BACKGROUNDBacterial lipopolysaccharide (LPS) can activate immunological cells to secrete various proinflammatory cytokines involved in the pathophysiological process of disseminated intravascular coagulation (DIC) during infection. In recent years, it has been found that bone marrow-derived mesenchymal stem cells (BMSCs) can affect the activity of these immune cells and regulate the secretion of proinflammatory cytokines. Here, we report the possible protective effect of BMSCs pre-treatment in LPS-induced DIC rat model and the mechanism.

METHODSForty-eight adult male rats were divided into five experimental groups and one control group with eight animals in each group. In the treatment groups, 0, 1'10(6), 2'10(6), 3'10(6), and 5'10(6) of BMSCs were injected intravenously for 3 days before LPS injection, while the control group was treated with pure cell culture medium injection. Then, the LPS (3 mg/kg) was injected via the tail vein in the treatment groups, while the control group received 0.9% NaCl. Blood was withdrawn before and 4 and 8 hours after LPS administration. The following parameters were monitored: platelets (PLT), fibrinogen (Fib), D-dimer (D-D), activated partial thromboplastin time (APTT), prothrombin time (PT), tumor necrosis factor-a (TNF-a), interferon-g (IFN-g), interleukin-1b (IL-1b), creatinine (Cr), alanine aminotransferase (ALT), creatinine kinase-MB (CK-MB), and endothelin (ET).

RESULTSCompared with the control group, a significant change of coagulation parameters were found in the experimental groups. The plasma level of the inflammatory mediator (TNF-a, IFN-g, IL-1b), organ indicator (Cr, ALT, and CK-MB), and ET in the experimental groups were much lower (P < 0.05) than that in the control group. Furthermore, some of these effects were dose-dependent; the statistical comparison of the plasma levels between the groups (from group 2 to group 5) showed a significant difference (P < 0.05), except the ALT and CK-MB levels (P > 0.05).

CONCLUSIONPre-treatment with BMSCs can attenuate organ dysfunction and inhibit systemic intravascular coagulation effectively via the regulatory effect on immune cells and proinflammatory cytokines in LPS-induced DIC rat model.

Alanine Transaminase ; metabolism ; Animals ; Blood Coagulation ; drug effects ; Bone Marrow Cells ; cytology ; Creatinine ; metabolism ; Interferon-gamma ; metabolism ; Interleukin-1beta ; metabolism ; Lipopolysaccharides ; pharmacology ; Male ; Mesenchymal Stromal Cells ; cytology ; physiology ; Rats ; Rats, Wistar ; Tumor Necrosis Factor-alpha ; metabolism

The use of mesenchymal stem cells (MSCs) has emerged as a potential new treatment for myocardial infarction. However, the poor viability of MSCs after transplantation critically limits the efficacy of this new strategy. The expression of microRNA-210 (miR-210) is induced by hypoxia and is important for cell survival under hypoxic conditions. Hypoxia increases the levels of hypoxia inducible factor-1 (HIF-1) protein and miR-210 in human MSCs (hMSCs). miR-210 positively regulates HIF-1alpha activity. Furthermore, miR-210 expression is also induced by hypoxia through the regulation of HIF-1alpha. To investigate the effect of miR-210 on hMSC survival under hypoxic conditions, survival rates along with signaling related to cell survival were evaluated in hMSCs over-expressing miR-210 or ones that lacked HIF-1alpha expression. Elevated miR-210 expression increased survival rates along with Akt and ERK activity in hMSCs with hypoxia. These data demonstrated that a positive feedback loop involving miR-210 and HIF-1alpha was important for MSC survival under hypoxic conditions.
Cell Survival ; Cobalt ; Gene Expression Regulation/*physiology ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit/genetics/*metabolism ; Mesenchymal Stromal Cells/drug effects/metabolism/*physiology ; MicroRNAs/*metabolism ; Oxygen/pharmacology ; *Oxygen Consumption ; RNA, Small Interfering/metabolism

BACKGROUNDAcute lung injury (ALI) and end-stage acute respiratory distress syndrome (ARDS) were among the most common causes of death in intensive care units. The activation of an inflammatory response and the damage of pulmonary epithelium and endotheliumwerethe hallmark of ALI/ARDS. Recent studies had demonstrated the importance of mesenchymal stem cells (MSCs) in maintaining the normal pulmonary endothelial and epithelial function as well as participating in modulating the inflammatory response and they are involved in epithelial and endothelial repair after injury. Here, our study demonstrates MSCs therapeutic potential in a rat model of ALI/ARDS.

