In vitro aging alters the gene expression and secretome composition of canine adipose-derived mesenchymal stem cells.

Introduction: Canine adipose-derived mesenchymal stem cells (cAD-MSCs) hold therapeutic promise due to their regenerative potential, particularly within their secretome. However, concerns arise regarding the impact of in vitro cultivation necessitated for storing therapeutic doses, prompting this study to comprehensively explore the impact of in vitro aging on gene expression and secretome composition. Methods: The study involved collecting abdominal adipose tissue samples from nine healthy female dogs, from which cAD-MSCs were extracted and cultured. Stem cells were validated through trilineage differentiation assays and flow cytometry immunophenotyping. Gene expression profiling using RT-qPCR array, and cAD-MSCs secretome LC-MS/MS analysis, were conducted at passages 3 and 6 to reveal gene expression and protein composition alterations during in vitro culture. Results and Discussion: The results demonstrate that the gene expression and secretome composition of cAD-MSCs were impacted by in vitro aging. Among many alterations in gene expression between two passages, two significant downregulations were noted in the MSC-associated PTPRC and IL10 genes. While the majority of proteins and their functional characteristics were shared between passages, the influence of cell aging on secretome composition is highlighted by 10% of proteins being distinctively expressed in each passage, along with 21 significant up- and downregulations. The functional attributes of proteins detected in passage 3 demonstrated a greater inclination towards supporting the regenerative capacity of cAD-MSCs. Moreover, proteins in passage 6 exhibited a noteworthy correlation with the blood coagulation pathway, suggesting an elevated likelihood of coagulation events. To the best of our knowledge, this study presents the first original perspective on the changes in secretome composition that occur when cAD-MSCs age in vitro. Furthermore, it contributes to broadening the currently restricted knowledge base concerning the secretome of cAD-MSCs. In conclusion, our findings show that the regenerative potential of cAD-MSCs, as well as their secretome, may be compromised by in vitro aging. Therefore, our study suggests a preference for earlier passages when considering these cells for therapeutic applications.

Cancer morbidity in Croatian cats: Retrospective study on spontaneously arising tumors (2009-2019).

One of the current leading causes of death in pet cats is neoplasia. The scarcity of data on tumor prevalence in cats from south-eastern Europe led to this retrospective study which reports an abundant collection of tumors diagnosed in cats living in Croatia. Archived histopathology findings (diagnosis, cell origin, and biological behavior), and epidemiological data (breed, sex, age, and anatomical site of the tumor) of the cats diagnosed with at least one tumor were collected and statistically analyzed. For the research, 2338 archived feline samples, obtained from 2009-2019, have been reviewed. In 659 samples (28.1%) one or more tumors were diagnosed. Tumors have most often affected non-pedigree cats, cats between 7 and 15 years of age, and female cats. Most tumors showed malignant behavior (85.7%). The most frequent diagnoses were mammary adenocarcinoma, lymphoma, and squamous cell carcinoma. Regarding origin, most samples were epithelial (46.9%), but mesenchymal (28.7%) and lymphoid tumors (17.0%) were also frequently found. Tumors most often occurred in the skin and subcutis, mammary gland (in females), digestive system, and hemolymphatic system (in males). Statistical analysis did not reveal breed, sex, and age predisposition, but female animals and non-pedigree cats were significantly more likely to develop malignant tumors. This detailed study gives insight into the most frequent tumors that occur in cats from Croatia, and the data reported here is likely applicable to other countries. This data will contribute to clinical veterinary oncology, as it presents the most extensive tumor index of cats in south-eastern Europe.

The impact and complete genome characterisation of viruses involved in outbreaks of gastroenteritis in a farrow-to-finish holding.

