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Mesenchymal stem cells (MSCs) are currently available for a range of applications and have become a good material for regenerative medicine, tissue engineering, and disease therapy. MSCs are self-renewing, multipotent progenitor cells with multilineage potential to differentiate into cell types of mesodermal origin, such as adipocytes, osteocytes, and chondrocytes, and exert potent immunosuppressive potentials. In the present review, we highlight the currently reported variations in the differentiation potential of MSCs from different tissue sources, the minimal criteria to define MSCs from various tissue environments, and provide a detailed description of MSCs surface markers. Furthermore, MSCs immunomodulatory features secrete cytokines and immune receptors which regulate the microenvironment in the host tissue also revisits in detail. We propose that there are likely more sources of MSCs waiting to be discovered. We need to Standardize MSCs characterization by selecting markers for isolation, cellular and molecular mechanisms involved in MSC-mediated immune modulation, and other functionalities of MSCs should be characterized prior to use in clinical applications.
Stem cells represent a novel cell type in the body. Stem cells have two features: the ability to differentiate along different lineages and the ability to self-renew and maintain tissue homeostasis. Stem cells can be multi-plied in large Stem cells are broadly classified based on their source into embryonic (hESCs) and adult (ASCs) stem cells. Bhartiya, (2013) Embryonic stem cells are pluripotent in nature and can be differentiated into 200 odd cell types where as the adult stem cells are isolated from adult body tissues and are multi-to unipotent in nature (Thomson et al., 1998). MSCs are adult stem cells which can be isolated from human and animal sources with the capacity to differentiate into meso-dermal lineage. MSCs and their multiline age were first found by Friedenstein via studies on the mouse bone marrow in the 1960s (Friedenstein et al., 1987; Frie-denstein et al., 1976; Friedenstein, 1966).
Table 1: Summary of hMSCs tissue sources and In-vitro differentiation potentials.
The multiline age differentiation potential of adult human MSCs from bone marrow was described by Pittenger and group. They have been isolated from almost all tissues including per vascular area (Pittenger et al., 1999). In view of many studies till now, neither single definition nor a quantitative assay to help in the identification of MSCs in mixed population of cells is available; some biomarkers are available for identi-fication of human MSC (hMSC). In 2006 a set of markers and cell characteristics has been proposed by the International Society for Cellular Therapy has proposed minimum criteria to define MSCs as these cells (a) should exhibit plastic adherence; (b) possess specific set of cell surface markers, i.e. cluster of differentiation CD73, D90, CD105 and lack expression of CD14, CD34, CD45 and human leucocyte antigen-DR (HLA-DR); and (c) have the ability to differentiate in-vitro into adipocyte, chondrocyte and osteoblast (Crisan et al., 2008). The availability and culturally expandable in vitro with special genomic stability and less ethical issues, marking these incredible cells importance in cell regenerative therapy and medicine (Dominici et al., 2006; Ullah et al., 2015). The resent study is the con-cise review article to gather available the information about stem cell sources, identification makers, Paracrine secretion, Immunomodulation by MSCs and their importance in regenerative therapies.
Table 2: Minimal criteria of mesenchymal stem cells.
Human Mesenchymal Stem Cell (HMSC) Sources:
The bone marrow (BM) has been the prevailing source of MSCs in humans (Mushahary et al., 2018). How-ever, while BM is a rich source of hematopoietic stem cells, it constitutes only a rare MSC population (Li et al., 2016) BM-derived MSC (bmMSC) supply is the painful harvesting procedure marking their application in research and in the clinical setting limited. Over time, a number of other tissues have been identified as alternative sources for hMSC. Today, MSC can be iso-lated from multiple tissues (Ullah et al., 2015). The human MSC (hMSC) properties can vary greatly depending on multiple parameters including tissue source, isolation method and medium composition and several studies mentioned variations in the differen-tiation potential of MSC from different tissue sources. Table 1 summaries some of the currently used tissue sources and the respective confirmed differentiation potentials.
Table 3: Surface Marker Expression Profiles of Main MSC Types.
