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PubMed2006

Technology insight: In vitro culture of spermatogonial stem cells and their potential therapeutic uses.

Male germline stem cells--spermatogonial stem cells (SSCs)--self-renew and produce large numbers of differentiating germ cells that become spermatozoa throughout postnatal life and transmit genetic information to the next generation. SSCs are the only germline stem cells in adults, because all female germline stem cells cease proliferation before birth. In this article, we first summarize development of SSCs, and then the relation of SSCs to somatic stem cells in tissues and pluripotent stem cells in vitro, such as embryonic stem cells. Next, we describe a transplantation technique in which donor testis cells from a fertile male can be transplanted to the testes of an infertile male where they re-establish spermatogenesis and restore fertility. The transplantation technique has been used to study the biology of SSCs, which made possible the identification of external factors that support in vitro self-renewal and proliferation of mouse and rat SSCs. Since SSCs of all mammalian species examined, including human, can replicate in mouse seminiferous tubules following transplantation, the growth factors required for SSC self-renewal are probably conserved among mammalian species. Culture techniques should therefore soon be available for human SSCs. In the final section, we discuss current and potential approaches for using the transplantation technique and in vitro culture of SSCs in human medicine. Because assisted reproductive techniques to fertilize oocytes with round or elongated spermatids are available, clinical use of cultured human SSCs will be greatly facilitated by development of techniques for in vitro differentiation of SSCs to mature germ cells.

Animals↗

PubMed2005

Sperm and assisted reproduction.

When optimization of the male and female partner has failed, techniques of assisted reproduction may be employed. Their common rationale is to enhance the probability of fertilization by bringing the spermatozoa closer to the oocyte(s). Assisted reproductive techniques are not a curative treatment but they intend to circumvent some functional deficits in gametes. The choice of the most appropriate assisted-reproduction treatment for the individual couple is often based on the quality of the ejaculate or the source of the gametes. Intrauterine insemination (IUI) may be considered as the first-line approach when after semen preparation at least 1 x 10(6) spermatozoa can be harvested. IVF is to be considered when dealing with longstanding infertility in a couple with a normal fertility status in the female partner. The choice between IVF and intracytoplasmic sperm injection (ICSI) is difficult since performing 3 ICSI treatments can prevent one complete fertilization failure even in couples with borderline semen characteristics. When less than 1 x 10(6) progressively motile spermatozoa are available after preparation, ICSI may be the treatment of choice. ICSI can also be performed using surgically-retrieved spermatozoa. In men with obstructive azoospermia spermatozoa can be retrieved through different techniques. However, in azoospermic men with primary testicular dysfunction, excisional biopsies are to be preferred. But even then, spermatozoa may only be obtained in about half of these men when no preliminary selection has been made on the basis of a diagnostic biopsy. Patients with extreme oligozoospermia or azoospermia should be informed about the possible genetic background of their condition that is transmissible to their children and about the limitations of the currently available genetic screening methods. Although currently there is no convincing evidence that ICSI is associated with a substantial higher incidence of congenital malformations, we should not draw the premature conclusion that ICSI is safe. ICSI must be applied with caution and only when no other treatment option is available.

Humans↗

PubMed1998

Manual collection and characterization of semen from Asian elephants (Elephas maximus).

The implications of collecting semen from elephants for use in artificial insemination programs are profound in the context of propagating captive elephants. Using a manual manipulation technique, semen was collected and characterized from five adult Asian elephants (Elephas maximus) and ejaculate fluid was obtained from one castrated elephant bull. The penis was stimulated to protrusion and erection by rectal massage of the pelvic portion of the urethra. During an ejaculatory response, massage was also directed onto the area of the ampulla of the ductus deferens. Sperm rich ejaculates were usually collected as a result of each ejaculatory contraction. Ejaculates were evaluated for spermatozoal concentration and pH (when possible) and sperm rich fractions combined for determination of total volume. Mean total volume of each collection was 27.5+/-4.4 ml. Mean concentration of the first and second ejaculatory responses from a collection was 2.05+/-0.17 x 10(9) and 1.34+/-0.19 x 10(9) sperm/ml, respectively. Measurement of seminal pH revealed no significant differences between the fractions. Mean pH of the first and second ejaculatory responses were 7.05+/-0.07 and 7.04+/-0.13. This method of collecting elephant sperm can be utilized for semen evaluation of bulls of unknown reproductive status in conjunction with other evaluation techniques (i.e. ultrasonographic, endocrinologic). It also has the potential for providing valuable genetic material for genome resource banks and for use with assisted reproductive techniques like artificial insemination.

