Shenzhen Beike Cell Engineering Institute

Shenzhen, China

Shenzhen Beike Cell Engineering Institute

Shenzhen, China
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Guo D.,Zhengzhou University | Li D.,Zhengzhou University | Li J.,Zhengzhou University | Li Y.,P.A. College | And 3 more authors.
Neurological Research | Year: 2017

Object: The pathologies resulting from traumatic brain injury (TBI) have been thoroughly studied, but rarely have the effects of bleeding and coagulation in the early stage of TBI been considered. In this study, we investigated the effects of topical Surgiflo® application on brain injury in experimental TBI mice using S100β, MAP-2 and mNSS scores. Methods: TBI was induced by modified weight drop injury in male C57BL/6 mice. The mice were then randomly divided into (i) the sham group, (ii) TBI mice applied with saline (vehicle), and (iii) TBI mice applied with Surgiflo® in the same volume. Modified neurological severity scores (mNSS) were measured on days 0 (before surgery), 1, 3, 7, and 28 to evaluate neurologic functional deficits. At day 28, the mice were sacrificed, and the forebrains were sliced. The effects of Surgiflo® on microtubule-associated protein 2 and serum S100β protein were examined by immunohistochemistry and electro-chemiluminescence immunoassay. Results: Serum S100β protein levels were significantly elevated at different time points (24 h, 3 days, 7 days) in the TBI groups (p  <  0.01) compared to normal control groups. Surgiflo® induced a lower concentration of serum S100β protein levels at day 3 (p < 0.05) and day 7 (p < 0.05) compared to the TBI group applied with saline. H&E staining showed that Surgiflo® treatment led to a 45% decrease in cortical brain lesion volume and in subcortical white matter 28 days after TBI. Compared with the saline-treated group, the number of MAP2-positive cells was significantly increased in the perilesional area of the Surgiflo®-treated group. The Surgiflo®-treated group exhibited lower mNSS scores on days 7 and 28 than did the saline-treated group. Discussion: Surgiflo® treatment produced a significant decrease in serum S100β protein levels in TBI mouse models, which may lead to an improvement in the recovery of TBI models. © 2017 Informa UK Limited, trading as Taylor & Francis Group

Ichim T.E.,Medistem Inc. | Harman R.J.,Vet Inc. | Min W.-P.,University of Western Ontario | Minev B.,University of California at San Diego | And 7 more authors.
Cellular Immunology | Year: 2010

Since the days of Medawar, the goal of therapeutic tolerogenesis has been a " Holy Grail" for immunologists. While knowledge of cellular and molecular mechanisms of this process has been increasing at an exponential rate, clinical progress has been minimal. To provide a mechanistic background of tolerogenesis, we overview common processes in the naturally occurring examples of: pregnancy, cancer, oral tolerance and anterior chamber associated immune deviation. The case is made that an easily accessible byproduct of plastic surgery, the adipose stromal vascular fraction, contains elements directly capable of promoting tolerogenesis such as T regulatory cells and inhibitory macrophages. The high content of mesenchymal and hematopoietic stem cells from this source provides the possibility of trophic/regenerative potential, which would augment tolerogenic processes by decreasing ongoing inflammation. We discuss the application of this autologous cell source in the context of rheumatoid arthritis, concluding with some practical examples of its applications. © 2010 Elsevier Inc.

Zhao Z.F.,Xuchang Central Hospital | Zhao Z.F.,Zhengzhou University | Fan R.T.,Zhengzhou University | Yang Y.,Wei Erke Stem Cell Biotechnology Liaoning Co. | And 5 more authors.
Chinese Journal of Tissue Engineering Research | Year: 2013

BACKGROUND: Radiotherapy effect on of brain glioma is not ideal, and there are various possible factors that influence the therapeutic effect. OBJECTIVE: To investigate the in vitro radiosensitivity of brain cancer stem cells in brain glioma. METHODS: Glioma cells were isolated and seeded into the serum-free medium containing growth factors for culture. Glioma cells from the most active cell site were subcultured to 3-5 generations. Glioma cell vitality was assessed to determine the optimal dose of X-ray radiation. RESULTS AND CONCLUSION: Distinct difference in activities of proliferation could be found among cancer stem cells from different sites of glioma. X-ray dosages above 8 Gy had notable killing effect on brain cancer stem cells. Different irradiation dosage had different influence on cancer stem cells.

