TA-MSCs and TA-MSCs-EVs have stronger immunosuppressive effects than normal MSCs, including inhibition of anti-tumor immune cells and promotion of immunomodulatory cells, which effectively contribute to malignant tumor progression. Notably, MIF can also induce immunosuppression by regulating various cell types in TME, including T cells, NK cells, DCs, MDSCs, TAMs, and Tregs, triggering immune escape by not only preventing tumor cells from being killed by anti-tumor immune cells but also promoting formation of immunosuppressive cells or enhancing their function, and ultimately, tumor progression.
TA-MSCs, TA-MSCs-EVs and MIF inhibit anti-tumor immune cells
T cells. T cells, an important component of TME and the immune system, mainly exert anti-tumor effects. Extensive studies suggest that MSCs inhibit both T cells proliferation [
109] by secreting Prostaglandin E2 (PGE2) and programmed death ligand-1 (PD-L1) [
110] and T cells anti-tumor immunity [
38] by secreting immunosuppressive cytokines, such as IL-4 and IL-10 [
111], which are not necessarily related to the MSC source. TA-MSCs also promote the formation and maintenance of immunosuppressive TME via inhibition of anti-tumor T cells. For example, NB-MSCs have been shown to significantly suppress proliferation of activated T cells [
112]. Pancreatic cancer (PC)-MSCs and melanoma-MSCs exert similar effects on T cells [
38,
113]. In a previous study, LC-MSCs reduced the number of CD4
+ T cells producing IL-17 and cytotoxic CD8
+ T lymphocytes (CTLs) and suppressed the expression of cytotoxic molecules (Fas ligand (FasL), perforin (PFP) and CD107) in CTLs that mediate anti-tumor immune responses [
114]. Recent research has disclosed that acute myeloid leukemia (AML)-MSCs are able to block the production of leukemia-reactive CTLs via a novel COX2/PG/NR4A/WNT signaling pathway, resulting in attenuation of anti-tumor immunity [
115]. CD39 and CD73 ectonucleotidases on the cervical cancer-derived MSC (CeCa-MSC) surface efficiently hydrolyze ATP, ADP and AMP nucleotides to generate adenosine (Ado), which inhibits the proliferation and activation of CTLs as well as anti-tumor immune responses [
36].
Previous experiments clearly demonstrate that MSCs-EVs molecules inhibit differentiation and activation of T cells in addition to reducing T cells proliferation and interferon-γ (IFN-γ) release [
116]. Recently, MSCs-EVs were shown to significantly suppress CD4
+ T cells proliferation, IL-17 and IFN-γ levels and enhance TGF-β and IL-10 secreted by T cells [
117], which could be attributed to miR-223 delivery [
118]. MSCs-EVs also inhibit T cells proliferation and infiltration [
119] via TGF-β and adenosine signaling [
120] concomitant with a decrease in the percentage of CD4
+ and CD8
+ T cells subsets [
121]. Furthermore, both BC-MSCs and BC-MSCs-EVs could inhibit production of IFN-γ by CD4
+ and CD8
+ T cells, thus thwarting the anti-tumor immune response [
122].
MIF downregulates NKG2D to inhibit the antitumor immunity of CD8
+ T cells [
123], inducing immune escape of glioma cells [
124]. Furthermore, high levels of MIFs protect BC cells from immunogenic cell death (ICD) and inhibit the antitumor immune response mediated by T cells producing IFN-γ [
125]. Similarly, neuroblastoma cells secreting MIF inhibit T cell proliferation and induce cell death through an IFN-γ pathway, which eliminates activated T cells from TME, thus contributing to tumor cell escape from immune surveillance [
126]. In an earlier study by Zhou and co-workers, production of high amounts of MIF by neuroblastomas (NB) led to a decrease in the number and infiltration of CD8
+ and CD4
+ T cells, and ultimately, suppression of antitumor immunity [
127]. MIF has additionally been shown to inhibit the activation of CD4
+, CD8
+ T cells and CTLs in the tumor regions of cancer-bearing mice [
128]. Downregulation of MIF in BC is associated with a significant increase in infiltration and anti-tumor functions of CD4
+ and CD8
+ T cells [
129].
