Article / Mini Review
University of South Carolina School of Medicine Greenville, Greenville, South Carolina , USA
William E. Roudebush, PhD
Department of Biomedical Sciences
University of South Carolina School of Medicine
700 Grove Road
SC 29605 USA
31 March 2021 ; 14 April 2021
Platelet-activating factor (alkylacetylglycerolphosphocholine; PAF) is a potent signaling phospholipid which has been found in numerous cell types in every physiological system studied to date. In reproduction, PAF is found to have a variety of roles, for example: in ovulation, sperm function, and early preimplantation development. The goal of this mini review is to highlight PAF’s synthesis, mechanism of action and its functional role in sperm physiology. PAF functions via a G protein coupled receptor mediated pathway, which ultimately increases intracellular calcium levels to enhance sperm motility required for fertilization. Exogenous PAF was also found to increase fertilization potential of spermatozoa in cases of non-male factor infertility. Finally, the mini review explores various lifestyle factors that could potentially affect PAF levels and fertility.
Platelet activating factor is a phospholipid mediator that belongs to a family of biologically active, structurally related alkyl phosphoglycerides . PAF is continuously produced by platelets, endothelial cells, macrophages, monocytes, and neutrophils in low quantity , as well as by keratinocytes where it functions for cutaneous inflammation . PAF is also found to be present in human sperm where it is found to have a positive relationship with sperm motility . PAF is produced by two distinct pathways namely: de novo pathway, which utilizes 1-alkyl-2-acetyl-sn-glycerol and CDP-choline as substrates of a DTT-insensitive phosphocholine transferase (PAF-PCT), and the remodeling pathway which requires the production of 1-alkyl-2-lyso-sn-glycero-3-phosphocholine (lysoPAF) via the enzyme phospholipase A2 .
The de novo pathway is mainly responsible for the physiological levels of PAF , which is kept low under normal conditions. The de novo pathway starts with the transfer of an acetyl residue onto a phospholipid, alkyl-lyso-glycerophosphate by the enzyme, acetyltransferase to form alkyl-acetyl-glycerophosphate ; Followed by the hydrolysis of alkyl-lyso-glycerophosphate to alkyl-acetyl-glycerol by the enzyme, phosphohydrolase . The final step involves the transfer of CDP-choline to alkyl-acetyl-glycerol by choline phosphotransferase to form the bioactive form of PAF . The remodeling pathway is initiated by the activation of phospholipase A2, which acts on 1-O-alkyl-2-arachidonoyl glycerophosphocholine to yield 1-alkyl-2-lyso-sn-glycero-3- phosphocholine (lyso-PAF) and free arachidonic acid . This is then followed by the transfer of an acetyl residue to lyso-PAF to produce PAF . In sperm, the enzymes lyso- PAF-acetyltransferase and PAF-acetylhydrolase have been shown to be necessary for PAF activation and deactivation respectively .
Human spermatozoa contain varying amounts of PAF, and it has been implicated to function as an autocrine mediator in the aspects of gamete and embryo physiology . PAF functions via a G-protein coupled receptor mediated signaling event  that affects intracellular calcium levels . Kazou, et al.  reported in mouse models that PAF enhances the acrosomal reaction required for sperm capacitation in the presence of calcium ions, further supporting the proposed PAF mechanism of action. There are PAF receptors on sperm cells , and they could be potential targets for therapeutics to improve sperm capacitation and subsequent fertilizing ability. PAF binds to special cell surface G-protein coupled receptors present on sperm cells inducing the formation of inositol triphosphate (IP3) and diacylglycerol (DAG), resulting in an increase of intracellular calcium levels . These findings support the importance of calcium (Ca2+) entry for sperm capacitation in the presence of PAF.
Since its discovery, PAF has been positively linked with increased fertility numerous times. Roudebush, et al.  found that boars with high fertility have a significant amount more PAF than boars with low fertility. Roudebush, et al.  reported that PAF levels were significantly higher (P < 0.01) during the breeding season of squirrel monkeys with a mean of 3.58 ng/10(6) spermatozoa in comparison to the nonbreeding season which had mean of 0.76 ng/10(6). Sengoku, et al.  demonstrated how an antagonist of PAF, called CV-3988, decreased the fertilization ability of human sperm in vitro by both decreasing sperm penetration ability towards hamster oocyte as well as its acrosome reaction. Interestingly, they not only found that PAF increased the penetration rates of human spermatozoa in a sperm penetration assay, but it also reversed the suppressive effects of CV-3988 on human sperm, aside from the penetration effects and the acrosome reactions. Given the positive effects PAF has on sperm motility, research into ways to improve endogenous levels of PAF could be useful in the treatment of infertility associated with low sperm motility. Marie, et al.  were able to stimulate the production of PAF by > 10-fold via the de novo pathway, by adding 2mM of oleic acid to Ehrlich ascite cells. This reveals an avenue to potentially increase the endogenous production of PAF.
Ricker et al  demonstrated how PAF significantly increased sperm motility by the addition of synthetic PAF at 3.69 x 10- 13 – 3.69 x 10-7 mol/L. In contrast the addition of lyso-PAF, which has no PAF biological activity, had no effect on the motility of sperm. Odeh, et al  went on further to conclude that a lower concentration of PAF enhances motility of stallion spermatozoa, whereas a higher concentration of PAF induces the acrosome reaction.