METHODSBone marrow derived MSCs were obtained from Sprague-Dawley (SD) rats and their differential potential was verified. ALI was induced in rats byoleic acid (OA), and MSCs were transplanted intravenously. The lung injury and the concentration of cytokines in plasma and lung tissue extracts were assessed at 8 hours, 24 hours and 48 hours after OA-injection.

RESULTSThe histological appearance and water content in rat lung tissue were significantly improved at different time points in rats treated with MSCs. The concentration of tumor necrosis factor-a and intercellular adhesion molecular-1 in rats plasma and lung tissue extracts were significantly inhibited after intravenous transplantation of MSCs, whereas interleukin-10 was significantly higher after MSCs transplantation at 8 hours, 24 hours and 48 hours after OA-challenge.

CONCLUSIONSIntravenous transplantation of MSCs could maintain the integrity of the pulmonary alveolar-capillary barrier and modulate the inflammatory response to attenuate the experimental ALI/ARDS. Transplantation of MSCs could be a novel cell-based therapeutic strategy for prevention and treatment of ALI/ARDS.

Acute Lung Injury ; chemically induced ; metabolism ; pathology ; therapy ; Animals ; Cell Differentiation ; drug effects ; Interleukin-10 ; metabolism ; Male ; Mesenchymal Stromal Cells ; cytology ; drug effects ; physiology ; Oleic Acid ; toxicity ; Rats ; Rats, Sprague-Dawley ; Tumor Necrosis Factor-alpha ; metabolism

BACKGROUNDBone marrow hematopoietic function suppression is one of the most common side effects of chemotherapy. After chemotherapy, the bone marrow structure gets destroyed and the cells died, which might cause the hematopoietic function suppression. Heme oxygenase-1 (HO-1) is a key enzyme of antioxidative metabolism that associates with cell proliferation and resistance to apoptosis. The aim of this study was to restore or resist the bone marrow from the damage of chemotherapy by the HO-1 expression of mouse mesenchymal stem cells (mMSCs) homing to the mice which had the chemotherapy-induced bone marrow suppression.

METHODSOne hundred and sixty female Balb/c mice (6-8-weeks old) were randomly divided into four groups. Each group was performed in 40 mice. The control group was intraperitoneally injected for 5 days and tail intravenously injected on the 6th day with normal saline. The chemotherapy-induced bone marrow suppression was established by intraperitoneally injecting cyclophosphamide (CTX) into the mice which performed as the chemotherapy group. The mMSCs were tail intravenously injected into 40 chemotherapically damaged mice which served as the mMSCs group. The difference between the HO-1 group and the mMSCs group was the injected cells. The HO-1 group was tail intravenously injected into the mMSCs that highly expressed HO-1 which was stimulated by hemin. The expression of HO-1 was analyzed by Western blotting and RT-PCR. Cell proliferation was measured using the 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay. Histopathologic examinations were performed 1 week after injection.

RESULTSCompared with the control group, the expression levels of HO-1 mRNA and protein were significantly higher in the HO-1 group (all P < 0.05), even obviously than the mMSCs group. CTX treatment induced apoptosis and inhibited proliferation. After injected, the white blood cell (WBC), red blood cell (RBC) and platelet (PLT) declined fast and down to the bottom at the 7th day. The bone marrow structure was destroyed incomplete. In vitro, the survival rate of cells in chemotherapy group was less than 50% after 24 hours. In contrast, mMSCs could do a favor to the cellular cleavage and proliferation. They slowed down the cell mortality and more than 50% cells survived after 24 hours. The effects of blocking apoptosis and bone marrow recovery could be more effective in the HO-1 group. In the HO-1 group, it had observed that the bone marrow structure became complete and the hemogram closed to normal at 7th day.