Tian Y.,Zhengzhou University | Hu X.,Jiangsu Public Technology Service Platform of Stem Cells and Biotherapy | Hu X.,Shenzhen Beike Cell Engineering Institute | Yang B.,Zhengzhou University | And 4 more authors.
Journal of Clinical Rehabilitative Tissue Engineering Research | Year: 2010

BACKGROUND: Human mesenchymal stem cells derived from Wharton's jelly (WJCs) display the characteristics of MSCs as defined by the International Society for Cellular Therapy. They can be differentiated into bone, cartilage, adipose, muscle, and neural cells. They can also support the expansion of other stem cells, be well-tolerated by the immune system, and have the ability to home to tumors. OBJECTIVE: To investigate biological changes of WJCs and brain tumor stem cells (BTSCs) co-cultured in vitro. METHODS: WJCs cultured by situ cultivation and BTSCs used enzyme digestion way respectively, and gathering the 3rd passage of WJCs though subculturing as well as BTSCs. Two kinds of cells co-cultured in 24-well plates in serum-free medium (SFM) without any growth factor. 3 and 7 days after co-cultured respectively, CD133 expression of suspension cells in the 24-well plates were identified by flow cytometry, and immunofluorescence was performed for Nestin and glial fibrillary acidic protein (GFAP) expression of adherent cells. Co-culture supernatant (CCS) re-suspended 3rd passage of BTSCs and cultured into 96-well plates at day 3, which were used to determine the difference in cell growth curve in both groups using a microplate reader. RESULTS AND CONCLUSION: With the cocultivation days increasing, the phenomenon that tumor sphere cells began to be decomposed, adherent and differentiated observed by an inverted microscope. BTSCs in the co-cultured group expressed GFAP and Nestin when adherent and differentiated. The higher degree of malignant brain tumor tissue used in culturing BTSCs was, the higher expression of CD133 in BTSCs was. CD133+ in BTSCs declined when co-cultured with WJCs. Growth curve of brain tumor stem cells cultured in CCS compared with in SFM at day 3, which indicates that the proliferation of BTSCs inhibited obviously. Results indicated that CD133+ expression and proliferative capacity of BTSCs went down and BTSCs underwent differentiation during the co-culture in vitro.

Ba Y.-T.,Zhengzhou University | Ba Y.-T.,Chinese Institute of Clinical Medicine | Guan F.-X.,Zhengzhou University | Xiang H.,Jiangsu Public Technology Service Platform of Stem Cells and Biotherapy | And 11 more authors.
Journal of Clinical Rehabilitative Tissue Engineering Research | Year: 2010

