Xiaochun Sun

Mesenchymal stem cells from adult human bone marrow differentiate into a cardiomyocyte phenotype in vitro.

A method for isolating adult human bone marrow mesenchymal stem cells (MSCs) was established, and the ability of human MSCs to differentiate into cells with characteristics of cardiomyocytes in vitro was investigated. Selected MSC surface antigens were analyzed by flow cytometry. The MSCs at Passage 2 were treated with 5-azacytidine to investigate their differentiation into cardiomyocytes. Characteristics of the Putative myogenic cells were determined by Immunohistochemistry and transmission electron and confocal microscopies. The expression of myogenic specific genes was detected by reverse transcriptase-polymerase chain reaction (RT-PCR), real-time quantitative PCR, and DNA sequencing. The MSCs were spindle-shaped with irregular processes and were respectively Positive for CD13, CD29, CD44, CD71 and negative for CD3, CD14, CD15, CD33, CD34 CD38, CD45, and HLA-DR. The myogenic cells differentiated from MSCs were positive for beta-myosin heavy chain (beta-MHC), desmin, and alpha-cardiac actin. When the pyogenic cells were stimulated with low concentration of K+ (5.0 mM), an increase in intracellular calcium fluorescence was observed. Myofilament-like structures were observed in electron micrographs of the differentiated myogenic cells. The mRNAs of beta-MHC, desmin, alpha-cardiac actin, and cardiac troponin T were highly expressed in the myogenic cells. These results indicate that 5-azacytidine can induce human MSCs to differentiate in vitro into cells with characteristics commonly attributed to cardiomyocytes. Cardiomyocytes cultured from bone marrow sources are potentially valuable for repairing injured myocardium.

Exosomes Derived From Akt-Modified Human Umbilical Cord Mesenchymal Stem Cells Improve Cardiac Regeneration and Promote Angiogenesis via Activating Platelet-Derived Growth Factor D

We have previously demonstrated the cardioprotective effects of exosomes derived from mesenchymal stem cells (MSCs). It is well known that the activation of Akt is involved in stem cell‐induced cardioprotection. In the present study, we investigated whether exosomes released from Akt‐overexpressing MSCs showed a beneficial effect on cardioprotection and angiogenesis. MSCs were collected from human umbilical cord (hucMSCs), and Akt was transfected into hucMSCs (Akt‐hucMSCs) by using an adenovirus transfection system. Exosomes were isolated from control hucMSCs (Exo) and Akt‐hucMSCs (Akt‐Exo). An acute myocardial infarction model was created by ligation of the left anterior decedent coronary artery (LAD) in rats. Various source exosomes (400 µg of protein) were infused via the tail vein immediately after LAD ligation. The cardiac function was evaluated by using echocardiography after different treatments for 1 and 5 weeks, respectively. Endothelial cell proliferation, migration, and tube‐like structure formation, as well as chick allantoic membrane assay, were used to evaluate the angiogenetic effects of Akt‐Exo. The results indicated that cardiac function was significantly improved in the animals treated with Akt‐Exo. In addition, Akt‐Exo significantly accelerated endothelial cell proliferation and migration, tube‐like structure formation in vitro, and blood vessel formation in vivo. The expression of platelet‐derived growth factor D (PDGF‐D) was significantly upregulated in Akt‐Exo. However, the angiogenesis was abrogated in endothelial cells treated with the exosomes obtained from MSCs transfected with PDGF‐D‐siRNA. Our studies suggest that exosomes obtained from Akt‐modified hucMSCs are more effective in myocardial infarction therapy through promoting angiogenesis. PDGF‐D plays an important role in Akt‐Exo‐mediated angiogenesis. Stem Cells Translational Medicine 2017;6:51–59

Mesenchymal stem cells derived from breast cancer tissue promote the proliferation and migration of the MCF‑7 cell line in vitro

Mesenchymal stem cells (MSCs) are critical in promoting cancer progression, including tumor growth and metastasis. MSCs, as a subpopulation of cells found in the tumor microenvironment, have been isolated from several tumor tissues, but have not been isolated from breast cancer tissue to date. Therefore, the purpose of this study was to isolate MSCs from primary human breast cancer tissue, and to study the effect of breast cancer MSCs (BC-MSCs) on the proliferation and migration of the MCF-7 cell line in vitro. MSCs were isolated and identified from primary breast cancer tissue obtained from 9 patients. The MCF-7 cell line was treated with 10 and 20% breast cancer-associated MSC (BC-MSC)-conditioned medium (CM) for 10–48 h, and changes in proliferation and migration were observed. Furthermore, we investigated the migration of 10 and 20% CM concentrations on MCF-7 through a scratch wound assay and a transwell migration assay. We successfully isolated and identified MSCs from primary breast cancer tissues. BC-MSCs showed characteristics similar to those of bone marrow MSCs, and possessed the capability of multipotential differentiation into osteoblasts and adipocytes. The results of the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed that 10 and 20% CM concentrations increased the proliferation of MCF-7 cells to different levels. The results also revealed a greater increase in different levels compared with the control group. In conclusion, MSCs were confirmed to exist in human breast cancer tissues, and BC-MSCs may promote the proliferation and migration of breast cancer cells.