CONCLUSIONSHO-1 played an important role in promoting the recovery of CTX-induced hematopoietic damage. We suggest that HO-1 is able to restore the functions of chemotherapy-induced hematopoietic damage.

Animals ; Apoptosis ; drug effects ; Blood Platelets ; drug effects ; Blotting, Western ; Bone Marrow ; drug effects ; enzymology ; Cell Proliferation ; drug effects ; Cells, Cultured ; Cyclophosphamide ; toxicity ; Erythrocytes ; drug effects ; Female ; Heme Oxygenase-1 ; genetics ; metabolism ; Leukocytes ; drug effects ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stromal Cells ; enzymology ; physiology ; Mice ; Mice, Inbred BALB C ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo explore the mechanism via which the epidermal growth factor (EGF) affects the migration of human amnion-derived mesenchymal stem cells (hAMSCs).

METHODSIn vitro cultured hAMSCs were divided into control (untreated), EGF group, inhibitor AG1478 + EGF group, inhibitor LY294002 + EGF group, and inhibitor U0126 + EGF group. The migration ability of hAMSCs in each group was measured using Transwell chamber. The expressions of phosphorylated EGFR (P-EGFR), phosphorylated AKT (P-AKT), and phosphorylated ERK1/2 (P-ERK1/2) as well as the expressions of metalloproteinase (MMP) -2 and MMP-9 were detected using Western blot analysis. The differentially expressed genes in the culture solutions in EGF groups and control group were analyzed with RNA-Seq technique.

RESULTSCells in EGF group had significantly stronger migration ability than in control group (P = 0.0361), inhibitor AG1478 + EGF group (P = 0.0113), inhibitor LY294002 + EGF group (P = 0.0169), and inhibitor U0126 + EGF group (P = 0.0293). EGF increased the phosphorylation levels of EGFR, AKT and ERK, and increased the expression of MMP-2. However, the increased expressions of P-AKT and P-ERK could be suppressed by AG1478 and LY294002. As shown by GO functional enrichment analysis and KEGG pathway analysis, EGF increased the transcription of genes, which were mainly involved in transcriptional regulation, protein modification, and apoptosis inhibition. Genes that were involved in the MARK pathway included DUSP5, IL1B, DUSP6, NGF, and HSPA2.

CONCLUSIONEGF-induced migration of hAMSCs may be mediated by the signaling pathways of PI3K and ERK, which needs MMP-2 expression and the co-expression of genes involved in transcriptional regulation, protein modification, and apoptosis inhibition.

Amnion ; cytology ; Cell Movement ; drug effects ; Cells, Cultured ; Epidermal Growth Factor ; pharmacology ; Humans ; Matrix Metalloproteinase 2 ; metabolism ; Matrix Metalloproteinase 9 ; metabolism ; Mesenchymal Stromal Cells ; metabolism ; physiology ; Proto-Oncogene Proteins c-akt ; metabolism

OBJECTIVETo investigate the effects of B7H4 on human bone marrow mesenchymal stem cells (HBMSC) mediating immune suppression.

METHODSThe expression of the negative immunoregulatory factor B7H4 on HBMSC were analyzed by RT-PCR and flow cytometry (FCM), respectively. The blocking experiment was used to detect the effects of B7H4 on HBMSC mediating suppression on PHA induced T cell activation, proliferation and cell cycle. HBMSC inhibiting T cell proliferation was examined by transwell cell culture system.

RESULTSB7H4 was highly expressed on HBMSC. Blocking the B7H4 expression by B7H4mAb significantly attenuated the inhibitory effects of HBMSC on T cell proliferation. Compared with that of the unblocking group, T cell stimulator index (SI) of the B7H4 blocked group was significantly increased (53 +/- 5 vs 15 +/- 8, P < 0.01) and the inhibitory effects of HBMSC on T cell cycle were weakened significantly through down-regulating the cell number in G(0)/G(1) phase \[(85.6 +/- 9.9)% vs (95.8 +/- 9.9)%\] and up-regulating those in S phase\[(5.8 +/- 3.2)% vs (2.3 +/- 2.2)%, P < 0.05\]. The suppressive effects of HBMSC on T cell proliferation were significantly weakened after separating HBMSC from T cells by transwell cell culture system. Compared with the cell to cell contact group, T cell SI was significantly increased (27 +/- 17 vs 15 +/- 3, P < 0.01).

