BPA is an endocrine-disrupting chemical that can induce a variety of adverse effects in mammals and other vertebrates and invertebrates 23. Worldwide, 50–80 million individuals are infertile, an estimate that is likely to increase considerably in the future. Several factors are known to underlie male infertility; exposure to environmental toxicants is one of these factors. Although it is not yet completely understood why the testis is so vulnerable to damage by environmental toxicants such as BPA, it is clear that toxicants affect many, if not all, mammalian organs in some adverse way 24.
In the present study, BPA was administered through a subcutaneous injection to adult rats. It is currently believed that the majority of BPA intake in humans occurs through oral ingestion and is thus subjected to hepatic first-pass metabolism, where ∼99% of orally ingested BPA by adults is rapidly metabolized to an inactive form before entry into the general circulation 25,26. In contrast, BPA delivery by a subcutaneous injection will enter the circulation before liver metabolization; thus, initial exposure levels of free BPA by this route might be markedly higher than current human circulating levels. This must be taken into consideration when evaluating BPA toxicity and its relevance to human health 27.
In this work, there was a statistically insignificant increase in body weight during the 8-week recovery period; also, the mean weights of testes in BPA-treated rats were insignificantly decreased throughout the study period. These results were in accordance with others who observed that successive intraperitoneal administration of BPA to adolescent male mice and rats 28,29 at a dose of 20 mg/kg/day for 4 weeks decreased the prostate and seminal vesicle weights (but not testis or epididymis weights) and also decreased serum testosterone and both liver and kidney weights. However, a significant decrease in testicular and epididymal weight was also reported to occur in adult male Wistar rats exposed orally to 0.2, 2, and 20 µg/kg/day of BPA for 45 days, whereas an increase in ventral prostate weight occurred at all doses 30,31.
Despite relative regaining of the normal morphological characteristics of the seminiferous tubules 8 weeks after BPA injection, residual structural defects in metamorphosis including acrosomal caps, the marginal rings of the exoplaxome, the postacrosomal sheath, and the ectopic ectoplasmic specialization remained; specialization of the caudal part of the spermatid also showed some anomalies. Similar deformities were observed at a dose of 20 µg/kg body weight of BPA by a subcutaneous injection to adult mice and Wistar rats for 6 days 32. Some authors 33,34 have proposed the presence of an AAM (Actin in the ectoplasmic specialization between the Sertoli cell and spermatids, Acroplaxome with marginal rings, a Manchette with perinuclear rings) complex that is responsible for nuclear shaping. Both the AAM complex and the postacrosomal sheath became affected in the spermatids under BPA treatment 32. The postacrosomal sheath serves as a cytoskeletal element to the nucleus and also triggers molecules for oocyte activation at the time of fertilization 35.
Spermatogenesis is a complex and stepwise cellular process that results in the production of ∼25 000 sperm each minute in healthy adult males, and it takes place within seminiferous tubules in the mammalian testis under the strict regulation of testosterone, follicle stimulating hormone, luteinizing hormone, and estradiol 17β 36.
In the current work, the morphological abnormalities that were detected during metamorphosis of spermatids were probably reflected on the ultrastructure of maturing sperm. These abnormalities involved cytoplasmic redundancy, mitochondrial content, outer fiber size and number, in addition to paucity of sperm stored in the epididymis. A decrease in daily sperm production and fertility in adult male rats was reported at oral doses of BPA between 20 and 200 000 µg/kg/day, and the maximum suppression of sperm production occurred at 20 µg/kg/day. At doses below 20 µg/kg/day, daily sperm production was not significantly different from that of the controls 37. It was suggested that there is a subpopulation of cells that are impacted by BPA and that a ∼40% decrease in daily sperm production is the maximum that occurs, irrespective of the dose of BPA administered above 20 µg/kg/day. Similarly, some data reported that after prenatal exposure to ethinylestradiol, maximum suppression of daily sperm production occurred at 2 ng/kg/day, with no further suppression occurring at higher doses 38. In addition, an injection (subcutaneous) of BPA (50 µg/animal, about 15–20 mg/kg/day) for the first 5 days after birth of mice resulted in a decrease in the percentage of moving sperm, an increase in the incidence of malformed sperm, and an increase in the number of ERα-positive cells in the epididymis of SHN strain mice at 10 weeks of age 39.
The ultrastructural adverse effects of BPA on the Sertoli cell population were insignificant and the intercellular cytoplasmic connecting bridges among them were present in all groups; this might indicate spared blood–testis barrier (BTB). Similar findings were reported by others 32,40. It was reported that the BTB is one of the tightest blood–tissue barriers in the mammalian body. It divides the seminiferous epithelium into basal and apical compartments. Meiosis I and II, spermiogenesis, and spermiation all occur in a specialized microenvironment behind the BTB in the apical compartment, but spermatogonial renewal and differentiation and cell cycle progression up to the preleptotene spermatocyte stage occur outside of the BTB in the basal compartment of the epithelium. The BTB is not a static ultrastructure. Instead, it undergoes extensive restructuring during the seminiferous epithelial cycle of spermatogenesis to allow the transit of preleptotene spermatocytes at the BTB. Recent findings have reported on the molecular mechanisms by which environmental toxicants including BPA induce testicular injury by their initial actions at the BTB to elicit subsequent damage to germ-cell adhesion, thereby leading to germ-cell loss, reduced sperm count, and male infertility or subfertility 41.