Markers for MSC Identification and Verification:
The International Society for Cellular Therapy (ISCT) published the minimal criteria for defining MSCs in 2006. The ISCT proposed positive and negative mar-kers that enabled researchers to distinguish MSCs from other cells in the bone-marrow compartment. The negative markers were selected to include surface anti-gens that are expressed by hematopoietic cells, while the positive markers were selected to include surface antigens that are absent from most hematopoietic cells. It is well established that cultured colonies of MSCs ex-press CD105, CD73, and CD90, but do not express CD45, CD34, CD14 or CD11b, CD19, and HLA-DR (Datta et al., 2011; Karystinou et al., 2009) Table 2. Some labeling strategies have also been used to successfully isolate MSCs enriched for markers such as STRO-1 (Saeedi et al., 2019; Gronthos et al., 2003; Kuroda et al., 2010; Psaltis et al., 2010) CD146 (Bensidhoum et al., 2004; Covas et al., 2008), SSEA-4 (da Silva Meirelles et al., 2015; Gang et al., 2007), CD271 (NGFR) (Battula et al., 2008; Vaculik et al., 2012) antigen 1 (MSCA-1). ISCT acknowledges that the criteria must be met with some flexibility, particularly as they relate to the expression of the negative marker, HLA Class II. Specifically, HLA Class II can be expressed by MSCs under certain conditions, such as cytokine stimulation. Therefore, cells that meet all other criteria, but are also positive for HLA Class II, can be designated as MSCs if an adjective is used to indicate that the cells were stimulated. According to the ISCT, CD34 is a negative marker of MSCs. However, some reports suggest that the CD34 negative status is an artifact of cell culture condition (Pilbauerová et al., 2019). In fact, several groups have shown that MSCs isolated from adipose tissue express CD34 at the time of isolation but lose expression while in culture (Lin et al., 2012; Quirici et al., 2010). Expression of CD34 by MSCs is also supported by the fact that the STRO-1 antibody (Clone STRO-1), which is commonly used to identify MSCs, was developed using CD34+bone marrow as the immunogen (Pachón-Peña et al., 2011). The debate over the use of CD34 as a negative marker raises the possibility that markers may vary depending on the MSC tissue source (Kim & Cho, 2013). For specific immunophenotypic patterns, the variety of tissue sources of peripheral stem cells that could be isolated by their lineage-specific surface markers are summarized in Table 3.
Mesenchymal Stem Cell Markers Co-expression:
Table 4: Anti-inflammatory mechanisms of MSCs.
The purity of MSCs can be increased by using more than one MSC marker for positive selection. As an example, a recent publication demonstrated that selec-tion of mesenchymal stem cells using CD271/NGF R, CD90/Thy1, and CD106/VCAM-1 resulted in a highly clonogenic population of cells (Simmons & Torok-Storb, 1991). Specifically, the addition of CD106 as a marker for positive selection led to isolation of cells with five times greater clonogenic potential compared to the cells isolated with CD271/and CD90 alone. The degree of co expression of surface markers on MSCs also studied (Rasmusson et al., 2003). The cells sub-sets detected for the presence of MSCA-1/TNAP, CD271/NGF R and CD56/NCAM from whole cell population of Human BM-MNCs were analyzed and reported that the CD271 expression detected and CD56 expression not detected cells expressed CD106 and CD146 whereas, CD271 and CD56 presences detected cells exclusively expressed CD166 (Mabuchi et al., 2013) CD271 and CD56 double positivity enriched SSEA-4 expression and MSCA-1 expression. The study conducted by Vaculik et al. (2012), explains the expression pattern of SSEA-4 in dermis was analogous to CD271. CD271 and SSEA-4 both co expressed with CD45 detected cells, in human dermis where as CD73 and CD105 are co expressed. The human dermis minor population of CD73 detected cells are not expressed CD90 (Battula et al., 2008). Dermis CD271 positive cells were also positive for CD73and CD105, whereas the majority of CD271 positive cells are CD90 negative (Battula et al., 2008). Several other studies have been performed recently aimed at achieving high-purity BM MSCs using a combination of CD271 and markers other than CD73, CD105, or CD90 (Pérez-Silos et al., 2016) for example, CD146 has attracted a lot of interest recently, by linking CD146 expression on MSCs with their pericyte topography and function (Battula et al., 2009; Pittenger et al., 1999). It was also reported that CD146 expression on CD271 positive MSCs correlates more with their in situ localization (Sacchetti et al., 2007). Maijenburg further reported that the distribution of CD271 and CD146 and subsets correlates with donor age. The main subset in pediatric and fetal BM was reported to be CD271 expression and CD146 expression, whereas the subset of CD271 expression and CD146 not expression detected population was dominant in adult marrow (Tormin et al., 2011). The endometrial MSC-like cells (eMSCs) can be purified on the basis of their co expression of two per vascular mar-kers, CD140b/platelet-derived growth factor receptor β (PDGFRβ) and CD146 (Maijenburg et al., 2012). The first novel single marker, W5C5 for isolation of endometrial MSC-like cells (eMSCs) (Schwab & Gargett et al., 2007; Masuda et al., 2012)
Immunomodulatory Properties of MSCS:
One of the main advantages of MSCs is their immuno-modulatory properties. MSCs grown in vitro have the ability to interact and regulate the function of the majority of effectors cells involved in the processes of primary and acquired immune response. Due to low expression of MHC I and lack expression of MHC class II along with co-stimulatory molecules, like CD80, CD40 and CD86, MSCs are unable to bring substantial all reactivity and these features protects MSCs from natural killer (NK) cells lysis (Masuda et al., 2012).