BACKGROUND: During culture of mesenchymal stem cells (MSCs) derived from Wharton's jelly of human umbilical cord. MSC morphology tends to become hypertrophic and irregular. MSCs were found to have higher rate of death and not be adapt to be adherent after passage. It is necessary to find a method to maintain its stability. OBJECTIVE: To isolate MSCs from human umbilical cord wharton's jelly by tissue block method, and to investigate basic fibroblast growth factor (bFGF) effects on biological characteristics of MSCs. METHODS: Human umbilical cord mesenchymal stem cells (hUCMSCs) were separated by tissue block method. Tissue fragments in control group were cultured in growth medium consisting of Dulbecco's Modified Essential /F12 Media (DMEM/F12) and 10% volume fraction of fetal bovine serum. Tissue fragments in experiment group were cultured in growth medium including 20 ug/L bFGF besides these in control group. Time of tissue blocks emigrating from cells and morphology of cells were observed. The medium was changed every 3-4 days. When the cells reached 80%-90% confluency, they were detached with 0.25% trypsin and were passaged at a ratio of 1: 2 or 1:3. RESULTS AND CONCLUSION: A few long spindle or flat fibroblast-like cells were presented firstly after 8-10 days of incubation in control group, while in experiment group it took 6-8 days. One week later, lots of long spindle or flat fibroblast-like cells like whirlpool around micro-mass were presented in two groups. In the first 3 passages, cell morphous were similar and cells were passaged at approximately equivalent time. After passage 3, cells in experiment group were easier to be adherent, lower rate of death and better proliferation ability (shown by growth curve) in comparison with these in control group. Flow cytometry revealed that cell cycle at the passages 3 and 6 showed the percentage of G0/G1 was more than 70% respectively. CD44 and CD29 were highly expressed on the surface of passages 3 and 6 cells, whereas the expression of HLA-ABC was less positive, but there was negative for CD34, CD45 and HLA-DR. Cells from Wharton's jelly of the human umbilical cords, which have the biological characteristics of MSCs, have been isolated by tissue culture method. bFGF can shorten the time that MSCs are presented firstly, improve its adherence and proliferation ability and maintain its morphological stability in some degree, moreover keep surface marker expression stability of MSCs.

Zhou C.-H.,Zhengzhou University | Zhou C.-H.,Henan Institution of Higher Education | Tian Y.,Henan Institution of Higher Education | Tian Y.,Zhengzhou University | And 10 more authors.
Journal of Clinical Rehabilitative Tissue Engineering Research | Year: 2010

BACKGROUND: Bone marrow mesenchymal stem cells have low immunogenicity and immunomodulatory effect, but there are seldom reports concerning the immunomodulatory effect of Wharton's jelly-derived mesenchymal stem cells of human umbilical cord and its mechanims. OBJECTIVE: To investigate the immunomodulatory effects and mechanisms of Wharton's jelly-derived mesenchymal stem cells of human umbilical cord on varient peripheral blood T lymphocytes. METHODS: Mesenchymal stem cells were isolateded from Wharton's jelly of human umbilical cord by tissue culture. T lymphocytes from human peripheral blood were stimulated by phytohemagglutinin and co-cultured with umbilical cord Wharton's jelly-derived mesenchymal stem cells and umbilical cord Wharton's jelly-derived mesenchymal stem cells supernatant respectively to measure A value following 72 hours of coculture using multifunctional microplate reader. Expression of cytokines including transforming growth factor-beta 1 (TGF-β1) and interferon-γ (IFN-γ) was evaluated by enzyme-labeled immunosorbent assay. RESULTS AND CONCLUSION: Wharton's jelly-derived mesenchymal stem cells could inhibite the proliferation of T lymphocytes induced by phytohemagglutinin. The proliferation inhibition rate was 56% (P < 0.01). Wharton's jelly-derived mesenchymal stem cells supernatant also had inhibitory effects on proliferation of T lymphocytes induced by phytohemagglutinin, in a dose-dependent fashion. The proliferation inhibition rates were 8.3% and 27% respectively in the 50% Wharton's jelly-derived mesenchymal stem cells supernatant and 100% Wharton's jelly-derived mesenchymal stem cells supernatant groups (P < 0.05). Wharton's jelly-derived mesenchymal stem cells significantly decreased γ-interferon secrted from T-lymphocytes (P < 0.05). The secretion of TGF-β1 was lower in the coculture of Wharton's jelly-derived mesenchymal stem cells and T lymphocytes group than Wharton's jelly-derived mesenchymal stem cells alone group (P < 0.05). These indicated that Wharton's jelly-derived mesenchymal stem cells and Wharton's jelly-derived mesenchymal stem cells supernatant have inhibitory effects on proliferation of T lymphocytes induced by phytohemagglutinin. The mechanims may be associated with cell contant and inhibition of γ-interferon secrted from T-lymphocytes.