The PAF effect on sperm to increase fertility was even more evident when Grigorious and colleagues  set up a study with fifty-two couples with unexplained infertility. The sperm of each male was treated with an exogenous mixture of PAF that totaled a concentration of 10-7 mol/L in sperm-washing medium. The sperm was prepared using the direct swim-up technique and a maximum of six IUI cycles with or without PAF treatment were given per couple. The clinical pregnancy rate after a maximum of six IUI cycles showed a significant difference with PAF treated sperm having a rate of 23.07% while non-treated PAF sperm was only 7.92% .
Other research – while agreeing with PAF’s positive effect on motility, acrosome reaction, and fertility – are more critical on its safety. Unsal, et al  study focused on PAF and a synthetic dimethylxanthine derivative, called Pentoxifylline effect on sperm DNA damage. Using a sperm chromatin dispersion (SCD) test they qualified sperm with large or medium sized halos as non-fragmented whereas sperm with small or no halos as significantly fragmented. Sperm that was treated with a baseline had a median SCD rate of 6%. Sperm that were treated with PAF had a median SCD rate of 10%. Sperm that were treated with pentoxifylline had a median SCD rate of 15%. It was concluded that even though DNA damage was higher in samples treated with pentoxifylline compared to PAF, both groups had increased DNA damage. It was still noted that PAF seemed to be more innocent when choosing viable sperm cells and in achieving sperm motility in vitro fertilization laboratory.
Wild and Roudebush explored the effects of exogenous PAF on intrauterine insemination (IUI) outcomes . The study involved collecting normal semen specimens from 60 men, preparing to undergo IUI. PAF was exposed to half of the samples collected, while the other half was not exposed. There was a 46.7% pregnancy rate among the group exposed to PAF, compared to the 16.7% pregnancy rate from the unexposed group . The study revealed that PAF treatment improved performance in motile sperm only and subsequent IUI pregnancy outcomes .
A similar study was performed by Stavroula, et al.  to outline the possible effects of sperm treatment with exogenous PAF on IUI in cases with mild male factor infertility. The study included 92 couples with mild male factor infertility, and each couple had 4 IUI cycles with or without exogenous PAF treatment. The results showed a comparable pregnancy rate of 12.24% vs 11.11% between the cases with and without PAF treatment respectively . It concluded that exogenous PAF does not improve clinical outcomes in cases of mild male factor infertility . Since the positive role of PAF is a receptor mediated event, suggests that this sub-population of infertile males have a defect in their respective PAF-receptors. This can be confirmed by the study of Levine, et al. , which demonstrated that abnormal spermatozoa have a different pattern of PAF receptor locations, which differ from its usual localization to the midpiece and equatorial regions of normal motile spermatozoa.
Jarvi, et al.  performed another study to illustrate the potential for PAF in conjunction with albumin could be used to improve sperm motility. Human spermatozoa were incubated with PAF, lyso-PAF or lysophosphatidylcholine concentrations, and varying concentrations of albumin, from 0% to 1.2%. Motility was then evaluated at different time periods from 5 to 240 minutes by computer motion assisted analysis. The results of the study illustrated that 50μM of PAF and 100μM of lyso-PAF, when supplemented with 0.3% albumin increased sperm linear velocity by 41% and 44% respectively, and the curvilinear velocity by 17% and 21% respectively. However, in the absence of albumin, neither PAF nor lyso-PAF induced any increase in sperm motion, when compared to control . Given these findings, the study poses a potential use for PAF and its metabolite in concert with albumin, as a potential treatment for asthenozoospermia .
There is little to no research associated with how lifestyle changes affect PAF directly; however, due to research that significantly correlates PAF with sperm fertility it can be indirectly associated. Jóźków, et al  looked at how several aspects of lifestyle can affect fertility beginning with three sports: cycling, running, and mountaineering. They found that cycling is the most troublesome and is associated with abnormal morphology and reduced motility of sperm due to the mechanical impact sustained from sitting on a saddle, gonadal overheating, wearing tight clothes, and hypogonadism [10,11]. High intensity running was found to decrease sperm density, motility, and morphology . Mountain trekkers were found to have lower sperm concentration, motility, and higher percent of abnormal and immature spermatozoa due to exposure to altitudes higher than 2000m .
Other examples of lifestyle factors that affect fertility is smoking, alcohol consumption, obesity, testicular heat, and vitamin intake. Smoking has been shown to have a strong correlation with % DNA fragmentation index (DFI) which is the most statistically robust test to predict fertility [14,15,16,17,18]. A DFI score of > 30% indicates infertility. Ramalu-Hansen, et al  found that heavy smokers have a 19% lower mean sperm concentration, and a 29% lower total sperm count than nonsmokers. Heavy alcohol consumption was found to have a significant correlation with infertility, while moderate alcohol consumption was found to have moderate impact on fertility . Obesity was found to be present in infertile men more frequently than in fertile men . It was also found that azoospermia and oligozoospermia occur more in obese men than non-obese men . Those who tend to be in environments where the heat of their testicle is more than normal people, like individuals who use the hot tub in abundance, tend to have a significant increase in DNA fragmentation which can lead to infertility. Vitamin C and Vitamin E intake; however, was shown to decrease DNA damage from 22.1% to 9.1% in 64 males with >15% damage .
The lack of available research on the direct effect of lifestyle changes on PAF opens up many avenues in future research projects. Currently, an indirect link is made, by correlating lifestyle behavior with PAF through sperm fertility. As more research is done to potentially correlate direct lifestyle behavior with PAF, it will be interesting to see how individuals can better increase their endogenous PAF concentration. Future projects can assess how exercise, nutrition, or general behavioral lifestyle can increase or decrease PAF concentration which can then lead to increased or decreased sperm fertility. This can allow for individuals struggling with specific fertility problems to have different avenues to fix these issues in a natural manner that goes beyond pharmaceutical products.
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