Extracellular vesicles (EVs) mediated intracellular communication plays an imperative role in the proper completion of different physiological events. Most of the bio-fluids are enriched with several subpopulations of EVs including exosomes and microvesicles (MVs), with the capacity of transferring different functional molecules (lipids, proteins, and nucleic acids) to target cells. Recipient cells upon receiving the signal molecules undergo different changes that positively affect the structural and functional integrity of the cells. This article was aimed to highlight the role of EVs secreted by gametes, the female reproductive tract, and the growing conceptus in the successful completion of different reproductive events related to gestation. EVs associated with the reproductive system are actively involved in the regulation of different physiological events including gamete maturation, fertilization, and embryo and fetal development. In the reproductive system, EVs mediated intracellular communication is not unidirectional but is rather regulated through crosstalk between the reproductive tract and the growing conceptus. These vesicles are secreted from the ovary, oviductal epithelium, endometrium, developing embryo, and the placenta. The cargo inside these vesicles exerts pleiotropic effects on both maternal and embryonic environments. A better understanding of the EVs-mediated crosstalk will be helpful in the development of useful tools serving both the diagnostic as well as therapeutic needs related to female fertility.

This study investigated the effects of conditioned medium (CM) from canine amniotic membrane-derived MSCs (cAMSCs) on dog sperm cryopreservation. For this purpose, flow cytometry analysis was performed to characterize cAMSCs. The CM prepared from cAMSCs was subjected to proteomic analysis for the identification of proteins present in the medium. Sperm samples were treated with freezing medium supplemented with 0%, 5%, 10%, and 15% of the CM, and kinetic parameters were evaluated after 4–6 h of chilling at 4 °C to select the best concentration before proceeding to cryopreservation. Quality-related parameters of frozen–thawed sperm were investigated, including motility; kinetic parameters; viability; integrity of the plasma membrane, chromatin, and acrosome; and mitochondrial activity. The results showed that 10% of the CM significantly enhanced motility, viability, mitochondrial activity, and membrane integrity (p < 0.05); however, the analysis of chromatin and acrosome integrity showed no significant differences between the treatment and control groups. Therefore, we concluded that the addition of 10% CM derived from cAMSC in the freezing medium protected dog sperm during the cryopreservation process.

Canine malignant mammary gland tumors present with a poor prognosis due to metastasis to other organs, such as lung and lymph node metastases. Unlike in human studies where obesity has been shown to increase the risk of breast cancer, this has not been well studied in veterinary science. In our preliminary study, we discovered that leptin downregulated cathepsin A, which is responsible for lysosomal-associated membrane protein 2a (LAMP2a) degradation. LAMP2a is a rate-limiting factor in chaperone-mediated autophagy and is highly active in malignant cancers. Therefore, in this study, alterations in metastatic capacity through cathepsin A by leptin, which are secreted at high levels in the blood of obese patients, were investigated. We used a canine inflammatory mammary gland adenocarcinoma (CHMp) cell line cultured with RPMI-1640 and 10% fetal bovine serum. The samples were then subjected to real-time polymerase chain reaction, Western blot, immunocytochemistry, and lysosome isolation to investigate and visualize the metastasis and chaperone-mediated autophagy-related proteins. Results showed that leptin downregulated cathepsin A expression at both transcript and protein levels, whereas LAMP2a, the rate-limiting factor of chaperone-mediated autophagy, was upregulated by inhibition of LAMP2a degradation. Furthermore, leptin promoted LAMP2a multimerization through the lysosomal mTORC2 (mTOR complex 2)/PH domain and leucine rich repeat protein phosphatase 1 (PHLPP1)/AKT1 (Serine/threonine-protein kinase 1) pathway. These findings suggest that targeting leptin receptors can alleviate mammary gland cancer cell metastasis in dogs.

