Reproduction is required for the survival of all mammalian species, and thousands of essential 'sex' genes are conserved through evolution. Basic research helps to define these genes and the mechanisms responsible for the development, function and regulation of the male and female reproductive systems. However, many infertile couples continue to be labeled with the diagnosis of idiopathic infertility or given descriptive diagnoses that do not provide a cause for their defect. For other individuals with a known etiology, effective cures are lacking, although their infertility is often bypassed with assisted reproductive technologies (ART), some accompanied by safety or ethical concerns. Certainly, progress in the field of reproduction has been realized in the twenty-first century with advances in the understanding of the regulation of fertility, with the production of over 400 mutant mouse models with a reproductive phenotype and with the promise of regenerative gonadal stem cells. Indeed, the past six years have witnessed a virtual explosion in the identification of gene mutations or polymorphisms that cause or are linked to human infertility. Translation of these findings to the clinic remains slow, however, as do new methods to diagnose and treat infertile couples. Additionally, new approaches to contraception remain elusive. Nevertheless, the basic and clinical advances in the understanding of the molecular controls of reproduction are impressive and will ultimately improve patient care.

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The biology of infertility: research advances and clinical challenges

Sperm DNA integrity is essential for the accurate transmission of genetic information. It has a highly compact and complex structure and is capable of decondensation-features that must be present in order for a spermatozoon to be considered fertile. Any form of sperm chromatin abnormalities or DNA damage may result in male infertility. In support of this conclusion, it was reported that in-vivo fecundity decreases progressively when > 30% of the spermatozoa are identified as having DNA damage. Several methods are used to assess sperm chromatin/DNA, which is considered an independent measure of sperm quality that may yield better diagnostic and prognostic approaches than standard sperm parameters (concentration, motility and morphology). The clinical significance of this assessment lies in its association not only with natural conception rates, but also with assisted reproduction success rates. Also, it has a serious impact on the offspring and is highly prognostic in the assessment of fertility in cancer patients. Therefore, screening for sperm DNA damage may provide useful information in cases of male idiopathic infertility and in those men pursuing assisted reproduction. Treatment should include methods for prevention of sperm DNA damage.

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Role of sperm chromatin abnormalities and DNA damage in male infertility

Extensive research in our center at the Cleveland Clinic indicates that the seminal oxidative stress test has diagnostic and prognostic capabilities beyond those of conventional tests of sperm quality or functions. An oxidative stress test can accurately discriminate between fertile and infertile men and identify patients with a clinical diagnosis of male-factor infertility who are likely to initiate a pregnancy if they are followed over a period of time. In addition, the test can help select subgroups of patients with infertility in which oxidative stress is a significant factor, and who may benefit from antioxidant supplementation. Incorporation of such a test into routine andrology laboratory practice may be of particular importance to the future management of male infertility. In recent years, the generation of reactive oxygen species (ROS) in the male reproductive tract has become a real concern because of their potential toxic effects at high levels on sperm quality and function. ROS are highly reactive oxidizing agents belonging to the class of free radicals (Aitken, 1994). A free radical is defined as ‘‘any atom or molecule that possesses one or more unpaired electrons’’ (Warren et al, 1987). Recent reports have indicated that high levels of ROS are detected in semen samples of 25% to 40% of infertile men (de Lamirande et al, 1995; Padron et al, 1997). However, a strong body of evidence suggests that small amounts of ROS are necessary for spermatozoa to acquire fertilizing capabilities (Aitken, 1999). Spermatozoa, like all cells living in aerobic conditions, constantly face the oxygen (O2) paradox: O2 is required to support life, but its metabolites such as ROS can modify cell functions, endanger cell survival, or both (de Lamirande and Gagnon, 1995). Hence, ROS must be continuously inactivated to keep only a small

https://onlinelibrary.wiley.com/doi/pdf/10.1002/j.1939-4640.2002.tb02324.x

Oxidative stress and male infertility: from research bench to clinical practice.