One week after BPA withdrawal, Leydig cells appeared unhealthy but after 4 weeks, these cells showed some signs of recovery. It was reported that 200 mg/kg BPA and 100 g/kg E2 significantly decreased Leydig cell numbers in the rat testis and BPA directly affected not only the Leydig cells but also the pituitary gland, but the former might be impaired at lower exposure concentrations than the latter 42. In addition, plasma-free testosterone levels were markedly decreased following 8 weeks of 12 mg/kg/day of BPA treatment of adult male mice compared with the control group, and abnormal multinucleated giant cells were found morphologically in seminiferous tubules in the testis following an 8-week BPA treatment at 120 µg/kg/day 43.
In the current work, much apoptotic bodies had desquamated into the lumen of the seminiferous tubules. It was reported that spermatogenesis is highly influenced by external stimuli, such as drugs, radiation, reproductive and somatic pathologies, seasonal breeding, temperature, and environmental pollutants including BPA, which increase the constitutive levels of apoptosis in germ cells 44. Several studies have suggested that increased germ cell apoptosis during spermatogenesis might explain decreased sperm production in patients with oligospermia and azoospermia 45.
After the withdrawal of BPA till the eighth week, the rough endoplasmic reticulum cisternae of prostatic parenchyma appeared distended with a homogeneous content whereas most of the secretory vesicles were empty; this might point to a BPA-induced metabolic error that probably resulted in delayed secretion from the acini in addition to the formation of prostatic concretions as seen in our results. Similarly, dilated cisternae of the endoplasmic reticulum, many protein-filled vacuoles, and large vacuoles containing altered cell organelles were suggestive of apoptosis induced by diabetes in prostate 46.
A growing body of evidence suggests that adult estrogen exposures can be carcinogenic to the prostate gland 47. Elevated estrogen levels during fetal life may affect the expression of genes involved in the morphogenesis of the gland and, in turn, result in persistent changes in the histological architecture of the gland 48. Alterations in the stromal compartment of prostatic tumors may enhance the invasive and/or malignant potential of the nascent epithelial tumor 49.
Penile erection depends on the balance and integration of neurotransmitters, vasoactive substances, endocrine factors, and tissue fibroelastic properties. It can be assumed that antiandrogen, estrogen, or xenoestrogen may affect erectile function 50. The erectile tissue of the penis is composed of elastic fibers, collagen fibers, smooth muscles, arteries, and veins, which are surrounded by a collagenous tunica albuginea. Trabecular smooth muscle tone has been considered as a major contributing factor to trabecular smooth muscle contractility; relaxation of the smooth muscle of the corpus cavernosum is crucial for the development of penile erection 51. For erection to occur, the penile arteries and sinusoids must be dilated, thus increasing the blood flow into the penis, and the distended sinusoids compress the venules against the tunica albuginea 52. In BPA-treated animals, the numbers of smooth muscle fibers were significantly increased within the trabeculae, thus compressing the sinusoidal spaces into anastomosing tunnels that were significantly narrowed, which might decrease the arterial blood flow and thereby affect penile erection. In addition, the thickness of tunica albuginea gained as a result of the marked increase of collagenous fibers might limit the expandability of the tunica albuginea and restricts the engorgement of the cavernosum, which might prevent penile enlargement and erection. Penile tunica albuginea represents an important part of the fibrous corpora cavernosa skeleton; its collagenous fibers, accompanied by elastic networks, are arranged in an outer, mainly longitudinal, and an inner circular layer 53. These fibers allow penile elongation during tumescence and an increase in girth during erection while providing adequate resilience for a rapid return to the relaxed nonerect state at detumescence 54,55. Loss of corporeal elasticity by decreased vascularity with a consequent increased ratio of cavernosal smooth muscle content and thickening of tunica albuginea might be a result of the xenoestrogenic effect of BPA, which adversely affects erectile homodynamic. Also, it was reported that the estrogen receptor pathway, the androgen receptor pathway, or both mediate estrogen-induced developmental penile disorders 56. Chronic exposure of low-dose BPA may result in erectile dysfunction in aging males or may exacerbate the aging process with consequent erectile dysfunction 50.
It is concluded that BPA inhibits spermatogenesis, increases the ratio of sperm anomalies, and exerts a potential harmful effect on erectile function. Some of the exposure to environmental toxicants can be controlled by personal choices, such as deciding not to smoke and choosing glassware over polycarbonate plastic ware; however, these choices cannot completely protect an individual from the harmful effects of environmental contaminants that might be the cause of subfertility or infertility.
Further studies on the effects of different doses of BPA on the physiology, types of collagen, and receptors in human erectile tissue should be carried out to identify the more precise effect of BPA on human erectile function.
The authors thank Qassim University for supporting this study (grant Sabic number SR-S-011-18).
There is no conflict of interest to declare.
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