Therapeutic Potential of Adipose-Derived Stem Cell-Conditioned Medium and Extracellular Vesicles in an In Vitro Radiation-Induced Skin Injury Model.

Radiotherapy (RT) is one of three major treatments for malignant tumors, and one of its most common side effects is skin and soft tissue injury. However, the treatment of these remains challenging. Several studies have shown that mesenchymal stem cell (MSC) treatment enhances skin wound healing. In this study, we extracted human dermal fibroblasts (HDFs) and adipose-derived stem cells (ADSCs) from patients and generated an in vitro radiation-induced skin injury model with HDFs to verify the effect of conditioned medium derived from adipose-derived stem cells (ADSC-CM) and extracellular vesicles derived from adipose-derived stem cells (ADSC-EVs) on the healing of radiation-induced skin injury. The results showed that collagen synthesis was significantly increased in wounds treated with ADSC-CM or ADSC-EVs compared with the control group, which promoted the expression of collagen-related genes and suppressed the expression of inflammation-related genes. These findings indicated that treatment with ADSC-CM or ADSC-EVs suppressed inflammation and promoted extracellular matrix deposition; treatment with ADSC-EVs also promoted fibroblast proliferation. In conclusion, these results demonstrate the effectiveness of ADSC-CM and ADSC-EVs in the healing of radiation-induced skin injury.

Wharton's Jelly Mesenchymal Stem Cell-Derived Extracellular Vesicles Reduce SARS-CoV2-Induced Inflammatory Cytokines Under High Glucose and Uremic Toxin Conditions.

Cytokine storm is recognized as one of the factors contributing to organ failures and mortality in patients with COVID-19. Due to chronic inflammation, COVID-19 patients with diabetes mellitus (DM) or renal disease (RD) have more severe symptoms and higher mortality. However, the factors that contribute to severe outcomes of COVID-19 patients with DM and RD have received little attention. In an effort to investigate potential treatments for COVID-19, recent research has focused on the immunomodulation functions of mesenchymal stem cells (MSCs). In this study, the correlation between DM and RD and the severity of COVID-19 was examined by a combined approach with a meta-analysis and experimental research. The results of a systematic review and meta-analysis suggested that the odd of mortality in patients with both DM and RD was increased in comparison to those with a single comorbidity. In addition, in the experimental research, the data showed that high glucose and uremic toxins contributed to the induction of cytokine storm in human lung adenocarcinoma epithelial cells (Calu-3 cells) in response to SARS-CoV Peptide Pools. Of note, the incorporation of Wharton's jelly MSC-derived extracellular vesicles (WJ-EVs) into SARS-CoV peptide-induced Calu-3 resulted in a significant decrease in nuclear NF-κB p65 and the downregulation of the cytokine storm under high concentrations of glucose and uremic toxins. This clearly suggests the potential for WJ-EVs to reduce cytokine storm reactions in patients with both chronic inflammation diseases and viral infection.

Rejuvenation of mesenchymal stem cells by extracellular vesicles inhibits the elevation of reactive oxygen species.

Aging induces numerous cellular disorders, such as the elevation of reactive oxygen species (ROS), in a number type of cells, including mesenchymal stem cells (MSCs). However, the correlation of ROS and impaired healing abilities as well as whether or not the inhibition of elevating ROS results in the rejuvenation of elderly MSCs is unclear. The rejuvenation of aged MSCs has thus recently received attention in the field of regenerative medicine. Specifically, extracellular vesicles (EVs) act as a novel tool for stem cell rejuvenation due to their gene transfer ability with systemic effects and safety. In the present study, we examined the roles of aging-associated ROS in the function and rejuvenation of elderly MSCs by infant EVs. The data clearly showed that elderly MSCs exhibited the downregulation of superoxide dismutase (SOD)1 and SOD3, which resulted in the elevation of ROS and downregulation of the MEK/ERK pathways, which are involved in the impairment of the MSCs' ability to decrease necrotic area in the skin flap model. Furthermore, treatment with the antioxidant Edaravone or co-overexpression of SOD1 and SOD3 rescued elderly MSCs from the elevation of ROS and cellular senescence, thereby improving their functions. Of note, infant MSC-derived EVs rejuvenated elderly MSCs by inhibiting ROS production and the acceleration of cellular senescence and promoting the proliferation and in vivo functions in both type 1 and type 2 diabetic mice.

