Swim up is the most popular technique of sperm preparation, in which sperm is selected on motility and ability to swim out the plasma of the semen. The goal of this research is to compare the influence of adding platelet rich plasma (PRP) and phosphate buffer saline in semen prepared by swim – up technique. This research included 119 semen sample evaluated according to sperms parameters stated in WHO and divided into 2 groups. The first group treated with equal quantity of phosphate buffer saline (250) μm contain 10% human serum albumin (HSA) while second group treated with equal quantity of phosphate buffer saline (250) μm contain 10% human serum albumin (HSA) and 2% platelet rich plasma. The samples are prepared by swim – up method. Applying swim up technique using-PBS+PRP (HAS) medium resulted in highest sperm parameters in a highly significant manner (p < 0.001). The values were far better in Swim -up PBS (HAS) with added 2% PRP than those produced by swim up –PBS (HAS) alone. The statistical analysis shows that the variations between Swim up-PBS (HAS) with addition 2% PRP media and swim -up PBS (HAS) were highly significant.
In about 40% of infertile couples, the husband was found to be either the single or a contributing cause of infertility and the systemic management ready for those couples with male infertility are restricted. 135 couples with infertility shared herein; the husbands required in vitro activation for their sperm samples, their wives however; had no fertility problem and were submitted ovarian induction protocol. However; Sperm preparation was done using both, the centrifugation indirect swim-up technique with Global medium and density gradient with ALLGrad solution. While ALLGrad 45% is a ready-to-be-used upper layer. There was an acceleration in seminal parameters after preparation using both various techniques. So, a highly significant increment (P≤0.001) in the percentages of progressive forward motility was in grades (A and B) and grade (A) progressive sperm motility, also a significant enhancement (P≤0.05) in percentage of morphologically normal sperms when applying density gradient mechanism in relation to centrifugation swim-up technique. This study concludes that the use of density gradient mechanism with ALLGrad solution and Global culture medium are more efficient in achieving better percentages of progressive forward motility grades (A and B), grade (A) progressive motility and morphologically normal values of spermatozoa than centrifugation indirect swim-up technique by using Global culture medium. However, it was found there was a non-significant difference when we use both techniques concerning other sperm parameters.
Background: The intra uterine insemination (IUI) still an easy, simple and effective method to overcome the infertility that resulted from different causes. Although the equipments and methods of assisted reproductive technologies (ART) were highly developed, the percentage of successful IUI live-birth not exceed 20-30%. Objective: The present study was aimed to elucidated the cut off values of in vitro sperm preparation technique, follicular number and size, endometrial thickness and hormonal profile that lead to successful pregnancy rate (PR)following IUI. Materials and Methods: In retrospective study, one hundred cases of infertile couples were in who were became pregnant following IUI at the Institute of Infertility Diagnosis and Assisted Reproductive Technologies, through the period from January 2007 to January 2010. Depending on the results of retrospective study, IUI was achieved for 100 infertile patients between August 2010and June 2011. The mean of significant prognostic variables were measured in both studies namely: age, type of infertility, type of sperm activation techniques, the protocol of ovulation induction medicine, the mean of hormonal status of Luteinizing Hormone (LH), Estradiol(E2), and number with diameter of follicles, endometrial thickness a cycle day before hCG injection. Results: The protocol of ovulation induction in IUI cycles by using clomiphene citrate(CC) and recombinant gonadotropin (rFSH) was significantly(p<0.05) improve the IUI results compared to CC alone or CC with PergonalS. The mean values obtained by ultrasonography namely; two follicles, and dominant follicle of 19.5mm in size with endometrium thickness of >9.5mm were gave significantly successful IUI outcome compared to other values. In prospective study similar results were obtained, therefore the overall pregnancy rate was 37% per cycle. Conclusion: The calculated data of successful IUI outcome from retrospective study gave a best pregnancy rate in prospective study which was 37%. Consequently, these results can be considered as guideline for physician who interest in assisted reproduction to have a success in IUI.
