Optimizing Mesenchymal Stem Cells for Regenerative Medicine: Influence of Diabetes, Obesity, Autoimmune, and Inflammatory Conditions on Therapeutic Efficacy: A Review

Introduction

Regenerative medicine is a branch of medicine based on the natural ability to repair damaged tissues by regenerating or replacing defective cells [1]. However, the extent of the damage or the type of tissue sometimes prevents effective regeneration [2,3]. In such situations, regenerative medicine uses its 2 main tools: biomaterials and stem cells [1]. Stem cells can accelerate the regeneration of damaged organs, even if this process would be very slow or nearly impossible under natural conditions [4,5].

The most promising type of stem cells for regenerative medicine appears to be mesenchymal stem cells (MSCs). Compared to other types of stem cells, they are present in most human tissues, and taking samples is not associated with any ethical concerns. Moreover, they retain high differentiation capacity with minimal risk of carcinogenesis [6]. In addition, MSCs operate through paracrine signaling, which is nonsynaptic release of small molecules, including cytokines and growth factors.

Their beneficial effects have already been proven; among other things, in the regeneration of bones, nervous system, cardiac muscle, liver, cornea, trachea, and skin [7], and they have also been shown to have anti-inflammatory effects in the treatment of autoimmune diseases [8–10] and sepsis [11,12].

MSCs have an enormous therapeutic potential. Currently, many clinical trials are being conducted on MSCs therapy [13], but the properties of these cells have not yet been fully explored. There are many factors that change the properties of MSCs – diseases, individual laboratory parameters, their source, and methods of harvesting and banking of cells [14,15], which make the use of MSCs very complex.

This article aims to review the characteristics, harvesting, banking, and paracrine signaling of MSCs and their role in diabetes, obesity, and autoimmune and inflammatory diseases, and potential role in regenerative medicine.

Characterization of Human Mesenchymal Stem Cells (MSCs)

MSCs are stem cells derived from the mesoderm. The criteria for recognizing MSCs are the fibroblastic shape of the cells, adhesion to a plastic medium, the presence of surface antigens CD73, CD90 and CD105 in the absence of CD34 and CD45, although some authors indicate the possibility of the presence of the CD34 antigen on the surface of MSCs [16,17].

MSCs also have the ability to differentiate into osteocytes, chondrocytes, and adipocytes [17]. MSCs have a great therapeutic potential due to their broad differentiation capabilities. They are classified as multipotent cells, although it has been observed that they may also exhibit pluripotent properties [18]. This makes it possible to grow most types of cells in the laboratory, which, when injected, can take over the function of degenerated cells [19–22].

Methods of Harvesting MSCs

MSCs are readily available because they are present in most human tissues. They are present in mature tissues such as bone marrow, adipose tissue, skin, peripheral blood, menstrual blood, muscle, and lung tissue, as well as in perinatal tissues, including umbilical cord, cord blood, and placenta. Although belonging to the same type, these cells vary in their potential to differentiate. Therefore, when selecting cells for therapy, it is crucial to consider not only the invasiveness of the collection procedure but also the variability of their properties [23].

After tissue collection, MSCs can be obtained through enzymatic methods or explant culture. In the explant culture method, the tissue source of the MSCs is washed and mechanically fragmented. The fragments obtained are then transferred to culture dishes containing the appropriate medium. In the enzymatic method, enzymatic digestion is performed to break down the tissue surrounding the MSCs [23].

The enzymatic method allows for the culture of isolated MSCs, ensuring they are not influenced by growth factors from the surrounding tissue. However, in this case, the cells may experience stress due to the action of digestive enzymes. In contrast, the explantation procedure offers a more stable environment, which enhances the migration of MSCs, but the surrounding tissues can influence the development of cultured cells [23,24]. Thus, the 2 methods yield different effects on the properties of MSCs, a factor that should be considered when choosing a procedure.

