Upstream or swim up processing technique: which one is more effective to select human sperm with high chromatin integrity

Background: sperm processing methods separate motile sperms with good morphology from dead and abnormal forms of sperms, immature germ cells, and non-sperm cells

Objective: the propose of this study was to compare the efficacy of upstream and swim-up processing techniques to separate sperms with the high quality especially in relation to sperm chromatin integrity

Materials and Methods: this experimental study used semen samples from 60 normozoospermic men. Specimens were divided into equal aliquots for processing by swim up [group A], and upstream [group B] methods and compare with control by raw semen [group C]. Sperm concentration, morphology, motility, DNA fragmentation and chromatin maturation were measured in these three groups

Results: the results revealed that sperm concentration in the swim up samples was significantly greater than upstream samples [p/= 0.4]. In addition, sperm DNA fragmentation and chromatin maturation were not significantly different between the three groups [p >/= 0.1]

Conclusion: according to results, apparently the upstream method had no significant efficiency to separate good quality sperms compare to swim up. Therefore, swim up seems to be a simple, inexpensive, reliable and widely available method with an efficient yield to separate motile sperm with good morphology and better chromatin integrity for insemination in the infertility clinics

Isolation and culture of human spermatogonial stem cells derived from testis biopsy

In cancer patients, chemo and radiotherapy can cause infertility by damaging spermatogenesis process. This process is based on self-renewal and differentiation of a rare population of the testicular cells called Spermatogonial Stem Cells [SSCs]. Scientists have tried to isolate, enrich and culture Human spermatogonial stem cells, hoping to resolve infertility problems in cancer recovered patients in the future. Spermatogonial stem cells were isolated and purified from human testicular biopsies sample consisting of at least 500,000 and at most 2,000,000 cells. Two enzymatic digestion steps were performed. Enriching methods, differential plating, and specific culture in serum-free medium with added growth factors: human GDNF, bFGF, EGF and LIF was performed on coated dishes. Human spermatogonial stem cell clusters were observed after 7 to 10 days in specific culture, then after several passages and successful expanding duration of 52 days, the cells were evaluated by three layer immunocytochemistry test [LSAB] to stain GPR125 protein as a surface marker in human spermatogonial stem cells. In current study human spermatogonial stem cell were isolated and expanded with the least manipulations in comparison with the other usual isolation methods like florescent or magnetic activated cell sorting. In contrast to the other SSCs isolation and culture methods, this system is based on the testicular biopsies against large samples, thus suggested method in this study is closer to clinical usage in the future

Evaluation of an aqueous-ethanolic extract from Rosmarinusa officinalis [Rosemary] for its activity on the hormonal and cellular function of testes in adult male rat

Rosmarinus officinalis has been used in traditional medicine extensively. This study evaluated the hormonal and cellular effects of Rosmarinus officinalis extract on testes of adult rats. Thirty male Wistar rats [in three groups] received 50 or 100 mg/Kg b.w of Rosmarinus officinalis extract [made from the plant's leaves, flower and stem] [treatment groups] and 10 mL/Kg b.w normal saline [control group] respectively, on a daily bases by gavage route for 60 days. Then, spermatological properties, histometric parameters and sperm dynamics, testis and body weight, testicular cell population and serum testosterone level were analyzed by an acceptable method. Results showed that the mean serum testosterone level was decreased significantly in both treatment groups [50 and 100 mg/Kg b.w] during the experiment time, compared with control group [p < 0.05]. However, Rosmarinus officinalis did not change the total count, motility and viability of sperm. In addition, Rosmarinus officinalis at both doses did not change body and testes weight and their ratio. Furthermore, Rosmarinus officinalis increased the number of Spermatogonia at both doses, Spermatocyte at doses of 50 mg/Kg b.w, Leydig cell and Spermatid at dose of 100 mg/Kg b.w significantly [p < 0.05]. Rosmarinus officinalis did not significantly affect the number of Spermatozoid and Sertoli cells. In conclusion, it seems that Rosmarinus officinalis may have some hormonal and cellular effects on the testes which can contribute the spermatogenesis process in rat. Rosmarinus officinalis may have antiandrogenic effect potentially indicating the possibility of developing herbal male contraceptive