The production, differentiation, and presence of male gametes represent inimitable challenges to the immune system, as they are unique to the body and appear long after the maturation of the immune system and formation of systemic self-tolerance Known to protect germ cells and foreign tissue grafts from autoimmune attack, the 'immune privilege' of the testis was originally, and somewhat simplistically, attributed to the existence of the blood-testis barrier Recent research has shown a previously unknown level of complexity with a multitude of factors, both physical and immunological, necessary for the establishment and maintenance of the immunotolerance in the testis Besides the blood-testis barrier and a diminished capability of the large testicular resident macrophage population to mount an inflammatory response, it is the constitutive expression of anti-inflammatory cytokines in the testis by immune and particularly somatic cells, that represents an essential element for local immunosuppression The role of androgens in testicular immune regulation has long been underestimated; yet, accumulating evidence now shows that they orchestrate the inhibition of proinflammatory cytokine expression and shift cytokine balance toward a tolerogenic environment Furthermore, the role of the testicular dendritic cells in suppressing antigen-specific immunity and T-lymphocyte activation is discussed Finally, the active role mast cells play in the induction and amplification of immune responses, both in infertile humans and in experimental models, highlights the importance of preventing mast cell activation to maintain the immune-privileged status of the testis

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The testis in immune privilege

https://www.clevelandclinic.org/reproductiveresearchcenter/docs/agradoc108.pdf

Effect of cigarette smoking on levels of seminal oxidative stress in infertile men: a prospective study☆

There are several mechanisms acting in synergism that can impair sperm characteristics of patients with accessory gland infection In some cases, conventional sperm variables are disturbed with oligo and/or asthenozoospermia In other patients, these sperm variables may appear normal, but the functional capacity of spermatozoa may be impaired In particular, changes in the composition of the sperm membrane may result in reduced acrosome reactivity and capacity to fuse with the oolemma, and oxidative damage of the sperm DNA may induce mutagenesis Changes in the biochemical make-up of seminal plasma can also reduce the in-vivo fertilizing capacity of spermatozoa, and infection-related disruption of the blood-testis barrier can induce the generation of anti-sperm antibodies and immunological infertility Many of these functional abnormalities will not become evident upon 'basic semen analysis', which explains why some authors are unable to link infection of the accessory sex glands to subfertility Also, functional and anatomical damage acquired as a result of infection is often permanent and not reversible by (antibiotic) treatment Clearly, there are many more aspects of male accessory gland infection that require investigation Available data should stimulate clinicians to place more emphasis on the prevention of infection-related infertility than on its treatment, as the latter is often unsuccessful

Mechanisms and effects of male genital tract infection on sperm quality and fertilizing potential: the andrologist's viewpoint

The presence of 2 million or more peroxidase-positive white blood cells per ml of semen, or the diagnosis of male accessory gland infection, is associated with important biochemical and biological changes in semen plasma and in the spermatozoa, reducing their fertilizing potential in vitro and in vivo (e.g., during intra-uterine insemination). In addition to the effects of reactive oxygen species, and its influence on the essential fatty acid composition of the sperm membrane, potentially unfavourable effects can occur through the intermediate of increased concentrations of certain cytokines, and decreased activity of enzymes such as alpha-glucosidase. In contrast, lower numbers of white blood cells may exert beneficial effects on spermatozoa thanks to the increased production of hepatocyte growth factor/scatter factor (a tissue repairing substance), and the stimulation of immuno-competent cells by particular cytokines (e.g., Interleukin-6).

Mechanisms of sperm deficiency in male accessory gland infection.

Summary. A case-control study of clinical, endocrine and seminal characteristics in 86 infertile men and a prospective study of conception rates during 804 follow-up cycles among 75 infertile couples with male immune infertility were performed. Genito-urethral infections or surgery, and palpable epididymal abnormalities were more common among the immunological cases than among the non-immunological infertile controls (n = 180), but sperm concentration, motility and morphology were similar. The spontaneous conception rate in 70 cases was 1.7% per cycle. Treatment by intra-uterine insemination (IUI) resulted in 8.6% pregnancies per cycle, conventional in vitro fertilization (IVF) had a 18.4% success rate per attempt, and two out of three couples treated by intracytoplasmic sperm injection (ICSI) attained pregnancy. Minimal sperm quality required for successful IUI was not lower than that recorded in couples with spontaneous conception.