MSCs are cultured using DMEM medium and fetal bovine serum (FBS) at 37°C, with 15% O2 concentration, 5% CO2 concentration, and 95% humidity [18]. The culture conditions themselves can also modify the properties of MSCs. For example, it has been shown that hypoxic conditions, as well as the use of activated platelet-rich plasma donors instead of FBS, can be better for MSC culture [25,26].

Banking of MSCs

Once isolated, MSCs can be directly used for therapy or banked. Storage of these cells is carried out at −120°C [25] with the addition of dimethyl sulfoxide (DMSO) to protect the cells from damage caused by the freezing process [27]. Banking MSCs only makes sense if it does not alter their properties. After thawing, MSCs should be as therapeutically effective as they were before freezing.

It has been shown that the process of freezing and thawing does not affect the morphology, expression of surface markers, or ability of MSCs to adhere to plastic substrates. These cells, after thawing, still meet the criteria for MSCs [28,29]. Results comparing fresh MSCs (F-MSCs) and thawed MSCs (T-MSCs) in terms of viability and differentiation ability are inconclusive [27,28]. However, it has been observed that T-MSCs can exhibit a reduced immunosuppressive potential [28]. On the other hand, both groups of cells – F-MSCs and T-MSCs – showed similar efficacy in the treatment of arthritis in rats, hind-limb ischemia in mice, and lung injury in rats [27]. Therefore, freezing MSCs can affect the cells, but likely does not impact their clinical utility.

Paracrine Signaling via Cytokines, Growth Factors, and Small Molecules

Undifferentiated MSCs can support the regeneration of surrounding tissues through secreted substances and exosomes. This is possible through the production of growth factors such as vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), insulin-like growth factor 1 (IGF1), fibroblast growth factor (FGF), placental growth factor (PIGF), nerve growth factor (NGF), glial cell derived neurotrophic factor (GDNF), epidermal growth factor (EGF), and transforming growth factor beta 1 (TGFβ). These factors stimulate growth of neighboring cells, have anti-apoptotic properties, and support angiogenesis [17].

MSCs exhibit immunomodulating effects. They secrete a number of substances such as cytokines, enzymes, and other proteins with anti-inflammatory properties [30,31]. In addition, they inhibit the proliferation of T lymphocytes and NK cells [17,32]. MSCs exhibit low expression of MHC class I and II antigens, and thus do not trigger an immune response after transplanting. This allows the use of allogeneic transplants [33,34].

The amount of secreted factors can vary depending on the health of the donor. This will affect the therapeutic properties of MSCs, so the awareness of how a given disease affects MSCs is crucial when choosing a cell source. This is especially important in autologous transplants, where the patient’s disease may be the reason for low therapy effectiveness [35]. In such a case, cells from an unrelated donor could prove more effective.

MSCs and Diabetes

Diabetes is a metabolic disease associated with elevated blood glucose levels. It is a disease that affects the entire body. It can cause complications in various organs and even entire systems [36]. Therefore, it also affects the properties of single cell types, such as MSCs.

MSCs derived from adipose tissue of patients with diabetic kidney disease do not show morphological differences compared to MSCs obtained from healthy people (H-MSCs). They have a fibroblastic shape, adhere to a plastic medium, and express the typical MSC antigens CD73, CD90, and CD105, with no expression of CD34 and CD45. These cells also retain the ability to differentiate into adipose, cartilage, and bone tissue cells [37].

The criteria for MSCs recognition are also met in MSCs from bone marrow obtained from people with type 1 diabetes [38]. Therefore, it can be concluded that diabetes in donors does not affect the morphology of MSCs. Surface antigens CD73, CD90, and CD105 are also present, but studies have shown that their number is reduced in type 2 diabetes [39].

It has been reported, however, that diabetes affects the paracrine signaling of MSCs. Diabetic kidney disease influences the anti-inflammatory properties of MSCs. It was observed that cells obtained from donors with diabetes show higher secretion of anti-inflammatory factors indoleamine 2,3-dioxygenase 1 (IDO) and prostaglandin E2 (PGE2) and lower secretion of IL6 compared to H-MSCs [37].