Mesenchymal stem cell-like cells from children foreskin inhibit the growth of SGC-7901 gastric cancer cells

Mesenchymal stem cells (MSCs) become a research hotspot in recent years because of their roles in regenerative medicine and tissue injury repair. However, the limited source for MSCs hampers its clinical application. In this study, we isolated and identified human mesenchymal stem cell-like cells from foreskin (hFMSCs) by explant culture. HFMSCs had similar morphology and immunophenotype to that of human bone marrow derived-mesenchymal stem cells. HFMSCs formed colonies after 9 days of inoculation and could be propagated for more than 50 passages. HFMSCs had a normal karyotype and high G0/G1 phase independent of passage number. Further, hFMSCs could be induced to differentiate into osteocytes and adipocytes. We found that the growth of SGC-7901 (human gastric adenocarcinoma) cells could be suppressed by simultaneous injection of hFMSCs in vivo. HFMSCs also inhibited SGC-7901 cell proliferation in vitro. HFMSC co-injection resulted in a decrease in PCNA-positive and an increase in apoptotic tumor cells. HFMSCs derived conditioned medium inhibited the expression of BCL-2 while increased the expression of BAX and caspase-3 in SGC-7901 cells. Taken together, our findings suggest that children foreskin is a new source for MSCs and hFMSCs could inhibit gastric cancer cell growth both in vitro and in vivo.

Mesenchymal stem cells isolated from human uterine cervix cancer tissues

In the present study, MSCs (mesenchymal stem cells) were successfully isolated and identified from hUCC (human uterine cervix cancer) tissues. The morphological appearance, immunophenotype, growth curve, cell cycle, cytogenetic features and differentiation potential of these cells were investigated. Results showed that cells isolated from the uterine cervix cancer tissues displayed fibroblast‐like morphology and grew into colonies. Immunophenotyping by flow cytometry revealed that the isolated cells were positive for CD13, CD29, CD44, CD105 and HLA‐I, while negative for CD10, CD14, CD31, CD34, CD38 and HLA‐DR. The cells kept a normal karyotype by chromosome analysis. At the third passage, the percentages of cells in G0‐/G1‐, 2‐/M‐ and S‐phase were 84.94, 8.36 and 6.71%, respectively. Under appropriate induction conditions, these cells can differentiate into osteogenic, adipogenic cells and hepatocytes. Taken together, MSCs were confirmed to exist in hUCC tissues, which may provide a new target for clinical cancer therapy.

miR-145 inhibits proliferation and migration of breast cancer cells by directly or indirectly regulating TGF-β1 expression

Studies have demonstrated low expression of miR-145 associated with cell proliferation and migration in a wide variety of tumors. Here, we studied the expression of miR-145 in relation to the occurrence and development of breast cancer. Total RNA from breast cancer tissue and corresponding adjacent normal tissue was extracted and used to detect miR-145 expression by quantitative real-time polymerase chain reaction (qRT-PCR). We also transfected breast cancer cells with hsa-miR-145 mimics, hsa-miR-145 inhibitor, mimics negative control (mimics NC) or inhibitor negative control (inhibitor NC). Cell proliferation was analyzed by colony formation assays and methyl thiazolyl tetrazolium assays. Cell proliferation in breast cancer cells was decreased after overexpression of miR-145 and increased following miR-145 suppression. Cell migration and invasion were assessed using Transwell and wound healing assays, respectively, and were also decreased after overexpression of miR-145 and increased after miR-145 suppression in breast cancer cells. Finally, western blot assays showed that overexpression of miR-145 inhibited expression of transforming growth factor-β1 (TGF-β1). Collectively, these data suggest that miR-145 may inhibit TGF-β1 protein expression which may in turn contribute to tumor formation.

