Incidences of autoimmune illness are still substantially lower in men than in women, yet males make 20 times more testosterone than women do. As a result, androgens are thought to have an impact on both the innate immune system’s development and operation as well as the adaptive immune system [38]. Loss of testicular immune privilege was observed in mice with AR deficiency, which is characterized by an increase in serum levels of inflammatory cytokines, such as IL-1b and tumor necrosis factor α (TNF-α), and the number of macrophages in circulation demonstrating the function of androgen in testicular immune privilege in Sertoli cells [39]. Testosterone levels are lower in rats with experimental autoimmune orchitis (EAO), a type of male immunological infertility.

Testosterone is the major androgen in the testes. Less than 3% of testosterone circulates in a free, bioavailable form. Sex hormone-binding globulin (SHBG) plays a crucial role in regulating the amount of unbound testosterone accessible to target tissues due to its high affinity for testosterone. Upon entering target cells, testosterone is converted into the most biologically active androgen, dihydrotestosterone (DHT), by the enzyme 5α-reductase, predominantly in the male reproductive organs. Additionally, testosterone can undergo metabolism by aromatase, leading to the production of estradiol. This conversion primarily occurs in adipose tissue, the hypothalamus, and hematopoietic cells [40]. In comparison to the EAO rat without testosterone replacement, testosterone therapy had a protective effect on disease development and the inflammatory response. It prevented the growth of macrophages and decreased the number of CD4+ T cells while increasing the number of regulatory T cells (Tregs) in the testes [41]. While having no effect on testicular macrophages, testosterone reduces the lipopolysaccharide (LPS)-induced inflammatory response on TNF-α mRNA expression in Sertoli cells and peritubular cells, which support spermatogenesis, spermatozoa transport, and testicular immune control [42]. Androgens affect the quantity and secretion of testicular immune cells, which in turn affects the function and reactions of innate immunity cells in mammals as well as the immunosuppressive effect on male reproduction. It’s evident that androgens influence the innate immune response through various mechanisms such as cell proliferation, cytokine secretion, and toll-like receptor expression. Interindividual genetic variations within innate immune genetic pathways, along with non-infectious environmental factors, could further influence the effects of androgens on the innate immune response [43]. The function and mechanism of androgens in controlling the testicular immunological state have yet to be fully understood.

As important physiological regulators in men, oestrogens (estrogens) have a positive impact on the immune system [38]. The mammalian immune system expresses two intracellular ER subtypes (ERα and ERβ) that control the formation of immune cells and the innate and adaptive immune systems [44]. Many different immune cell types (Figure 4), including T cells, B cells, macrophages, dendritic cells (DCs), and natural killer (NK) cells, express both ERα and ERβ [45]. In males, estrogen controls immunity and upholds immunometabolic function. The over expression of human aromatase genes in transgenic male mice (AROM+ animals) stimulated testicular macrophage activation; however, a rat model of EAO showed increased testicular macrophages, demonstrating the stimulating influence of oestrogens on the immunoregulation of male reproductive function [46]. Androgens and oestrogens have varied effects on various immune cell subsets, and sex steroids as a whole regulate the immune system. Generally speaking, androgens seem to have a stronger immunosuppressive effect than estrogen does on immune cells and immunological function. As a result, although oestrogens have immunoenhancing effects in the immune-privileged environment of the testes, androgens have immunosuppressive effects, which calls for more research.

Macrophages are immune cells that are active in most prevalent in the interstitial area of the testes. For instance, around 80% of the testicular leukocytes in rat testicles were macrophages. According to a recent study on human testes, resident macrophages in the testes make up about 62% of the testicular myeloid cells and express the M2 macrophage marker (CD163) at levels six times greater than those of blood monocytes [52]. In a stable state, these macrophages can sustain themselves for extended periods of time without needing blood monocyte replenishment. Leydig cells and interstitial macrophages have intimate interactions, which may help testosterone production. For instance, when Leydig cells were cultivated in the medium that had been conditioned by testicular macrophages, the production of testosterone increased dramatically, although it had less of an impact on the conditioned medium from peritoneal macrophages [53]. By interacting with the AR in Sertoli cells, testosterone controlled the expression of downstream genes such as reproductive homeobox 5 (Rhox5) and then controlled spermatogenesis processes such as maintaining spermatogonial numbers and BTB, having spermatocytes complete meiosis, adhering elongated spermatids to Sertoli cells, and releasing mature spermatozoa [54].