CONCLUSIONHBMSC highly express B7H4, which plays an important role in the suppressive effects of HBMSC on T cell proliferation.

B7-1 Antigen ; metabolism ; physiology ; Bone Marrow Cells ; immunology ; metabolism ; Cell Cycle ; immunology ; Cell Proliferation ; Cells, Cultured ; Humans ; Lymphocyte Activation ; drug effects ; immunology ; Mesenchymal Stromal Cells ; immunology ; metabolism ; Phytohemagglutinins ; pharmacology ; T-Lymphocytes ; cytology ; drug effects ; immunology ; V-Set Domain-Containing T-Cell Activation Inhibitor 1

OBJECTIVETo evaluate whether human mesenchymal stem cells (hMSCs) administration alter the clinical course of hyperoxia-induced lung injury.

METHODShMSCs were obtained from bone marrow aspirates from healthy donors after informed consent was signed, hMSCs were separated, cultured, amplified, identified and labeled with BrdU. For BrdU labeling, a sterile stock solution was added to the culture medium 48 h before the end of culture, at a final concentration of 10 micromol/L. Thirty-two 3-day old SD rats from four litters were randomly divided into four groups, as hyperoxia exposed + hMSC group (A), air-exposed + hMSC group (B), hyperoxia exposed group (C), and air-exposed group (D). The rats from the group A and the group C were placed in a sealed Plexiglas chamber with a minimal in- and outflow, providing six to seven exchanges per hour of the chamber volume and maintaining O2 levels above 95%, while the rats in the group B and the group D were only exposed to room air. Seven days later, all of them were taken out of the chamber, rats in the group A and B were injected intraperitoneally with hMSCs (1 x 10(5) in 50 microl of PBS) immediately, while the rats in the group C and D were only treated with 50 microl of PBS 3 days later. All the animals were sacrificed by an injection of sodium pentobarbital (120 mg/kg), perfused with cold 0.9% NaCl, and the left lungs were removed, the upper lobes of which were ground as tissue homogenates and used for ELISA, while the inferior lobes were stored at -70 degrees C until use for RT-PCR. The right lungs were fixed in situ for 2 h by the intratracheal instillation with 10% neutral formalin and then postfixed for 24 h. Sagittal sections (4-microm) of paraffin-embedded middle lobe and upper lobe of the right lung were used for immunohistochemistry and histology, respectively.

RESULTS(1) There was a significant difference in the value of RAC (raditive alveoli coant) among the 4 groups (11.145 +/- 1.331, 13.941 +/- 0.985, 9.595 +/- 0.672, 14.819 +/- 1.080, F = 43.234, P = 0.000). RAC in group A and C were significantly reduced compared with subjects in group D (P < 0.05, P < 0.05); and there was also a significant difference between group A and group C (P < 0.05), but not between group B and D subjects (P > 0.05). (2) There were significant differences in the levels of both TNFalpha and TGFbeta(1) in the homogenate of lungs among the 4 groups (142.933 +/- 24.017, 79.033 +/- 11.573, 224.088 +/- 41.915, 76.500 +/- 10.373, F = 59.970, P = 0.000; 1726.484 +/- 91.086, 1530.359 +/- 173.441, 2047.717 +/- 152.057, 1515.777 +/- 131.049, F = 24.977, P = 0.000). The levels of TNFalpha and TGFbeta1 were significantly elevated in both group A and group C when compared with subjects in group D (P < 0.05 for both). Concentrations of TNFalpha and TGFbeta1 were both significantly decreased in group A versus group C (P < 0.05 for both). There was no significant difference between group B and D subjects in the fields of TNFalpha and TGFbeta(1) (P > 0.05 for both). (3) BrdU-labelled cells were observed at alveolar wall and bronchioles in both group A and group B, and there was a significant difference in BrdU-labeled cells between two groups (0.230 +/- 0.026, 0.190 +/- 0.015; t = 3.769, P = 0.002), but none was found in group C and group D. Electrophoresis of the PCR products showed a 224 bp band, specific for Alu mRNA, in 7 of 8 rats of group A and 5 of 8 rats of group B, respectively, but no such band was found in group C and group D.

