Recent work has suggested that environmental chemicals, including those contained in cigarette smoke, can have adverse effects on the exposed individuals as well as their future progeny. The mechanisms underlying transmission of environmentally induced phenotypes through the germ line are not well understood. However, a predominant process appears to be the establishment of permanent heritable epigenetic alterations, and a number of studies have implicated microRNAs in such processes. Here, we show that cigarette smoke induces specific differences in the spermatozoal microRNA content of human smokers compared with non-smokers, and that these altered microRNAs appear to predominantly mediate pathways vital for healthy sperm and normal embryo development, particularly cell death and apoptosis. microRNA-mediated perturbation of such pathways may explain how harmful phenotypes can be induced in the progeny of smokers.

What are the levels of anandamide (N-arachidonoylethanolamide, AEA) in human seminal plasma and how are these related to abnormal spermatozoa? Seminal plasma AEA levels were lower in men with asthenozoospermia and oligoasthenoteratozoospermia compared with normozoospermic men. AEA, a bioactive lipid, synthesized from membrane phospholipids may signal through cannabinoid receptors (CB1 and CB2) to regulate human sperm functions and male reproduction by modulating sperm motility, capacitation and the acrosome reaction in vitro. Local AEA levels are regulated by the synthetic and degradative enzymes, NAPE-PLD and FAAH, respectively. How the deregulation of this endogenous signalling pathway affects human sperm function(s) is not clear. This was a cross-sectional study of 86 men presenting at an infertility clinic for semen analysis over a period of 2 years. AEA was quantified, by ultra-high performance liquid chromatography-tandem mass spectrometry, in seminal plasma from 86 volunteers. Using qRT–PCR, CB1, CB2, NAPE-PLD and FAAH transcript levels were determined in spermatozoa from men with normozoospermia, asthenozoospermia, oligoasthenoteratozoospermia and teratozoospermia. Normal spermatozoa were exposed in vitro to methanadamide (meth-AEA) to determine its effect on sperm motility, viability and mitochondrial activity. Seminal plasma AEA levels (mean ± SEM) were significantly lower in men with asthenozoospermia (0.080 ± 0.01 nM; P < 0.05) or oligoasthenoteratozoospermia (0.083 ± 0.01 nM; P < 0.05) compared with normozoospermic men (0.198 ± 0.03 nM). In addition, the levels of spermatozoal CB1 mRNA were significantly decreased in men with asthenozoospermia (P < 0.001) or oligoasthenoteratozoospermia (P < 0.001) compared with normozoospermic controls. Supra-physiological levels of meth-AEA decreased sperm motility and viability, probably through CB1-mediated inhibition of mitochondrial activity. The inhibitory effect of meth-AEA was only shown in vitro and may not reflect what happens in vivo. As the regulation of the endocannabinoid system appears to be necessary for the preservation of normal sperm function and male fertility, there may be implications for the adverse reproductive consequences of marijuana use. Exocannabinoids, such as Δ⁹-THC, are likely to compete with endocannabinoids at the cannabinoid receptors, upsetting the finely balanced endocannabinoid signalling system. The importance of the endocannabinoid system makes it an attractive target for pharmacological interventions to control male fertility. This work was funded in part by miscellaneous educational funds from the University Hospitals of Leicester National Health Services Trust to support the Endocannabinoid Research Laboratory of University of Leicester. The authors declare no competing interests.