Journal of Reproduction & Infertility
Year:2007 | Volume:7 | Issue:4 (29)|Papers Count:14

Introduction: Estrogen, as one of the most important female hormones, plays an essential role in fertility cycle and its receptors mediate estrogen actions in target tissues. Several polymer-phisms of estrogen receptor (ER) alpha gene have been associated with different pathological conditions. In this study, the hypothesis that polymorphisms in ER alpha gene in women may be associated with their ovarian response to ovulation stimulation during in-vitro fertilization (IVF) was evaluated. Materials & Methods: The prevalence of PvuII polymorphism of ER alpha gene in 102 infertile women undergoing IVF treatment was evaluated and its relationship to the mean number of follicles and oocytes, follicles to oocytes ratios, mean serum estrogen concent-rations, the number of obtained embryos and pregnancy rates was determined. In each patient PP, Pp, or pp genotypes were identified by using PCR-RLFP technique. Statistical analysis was done through χ², Kruskal Wallis and Mann-Whitney by using SPSS software, version 13. P-values smaller than 0.05 were considered significant.Results: Based on the results of the study, 31.4% of the patients showed pp, 39.2% Pp and 29.4% PP genotypes. The percentages of male and female infertility etiologies, mean number of follicles, oocytes, and embryos in all three groups had no significant differences. Conclusion: Investigating the role of different genes in response to different medications (Pharmacogenetics) is very important in choosing proper treatment methods. This study indi-cated the importance of one of the genes effective in ovarian response to ovulation induction in IVF treatments, but noticing the role of different endocrine and paracrine factors in ovulation induction and proper oocyte maturation, further studies are needed to find the importance of other effective genes in ovaries and to determine other ER mutations and prevalence of ER polymorphism in the fertile women's population.

Introduction: Considering the 5% prevalence of abortion in women, its devastating psycholo-gical effects on affected people’s lives and as tendency for clotting is one of the causes of recur-rent spontaneous abortion (RSA), this study was designed to compare plasminogen activator inhibitor-1 (PAI-1), angiotensin converting enzyme (ACE) and blood coagulation factor XIII (FXIII) gene polymorphisms in Iranian patients with RSA and healthy women.Materials & Methods: 120 patients with recurrent abortions (at least two) as cases and 112 healthy female controls without a history of abortion were entered into the study. In order to characterize PAI-1 (4G/5G), ACE (D/I) and FXIII (Val 34 Leu) polymorphisms, a polymerase chain reaction followed by digestion with restriction enzymes (PCR-RFLP) was designed. For the statistical analysis, SPSS software version 11.2 was used and t-test, Chi-square and Fisher’s exact tests were calculated.  P-values <0.05 were considered as significant.Results: Homozygosity for PAI-1 4G polymorphism was seen in 16 cases, (14.4%), in contrast to two persons in the control group (2%), (p = 0.001) and patients with homozygote 4G mutation were significantly more prone to RSA in contrast to others (OR: 8.2, % 95 CI: 1.8- 36.5). 38 patients, (29.5 %), and 25 people from the control group, (26.6%), were homozygote (D/D) for ACE polymorphism, depicting no statistically significant difference. Only two patients and one person from the control group had homozygosity (34leu) for FXIII polymorphism.Conclusion: 4G/4G polymorphism for PAI-1 gene could be a thrombophilic mutation leading to abortion. Analysis of this mutation and other suspected factors such as MTHFR and FV Leiden is recommended in patients with RSA. In this study, there were no significant associations between ACE and FXIII with RSA.

