Differences in the Endocannabinoid System of Sperm from Fertile and Infertile Men

Male infertility is a major cause of problems for many couples in conceiving a child. Recently, lifestyle pastimes such as alcohol, tobacco and marijuana have been shown to have further negative effects on male reproduction. The endocannabinoid system (ECS), mainly through the action of anandamide (AEA) and 2-arachidonoylglycerol (2-AG) at cannabinoid (CB(1), CB(2)) and vanilloid (TRPV1) receptors, plays a crucial role in controlling functionality of sperm, with a clear impact on male reproductive potential. Here, sperm from fertile and infertile men were used to investigate content (through LC-ESI-MS), mRNA (through quantitative RT-PCR), protein (through Western Blotting and ELISA) expression, and functionality (through activity and binding assays) of the main metabolic enzymes of AEA and 2-AG (NAPE-PLD and FAAH, for AEA; DAGL and MAGL for 2-AG), as well as of their binding receptors CB(1), CB(2) and TRPV1. Our findings show a marked reduction of AEA and 2-AG content in infertile seminal plasma, paralleled by increased degradation: biosynthesis ratios of both substances in sperm from infertile versus fertile men. In addition, TRPV1 binding was detected in fertile sperm but was undetectable in infertile sperm, whereas that of CB(1) and CB(2) receptors was not statistically different in the two groups. In conclusion, this study identified unprecedented alterations of the ECS in infertile sperm, that might impact on capacitation and acrosome reaction, and hence fertilization outcomes. These alterations might also point to new biomarkers to determine male reproductive defects, and identify distinct ECS elements as novel targets for therapeutic exploitation of ECS-oriented drugs to treat male fertility problems. 

Sperm DNA damage has a negative association with live birth rates after IVF.

Sperm DNA damage has a negative impact on pregnancy rates following assisted reproduction treatment (ART). The aim of the present study was to examine the relationship between sperm DNA fragmentation and live-birth rates after IVF and intracytoplasmic sperm injection (ICSI). The alkaline Comet assay was employed to measure sperm DNA fragmentation in native semen and in spermatozoa following density-gradient centrifugation in semen samples from 203 couples undergoing IVF and 136 couples undergoing ICSI. Men were divided into groups according to sperm DNA damage. Following IVF, couples with <25% sperm DNA fragmentation had a live-birth rate of 33%; in contrast, couples with >50% sperm DNA fragmentation had a much lower live-birth rate of 13%. Following ICSI, no significant differences in sperm DNA damage were found between any groups of patients. Sperm DNA damage was also associated with low live-birth rates following IVF in both men and couples with idiopathic infertility: 39% of couples and 41% of men with idiopathic infertility have high sperm DNA damage. Sperm DNA damage assessed by the Comet assay has a close inverse relationship with live-birth rates after IVF.

Sperm DNA damage has a negative impact on assisted reproduction treatment outcome, in particular, on pregnancy rates. The aim of the present study was to examine the relationship between sperm DNA fragmentation and live-birth rates after IVF and intracytoplasmic sperm injection (ICSI). The alkaline Comet assay was employed to measure sperm DNA fragmentation in native semen and in spermatozoa following density-gradient centrifugation in semen samples from 203 couples undergoing IVF and 136 couples undergoing ICSI. Men were divided into groups according to sperm DNA damage and treatment outcome. Following IVF, couples with <25% sperm DNA fragmentation had a live birth rate of 33%. In contrast, couples with >50% sperm DNA fragmentation had a much lower live-birth rate of 13% following IVF. Following ICSI, there were no significant differences in levels of sperm DNA damage between any groups of patients. Sperm DNA damage was also associated with the very low live-birth rates following IVF in both men and couples with idiopathic infertility: 39% of couples and 41% of men have high level of sperm DNA damage. Sperm DNA damage assessed by the Comet assay has a close inverse relationship with live-birth rates after IVF.

The place of sperm DNA fragmentation testing in current day fertility management

In this debate article, I am going to set out the case that sperm DNA fragmentation testing is essential in current day fertility management because-
• Our current semen analysis testing is unfit for purpose
• Sperm DNA damage testing has strong associations with every fertility check point
• Sperm DNA damage testing has strong associations with miscarriage
• Sperm DNA testing can explain ‘unexplained’ infertility
• There are reasons why sperm with poor DNA are successful in ICSI
• There are no non-invasive sperm function tests that provide the same information
• We need to take a fresh look at the ‘evidence’ against sperm DNA testing
• We have no reason to wait. There are benefits for clinics and couples alike.

