Abstract
Simple SummaryIn this study, we have tried to exploit the potential role of exosomes-derived from canine adipose-derived mesenchymal stem cells in the protection and repair of damaged sperm during cryopreservation. The treatment of sperm with an optimal level of exosomes resulted in improved post-thaw semen quality. This improvement was due to the exosomal proteins that protect the sperm against oxidative damage and induce the repair of membranes and chromatin. The effect of exosomal proteins was also confirmed by higher expression of genes related to the repair of membranes and chromatin accompanied by the lower expression of a gene associated with reactive oxygen species production.Freezing decreases sperm quality, ultimately affecting fertilizing ability. The repair of freeze-damaged sperm is considered crucial for improving post-thaw viability and fertility. We investigated the effects of exosomes derived from canine adipose-derived mesenchymal stem cells on dog sperm structure and function during cryopreservation. The pooled ejaculate was diluted with buffer, without (Control), or with exosomal proteins (25, 50, or 100 µg/mL). Using fresh semen, the determined optimal exosomal protein concentration was 50 µg/mL (Group 2) which was used in further experiments. Post-thaw sperm treated with exosomes were superior to control (p < 0.05) in terms of motility (56.8 ± 0.3% vs. 47.2 ± 0.3%), live sperm percentage (55.9 ± 0.4% vs. 45.4 ± 0.4%), membrane integrity (55.6 ± 0.5% vs. 47.8 ± 0.3%), and acrosome integrity (60.4 ± 1.1% vs. 48.6 ± 0.4%). Moreover, expression of genes related to the repair of the plasma membrane (ANX 1, FN 1, and DYSF), and chromatin material (H3, and HMGB 1) was statistically higher in exosome-treated sperm than control, but the expression of the mitochondrial reactive oxygen species modulator 1 gene was significantly higher in control. Therefore, exosomal treatment may improve the quality of post-thaw dog semen through initiating damaged sperm repair and decreasing reactive oxygen species production.
Highlights
The freezing of mammalian sperm is considered imperative for the preservation of genetic material from individuals with superior breeding value or threatened with extinction from some disease or natural disaster
Sperm treated with 50 μg/mL exosomal protein exhibited higher motility (77.8 ± 0.2%) than untreated controls and those treated with the other concentrations
Treatment with exosomal proteins maintains the integrity of the plasma membrane as well as the chromatin material of canine sperm during cryopreservation
Summary
The freezing of mammalian sperm is considered imperative for the preservation of genetic material from individuals with superior breeding value or threatened with extinction from some disease or natural disaster. The use of cryopreserved semen reduces the problems concomitant with natural breeding, animal. The freezing process can exert certain detrimental changes in the morphology of sperm, resulting from thermal, mechanical, chemical, osmotic, and oxidative damage [5,6]. These changes cause lower post-thaw sperm motility, decrease the integrity of the plasma and acrosomal membrane [7], and damage DNA, decreasing the fertilizing capability of sperm [8]. The factors responsible for reduced fertility of post-thaw sperm include ice formation, high osmotic pressure [9], reactive oxygen species (ROS) generation [6,10,11], and apoptotic pathway activation [12]
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