A testosterone derivative called DHT affects the amounts of enzymes and transporters involved in glycolysis, such as lactate dehydrogenase and phosphofructokinase 1, which has an impact on glucose metabolism in rat Sertoli cells [55]. The retinoic acid biosynthetic enzymes aldehyde dehydrogenase 1A2 (ALDH1A2) and retinol dehydrogenase 10 (RDH10), as well as colony stimulating factor 1 (CSF1), may be expressed by testicular macrophages to encourage the proliferation and differentiation of spermatogonial cells. Spermatogonia development would be disturbed in testes lacking in macrophages after ALDH1A2 and RDH10 levels decreased [56]. It is unknown if the immunosuppressive status in the testicular interstitial space was partially caused by the nutritional composition. According to the aforementioned data, testicular macrophages may increase oxidative metabolism while decreasing anaerobic metabolism in order to retain M2 macrophage phenotype. However, further research is required to fully demonstrate the role of macrophages in spermatogenesis and their pattern of metabolism in the testes.

Under physiological circumstances, testicular DCs, a tiny immune cell population within the testes, are phenotypically immature and functionally tolerogenic DCs. Under physiological circumstances, they are connected to the adaptive immune system of the testes and play a role in the inactivation of effector T cells and the formation of Tregs [57]. Additionally, it’s possible that immature DCs play a role in the identification of typical sperm antigens and the establishment of immunological tolerance to safeguard sperm cells under physiological circumstances. For the growth of immature DCs, FAO and oxidative phosphorylation (OXPHOS) are crucial catabolic processes. Notably, substances that support the OXPHOS pathway, such as vitamin D3, may support tolerogenic DC (acquired tolerant status) phenotype and function, including the generation of IL-10 [58]. IDO (indoleamine 2,3-dioxygenase), an enzyme that catalyzes’ the metabolism of tryptophan and ultimately produces kynurenine, is typically kept at a basal level in DCs. Kynurenine has been discovered to trigger the development of Foxp3+ (Forkhead box protein 3 gene) Tregs and the overexpression of programmed cell death-1 (PD-1) receptor on activated CD8+ T cells, both of which were linked to immunosuppression. Kynurenine serves as an endogenous ligand for aryl hydrocarbon receptors on T cells [59]. In tumors and the uterus during pregnancy, both of which are immunologically privileged sites like the testes, IDO is said to induce DCs to start immune tolerance and stimulate the production of Tregs [60]. IDO was only weakly expressed by testicular DCs, and EAO rats had decreased IDO activity. These variations could be explained by the Sertoli cells’ indirect inhibition of LPS-induced DC maturation through a paracrine action. The expression of inducible nitric oxide synthase (iNOS), cyclooxygenase 2 (COX2), and IDO is reduced in bone marrow-derived DCs when they are cocultured with Sertoli cells from mouse testes that released galectin-1, whereas the levels of IL-10 and TGF-b1 are dramatically elevated in LPS-stimulated DCs, providing evidence in favor of this theory [61]. By upregulating co-stimulatory proteins like CD40, CD80, and CD86, inflammatory cytokines, and major histocompatibility complex (MHC) class-II, maturing DCs under pathological conditions overcome immune tolerance, which leads to autoimmunity and male infertility [62]. The production of iNOS, a reactive nitrogen species that can limit mitochondrial respiration and reduce OXPHOS, is likewise increased in mature DCs stimulated by LPS [63].