TEENAGERS AND ILLICIT DRUG-USE - EXPANDING THE USER VS NONUSER DICHOTOMY

The present study examines proximal and distal factors associated with the use and non-use of illegal substances within a sample of 860 teenagers in North Wales. Arguing that there is predictive utility in expanding the traditional 'users vs non-users' design dichotomy, four groups are identified-resistant and vulnerable non-users and experimental and repeated users. 'Person' variables (life satisfaction, deviance, hopelessness and drug-related attributions) appeared to primarily differentiate the vulnerable group from their resistant counterparts and identify this, as yet non-using group, with user samples. It is suggested that these variables might represent 'risk' factors for illicit substance use and that the group design employed suggests they precede, rather than follow as a consequence of, illicit drug use. Like their resistant counterparts however, the vulnerable group are differentiated from user samples on some lifestyle and context indices. It is argued that these represent 'protective' influences in an otherwise at-risk group of non-users. Variables associated with an escalation of illicit drug use are discussed in considering the differences between the experimental and repeated user groups. Apart from the more proximal factor of drug-related attributions, 'person' variables appeared less involved here. Repeated users did however, tend to use a greater number of drugs, have a greater proportion of friends who also used illegal substances and significantly fewer had a Welsh cultural identity.

The impact of sperm DNA damage in assisted conception and beyond: recent advances in diagnosis and treatment

Abstract Sperm DNA damage is a useful biomarker for male infertility diagnosis and prediction of assisted reproduction outcomes.
It is associated with reduced fertilization rates, embryo quality and pregnancy rates, and higher rates of spontaneous miscarriage
and childhood diseases. This review provides a synopsis of the most recent studies from each of the authors, all of whom have major
track records in the field of sperm DNA damage in the clinical setting. It explores current laboratory tests and the accumulating body
of knowledge concerning the relationship between sperm DNA damage and clinical outcomes. The paper proceeds to discuss the
strengths, weaknesses and clinical applicability of current sperm DNA tests. Next, the biological significance of DNA damage in
the male germ line is considered. Finally, as sperm DNA damage is often the result of oxidative stress in the male reproductive tract,
the potential contribution of antioxidant therapy in the clinical management of this condition is discussed. DNA damage in human spermatozoa is an important attribute of semen quality. It should be part of the clinical work up and properly controlled trials
addressing the effectiveness of antioxidant therapy should be undertaken as a matter of urgency.

Two potential clinical applications of the alkaline single-cell gel electrophoresis assay .1. Human bladder washings and transitional cell carcinoma of the bladder .2. Human sperm and male infertility

Part 1: The alkaline single-cell gel electrophoresis (comet) assay was used to analyse the integrity and DNA content of exfoliated cells extracted from bladder washing specimens from 9 transitional cell carcinoma patients and 15 control patients. DNA damage, as expressed by % tail DNA and tail moment values, was observed to occur in cells from both control and bladder cancer samples. The extent of the damage was, however, found to be significantly greater in the cancer group than in the control group. Comet optical density values were also recorded for each cell analysed in the comet assay and although differences observed between tumour grades were not found to be statistically significant, the mean comet optical density value was observed to be greater in the cancer group than in the control population studied, These preliminary results suggest that the comet assay may have potential as a diagnostic tool and as a prognostic indicator in transitional cell carcinoma, Part 2: Baseline DNA damage in sperm cells from 13 normozoospermic fertile males, 17 normozoospermic infertile males and 11 asthenozoospermic infertile males were compared using a modified alkaline comet assay technique. No significant difference in the level of baseline DNA damage was observed between the 3 categories of sperm studied; however the untreated sperm cells were observed to display approximately 20% tail DNA. This is notably higher than the background DNA damage observed in somatic cells where the % tail DNA is normally less than 5%. Sperm from the 3 groups of men studied were also compared for sensitivity to DNA breakage, using the modified alkaline comet assay, following X-ray irradiations (5, 10 and 30 Gy) and hydrogen peroxide treatments (40, 100 and 200 mu M). Significant levels of X-ray-induced damage were found relative to untreated control sperm in the two infertile groups following 30 Gy irradiation. Significant damage in hydrogen peroxide-treated sperm was observed in sperm from fertile samples, at 200 mu M and in infertile samples at 100- and 200-mu M doses relative to controls. These results therefore indicate that fertile sperm samples are more resistant to X-ray- and hydrogen peroxide-induced DNA breakage than infertile samples. Further studies involving greater numbers of individuals are currently in progress to confirm these findings.