Animals↗

PubMed2000

[Various features of neonatal outcome in assisted fertilization pregnancies].

Assisted pregnancies have been often considered as high risk pregnancies because of several factors as increased maternal age, primary sterility, pharmacological treatments, embryos and gametes manipulation in the first weeks of life. Many studies showed high rates of perinatal morbidity (especially low birth weight and prematurity), and an increase of maternal morbidity and mortality. The present study was designed to analyse correlation between morbidity and twinning rate, and to show the distribution of birth weight in a population of neonates after assisted reproductive techniques (ART). We studied 352 newborns (168 singletons, 126 twins, 42 triplets, 16 quadruplets) from assisted pregnancies; morbidity was 7.1% in singletons, 11.9% in twins, 21.4% in triplets, raising to 56.3% in quadruplets. Our experience suggests that multiple gestations after ART seem to have an increased incidence of neonatal complications.

Humans↗

PubMed2006

Non-invasive genetic diagnosis of male infertility using spermatozoal RNA: KLHL10 mutations in oligozoospermic patients impair homodimerization.

Infertility affects an estimated 7% of men worldwide, nearly a quarter of whom are diagnosed as idiopathic. The genetic etiologies of idiopathic male infertility are unknown, partly due to lack of simple diagnostic techniques. Moreover, the transmission risk of such genetic defects to offspring born from assisted reproductive techniques is increasingly becoming a concern for physicians and infertile couples. We explored the feasibility of obtaining full-length mRNAs from transcriptionally inert human spermatozoa in semen as a non-invasive diagnostic tool for identifying germline mutations in candidate infertility-associated genes. The efficacy of reverse-transcription PCR on spermatozoal RNA from infertile patients with wide-ranging sperm concentrations varied between 91 and 99% for multiple haploid germ cell-expressed genes. Using this methodology, we identified seven oligozoospermic patients with missense and splicing mutations in the germ cell-specific gene, KLHL10. Three of 270 (1.1%) severely oligozoospermic patients (<10(6) sperm/ml) harbor KLHL10 alterations that were absent in 394 controls and exhibited significant association (P=0.02). Two KLHL10 missense mutations (A313T and Q216P) resulted in impaired homodimerization with the wild-type protein in yeast interaction assays, suggesting a functional deficiency. This study demonstrates the utility of this approach for analysis of haploid germ cell-expressed genes regulating post-meiotic events including sperm maturation, motility and fertilization. The development of non-invasive techniques to analyze genetic defects of human spermatogenesis, previously possible only with invasive testis biopsies, provides important diagnostic and therapeutic implications for reproductive medicine.

Amino Acid Sequence↗

PubMed2000

Clinical results of the intra cytoplasmic sperm injection (ICSI) at Baqai Institute of Reproduction and Developmental Sciences (BIRDS).

OBJECTIVE: To analyse the results of the new revolutionary technique of Intra Cytoplasmic Sperm Injection (ICSI) for male infertility. Furthermore, to find out the ways of reducing cost of the expensive methods of Assisted Reproductive Techniques. DESIGN: This is an analytical study of results of 71 patients (86 Cycles) of ICSI performed in first year of its launching in Baqai Institute of Reproduction and Developmental Sciences (BIRDS) from May 1997 to April 1998. Men with semen reports of parameters less than WHO criteria were treated including Azoospermic men. RESULTS: After ICSI procedure a fertilisation rate of 58.9% was achieved in 86 cycles. A total of 17 clinical pregnancies were had from the transfer of fresh embryos. This includes three pregnancies from Surgical Sperm Collection in azoospermic men. Cost could be reduced by avoiding high dosage of drugs and by cutting out serial serum estrogen levels. Ultrasonic assessment for follicle monitoring is equally satisfactory. CONCLUSION: ICSI proved to be the only successful treatment for men with poor semen quality. It is giving nearly 20% chances of fatherhood in a man who was labeled infertile previously. Results are promising and will be improving with time. The cost could be reduced to an extent by cutting down drug doses and laboratory investigations without compromising the results.