Background: The procedures of assisted reproductive technologies have been developed during recent years and have been revealed the need for new suitable and effective techniques of sperm treatment in the laboratory. Objectives: This study is designed to normozoospermic and asthenozoospermic compare the semen outcomes of two in vitro sperm activation techniques and evaluate their efficiency, namely; Glass wool filtration technique and a new technique of Glass wool filtration combined with Pentoxifylline. Subjects, Materials and Methods: Fifty five semen samples were collected from the patients, fifteen were obtained from normozoospermic men, forty were obtained from asthenozoospermic men who at tend the infertility clinic were involved in this study. Sperm parameters were assessed according to WHO (2010 and 1999). Post-activation of each sample were divided into two aliquots, the first one : using Glass wool filtration technique , the second : using Glass wool filtration technique with Pentoxifylline. Results: A significant increase of sperm motility, progressive sperm motility and morphologically normal sperm when using Glass wool filtration technique combined with Pentoxifylline as compared to Glass wool filtration technique. Regarding sperm agglutination and Round cells count the study showed the best results by decreasing their levels when using Glass wool filtration technique combined with Pentoxifylline as compared to Glass wool filtration technique alone. Conclusions: From the results of the present study, the sperm function parameters outcome when using glass wool filtration technique combined with Pentoxifylline were superior to other technique since it yields better sperm function parameters rather than the outcome of glass wool filtration ; respectively especially when using a low quality of semen samples such as decreased sperm function parameters (sperm motility) so it was fit in cases of asthenozoospermia which taken as an example of infertile group within this study.
Background: Semen preparation techniques were developed to separate motile sperm that are morphologically normal from seminal plasma to optimize successful assisted reproductive technology cycles which seem to be the most effective options in cases of a male factor infertility. Objective: The objective of the present study was to compare between outcomes of two sperm preparation techniques for asthenozoospermic patients includes:1-Centrifugation swim up technique.2-Glass wool filtration technique. Subjects, Materials and Methods: Fifty three infertile males were participated in this study during their attendance to the Infertility Clinic at High Institute for Infertility Diagnosis and Assisted Reproductive Technologies; Al-Nahrain University. Semen samples were collected and SFA was done according to WHO (2010 and 1999). Each semen sample was divided into two aliquots. The first one prepared using centrifugation swim up technique, while the other one prepared using glass wool filtration technique then sperm parameters were assessed for both techniques and the results were statistically analyzed. Results: After in vitro sperm activation using both techniques, there was significant reduction (P<0.05) in the sperm concentration, significant improvement (P<0.05) in the percentages of sperm motility and morphologically normal sperm when compared to pre-activation. Present study appeared that the glass wool filtration technique resulted in significantly (P<0.05) better results for sperm concentration and total number of progressive motile sperm than the swim-up technique. The present study proved that there was significant (P<0.05) improvement in sperm parameters (increment for sperm motility (%), progressive sperm motility (%), total number of progressive motile sperm and normal sperm morphology (%), while reduction for round cell count and sperm agglutination percent) for all cases using glass wool filtration techniques. In contrast, there was failure of sperm activation for 10 cases using centrifugation swim up technique. Conclusions: From results of the present study, the sperm parameters outcomes using glass wool filtration technique was superior to the outcomes of centrifugation swim up technique when prepare semen of asthenozoospermic patients.
ackground: Pretreatment of semen samples of the infertile patients and in vitro preparation techniques with the certain sperm stimulant agents prior to use for assisted reproductive techniques (ART) has been reported to have beneficial effects on pregnancy rates. Objectives: The objective of this study is to improve certain sperm function parameters in vitro of infertile patients semen by using novel combination motility stimulants substances that improved medium to be used for ART in future. Patients and Methods: One hundred infertile men involved in the current study. They were divided into four groups. Each semen sample was divided into four portions. One part was considered as a control and in vitro activated by using culture medium only. The other three portions were considered as treated groups and activated in vitro by adding pentoxifylline(PX,1mg/ml) and/or L-carnitine (LC,0.5mg/ml) to the culture media. Certain sperm function parameters were examined before and following in vitro activation using layering and sedimentation techniques. Results: The results showed a highly significant (P<0.001) improvement in the sperm concentration and percentage of progressive sperm motility grade(A), grade (B) and grade (A+B) with a highly significant (P<0.001) increase in the percentage of morphologically normal sperm (MNS) after addition of PX and/or LC to the culture medium in comparison with control medium using layering and sedimentation(layering under paraffin) techniques in the infertile patients. Conclusion: The results of this study indicate that adding 20%PXand/ or LC to the culture medium for in vitro sperm activation using layering and sedimentation techniques leads to an improvement in certain sperm function parameters.
