Association between JMJD1A expression and sperm retrieval in non-obstructive azoospermic patients

Identification of molecular markers which can predict the outcome of sperm retrieval non-invasively in patients with non-obstructive azoospermia [NOA] are valuable in clinical andrology. Jumonji domain-containing 1a [JMJD1A] is a significant epigenetic regulator during spermatogenesis, which plays an important role in the differentiation of post-meiotic germ cells into mature spermatozoa. We therefore aimed to examine the potential association between JMJD1A expression and the outcome of sperm retrieval in patients with NOA. Testicular biopsy specimens from 50 NOA patients with either successful sperm retrieval [sperm+, n=22] or failed sperm retrieval [sperm-, n=28] were collected and then examined for JMJD1A expression by reverse transcription-quantitative polymerase chain reaction [RT-qPCR]. In addition, conventional clinical parameters including luteinizing hormone, follicle-stimulating hormone, testosterone, age, and testicular volume were compared between the two NOA groups. The expression of JMJD1A in the sperm+ group was significantly higher than in the sperm- group [P<0.001], however, no significant difference was observed between the two groups in clinical parameters. The receiver operating characteristic [ROC] curve of JMJD1A expression in predicting the sperm retrieval outcome showed a sensitivity of 90.91% and a specificity of 89.29% with significant discriminatory ability between the sperm+ and sperm- groups [area under the ROC curve [AUC]= 0.91]. This study demonstrates a significant association between the expression of JMJD1A and the success of sperm recovery in patients with NOA, and thus suggests that JMJD1A expression quantification in testicular biopsies may be a valuable biomarker along with conventional parameters in predicting the presence of spermatozoa

[Introduction of a new method for mono-specific antibody production by sequential use of recombinant proteins and synthetic peptides [PrIPeP model]]

Objective: This study attempted to generate monospecific antibodies through immunization with recombinant proteins and subsequent purification by synthetic peptides [the PrIPeP model]

Methods: The SRY gene was cloned on a pet-28a vector and the recombinant protein was expressed in the Escherichia coli [E.coli] BL21 strain. The purified antigen was emulsified in Freund's adjuvant and injected into rabbits according to a standard time table. Then, a specific peptide was designed, synthesized, and conjugated to sepharose 4B to generate an affinity purification column. As a control, the peptide was conjugated to KLH and used for immunization, as above. Antisera against the conjugated peptide [Pepantisera] and SRY recombinant protein [Pro-antisera] were evaluated by ELISA and subsequently subjected to the affinity purification column. Sensitivity and specificity of the purified antibodies against SRY recombinant protein as well as negative controls [recombinant HSFY, RBMY, and RPSFY] were assessed by Western blot analysis

Results: Titration by ELISA confirmed proper immunization and specificity of both antigens. Western blot analysis validated the specificity and sensitivity of the IgG class purified antibodies

Conclusion: By applying the PrIPeP model, it is possible to develop antibodies against the native structure of a protein whilst avoiding challenges of peptide-carrier protein conjugation

Predictive factors of successful microdissection testicular sperm extraction in patients with presumed sertoli cell-only syndrome

To evaluate predictive factors of successful microdissection-testicular sperm extraction [MD-TESE] in patients with presumed Sertoli cell-only syndrome [SCOS]. In this retrospective analysis, 874 men with non-obstructive azoospermia [NOA], among whom 148 individuals with diagnosis of SCOS in prior biopsy, underwent MD-TESE at Department of Andrology, Royan Institute, Tehran, Iran. The predictive values of follicle stimulating hormone [FSH], luteinizing hormone [LH], and testosterone [T] levels, testicular volume, as well as male age for retrieving testicular sperm by MD-TESE were analyzed by multiple logistic regression analysis. Testicular sperm were successfully retrieved in 23.6% men with presumed SCOS. Using receiver operating characteristic [ROC] curve analysis, it was shown that sperm retrieval rate in the group of men with FSH values >15.25% was 28.9%. This was higher than the group of men with FSH

Enzymatic digestion improves testicular sperm retrieval in non-obstructive azoospermic patients