Introduction: Using Assisted Reproductive Techniques (ART) may result in the birth of a neonate and help solve the infertility problem. However, 10-15% of oocytes fail to fertilize and this is called a Total Fertilization Failure (TFF). This failure may impose financial burden on the infertile couples and be time consuming to IVF team. The aim of this study was to determine the relationship between chromosomal abnormalities of unfertilized eggs and female hormones, FSH and LH.Materials & Methods: Unfertilized oocytes from IVF treatment cycles were prepared by Tar-kowski method for chromosomal studies and after staining by Giemsa method, they were chro-mosomally analyzed. For hormonal evaluations of FSH and LH, ELISA was used. The results were analyzed by ANOVA and correlation coefficient calculations through using SPSS software and p-values <0.05 were considered significant. Results: From 52 treatment cycles, 362 oocytes were left for IVF from which 285 oocytes were fertilized and 77 oocytes were unfertilized. Considering the elimination of 26 fertilized oocytes chromosomal analyses were done on 51 ova and sixteen oocytes from nine treatment cycles had chromosomal abnormalities. Most abnormalities were in the C group and the minimum were observed in the G group of the standard classification of chromosomes. There were no significant relationships between chromosomal abnormalities and FSH, LH levels (with mean values of 16.22±8.1IU/L and 7.74±3.8 IU/L respectively), age and duration of infertility.Conclusion: The results of this study showed that there were no relationships between chromo-somal abnormalities of oocytes and female factors. Therefore, it seems that for the identification of the main causes of TFF in infertility cycles, it is necessary to consider other related factors such as sperm function, when there is no oocyte or female hormone abnormalities.

Introduction: Stem cell therapy seems promising for the treatment of incurable diseases such as injured myocardium. Directed in vitro differentiation of embryonic stem cells (ES) to cardiomyo-cytes is invaluable. TGFβ-2 and BMP-2, two different growth factors from the transforming growth factor-β superfamily (TGF-β), exhibit their effects on growth, immigration and different-tiation and other cellular functions during embryonic development. In this study, the effects of these factors on differentiation of ES cells into cardiomyocytes were evaluated.Materials & Methods: Murine embryonic stem cells (CCB derived from mouse strain 129) were used for cardiomyocyte differentiation. To increase the effects of the factors, ES cells were grown on inactivated fibroblasts for 24 hours in differentiating culture medium containing 8 ng/ml of TGF-β2 and 5 ng/ml of BMP-2 while decreasing serum concentration from 20% to 7.5%. Later, embryoid bodies (EB) were formed in 800-cells hanging drops and suspension from ES cells and eventually, EBs were cultured on gelatin-coated plates. After continuation of these steps, ES cells differentiated into beating cells. To determine the effects of the growth factors on ES cell differen-tiation into cardiomyocytes, expression of primary transcribing factors, MEF2, Nkx2.5, and specific cardiac genes including MHC, ANF, MLC-2v were assessed by semiquantitative RT-PCR and expression of desmin and α-actinin in differentiated cardiomyocytes was studied by western blotting and immunostaining.Results: After completion of the differentiation process, the EBs in the experimental groups exhibited beating compared with the control group. Considering the quantitative comparison of the dectrophoretic bands of RT-PCR and PCR products of the plated EBs in different media, it was seen that TGF-b2 from BMP-2 and both factors together, compared to each one alone, is more effective on differentiation of embryonic stem cells into cardiomyocytes. In addition, in plated EBs, in a medium containing both factors, the expression of desmin and a-actinin were determined by western blotting and the presence of cardiomyocytes, by marking desmin, by immunohistochemical staining.Conclusion: The results showed that the synchronized effects of both factors, while decreasing serum concentration during ES and EB cell cultures, increases differentiation of ES cells into cardiomyocytes.

Introduction: Embryonic stem (ES) cells are pluripotent cells that can differentiate into all types of specific adult cells and are used for cell therapy in certain diseases. It seems that the inductive factors may be used to produce neurons and gliocytes out of ES cells for the treatment of some nervous system diseases in future. In this research, pluripotent and undifferentiated murine embryonic cells were cultured and effects of the growth factors NGF, RA and Shh were studied for the induction of neural and glial differentiations.Materials & Methods: CCB ES cell line derived from  mouse strain 129 were cultured on inactivated embryonic fibroblasts from C57/BL6 mice and embryonic bodies (EBs) were pre-pared and transplanted onto culture plates covered by fibronectin and growth factors NGF, Shh and RA with respective concentrations of  50 and 100 ng/ml, 300 and 500 ng/ml and 1 μM and  in some groups bFGF with a concentration of 20 ng/ml were added to the specific culture medium for neural cell precursors to induce cell differentiation into neural cells. For studying differentiation into many types of neurons and oligodendrocyte, gene expression of specific neural genes such as nestin, Nkx2-2, Nurr1, S100 and Olig-2 were assessed by RT-PCR and immunocytochemistry assays were used to confirm the presence of MAP-2 protein.Results: In this study, genetic evaluations showed that each of the previously mentioned growth factors activate some molecular mechanisms that have essential effects on differentiation into different kinds of nervous system cells by affecting the expression of marker genes. Cytochemistry of these cells by monoclonal MAP-2 antibody showed the resultant cells have dendrites that are specific for neural cells or neurons.Conclusion: The findings indicated that pluripotent and undifferentiated CCB ES cells underwent differentiation into neural cells and oligodendrocytes under the effects of NGF, Shh and RA growth factors and this neural induction was confirmed by the presence of molecular and antigenic markers in these cells. Each of these factors was effective in the induction of differentiation of specialized neural cells and even different concentrations of the factors induced particular cell differentiations.