Co-occurrence of diabetes affects the formation of new blood vessels by MSCs. MSCs obtained from people with type 1 diabetes show lower HGF expression compared to H-MSCs [38]. MSCs obtained from rats with induced diabetes showed a significant reduction in Krueppel-like factor 2 (KLF2), which impairs the pro-angiogenic properties of these cells [40]. Other studies showed that transplantation of MSCs obtained from mice with type 2 diabetes improves blood supply to the limbs, but to a lesser extent than MSCs obtained from healthy mice [41].

Diabetes also influences the direction of MSCs differentiation. It was demonstrated that cells obtained from mice with type 2 diabetes are more likely to transform into adipose tissue cells compared to H-MSCs, which limits their angiogenic potential [41].

Another study showed that IGF1 expression is higher in cells obtained from patients with type 2 diabetes [39], while type 1 diabetes reduces FGF3, HGF, epidermal growth factor receptor (EGFR), and fibroblast growth factor receptor 1 (FGFR1) expression [42].

The influence of type I diabetes on the anti-inflammatory properties of MSCs was also investigated. However, the results are not conflicting. One study on people with newly-diagnosed diabetes showed no differences in the expression of genes such as programmed cell death 1 ligand 1 (PDL1), IL10, or TNF alpha induced protein 6 (TSG6) among H-MSCs and diabetic MSCs, and no differences were observed in the immunosuppressive capacity of peripheral blood mononuclear cells (PBMC) [38]. However, the results of other studies indicate that vascular cell adhesion molecule 1 (VCAM1), C-X-C motif chemokine ligand 12 (CXCL12), C-C motif chemokine ligand 2 (CCL2), CCL24, CXCL5, and CCL7 show lower expression in diabetic MSCs, whereas CCL13, CCL15, CXCL16, CCL3L3, C-C motif chemokine receptor 3 (CCR3), and C-X-C motif chemokine receptor 5 (CXCR5) show higher expression in diabetic MSCs [42]. Therefore, it is impossible to clearly determine the impact of type I diabetes on the immunosuppressive properties of MSCs. One can only speculate that the initial phase of type I diabetes development does not affect the anti-inflammatory properties of MSCs, which changes as the disease progresses.

While studying the effect of type II diabetes on the anti-inflammatory properties of MSCs, it was observed that the diabetic environment increases the expression of anti-inflammatory genes and decreases the expression of pro-inflammatory genes, which leads to an increase in the number of M2 macrophages and a decrease in the number of M1 macrophages. Therefore, it can be concluded that type II diabetes can activate the anti-inflammatory properties of MSCs [43].

For the MSCs therapy to be effective, cells must be transplanted in sufficient numbers. It is also important that after injection, MSCs go directly to the damaged location and that their survival be high enough to cause positive effects of therapy. Therefore, it is important to assess the dependence of MSCs proliferation, migration, and survival on the coexistence of various diseases.

It was observed that MSCs from adipose tissue from patients with diabetic kidney disease show a reduced ability to migrate, but their proliferation is higher [37]. MSCs from umbilical cords obtained from women with gestational diabetes show lower proliferation and faster aging in comparison to H-MSCs [44]. MSCs from bone marrow from patients with type I diabetes do not show differences in their ability to proliferate compared to H-MSCs [42]. It was also shown that MSCs from patients with type 2 diabetes have higher levels of pro-apoptotic markers than H-MSCs, which results in lower proliferation [39].

In conclusion, different types of diabetes do not affect the morphology and basic recognition criteria of MSCs. It was shown that MSCs from diabetic donors may have stronger immunomodulatory properties, but weaker pro-angiogenic properties. In addition, diabetes affects the migration, proliferation, and survival rate of MSCs (Table 1).