Above studies have shown that TA-MSCs, TA-MSCs-EVs and MIF enhance the immunosuppressive activity of tumor microenvironmentnot by inhibiting the number and function of CD4+ and CD8+ T cells and reducing the expression of IFN-γ in T cells, which can significantly inhibit the anti-tumor immune responses of T cells.
Nature killer (NK) cells. NK cells are major tumor suppressor cells. The CD56
bright subsets play an immunomodulatory role through secretion of cytokines and CD56
dim subsets exert cytotoxic effects through cell degranulation [
130]. Recent studies have reported negative effects of MSCs on NK cells proliferation. Conditioned medium (CM) of MSCs has been shown to induce a significant reduction in the number of NK cells in metastatic tissues of lung cancer, leading to inhibition of anti-tumor cytotoxicity [
114]. TA-MSCs also inhibit NK cells-mediated anti-tumor immunity through simultaneously suppressing immunomodulatory factor secretion and degranulation of NK cells [
131]. LC-MSCs induce a decrease in NK cells number while significantly attenuating anti-tumor cytotoxicity in an NO- and indoleamine 2,3-dioxygenase (IDO)-dependent manner [
114]. Furthermore, by secreting IL-6 and PGE2, squamous cell lung carcinoma-derived MSCs (SCC-MSCs) not only regulate the immunomodulatory effects of CD56
bright NK cells through inhibition of IFN-γ and TNF-α secretion but also downregulate NK cells-activating receptors and block CD56
dim-mediated NK cells degranulation to inhibit the cytotoxicity of NK cells [
37]. MSCs-EVs molecules additionally inhibit proliferation, activation, and cytotoxicity of NK cells through activation of downstream TGF-β/Smad2/3 signaling mediated by latency-associated peptide (LAP), thrombospondin 1 (TSP1) and TGF-β delivery [
132].
Besides, MIF inhibits antitumor immunity through downregulating NKG2D on NK cells at the transcriptional level, in turn, impairing NK cells cytotoxicity towards ovarian cancer cells [
123,
124]. Melanoma-derived MIF suppresses NK cells-mediated killing of uveal melanoma cells, thus maintaining an immunosuppressive TME [
133]. A significant negative correlation between serum MIF and NK cells levels has recently been reported in ovarian cancer patients before and after chemotherapy, suggesting that MIF inhibits the production and activation of NK cells [
134]. On the other hand, Loyon et al
. shown that IL-21-induced NK cells affect CD4
+T cell priming by secreting MIF [
135]. Additionally, hypoxia is reported to promote secretion of MIF in NK cells and induce apoptosis of leukemia target cells [
136]. Current researchs show that TA-MSCs, TA-MSCs-EVs and MIF inhibit NK cells cytotoxicity and degranulation, but there are few studies on the specific mechanisms, which need to be further elucidated.
Dendritic cells (DCs). DCs are the most powerful antigen-presenting cells that activate anti-tumor immunity through stimulating naive T cells and specific T cells proliferation to inhibit tumor progression during the early stages of tumorigenesis. LC-MSCs significantly reduce the number of DCs by producing tumor necrosis factor (TNF)-α [
114]. Melanoma-MSCs suppress the expression of cystathionase in DCs through the IL-10-STAT3 pathway, thus blocking export of cysteine from DCs to T cells, leading to reduced proliferation and effector function of T cells [
38,
137]. MSCs also inhibit maturation and typical functions of DCs (such as IL-12 production and the ability to prime T cells) through release of PGE2 [
138] and TNFα-stimulating gene (TSG)-6 [
139]. Regulatory DCs (regDCs), a distinct DCs population differentiated from mature DCs (mDCs), is characterized by limited T cells proliferation, high endocytotic capacity, low immunogenicity, and strong immunoregulatory function [
140]. MSCs induce differentiation of mDCs into regDCs via paracrine hepatocyte growth factor (HGF) [
141]. Additionally, regDCs are generated by MSCs from hemopoietic progenitor cells (HPCs) synergistically via the Notch and TGF-β signaling pathways [
142]. Zhao et al. [
143] showed that CML-MSCs induce differentiation of mDCs into regDCs, which inhibit proliferation of T cells through both TGF-β1 and production of Tregs or T cells anergy. Shahir and co-workers [
144] reported that MSCs-EVs inhibit DCs maturation and IL-6 release while increasing IL-10 and TGF-β release.