Lymphocytes made up 10–20% of the total leukocyte population and were sparsely distributed across the interstitial space in healthy human and rat testes, while B lymphocytes were not present [64]. However, the number of lymphocytes, including Tregs and effector T helper cells (Th1 and Th17), rises dramatically in the testicular interstitial space after inflammation brought on by infection or autoimmunity. T cells in rat testes are primarily of the CD8+ subset, in contrast to peripheral blood where CD4+ T cells were the predominate lymphocyte subset. A sizable population of NK cells was also discernible [65]. Foxp3+ under physiological settings, rat, mouse, and human testes include the potent immunosuppressive cells known as Tregs, which may explain the immunosuppressive properties of testes [66]. The CD8+ and CD4+ T-cell subsets’ ability to activate may be inhibited by testosterone. Ethane dimethane sulfonate dramatically raised CD8+ and CD4+ T cell numbers by killing Leydig cells. Lower than baseline amounts of CD8+ and CD4+ T cells would result from the addition of testosterone working with recovering Leydig cells [67]. In order to oxidase exogenous FAs, which is slowed by increased glycolysis, mitochondrial FAO and OXPHOS are crucial for Treg production and suppressive activities. Adenosine monophosphate-activated protein kinase (AMPK) and transcription factor Foxp3 are two regulators of Tregs suppressive function that work by preventing glycolysis and increasing FAO by blocking motor activity [68]. To encourage proliferation and counteract their suppressive effect, Tregs up regulate anaerobic glycolysis and the expression of glucose transporter 1 (GLUT1). A greater number of effector T cell subsets can overcome the inhibitory impact of Tregs by generating proinflammatory cytokines including TNF-α and IL-6 in inflammatory testes, despite higher CD4+ and CD8+ Tregs and TGF-b expression [69]. Thus, excessive T cell activation in autoimmune conditions may not be controlled by Tregs alone.

When T cells are activated, the metabolism of the cell undergoes a dynamic metamorphosis that helps the cell defend itself from infections and coordinate the actions of other immune cells. T cells engage in anaerobic glycolysis, a process typically seen in highly proliferative cells where glucose is digested into lactate rather than oxidized in mitochondria, to meet the bioenergetic requirements for activation. Recent research has described the dynamic profile of CD4+ and CD8+ T cells following activation and shown that glucose plays a critical role in sustaining effector responses and immune cell proliferation [70]. Numerous inhibitory molecules, including IDO and PD-1, change the metabolism of T cells by lowering the expression of GLUT1, which consequently prevents the uptake of glucose. IDO and PD-1 inhibition is also seen in NK cells, however, it is unclear if this affects the ability of these cells to exert effector function by changing their metabolism [71]. Sertoli cells may also play a role in the immunosuppression of the testicular immune privilege.

Sertoli cells may also play a role in the immunosuppression of the testicular immune privilege. Extracellular ATP may operate as a dangerous chemical to peritubular cells upon immune cell activation by activating their purinoceptors (P2RX4 and P2RX7). The expression of pro-inflammatory chemicals like IL-6 and chemokine (C-C motif) ligand 7 (CCL7) is then increased in peritubular cells as a result of ATP-mediated processes [72]. Evidence suggests that preventing the synthesis of glycolytic ATP may prevent IL-33-induced bone marrow-derived mast cell activation and the generation of the proinflammatory factors IL-6 and TNF. Therefore, mast cells’ metabolic byproducts may contribute to testicular inflammation [73]. Immune cells’ phenotypic and immunoregulatory functions are established by both their surroundings and metabolic reprogramming. In contrast, inflammatory phenotype cells employ glycolysis as a primary source of energy more than mitochondrial OXPHOS for quick pathogen elimination. Glucose and FA metabolism primarily promote cell differentiation into immunologically tolerant cell morphologies. Inflammation, autoimmune diabetes, metabolic syndrome, and even male infertility may be brought on by the interruption of metabolic reprogramming. Therefore, investigating the roles of immune cell metabolism in the testes is crucial for learning more about the cellular and molecular mechanisms behind male infertility. And this might make it easier to create brand-new anti-inflammatory medicines that target immunometabolism.