Lead acetate reduces the ability of human umbilical cord mesenchymal stem cells to support hematopoiesis in vitro

Plumbum (Pb) is a heavy metal toxin that causes many pathophysiological effects in various systems of the human body. It has previously been reported that excessive lead trioxide causes hematopoietic system toxicity. Mesenchymal stem cells (MSCs), as cells with self-renewal and multipotent differentiation potential, play a supportive role in hematopoietic function. Lead is well known to interfere with hemoglobin synthesis and affect erythrocyte morphology and survival. MSCs and the cytokines secreted by MSCs are the important components of the hematopoietic microenvironment. Thus, we hypothesized that lead may cause damage to MSCs, which may provide a new understanding of the mechanism of lead toxicity in the hematopoietic system. In the present study, cell count, MTT assay, apoptosis assay, osteogenic differentiation, cell histochemical staining, cell cycle analysis, colony forming assay and RT-PCR were used. The results showed that the proliferation of umbilical cord MSCs (UCMSCs) was affected if the concentrations of lead were higher than 10 µM. Following osteogenic differentiation, the rate of alkaline phosphatase and Von Kossa stain positivity in the experimental group was lower than that in the control group. In conclusion, these results demonstrate that lead suppresses the self-renewal and multipotent differentiation potential of UCMSCs, and induces an adverse effect on the expression of UCMSCs cytokines. MSCs also have a hematopoiesis-promoting function that is capable of supporting colony formation of bone marrow cells. Furthermore, we found that following treatment with lead acetate, the supportive function of UCMSCs on colony formation was inhibited. Taken together, lead acetate has a toxic effect on the self-renewal, multipotent differentiation potential and hematopoiesis-promoting function of UCMSCs.

Nicotine-enhanced stemness and epithelial-mesenchymal transition of human umbilical cord mesenchymal stem cells promote tumor formation and growth in nude mice

Cigarette smoking is a well-known risk factor in the development and progression of malignant diseases. Nicotine, the major constituent in cigarette smoke, has also shown negative effects on stem cells. Mesenchymal stem cells (MSCs) have been widely demonstrated to migrate into tumors and play key roles in cancer progression. However, the mechanisms by which nicotine impacts MSCs and tumorigenesis of lung cancer are still undetermined. In this study we investigated the effects of nicotine on human umbilical cord mesenchymal stem cells (hUC-MSCs) and the impacts of nicotine-treated hUC-MSCs on tumor formation and progression. We found that nicotine has a toxic effect on hUC-MSCs and changes the morphology, inhibits proliferation and promotes apoptosis of hUC-MSCs in a dose-dependent manner. Nicotine-treated hUC-MSCs produce higher level of IL-6. Moreover, nicotine promotes migration, stemness and epithelial-mesenchymal transition (EMT) of hUC-MSCs by inhibiting E-cadherin expression and upregulating mesenchymal markers such as N-cadherin and Vimentin, leading to the induction of stem cell markers Sox2, Nanog, Sall4, Oct4 and CD44. Migration and proliferation of non-small cell lung cancer A549 cells and breast cancer MCF-7 cells are promoted after their coculture with nicotine-treated hUC-MSCs in a cell-cell contact-independent manner. Furthermore, nicotine-treated hUC-MSCs promote tumor formation and growth of A549 cells in nude mice. These studies demonstrated that the enhanced stemness and EMT of hUC-MSCs induced by nicotine are critical for the development of tobacco-related cancers.

Effect of idebenone on bone marrow mesenchymal stem cells in vitro

In recent years, stem cell research has continued to benefit from its crossover with chemistry, particularly the investigation of small molecular drugs modulating specific targets to regulate stem cell fate. Idebenone (IDB) is a yellow crystalline powder that is used in the treatment of chronic cerebrovascular diseases. The objective of the present study was to examine whether IDB had an influence on bone marrow‑derived mesenchymal stem cells (BMSCs) extracted from the bone marrow of Sprague‑Dawley rats. The effects of IDB on cell proliferation, cell cloning and migration were investigated. Cell cycle, apoptosis, DAPI nuclear staining and senescence‑associated β‑galactosidase (SA‑β‑gal) staining were also examined. The results revealed that IDB at suitable concentrations enhanced cell cloning capacity, promoted the proliferation of BMSCs, delayed cellular senescence, and inhibited cell apoptosis and migration. Western blot analysis indicated that IDB increased the expression of B‑cell lymphoma 2 (Bcl‑2), signal transducer and activator of transcription‑3, Nanog, octamer‑binding transcription factor 4, E‑cadherin, proliferating cell nuclear antigen, cyclinD1 and cyclinD3, and decreased the expression of Bcl‑2‑associated X protein, cleaved caspase‑3, N‑cadherin, vimentin and α‑smooth muscle actin. In conclusion, these experiments confirmed that IDB in low doses had no toxic effect and may exert protective effects on BMSCs.