MSCs and Obesity

Obesity is another disorder that affects MSCs functionality. MSCs obtained from obese patients (O-MSCs) meet the criteria for MSC recognition, but obesity affects their properties. Lower proliferation was observed in O-MSCs [45,46]. Moreover, MSCs obtained from non-obese patients have higher expression of the pro-angiogenic VEGF, improve vascular remodeling, and lower blood pressure in kidneys [47]. Additionally, exosomes obtained from obese patients contain reduced amounts of VEGF, matrix metallopeptidase 2 (MMP2), and microRNA 126 (miR126) - pro-angiogenic factors [48].

A mouse study compared the therapeutic effects of O-MSCs and H-MSCs in a multiple sclerosis model, showing that O-MSCs cannot improve the motor functions of mice with multiple sclerosis, but a therapeutic effect was observed in H-MSCs. H-MSCs more strongly inhibited the demyelination of nerve cells and had anti-inflammatory effects [35]. These stronger anti-inflammatory properties of H-MSCs are also confirmed by the results of other studies, which showed lower expression of inflammatory markers IL1α, monocyte chemoattractant protein-1 (MCP1), IL6, and plasminogen activator inhibitor-1 (PAI1) compared to O-MSCs [47]. In the case of O-MSCs, an increase in proliferation and stimulation of T cell maturation was also demonstrated, which was not observed with H-MSCs. Moreover, O-MSCs have higher expression of pro-inflammatory cytokines after stimulation with IFNγ [35]. H-MSCs, unlike O-MSCs, inhibit the development of multiple sclerosis when administered in the initial phase, and are also effective at the peak of the disease. Therefore, MSCs obtained from obese patients may not be effective in the treatment of multiple sclerosis [35].

In addition to better pro-angiogenic and anti-inflammatory properties, H-MSCs reduce fibrosis and oxidative stress in the kidneys [47]. MSCs obtained from obese mice age faster, and the composition of their secretome differs from proteins secreted by cells from healthy donors. For example, MSCs collected from mice on a high-fat diet secrete proteins involved in the repair of DNA double-strand break, which may indicate the existence of greater genotoxic stress in MSCs from obese patients [46].

Obesity is one of the factors coexisting with metabolic syndrome (MS). This syndrome affects the properties of MSCs extracellular vesicles, and the occurrence of MS increases the expression of genes involved in the synthesis and release of exosomes, reduces the size of vesicles, and changes their protein composition [49,50].

Another study noted differences in miRNAs. Exosomes obtained from animals with MS contain miRNAs involved in the development of obesity and diabetes, whereas exosomes obtained from animals without MS contained miRNAs supporting cell migration, synapse synthesis, and pro-angiogenic miRNAs [51].

Other studies suggest that exosomes obtained from animals without MS have stronger regenerative properties due to proteins that support cell proliferation and differentiation [52].

A study of stem cells derived from milk collected from obese patients showed reduced expression of genes from the family of apoptosis inhibitors, such as baculoviral IAP repeat containing 5 (BIRC5) and BIRC6, which play an important role in cell apoptosis and cell division, which confirms the possible impact of obesity on the decrease in the proliferative capacity of stem cells [18]. Taking into account the possible impact of BIRC5 on maintaining the pluripotency state of stem cells, reduced BIRC5 expression probably also affects the differentiation abilities of these cells [18,53].

MSCs from MS patients are less susceptible to TNFα-stimulated apoptosis [52], which may be related to the fact that MS affects the pro-inflammatory properties of MSCs exosomes by inducing the expression of TNFα, MCP1, IL6, and IL1β [52].

To sum up, MS and isolated obesity do not affect the morphology of MSCs, but they reduce the angiogenic potential of MSCs and can induce the pro-inflammatory nature of the cells. Obesity reduces MSCs proliferation, increases genotoxic stress, and stimulates faster cell aging. MS reduces cell migration and decreases the size of extracellular vesicles secreted by MSCs while increasing their number (Table 2).

