Background: Citrullus colocynthis (CC) is an herbal medicine used in the treatment of a wide range of diseases. Recently, It has been studied on in vitro activation of mice sperms with no research concern the effect of this plant on the DNA normality following in vitro preparation of human semen samples. Objectives: The present study was aimed to examine the effect of adding CC in the medium used for in vitro sperm activation of asthenospermic men and related factors and to found out the effect of CC on the DNA status following the activation. Materials and Methods: Citrullus colocynthis aqueous extract (0.01mg/ml culture medium) was used for in vitro sperm activation . One hundred and ten semen samples were divided into two groups, the first group activated in vitro by adding CC extract to the culture medium Ham's F-12 (the treated group) and, the second group sperm activated in vitro with Ham's F-12 alone ( the control group). Results: In vitro activation of sperms with aqueous extract of CC has shown positive effect on sperm concentration, sperm motility, and grade activity of progressive forward movement. There was a significant (P<0.05) increase in certain sperm function parameters of asthenozoospermic in treated group compared with the control group .The study showed that the DNA fragmentation resulted from treated group were lower than that of control group. Conclusions: According to the results of present study, adding aqueous extract of CC to the human sperms activation culture medium leads to an improvement in certain sperm function parameters of asthenozoospermic men with no harmful effect on the human sperms DNA with decrease in DNA denaturation.
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.
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.
Background: Citrullus colocynthis(CC) is an herbal medicine used in the treatment of a wide range of diseases. Recently, the effect of this plant on the reproductive system has been studied.However, its role in vitro still unclear. Objective: The present study was designed to investigate the possibility of using CC extract for in vitro sperm direct activation technique, in vitro fertilization(IVF) and early embryonic development using the mice as a model for mammals. Materials and Methods: Citrullus colocynthis extract (0.05mg/ml culture medium) was used for in vitro direct sperm activation technique. The same medium was used for oocytes insemination and for culturing the embryos after 24-48 hours of insemination. The oocytes were collected from superovulated female mice and divided into two groups: first group, 343 oocytes were inseminated and cultured in CC- free Ham's F-12 medium (the control group). The second group, 345 oocytes were inseminated and cultured in 10% CC - Ham's F-12 medium (treated group). Each 4 oocytes were inseminated with the same sperm concentration (1-2 x 10^5 sperm/IVF well). The fertilization rate was recorded after 24 hours of insemination, while embryonic development rate was recorded after 24 and 48 hours of insemination. Results: In vitro activation of epididymal sperms with 10%CC has shown positive effect on sperm concentration, sperm motility, and grade activity of progressive forward movement. There was a highly significant (P<0.004) increase in FR of the treated group (61.5%) compared to control group (50.4%) after 24 hours of insemination. Embryonic developmental rate was significantly increased after 24 and 48 hours of insemination in treated group compared to control group. Conclusion: It concluded that adding the 10%CC to the culture medium of the epididymal sperm and in vitro inseminated lead to improvements in certain sperm function parameters and sustain the FR and early embryonic development rate.
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: There are many factors and media affecting sperm characters which are defined by classical parameters namely, concentration, motility and morphology, according to standard World Health Organization (WHO). Objectives: To study the effect of incubation time on certain sperm function parameters following in vitro centrifugation technique by sperm wash, FertiCult medium for the purpose of assisted reproductive techniques (ART) in couples presenting sub-fertility. Methods: Two groups of 40 semen samples (group 1: fertile men with normal semen parameters and group 2: abnormal semen parameters) underwent sperm activation test (SAT) in the ART laboratory, High Institute of Infertility Diagnosis & ART/Al-Nahrain University. Semen analysis was assessed in standard WHO (1999). Wash and spin activation technique by FertiCultTM medium was done for all samples. The tubes which was sealed, inclined vertically and kept at 37°C for 30 minute. The supernatant contained actively motile sperms was transferred to another test tube to separate that active sperm from pellet and record sperm parameters after. Results: There is a significant (p<0.05) improvement in certain sperm parameters after 30min. of incubation following SAT in both groups compared to before activation and following 30min. incubation in SAT. Certain sperm characters are significantly decline in group 2 after 1 hr. incubation compared to before activation. Conclusions: Sperm activation test is very important method to perform before undergone artificial insemination (AIH). Incubation of prepared sperm in vitro by sperm wash, FertiCult medium is benefit for the first half hour to all semen samples to have a success in artificial insemination.
Background: The increasing prevalence of infertility cases is becoming a major public health problem in developing countries due to changes in diet and lifestyle. Objectives: Black Carrot Extract BCE (Daucuscarota) plays an important role as fertility agent due to its antioxidant and anti-proliferative properties. Therefore, the aim of the present study was to identify the effects of different concentrations of BCE on the outcome of in vitro human sperm activation of infertile patients. Methods: Thirty two semen samples were prepared which served as a control, using direct swim-up technique. Modified Earl's medium (MEM) was used only in group 2 (G2) and two concentrations of BCE (10µg/1ml and 20µg/1ml) were added to culture media MEM of groups 3 (G3) and 4 (G4) for in vitro sperm activation (ISA). Results: A highly significant (P<0.001) correlation was noticed between G4 with G2 and G3 in progressive motility when activation by adding 20µg/1ml of BCE to culture media as compared to G3 (which adding 10µg/1ml of BCE to culture media) and G2 (activation with culture media only). Moreover, swim-up activation technique with culture media and by both adding (10µg/1ml and 20µg/1ml of BCE respectively) to culture media enhancement in the (sperm motility, progressive sperm activity and sperm morphology). Conclusion: The high concentration of BCE is effective in the enhancement of progressive sperm activity during in vitro human sperm activation.