[Fertility preservation in men after cancer treatment; a review article]

Some cases of male infertility are due to the destructive side-effects of anticancer treatment methods such as chemo and radiotherapies on germ cell lines. The increase in the survival rate of cancer patients who undergo treatment, especially children, has drawn attention to fertility preservation. The most common and effective technique in preserving male fertility is sperm freezing and its subsequent IVF. Children cannot efficiently produce sperm because of their spermatogonial immaturity. One of the strategies to maintain fertility in these patients is to preserve the testes or the germ cells by freezing them for their later maturation and production of fertile sperm, although the state in which the spermatogonia may not undergo maturation is one of the main obstacles faced in this method. Therefore, scientists have attempted to transplant cryopreserved testis tissues or produce in vitro-matured spermatozoa in this group of patients upon anticancer treatment. In this study we reviewed the germ cell biology, the side-effects of chemo and radiotherapies on germ cells and fertility preservation techniques in adults and children undergoing anticancer treatment

[Common techniques for preserving fertility in girls and young women undergoing cancer treatment]

Current protocols for cancer treatment could lead to the failure of ovarian function and subsequent infertility in women. Therefore, utilizing ways to preserve fertility in these individuals seem to be essential. In this review, the full-text of articles which were accessible and had been published during 1976 to 2009 about different methods of female fertility preservation were collected and studied through various online databases such as PubMed, Science Direct, etc. According to the reviewed articles, there are several methods for fertility preservation in women, including ovarian transposition and oocyte, embryo and ovarian cortex cryopreservation. Ovarian transposition is not useful for preserving fertility in women who undergo chemotherapy. Embryo and oocyte cryopreservations require a delay before starting treatment. Metaphase II oocytes are high-volume and fully-differentiated cells which may sustain injury due to the freezing process restricting the number of collected oocytes and reducing the chances of fertility. On the other hand, ovarian stimulation and oocyte collection are not practical in young patients, especially in underage girls. In addition to the restrictions on the number of collected embryos and the raised legal and ethical issues, embryo crypreservation is limited to adults and married women. In comparison to other methods, cryopreservation of the ovarian cortex seems to be more appropriate as ovarian tissue is resistant to cryopreservation and it is easy to be collected by laprascopy, making it practical for use in premature girls. Furthermore, the large number of follicles in the ovarian tissue increases the chances of fertility preservation in women. In general, several parameters including the type, time and duration of treatment, cancer type, age and marital status determine the efficacy of each method

effect of human gonadotropin treatment on recipient mouse germ cell proliferation following spermatogonial stem cell transplantation of neonatal donor mice

Spermatogonia are the male germ line stem cells whose life long expansion is needed for permanent production of spermatozoa. The present study was designed to examine the effect of hCG treatment on germ cell proliferation following stem cell transplantation in mice. Spermatogonial stem cells were isolated from neonatal mice testes and characterized by alkaline phosphatase, immunoreactivity and morphological analysis. hCG was injected into normal and cell transplanted mice. We then evaluated the testosterone levels and cell number in normal mice. After that, cyclin B1 gene expression was investigated in transplanted mice. Different doses of busulfan were injected to investigate the effects of chemotherapy on morphological criteria and preparation of recipient mice for transplantation. In this report we show proliferative potential of spermatogonial stem cells after cytotoxic treatment, transplantation efficiency by semi-quantitative RT-PCR, and hCG effect on stem cell regeneration in normal mice and following cell transplantation. The results indicate the spermatogonial stem cells can proliferate after transplantation, and the efficiency of their transplantation depends on hormonal treatment. Therefore, hormonal treatment after stem cell transplantation will be a powerful avenue for increasing the efficiency of transplantation and fertility restoration

[Effect of Environmental Risk Factors on Human Fertility]