Wang C.-C.,Zhengzhou University | Wang C.-C.,Chinese Institute of Clinical Medicine | Bo Y.,Chinese Institute of Clinical Medicine | Bo Y.,Zhengzhou University | And 9 more authors.
Journal of Clinical Rehabilitative Tissue Engineering Research | Year: 2010

BACKGROUND: APP gene is closely associated with the onset of Alzheimer' disease. Intravenous transplantation of human amniotic mesenchymal stem cell (AMSCs) can promote the learning and memory improvement in transgenic APP+ mice with Alzheimer' disease. OBJECTIVE: To study whether the AMSCs can transfer into the brain tissue of Alzheimer's disease mice, and differentiate into the neural cell, then cure the disease after the human AMSC transplantation via tail venous injection. METHODS: Human AMSCs were isolated in vitro sterilely. At the third passage, 0.5 mL single cell suspension at 1 × 109/L was obtained and transplanted by tail venous pathway in transplantation group mice, Mice in the control group were injected with an equal volume of saline. APP-gene mice in the normal group were left intact. 5′-bromo-2-deoxyuridine (BrdU) labeled third-generation AMSCs expression was detected in mice brain tissue by immunohistochemical method. Glial fibrillary acidic protein (GFAP), Nestin and neuron specific enolase (NSE) expression was measured in the brain tissue of mice from each group. RESULTS AND CONCLUSION: Under an optical microscope, a majority of nuclei in the brain tissue of mice from transplantation group were stained blue, but some nuclei were stained brown, positive for BrdU. Compared with control group, the expression of GFAP in the brain tissue of transplanted mice was increased about 4 times, even more than in the normal group (P < 0.05). The expression of Nestin in the brain tissue of transplanted mice was increased about 10%, but still lower than the normal group nearly 20% (P < 0.05). NSE expression was decreased by 1/3, but still higher compared with the normal group (P < 0.05). Above-mentioned results have shown that human AMSC transplantation for treating Alzheimer's disease takes place by AMSCs homing to APP+ mouse brain tissue and differentiating into neural cells.

Zhao Q.,Zhengzhou University | Zhao Q.,Zhengzhou Peoples Hospital | Li Q.-R.,Zhengzhou University | Du Y.,Zhengzhou University | And 3 more authors.
Journal of Clinical Rehabilitative Tissue Engineering Research | Year: 2011

BACKGROUND: T cells are believed to play an important role in anti-infection, anti-tumor and immune function. However, the mechanism underlying the differentiation and development remains poorly understood. OBJECTIVE: To investigate the distribution of T cells in nude mice that are jointly transplanted human thymus and cord blood and the reconstruction of the immune function. METHODS: Thirty Balb/c nu/nu nude mice were randomly divided into two groups: an experimental group and a control group. In the experimental group, human thymus tissue was transplanted into the renal capsule of nude mice. Two weeks later, freshly isolated human cord blood CD34 + cells suspension was back perfused into the nude mice via the vein. In the control group, CD34 + cells transplantation was performed directly without thymus transplantation. After 60 days of breeding, the immune function of nude mice was detected in two groups. RESULTS AND CONCLUSION: Human thymus tissue in the renal capsule of nude mice survived and expressed CD3 and HLA-DR molecule. In the experimental group, CD3 + cells which distributed in the form of dots were observed in the mouse spleen. The proportion of CD3 +, CD4 +, CD8 +, and CD4 +CD25 + cells were significantly higher in the experimental group than in the control group. The nude mice from the experimental group rejected human gastric cancer BGC823 cells, while those from the control group did not. These findings demonstrated that combined human thymus and CD34 + cell transplantation allow nude mice to acquire T cell-mediated cellular immune function and possess the ability of anti-tumor.