Intrauterine insemination (IUI), it is a type of artificial insemination, used to treat infertility by which the Sperm that have been prepared and placed directly into the woman’s uterus around the time of ovulation. The main purpose of this procedure is to enable the sperm to swim into the fallopian tubes and fertilize a ready egg in order to lead to a natural pregnancy. IUI can be coordinated with the normal cycle with fertility or medications depending on the reasons for infertility The IUI outcome still low about 20-25% so adding number of biochemical factors to the culture medium which are essential to regulate the fertility potential of spermatozoa, for example, platelet-activating factor (PAF) which is a unique and novel signaling phospholipid that has multiple biologic properties have been tried. To determine the effect of adding the PAF into the culture medium to prepare and activate the semen samples of non-male factor infertility (normal semen analyses). This study was carried out in the Infertility Unit, AL-Hussein Teaching Hospital, Al-Nassirya City, through the period from 30th of October 2020 till 17th of April 2021. The study included thirty couples who are complaining from infertility (they failed to conceive after one year at least of regular unprotected intercourse. The frequency distribution of enrolled infertile men with normozoospermia Baseline characteristics of seminal fluid analysis of males with normozoospermia those with PAF are Semen volume ml, pH, Sperm concentration before million/ml, Sperm motility %, morphologically normal sperm %, were Mean ±SD (3.10 ±1.23, 7.39 ±0.03, 46.67 ±15.31, 1.33 ±3.52, 36.00 ±5.07) respectively. Certain sperm parameters were highly improved after in vitro activation by PAF medium of Normozoospermic samples.
Background: The sperm plasma membrane activity and absence of antisperm antibodies in infertile patients ejaculate significantly associated with fertilization potential of human spermatozoa. The sperm viability detected by hypoosmotic swelling test (HOST) and autoimmune infertility detected by antisperm antibody assay presented diagnostic and prognostic tools for male infertility factors. Objective: This study was devised to compare the results of hypoosmotic swelling test and antisperm antibody assay for normozoospermic men and infertile patients affected by autoimmune infertility. Methods: Thirty semen samples were collected by masturbation and prepared by direct layering technique. Hypo-osmotic swelling test was performed by mixing 0.1 ml of semen with 0.9 ml of a 150 mOsm/L NaCl as a hypo-osmotic solution and direct immunobead assay were used to determine the presence of Antisperm Antibodies (ASA) bound on sperm surface. However, sperm concentration, sperm motility, and normal sperm morphology were evaluated according to World Health Organization (WHO) criteria and subjected to HOS test and ASA assay. The sperm prepared and incubated for 30 minute in 5% CO2 at 37°C after in vitro sperm preparation. Results: The results of the present study reported that the percentage of HOS test score in antisperm antibodies positive sperm samples (52.30 ±5.2) was significantly lower than that noticed in antisperm antibodies negative (70.30 ± 3.4) sperm samples. Conclusion: It was concluded that the use of HOS test as a simple and diagnostic and prognostic tools for identify male infertility. Further studies are suggested to assess the effect of ASA on sperm plasma functional integrity after ICSI and in vitro fertilization and embryo transfers (IVF-ET).
Background: Many studies conducted on the pentoxifylline (PX) and Glycyrrhiza glabra (G.glabra) as motility stimulants showed positive effect on sperm preparation in vitro improving the progressive forward movement. However, it was not known the impact of these stimulants on the nature of genetic material, especially after cryopreservation. Objective: The experiment was designed to find out any harmful effect of a medium containing a mixture of PX and aqueous extract of G.glabra on the sperm DNA before and after cryopreservation. Materials and Methods: Fifty mature fertile male mice 8-12 weeks old were used. Epididymal sperms were obtained from caudal region and prepared in vitro by direct activation technique using four media namely; PX alone, G.glabra alone, a mixture of PX with G.glabra and Ham's F-12 (as control medium). Certain sperm function characteristics were examined as well as evaluation of DNA integrity by acridine orange test and comet assay before and after cryopreservation. Results: The results showed a highly significant (P<0.001) improvement in the sperm forward movement, with a highly significant (P<0.001) decrease in DNA abnormality using acridine orange test and comet assay, following in vitro activation by the four media compared to before activation. Following in vitro activation and cryopreservation, the results of epididymal sperm showed a highly significant (P<0.001) decrease in DNA abnormality by using PX medium alone, G.glabra medium alone and the mixture of PX and G.glabra compared to control medium and the results before cryopreservation. Conclusion: According to the results of present study, it has been found no toxic effect of medium containing PX and/or G.glabra on the epididymal sperms DNA after activation in vitro and cryopreservation using acridine orange test and comet assay. The best result obtained when using a medium containing a mixture of PX+ G.glabra together. These data could be applied for preparation and activation of epididymal and testicular sperms obtained from obstructive azoospermic men.