Adult↗

PubMed1996

Collection of semen from marmoset monkeys (Callithrix jacchus) for experimental use by vaginal washing.

Assisted reproductive techniques make an important contribution to the conservation of endangered primate species. In our laboratories marmoset monkeys (Callithrix jacchus) are used as a model species for developing assisted reproductive technologies for New World primates. The studies require a reliable method for collecting functional sperm from these small animals. For this purpose a minimally invasive procedure, vaginal washing after natural mating, was evaluated for its suitability as a routine method of obtaining ejaculated sperm of high quality. The objective of the first series of tests was to identify a behavioural pattern which was easily discernible by the observer and provided a reliable indication that ejaculation had occurred. In a second series of tests the influence of length of separation of the male prior to copulation on the quality of the ejaculate was evaluated. Six adult ovariectomized and five adult intact females were used for vaginal washing, while their mates served as donors of ejaculates. Matings at specific times were achieved by separating the males from their females for a certain time period and subsequently introducing either their own mates or unfamiliar males to the females. After each observed mating vaginal washing was performed on the unsedated females. The seminal samples obtained were analysed for sperm concentration, the proportion of motile sperm and the proportion of live sperm. The results have shown (a) that ejaculation was indicated with a reliability of 86.1% by the behaviour of the female, who terminates a copulation by moving away from her partner; (b) that separation of the male for up to six days had no negative effect on the sperm quality; (c) that the sperm samples were of high quality. Vaginal washing after natural matings appears to be a practical, reliable and gentle method for routinely collecting ejaculates from marmoset monkeys and possibly other New World primates.

Animals↗

PubMed1996

Preparation and recovery of frozen-thawed bovine spermatozoa via various sperm selection techniques employed in assisted reproductive technologies.

A number of semen manipulative techniques are currently available to remove the undesirable spermatozoa, debris and other factors and to increase sperm quality. The use of motility stimulants such as caffeine or others could optimize the recovery and quality of frozen-thawed spermatozoa processed by a variety of sperm selection techniques. Frozen-thawed specimens from 5 bulls were slowly diluted and washed with Ham's F-10 medium containing 3% BSA (w/v) and 0 or 2 mM caffeine. Aliquots containing approximately 50 x 10(6) total sperm cells were used for conventional sperm wash, swim-up, Percoll density gradient centrifugation (80, 70, 55 and 40% Percoll gradients) and Sephadex (SpermPrep I) filtration. Quantitative and qualitative characteristics of selected spermatozoa included: total sperm (x 10(6)), percentage and grade (0 to 4) of motility, percentage of spermatozoa with coiled tails and response to the hypoosmotic swelling (HOS) test (percentage of swollen spermatozoa). When compared to washed specimens, fewer spermatozoa were recovered via the swim-up, Percoll and SpermPrep I filtration methods. Quantitative and qualitative characteristics of these spermatozoa improved further after processing with Ham's F-10 containing 2 mM caffeine, followed by selection via the various techniques. Enhancement of sperm motility, in conjunction with the most appropriate sperm selection technique, represents an efficient method for the recovery of spermatozoa with improved qualitative characteristics.

Journal Article↗

PubMed1996

The outcome of multiple pregnancy.

The incidence of multiple pregnancy and delivery has increased dramatically over the past 10-15 years in many developed countries of the world. Data for England and Wales show that between 1980 and 1993 the twin maternity rate increased by approximately 25% and the triplet and higher order maternity rate more than doubled. Similar trends have been reported elsewhere. The majority of these increases have been linked to the use of ovarian stimulants and assisted reproduction techniques, and multiple pregnancy must be considered to be one of the most important adverse outcomes in current methods of infertility treatment. Obstetric complications associated with multiple pregnancy include prenatal screening problems and increased incidence of pregnancy-induced hypertension, antepartum haemorrhage, preterm labour and assisted or surgical delivery. Neonatal problems include low birthweight and increased prevalence of congenital malformations. Compared with singletons, neonatal mortality was seven times higher in twins and > 20 times higher in triplets and higher order births in England and Wales in 1991. Survivors also suffer higher rates of cerebral palsy and other neurological impairments. Most studies of pregnancies and babies resulting from assisted reproduction have demonstrated similar, if not higher, risks of adverse obstetric and neonatal outcomes for multiple births compared with national expectations. A poorer outcome in multiple pregnancy, especially in triplet and higher order pregnancy, supports the replacement of two good quality embryos in assisted reproduction treatment cycles.