A number of researchers suggest that MIF secreted by glioblastoma inhibits migration and maturation of DCs, suppressing the anti-tumor immune response [
145]. MIF not only suppresses DCs maturation and activation but also significantly impairs their ability to activate cytotoxic T cells killing function, facilitating metastatic melanoma progression [
146]. Furthermore, MIF has been shown to inhibit migration of both immature DCs (iDCs) and mDCs, meanwhile impair the expression of co-stimulatory markers [
145]. In a mouse model of BC, MIF depletion led to an increase in the abundance and activation of DCs, further confirming that MIF mediates tumor growth promotion through DCs inhibition [
125]. Interestingly, inhibition of MIF resulted in functional reversion of MDSCs from an immunosuppressive to immunostimulatory DCs-like phenotype [
147].
TA-MSCs, TA-MSCs-EVs and MIF show similar inhibitory effects on DCs, they can inhibit DCs recruitment into immunosuppressive TME, meanwhile suppress maturation and activation of DCs, thus indirectly promoting tumor progression.
T helper cells (Th). Th is one of the T cells subsets that includes Th1, Th2 and Th17 (IL-17-producing effector T cells). Th1 and Th2 are often in a state of balance and disruption of this equilibrium is proposed to affect tumor progression. Th2 mainly plays a promote-tumor role while Th1 and Th17 exert anti-tumor effects [
148,
149]. In an earlier study, in vivo injection of multipotent MSCs derived from human-induced pluripotent stem cells (huiPS-MSCs) significantly reduced Th1 in mouse spleen and peripheral blood mononuclear cells (PBMCs) [
150]. Similarly, GC-MSCs show the ability to not only reduce the Th17 level in PBMCs but also inhibit Th17 proliferation [
151].
MSCs-EVs also promote transformation of Th1 into Th2 and reduce the ability of T cells to differentiate into Th17 [
152]. Another recent study has reported that MSCs-EVs inhibit the differentiation of tumor-suppressing Th1 and Th17 [
117].
Multiple studies have demonstrated that the MIF level in TME is positively correlated with Th2 and negatively correlated with Th1, although the specific mechanisms are yet to be clarified [
153‐
155]. A high level of MIF in TME is proposed to induce a change in the balance of Th1/Th2 differentiation, leading from Th to Th2 phenotypic differentiation [
154]. In addition, MIF significantly enhances lymphocyte production of Th2 cytokines, such as IL-2, after antigen stimulation [
156]. In nasopharyngeal carcinoma, MIF promotes the formation and migration of Th17 cells mediated by the MIF-CXCR4 axis and dependent on the mTOR pathway [
157].
Studies have shown that TA-MSCs, TA-MSCs-EVs and MIF affect the proportion balance of Th phenotype and promote the transformation of Th into promote tumor Th2 phenotype, however, the specific mechanism of action remains to be elucidated.
TA-MSCs, TA-MSCs-EVs and MIF promote immunosuppressive cells
Myeloid-derived suppressor cells (MDSCs). MDSCs, an immunosuppressive cell type in TME, inhibit anti-tumor immunity through a variety of mechanisms. Zhao et al
. [
158] confirmed that MSCs effectively facilitate accumulation of MDSCs to TME, as reported for TA-MSCs. CML-MSCs enhance immunosuppressive MDSCs production and aggregation, promoting tumor progression via upregulating immunomodulatory arginase-1 (Arg-1), IL-6, IL-1β, COX-2, and TNF-α in MDSCs and inhibiting anti-tumor immunity [
40].