CONCLUSIONhMSCs administered by intraperitoneal injection could be implanted in the lungs of newborn rats, and they could effectively protect the rats against damage to the lungs caused by hyperoxia.

Animals ; Animals, Newborn ; Bone Marrow Cells ; drug effects ; Bromodeoxyuridine ; pharmacology ; Cell Communication ; Cell Differentiation ; drug effects ; Cells, Cultured ; Hematopoietic Stem Cells ; Humans ; Hyperoxia ; metabolism ; Infant, Newborn ; Lung ; pathology ; Lung Injury ; pathology ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stromal Cells ; drug effects ; physiology ; Oxygen ; metabolism ; Pulmonary Alveoli ; pathology ; Rats ; Rats, Sprague-Dawley ; Reverse Transcriptase Polymerase Chain Reaction ; Transforming Growth Factor beta ; analysis ; Tumor Necrosis Factor-alpha ; analysis

Midbrain dopamine (DA) neurons play an essential role in modulating motor control. Defects in central DA neurons affect a wide range of neurological disorders including Parkinson's disease (PD). The greatest motivation in the field has been the potential use of DA neurons for cell transplantation therapy in Parkinsonian patients. Recent studies indicated that BMSCs could differentiate into DA neurons in vitro as neural stem cells (NSC) and embryonic stem cells (ESC) could. However, there are no direct evidences about functional DA neurons derived from BMSCs. According to the protocols which had been applicated in inducing neuronal stem cells and embryonic stem cells differentiate into DA neurons in vitro, the present study provides a protocol by using 50 micromol/L brain derived neurotrophy factor (BDNF), 10 micromol/L forskolin (FSK) and 10 micromol/L dopamine (DA) to induce BMSCs differentiate into DA neurons. After 2 weeks of differentiation, the cells expressed the character of neurons in ultrastructure. RT-PCR discovered mRNA of NSE (neuron specific enolase), Nurr1, Ptx3, Lmx1b and Tyrosine hydroxylase (TH) were positive. Immunocytochemistry staining indicated the ratio of TH-positive neural cells was significantly increased after induced 2 weeks (24.80 +/- 3.36) % compared to that of induction of 3 days (3.77 +/- 1.77) %. And the DA release was also different between differentiated and undifferentiated cells detected by high performance liquid chromatography (HPLC). That is to say BDNF and FSK and DA can induce BMSCs differentiate into DA neurons in vitro, and the transdifferentiated cells express mature neurons characters. BMSCs might be a suitable and available source for the in vitro derivation of DA neurons and cell transplantation therapy in some central neural system diseases such as PD.
Adult ; Aged ; Bone Marrow Cells ; cytology ; metabolism ; ultrastructure ; Brain-Derived Neurotrophic Factor ; pharmacology ; Cell Transdifferentiation ; drug effects ; genetics ; physiology ; Cells, Cultured ; Chromatography, High Pressure Liquid ; Colforsin ; pharmacology ; Dopamine ; metabolism ; pharmacology ; Female ; Humans ; Immunohistochemistry ; Male ; Mesenchymal Stromal Cells ; cytology ; metabolism ; ultrastructure ; Microscopy, Electron, Transmission ; Middle Aged ; Neurons ; cytology ; metabolism ; ultrastructure ; Phosphopyruvate Hydratase ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Tyrosine 3-Monooxygenase ; genetics ; metabolism ; Young Adult

BACKGROUNDExposure of cells to sublethal concentrations of hydrogen peroxide (H2O2) can alleviate subsequent oxidative stress-induced apoptosis. We assessed the effects of H2O2 preconditioning on the therapeutic potential of human umbilical cord Wharton's Jelly mesenchymal stem cells (WJ-MSCs) in a murine model of myocardial infarction.