Infertility is believed to be part of the various medical problems that has increased up to 50% since 1955 in the world and 10-15% of couples are already suffering from it. Development of industrialization and urbanization in human societies have dramatically changed the human life style and led gradually to the increase of various environmental risk factors. Humans are exposed to polluted air, containing harmful elements such as lead, drinking water which is frequently contaminated by different noxious materials like arsenic, chromium, benzene, agricultural water and soil containing pesticides and chemical fertilizers - which subsequently will produce contaminated crops - use of hormones and drugs in animal husbandries and presence of their residues such as steroidal hormones in meat and dairy products, the ever-growing use of synthetics and preservatives in food industry. Furthermore, poor dietary habits and malnutrition, consumption of diets deficient in antioxidants, zinc, selenium and copper, adverse effects of some pharmaceutical products and chemical agents such as ketoconazole and dioxin, daily exposure to harmful radiations such cosmic, ultraviolet and X rays, electromagnetic waves emitted from telecommunication transmitters and cell-phones, physical and psychological stresses in living and working environments, smoking, stationary life-style, obesity, and the increasing age of marriage all are the factors which can directly and indirectly affect human fertility. Although measuring environmental hazards and studying their effect on fertility reduction is difficult due to their multifactorial and diverse nature, the problem still remains indeterminate and more studies are required to draw a strong conclusion. The purpose of this review was to study the effects of environmental risk factors, especially emerging risk factors, on decreasing human fertility

Induction of endometriosis by implantation of endometrial fragments in female rats

Endometriosis is defined as the growth of endometrial tissues in ectopic places outside the uterus. This disease has an important effect on the health and fertility of affected women. It's etiology is not clearly known. For better understanding the pathophysiology of this disease, many researchers study on several aspects of the disease on animals. In this experimental study endometriosis was induced in female rats surgically and then its side effects were investigated with special focus on adhesion formation that is a major problem in women with this disease. In Protestrous phase, female rats were randomly divided into two groups. In both groups, under intra peritoneal anesthesia, laparotomy was done and left horn and associated fat were removed. In experimented group [A], the removed endometrium was cut to six square pieces [2mm each] and they were sutured to the peritoneum, near ovaries and subcutaneous. In sham group [B], the same procedure was done for the fat tissues around the removed horn and the pieces were sutured to the same places. After 8 weeks, in Protestrous phase, clinical adhesion and size of implants were evaluated. The total mean size of implants was calculated in each group, and this was significantly larger in experimented group [25.4 mm versus 2 mm p=0.000]. The mean diameter of implants that calculated for each site of implantation in experimented group were significantly larger in left peritoneum [p=0.002], followed by right [p=0.000] and left [p=0.000] ovaries. The endometrial tissues grew in 100% of implants in subcutaneous area. Clinical adhesions [Score. 2] were detected in 7 out of 10 in experimented group and in 2 out 10 in control group. The number of Esterous cycle were similar in both groups. Our study showed that after inducing endometriosis by surgical approach, only endometrial implants grew as a cystic structures and this is a unique aspect of endometrial cells. Our results showed that endometriosis had a direct effect on adhesion formation, not surgery alone and induction of this disease didn't have any adverse effect on ovarian function in female rats

Anti-zona pellucida antibodies in infertile patients in relation to multiple puncture of ovaries and unexplained infertility

Auto antibodies to zona-pellucida [AZA] seem to be important autoantibodies implicated in reproduction, with substantial role in both endocrine and reproductive functions of the human ovary. There are some debates on the relation of AZA with infertility, repeated In Vitro Fertilization [IVF] attempts, and outcome of it. In this study, we assessed the presence of AZA in the follicular fluids [FFs] of women who underwent intra cytoplasmic sperm injection [ICSI], in relation to etiology of infertility and multiple puncture of ovaries. In this prospective study, follicular fluids were evaluated from 96 infertile women, [19-40 years old, 31.5 +/- 5.1], who were c and idates for ICSI based on the etiology of infertility. From these 80 women had explained infertility whereas 16 had unexplained infertility. All FFs were evaluated for presence of AZA by ELISA test. Twenty patients [20.8%] were positive for AZA in follicular fluid. In patients with unexplained infertility, AZA antibody in follicular fluid, was significantly higher than the group with proven etiology of infertility [p=0.001]. In addition, 20.4% of patients who had been punctured previously showed AZA in their FFs which is statistically similar to the patients who were punctured for the first time. The high incidence of AZA in infertile women, especially women with unexplained infertility has to be considered. Relation of the presence AZA and repeated puncture of ovaries is still debatable. Determinations of AZA are highly recommended in evaluation of infertile couples especially in patient with unexplained infertility