the concomitant effects of aging, tidal volume (TV), and respiratory rate (RR) on these velocities have not been quantitatively investigated. Methods: We evaluated 12 normal male volunteers (Group I) aged 20-26 years (mean: 22.8) and 8 normal subjects aged 41 to 54 years old (mean: 45.9) (Group II). Using DTI we measured peak early (E-a) and late (A(a)) velocities of longitudinal axis expansion at lateral and medial MVA. Doppler mitral and tricuspid flow velocities were measured: peak early (E) and late (A) inflow velocity, early (E-i) and late (A(i)) flow integral, and deceleration time of peak early mitral flow velocity (DT). Respiratory cycles were simultaneously recorded at RR of 9, 12, 15, and 18 cycles/min and TV of 600 and 900 mL during respiration (RESP). Results and conclusions: (1) E, A, and A(i) in MV had negligible change during respiration, but E-i was significantly reduced during inspiration; (2) DT reduced slightly with inspiration, but the change was significant only with TV of 900 mL; (3) an important increase of E in right ventricular flow was observed during inspiration; (4) variations of RR and TV did not significantly influence right and left ventricular inflow in normal subjects, in the conditions of this investigation; (5) a significant increase of E-a at medial MVA was documented during inspiration only in young subjects; (6) a significant decrease of A(a) at medial MVA was observed during inspiration in both groups of volunteers; (7) RR and TV did not influence MVA velocities in young and adult subjects; (8) a consistent reduction in E-a and a significant increase in A(a) were observed with increasing age; (9) these changes were more conspicuous and consistent than those documented in ventricular filling when young and middle-age men are compared, suggesting that the DTI is more sensitive to detect changes in diastolic function; and (10) in addition, these data suggest that, for evaluation of diastolic function, in clinical context, it is not necessary to control rigorously RR or TV.

Histologic Study of Urethral Extracellular Matrix and Collagen From Aging and Long-Term Alloxan-induced Diabetic Male Rats

OBJECTIVES To evaluate the histological alterations of extracellular matrix in long-term alloxan-induced diabetes and aging urethras of male rats with descriptions of total connective tissue, muscle layer and collagen types I and III relative amounts. METHODS Histologic evaluations were performed in 3 animal groups: group 1, 8 weeks old; group 2, 44 weeks old; and group 3, 44 weeks old with alloxan-induced diabetes. The muscle layer thickness, extracellular matrix fibrosis, and collagen were quantified on digital images of the urethral samples. RESULTS A higher total thickness and muscle layer thickness and higher connective tissue and collagen content were observed in the urethras of group 3. No changes in the collagen type III/I ratio were found in the urethra of groups 2 and 3. CONCLUSIONS Our results suggest that the morphologic alterations of the urethra should also be considered in long-term studies of diabetic lower urinary tract dysfunction. These morphologic alterations due to diabetes differ from the changes induced by aging itself and could represent a final stage in decompensate urethras. Further studies are necessary to establish the real influence of the urethral morphologic changes on lower urinary tract diabetes dysfunction. UROLOGY 77: 510.e6-510.e11, 2011. (C) 2011 Elsevier Inc.

Isolation and genetic characterisation of Toxoplasma gondii from a red-handed howler monkey (Alouatta belzebul), a jaguarundi (Puma yagouaroundi), and a black-eared opossum (Didelphis aurita) from Brazil