Li Y.,Zhengzhou University | Li Y.,Stanford University | Hu J.,Stanford University | Guan F.,Zhengzhou University | And 6 more authors.
Oncology Reports | Year: 2013

Most human tumor cells, including glioblastoma multiforme (GBM) cells, have aberrant control of cell aging and apoptosis. Subcytotoxic concentrations of oxidative or stress-causing agents, such as hydrogen peroxide, may induce human cell senescence. Thus, induction of tumor cells into premature senescence may provide a useful in vitro model for developing novel therapeutic strategy to combat tumors. In the present study, we assessed the molecular mechanism(s) underlying senescence in GBM cells induced by copper sulfate. Following pretreatment with subcytotoxic concentrations of copper sulfate, U87-MG tumor cells showed typical aging characteristics, including reduced cell proliferation, cell enlargement, increased level of senescence-associated β-galactosidase (SA β-gal) activity, and overexpression of several senescence-associated genes, p16, p21, transforming growth factor β-1 (TGF-β1), insulin growth factor binding protein 3 (IGFBP3) and apolipoprotein J (ApoJ). We further demonstrated that the Bmi-1 pathway was downregulated in GBM cells in parallel with the induced senescence. The present study for the first time demonstrates the ability of copper to induce GBM cell senescence by downregulating Bmi-1.

Chen H.-P.,Zhengzhou University | Li Q.-R.,Zhengzhou University | Zhang J.,Zhengzhou University | Du Y.,Zhengzhou University | And 4 more authors.
Journal of Clinical Rehabilitative Tissue Engineering Research | Year: 2010

BACKGROUND: Further studies are needed to understand the cytobiological character, functional regulation, gene changes and expression difference of CD34+ and CD133+ stem cells induced by basic fibroblast growth factor (bFGF) using gene chip.OBJECTIVE: To compare the differences of gene expression and the response to bFGF of human umbilical cord CD34 + and CD133+ cells, and to explore gene expression changes of bFGF-induced umbilical cord CD34+ and CD133+ hematopotic stem cells/hemapoietic progenitor cells in vitro. METHODS: Human umbilical cord blood CD34+ and CD133+ cells were isolated and purified by MiniMACS immunomagnetic beads selection. The CD34+ and CD133+cells were cultured for 10 to 15 days in DMEM/F12 medium, supplemented with bFGF and B27. Total RNA from these cells was extracted and the genetic level of these cells was performed using Oligo GEArray® chip and GEArray software. Selected rate of CD34+ and CD133+ hematopoietic stem cells was detected using flow cytometry. CD34+ and CD133+ cell morphological changes were measured before and after bFGF induction. The concentration and purity of RNA were determined by agarose gel electrophoresis degeneration. Gene-chip test results were analyzed. RESULTS AND CONCLUSION: ?The 20 samples of cord blood were isolated and purified respectively, CD34+ cell purity (77.52±5.06)%, recovery rate (2.74±1.59)%; CD133+ cell purity (79.16±3.37) % and the recovery (1.12±0.94)%. ?The newselected CD34+ cells were spherical. Following induced by bFGF for 15 days, the majority of cell morphology did not find significant changes, some cells were adherent growth, and protruding and spindle cells were seen. CD133+ cells were spherical, by bFGF in cultured 15 days later, cells were significantly amplified, the round shape changed into an irregular shape, and some cells were adherent growth. ?The total RNA of CD34+ stem cells before and after incubation was respectively 2 236 ng and 1 796 ng. The total RNA of CD133 + stem cells before and after induction was respectively 2 518 ng and 2 191 ng. ?In the detection of 263 genes related to stem cells, two-fold differences of 10 genes in umbilical cord blood CD34+ cells and CD133+ cells, including five kinds of genes expression were higher in the former than the latter, five kinds of genes expression were lower in the former than the latter. After bFGF-induced culture, 32 kinds of gene expression of CD133+ cells were significantly higherthan CD34+ cells. Among detected 263 genes, no gene was lower than CD34+ cells. There were only a few gene expression differences of fresh-separated cord blood CD133+ cells and CD34+ cells. The response of CD133 + cells to bFGF was significantly stronger than CD34+ cells, which manifested cell cycle regulation, signal transduction and differentiation, gene expression enhanced.

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