MSCs and Autoimmune Diseases

MSCs can inhibit the inflammatory reaction, and they reduce oxidative stress, which can be used in treating autoimmune diseases. In addition, they support the process of angiogenesis, thus accelerating the regeneration of already damaged organs, and they have a neuroprotective effect, so they might be used in the treatment of, for example, multiple sclerosis. MSCs seem to be an effective tool in counteracting autoimmunization and reducing its effects [55]. Before using them, however, it is worth assessing whether autoimmune diseases in any way affect the properties of MSCs and whether autologous transplants will be as effective as H-MSCs transplants.

Studies that assessed the properties of MSCs obtained from patients with various autoimmune diseases (AD-MSCs) showed that they do not differ from H-MSCs in terms of morphology, immunophenotype, multipotency ability, or proliferation intensity. It was also noted that AD-MSCs retain the ability to inhibit PBMC proliferation [56].

More detailed studies were conducted that evaluated MSCs obtained from patients with specific autoimmune diseases. Studies of MSCs obtained from bone marrow from patients with multiple sclerosis met the criteria for the MSCs recognition. However, multiple sclerosis was shown to reduce the amount of CD105 and CD73 on MSCs. Moreover, MSCs obtained from patients with multiple sclerosis (SM-MSCs) resemble senescent cells [57] and are characterized by lower proliferation [58]. Differences in the expression of many genes between H-MSCs and SM-MSCs were also observed. Multiple sclerosis was shown to result in lower expression of TGFβ1, HGF, IL10, IL6, interferon gamma receptor 1 (IFNGR1), and IFNGR2. SM-MSCs secrete lower concentrations of IL10 and TGFβ than H-MSCs. The same study noted that H-MSCs inhibited T cell proliferation more strongly than SM-MSCs. It can therefore be concluded that SM-MSCs have weakened immunomodulatory properties and, due to lower HGF expression, their ability to stimulate angiogenesis is also impaired [57]. Moreover, it was observed that SM-MSCs secrete lower amounts of antioxidants compared to H-MSCs and are less resistant to induced stress [59].

In another experiment, neurons were co-cultured with MSCs. Neuronal survival was lower when cultured with SM-MSCs compared to cultures with H-MSCs [60].

Another autoimmune disorder is systemic sclerosis (SS). SS does not affect the morphology of MSCs obtained from adipose tissue. MSCs express the same surface antigens, but it was noticed that the expression of CD146, which is considered to be a marker of multipotency, is lower. However, no difference was observed in the ability to transform into bone cells and adipose tissue cells between H-MSCs and MSCs obtained from SS patients (SS-MSCs). Moreover, SS-MSCs retain the ability to inhibit lymphocyte proliferation and create new endothelial tubes [61].

However, other studies showed that SS-MSCs from bone marrow, differentiated into endothelial cells, form endothelial tubes more slowly than H-MSCs. This may be caused by a reduced amount of VEGFR2, which is involved in angiogenesis, and a reduced amount of CXCR4 [62], which supports organ regeneration [63].

Yet another study found that SS-MSCs from bone marrow show increased expression of VEGFA and VEGFR2 compared to H-MSCs. Endothelial cells co-cultured with SS-MSCs were observed to support the formation of new endothelial tubes, exactly like H-MSCs [64].

Thus, SS-MSCs differentiated into endothelial cells form new vessels more poorly than H-MSCs, but SS-MSCs co-cultured with endothelial cells support new vessel formation at the same level as H-MSCs. From 2 apparently contradictory studies, it can be concluded that SS-MSCs are more effective in supporting endothelial cells in the formation of new vessels than cells differentiated into endothelial cells. This confirms the hypothesis that in terms of SS, the use of paracrine properties of MSCs is more beneficial in therapy than differentiating MSCs into the desired cells.