Toxoplasma gondii isolates are highly diverse in domestic animals from Brazil. However, little is known about the genetics of this parasite from wild mammals in the same region. Reveal genetic similarity or difference of T. gondii among different animal populations is necessary for us to understand transmission of this parasite. Here we reported isolation and genetic characterisation of three T. gondii isolates from wild animals in Brazil. The parasite was isolated by bioassay in mice from tissues of a young male red handed howler monkey (Alouatta belzebul), an adult male jaguarundi (Puma yagouaroundi), and an adult female black-eared opossum (Didelphis aurita). The monkey and the jaguarundi had inhabited the Zoo of Parque Estadual Dois Irmaos, Pernambuco State, Northeastern Brazil, for 1 year and 8 years, respectively. The wild black-eared opossum was captured in Sao Paulo State, Southeastern Brazil, and euthanised for this study because it was seropositive for T. gondii (titre 1:100 by the modified agglutination test, MAT). Ten PCR-RFLP (Polymerase Chain Reaction-Restriction Fragment Length Polymorphism) markers, SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico, were used to genotype the isolates. T. gondii was isolated from the brain and heart homogenate of the monkey, the muscle homogenate of the jaguarundi, and the heart homogenate of the black-eared opossum. This was the first isolation of T. gondii from a neotropical fetid from Brazil. The isolate from the monkey (TgRhHmBr1) was not virulent in mice, whereas the isolates from the jaguarundi (TgJagBr1) and the black-eared opossum (TgOpBr1) were virulent in mice. The genotype of the isolate from the monkey has been identified in isolates from a goat and ten chickens in the same region of Brazil, suggesting that it may be a common lineage circulating in this region. The genotypes of the isolates from the jaguarundi and the black-eared opossum have not been previously reported. Although there are already 88 genotypes identified from a variety of animal hosts in Brazil, new genotypes are continuously being identified from different animal species, indicating an extremely high diversity of T. gondii in the population. (C) 2010 Elsevier B.V. All rights reserved.

Correlation between mastitis occurrence and the count of microorganisms in bulk raw milk of bovine dairy herds in four selective culture media

Milk is the normal secretion of the mammary gland, practically free of colostrum and obtained by the complete milking of oil(? or more healthy animals. Mastitis is all inflammatory process of the mammary gland and it may cause alterations in the milk. The present work aimed to verify whether it is possible, by means of the counts of microorganism in the bulk raw milk ill four selective culture media, to establish a correlation with the occurrence of mastitis and therefore, to monitor this disease in bovine dairy herds. The following selective culture media were Used: KF Streptococcus Agar, Edwards Agar, Baird-Parker Agar, Blood Agar plus potassium tellurite. Spearman`s correlation coefficient was calculated in order to compare the Occurrence of mastitis (percentage) in each herd with respective selective culture media counts of microorganisms in bulk raw milk. Thirty-six possibilities were analysed (Tamis and CMT-positive rates were compared with the log-transformed count in four selective Culture media) and there was a negative correlation between Tamis 3 and the Baird-Parker Agar plate count. The total results of microbiological tests showed that there were three correlations of the counts in selective culture media. Fifty-two possibilities were analysed and there was a negative correlation between no-bacteria I-growth mastitis rates and log(10) of KF Streptoccocus Agar plate Count and there were two positive correlations between coagulase-positive staphylococci and log(10) of Baird-Parker Agar plate count and Blood Agar plus potassium tellurite plate count.

Prenatal lipopolysaccharide reduces motor activity after an immune challenge in adult male offspring

Prenatal lipopolysaccharide (LPS) exposure causes reproductive, behavioral and neurochemical injuries in both the mother and pups. Previous investigations by our group showed that prenatal LPS administration (100 mu g/kg, i.p.) on gestational day 9.5 impaired the male offspring`s social behavior in infancy and adulthood. In the present study, we investigated whether these social behavioral changes were associated with motor activity impairment. Male rat pups treated prenatally with LPS or not were tested for reflexological development and open field general activity during infancy. In adulthood, animals were tested for open field general activity, haloperidol-induced catalepsy and apomorphine-induced stereotypy; striatal dopamine levels and turnover were also measured. Moreover, LPS-treated or untreated control pups were challenged with LPS in adulthood and observed for general activity in the open field. In relation to the control group, the motor behavior of prenatally treated male pups was unaffected at basal levels, both in infancy and in adulthood, but decreased general activity was observed in adulthood after an immune challenge. Also, striatal dopamine and metabolite levels were decreased in adulthood. In conclusion, prenatal LPS exposure disrupted the dopaminergic system involved with motor function, but this neurochemical effect was not accompanied by behavioral impairment, probably due to adaptive plasticity processes. Notwithstanding, behavioral impairment was revealed when animals were challenged with LPS, resulting in enhanced sickness behavior. (C) 2010 Elsevier B.V. All rights reserved.

