Cis, a chemotherapy drug used to treat several cancers, causes damage to tissue in the testes despite its potent antitumoral activity. Because chemotherapy agents attack dividing cells, the spermatogenesis process is particularly affected. Previous studies reported that exposure to Cis reduced reproductive function. (2, 5, 11, 12, 14, 32) This study's histological, immunohistochemical, and biochemical results demonstrated Cis's damaging impact on testicular tissue.
Antitumoral drugs that destroy cancer cells, like Cis, damage tissue by causing cells to undergo autophagy and apoptosis. (1–3, 6, 10–14, 22, 27, 28) Exosomes of BM-derived MSCs have been shown to have therapeutic potential in different articles studying tissue damage and degenerative disorders. (15, 19, 33–35) Exosomes function as extracellular organelles with a paracrine/endocrine role, transporting proteins, miRNAs, and enzymes to target cells. (18) Many studies have reported that MSC exosomes have antiapoptotic, anti-inflammatory, and regenerative effects. (17, 18, 33–35)
Cis therapy changed the testicular morphology, induced autophagy, and decreased hormone levels, sperm count, and oxidative stress. A single dose of exo-therapy was shown to have considerable regenerative and antioxidant effects, as well as increased autophagic activity. Hormone levels increased significantly, although sperm motility and count were unaffected. The Cis group exhibited degeneration, atrophied tubular epithelium, cavities, and dilatation of seminiferous tubules. A decrease in tubule spermatogenic cells, germ cell infiltration into the lumen, and interstitial edema were reported. These findings were supported by a decrease in the JTBS score and tubule diameter. In studies, it was reported that seminiferous tubule diameters, germinal epithelial thickness, and testosterone synthesis decreased in Cis-treated rats; testicular atrophy and infertility may develop accordingly. (36–38)
Exosomes are highly effective active paracrine components that play a vital role in cell communication and have a high potential for tissue repair (23). Exosomes establish cell-cell interactions by delivering proteins, miRNAs, and enzymes to the target cell. Exosomes, as extracellular organelles, thus serve a paracrine/endocrine role. (18) Exosomal proteins and microRNAs have a wide range of biochemical and physiological functions, including communication, structure and interactions, inflammation, exosome biogenesis, tissue repair and regeneration, and metabolism. (16, 17, 34) Exo treatment reduced seminiferous tubule degeneration and restored germinal epithelial cell structure. Edema continues in this group. Additionally, the JTBS socket and tubule diameters increased in this group.
Cis, an alkalizing chemical, cross-links to the inner and outer chains of DNA, generating 'adduct' forms that can be seen as twisted DNA chains. Sperm are presumably impacted by this mechanism, as is the testicular structure. (36) In studies, Cis treatment caused a significant decrease in sperm count and sperm motility in rat testes. (27, 37, 38) In this study, sperm motility and count were significantly decreased by Cis therapy. A significant increase in sperm count was detected in the Cis + Exo group that received Exo therapy, but sperm motility did not significantly increase. Sperm count and motility increased when Exo was given as only one dose on the third day following Cis therapy, but motility did not significantly increase.