By carrying high levels of TGF-β, C1q, BC-MSCs-EVs not only enhance the immunosuppressive activity of MDSCs and induce conversion to type M2 macrophages expressing high levels of PD-L1, but also reduce PD-1 expression in infiltrating T cells through upregulation of TGF-β in MDSCs, thus inhibiting the anti-tumor immune response [
122].
Huang et al. [
159] demonstrated that MIF knockout inhibits not only recruitment of MDSCs but also tumor growth and metastasis, supporting the theory that MIF promotes tumor progression by exerting effects on MDSCs. MIF is considered to be a determinant of melanoma MDSCs differentiation and immune suppression [
160]. MIF induces differentiation and stimulates chemotaxis of MDSCs through activating PI3K/AKT and p38/MAPK pathways in head-and-neck squamous cell carcinoma (HNSCC) [
161]. MyD88-dependent MAPK and NF-κB pathways are also involved in MIF-CXCR2-mediated recruitment of MDSCs to bladder cancer TME [
162]. A decrease in MIF in mouse BC tissue results in significant reduction of circulating MDSCs and its suppressive cytokines along with inhibition of growth and metastasis of BC in vivo, further indicating that MIF promotes tumor progression by regulating the number and function of MDSCs [
129]. Interestingly, MIF secreted by CSCs is reported to enhance immunosuppression mediated by MDSCs through binding the CXCR2 receptor, which facilitates glioblastoma immune evasion [
106]. A further study suggests that MIF relies on its tautomerase activity to promote myeloid cell differentiation into mononuclear MDSCs (mMDSCs), promoting the formation of immunosuppressed TME and consequently, tumor growth and metastasis [
163].
The effects of TA-MSCs, TA-MSCs-EVs and MIF on MDSCs are reflected in promoting the aggregation of MDSCs in immunosuppressive TME and enhancing the function of MDSCs by upregulating the expression of immunoregulatory cytokines in MDSCs.
Regulatory T cells (Tregs). Tregs represent a special subgroup of T cells that play a pivotal tumor-promoting role. Tregs induce tumor progression by inhibiting anti-tumor immunity and are linked to poor prognosis [
164]. MSCs have the capacity to generate Tregs [
111,
150] and enhance their tumorigenic activity [
110]. A recent study indicates that AML-MSCs and myelodysplastic syndrome (MDS)-MSCs efficiently induce Treg generation associated with sustained leukemic cell viability and proliferation [
165]. BC-MSCs and GC-MSCs induce generation of Tregs and their production of IL-10, IL-17 and TGF-β, in turn, facilitating tumor cell progression [
166,
167]. Mechanistically, IL-15 derived from GC-MSCs stimulates Tregs through activation of STAT5 in CD4
+ T cells and upregulation of PD-1 [
79].
Several reports have confirmed that MSCs-EVs enhance proliferation of Tregs and their immunosuppressive cytokines, including IL-10 [
152,
168]. MSCs-EVs are reported to promote proliferation and immunosuppressive capacity of Tregs via upregulating IL-10 and TGF-β1 secreted from PBMCs [
169]. Although the issue of whether TA-MSCs-EVs affect Tregs remains to be definitively established, findings to date clearly support the theory that TA-MSCs-EVs promote immunosuppressive TME formation by increasing the number and function of Tregs.
The use of exogenous MIF can increase the number of IL-10-producing Tregs via Toll-like receptor 2 [
170] in the colon and peritoneal cavity of mice [
171]. MIF promotes tumor growth by increasing Tregs production through modulation of IL-2 in a colon cancer model mouse model [
172]. Previous studies have identified a role of MIF in recruitment of Tregs [
173]. The use of MIF receptor antagonists led to a significant reduction in the number of Tregs in metastatic melanoma tissues in mice [
146], indicating that MIF promotes tumor development via effects on Tregs.
Similar to the effect on MDSCs, TA-MSCs, TA-MSCs-EVs and MIF promote the production and recruitment of Tregs in immunosuppressive TME and enhance the expression of immunomodulatory factors, thus enhancing the tumor-promoting effect of Tregs.