Type 2 Diabetes Mellitus Induced Paracrine Effects on Breast Cancer Metastasis Through Extracellular Vesicles Derived from Human Mesenchymal Stem Cells.

Cancer metastasis is the leading cause of mortality among breast cancer patients. Type 2 diabetes mellitus (T2DM) has been suggested as a risk factor of breast cancer; however, whether or not T2DM is associated with breast tumor metastasis remains unclear. In this study, we examined the involvement of T2DM with breast cancer metastasis by a combined approach of a meta-analysis and experimental research. The results of a systematic review and meta-analysis suggested that diabetes significantly increases the risk of lymph node metastasis by 1.10-fold (P < 0.01). Consistently, our data from experimental research showed that T2DM induced paracrine effects of mesenchymal stem cells (MSCs), a key contributor to cancer progression, to stimulate metastasis of breast cancer cells (BCCs) by two independent mechanisms. First, T2DM induced the excess secretion of interleukin 6 (IL6) from MSCs, which activated the JAK/STAT3 pathway in BCCs, thus promoting the metastasis of BCCs. Second, beside the EGR-1-/IL6-dependent mechanism, T2DM altered the functions of MSC-derived extracellular vesicles (EVs), which are highly associated with the metastasis of BCCs. Our present study showed that T2DM is a risk factor for breast cancer metastasis, and MSC-derived EVs might be useful for developing a novel anti-breast cancer therapy strategy.

Dynamic Changes of Mouse Embryonic Stem Cell-Derived Neural Stem Cells Under In Vitro Prolonged Culture and Hypoxic Conditions.

Embryonic stem (ES) cells have been utilized as an excellent model for the study of neural development. However, the dynamic changes of ES cell-derived neural stem cells (ES-NSCs), under the effects of prolonged cell culture and hypoxic conditions, are still obscured. In the present study, using the combination of serum-free culture of embryoid body-like aggregates (SFEB) culture and cell sorting by Sox-1, the ES-NSCs were easily isolated and showed in vitro temporal neural specification, which resulted in distinct cell fates after neural differentiation. Early passaged ES-NSCs gave rise to neurons, whereas late-passaged ES-NSCs gave rise to glial cells, similar to the in vivo dynamic changes during the neural development. Remarkably, hypoxia treatment induced the neural differentiation of ES-NSCs but did not affect the cell fate. Under hypoxic conditions, early passaged ES-NSCs showed the upregulation of neuronal markers, whereas late-passaged ES-NSCs showed the upregulation of a glial marker. In addition, the knockdown of the hypoxia-inducible factor 1α expression impaired the neuronal differentiation of early passaged ES-NSCs under hypoxic conditions. These data demonstrated the distinct effects of prolonged culture and hypoxic stimuli on the neural differentiation of ES-NSCs; prolonged culture was involved in the cell fate after neural differentiation, while hypoxia treatment efficiently promoted neural differentiation.

Glucocorticoid Impaired the Wound Healing Ability of Endothelial Progenitor Cells by Reducing the Expression of CXCR4 in the PGE2 Pathway.