In patients with non-obstructive azoospermia [NOA], vital spermatozoa from the tissue is obtained from testes by enzymatic treatment besides the mechanical treatment. To increase the sperm recovery success of testicular sperm extraction [TESE], with enzymatic digestion if no sperm is obtained from testis tissue by mechanical method. Tissue samples were collected from 150 men who presented with clinical and laboratory data indicating NOA by means of TESE and micro dissection TESE methods. Initially, mature spermatozoa were examined for by mechanical extraction technique shredding the biopsy fractions. In cases whom no spermatozoa was observed after maximum 30 min of initial searching under the inverted microscope, the procedure was followed by enzymatic digestion using DNaseI and collagenase type IV. Surgery type, pathology, AZF, karyotype, hormones and testis size were compared in patients. Of 150 cases with NOA, conventional mincing method extended with enzymatic treatment yielded successful sperm recovery in 13 [about 9%] patients. Comparison of parameters revealed that level of FSH and LH were significantly different [p=0.04 and 0.08 respectively] between two groups that response negative and positive to enzymatic digestion. The combination of conventional TESE and enzymatic digestion is an effective method to recover spermatozoa. The benefit of the mincing combined with enzyme to sperm retrieval for NOA firstly shorten the mechanical searching time, leading to minimizing further cellular damage as well as exposure to external conditions, and secondly reduce the number of cases with sperm recovery failures. Also, the serum level of FSH and LH are factors that influence the chance of sperm retrieval

[Predictive value of semen parameters and age of the couple in pregnancy outcome after Intrauterine insemination]

Intrauterine insemination [IUI] is one of the most common methods in infertility treatment, but its efficiency in infertile couples with male factor is controversial. This study is a retrospective study about correlation between semen parameters and male and female age with successful rate of IUI in patients attending to Royan Institute. A total of 998 consecutive couples in a period of 6 months undergoing IUI were included. They were classified into two groups: couples with successful and unsuccessful pregnancy. Main outcome was clinical pregnancy. Data about male and female ages and semen analysis including concentration, total sperm motility, class A motility, class B motility, class A+B motility and normal morphology was extracted from patients' records. Semen samples were collected by masturbation or coitus after 2 to 7 days of abstinence. Their female partners were reported to have no chronic medical conditions and have normal menstrual cycles. One hundred and fifty seven of total 998 cycles [15.7%] achieved pregnancy. The average of female age in successful and unsuccessful group was 28.95 +/- 4.19 and 30.00 +/- 4.56 years, respectively. Mean of male age was 33.97 +/- 4.85 years in successful group and 34.44 +/- 4.62 years in unsuccessful group. In successful and unsuccessful groups, average of sperm concentration was 53.62 +/- 38.45 and 46.26 +/- 26.59 [million sperm/ml], normal morphology of sperm was 8.98 +/- 4.31 [%] and 8.68 +/- 4.81 [%], sperm total motility was 47.24 +/- 18.92 [%] and 43.70 +/- 20.22 [%] and total motile sperm count was 80.10 +/- 63.61 million and 78.57 +/- 68.22 million, respectively. There was no significant difference in mean of females' age and males' age between successful and unsuccessful groups [P<0.05]. In addition, there was no significant difference in semen parameters including concentration, total sperm motility, class A motility, class B motility, class A+B motility and normal morphology between two groups. It was shown that common semen analysis and male and female ages cannot predict IUI outcome.

Effects of sperm chromatin integrity on fertilization rate and embryo quality following intracytoplasmic sperm injection

Sperm chromatin integrity has been being recognized as an important factor in male fertility. During normal fertilization, high quality sperm with intact chromatin are selected through natural selection in journey from vagina to fallopian tube. However, using Assisted Reproductive Techniques, particularly ICSI, the natural selection is bypassed. Therefore sperm with DMA breakage have the opportunity to fertilize the egg which may lead to decreased embryo quality and implantation rate. The aim of this study was to evaluate the effects of sperm chromatin integrity on ICSI outcomes. A total of 200 semen samples were collected from couples undergoing ICSI and were analyzed according to WHO criteria. Each sample was evaluated for sperm chromatin integrity using four cytochemical assays and semen processing by swim up method. The ICSI was carried out according to a long-term pituitary down-regulation protocol. The correlation between sperm parameters, sperm chromatin integrity and ICSI outcomes [fertilization rate and embryo quality] was examined. The mean number of oocyte, fertilization rate and cleavage embryos per cycles was 7.5 +/- 5.0, 74.O6% +/- 25 and 5.4 +/- 3.6, respectively. There was not significant correlation between the results of chromatin assays [AO, AB, TB, and CMA3] and fertilization outcomes following ICSI. The fertilization rate was significantly higher for a group with less than 10% chromatin abnormality [p<0.05]. Sperm chromatin integrity is essential for successful fertilization, embryo development and normal pregnancy. A protamine deficiency appeared to affect fertilization rate and embryo quality. However, the presence of confounding factors such as selection of spermatozoa according to normal morphology may influence the effect of sperm chromatin status on ICSI outcomes