Semen Analysis of Total Motile Sperm Count Based on the 1999 and 2010 WHO Criteria.

OBJECTIVE: Approximately 15% of the couples suffer from infertility. Half of the cases of infertility are due to male factors. Several sperm function tests have been proposed to evaluate male fertility, but sperm analysis is still the first and most important diagnostic test for male infertility. The prognostic value of semen characteristics such as concentration, morphology and motility markers are often confused with male infertility. Evaluation of seminal parameters and classification for normality remains a frequent topic of discussion. METHODS: This study evaluated 477 semen samples from men undergoing investigation or infertility treatment between 2011 and 2015. RESULTS: The spermograms of 401 patients were deemed abnormal based on the 1999 World Health Organization (WHO) criteria; the number changed to 223 when the spermograms were assessed based on the 2010 WHO criteria and to 200 when Total Motile Sperm Count (TMSC) was used as the criterion. Sperm morphology was the item in the criteria that most significantly changed spermogram classification. Normality parameters became less rigid from 1999 to 2010, thereby significantly changing the proportion of individuals no longer described as infertile/subfertile. CONCLUSIONS: The classification based on TMSC could not differentiate between fertile and infertile subjects for not taking sperm morphology into account. Nevertheless, it may be helpful in cases where intrauterine insemination is indicated.

Safety of brilliant cresyl blue staining protocols on human granulosa and cumulus cells.

The selection of human immature oocytes destined for in vitro maturation (IVM) is performed according to their cumulus-oocyte complex (COC) morphology. In animal models, oocyte pre-selection with brilliant cresyl blue (BCB) staining improves fertilization and blastocyst rates and even increases the number of calves born. As the granulosa cells and cumulus cells (GCs and CCs) have a close relationship with the oocyte and are available in in vitro fertilization (IVF) programs, applying BCB staining to these cells may help to elucidate whether BCB shows toxicity to human oocytes and to determine the safest protocol for this dye. GCs and CCs were isolated from 24 patients who underwent controlled ovarian stimulation. After 48 h, cells were exposed to: Dulbecco's Modified Eagle Medium (DMEM) with or without phenol red, DPBS and mDPBS for 60 min; 13, 20 and 26 µM BCB for 60 min; and 60, 90 or 120 min to 13 µM BCB. Cellular viability was tested using 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) and trypan blue assays. The 20 and 26 µM BCB exposures resulted in lower cell viability, similar to when cells were exposed to BCB for 90 or 120 min. GCs and CCs viabilities were equal among control group and 13 µM BCB group after 60 min. BCB staining was not toxic to GCs and CCs when the regime of 13 µM BCB for 60 min was used. Due to the close molecular/biochemical relationship between these cells and the gamete, we propose that it is unlikely that the use of BCB could interfere with the viability/health of human oocytes.

Adequacy of ovarian diathermy under ultrasound control: an experimental model.

BACKGROUND: To develop a minimally invasive ovarian cauterization technique under transvaginal ultrasound control and evaluate the safety and feasability of monopolar cauterization to cause ovarian injury using female cattle of reproductive age as an experimental model. METHOD: Experimental study in a university research center was performed. Eleven female bovines of reproductive age were submitted to monopolar transvaginal ovarian cauterization. The right ovary (RO) was punctured at four sites and 40 W was applied for 5 s at each point, resulting in a total of 800 J (Joules) of thermal energy. In the left ovary (LO), the procedure was similar, with the same time and 80 W, resulting in a thermal energy of 1600 J. Macroscopic and microscopic lesions were assessed. RESULTS: Of 22 ovaries punctured, 20 were cauterized and exhibited macroscopic and typical microscopic lesions. No lesions could be found in the needle path. The measures of the areas of microscopic electrocautery lesions calculated estimating a cylindrical volume showed a median of 1.12% in the right ovary and 1.65% in the left ovary. When the estimate was calculated by spherical shape, the medians were 1.77% in the right ovary and 3.06% in the left ovary. There was a statistically significant difference in these two estimates (sphere, p = 0.008; cylinder, p = 0.021). CONCLUSION: The experimental animal model described for transvaginal ultrasound-guided ovarian needle cauterization seems to be feasible. The ovaries were successfully cauterized without injuries in needle path and more energy resulted in significantly more thermal lesion. The safety and effectiveness of this technique, theoretically less invasive than current ovarian drilling methods, could be tested in anovulatory women with PCOS.

Proporção macho: fêmea de embriões bovinos cultivados na presença ou ausência de glicose após FIV com espermatozóides selecionados por Swim-up ou Gradiente de Percoll

No sistema de PIV em bovinos, tem sido obtida uma elevada porcentagem de embriões machos. Este experimento foi realizado para determinar se a presença de glicose no meio de cultivo afeta a proporção macho:fêmea (M:F) dos embriões bovinos PIV a partir da FIV com espermatozóides preparados pelos métodos do swim-up (S) ou do gradiente de Percoll (P). Após a MIV, os COCs foram divididos em dois grupos e inseminados com espermatozóides preparados por um dos métodos. Os zigotos foram cultivados em meio com ou sem 5,56mM de glicose, totalizando 4 tratamentos: S-Gli, S+Gli, P-Gli e P+Gli e 48h após a inseminação, os embriões de cada tratamento foram submetidos à sexagem por PCR (n=845). O efeito da glicose no meio de cultivo sobre a proporção M:F dos embriões PIV a partir dos dois métodos foi semelhante (teste do c²), resultando em uma porcentagem de machos menor do que 50 por cento no estágio de 2-C (S: 30,8 por cento; P: 23,8 por cento: P<0,01) e maior do que 50 por cento no estágio de 8-C (S: 79,4 por cento; P: 68,8 por cento: P<0,01). Estas porcentagens foram diferentes (P<0,05) das observadas quando os embriões foram cultivados sem glicose, tanto no estágio de 2-C (S: 48,5 por cento; P: 41,5 por cento) como no de 8-C (S: 62,5 por cento; P: 50,8 por cento). A presença de glicose não afetou a proporção M:F no total de embriões produzidos (S: 56,7 por cento; P: 49,0 por cento), que foi semelhante à observada na ausência de glicose (S: 55,7 por cento; P: 46,2 por cento). Portanto, a glicose exacerbou a diferença na velocidade de desenvolvimento entre os embriões machos e fêmeas.