Another important feature of MSCs is their influence on tissue fibrosis. SS-MSCs show higher expression of alpha-smooth muscle actin (αSMA), collagen type I alpha 1 chain (Col1A1), and Col1A2, which may stimulate this process [64]. Compared to H-MSCs, SS-MSCs have lower proliferation, greater sensitivity to genotoxic stress, and increased levels of markers of cellular senescence. There were no significant differences in the inhibition of PBMC proliferation by SS-MSCs and H-MSCs. Higher IL6 expression and no difference in TGFβ levels were observed in SS-MSCs, while TGFβ expression significantly increased after SS-MSCs were cultured with PBMCs compared to H-MSCs/PBMCs [65]. However, another study showed higher expression of one of the TGFβ receptors, transforming growth factor β receptor II (TBRII), in SS-MSCs [66].

Another study found that rheumatoid arthritis (RA) does not significantly affect the properties of MSCs (RA-MSCs), and RA-MSCs retain their morphology and function. No significant differences were found in the levels of angiogenic or inflammatory cytokines. Only a reduction in the proliferative potential was observed, which does not exclude the use of RA-MSCs in therapy [67].

Another study assessed the properties of MSCs from patients with systemic lupus erythematosus (SLE-MSCs). This disease does not affect the morphology, immunophenotype, or differentiation capacity of MSCs. However, lower proliferation, DNA damage, and p53 and p16 activation were observed in SLE-MSCs, which results in faster cell aging. Moreover, a weakening of anti-inflammatory functions was demonstrated. SLE-MSCs had higher expression of IL6 and IL8, as well as higher secretion of corresponding cytokines and lower expression of TGF-β, IDO1, and LIF interleukin 6 family cytokines (LIF) [68].

MSCs obtained from patients with psoriasis (P-MSCs) do not show morphological or immunophenotypic changes. They also retain the ability to differentiate. However, lower proliferation, more frequent apoptosis, and reduced expression of genes involved in inflammatory processes were noticed [69].

In summary, there is no single specific pattern depicting the impact of autoimmune diseases on the properties of MSCs. Depending on the disease, different changes in MSCs properties are observed (Table 3).

MSCs and Inflammatory Diseases

MSCs perform immunomodulating functions. They regulate the proliferation of lymphocytes and secrete a number of substances with anti-inflammatory effects. MSCs have an impact on the course of inflammation, but the inflammatory environment induces changes in MSCs properties, too. In the case of MSCs from bone marrow, it was shown that an inflammatory environment can increase the amount of secreted NGF, GDNF, and VEGF [70], which increases the repair potential of nerve tissue. Inflammation was also observed to enhance the immunoregulatory properties of MSCs. Inflammation decreases cytochrome C oxidase I (COX1) expression and increases COX2 expression, and it also increases PGE2 secretion. Moreover, it increases HGF expression and reduces TGFβ expression [71].

On the other hand, due to inflammation, the expression of IL6, IL8, IL1β, IL1Ra, TNFα, and CCL5 increases in MSCs from adipose tissue, which can intensify inflammation [72].

Other studies on MSCs from adipose tissue showed that inflammation slightly increases the production of HGF and COX2 and significantly increases the expression of IDO. Moreover, the inflammatory environment was observed to increase the expression of genes involved in the antiviral and antibacterial protection of MSCs. These studies also confirm the increase in the expression of pro-inflammatory cytokines – IL8 and IL1β and the TNF family [73].

Activated PBMCs increase the expression of HLA class I on MSCs, while pro-inflammatory cytokines increase the expression of HLA class II. Thus, it can be assumed that inflammation will increase the immunogenicity of MSCs; therefore, they will not be suitable for transplantation from an unrelated donor [73].

An increase in the expression of the chemokines CXCL1, CXCL6, CXCL9, CXCL10, and CXCL11 was also observed. Additionally, it was noticed that because of pro-inflammatory cytokines, the expression of 13 types of collagens is reduced, which may lower the risk of fibrosis [73].