Tumor-associated macrophages (TAMs). TAMs present in TME are influenced by various factors. These macrophages are usually induced and polarized to the tumor-promoting M2 phenotype. A number of studies indicate that TA-MSCs affect the quantity and function of TAMs. For example, GC-MSCs secrete IL-6 and IL-8 to activate the JAK2/STAT3 signaling pathway in TAMs and promote polarization to M2 phenotype, leading to enhanced proliferation and metastasis of GC cells [
167]. In turn, EVs isolated from GC cells facilitate TAMs recruitment by activating NF-kB signaling in GC-MSCs while enhancing the phagocytic function of TAMs, upregulating IL-6 and IL-8 secretion, indicating the existence of a feedback loop between tumor cells and TA-MSCs [
167]. Chemokines secreted by ovarian cancer (OC)-MSCs promote polarization of TAMs to M2 phenotype and chemotherapy resistance of OC cells [
174]. Melanoma-MSCs have also been shown to induce TAMs polarization to M2 phenotype, stimulating angiogenesis and tumor progression [
175].
Consistent with the above findings, MSCs-EVs have been shown to promote TAMs M2 polarization mediated by miR-21-5p [
81]. Interestingly, MSCs-EVs could be efficiently internalized by TAMs, eliciting a switch from M1 to M2 phenotype [
176]. In addition, BC-MSCs-EVs promote the polarization of M0 to M2 phenotype, upregulation of PD-L1 expression in M2 macrophages, and ultimately, growth and metastasis of BC [
122].
Research on metastatic melanoma shows that MIF interacts with CD74 on TAMs, which triggers activation of AKT, ERK and downstream signal pathways and increases the expression of immunosuppressive factors in TAMs, including TGF-β, IL-10, IL-6, arginase-1 and PD-L1, enhancing their tumor-promoting effects [
146]. MIF also triggers TAMs polarization into M2 tumor-promoting phenotypes via CD74 and CXCR7 signal transduction, which not only increases the pro-angiogenic potential of TAMs but also promotes MM cell survival, proliferation, tumor growth and metastasis [
177]. Similar results have been obtained in lung cancer [
178] and melanoma [
179]. In addition, MIF promotes the recruitment and infiltration of macrophages in mice [
151], potentially mediated by the MIF-dependent chemokines monocyte chemotactic protein-1 (MCP-1), CXCL10, and macrophage inflammatory protein 2 (MIP2) [
180]. Conversely, a study on CLL mice reported that the absence or inhibition of MIF reduced the number and migratory activity of TAMs, leading to changes in their distribution, concomitant with increased apoptosis of CLL cells [
181]. The promotor effect of macrophage recruitment is proposed to be related to the tautomerase activity of MIF [
182].
These studies suggest that TA-MSCs and TA-MSCs-EVs promote the polarization of TAMs to the tumor-promoting M2 phenotype and increase the expression of immunosuppressive cytokines, MIF may play a role as a mediator in this process.
Neutrophils. Interactions of TA-MSCs with neutrophils are also reported to stimulate tumor progression. IL-6 secreted by GC-MSCs induces neutrophils chemotaxis by activating the STAT3-ERK1/2 signal cascade. Activated neutrophils promote expression of IL-8, TNF-α and CCL2, which protect against spontaneous apoptosis [
183]. Furthermore, activated neutrophils induce MSCs to differentiate into CAFs, enhancing the growth and metastasis of GC [
183]. Similarly, smoldering MM (SMM)-MSCs activate neutrophils and induce an immunosuppressive phenotype through TLR4 signaling. Activated neutrophils not only recruit immunosuppressive Tregs but also induce CD8
+ T cells apoptosis via upregulation of reactive oxygen species (ROS), which promotes immunosuppressive TME formation and tumor progression [
131]. Furthermore, BC-MSCs recruit CXCR2
+ neutrophils into TME, inducing a significant increase in expression of metastasis-related genes (CXCR4, CXCR7, MMP12, MMP13, IL-6 and TGF-β) in tumor cells and consequent metastasis [
184].