Immune cell’s metabolic reprogramming is primarily responsible for controlling both their phenotypic and adaptability. Given the abundance of immune cells in the testes, pro-inflammatory stimuli that activate and polarize macrophages cause metabolic reprogramming, which further shifts the balanced mitochondrial metabolism in favor of reactive oxygen species (ROS) generation [74]. In vitro Sertoli cell function may be regulated by the IL-1 system, and immature rats that are 21 days old have been shown to have reduced testicular steroidogenesis when exposed to IL-1b [75]. Due to the lipid peroxidation of the cytomembrane, DNA damage, protein oxidation, and enzyme inactivation caused by elevated ROS levels, male infertility is the end outcome [76]. Not only are macrophages among the immune cells that secrete certain types of cytokines in response to stimulations, but these cytokines may also have an opposite effect on immune cells’ metabolic processes and functions. Therefore, immune cells may modify cellular metabolism upon activation, resulting in the release of cytokines that may impair the immune cells’ normal activity in the testes [77].

The male reproductive system has a variety of immune cells that release cytokines in response to infections, external antigens, and chronic inflammation. The earliest sign of innate host defense against inflammation of the reproductive tract comes from the release of cytokines. One of the common etiological causes of human subfertility and infertility is orchitis [78].

In autoimmune orchitis, the immune system’s recruitment was followed by its activation and an increase in the production of pro-inflammatory cytokines including IL-1 and TNF-α. These diverse proinflammatory cytokines alter BTB permeability, which allows them to penetrate the seminiferous epithelium and trigger the death of germ cells. There is mounting evidence that semen contains higher concentrations of cytokines including IL-1b, IL-6, and IL-8, which are frequently linked to inflammation and infection of the male reproductive system [79]. It has been discovered that inflammation in the male reproductive system raises the levels of ROS, which causes cell damage. Concurrently, both necrosis and apoptosis ROS may also contribute to sperm and spermatogenic cell damage, and also harms Sertoli cells and Leydig cells in the testes, which adds to the failure of spermatogenesis [80].

Hyperinsulinemia, hyperlipidemia, hyperleptinemia, and chronic inflammation are features of metabolic syndrome and obesity. By dysregulating the HPT axis and immunological activities in male reproduction, these conditions may also directly impact testicular functions [81]. In obese males, lower testosterone levels are linked to lower LH and worsened semen quality (including lower sperm count and motility as well as morphologically normal sperm) [82]. An obesogenic environment triggers Th1-lymphocyte and M1-macrophage chronic inflammatory responses that eventually lead to systemic inflammation in the male reproductive system and a decline in immune regulatory cells and cytokines [83]. Furthermore, obesity has been linked to elevated ROS in semen, which has a negative effect on the quality and functionality of semen. Obesity and metabolic syndrome have similar effects on male reproductive system and diabetes mellitus (DM) also alters testicular metabolism, which eventually has an impact on spermatogenesis [84].