Sperm motility in Solea senegalensis: effect of temperature, salinity, pH and ovarian fluid

A análise da mobilidade seminal é uma ferramenta importante para reprodução em aquacultura. Esta é uma técnica in vitro que auxilia a estabulação, manutenção e selecção de lotes de reprodutores. A análise de mobilidade seminal pode tornar-se potencialmente uma ferramenta para o melhoramento das condições do ambiente de fertilização. A utilização do software CASA (Computer Assisted Sperm Analysis) revolucionou a descrição e quantificação específica da mobilidade seminal. A maioria da informação recolhida sobre mobilidade de sémen de peixes baseia-se em espécies de água doce, pelo que é crucial conhecer as condições óptimas de activação da mobilidade de espermatozóides para novas espécies de de água salgada de interesse em aquacultura tal como Solea senegalensis. A optimização das condições de fertilização desta espécie é particularmente importante já que os lotes de reprodutores em cativeiro podem desenvolver disfunções reprodutoras. Este trabalho teve como objectivo realizar a avaliação das condições óptimas de activação da mobilidade do sémen em S. senegalensis em termos de temperatura, salinidade e pH. O segundo objectivo foi realizar a avaliação da influência de fluido ovárico homólogo (S. senegalensis) e heterólogo (Epinephelus marginatus) na mobilidade seminal de S. senegalensis. Deste modo foram realizados dois conjuntos de experiências: 1) mobilidade de sémen de 7 machos analisado através do CASA em diferentes temperaturas, salinidades e pH, 2) mobilidade de sémen de 8 machos activados na presença de diferentes concentrações de fluido ovárico. Os parâmetros do CASA foram registados e posteriormente analisados através de médias e cluster analysis. Concluiu-se que temperaturas mais elevadas (20 ºC) e baixas salinidades (25 ‰ e 30 ‰) da solução de activação ocorre um melhoramento das características de mobilidade seminal, tal como a velocidade. A presença de fluido ovárico em baixas concentrações melhora as características da mobilidade seminal assim como a longevidade dos espermatozóides. O fluido ovárico é consequentemente um factor que estimula a mobilidade seminal que tem sido negligenciado em estudos anteriores. Este estudo demonstrou que durante a época de reprodução a temperatura da água (20 ºC) e a salinidade (25 ‰ e 30 ‰) no tanque são os principais factores que melhoram a activação da mobilidade do sémen, sendo consequentemente uma contribuição importante para compreender a dinâmica do processo de fertilização em S. senegalensis.

Potential biomarkers on gametogenesis process and in sperm quality of european EEL (Anguilla anguilla)

Dissertação de mestrado, Aquacultura, Faculdade de Ciências e Tecnologia, Universidade do Algarve, 2015

Development and standardization of a protocol for sperm cryopreservation of two important commercial oyster species

Dissertação de mestrado, Aquacultura e Pescas, Faculdade de Ciências e Tecnologia, Universidade do Algarve, 2015

Why do females mate with multiple males ? The sexually selected sperm hypothesis

One debated issues in evolutionary biology is, why in many species females mate with multiple males. Several hypotheses have been put forward, yet the benefits of multiple mating (here defined as mating with several males) remain unclear in many cases. The sperm sexual selection (SSS) hypothesis has been developed to account for the widespread occurrence of multiple mating in females. It argues that multiple mating by females may rapidly spread, when initially a small fraction of the females mate multiply, and if there is a heritable difference among males in one or several of the four characteristics: (1) the quantity of sperm they produce; (2) the success of their sperm in reaching and fertilizing an egg; (3) their ability to displace the sperm that females stored during previous mating; and (4) their ability to prevent any other male from subsequently introducing sperm (e.g., differential efficiency of mating plugs).