Infertility has been defined as the disability to fulfill pregnancy after twelve months of regular sex coition, with no contraceptive taken. Male factor infertility was explained in terms of low spermatozoa concentration (oligozoospermia), or morphology (teratozoospermia), impaired motility (asthenozoospermia), or in some cases, total lack of spermatozoa in the ejaculate (azoospermia). In several cases, a collection of one, or more of these sperm variables defects may be noticed, evaluation these factors by the initial and most important step is seminal analysis. Our study aims to study some sperm characteristics in asthenozoospermia men in comparison with normozoospermia men before and after glass wool and Sephadex activation. This study involved 60 semen samples collected from male patients that came to male infertility clinic at Al-Nahrain University; the recruited semen samples were divided into 2 groups, (40 asthenozoospermic and 20 normozoospermic subjects). After collected semen samples, and assessed analysis of seminal fluid. Each semen samples were divided into 3 aliquots. The first aliquot prepared was In-Vitro to assessed sperm characterization before activation, the second part using glass wool filtration (GWF) technique, while the last part was prepared using Sephadex. Both techniques resulted equally in reducing sperm concentration, were equally effective in upgrading sperm motility and in minimizing round cell count; however, Sephadex was superior to glass wool method in upgrading the percentage of morphologically normal sperm. Both Sephadex and glass wool techniques has been proved effective to improve semen quality.
Phosphatidylserine (PS), exposed on viable sperm, is required for fertilization as it is recognized by specific receptors located on the microvilli of the oocyte to promote sperm: egg fusion. The heads of viable and motile sperm exhibit PS. With (PS) exposure progressively increasing during sperm transit through the epididymis. PS asymmetry on the leaflets of plasma membrane is arranged in a complex way, via multiple proteins functioning in opposite directions; further, PS is also essential for many intracellular trafficking events. The aim of this study was to evaluate the phosphatidylserine PS level in seminal plasma of infertile men before and after activation using Direct Swim Up (DSU) and Indirect Swim Up (IDSU) activation method and make a comparison between them. Forty infertile men were included in this study during their attendance to the infertility clinic at High Institute for Infertility Diagnosis and Assisted Reproductive Technologies, Al-Nahrain University. Normozoospermic or mild asthenozoospermic semen samples were used to choose the cases, according to WHO manual standards (1999 -2010). The collected semen samples obtained were divided into three parts: The first part prepared was as an in vitro sperm characterization before activation while the second and third parts were after activation using direct and indirect swim up techniques respectively. The level of seminal plasma phosphatidylserine was increased after the activation techniques from the baseline level before activation. Comparison of Seminal plasma phosphatidylserine levels between direct and indirect sperm activation technique were significantly higher in direct swim up activation technique than indirect one. Phosphatidylserine (PS) in semen plasma of infertile men was significantly increased after activation using Direct Swim Up DSU preparation method.
Background: Pretreatment of the semen samples of the asthenozoospermia patients with certain sperm stimulant agents prior to use for assisted reproductive techniques (ART) has been reported to have beneficial effects on fertilization rates. Objectives: Investigating the influence of addition of different concentrations of oxytocin (OT) to semen samples of asthenozoospermic patients for in vitro sperm activation as a new method for stimulation of sperm motility in these patients and exploring the concentration of OT that harvests amaximal numberofmotile sperms. Methods: Simple layer technique for in vitro sperm activation using culture medium enriched with OT was studied in asthenozoospermic patients (No. 90). Semen samples were examined before sperm activation, and then they were divided into three equal groups. In the first group, three concentrations of OT (2, 4, and 6 IU/ml) were applied. Based on the results of the mentioned work, OTat the concentration of 2 IU/mlwas applied with sperm preparation medium (SPM) and without in the second and third groups, respectively. Results: Results showed that there was a significantimprovementin the percentageof sperm motilityand spermgrade activity of the OT-treated samples atthe concentration of 2IU/mlas comparedto the control samples. Moreover, there was a tendency for a decrease in sperm grade activity with increase OT concentration. The outcome of sperm activation in presence of SPM revealed a significant increase in the sperm motility characteristics (the percentage of sperm motility and sperm grade activity) in the control and OT-treated samples and the best improvement in these sperm motility characteristics was observed in the OT-treated samples. When SPM was absent, a significant improvement in the sperm motility characteristics was observed only in the OTtreated samples. Conclusion: The results of this study indicate that OT at the concentration of 2 IU/ml has got a stimulatory effecton sperm motility characteristics; also, the use of SPM succeeds to supportthis effect.