A redox imbalance caused by excessive ROS generation results in the synthesis of a lot of inflammatory cytokines [87]. Male infertility is linked to oxidative stress brought on by excessive ROS generation and chronic inflammatory conditions that manifest in the body, such as obesity and metabolic syndrome. For men with subfertility, blocking ROS production is regarded as a first line of treatment [80]. ROS play a critical role in various aspects of male reproductive functions, including supporting spermatogenesis, maintaining sperm viability, facilitating maturation, hyper activation, and capacitation, enhancing sperm motility, and triggering the acrosome reaction [88]. It is necessary to use the best management techniques, which include reducing the increased inflammatory state and changing one’s lifestyle with the proper intervention [83]. Adopting a healthy lifestyle, which includes sufficient eating, useful exercise, and successful weight control, stands out as the most important strategy for managing metabolic diseases and ensuring excellent health for male fertility. Metabolites and neutraceuticals like vitamin C, vitamin E, beta-carotene, magnesium, selenium, and zinc that are administered concurrently enhance the immune system’s regulatory mechanisms [89], causing anti-inflammatory activities that might be due to the antioxidant nature. Common antioxidants increase basic semen parameters and lower ROS and sperm DNA fragment levels, all of which have a positive impact on male fertility [90]. The heterogeneous nature of the study designs, including the type of antioxidant, dosages, treatment period, sample size, and included criteria of participants with either poor semen quality or high levels of oxidative stress, may contribute to the conflicting findings of various studies about using antioxidants to improve semen quality. However, there is a lot of variation in the claimed antioxidant effect on semen parameters, and the Cochrane review found it difficult to draw any conclusions regarding how well sperm quality is improved by antioxidants [91]. The effects of antioxidant therapy on male infertility are still debatable. The impact of antioxidant supplementation on male fertility requires further research, particularly with bigger sample sizes and well-designed randomized controlled trials.

According to reports, the hypoglycaemic drug metformin, which is typically used as the first-line therapy for the management of type II DM (T2DM), directly lowers chronic inflammation and ROS while also enhancing insulin sensitivity, hyperglycemia, and dyslipidemia [92]. Additionally, metformin has been shown to ameliorate poor B cell function associated with T2DM in vivo, reduce B cell-intrinsic inflammation in vitro, and block antigen-induced T cell proliferation in an AMPK independent way [93]. Metformin modulates fertility status and sperm quality, particularly in T2DM, by 1) restoring the structure and weight of the testes, epididymis, and seminal vesicles, 2) inhibiting germ cell apoptosis by enhancing the nutritional function of Sertoli cells to produce, and 3) increasing sperm count, motility, and normal morphogenesis. However, the effects of metformin on male reproductive function and fertility are yet unclear. Metformin may have a negative impact on the male reproductive system, either in non-diabetic or non-T2DM circumstances [94]. Since mammalian target of rapamycin (mTOR) signaling pathway is a master regulator of cell growth and metabolism and several human diseases such as cancer, diabetes, obesity, neurological diseases, and genetics caused by disorders due to the deregulation of the mTOR pathway. mTOR is blocked by rapamycin, which also promotes tolerance, the production of memory T cells, and tissue-resident macrophages [95]. Studies have also shown that the stimulation of the mTOR signaling pathways in rat models of chronic nonbacterial prostatitis increases inflammatory immunological responses by inhibiting NF-кB and IL-1b, while rapamycin treatment reverses these effects [96].

A testicular immunosuppressive microenvironment is maintained by high testosterone levels. This is primarily related to the inhibited NF-кB signaling pathway and reduced production of proinflammatory cytokines, which result in maintained M2-like macrophage phenotype and lower systemic inflammation [50]. Some studies have attempted to regulate testosterone in an EAO animal because of the immunosuppressive effects of testosterone. Additionally, they discovered that testosterone therapy dramatically slows the evolution of inflammation. This is accomplished by preventing the influx of inflammatory immune cells and encouraging the growth of capable CD4+CD25+Foxp3+ Tregs in the testes [41, 96]. Aromatase inhibitors raise testosterone levels in obese men and enhance spermatogenesis and sperm quality, although there are still few large-scale, evidence-based researches on the treatment of male fertility [97]. Although spermatogenesis requires an acceptable concentration of testosterone, testosterone therapy that is too aggressive typically worsens male reproductive indices through a negative feedback process, and should not be utilized in the management of fertility [98]. Based on Tregs’ immunosuppressive characteristics, medications that stabilize Tregs [such as glucocorticoids and nonsteroidal anti-inflammatory drugs (NSAIDs)] or medications that encourage Tregs to differentiate (like TGF-b and all-trans retinoids) are anticipated to maximize the efficacy of Tregs-based therapies for autoimmune disease suppression [69].