MSCs-EVs molecules have protective effects on neutrophils phagocytosis capacity and lifespan [
185] as well as ROS production, along with inhibitory effects on neutrophils apoptosis [
186].
The ability of MIF to promote neutrophils recruitment is documented in the literature [
187]. MIF is highly expressed in head-and-neck cancer (HNC) and stimulates functions of neutrophils by enhancing their CXCR2-dependent recruitment and survival and release of CCL4 and MMP9, in turn, promoting tumor progression [
188]. Neutrophils infiltration could also be induced by MIF produced by leukemia cells [
189]. In an in vivo study, MIF-deficient mice showed markedly reduced neutrophils infiltration, tumor incidence and angiogenesis upon chronic UVB exposure [
73]. Clinically, MIF and neutrophils counts are significantly positively correlated in various tumors, such as HCC [
190] and GC [
191], indicating that MIF in TME is potentially responsible for neutrophils recruitment. In addition, MIF has been shown to significantly inhibit neutrophils apoptosis via interacting with CXCR2 [
192].
TA-MSCs, TA-MSCs-EVs and MIF mainly promote the recruitmention in TME, survival and secretory function of neutrophils, which not only inhibits the spontaneous apoptosis of neutrophils but also suppresses the killing of tumor cells by other anti-tumor immune cells.
Cancer-associated fibroblasts (CAFs). CAFs are involved in almost all stages of tumor progression and contribute significantly to tumor invasion, angiogenesis, regulation of TME metabolism, immune cell recruitment and reprogramming as well as chemotherapy resistance [
193]. TA-MSCs can be induced to differentiate into CAFs by tumor cells or other components of TME to promote tumor progression. For example, in BC and CRC, MSCs are specifically recruited into TME where they are induced to differentiate into CAFs via paracrine pathways, such as TGF-β and PDGFR-β [
194,
195]. Tumor-educated blood platelets (TEP) also induce MSCs differentiation into CAFs via stimulation of TGF-β expression [
196]. In bladder cancer, TGF-β1 receptor and Smad2 are involved in differentiation of MSCs to CAFs, in turn, promoting tumor growth in vivo [
197]. TGF has therefore been identified as the key signal in transformation of MSCs into CAFs. Lactate secreted by pancreatic cancer (PC) cells facilitates transformation of MSCs to CAFs by inducing an increase in 5-hydroxymethylcytosine (5hmC) levels [
198]. Hepatoma-derived growth factor (HDGF) secreted by GC cells infected with
H. pylori recruits MSCs and induces their differentiation into CAFs, leading to enhanced survival and invasive ability [
199]. In addition, EVs derived from MM cells induce transformation of MSCs into CAFs with increased IL-6 secretion via delivery of miR-21 and miR-146a [
200]. Chemotherapy agents, such as cytarabine and daunorubicin, are also reported to promote phenotypic transition from MSCs to CAFs [
201].
TA-MSCs, TA-MSCs-EVs and MIF promote tumor development through regulation of other factors
Tumor angiogenesis. Tumor angiogenesis is a necessary condition for rapid tumor growth and metastasis. An earlier study utilizing high-resolution isoelectric focusing-coupled liquid chromatography tandem mass spectrometry (HiRIEF LC–MS/MS) achieved full characterization of 1927 proteins in MSCs-EVs, including several potential paracrine factors related to angiogenesis, such as PDGF, epidermal growth factor (EGF), FGF and NF-kB signaling pathways [
202]. LC-MSCs promote tumor angiogenesis by secreting various pro-angiogenic factors (ASPN, Clusterin, vascular endothelial growth factor (VEGF), IL-8, Ang and PDGF-BB) [
42]. Thyroid hormones additionally upregulate angiogenesis-related factors, such as Ang and insulin-like growth factor 1 (IGF1), and stimulate VEGF signaling in hepatocellular carcinoma (HCC)-MSCs via alpha(V)beta(3) integrin (avb3), which promotes tumor angiogenesis in vivo and in vitro [
203]. Moreover, the pro-angiogenic effect of GC-MSCs is mediated by the NF-κB/VEGF pathway, which may also be involved in regulation of VEGF expression in tumor cells [
204].