Introduction: Endometriosis, as one of the main causes of infertility in women, has devoted a lot of research to itself. Three-dimensional culture of human endometrial tissue has been known as a functional model for endometriosis. Statins have a variety of different effects on angiogenesis and they have been suggested as a candidate for the inhibition of this disease. Therefore, the aim of the present study is to determine inhibitory or promotory effects of different concentrations of Atorvastatin on the growth of cultured human endometrium.Materials & Methods: 8 endometrial samples were cut into 1x1 mm pieces and were cultured in 24-well culture plates. To establish the desired three-dimensional model, 0.5 ml of fibrinogen solution in M199 medium, with a concentration of 3mg/ml and 10µL of thrombin were added to each well. After formation of the jelly, the endometrial fragments were placed on it and a second jelly layer was added. The endometrial tissue of each specimen was cultured in four 5-piece layers. The first row was regarded as control and to the second, third and forth rows, 0.1, 1.0 and 10µM concentrations of the medication were added respectively. Every 3 days, endometrial pieces and their probable changes were observed by an invert microscope (Motic-AE31) and the culture media were changed. The tissue fragments were also photographed on days 1, 7, 14 and 21. At the end of the study, the cultures were graded according to their cellular outgrowths, invasion of deposited cells into the jelly and growth of endometrial glands and images related to the 3-D cultures were analyzed. To confirm the growth of cellular structures into the jelly, paraffin embedded cuts of the cultured specimen were immunohistochemically stained by anti-vimentin, anticytokeratin and anti-CD31. The data were analyzed by SPSS statistical software (Version 11.5). The statistical analysis related to the grading of wells was done by Freedman test and the statistical analysis of the images was performed by one-way ANOVA.Results: Growth changes were 82.5% in the control group and 87.5% in the 0.1µM concentration group, depicting significantly higher values compared to the control group (p<0.05). The changes in 1µM and in 10µM  groups were 50% and  5%  respectively which had  significant differences with that of the control group (p<0.001).Conclusion: Different concentrations of Atorvastatin have different effects on the growth of cultured endometrial tissue, 0.1µM concentration increases growth of cultured human endomet-rium, while 1µM and 10µM can decrease growth of the tissue. Further animal studies on this subject are suggested.