Prenatal Lipopolysaccharide Reduces Social Behavior in Male Offspring

Objective: This study investigated the relationship between maternal sickness behavior during pregnancy and offspring development and behavior. Methods: Pregnant Wistar rats were administered with lipopolysaccharide (LPS, 100 mu g/kg, i.p.) on gestation day (GD) 9.5. Dams` sickness behavior was analyzed, and at birth, offspring number and weight were evaluated. Male offspring was evaluated through physical development, play behavior, adult social interaction, plus maze studies and morphological analysis of the brain. Results: Results, with respect to the control group, showed that: (1) LPS decreased general activity, food intake, and weight gain in dams, but no pyrexia was observed following treatment; (2) LPS reduced litter size, but no alterations in physical development were observed; (3) LPS reduced play behavior parameters in baby rats; (4) LPS decreased adult social interaction; (5) no alterations were observed between groups on plus maze studies; (6) no differences were observed between groups on morphological analyses of the brain. Conclusion: These data reveal that LPS administered on GD 9.5 impaired male offspring`s social behavior in infancy and adulthood. These results may be related to an alteration in motivational states or/and increased anxiety. Copyright (C) 2010 S. Karger AG, Basel

Prenatal Lipopolysaccharide Exposure Affects Maternal Behavior and Male Offspring Sexual Behavior in Adulthood

Objective: This study investigates the effects of prenatal lipopolysaccharide (LPS) exposure on the maternal behavior of pregnant rats and the physical development and sexual behavior of their male offspring in adulthood. Methods: For two experiments, pregnant rats were injected with LPS (250 mu g/kg, i.p.) on gestation day (GD) 21. In the first experiment, the maternal behavior (postnatal day, PND, 6) and the dam`s open-field general activity (PND7) were evaluated. In the second experiment, the maternal pre- and postnatal parameters, the pup`s development, the offspring`s sexual behavior in adulthood, and the pup`s organ weights were assessed. Results: Compared to the control group, the LPS-treated dams presented reduced maternal behavior, decreased general activity, a smaller body weight difference between GD21 and PND1, a greater number of perinatal deaths, and smaller litters. For the male pups, LPS treatment resulted in a decreased body weight on PND2, whereas the anogenital distance and the day of testis descent were not modified. The male sexual behavior was impaired by prenatal LPS. Particularly the number of ejaculating animals was reduced. The testis weight was also lower in the prenatally LPS-treated rats than in the control rats. Conclusion: We propose that prenatal LPS exposure on GD21 acts as an imprinting factor that interferes with the programming of brain sexual determination in offspring. Copyright (C) 2009 S. Karger AG, Basel

Ivermectin impairs sexual behavior in sexually naive, but not sexually experienced male rats

Ivermectin (IVM) is an antiparasitic drug, widely used in domestic animals. In mammals, IVM act as a GABA agonist. This neurotransmitter has an important role in the regulation of sexual behavior. Thus, this study sought to investigate the effects of various medically relevant doses IVM on the sexual behavior of male rats. In particular, we also wished to examine if previous sexual experience modulated responses to IVM. In the first experiment, the sexual behavior of inexperienced male rats was analyzed after they received 0.2, 0.6, 1.0 or 2.0 mg/kg IVM, 15 mm prior to behavioral testing. In the second experiment, the effects of four previous sexual experiences on IVM treated rats (1.0 or 2.0 mg/kg, 15 min prior to the 5th session) were assessed. The standard therapeutic dose (0.2 mg/kg) did not impair the sexual behavior of inexperienced male rats. At a more concentrated dose (0.6 mg/kg), which is still within the therapeutic range, the appetitive phase of sexual behavior of inexperienced male rats was impaired. Likewise, 1.0 mg/kg impaired the appetitive phase. Previous sexual experience blocked almost entirely this sexual impairment, suggesting that previous sexual experience exerts a positive effect in attenuating the sexual impairment produced by IVM treatment. Therefore, the standard therapeutic dose of IVM can be used without producing side effects on sexual behavior. Use of more concentrated therapeutic doses is not recommended during reproductive periods, unless the animals have had previous sexual experience. (C) 2010 Elsevier Ltd. All rights reserved.