Background: Semen cryopreservation is a useful tool for preserving fertility for men involved in ART program who have been diagnosed with cancer and will undergo chemotherapy, radiotherapy or testicular surgery. Objective: The objective of this study to evaluate the effect of honey bee supplemented to cryoprotectant medium on post-thaw sperm DNA integrity ,morphology and round cells. Subjects ,Materials and Methods: Thirty semen samples were collected from 30 infertile patients. After assessment of semen analysis, semen samples were divided into 3 aliquots (0.7ml for each sample) and mixed with 1 ml of cryopreservation solution (G1,control) alone, enriched with 5% honey bee(G2) with 10% honey bee (G3) for cryopreservation. Current study used the acridine orange test to investigate the effect of honey bee in two concentration (5%, 10%) on the integrity of sperm chromatin structure pre-and post cryopreservation. Cryopreservation was done at -196 Co in liquid nitrogen and thawing was performed after six months. Direct swim up technique was used for in vitro sperm preparation post-thawing, sperm parameters were assessed and data were statistically analyzed pre-and post-thawing. Results: The results show there was a significant(P<0.05) increase in the percentage of morphologically normal sperm for all groups post-thawing, particularly for G3 group that was significantly (P<0.05) increased as compared to G1 and G2 groups, In contrast, non-significant differences (P>0.05) were observed between G1 and G2 groups. Sperm DNA fragmentation percentage (%) was significantly(P<0.05) increased in G1 post thawing without any additives as compared to pre-cryopreservation , where as in G2 and G3 sperm DNA fragmentation were increased but not significantly as post-cryopreservation groups. Also the results show that there were no significant differences between G2 and G3 whereas G3 gives better result as compared to G1 with significant differences. Round cells counts for all groups of post cryopreservation were significantly(P>0.05) decreased as compared to pre-cryopreservation. Non significant (P>0.05) differences in the round cells counts among all the groups of post -thawing in spite of G3 group has the lowest mean. Conclusions: From these results we concluded that supplementation of 10% of honey bee to the freezing cryoprotectant medium gives best protection to the morphology and DNA integrity of human sperm.
For conception and the development of healthy embryos, sperm DNA integrity is crucial. According to a growing body of studies, there is a strong correlation between sperm DNA damage and male infertility. Among the new medicines being developed in the medical field, the application of Platelet Rich Plasma (PRP) in human reproduction has yet to be examined. A total of 100 semen samples were used in the current experimental investigation. From November 2020 to June 2021, the research was conducted at the High Institute for Infertility Diagnosis and Assisted Reproductive Technologies. Masturbation was used to get an ejaculated semen sample. After semen analysis, the samples were separated into two equal parts, one without autologous PRP and the other with 2% autologous PRP, with the DNA fragmentation assessed using the Sperm Chromatin Dispersion Test. There was highly significant reduction in DNA fragmentation index (p < 0.001). The mean sperm DNA integrity was reduced after adding PRP (33.85±16.73 vs 38.55±16.64), Mean (± SE). PRP has been shown to improve human sperm DNA integrity.
Infertility is the inability of a sexually active non-contraceptive couple to achieve pregnancy in one year. Sperm motility is one of the most important prerequisites to achieving fertilization and pregnancy . The objective of the current study was to improve certain sperm function parameters following in vitro activation of men suffering from asthenozoospermia by using a novel combination of motility stimulant substances, namely Maca, L-carnitine, and Pentoxifylline, to be used for assisted reproductive technologies programs in the future.Ninety asthenozoospermic patients were involved in the current study. Each sample was divided into two equal portions. One part was considered as a control by using Ham’s F12 medium, and the other part was considered as a treated group by adding medium content, a mixed combination of Maca powder extracts (Lepidium meyenii) (M)1mg/ml, Pentoxifylline (PTX) 10 mg /ml, and L-Carnitine (LC) 0.5mg /. Certain sperm function parameters were examined before and following in vitro preparation using the swim-up technique according to WHO guidelines.The results showed a significant (p<0.001) elevation in active sperm motility grade A, B, and progressive motility following in vitro preparation by Ham’s F12 media and mixed culture medium compared to before activation. The morphologically normal sperm percentage was significantly higher in mixed culture medium compared to control medium, and before activationWhile the results of DNA normality by using mixed culture and Ham’s F12 media were statistically shown a significant (p<0.001) reduction compared with before activation . It is concluded that the combination medium have a superior effects on certain sperm function parameters and DNA normality enhance its usage in sperm activation by swim up technique for ART programs.