It is concluded that immunological infertility, as diagnosed by a positive mixed antiglobulin reaction (MAR) test, is associated with a low spontaneous conception rate. Techniques of assisted reproduction are useful in the management of couples with infertility resulting from this pathology, but sperm characteristics required for successful IUI are much higher than those needed for successful IVF. A strategy for the logical management of this pathology is proposed.

Clinical and biological aspects of male immune infertility: a case-controlled study of 86 cases.

The purpose of a debate contribution is to stir up controversy and discussion and a recent paper (Helmerhorst et al. 1999) has done exactly that. However aside from several criticisms already highlighted (Bronson 1999) it seems that the view-point of Helmerhorst et al. (1999) is biased among other things by the selection of references. Both Helmerhorst et al. (1999) and Bronson (1999) fail to mention to mixed agglutination reaction (MAR) test using latex beads instead of red blood cells (SpermMar test; Fertipro Beernem Belgium); (Jager et al. 1978; Vermeulen and Comhaire 1983). In contrast to the polyacrylamide beads used in the immunobead test (MacMillan and Baker 1987) the latex particles have a uniform diameter of 2 µm. In the SpermMar test for immunoglobulin (Ig)G the particles are coated with IgG and they do attach to the region of the spermatozoa where the antisperm antibodies are located. This attachment is obtained by adding strong and highly specific anti-IgG to the mixture of fresh untreated spermatozoa and latex beads (Rasanen et al. 1994). There are also SpermMar tests for IgA and IgM. (excerpt)

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Antisperm antibodies: Use of the mixed agglutination reaction (MAR) test using latex beads

The present study was undertaken to assess the relationship between the results of conventional semen analysis and the sperm motility index (SMI) as measured by the sperm quality analyser (SQA), and to evaluate these in relation to the fertilization and/or pregnancy outcome of assisted reproduction. SMI determinations and conventional semen analyses were performed on 223 samples from subfertile men in two laboratories in Leuven (n = 136) and Antwerp (n = 87), and on spermatozoa prepared on a Percoll gradient (n = 136) used for treatment of male factor infertility in 57 cycles of intrauterine insemination (IUI), 44 attempts at in vitro fertilization (IVF) and 31 attempts at intracytoplasmic sperm injection (ICSI). SMI values for native semen correlated significantly with sperm concentration, motility and morphology. Multiple regression analysis revealed sperm concentration after preparation, and the concentration of motile spermatozoa with normal morphology and SMI (before preparation) to be the independent determinants for SMI after preparation. SMI values were significantly higher after, than before, preparation (p < 0.0001). In regular IVF (n = 44) the percentage of fertilized oocytes correlated significantly (p < 0.05) with sperm motility (A + B%, r = 0.33), with the percentage of spermatozoa with normal morphology (r = 0.46) before preparation, with the values of SMI both before and after preparation (r = 0.54, r = 0.48), with sperm concentration (r = 0.34) and with the motile sperm concentration (r = 0.29) after preparation. For the occurrence of pregnancy (all treatment methods), comparison of areas under ROC curves (AURC) indicated motile sperm concentration after preparation, as well as SMI both before and after preparation, to have the highest AURC, with no significant difference between these values as far as predictive power was concerned. These results indicate that the SQA allows for rapid evaluation of sperm characteristics and of the effectiveness of sperm preparation techniques. However, it is not superior to conventional semen analysis in predicting the outcome of assisted reproduction.

A Mahmoud

Papers

Mechanisms and effects of male genital tract infection on sperm quality and fertilizing potential: the andrologist's viewpoint

Mechanisms of sperm deficiency in male accessory gland infection.

Clinical and biological aspects of male immune infertility: a case-controlled study of 86 cases.

Antisperm antibodies: Use of the mixed agglutination reaction (MAR) test using latex beads

Performance of the sperm quality analyser in predicting the outcome of assisted reproduction