The inflammatory environment was observed to increase MSCs diameter [73]. This was also confirmed in studies on MSCs from bone marrow. Here, it was also observed that pro-inflammatory cytokines, IFNγ and TNFα, change the morphology of MSCs. MSCs affected by IFNγ increase their size and change their shape into a more irregular one, and in the case of TNFα, the cells become more spindle-shaped. However, no changes in morphology were observed in the umbilical cord MSCs [74]. The same study showed that bone marrow MSCs in an inflammatory environment reduced IFNγ more effectively than control MSCs. Additionally, it was proven that TNFα reduces HGF secreted by MSCs from the umbilical cord [74], and that the presence of TNFα increases MSCs proliferation, whereas IFNγ decreases MSCs proliferation [74]. This indicates an ambiguous impact of inflammatory factors on the number of MSCs.

In conclusion, the above studies indicate that inflammation can enhance the pro-inflammatory effects of MSCs, although this effect is ambiguous (Table 4). Differences were also found depending on the source of MSCs.

Other Factors and Medical Conditions

Diseases that have been poorly discussed that change the properties of MSCs include renal vascular disease, hypothyroidism, and hypertension. However, it is not only the occurrence of pathology that affects MSCs; individual characteristics such as age, sex, and laboratory parameters were also observed to affect the behavior of MSCs.

Renal vascular disease does not affect the typical characteristics of MSCs (RVD-MSCs) – cells retain their morphology and immunophenotype, but the secretion capabilities of MSCs change. Compared to H-MSCs, RVD-MSCs secreted less VEGF but more HGF. Additionally, RVD-MSCs show a lower ability to migrate and contain partially damaged DNA [75].

Differences in the expression of inflammatory cytokines were also noticed between H-MSCs and MSCs obtained from the umbilical cord in patients with hypertension (Hy-MSCs). Hy-MSCs show higher IL1A expression and lower IL6R expression. Moreover, MSCs isolated from the umbilical cord obtained from patients with hypertension have higher expression of the VEGFA and VEGFC genes [76]. Additionally, higher expression of TSG6 [14] and higher expression of VEGFA [76] were observed in the case of umbilical cord MSCs collected from women with hypothyroidism.

Interestingly, differences in the paracrine signaling of MSCs were observed between male and female donors. MSCs obtained from female rats secrete higher amounts of VEGF, but this did not seem to improve the angiogenesis process. Additionally, female MSCs produce more IL10, which results in stronger anti-inflammatory properties [77].

A significant influence of patient age on the properties of MSCs was observed. The length of the telomeres of these cells decreased with age, which may affect the viability of MSCs. Age-dependent changes in the ability to create new blood vessels were also demonstrated. MSCs from older patients stimulate capillary formation less strongly than MSCs from younger patients, perhaps because younger MSCs secrete greater amounts of VEGF, PIGF, HGF, and angiogenin [78].

Donor age also influences the neuroprotective properties of MSCs. In neurons cultured from MSCs, nerve cell survival decreases with increasing age of the MSCs donor, and with age, the resistance of MSCs to NO-mediated neurotoxicity decreases [60].

In MSCs collected from the umbilical cord, the expression of genes encoding pluripotency factors such as SRY-box transcription factor 2 (SOX2) or POU domain, class 5, transcription factor 1 (POU5F1) decreases with the age of the patient [79,80], and the expression of apoptosis inhibitors in umbilical cord MSCs decreases with patient age [81].

In previous studies, we have shown that the expression of some genes that are responsible for the therapeutic properties of MSCs is associated with various physicochemical parameters. The study assessed MSCs from the umbilical cord. For example, it has been shown that the expression of TSG6, a gene with strong anti-inflammatory properties, depends on the pH of umbilical cord blood. Moreover, the pH of umbilical cord blood affects the expression of IL1A and IL6, which are associated with pCO2 level. In turn, pO2 affects the expression of IL6. Other parameters that may influence the expression of inflammatory cytokines include WBC, RBC, PLT, and hemoglobin levels [14]. Also, the expression of one of the key stem cell pluripotency factors, SOX2, in the examined umbilical cord MSCs is dependent on the physicochemical parameters of cord blood, such as pH, pCO2, and pO2. Increased levels of SOX2 gene expression are observed in MSCs in the more acidic environment of cord blood [79]. Expression of genes encoding apoptosis inhibitors BIRC2, BIRC3, and BIRC5 in umbilical cord MSCs depends on the pH of umbilical cord blood, and an increase in the expression of these genes was also noted at the more acidic pH of umbilical cord blood [81].