Female↗

PubMed2026

Hysteroscopic platelet-rich plasma and medically assisted reproduction outcomes: a systematic review and SWOT analysis.

BACKGROUND: Platelet-rich plasma (PRP) has been proposed as an adjuvant treatment in reproductive medicine. While most evidence refers to blind intrauterine instillation, subendometrial administration under hysteroscopic guidance allows targeted delivery under direct visualisation. This systematic review aimed to synthesise the available evidence on hysteroscopic PRP administration and its impact on clinical medically assisted reproduction (MAR) outcomes. METHODS: A systematic search was conducted from inception to December 2025 across major databases. Studies were included if they evaluated hysteroscopic PRP administration in women undergoing MAR, comparing reproductive outcomes between treated and control groups. RESULTS: Out of 142 records, 3 studies met the inclusion criteria. Study populations were heterogeneous and included women with refractory thin endometrium and/or a history of implantation failure. Hysteroscopic PRP administration protocols varied in timing, technique, and dosage. In a prospective case-control study, hysteroscopic intraendometrial PRP injection at a depth of 2-3 mm in the four uterine walls, using an ovum aspiration needle, on days 11-13 of the cycle prior to euploid frozen embryo transfer (ET), was associated with higher implantation (IR), clinical pregnancy (CPR), and live birth rates (LBR) compared with standard therapy. Conversely, no significant differences in CPR, miscarriage rate, or LBR were observed in an observational study evaluating a single intraendometrial PRP injection (35-40 mL, 2-3 mm depth), administered via endoscopic needle on days 6-8 of the menstrual cycle preceding frozen ET, alone or after electrical impulse therapy. A randomised controlled trial in women undergoing intrauterine insemination reported a significant improvement in CPR following hysteroscopic subendometrial PRP instillation in the four uterine walls (1.0 mL each). CONCLUSIONS: Current literature on hysteroscopic PRP administration in reproductive medicine is limited, and robust conclusions cannot yet be drawn. Well-designed randomised controlled trials with standardised protocols are needed to clarify its clinical role.

Humans↗

PubMed1996

Male fertility following spinal cord injury: facts and fiction.

Although most spinal cord injuries (SCIs) occur to males of parenting age, myths abound as to whether men can biologically father children after SCI. Following SCI, most men experience impairments in erectile and ejaculatory function. Semen can be obtained from almost all men with SCI through the use of vibratory or electrical stimulation. Their ejaculates often have normal sperm counts but more immotile sperm than found in men without SCI. This condition does not seem to be caused by lifestyle factors (such as elevated scrotal temperature, ejaculation frequency, and method of bladder management) but may be related to factors within the seminal plasma. Sperm from men with SCI may be used in assisted reproductive techniques to attempt biological fatherhood. Health care providers, including physical therapists, should encourage men with SCI seeking biological fatherhood to be evaluated at a center dedicated to assisting men with SCI in this specialty area.

Body Temperature↗

PubMed2001

Need for sperm retrieval and cryopreservation at vasectomy reversal.

