Review articleAnalysis of MSCs' secretome and EVs cargo: Evaluation of functions and applications
Graphical abstract
Introduction
Mesenchymal stem cells (MSCs) are multipotent and undifferentiated cells with the ability to differentiate into several mesenchymal lineages such as osteoblasts, chondrocytes, adipocytes, myocytes [1], [2] and also neuron-like [3]. Although it is known that MSCs are responsible of several cell functions, such as pro-angiogenesis [4], anti-apoptosis [5], neuro-protection and regulation [6], anti-inflammation [7], immunomodulation [8] and others, several studies showed that these functions are mainly exerted by MSCs secreted paracrine factors [9], [10], [11], [12], [13].
In fact, cells need to communicate between each other and to exchange information not only through a direct cell-cell interaction but also through an indirect way as the endocrine, autocrine and paracrine signalling [14]. In particular, paracrine signalling can occur through a set of soluble factors and/or extracellular vesicles (EVs) secreted by the cells in the culture medium, called ‘conditioned medium’. The set of secreted factors with EVs is called ‘secretome’.
Amongst the main components of the soluble fraction, there are proteins, cytokines, chemokines and growth factors [15], whilst EVs are distinguished into exosomes, microvesicles (MVs) and apoptotic bodies, according to their size and biogenesis. About that, exosomes are 30–150 nm in size and derive by multivesicular bodies generated by the early endosomes, whilst microvesicles are 100–1000 nm in size and derive by plasma membrane shedding. Both of them are enriched in small and long non-coding RNAs, mRNAs, lipid and proteins which exert specific functions in the recipient cells [16], [17]. Finally, apoptotic bodies are 1–3 μm in size and derive from apoptotic cells that disassemble themselves and close their content into vesicles, protecting the neighbouring cells. They contain cytosolic fragments and proteins as well as nuclear fragments, such as DNA and histones, or even organelles [18], [19].
It has been proved that the secretome has a higher safety profile if compared to cell engrafting due to its reduced possibility of neoplastic transformation [20] and a better ease of storage with the use of natural and non-toxic agents such as the trehalose, a natural disaccharide found in many foods [21]. Moreover, in the last few years, the analysis of factors contained in the secretome and/or extracellular vesicles made possible for researchers to engineer EVs for a controlled drug delivery [22], [23], [24], [25] or also to defeat viruses [26] but also to evaluate the risks and/or benefits of exposure to certain substances (e.g., chemical agents, environmental pollutants, etc.) [27], [28].
This review aims to summarize current literature on secretome derived from different MSCs with the purpose of creating a compendium of the most recently discovered proteins and non-coding RNAs as components of the soluble fraction or EVs.
Starting from these goals, the applications of the secretome from MSCs from different sources, analyzed in its totality or in its specific components, have been characterized in a broad range of physiological and pathological conditions both in in vitro and in vivo studies.
Section snippets
Angiogenesis
Tissue and organ regeneration is nowadays a field of interest for many worldwide researchers. Although cell applications, together or not with a scaffold, were interestingly investigated [29], [30], [31], [32], [33], recently the attention was focused on the cells released paracrine factors and their regenerative properties. For example, studies concerning the secretome characterization from bone marrow MSCs (BM-MSCs) showed that the highest percentage of secreted proteins were involved in cell
Cancer
Cancer diseases are nowadays amongst the most diffused and lethal diseases in the world. Moreover, several studies predicted an increase in the number of affected people by 2040 [59], [60]; thus, it can be important to identify new tumor markers as well as to characterize cancer cells secretome to provide a tool to target specific cancer pathways. In this regard, several studies analyzed the secretome of cancer cells to find some specific factors that could be considered as biomarkers of
Cardiovascular diseases
Cardiovascular diseases (CVDs) are heart and circulatory system disorders with an extremely high incidence. CVDs are classified as chronic diseases and evolve gradually over a lifetime, generating numerous symptoms in the patients. They can also lead to sudden death. EVs seem to be excellent candidates for the treatment of myocardial damage, as shown by some recent studies. The treatment with MSC-EVs by hypoxic cardiomyocytes caused a marked improvement in cell invasion, proliferation and
Autoimmune diseases
Autoimmune diseases are characterized by a misreaction of the immune system, which attacks and destroys healthy tissues in the body by mistakenly recognizing them as foreign. These disorders are multifaceted and the causes are not entirely clear but several external factors seem to play a role. Recently, it was discovered that EVs derived from mesenchymal stem cells play a role in the progression or otherwise of the disease. In this regard, over-expressing programmed cell death-ligand
Characterization of secretome from fetal MSCs
Several studies have already proved that there are important differences amongst adult and fetal MSCs. Indeed, fetal MSCs showed better abilities as immune-modulators [79] and are more proliferative with longer telomeres and less predestined, with an higher expression of the pluripotency markers than adult ones [80], [81]. Moreover, they showed enhanced neuroprotective abilities also in in vivo models [82].
Due to the differences in the cell properties at different stages of life, it was easy to
Analysis of secretome from 3D cultured MSCs
Several scientific studies proved that cells cultured under 3D conditions were able to better mimic the properties of their native tissues [87], [88] both in oncological and regenerative studies. In this last field, until ten years ago, mesenchymal stem cells were only cultured in 2D adhesion conditions. Recently, several studies were performed on 3D cultures, demonstrating their effective superiority. Although to date there is still no standardized technique to obtain 3D spheroids, many
Secretome and epigenetic modifications
In recent years, science has focused much of its interest on studying the environmental effects on cells. The outdoor and indoor air pollution is amongst the increasingly significant issues, resulting in the death of more than 6 millions of people [100]. Moreover, the exposure to chemical compounds (for example phthalates and phenols, perfluorinated chemicals, ethyl paraben) as well as ionizing radiations, metals or cigarette smoke, lead cells to side effects [101]. It was already known that
Conclusion
Mesenchymal stem cells are multipotent cells with different functions [1], [2], [3]. Recently, several studies attributed these abilities to their secreted factors that act in a paracrine manner on the neighbouring cells [9], [10], [11], [12], [13]. In fact, paracrine signalling happens through the so-called “secretome”, formed by soluble factors, released by the cells in the culture medium, and extracellular vesicles (EVs), which are vehicles of small and long non-coding RNAs, proteins and
Funding
This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.
Data availability
No data was used for the research described in the article.
CRediT authorship contribution statement
Valentina Urrata: Conceptualization, Writing – original draft, Writing – review & editing. Marco Trapani: Writing – original draft. Mara Franza: Writing – original draft. Francesco Moschella: Supervision, Writing – review & editing. Anna Barbara Di Stefano: Conceptualization, Supervision, Writing – original draft, Writing – review & editing. Francesca Toia: Supervision, Writing – review & editing.
Declaration of competing interest
The authors declare that there are no conflicts of interest.
Acknowledgements
Valentina Urrata, PhDst is supported, for this research, by the University of Palermo (IT), Doctoral Course of Experimental Oncology and Surgery, Cycle XXXVI.
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Equal contribution, joint co-last authors.