Sperm cryopreservation is a widely used assisted reproductive technology for canine species. The long-term storage of dog sperm is effective for the breeding of dogs living far apart, scheduling the time of artificial insemination that suits the female, and preventing diseases of the reproductive tract. However, spermatozoa functions are impaired during the freeze–thaw processes, which may decrease reproductive performance. Numerous attempts have been made to restore such impairments, including the use of cryoprotectants to prevent the damage caused by ice crystal formation, and supplementation of antioxidants to reduce reactive oxygen species generation due to osmotic stress during the procedure. Egg yolk derivatives, antioxidants, and, more recently, mesenchymal stem cells (MSCs) and their derivatives have been proposed in this research field. This review article will summarize the current literature available on the topic.

Exosomes have been discovered in 1983 (Pan and Johnstone, 1983) and are defined as 40-100 nm extracellular vesicles originating from the fusion of multi-vesicular bodies, also referred to as late endosomes, with the plasma membrane. They are known to mediate cell-to-cell communication as they can carry proteins, nucleic acids, and lipids, providing through that cargo a paracrine effect to the recipient tissues or cells (Mitchell et al., 2019). Thus, stem cell-derived exosomes have been used as a shuttle in cell-free therapies for the treatment of different conditions such as kidney diseases (Dorronsoro and Robbins, 2013) or osteoarthritis (Zhang et al., 2019). Among stem cell types, perinatal tissue-derived stem cells are a good alternative to embryonic stem cells as they are obtained without major ethical issues since they come from discarded fetal annexa, but the use of their secretome is still not common as there are over 30 clinical trials using perinatal tissue-derived stem cells but less than 10 trials using their derived exosomes (https://clinicaltrials.gov/). Among the perinatal tissues, the amniotic membrane is a good source of mesenchymal stem cells (Alviano et al., 2007) but the use of these stem cells or their derived exosomes is not as spread out as the other stem cells types, although they have the same properties and their secretome is as rich in growth and anti-apoptotic factors (An et al., 2017).

The oral cavity is the second-largest habitat for microorganisms, and a well-balanced oral microbiome contributes to preventing dental disorders caused by pathogenic bacteria. Since humans and dogs have different lifestyles and oral microbiome structures, the present study aimed to develop novel probiotics for dogs. A total 53 Lactobacillus spp. were isolated from healthy dogs, and nine isolates were identified as Lactobacillus acidophilus according to 16S rRNA gene sequencing. According to the high antimicrobial activity against the dental caries-causing bacterium Streptococcus mutans, single or three mixed strains were orally administered to dogs for 4 weeks with concentration of 108-109 CFU/day. Intraoral swab samples were collected before and after the administration, and changes of oral pathogen were analyzed using quantitative PCR. Among them, Porphyromonas gingivalis, a critical factor of periodontitis, was significantly reduced in the single-strain administered group. Based on the acid and bile salts tolerance characteristics of isolates, systemic effects were also analyzed by comparing serum immunoglobulin and reproductive ability before and after the administration. However, no significant changes were observed in the serum IgG level and sperm quality. Overall, these in vitro and in vivo results suggest that L. acidophilus isolates from dogs, especially L. acidophilus MJCD175, could be promising probiotic candidates to support oral health without systemic adverse effects in dogs.

Although several methods have been developed to improve male fertility and sperm quality, subfertility remains a primary clinical issue in male reproduction worldwide. The aim of this study was to determine the effects of the oral administration of three commensal Lactobacillus spp. on healthy normozoospermic dogs and the qualitative parameters of their sperm. Three weeks of supplementation induced a significant decrease of two phyla, Proteobacteria and Tenericutes, and an increase of phylum Firmicutes. At the species level, the number of Fusobacterium perfoetens and Anaerobiospirillum succiniciproducens decreased, while Limosilactobacillus reuteri increased. Parallel to these results, qualitative sperm parameters such as total and progressive motility, acrosome integrity, and other kinematic parameters were significantly enhanced after commensal lactobacilli supplementation. In addition, we showed that Firmicutes were positively correlated with sperm qualitative parameters, while Proteobacteria, F. perfoetens, and A. succiniciproducens were negatively correlated. Considering the similarities between the gut microbiome of dogs and humans, these results provide more insight into how gut microbiota regulation could improve male sperm quality in both species.

No abstract