*4.6. MSCs in Radiation-Induced Heart Injury*

Apart from the lung, thoracic irradiation also induces heart injury, namely, radiationinduced heart disease (RIHD). RIHDs, such as myocardial, coronary artery, pericardial, valvular, and conduction system diseases, have been observed in breast cancer and Hodgkin's lymphoma patients [132,133]. These manifestations had a 50% cumulative incidence during 40 years of follow-up in an epidemiological study [132]. RIHD often involves vascular endothelial dysfunction [134], hypertrophy [135], and fibrosis [136]. The

underlying mechanisms of RIHD remain mostly indistinct, but the roles of DNA damage, inflammation, oxidative stress, and epigenetic regulation in RIHD have been well illustrated. For the treatment of RIHD, conventional statins and angiotensin-converting enzyme inhibitors are still the first-chosen drugs clinically. With increasing interest in MSCs regeneration therapy, scientists are paying attention to the application of MSCs in RIHD. Vascular injury is the most common feature of RIHD. BM-MSCs (1 <sup>×</sup> <sup>10</sup>6/kg) transplantation via tail vein can attenuate radiation-induced artery inflammation and oxidative stress [137]. The repair effect was attributed to the modulation of a series of cytokines and the differentiation potential of MSCs into endothelial cells facilitating vascular regeneration [138]. Additionally, vascular injury is usually accompanied by myocardial fibrosis and cardiac remodeling. Encouragingly, in a RIHD rat model, BM-MSCs (1.5 <sup>×</sup> <sup>10</sup><sup>6</sup> ) transplantation via caudal vein improved myocardial fibrosis and inflammation, which were related to DNA repair and downregulated PPAR-α, PPAR-γ, TGF-β, IL-6, and IL-8 [139]. As mentioned above, MSCs-CM is beneficial to radiation injury repair owing to the paracrine effect. Chen et al. assessed the therapeutic effect of human UC-MSCs-CM on radiation-induced myocardial fibrosis. They found that irradiated human cardiac fibroblasts cultured with UC-MSCs-CM showed greater viability [140]. Inhibited NF-κB activity decreased expression of several pro-fibrotic cytokines, including TGF-β1, IL-6, and IL-8, followed by mitigated collagen deposition and fibrosis [140]. Meanwhile, changes in oxidation markers (malondialdehyde) and antioxidant enzyme levels reflected reduced oxidative stress [140]. However, specific nutritional factors released by MSCs and involved in myocardial protection from IR were not clarified [140]. Thus far, there are few MSCs therapy attempts to manage RIHDs, and abundant evidence is lacking for proving its efficacy. The data on myocardial regeneration suggest that the MSCs therapy is potentially therapeutic to treat RIHD.

#### **5. Challenges and Future Perspectives of MSCs Therapy**

Although the MSCs have powerful tissue repair capacity due to their paracrine and immunomodulation activity, huge barriers hinder their clinical application. Here, we will focus on safety and efficacy, the two most concerning aspects.

Currently, the relationship between MSCs and tumor has been attracting increased attention. The tumor consists of many types of cells involving a complex pathological environment. Cancer stem cell (CSC) is a kind of multipotent stem cell with great selfrenew and differentiation capability in the tumor tissue. Like normal stem cells in the body, CSC is also indispensable for supporting tumor progression, inducing tumorigenesis, maintaining tumor growth, and promoting metastasis [141]. The tumor involves a chronic inflammatory process that recruits endogenous or exogenous MSCs [142,143]. Homed MSCs promote angiogenesis [144] and interact with CSC enhancing the growth [145] and chemoresistance [146] of CSC. The tumor exploits MSCs' unique immunosuppression nature, allowing malignant cells to escape recognition and clearance by the immune system [147–149]. It is reported that once exposed to the tumor microenvironment, MSCs would be reprogrammed and become "allies" of tumor cells, accelerating tumor progress, and invading surrounding normal tissue [149–151]. Interestingly, Chen et al. found the engulfment of stromal cells by cancer cells in human breast tumors, and these engulfing breast cancer cells exhibited gene features of MSCs [152]. However, contradictory outcomes about the cancer-promoting effect of MSCs were presented in other studies [153]. For example, several groups found that co-cultured MSCs inhibited melanoma growth by inducing cell apoptosis [154,155]. Colorectal cancer progression could also be attenuated through the intravenous injection of BM-MSCs (1 <sup>×</sup> <sup>10</sup><sup>7</sup> ) [156]. The bidirectional effects of MSCs on tumor development motivate scientists to ascertain more precise mechanisms underlying MSCs and tumor tissue interaction. Unfortunately, it seems that the protumorigenic effect is dominant due to more substantial preclinical evidence. Therefore, MSCs-based therapy must be performed with great caution in clinics, especially with regards to radiation injury patients with malignancy history.

On the premise that security can be guaranteed, investigators need to seek appropriate protocols by which MSCs therapy remedy would maximize radiation repair efficiency. Many questions need to be discussed, for example, how do we determine the selection of the MSC population considering heterogeneity? In addition, the most effective delivery dose and pattern are required to ensure a high retention rate and therapy efficacy. Indeed, different organizational origins give rise to MSCs heterogeneity reflected by diversities of proliferation and differentiation capability, paracrine potential, and immunomodulatory effect [46–48]. Despite the minimal criteria mentioned above, it is difficult to sort out homogeneous MSCs. Apart from shared surface CD antigens, there are no additional markers to identify each type of tissue-derived MSCs [157]. Such heterogeneity can lead to the deviation of actual results from expectation and become a significant obstacle to selecting MSCs for clinical usage [158]. Because of the heterogeneity, each MSC population may have distinct therapeutic effects on the same tissue injury. It is necessary to search for the most potent MSC population for radiation injury of a specific tissue. On the other hand, different laboratories have their respective protocols of MSCs isolation, culture, and expansion procedures, causing MSCs heterogeneity and the following difference in quality. Therefore, MSCs management system should be standardized as much as possible. This can reduce heterogeneity caused by different treating conditions and increased comparability among different research results, thus providing valuable clinical guidance of MSCs application. Apart from heterogeneity, the effective dose range and cell delivery route must be emphasized and discussed. A dose gradient experiment of MSCs therapy in radiation injury models should be carried out to find both safe and efficient dose range [159]. In a study of radiation-induced artery injury, a high dose of BM-MSCs (1 <sup>×</sup> <sup>10</sup>7/kg) showed greater therapeutic potential in irradiated mice than a low dose of BM-MSCs (1 <sup>×</sup> <sup>10</sup>6/kg) [137]. Additionally, different injection patterns, including whole-body infusion via a vein or local interventional injection, will affect the homing of MSCs to injured sites [160]. Thus far, our understandings of the therapeutic effect of MSCs in mitigating radiation injury and the underlying mechanism are basically from preclinical trials. The transition of MSCs administration from animal to clinical studies still requires lots of effort.
