In this study, we performed a two-sample MR analysis to investigate the causal relationship between male fertility and gut microbiota. The results of this study indicate that Ruminiclostridm6, Prevotella9 and Lachnospiraceae NC2004 group are key gut bacterial species that affect male infertility, abnormal sperm and erectile dysfunction, respectively.
There are various causes of male infertility, and the fundamental cause is that the sperm production process is blocked, resulting in no sperm or mature sperm production [11]. Disruption of microbiota composition or function and subsequent altered host-microbiota interactions perturb the stability of multiple organ systems, including the body [12], while also affecting spermatogenesis [13]. Intestinal flora can act on the male reproductive system through trace elements, metabolism, inflammatory response, and nerve function [14], which is potentially related to male infertility.
Various trace elements in the body can cause abnormalities in sperm. Abnormal gut microbiota and hormone synthesis caused by iron deficiency can indirectly impair spermatogenesis [15]. At the same time, sperm motility needs to increase the concentration of intracellular calcium ions [16]. Hadwan et al. [17] found that semen volume and sperm motility of infertile men with asthenospermia increased significantly after 3 months of daily zinc supplementation at 220 mg. The regulation of intestinal microbiota will change the level of bile acids and further affect the absorption of vitamin A by the host intestine, leading to impaired spermatogenesis [18]. It has been proved that vitamin D supplementation can improve serum 25-OH-Vitamin D3 levels, and become a potential effective method for the treatment or prevention of male semen quality [19]. Various trace elements are involved in the function of the various organs of the human body and can affect male fertility.
The gut microbiota can mediate the metabolite SCFA to affect insulin resistance and further indirectly regulate male fertility. Studies have shown that insulin resistance index is negatively correlated with semen volume and the percentage of progressively motile sperm [20, 21]. In addition, spermine is used to maintain normal reproductive physiology [22] and can be synthesized by endogenous polyamine metabolism or obtained from dietary intake and gut microbiota, such as actinomycetes, fimmicutes, Proteobacteria and Bacteroides [23]. Spermine producing gut bacteria can protect the tests from toxic exposure [24], improve seminiferous tubules, germ cell counts, and promote the expression of genes involved in spermatogenesis. If spermatozoa are deficient, testicular dysfunction can be improved by supplementation with spermatozoa or spermatozoa-producing gut microbes.
Intestinal dysbiosis increases the number of pro-inflammatory bacteria, causes intestinal mucosal injury, high intestinal permeability and intestinal leakage, causes a large number of bacterial metabolites to enter the blood circulation, changes serum cytokine levels, leads to excessive immune response and chronic inflammatory state, and induces endothelial damage and blood-testis barrier (BTB) destruction [25]. It may affect the hormone production of the testis and disturb the production and motility of sperm [26]. Bacterial translocation affects male reproductive capacity. On the other hand, when the flora is disordered, the pathogenic bacteria will release endotoxin into the blood after autolysis, which can cause index [27]. The increase of endotoxin level leads to male infertility, which has attracted wide attention [28]. It has been fully confirmed that endotoxemia has a causal relationship with decreased sperm motility and spermatogenesis disorders [29]. DING et al. [30] found that the increase in the abundance of Prevotella in the phylum Bacteroidetes of the intestinal flora was positively correlated with the increase in circulating endotoxin, which was a potential pathogenic bacteria. Endotoxemia and epididymal inflammation are the final causes of down-regulation of testicular related genes [31]. The "leaky-gut" hypothesis also links endotoxin to hypogonadism, in which endotoxin can inhibit steroid hormone synthesis in Leydig cells and reduce pituitary luteinizing hormone drive, resulting in decreased testosterone production and ultimately reduced spermiogenesis [32].
There is also a significant correlation between gut microbiota and behavioral emotions [33]. However, Pan et al. [34] found that there may be a correlation between GAD and semen quality in male infertility patients. Therefore, intestinal flora imbalance may also affect the emotional processing center located in the hypothalamus through neurotransmitters [35], leading to anxiety and depression, increasing plasma and semen oxidative parameters, causing oxidative damage and damaging sperm quality [36]. In addition, neurotransmitters produced by an imbalance in the gut flora stimulate the HPGA of the hypothalamic-pituitary-gonadal axis, which inhibits the secretion of gonadotropin-releasing hormones LH and FSH, resulting in a reduction in testosterone secretion. In addition, experiments in mice have confirmed that the intake of beneficial flora can change the emotional behavior of the body and the expression of central γ-aminobutyric acid receptor through the vagus nerve [37, 38], and affect HPGA. Alterations in the gut microbiota may also lead to increased serum TMAO levels and promote vascular inflammation, which in turn leads to cavernous endothelial and smooth muscle cell damage and ultimately to the progression of erectile dysfunction [39].