Introduction: Based on supportive aids of WHO in the field of public health, family planning and reproductive health, consumption of herbal drugs has been considered as an alternative to synthetic contraceptive drugs. Centella asiatica has been traditionally used for the treatment of different types of diseases for thousands of years in various parts of the world including Eastern Asia, China and India. Considering the dispersal of this plant in Bandar Anzali’s wetlands and lack of research on the effects of this plant on reproductive system in laboratory models, this study was carried out to evaluate the effects of Centella asiatica extract on spermatogenesis and testical tissue of rats.Materials & Methods: At first, the plant was collected and its extract was prepared through percolation process. Some 8 to 10-week old male Wistar rats, weighing about 200-250 grams, were selected randomly and were divided into 6-member subgroups. The rats in the control group received water, the sham group received Tween solvent and the third to the sixth groups received non-fatal doses of 10, 50, 80 and 100mg/kg of the crude extract for 60 days respectively. On the 61st day, the rats were dissected and their testes were taken out and weighed for probable effects of the extract on the organ¢s weight. Then sperm parameters and histological evaluations of the testes were done. The results were analyzed by the use of SPSS and Pharm softwares by calculation of one-way ANOVA while considering p<0.05 as the significance level. Results: The LD50 of the plant was calculated as 500mg/kg, with a confidence interval of 1.9- 2.2 in male rats. The non-lethal doses of 10, 50, 80 and 100 mg/kg of Centella extract were considered for histological and spermatogonial evaluations. There were significant increases in body and testis weight in rats receiving the extract compared to the control and sham groups. There were histological changes during spermatogonial evolution such as degeneration of spermatozoa and interstitial congestion in some tubules and sperm analysis showed a meaningful decrease in the number of spermatozoa (p<0.01), motile sperms (p<0.001) and epididymal sperm storage (p<0.001) compared to the sham and control groups but there were no changes in sperm morphology.Conclusion: Based on the above results, it seems that Centella asiatica can be used as a temporary contraceptive agent in animals. Nevertheless, more biochemical and molecular research on the contraceptive effects of this extract is needed to determine its economical benefits and prepare an appropriate formulation.

Introduction: Pre-eclampsia is one of the complexities of maternal and neonatal health. There are several similarities between pre-eclampsia and atherosclerosis. The relation between chronic Chlamydia pneumoniae infections with atherosclerosis has been shown previously. This study has been done to determine the connection between pathogenesis and onset time of pre-eclampsia and chronic Chlamydia pneumonia infection.Materials & Methods: This cross-sectional study was done on 88 women with normal pregnancy outcomes, 72 patients with late-onset pre-eclampsia and 16 patients who developed early-onset pre-eclampsia in Zahedan in the years 2004-2005. Anti-Chlamydia IgG and its titer were checked for in the cases and controls by ELISA method. After entering the demographic data and IgG status of  the study population, statistical analysis was performed using SPSS software by one-way ANOVA and Chi-Square test, at a significance level of p<0.05.Results: The results showed that there were no difference in seropositivity among the three groups (p=0.4). There were also no differences between early-onset and late-onset pre-eclampsia (p=0.5), early-onset pre-eclampsia and control group (p=0.32) and late-onset pre-eclampsia and control group (p=0.25) in that regard. There was a significant difference in IgG titers among the three groups (p<0.0001), between early-onset and late-onset pre-eclampsia (p<0.0001) and between early-onset pre-eclampsia and control group (p<0.0001), but there were no differences between those with late-onset pre-eclampsia and the control group (p=0.98).Conclusion: The results of this study showed that chronic infection with Chlamydia pneumoniae could be related to early-onset pre-eclampsia. This study can be a support for evaluating the relationship between pre-eclampsia and subsequent atherosclerosis. Therefore, it is suggested that use of macrolides be part of the preventive programs in high-risk individuals for pre-eclampsia.

Introduction: Infertility and azoospermia are among the clinical presentations of males with impalpable testis. The management of non-palpable testis especially in adult patients remains controversial. Routine imaging methods like sonography, CTS and MRI are not sensitive enough to detect impalpable testes and have high false negative results. Therefore, it is necessary to perform laparoscopic examination in such patients. Although a few studies have reported benefits of laparoscopic examinations in the management of such cases, but it is still subject to many controversies showing the need for more investigations in this area. The aim of this study is to evaluate the role of laparoscopy in the diagnosis and treatment (mostly orchiectomy) of impalpable testis in azoospermic men.Materials & Methods: This is a case series study in which the files and laparoscopy movies of patients who had attended Avesina Infertility Clinic form February 2006 to October 2005 and had indication for laparoscopy were retrospectively analyzed. During the study twenty testicular units from 12 married patients with primary infertility and azoospermia were studied. After preliminary evaluations, informed consent for probable orchiectomy was taken from the patients. After preoperative preparations, diagnostic/surgical laparoscopies were performed. In cases of blind ending vessels or vanishing testes, the procedure was terminated. Orchiopexy or orchiectomy was performed based on testis size, distance from internal ring and patient’s age.Results: Twenty testicular units from 12 patients with unilateral or bilateral cryptorchidism were studied. The mean age of patients at the time of attendance and laparoscopy was 37 years (33-43). Eight patients presented bilateral non-palpable testes and four patients with palpable one-sided atrophic testis, two patients had natural descents and two had undergone postpubertal orchiopexy. From the 20 impalpable testes undergoing the procedure, 5 (25%) were absent, 3 vanishing testes and 2 previously done orchiectomies. 15 out of the 20 testes (60%) were located in intraabdominal cavity and were managed successfully by the means of laparoscopic orchiopexy or orchiectomy. No laparoscopy related complications occurred. All testicular biopsies from orchiopexy or orchiectomy were sent to a pathology laboratory. Three of the 15 biopsies were reported as atrophic testis, three as immature testis and 8 of them had germ cell aplasia.Conclusion: The data suggest that laparoscopy can be considered as a safe and effective method for the management of non-palpable testis in adults. The patients may experience less pain and benefit from better cosmetics, early discharge and rapid recovery.