Viral enteric pathogens continuously burden intensive pig farming, causing gastrointestinal diseases of epidemic and endemic nature. The present study investigated two diarrhoea outbreaks on a large farrow-to-finish holding and subsequent circulation of outbreak-related enteric viruses. These viruses were characterised by whole genome sequencing, and statistical evaluation of the impact on specific production metrics was performed. The results provided evidence that the Porcine epidemic diarrhoea virus-swine enteric coronavirus (PEDV-SeCoV) S gene recombinant strain was responsible for the first outbreak, whilst Rotavirus A (RVA) in a mixed infection with Rotavirus B (RVB) and porcine kobuvirus (PKV) probably caused the second diarrhoea outbreak. Whole genome characterisation revealed a porcine origin of all viruses involved and significant heterogeneity of RVB strain, proposing four novel genotypes and changes in RVB VP1 genotype classification. The statistical evaluation confirmed only a minor disturbance in the number of weaned pigs per sow, with statistical forecasting showing positive trends. A follow-up study corroborated the endemicity of RVA and PKV, in contrast to PEDV-SeCoV. Punctual, comprehensive and timely investigation of diarrhoea outbreaks is a prerequisite for applying adequate pig health and biosecurity management. Calculating such outbreaks' impact on production metrics can potentially shape future decisions on management improvements.

An Outstanding Role of Adipose Tissue in Canine Stem Cell Therapy.

Adipose tissue, previously known as connective tissue with a role in energy storage, is currently changing the course of treatments in veterinary medicine. Recent studies have revealed one particularly impressive function among all the newly discovered functions of adipose tissue. The interactive cells hosted by adipose tissue, the stromal vascular fraction (SVF), and their role in treating numerous diseases have provided a prospective course of research with positive outcomes in regenerative veterinary medicine (RVM). This review describes the main features of adipose tissue, emphasizing an eclectic combination of cells within the SVF and its thus far researched therapeutic possibilities in canine RVM. An afterwards focus is on a highly researched component of the SVF, adipose-derived mesenchymal stem cells (ASCs), which were shown to have an extraordinary impact relying on several proposed mechanisms of action on mitigating pathologies in canines. Furthermore, ASC therapy showed the most significant results in the orthopaedics field and in neurology, dermatology, ophthalmology, gastroenterology, and hepatology, which elevates the possibilities of ASC therapy to a whole new level. Therefore, this review article aims to raise awareness of the importance of research on cellular components, within abundant and easily accessible adipose tissue, in the direction of regenerative therapy in canines, considering the positive outcomes so far. Although the focus is on the positive aspects of cellular therapy in canines, the researchers should not forget the importance of identifying the potential negative aspects within published and upcoming research. Safe and standardized treatment represents a fundamental prerequisite for positively impacting the lives of canine patients.

The Expression Pattern of Surface Markers in Canine Adipose-Derived Mesenchymal Stem Cells.

The influence of cultivation on the expression pattern of canine adipose-derived mesenchymal stem cells (cAD-MSCs) surface markers, contributing to, among others, the promotion of growth, proliferation, differentiation and immunomodulatory mechanisms of an excellent therapeutic, is still unknown. To fill the gap, we investigated CD90, CD44, CD73, CD29, CD271, CD105, CD45 and CD14 patterns of expression at the protein level with flow cytometry and mRNA level using a real-time polymerase chain reaction array. Gentle variations of expression occurred during cultivation, along with increased CD90, CD44 and CD29 expression, low and decreasing CD271 and CD73 expression and a decrease of initially high CD105. As expected, CD45 and CD14 were not expressed by cAD-MSCs. Interestingly, we discovered a significant decrease of CD73 expression, compared to early (P1-P3) to late (P4-P6) passages, although the CD73 gene expression was found to be stable. The percentage of positive cells was found to be higher for all positive markers up to P4. As CD73's one important feature is a modulation from a pro-inflammatory environment to an anti-inflammatory milieu, the expression of CD73 in our conditions indicate the need to consider the time cells spend in vitro before being transplanted into patients, since it could impact their favourable therapeutical properties.