PURPOSE: Controversy exists on whether to obtain sperm for cryopreservation routinely at vasectomy reversal. With recent improvements in in vitro fertilization with intracytoplasmic sperm injection, it is now possible to obtain a small amount of testicular tissue for cryopreservation in the event of reversal failure. However, to our knowledge no studies exist of who is most likely to benefit from this procedure. MATERIALS AND METHODS: We reviewed 84 consecutive vasectomy reversals performed by 1 surgeon (J. I. S.) between July 1996 and March 2000 with followup available for 77. We grouped cases by procedure as vasovasostomy, vasoepididymostomy and vasovasostomy with vasoepididymostomy as well as bilateral or unilateral. Sperm was retrieved at reversal in 15 of 46 vasovasostomy (none used), 11 of 18 vasoepididymostomy (3 used) and 13 of 20 vasovasostomy with vasoepididymostomy (none used) cases. RESULTS: The overall anastomotic patency rate after unilateral or bilateral vasovasostomy, unilateral vasovasostomy with contralateral vasoepididymostomy and unilateral or bilateral vasoepididymostomy was 96%, 83% and 57%, respectively. The natural pregnancy rate without in vitro fertilization was 57%, 50% and 14%, respectively. The most recent vasoepididymal anastomoses were performed by the Berger triangulation technique with a 78% patency and 25% pregnancy rate. Only 8% of men with banked sperm eventually used it for assisted reproductive techniques, in whom unilateral or bilateral vasoepididymostomy failed in all. CONCLUSIONS: We currently do not recommend routine sperm retrieval for cryopreservation in men who undergoing vasovasostomy. We encourage men who require bilateral vasoepididymostomy to bank sperm at reversal. In men who undergo vasovasostomy with vasoepididymostomy we base the decision on preoperative counseling and intraoperative findings.

Cryopreservation↗

PubMed1997

Advances in the evaluation and treatment of the infertile man.

Numerous advances in technology have been made in the last several years in the diagnosis and treatment of the infertile man. Using case presentations, this article highlights these important new developments and reviews the basics of a comprehensive male infertility evaluation. The use of ultrasound as a critical, indispensable, yet noninvasive tool in the evaluation of possible male reproductive tract obstruction is discussed. Since assisted reproductive techniques (ARTs) have become increasingly important in the management of the otherwise untreatable infertile man, specialized testing of sperm function, e.g., the sperm penetration assay (SPA), and strict morphology assessment are often useful prior to the initiation of some of these assisted reproductive procedures such as intrauterine insemination (IUI) or in vitro fertilization (IVF). Testicular touch preparation cytology and image analysis are also important additions to routine testicular biopsy for quantifying the extent of spermatogenesis. Finally, the revolutionary micromanipulation procedure of intracytoplasmic sperm injection (ICSI) has forever changed the practice of andrology by enabling men previously thought to be irreversibly infertile the chance to initiate their own biologic pregnancy.

Humans↗

PubMedDate not supplied

Indications for corticosteroids prior to epididymal sperm retrieval.

OBJECTIVE: To evaluate the effectiveness of low-dose intermittent prednisone treatment designed to reduce antisperm antibodies for men prior to epididymal sperm retrieval and in vitro fertilization. STUDY DESIGN: This was a retrospective review of 75 subfertile men with reproductive obstruction due to congenital absence of the vas deferens, or other reproductive tract obstruction, who underwent a total of 98 episodes of microsurgical epididymal sperm aspiration (MESA) with in vitro fertilization. RESULTS: For couples in whom the man was pretreated with prednisone, better fertilization rates (39% vs. 21%, P < .0001) and pregnancy rates (48% vs. 26%, p = 0.06) were obtained than in couples in whom the man was not pretreated. The benefit of prednisone treatment was most noticeable for men with preoperatively detectable antisperm antibodies. Prednisone-treated men with preoperative antisperm antibodies showed improved fertilization rates (40% vs. 9%, p = 0.005). Men without antibodies who were treated with prednisone showed a decrement in fertilization rate, from 73% to 24% (fertilizations per oocyte, P < .0005). Success of prednisone treatment could not be correlated with specific assisted-reproduction technique (IVF, PZD, SuZI, or ICSI). CONCLUSIONS: Preoperative evaluation of antisperm antibody status and treatment of antibody-positive men prior to epididymal sperm retrieval may play a role in improvement of the fertilization and pregnancy results achieved by centers that provide MESA with assisted reproduction. These results are most important when MESA is performed without the assistance of intracytoplasmic sperm injection. Empiric treatment with prednisone may be detrimental to the fertility of men who have no antisperm antibodies.

Adult↗

PubMed1994

Pentoxifylline potentiates ionophore (A23187) mediated acrosome reaction in human sperm: flow cytometric analysis using CD46 antibody.