Moxidectin interference on sexual behavior, penile erection and hypothalamic GABA levels of male rats

The moxidectin (MXD) is an antiparasitic drug used in domestic animals. The mechanism of action, in mammals, involves GABA, a neurotransmitter with an important role in the sexual behavior control. Presently, the effects of 0.2 mg/kg therapeutic dose were studied on sexual behavior, sexual motivation, penile erection and central GABA levels. Sexual behavior results showed increased latencies to the first mount and intromission as well as in inter-intromission interval; a reduction in total mounts was detected on the drug post-treatment. No difference was observed between sexual motivation of control and experimental animals. MXD treatment reduced penile erection and hypothalamic GABA levels. The results suggest that MXD reduced sexual behavior and penile erection by an action on the hypothalamic GABA system. Probably, the lack of effects in the motivational test and the increased mount and intromission latencies as well as decreased total mounts could be explained as a consequence of reduced male rat erection process. (C) 2007 Elsevier Ltd. All rights reserved.

Serum Starvation and Full Confluency for Cell Cycle Synchronization of Domestic Cat (Felis catus) Foetal Fibroblasts

Nuclear transfer of domestic cat can be used as a tool to develop reproductive biotechnologies in wild felids. The importance of cell cycle phase during the nuclear transfer has been a matter of debate since the first mammalian clone was produced. The cell cycle phase of donor cells interferes on maintenance of correct ploidy and genetic reprogramming of the reconstructed embryo. The use of G0/G1 arrested donor cells has been shown to improve nuclear transfer efficiency. The present study was conducted to test the hypothesis that domestic cat foetal fibroblasts cultured up to the fifth passage and submitted to full confluency provide a higher percentage of cells at G0/G1 stage than fibroblasts cultured in serum starved media. Results demonstrated that serum starvation increased (p < 0.05) the percentage of G0/G1 fibroblasts when compared with control. Moreover, the combined protocol using confluency and serum starvation was more efficient (p < 0.05) synchronizing cells at G0/G1 stage than serum starvation or confluency alone for the first 3 days of treatment. In conclusion, serum starvation and full confluency act in a synergistic manner to improve domestic cat foetal fibroblast cell cycle synchronization at the G0/G1 stage.