Introduction: Reproductive health and paying attention to its different dimensions in national and international levels are the essential steps in providing social and family health, with an emphasis on women's health. Regarding the new applications of the term, "Reproductive Health" is dependent upon all aspects of girls' and women's lives and health. The point that women should enjoy their reproductive health, reproductive rights are considered as parts of human rights and also considering Islam’s support for this issue, the present research was carried out to determine women’s knowledge, attitude and behavior on reproductive health subjects.Materials & Methods: This is a cross-sectional and analytical-descriptive study carried out in seven central cities of Iran in 2005. The subjects consisted of 840 married women, aged 15-49 years, who attended urban and rural health centers and were selected in a 3-stage sampling method. The instrument for data collection was a questionnaire completed through interviews. The data were entered into and analyzed by SPSS, version 11.5. Applied statistical tests in this research were Kruskal- Wallis non parametric test, c2 and Pearson correlation test.Results: According to the results, 62% of the women were familiar with the term reproductive health and 66.4% of them used effective contraceptive methods, although they obtained an average of 9.21 (from a maximum mark of 14 for knowledge), just 18% of them knew the appropriate time for Pap smear and breast self-exam and also there were many misconceptions on HIV/AIDS. Kruskal-Wallis test showed a significant difference between awareness of women in various cities with their educational achievements (p<0.05). Women's attitude about reproductive health was good but there were significant differences among women from different cities (p<0.05). The women’s behavior in many categories, such as Pap smear, breast self-exam and use of safe contraception methods need to be enhanced.Conclusion: Regarding the results and importance of women's reproductive health and rights, as well as instructive advocacy of Islam on the matter, promoting women's knowledge, attitude and behavior on reproductive health must be included on the agenda of health policy makers and non-governmental organizations in studied cities.

Leukocytes in the male genital tract and seminal fluid play a complex and dynamic role. Leuko-cytes are found in virtually every ejaculate and function at multiple levels. Presence of high leuko-cyte counts in seminal fluid is an important indicator of male genital tract infection or inflamma-tion. In many studies, leukocytospermia has been associated with male infertility. Decreased sperm count and motility, as well as increased abnormal sperm morphology and a high frequency of immature germ cells, have been reported in men with leukocytospermia. According to the World Health Organization, leukocytospermia is defined as the presence of 1 x 106 WBCs per milliliter of semen. Methodological problems have been found to interfere in differentiating WBCs from immature germ cells that are present in the seminal fluid. For clinical purposes, peroxidase staining is the most practical method of leukocyte detection in semen. A large number of studies have shown a wide range of leukocytospermia incidence in infertile men (from 2% to 35%). The majority of studies with the largest number of cases have estimated the prevalence of leukocytospermia to be between 12 and 20% among all infertile men. The assumption that leukocytospermia is merely the result of a subclinical male genital tract infection is not correct. Environmental factors such as smoking, alcohol consumption, and marijuana use, increase the number of WBCs in semen. Prolonged abstinence and certain sexual practices, such as anal sex, may also cause leukocytospermia. Nowadays, numerous scientific evidences refer to leukocytes and their products as effective factors on sperm and their functions. It is thus strongly believed that diagnosis of leukocytospermia and its causative agents as well as its treatment play a crucial role in evaluation of infertile couples.