In mesenchymal stem cells isolated from the umbilical cord, the type of delivery, the number/sequence of pregnancies, and the date of delivery are also important for the functionality of these cells. In premature births, increased expression of VEGFB [76] and increased expression of SOX2 [79] are observed in MSCs of the umbilical cord. Stem cells collected from umbilical cords from patients giving birth naturally and in those giving birth for the first time have increased expression of genes encoding key pluripotency factors [79–81]. In stem cells obtained from breast milk, the type of delivery, the date of delivery, and the number of pregnancies and deliveries are important for the secretory properties of these cells [18].

Paracrine Properties of MSCs Depending on Their Source

MSCs are present in most human body tissues, which gives us many potential sources for obtaining them. The most popular sources include MSCs from bone marrow (BM-MSCs), adipose tissue (AD-MSCs), umbilical cord (UC-MSCs), and umbilical cord blood. Before selecting a place to take a sample of MSCs, one should take into account the invasiveness of the procedure and whether the cells from this particular place will meet therapeutic expectations.

One study examined differences in gene expression between MSCs obtained from lungs, adipose tissue and bone marrow. It was observed that the expression of FGF10 and HGF was the highest in lung MSCs, while FGF7 expression was the lowest in these cells [82].

Other studies demonstrated that UC-MSCs, due to differences in gene expression, may have a greater pro-angiogenic potential compared to BM-MSCs, while BM-MSCs may have a more beneficial effect on the development of the skeletal system than UC-MSCs [83].

However, another research group reached different conclusions. Here, the properties of mature MSCs were compared to those of developmentally younger MSCs. For this purpose, AD-MSCs and BM-MSCs were used as mature cells and UC-MSCs as younger cells. This study demonstrated that the degree of cell development and thus their source have a significant impact on the properties of MSCs. It was shown that MSCs from mature tissues have stronger pro-angiogenic properties, which may be due to the more intense secretion of FGF, PDGF, and VEGF. In turn, young MSCs have stronger immunomodulatory properties. This study explored the cell secretome, but not their expression level [84].

The situation changed when properties were compared among different groups of MSCs divided by source. It was then observed that BM-MSCs have the greatest immunomodulatory potential, followed by UC-MSCs, and AD-MSCs have the weakest immunomodulatory properties [84].

Moreover, the study investigated the effects of IFNγ and TNFα on the behavior of BM-MSCs and UC-MSCs, finding that IFNγ increases the expression of HLA-DR in BM-MSCs. However, no expression of this gene was detected in UC-MSCs. This may indicate that UC-MSCs are less immunogenic than BM-MSCs. On the other hand, BM-MSCs were shown to secrete more PGE2, which may result in stronger immunomodulatory properties of these cells compared to UC-MSCs [74].

Future Directions – Regenerative Medicine

Regenerative medicine is a field that will enable the introduction of modern therapies based on stem cells in the future. MSCs have great therapeutic potential, but their properties depend on many variables (Table 5). It is necessary to conduct further research that will provide new information about how other diseases and the patient’s condition affect the properties of MSCs. Furthermore, a more accurate comparison of cells obtained from different sources is needed. This will make it possible to select cells for specific needs, which will make the therapy more targeted and ensure its greatest possible effectiveness.

Conclusions

Pathological factors such as diseases occurring in the patient, as well as physiological conditions and individual factors, affect paracrine signaling and thus the functionality of stem cells and their therapeutic potential. Knowledge of clinical factors allows for better use of the potential of stem cells.