The study was designed to investigate the effects of pentoxifylline on the acrosome reaction of human spermatozoa in vitro, and to determine whether the reaction is differently modulated after sperm selection by multiple tube swim-up and Percoll buoyant density centrifugation. The acrosome reaction was induced in vitro by using calcium ionophore (A23187) and was detected by measuring the fluorescence of FITC-conjugated goat anti-mouse immunoglobulin bound to CD46 antibody (which binds to the CD46 receptor site on the inner acrosomal membrane) by flow cytometry. Spermatozoa separated on Percoll displayed significantly lower spontaneous acrosome reactions (P = 0.002) than did those separated by the swim-up technique. Pentoxifylline did not, by itself, induce acrosome reaction, but after induction with ionophore, it significantly increased the reaction (P = 0.003) and this increase was seen to be greater when Percoll separation was used as compared to the swim-up technique (P = 0.0002). We therefore conclude that Percoll selection of motile spermatozoa together with pentoxifylline treatment may be of value in assisted reproductive techniques, as an increased ARIC score arose after both treatments, and that flow cytometry allows a precise and rapid quantification of the acrosome reaction.

Acrosome↗

PubMed2004

Preimplantation genetic diagnosis and the welfare of the child.

Preimplantation genetic diagnosis is a form of very early prenatal diagnosis. The technique combines assisted reproductive technology with molecular genetics and cytogenetics to allow the identification of abnormalities in embryos prior to implantation. Since its introduction in 1990 this approach has been applied to an increasing number of single gene disorders, chromosomal rearrangements, and more recent indications such as aneuploidy screening and HLA matching. Since its inception the technology has attracted much attention: geneticists have expressed concerns about the robustness and validity of diagnosis based on single cell analysis, perinatologists were anxious about the effect of embryo biopsy on normal fetal development; and philosophers and ethicists have argued the cases for and against embryo selection. This article attempts to highlight the difficult choices and ethical challenges confronting patients and clinicians in an effort to balance the recognition of parental autonomy with the obligation of clinics to consider the welfare of any child born as a result of this treatment.

Child↗

PubMed2000

Evidence for major histocompatibility complex-mediated effects on spermatogenesis in humans.

Studies in fertile and infertile populations suggest an influence of genes of the major histocompatibility complex (MHC) on reproduction, although it remains unresolved if MHC-mediated effects on fertility are based on direct immunological or non-immunological effects of human leukocyte antigen (HLA) genes or rather on defects in MHC-associated non-HLA genes which affect gamete quality and embryonic development. We analysed allele frequencies for HLA class II loci DQA1, DQB1 and DRB1 and HLA class II haplotype frequencies in couples with tubal and andrological infertility who were treated with assisted reproductive techniques. Males with severe andrological infertility had significantly different allele frequencies for all three HLA loci when compared to males with normozoospermia, whereas no difference was found in the females. Differences in allele frequencies were stronger when only males whose partners achieved pregnancies after assisted reproduction treatment were compared. In those subgroups, we could also observe significant differences in three locus HLA class II haplotype frequencies. In summary, patients with male factor infertility differ in their HLA class II allele constitution from males with normozoospermia, which suggests that genes identical to or located in close vicinity to HLA class II genes may influence spermatogenesis and male gamete function.

Alleles↗

PubMed1996

[Modern fertilization techniques].

In Germany, some 17.2% of married couples are unintentionally childless. Of these, about one-third have a male factor infertility (with disordered spermatogenesis), which in part is unresponsive to drug treatment. In such cases, assisted reproductive techniques are giving childless couples new hope. The techniques are based on two principles: 1) in vitro improvement in semen quality in the laboratory, and 2) the overcoming of natural barriers to fertilization. The latter includes intra-uterine insemination (IUT), in vitro fertilization and embryo transfer (IVF/ET), gamete intrafallopian transfer (GIFT), subzonal sperm insemination (SUZI), zona pellucida drilling, and intracytoplasmatic injection of sperm (ICSI). ICSI has also used epididymal spermatozoa obtained by microsurgical aspiration (MESA) and sperm extracted from testicular biopsies (TESA). These methods have their own specific indications, contraindications and pregnancy rates. However, assisted conception techniques should not be considered outside the overall situation of the couple, including ethical, sociological, physiological and general medical problems.

Female↗