Background: Endothelial progenitor cells (EPCs) can be used to treat ischemic disease in cell-based therapy owing to their neovascularization potential. Glucocorticoids (GCs) have been widely used as strong anti-inflammatory reagents. However, despite their beneficial effects, side effects, such as impairing wound healing are commonly reported with GC-based therapy, and the effects of GC therapy on the wound healing function of EPCs are unclear. Methods: In this study, we investigated how GC treatment affects the characteristics and wound healing function of EPCs. Results: We found that GC treatment reduced the proliferative ability of EPCs. In addition, the expression of CXCR4 was dramatically impaired, which suppressed the migration of EPCs. A transplantation study in a flap mouse model revealed that GC-treated EPCs showed a poor homing ability to injured sites and a low activity for recruiting inflammatory cells, which led to wound healing dysfunction. Impairment of prostaglandin E2 (PGE2) synthases, cyclooxygenase (COX2) and microsomal PGE2 synthase 1 (mPEGS1) were identified as being involved in the GC-induced impairment of the CXCR4 expression in EPCs. Treatment with PGE2 rescued the expression of CXCR4 and restored the migration ability of GC-treated EPCs. In addition, the PGE2 signal that activated the PI3K/AKT pathway was identified to be involved in the regulation of CXCR4 in EPCs under the effects of GCs. In addition, similar negative effects of GCs were observed in EPCs under hypoxic conditions. Under hypoxic conditions, GCs independently impaired the PGE2 and HIF2α pathways, which downregulated the expression of CXCR4 in EPCs. Our findings highlighted the influences of GCs on the characteristics and functions of EPCs, suggesting that the use of EPCs for autologous cell transplantation in patients who have used GCs for a long time should be considered carefully.

Aging impairs beige adipocyte differentiation of mesenchymal stem cells via the reduced expression of Sirtuin 1.

In the body, different types of adipose tissue perform different functions, with brown and beige adipose tissues playing unique roles in dissipating energy. Throughout life, adipocytes are regenerated from progenitors, and this process is impaired by aging. One of the progenitors of adipocytes are mesenchymal stem cells (MSCs), which have recently become a promising tool for stem cell therapy. However, whether or not aging impairs the brown/beige adipocyte differentiation of adipose tissue-derived MSCs (AT-MSCs) remains unclear. In the present study, we isolated AT-MSCs from two different age groups of donors (infants and elderly subjects) and examined the effects of aging on the AT-MSC brown/beige adipocyte differentiation ability. We found that none of the AT-MSCs expressed Myf5, which indicated the beige (not brown) differentiation ability of cells. Of note, an inverse correlation was noted between the beige adipocyte differentiation ability and age, with AT-MSCs derived from elderly donors showed the most severely reduced function due to induced cellular senescence. The impaired expression of Sirtuin 1 (Sirt1) and Sirt3 proved to be responsible for the induction of senescence in elderly AT-MSCs; however, only Sirt1 was directly involved in the regulation of beige adipocyte differentiation. The overexpression of Sirt1 impaired the p53/p21 pathway, thereby preventing elderly AT-MSCs from entering senescence and restoring the beige differentiation ability. Thus, our study represents the important role of Sirt1 and senescence in the regulation of beige adipocyte differentiation during aging.

Elevated Expression of Dkk-1 by Glucocorticoid Treatment Impairs Bone Regenerative Capacity of Adipose Tissue-Derived Mesenchymal Stem Cells.

Glucocorticoids are steroid hormones used as anti-inflammatory treatments. However, this strong immunomodulation causes undesirable side effects that impair bones, such as osteoporosis. Glucocorticoid therapy is a major risk factor for developing steroid-induced osteonecrosis of the femur head (ONFH). Since ONFH is incurable, therapy with mesenchymal stem cells (MSCs) that can differentiate into osteoblasts are a first-line choice. Bone marrow-derived MSCs (BM-MSCs) are often used as a source of stem cell therapy for ONFH, but their proliferative activity is impaired after steroid treatment. Adipose tissue-derived MSCs (AT-MSCs) may be an attractive alternative source; however, it is unknown whether AT-MSCs from steroid-induced ONFH (sAT-MSCs) have the same differentiation ability as BM-MSCs or normal AT-MSCs (nAT-MSCs). In this study, we demonstrate that nAT-MSCs chronically exposed to glucocorticoids show lower alkaline phosphatase activity leading to reduced osteogenic differentiation ability. This impaired osteogenesis is mediated by high expression of Dickkopf1 (Dkk-1) that inhibits wnt/ß-catenin signaling. Increased Dkk-1 also causes impaired osteogenesis along with reductions in bone regenerative capacity in sAT-MSCs. Of note, plasma Dkk-1 levels are elevated in steroid-induced ONFH patients. Collectively, our findings suggest that glucocorticoid-induced expression of Dkk-1 could be a key factor in modulating the differentiation ability of MSCs used for ONFH and other stem cell therapies.