Reproduction in the temperate crayfish, Orconectes rusticus: intermale aggression, copulatary behaviour, and sperm competition

The chelipeds of Orconectes rusticus are sexually dimorphic; males possessing the larger. Males use their chelae in intermale aggressive interactions, both to threaten, and assault opponents. In dyadic interactions males with larger chelae were dominant over otherwise physically similar opponents. A high frequency of attack behaviour, coupled with a low frequency of threats during these interactions indicates that actual physical contact is required for opponent assessment. Large clawed males oriented females into the copulatory position faster than small clawed males. Females more frequently escaped the precopulatory-grasp attempts of small clawed males. Additionally, male-female pairs that included a large clawed male remained in copula longer than pairs that included a small clawed male. Sperm of the second male to mate took precedence over the sperm of the primary male. Sperm precedence was incomplete; about 900/0 paternity accrued to the second male.

Sperm utilization patterns in Gryllus integer (orthoptera: gryllidae

Sperm competition is the competition for fertilizations between ejaculates, within a female, following multiple mating. There are four sperm utilization or precedence patterns: first male precedence, where the first male to mate fertilizes most of the eggs laid by a female; last male precedence, where the last male to mate fertilizes most of the eggs laid by a female; "all-or-none" pattern, where sperm from either male fertilizes all the eggs laid by a female but which male's sperm that is used is random; or sperm mixing, where sperm from each male is used equally in fertilizing eggs laid by a female. Intermediate utilization patterns are also possible. Sperm competition occurs in a wide variety of insect species as well as other animals. This study was undertaken to study sperm competition in the field cricket, Gryllus integer. Four experiments were conducted: a radiation and sterilization experiment, a diapause experiment, and 2 competition experiments. It was found that 7,000 rad of gamma radiation sterilized adult ~ integer males. There was no diapause in the laboratory in ~ integer eggs. In the first competition experiment, three groups of females were used: females mated with a normal male, then with a second normal male (NN group); females mated with a normal male, and then with a sterile male (NR group); and females mated with a sterile male, and then with a normal male (RN group). The results obtained from this experiment showed that the mean proportion of eggs hatched was significantly different between 3 groups of females, with the proportion hatched much greater in the NN group than in either the NR or RN groups. The pattern for the proportion of eggs hatched following a double mating most closely resembled a pattern expected if sperm mixing is occurring. Results obtained in the replicate competition experiment showed that the mean proportion of eggs hatched for the females in the NR group was significantly lower than the proportion hatched in the other two groups. This also supports a model of sperm mixing as a precedence pattern. Values calculated following Boorman and Parker (1976), for the proportion of eggs fertilized by the second male to mate following a double mating, were 0.57 in competition experiment 1 and 0.62 in the replicate. These values indicate that sperm mixing occurs in~ integer.

Health, Education and Employment in a Forward-Backward Dichotomy Based on Standard of Living Index for the Tribes in Kerala

The Paper unfolds the paradox that exists in the tribal community with respect to the development indicators and hence tries to cull out the difference in the standard of living of the tribes in a dichotomous framework, forward and backward. Four variables have been considered for ascertaining the standard of living and socio-economic conditions of the tribes. The data for the study is obtained from a primary survey in the three tribal predominant districts of Wayanad, Idukki and Palakkad. Wayanad was selected for studying six tribal communities (Paniya, Adiya, Kuruma, Kurichya, Urali and Kattunaika), Idukki for two communities (Malayarayan and Muthuvan) and Palakkad for one community (Irula). 500 samples from 9 prominent tribal communities of Kerala have been collected according to multistage proportionate random sample framework. The analysis highlights the disproportionate nature of socio-economic indicators within the tribes in Kerala owing to the failure of governmental schemes and assistances meant for their empowerment. The socio-economic variables, such as education, health, and livelihood have been augmented with SLI based on correlation analysis gives interesting inference for policy options as high educated tribal communities are positively correlated with high SLI and livelihood. Further, each of the SLI variable is decomposed using Correlation and Correspondence analysis for understanding the relative standing of the nine tribal sub communities in the three dimensional framework of high, medium and low SLI levels. Tribes with good education and employment (Malayarayan, Kuruma and Kurichya) have a better living standard and hence they can generally be termed as forward tribes whereas those with a low or poor education, employment and living standard indicators (Paniya, Adiya, Urali, Kattunaika, Muthuvans and Irula) are categorized as backward tribes