Moreover, it is observed that human BM-MSCs were not recognized by NK cells, as they expressed HLA-DR molecules (Rasmusson et al., 2003) MSCs exert immunomodulatory effects by inhibiting the com-plement-mediated effects of peripheral blood mono-nuclear cell proliferation (Spaggiari et al., 2006; Tu et al., 2010) blocking apoptosis of native and activated neutrophils, as well as reducing the number of neutrophils binding to vascular endothelial cells, limiting the mobilization of these cells to the area of damage (Moll et al., 2011; Cassatella et al., 2011). In response to inflammatory molecules such as interleukin-1 (IL-1), IL-2, IL-12, tumor necrosis factor-a (TNF-α) and inter-feron-gamma (INF-ɣ), MSCs secrete an array of growth factors and anti-inflammatory proteins with complex feedback mechanisms among the many types of immune cells summarize in Table 4 (Munir et al., 2015; Shi et al., 2015; Cagliani et al., 2017; Weiss & Dahlke, 2019; Maria et al., 2017; Zhao et al., 2016).
The key immunomodulatory cytokines include prostaglandin 2, TGF-b1, HGF, SDF-1, nitrous oxide, indoleamine 2,3-dioxygenase, IL-4, IL-6, IL-10,IL-1 receptor antagonist and soluble tumor necrosis factor-a receptor (Murphy et al., 2013). MSCs prevent proliferation and function of many inflammatory immune cells, including T cells, natural killer cells, B cells, monocytes, macrophages and dendritic cells. MSCs can block the differentiation of CD34+ cells isolated from the bone marrow or blood monocytes into mature dendritic cells both by direct contact as well as by secreted paracrine factors (Nauta et al., 2006; Jiang et al., 2005). They inhibit the transformation of immature dendritic cells into mature forms and limit the mobilization of dendritic cells to the tissues. (Su et al., 2011) Due to the influence of MSCs, M1 (pro-inflammatory) macrophages are transformed into M2 type cells with an anti-inflammatory phenol-type, and the interleukin (IL)-10 secreted by them inhibits T-cell proliferation (Chen et al., 2014; Gao et al., 2014).
In vitro studies have demonstrated a direct immune-modulatory effect of MSCs on lymphocytes by suppression of activated CD4+ and CD8+ T cells and B-lymphocytes was observed (Sharif et al., 2019; Glennie et al., 2005). In addition, MSCs reduce the level of pro-inflammatory cytokines synthesized by T-lymphocytes, such as tumor necrosis factor (TNF)-α and interferon (IFN)-γ (Yañez et al., 2006) and increase synthesis of anti-inflammatory cytokines, for example, IL-4. MSCs also have the ability to limit the synthesis of immune-globulins like immunoglobulin (Ig) M, IgG, and IgA classes secreted by activated B cells, thereby blocking the differentiation of these cells to plasma cells (Corcione et al., 2006). MHC class I chain-like gene A (MICA) together with TLR3 ligand and other immune-regulatory proteins kept the MSCs safe from NKs invasion (Giuliani et al., 2014). Together with other properties, these immunomodulatory features makes MSCs one of the feasible stem-cells sources for performing cell transplantation experiments.
Furthermore, it is also important to note that MSCs from different sources may differ in their mechanisms and capacities for immune modulation (Mattar & Bieback, 2015). Because of their trophic and immune-modulatory functions, MSCs are generally considered to possess greater advantages in cell-based regenerative medicine, MSCs an important regulator of immune tolerance and attractive therapeutic target for limiting autoimmune inflammation.
Mesenchymal stem cells have been isolated from a wide range of species and tissues using several techniques. MSCs are isolated as a heterogeneous population of cells that differ in growth kinetics and differentiation potentials. A large number of markers have been brought forward to facilitate the isolation of MSCs from their surrounding environment or the selection of MSCs with high stemness. With their ability to differentiate into multiple lineages, secrete factors related to immune regulation, and migrate to-ward sites of inflammation, All these properties of MSCs make them distinct from other stem cells and can be used in future cell replacement therapy and many other clinical implications. In this review, we concisely bring up the current data available for MSCs isolation sources, characterization markers and its immunomodulatory properties. The future MSCs research should focus on finding more suitable markers to isolate the source-specific MSCs, basic understanding of growth regu-lators in differentiation and trans-differentiation and its immunomodulatory properties to modify the host immune environment.
The authors would like to express their gratitude Pakistan Council of Scientific and Industrial Research for their constructive cooperation throughout the research work.
The authors wish to confirm that there are no conflicts of interest associated with this publication and there has been no significant financial support for this work that could have influenced its outcome.
Academic Editor:
Md. Ekhlas Uddin Dipu, Department of Biochemistry and Molecular Biology, Gono Bishwabidalay, Dhaka, Bangladesh.
Pharmaceutical Research Center, Pakistan Council of Scientific and Industrial Research (PCSIR), Laboratories Complex, Karachi-75280, Pakistan.
Abbasi K, Iqbal S, Bano S, Siddiqui K, and Muthiah L. (2021). More to explore; the mesenchymal stem cells (MSCs) major tissue sources, known surface markers and its immunomodulation properties. Am. J. Pure Appl. Sci., 3(4), 85-97. https://doi.org/10.34104/ajpab.021.085097
