Proteomic Analysis of the Secretome and Exosomes of Feline Adipose-Derived Mesenchymal Stem Cells

Abstract

Mesenchymal stem cells (MSCs) have been shown to have therapeutic efficacy in different complex pathologies in feline species. This effect is attributed to the secretion of a wide variety of bioactive molecules and extracellular vesicles, such as exosomes, with significant paracrine activity, encompassed under the concept of the secretome. However, at present, the exosomes from feline MSCs have not yet been studied in detail. The objective of this study is to analyze and compare the protein profiles of the secretome as a whole and its exosomal fraction from feline adipose-derived MSCs (fAd-MSCs). For this, Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and Protein–Protein Interaction Networks Functional Enrichment Analysis (STRING) were utilized. A total of 239 proteins were identified in the secretome, and 228 proteins specific to exosomes were identified, with a total of 133 common proteins. The proteins identified in the secretome were located in the extracellular regions and in the cytoplasm, while the exosomal proteins were located mainly in the membrane, cytoplasm and cytosol. Regarding function, in the secretome, proteins involved in different metabolic pathways, in pathways related to the immune system and the endocrine system and in the processing of proteins in the endoplasmic reticulum predominated. In contrast, proteins specific to exosomes were predominantly associated with endocytosis, cell junctions, platelet activation and other cell signaling pathways. The possible future use of the secretome, or some of its components, such as exosomes, would provide a non-cell-based therapeutic strategy for the treatment of different diseases that would avoid the drawbacks of cell therapy.

Mesenchymal Stem Cell Exosomes Derived from Feline Adipose Tissue Enhance the Effects of Anti-Inflammation Compared to Fibroblasts-Derived Exosomes

“…The chemokine RANTES participates in the regulation of perivascular inflammation, vascular malfunction, and T-cell accumulation in cardiovascular disease [ 53 ]. Recently, exosomes and secretomes were separated from feline AD-MSCs, and proteomics KEGG and GO analyses were conducted to address the possibility of using exosomes as therapeutic substances [ 54 ]. Several studies reported the analysis and characterization of exosomes derived from canine-species MSCs and presented differential expression of cytokines and growth factors in adipose tissue- and bone-marrow-derived MSC exosomes [ 55 ].…”

Altered Proteomic Profile of Adipose Tissue-Derived Mesenchymal Stem Cell Exosomes from Cats with Severe Chronic Gingivostomatitis

“…Further, fAd-MSCs of each donor were isolated and characterized according to a protocol previously optimized within our group [ 15 , 29 ]. Briefly, adipose tissue was treated with type I collagenase (Merck, Darmstadt, Germany) and the isolated fractions were seeded in T-175 culture flasks containing DMEM supplemented with 10% fetal bovine serum (FBS), 2.5 mM L-glutamine, 100 U/mL penicillin, 100 µg/mL streptomycin, and 1.25 µg/mL fungizone (Merck, Darmstadt, Germany).…”

“…The exosomes from each individual donor were processed separately. The isolation of the exosomes was conducted by ultracentrifugation and their characterization afterwards was conducted following a protocol previously used by our group [ 13 , 15 ]. Briefly, the cells of each donor were seeded at a density of 5.7 × 103 cells/cm 2 in three T-175 flasks in DMEM supplemented with 10% fetal bovine serum (FBS), 2.5 mM L-glutamine, 100 U/mL penicillin, and 100 µg/mL streptomycin (Merck, Darmstadt, Germany).…”

“…The exosome cargo is involved in communication between cell lineages and has a pleiotropic effect [ 13 , 14 ]. Our group was the first to describe the exosomes derived from feline adipose tissue-derived MSCs (fAd-MSC), demonstrating that they play a role in several biological functions [ 15 ].…”

Secretory Profile of Adipose-Tissue-Derived Mesenchymal Stem Cells from Cats with Calicivirus-Positive Severe Chronic Gingivostomatitis

“…However, it is currently assumed that the mechanism of action of the MSCs to maintain homeostasis and to modulate the responses of the immune system is mainly paracrine and relies on the production and release of a great variety of biomolecules with different biological functions. Altogether, these components secreted by the cells, which include growth factors, cytokines, and extracellular vesicles, among others, are known as the secretome [11][12][13]. In some human pathologies, such as lupus, type 2 diabetes, or metabolic syndrome, it has been shown that the MSCs of the patients are dysfunctional and may participate in the etiopathogenesis of the diseases [14][15][16][17].…”