METHODSWJ-MSCs were incubated in the media for 2 hours with or without 200 µmol/L H2O2. Mice underwent left anterior descending coronary artery ligation, and received injection of phosphate buffered saline, 1×10(6) WJ-MSCs, or 1×10(6) H2O2 preconditioned WJ-MSCs 3 hours later via tail vein. Echocardiography was performed 0, 7, 14 and 28 days after surgery, and the mice were euthanized on day 28 for histological analysis. In vitro cytokine concentrations in the WJ-MSC cell supernatant were measured by enzyme-linked immunosorbent assay (ELISA). The effect of WJ-MSC cell supernatant on the migration and proliferation of endothelial cells were observed by transwell migration and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazoliumbromide (MTT) assays.

RESULTSEchocardiographic measurements revealed a significant improvement in the left ventricular contractility of the WJ-MSCs-H2O2 group compared to the WJ-MSCs group. Histological analysis revealed increased neovascularization and reduced myocardial fibrosis in the WJ-MSCs-H2O2 group compared to the WJ-MSCs group. Pretreatment of WJ-MSCs with H2O2 increased the secretion of interleukin-6 (IL-6) into the cell culture supernatant by approximately 25-fold. The culture supernatant from WJ-MSCs-H2O2 significantly increased the migration and proliferation of endothelial cells; these effects could be blocked using an anti-IL-6 antibody.

CONCLUSIONSThis study demonstrates that H2O2 preconditioning significantly enhanced the therapeutic potential of WJ-MSCs, possibly by stimulating the production of IL-6 by WJ-MSCs, which may cause migration and proliferation of endothelial cells and increase neovascularization.

Animals ; Cell Movement ; physiology ; Echocardiography ; Enzyme-Linked Immunosorbent Assay ; Humans ; Hydrogen Peroxide ; pharmacology ; Immunohistochemistry ; Interleukin-6 ; metabolism ; Male ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stromal Cells ; drug effects ; metabolism ; Mice ; Mice, Inbred C57BL ; Myocardial Infarction ; pathology ; therapy ; Reactive Oxygen Species ; metabolism ; Wharton Jelly ; cytology

OBJECTIVETo explore the role of stromal-derived factor-1 (SDF-1) in the migration of mesenchymal stem cells (MSCs) and the underlying signal transduction mechanism.

METHODSRat bone marrow-derived MSCs were infected with 100 ml recombinant adenovirus containing human SDF-1alpha gene (Ad-hSDF-1alpha), and the cell migration changes were observed at 1, 2, and 3 days after the infection. Twelve hours after Ad-hSDF-1alpha transfection, the MSCs in separate cultures were treated with wortmannin (50 nmol/L), LY294002 (10 mmol/L), PD98059 (50 mmol/L), U73122 (10 mmol/L), AMD3100 (0.1 g/L), or verapamil (50 nmol/L), respectively, and the signal transduction pathways involved in MSC migration were analyzed.

RESULTSThe MSCs grew in colonies after transfection with Ad-hSDF-1alpha, but this growth pattern was substantially attenuated after treatment with wortmannin, LY294002, PD98059, U73122, AMD3100 and verapamil, among which U73122 and AMD3100 treatments resulted in the most conspicuous inhibitory effects.

CONCLUSIONThe effect of SDF-1 in promoting MSC migration is related to mitogen-activated protein kinase, phosphatidylinositol phospholipase C, and protein kinase signal pathways.

Adenoviridae ; genetics ; Animals ; Cell Movement ; genetics ; physiology ; Cells, Cultured ; Chemokine CXCL12 ; genetics ; physiology ; Enzyme Inhibitors ; pharmacology ; Genetic Vectors ; genetics ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Mitogen-Activated Protein Kinases ; antagonists & inhibitors ; metabolism ; Rats ; Rats, Wistar ; Signal Transduction ; drug effects ; Transfection ; Type C Phospholipases ; antagonists & inhibitors ; metabolism