MSCs can additionally promote tumor vasculogenesis through generation of pericytes. For instance, PDGF-B derived from BC and PC cells promotes the transformation of MSCs into mature pericytes via interactions with neuropilin-1 (NRP-1) in MSCs [
205]. Similarly, stromal cell-derived factor (SDF)-1α and PDGF-B released by tumor cells bind CXCR4 and PDGFR-β on MSCs, respectively, resulting in differentiation into pericytes, leading to vasculogenesis and tumor recurrence [
206].
MSCs-EVs also transport angiogenic miRNA to vascular endothelial cells, which can promote angiogenesis [
207]. Other studies have reported that MSCs-EVs increase VEGF expression in tumor cells by activating ERK1/2 [
208] and NF-kB pathways [
202], driving angiogenesis and tumor progression.
The serum MIF level in patients with esophageal squamous cell carcinoma (ESCC) is associated with that of VEGF and vascular density in tumor tissue [
209]. MIF level in BC tissue is also positively correlated with IL-8 expression and tumor microvessel density (MVD) [
34]. Furthermore, co-expression of MIF and its receptor CD74 in NSCLC is associated with greater tumor angiogenesis and angiogenic CXC chemokine levels [
210]. Consistent with these findings, MIF is reported to promote tumor angiogenesis by upregulating VEGF in UVB-induced NMSC cells [
73]. Additional studies showed that exogenous MIF induces BC cells to secrete VEGF and IL-8 [
34]. Subsequently, similar results were reported in human rhabdomyosarcoma (RMS) [
211] and intestinal tumor [
212]. Importantly, inhibition of MIF expression in melanoma stroma decreases the response of tumor cells to hypoxia, suppressing VEGF expression and MVD in tumor tissue [
213]. These results support the theory that MIF facilitates tumor growth by inducing angiogenesis.
Mechanistically, overexpression of MIF in NSCLC increases the phosphorylation level of JNK, c-Jun, and subsequent activity of the transcription factor AP-1 in a CD74-dependent manner, which promotes tumor angiogenesis by increasing expression of the angiogenic factors CXCL8 and VEGF [
214]. MIF enhances tumor angiogenesis through activating the MAPK signal pathway by enhancing phosphorylation of p38-MAPK and p44/42 and expression of VEGF-C in BC cells [
215]. MIF also promotes vasculogenic mimicry (VM) formation through the CXCR4-Akt-EMT pathway in glioblastoma to support malignant tumor progression [
84].
The above studies indicate that TA-MSCs, TA-MSCs-EVs and MIF are correlated in promoting tumor angiogenesis. Both TA-MSCs and TA-MSCs-EVs can up-regulate the expression levels of angiogenic cytokines in tumor cells, MIF may play an important role in the process because it also upregulates angiogenic cytokines by binding to CD74 receptors and activating related signaling pathways.
PD-1 and PD-L1. PD-L1, one of the most important immune checkpoints, continuously inhibits the activation, proliferation and anti-tumor functions of T cells after binding PD-1, making it impossible to effectively recognize and kill tumor cells and resulting in tumor immune escape. Both PD-L1 and PD-1 are expressed in MSCs, although differences in expression levels in MSCs from different sources have been reported [
111,
216]. TA-MSCs inhibit the killing effect of T cells on tumor cells via downregulating PD-1 and upregulating PD-L1 expression in TME, leading to tumor progression. PCa-MSCs significantly enhance PD-L1 expression under stimulation of IFN-γ and TNF-α, leading to inhibition of T cells proliferation and function [
113]. Pro-inflammatory cytokines secreted by CD4
+ T cells also upregulate PD-L1 in GC-MSCs through activation of the STAT3 pathway [
217]. IL-8 secreted by GC-MSCs has been shown to upregulate PD-L1 in GC cells through the STAT3/mTOR-c-Myc axis, enhancing the cytotoxicity of CD8
+ T cells against GC cells [
218].