Because exosome uptake is dependent on intracellular and microenvironmental acidity and tissue injury is frequently characterized by tissue acidosis, exosomes would be preferentially endocytosed by cells in an injured tissue, implying that exosomes could be home to injured tissues. (35, 39) Furthermore, exosomes are small, nanometer-sized particles that can easily be carried via blood and other bodily fluids, as indicated by the quantity of exosomes seen in most biological fluids, and MSC exosomes can interact with cells in a paracrine and endocrine way. As a result, exosomes have the biophysical capacity to orchestrate MSC interactions with a variety of cell types in both local and distant environments. However, the ability to evoke appropriate cellular responses after tissue injury in order to restore and maintain tissue microenvironment homeostasis would be dependent on the biochemical capability of its protein and RNA cargo. (33)
Autophagy, which is essential for cellular adaptability or survival, can be triggered by a variety of stressors, including metabolic stress, endoplasmic reticulum (ER) stress, and chemotherapeutic drugs. (40) An intense energy is expended in the process of sprematogenesis. In this process, starvation, chemical exposure and radiation negatively affect this process. The connection between apoptotic and autophagic molecular mechanisms in these pathophysiological conditions has been shown by the studies carried out. (41, 42) Cis-induced oxidative stress and tissue acidosis in testicular tissue stimulated the apoptotic pathway and autophagy process. Autophagy induced by Cis in testis was detected by increased Beclin-1, LC3-2 and p62 proteins. Exo therapy at a single dose significantly improved in tissue repair via regulating Cis-induced autophagy. Previous studies have showed that exos protected against testicular ischemia/reperfusion injury in rats through antioxidant, anti-inflammatory, and antiapoptotic mechanisms (43, 44). It has also been reported that antitumoural drugs trigger the autophagy process. (22) Exosomes have been shown to have immunoregulatory, anti-inflammatory, regenerative, and anti-apoptotic capacities in therapeutic applications, however it is unclear whether or if such characteristics regulate autophagy. (19)
Oxidative stress, defined as an imbalance between free radicals and the antioxidant defense system, is crucial in disease development. (45) Chemotherapy drugs cause mitochondrial damage. This results in an increase in reactive oxygen species and, as a result, an increase in oxidative stress. (46) This accumulation of ROS would operate as a cell chemoattractant, attracting MSCs and exosomes to the damage site. (47) As a result of the accumulation of ROS at the injury site, MSCs and their exosomes are drawn to the area by this chemoattractant. (34, 48) The levels of antioxidant enzymes such as GSH-PX, MDA, SOD, and CAT were evaluated in male rats to evaluate the therapeutic effects of Exo on testicular toxicity caused by Cis-induced oxidative stress. The Cis-induced group's GSH-PX tissue concentration, SOD, and CAT activities in testicular homogenate were significantly lower than the control group. Cis increased MDA levels, a marker of testicular lipid peroxidation. TNF-α, an inflammatory marker in the testis, was significantly increased. The results confirmed oxidative stress, which is similar to previous research on gonadal damage caused by cisplatin.(2, 3, 12, 27, 28, 36) The exo-treated Cis group's GSH-PX concentration and SOD and CAT activities improved significantly. MDA levels were significantly reduced. Single doz exo caused protection against the up-regulation of testicular TNF-α levels in Cis-inducted rats. However, no significant decrease in TNF-α level was obtained. TNF-α plays an active role as an inflammatory cytokine in Cis-induced testicular damage. (49, 50) An increase in TNF-α levels results in impaired spermatogenesis and decreased testosterone levels. Increased ROS production following damage to mitochondria contributes as well to this process by initially apoptosis. (51) Studies have shown that exosomes are able to penetrate the body's barrier and reach the damaged area. (52) Exosomes with antiapoptotic, anticancer, and antioxidant qualities have been reported to lower oxidative stress in the area of damage. (19, 44, 53, 54)
The primary functions of the testicles are the production of testosterone and sperm. (55) Leydig cells in the interstitial tissue of the testes are responsible for the secretion of testosterone. (56) In this study, the group that received Cis testosterone had lower testosterone levels. The decrease in SF-1 and StAR protein expression supports the decrease in testosterone levels. NHB is a key hormone that regulates reproductive function. The INHB hormone is a key regulator of FSH, which is released by sertoli cells in the testis. (57, 58) FSH stimulates the seminiferous tubules and the Sertoli cells. It is in charge of starting the spermatogenesis process. Testosterone is required for spermatogenesis to continue. (59)
INHB and FSH secretion become negatively correlated during secondary Sertoli cell development and spermatogenesis. Decreased serum INHB indicates sperm damage and infertility. As a result, it is used to measure male spermatogenesis (60, 61). Inhibins also serve as growth factors in Leydig cells via paracrine mechanisms. INHB, in conjunction with testosterone, regulates spermatogenesis indirectly. (62) Previous research has shown that CP intake has a direct effect on the Leydig cells, resulting in cellular death and a decrease in secretory activity. (28, 63) Because of damaged seminiferous tubule epithelium and low testosterone levels, serum INHB levels were considerably lower in the Cis group compared to the control group. INHB levels significantly increased in the exo-treated group after Cis. This recovery process was supported by tissue regeneration in the seminiferous tubule and interstitial areas.