Background: The effect of L-carnitine (LC) on the male infertility have been previously reviewed. However there are few studies about the effect of LC on sperm function parameters in vitro. Objectives: The objective of this study was to improve certain sperm function parameters in vitro of asthenozoospermic infertile men using LC as a motility stimulant substance which may enhance the medium used for assisted reproductive technologies (such as artificial insemination) in future. Methods: Semen was collected from 50 infertile men involved in the current study. Each semen sample was divided into two portions. One part was considered as a control and in vitro activated by using culture medium only. The other portion was considered as treated group and in vitro activated by adding LC (0.5mg) to the culture media. Certain sperm function parameters were examined before and following in vitro activation using simple layer technique. Results: The results revealed highly significant increment (P<0.01) in the percentage of total sperm motility grades (A+B+C) and progressive sperm motility grade (A) with a significant improvement (P<0.05) in the percentage of progressive sperm motility grade (B) and morphologically normal sperm when using LC medium in comparison with control medium after 30 minutes incubation. Conclusion: It is concluded from the results of the present study that adding 20% LC to the culture medium ofthe in vitro sperm activation leads to an improvement in certain spermfunction parameters.
Background: Many studies has been found that the pentoxifylline (PX) and L-carnitine (LC) as motility stimulants showed a positive effect on the activation of sperm in vitro and improve the forward movement after cryopreservation. Objectives: The present study aim to investigate the possibility of using the motility stimulants(i.ePX,LC and mixture of PX with LC medium) for in vitro sperm activation after cryopreservation of epididymal sperms and their effects in vitro on fertilization rate (FR) and early embryonic development (ED) following in vitro fertilization (IVF) process as a model for obstructive azoospermia in men. Methods: Fifty mature male and Fifty mature female mice, 8-12 weeks old were used. Epididymal sperms were obtained from caudal region and in vitro direct activation technique was done after cryopreservation by using four media namely; PX alone, LC alone, a mixture of PX with LC and Ham's F-12 (as control media). The four media were used for IVF of mature ova and FR with ED were recorded. Results: In vitro activation of epididymal sperms with PX, LC and mixture of PX with LC has shown positive effect on sperm concentration, sperm motility, and grade activity of progressive forward movement after cryopreservation. There was a significant (P<0.05) increase of FR in treated media compared with the control medium after 24 hours of insemination. The study showed that the quality and quantity of embryos. Conclusion: It is concluded from the results of the present study that adding PX, LC and mixture of PX with LC medium to the culture medium of cryopreserved epididymal sperms and IVF leads to an improvement in certain sperm function parameters, supports the FR and early ED rate.
Background: Glycyrrhiza glabra(Gg) is a herbal medicine used in the treatment of different syndromes. Recently, the effect of this plant on the reproductive system has been studied. However, its role in in vitro still not clear. Objectives: The present study aims to investigate the possibility of using Glycyrrhiza glabra extract for in vitro sperm direct activation, and its effect on in vivo fertilization rate (FR) and early embryonic development (ED) following intra-peritoneal insemination (IPI). The mice were used as an experimental model for mammals. Materials and Methods: Glycyrrhiza glabra extract (2 mg/ml culture medium) was used for in vitro direct sperm activation technique. The female mice were divided into two groups: spontaneously ovulated (SOM) and superovulated groups (SuOM). Each group was divided into two subgroups, the first: the IPI was accomplished by epididymal sperm activated in vitro by adding 20%Gg to the culture medium. The second subgroup: the IPI is accomplished without adding Gg to the culture medium. Results: In vitro activation of epididymal sperms with 20% Gg has shown positive effects on sperm concentration, sperm motility, and grade activity of progressive forward movement. There was a significant (P<0.05) increase in FR with adding 20%Gg compared with that free of Gg of spontaneously ovulated groups after 24 hours of insemination. The FR in the right oviduct showed highly significant (P<0.001) increment compared to the left. Embryonic development rate significantly ( (P<0.05) increased after 24 and 48 hours of insemination in grouping with adding Gg. The study showed that the quantity and quality of embryos generated from IPI with adding Gg were higher than that of groups inseminated without adding Gg. Conclusion: It is concluded from the results of the present study that adding the 20% Gg to the culture medium of the epididymal sperm and IPI leads to an improvement in certain sperm function parameters and supports the FR in SOM and SuOM with an increase in the embryonic development rate. These data can be utilized for artificial insemination programs.
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.
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.
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.