NONINVASIVE MONITORING OF ANDROGENS IN MALE AMAZONIAN MANATEE (TRICHECHUS INUNGUIS): BIOLOGIC VALIDATION

The Amazonian manatee (Trichechus inunguis) is endemic in the Amazonian basin and is the only exclusively fresh water sirenian. Historically hunted on a large scale, this species is now considered endangered, and Studies on the reproductive physiology are critical for the improvement of reproductive management of captive and wild Populations of manatees. The aim of this Study was to verify the viability of androgen measurement in saliva, lacrimal, urine, and fecal samples of the Amazonian manatee by conducting a hormone challenge. Two adult male manatees (A-1 and A-2) were Submitted to an experimentation protocol of 12 day (D1 to D10). On D0, the animals received an intramuscular injection of gonadotropin-releasing hormone (GnRH)-analogue. Salivary, lacrimal, urinary, and fecal samples were collected daily (between 0800 hours and 0900 hours) and frozen at -20 degrees C until assayed. Fecal samples were lyophilized, extracted with 80% methanol, and diluted in buffer before the radioimmunoassay (RIA). Urine samples underwent acid hydrolysis and were diluted in depleted bovine serum. Salivary and lacrimal samples were assayed without the extraction step. Hormonal assays were conducted with a commercial testosterone RIA kit. An androgen peak (>median + 2 interquartile range [IQR]) was observed in all matrices of both animals, although it was less prominent in the lacrimal samples of A-2. However, the fecal androgen peak (A-1 peak = 293.78 ng/g dry feces, median [IQR] = 143.58 [32.38] ng/g dry feces; A-2 peak = 686.72 ng/g dry feces, median [IQR] = 243.82 [193.16] ng/g dry feces) occurred later than urinary (A-1 peak = 648.16 ng/mg creatinine [Cr], median [IQR] = 23.88 [30.44] ng/mg Cr; A-2 peak = 370.44 ng/mg Cr, median [IQR] = 113.87 [117.73] ng/mg Cr) and salivary (A-1 peak = 678.89 pg/ml, median [IQR] = 103.69 [119.86] pg/ml; A-2 peak = 733.71 pg/ml, median [IQR] = 262.92 [211.44] pg/ml) androgen peaks. These intervals appear to be correlated with the long digesta passage time in this species. The salivary and urinary peaks were closely associated. These results demonstrate that androgen concentrations in saliva, urine, or feces samples reflect reliably physiologic events and are a powerful tool for noninvasive reproductive monitoring of Amazonian manatees.

Effect of culture media on porcine embryos produced by in vitro fertilization or parthenogenetic activation after oocyte maturation with cycloheximide

This study evaluated the effects of reversible meiotic inhibition and different culture media (PZM3 or NCSU23) on production of porcine embryos by either in vitro fertilization (IVF) or parthenogenetic activation (PA). Oocytes from abattoir-derived ovaries were allocated into two groups for maturation: CHX (5 mu g/ml cycloheximide for 10 h) or Control (no CHX). The percentage of metaphase II (MII) oocytes was determined at 36, 40 or 44 h of in vitro maturation. For IVF and PA, denuded oocytes were fertilized with purified sperm for 6 h or activated by electric stimuli. Zygotes were then subdivided into two culture groups: NCSU23 or PZM3. No effect of treatment with CHX and culture media was observed on cleavage (D3) and blastocyst (D7) rates in IVF and PA groups. There are no differences of quality or development rates between IVF-derived embryos cultured in NCSU23 or PZM3. However, we observed high quality PA embryos in PZM3 compared with NCSU23. Maturation arrest with CHX decreased the average blastocyst cell number in IVF while it was increased in PA embryos. As older oocytes are more effectively activated, CHX-blocked oocytes reached the mature stage faster than the control group. In conclusion, the CHX treatment for 10 h, followed by oocyte maturation for 40 h, is an efficient protocol to produce high quality parthenote embryos, especially when they are cultured in PZM3. However, this protocol is not satisfactory for IVF embryos production. In this case, a shorter maturation period could provide better embryo quality.

Redescription of the female, description of the male, and several new records of Amblyomma parkeri (Acari: Ixodidae), a South American tick species

The tick Amblyomma parkeri Fonseca and Arago was described in 1952, based on female and immature ticks collected in the states of So Paulo and Santa Catarina, Brazil. Thereafter, there has been no further report of A. parkeri, and the male has remained unknown. Herein, we examined ticks collected on porcupines from a locality in the state of So Paulo. Some of the ticks were identified as Amblyomma longirostre (Koch, 1844), whereas others as A. parkeri, including male specimens, for which we provide the first description. We also provide additional reports of A. parkeri after examining collections of A. longirostre and Amblyomma geayi Neumann, 1899 from different tick collections. Morphological evidence to support the original description of A. parkeri is presented, supported by molecular analyses of portions of the 16S rRNA and 12S rRNA mitochondrial genes. Morphological particularities to separate A. parkeri, A. longirostre, and A. geayi are provided.