In addition, TGF-β and C1q contained in BC-MSCs-EVs induce upregulation of PD-L1 in MDSCs and M2 macrophages along with downregulation of PD-1 in infiltrating T cells, which maintains the immunosuppressive TME, thus overcoming the T cells-mediated anti-tumor immune response [
122].
MIF interacts with its receptor CD74 to activate the IFNγ-JAK-STAT pathway, resulting in significant upregulation of PD-L1 in melanoma cells, which aids in tumor cell escape from the immune response and maintenance of immunosuppressive TME [
219]. Data from a recent study indicate that MIF secreted by MM cells augments CD84 expression in TME, leading to upregulation of PD-L1 on MDSCs and suppression of T cells function, consequently promoting MM progression [
220]. Neutralization of MIF is reported to inhibit PD-L1 expression in colon cancer-bearing mice, thereby suppressing tumor progression [
128].
In terms of mechanism, TA-MSCs, TA-MSCs-EVs and MIF reduce the expression of PD-1 in immunosuppressive TME and PD-L1 in tumor cell and immunosuppressive cell, thereby inhibiting anti-tumor immunity and promoting tumor cell immune escape.
Fusion of MSCs and tumor cells. Multiple studies have shown that self-fusion and allogeneic fusion of tumor cells potentially give rise to the metastatic phenotype by generating widespread genetic and epigenetic diversity, which provides novel therapeutic opportunities [
221]. Early reports have demonstrated the spontaneous formation of heterotypic hybrids between MSC and BC cells in vitro with predominantly mesenchymal morphological characteristics, mixed gene expression profiles and tumorigenicity in immunodeficient mice [
222]. Although fusion of MSCs and BC cells is a rare event, hybrid cells exhibit stronger telomerase activity and proliferation ability, significant regulation of genes involved in EMT and increased expression of metastasis-associated S100A4 genes relative to parental cells. These results have been further confirmed in vivo [
223]. Polyethylene glycol (PEG) stimulates fusion of human umbilical cord MSCs (UC-MSCs) and GC cells in vitro. The hybrid cells express high levels of stemness factors OCT4, Nanog, SOX2 and Lin28, resulting in enhanced migration, proliferation, and growth of gastric xenograft tumors in vivo [
21]. Co-cultured MSCs and glioma stem cells can also fuse in vitro and these hybrid cells have been shown to promote tumor angiogenesis in vivo and ex vivo and tumorigenicity in vivo [
224].
Malignant transformation of TA-MSCs. TA-MSCs promote tumorigenesis through spontaneous or malignant transformation induced by tumor cells or TME. Numerous studies have confirmed that after malignant transformation, TA-MSCs acquire tumor cell characteristics, such as increased proliferation, invasion/migration and pro-angiogenesis abilities, suppression of cellular senescence, and upregulation of protein and mRNA levels of tumor-related markers (glial fibrillary acidic portein (GFAP), CD133, Nestin and c-Myc) [
225‐
227]. Malignant transformation of MSCs is affected by multiple factors, including microRNAs, oncogene expression, and methylation status [
225,
228,
229].
In terms of the underlying mechanisms, Vishnubalaji et al
. [
230] showed that the Lin28b/let-7 axis is involved in the malignant transformation of hBM-MSCs. Akt, STAT3 and Wnt-β-catenin pathways are additionally associated with malignant transformation of MSCs [
225,
226]. Moreover, low Rb and high c-Myc expression induce the expression of osteosarcoma-related markers, such as alkaline phosphatase (ALP), osteonectin and osteocalcin, which are related to malignant transformation of BM-MSCs into osteosarcoma (OS) cells [
225]. The OS phenotype triggered by malignant transformation of hMSCs may be linked to overexpression of the activator protein-1 (AP-1) complex [
231]. MSCs infected by Kaposi's sarcoma-associated herpesvirus (KSHV) could induce malignant transformation into Kaposi sarcoma cells by promoting mesenchymal-to-endothelial transition (MET) [
232]. Malignant transformation of MSCs is also related to hypermethylated in cancer 1 (HIC1) and Ras-association domain family member 1A (RassF1A) methylation, which leads to decreased expression of tubulin [
229].