SDF-1 improves wound healing ability of glucocorticoid-treated adipose tissue-derived mesenchymal stem cells.

Glucocorticoids cause the delayed wound healing by suppressing inflammation that is required for wound healing process. Adipose tissue-derived mesenchymal stem cells (AT-MSCs) play an important role for wound healing by their cytokine productions including stromal derived factor 1 (SDF-1). However, it has not been clear how glucocorticoids affect the wound healing ability of AT-MSCs. In this study, we found that glucocorticoid downregulated SDF-1 expression in AT-MSCs. In addition, glucocorticoid-treated AT-MSCs induced less migration of inflammatory cells and impaired wound healing capacity compared with glucocorticoid-untreated AT-MSCs. Of note, prostaglandin E2 (PGE2) synthesis-related gene expression was downregulated by glucocorticoid and PGE2 treatment rescued not only SDF-1 expression in the presence of glucocorticoid but also their wound healing capacity in vivo. Furthermore, we found SDF-1-overexpressed AT-MSCs restored wound healing capacity even after treatment of glucocorticoid. Consistent with the results obtained from glucocorticoid-treated AT-MSCs, we found that AT-MSCs isolated from steroidal osteonecrosis donors (sAT-MSCs) who received chronic glucocorticoid therapy showed less SDF-1 expression and impaired wound healing capacity compared with traumatic osteonecrosis donor-derived AT-MSCs (nAT-MSCs). Moreover, the SDF-1 level was also reduced in plasma derived from steroidal osteonecrosis donors compared with traumatic osteonecrosis donors. These results provide the evidence that concomitant application of AT-MSCs with glucocorticoid shows impaired biological modulatory effects that induce impaired wound healing.

Uremic Toxins Affect the Imbalance of Redox State and Overexpression of Prolyl Hydroxylase 2 in Human Adipose Tissue-Derived Mesenchymal Stem Cells Involved in Wound Healing.

Chronic kidney disease (CKD) results in a delay in wound healing because of its complications such as uremia, anemia, and fluid overload. Mesenchymal stem cells (MSCs) are considered to be a candidate for wound healing because of the ability to recruit many types of cells. However, it is still unclear whether the CKD-adipose tissue-derived MSCs (CKD-AT-MSCs) have the same function in wound healing as healthy donor-derived normal AT-MSCs (nAT-MSCs). In this study, we found that uremic toxins induced elevated reactive oxygen species (ROS) expression in nAT-MSCs, resulting in the reduced expression of hypoxia-inducible factor-1α (HIF-1α) under hypoxic conditions. Consistent with the uremic-treated AT-MSCs, there was a definite imbalance of redox state and high expression of ROS in CKD-AT-MSCs isolated from early-stage CKD patients. In addition, a transplantation study clearly revealed that nAT-MSCs promoted the recruitment of inflammatory cells and recovery from ischemia in the mouse flap model, whereas CKD-AT-MSCs had defective functions and the wound healing process was delayed. Of note, the expression of prolyl hydroxylase domain 2 (PHD2) is selectively increased in CKD-AT-MSCs and its inhibition can restore the expression of HIF-1α and the wound healing function of CKD-AT-MSCs. These results indicate that more studies about the functions of MSCs from CKD patients are required before they can be applied in the clinical setting.