Intracellular flora imbalance can lead to male infertility due to trace element deficiency, abnormal metabolic regulation, and intestinal inflammation due to endotoxemia. It can also affect male fertility by affecting the host's cognitive mood or by modulating reproductive function through neural networks. The combination of the above factors is the internal mechanism of male infertility.
Ruminiclostridm6 is closely related to a variety of digestive disorders. It is important in digestion, absorption and metabolism, and can be involved in the synthesis of secreted polysaccharides. In this study, Ruminiclostridm6 was found to be significantly and negatively associated with male infertility. If the numbers of these bacteria increase, they can induce dendritic cells to secrete inflammatory factors. The activated macrophages and dendritic cells infiltrating the epididymal cavity may capture normal sperm and induce immune damage [40], affecting the male reproductive function. Prevotella9 is strongly associated with abnormalities in male sperm. Ning Ding et al. found that the abundance of Bacteroides and Prevotella9 was negatively correlated with sperm motility through the fecal microbiota transfer technique[30]. Transplantation of fecal microbiota from high-fat diet mice into ND (normal diet) mice led to a significant increase in Bacteroides and Prevotella in the gut [41], resulting in local inflammation and endotoxemia, interfering with sperm epigenetics affecting spermatogenesis, and causing metabolic disorders [30, 42]. This is consistent with the results of the present study. Obesity and metabolic disorders affect the gut microbial ecosystem. This is due to a significant increase in Bacteroides, Prevotella, Rickettsia, and Lactobacillus in the gut microbiota caused by a high-fat diet, resulting in a concomitant increase in endotoxin levels in the circulation. Endotoxin significantly increases inflammatory factors and monocyte chemotactic factors in the epididymis and then inhibits the expression of testicular meiosis-related genes, resulting in a decrease in sperm count. This suggests that the gut microbiota of men who eat a high-fat diet may affect fertility. Prevotella9 helps break down proteins and carbohydrates. A low-fat, high-fiber diet has been suggested to improve glucose metabolism and insulin resistance in male infertile patients, and further improve male sperm abnormalities. These studies have revealed that gut microbiota imbalance may affect the mood of male infertility patients and worsen their fertility decline. The Lachnospiraceae NC2004 group was found to be potentially beneficial and its reduction was associated with diabetes. The bacterium is closely associated with erectile function in men. The underlying mechanism may be that the gut flora, with the help of the nervous system, regulates changes in metabolic signal transduction in the body. After insulin resistance occurs, vascular nitric oxide synthesis disorder and insulin-induced vasodilation dysfunction occur one after another, both of which may be the pathogenic factors of male erectile dysfunction [43]. In addition, nitric oxide, as an important substance to regulate penile erection and arose brain sexual desire, its content will also be affected when intestinal flora is dysregulated, and then affect penile erection [44]. Studies and evidence on this bacterium are insufficient, and in the future transgenic probiotics could be developed for the treatment and prevention of male sexual dysfunction.
Animal studies have also shown that fecal microbiota transplantation can treat infertility and improve sperm quality, suggesting another link between gut dysbiosis and fertility. The improved gut microbiota by alginate-oligosaccharide AOS can be used to promote T2D-reduced semen quality and male subfertility associated with metabolic diseases [45, 46]. Transplantation of pre-diabetic sheep faeces into mice revealed elevated sphingosine levels and impaired spermatogenesis. As a potential prebiotic agent, melatonin can be used to treat male infertility by improving blood glucose and insulin resistance levels [47]. Another study in mice by Zhang et al. [8] also found that FMT improved spermatogenesis, increased sperm concentration, improved motility, increased levels of testicular antioxidants and key reproductive proteins, and increased expression of genes related to spermatogenesis [48].
Prebiotics and probiotics can, to a certain extent, increase the proportion of germ cells, promote the expression of spermatogenic genes, improve transcription factors in vivo, and improve the testicular microenvironment, thereby increasing sperm concentration and sperm motility and improving sperm quality [49, 50]. The results of the current study provide new insights into the mechanisms of action of beneficial gut microbiota in treating men with high risk of infertility who should be treated individually and in combination with their own gut microbiota composition. To find the intestinal microbiota associated with male infertility, with the aim of identifying transgenic probiotics, it is necessary to explore microbiota therapy (FMT) to prevent and treat current fertility problems by regulating body homeostasis and protecting intestinal mucosal barrier.
This study is the first to investigate the causal relationship between male fertility and gut microbiota using MR methods, and various sensitivity analyses were performed to assess the robustness of the analysis. As suggested by the above results, abnormal gut microbiota can emerge to affect male fertility. The limitation of this study is that the target population only includes the European population, which affects the extrapolation of the conclusions. At present, most of the studies on the role of intestinal flora in male infertility focus on animal experiments, and there is a lack of large-scale randomized controlled trials to provide clinical evidence for further exploring the mechanism of intestinal flora in the intervention of male fertility and finding reasonable transgenic probiotics for the treatment of male infertility.