Applied sciences  
Article  
Pregnancy Achievement by Medical Assisted Reproduction Is Correlated to the G Protein-Coupled Receptor 30 mRNA Abundance in Human Spermatoaa  
[PERSON] 1,2,3,4,5,6, [PERSON] 5, [PERSON] 6, [PERSON] 3,6,7, [PERSON] 6,8,8,  
[PERSON] 1,2,9,10,10, and [PERSON] 4,*  
Footnote 10: https://wwwmale fertility [1; 2]. E2 can mediate genomic effects through specific estrogen receptors (ER\(\alpha\) and ER\(\beta\)), which are widely distributed in different tissues and particularly in the cells of the male reproductive system [3]. Recently, a non-classic receptor, known as the G protein-coupled receptor 30 (_GPR30_), was also found to be essential for mediating the non-genomic rapid effects of E2 on the human testis, being present in Sertoli cells, Leydic cells, spermatagonia, spermatocytes, round spermatids, and spermatozoa [4; 5]. As androgens, E2 can induce a negative regulation of the hypothalamus-pituitary-gonadal (HPG) axis, promoting the down-expression of the gonadotropin-releasing hormone, as well as the release of luteinizing hormone (LH) and follicle-stimulating hormone (FSH). The pituitary sex hormones, LH and FSH, play crucial roles in the maintenance of both steroidogenesis and spermatogenesis [6]. However, the effects of E2 on spermatozoa, particularly the ones mediated by _GPR30_, remain to be elucidated.  
As a haploid cell, sperm DNA is highly condensed, allowing for its safe delivery into the oocyte. In addition, the closed conformation of the genetic material leads to the arrest of almost all transcription activity [7]. Interestingly, mature spermatozoa carry into the oocyte a pool of mRNAs. The origin and function of these mRNAs are still unknown [7; 8]. Nonetheless, recent studies suggest that the male metabolic state can influence the offspring's health. [PERSON] and colleagues reported that, in mice, the induction of obesity by the consumption of a high-fat diet by F0 males altered the germ cell DNA methylation pattern, the expression of testicular mRNAs, and the miRNA sperm content in the males of subsequent generations, even though these animals were fed with a standard diet [9]. Several studies from [PERSON] and colleagues came to a similar conclusion [10; 11; 12]. The authors reported high-fat-feeding male mice (F0) affected the testicular metabolome and function of their sons (F1) and grandsons (F2) [10]. Sperm quality was affected in all generations, even though F1 and F2 males were fed with a standard diet [10; 11]. Interestingly, with a reversion in the diet of the F0 animals from a high-fat to a standard diet, the testicular content in fatty acids was still irreversibly affected, as well as sperm defects [12]. This means that the effects promoted by the high-fat diet could be irreversible and still passable to subsequent generations [12]. Although these studies did not identify RNAs as being responsible for the transmission of these metabolic defects through the generations, another study reported solid evidence of such a finding. [PERSON] and colleagues isolated transfer-RNA-derived small RNAs (tsRNAs) from obese mice sperm. These tsRNAs were later injected into healthy developing embryos, which resulted in the alteration of the expression of genes associated with metabolic regulation [13]. Furthermore, other factors, such as paternal age and obesity, are known to have several detrimental effects on males' fertility (for further details, see [14; 15]). Several studies have already linked high paternal body mass indexes (BMI) and decreased live birth outcomes after medically assisted reproduction treatments (ART) [16; 17; 18]. Other studies have reported a similar trend for advanced paternal age and ART outcomes [19; 20].  
The present study aimed to detect and quantify the abundance of _GPR30_ mRNA in human spermatozoa and to determine its impact on the outcome of ART. Furthermore, we aimed to elucidate if the abundance of this transcript in spermatozoa is associated with paternal BMI and age.