Human bone morphogenetic protein-2 gene transfer induces human mesenchymal stem cell proliferation and differentiation in vitro

To identify eukaryotie expression veetor of human bone morphogenetie protein 2 peDNA3/BMP2, verify its expression in transfected hulnan mesenchynlal stem cells(hMSCs) and the effect on hMSCs differentiation. Methods: The BMP2 gene was cloned into a eukaryotic expression veetor pcDNA3. Transfeeted the reeombinant into hMSCs by liposome, Lmmunnohistochemistry and it situ hybridization methods were used to identify the expression of BMP2 mRNA and protein; ALP and Von Kossa stains were performed to identify the BMP2 gene differentiated effect on the hMSCs. Resuits: The pcDNA3/BMP2 fragments were as large as theory. BMP2 mRNA and protein were espressed and synthesized both in 48 h and 4 weeks after transfeetion, the ALP and Ca deposit eshibitiio, which marked the osteogenie lineage of hMSCs, were enhanced and Sped. Conclusion: Transfeetion of pcDNA3/BMP2 is able to provide transient and persistent expressionin hMSCs, and promote the MSCs differentiation to osleogenic lineage.     

Effects of varied interferons in combination with all-trans retinoic acid （ ATRA ） on proliferation and differentiation of ATRA-resistent APL cell

Objective：To investigate the effects and mechanisms of interferon in combination with alltrans retinoic acid （ATRA） on proliferation and differentiation of ATRA-resistent APL cell. Methods ：After MR2 cells （ATRA-resistance cell line） were treated with IFN-α, IFN-γ and ATRA alone or IFN-α and IFN-γ in combination with ATRA respectively. The cell proliferation was tested by MTT test and the cell differentiation was tested through light microscope by NBT test and flow cytometry （FCM）. The expres sion of promyelocytic leukemia （PML） protein was observed by indirect immune fluorescent method. Results： Both IFN-α and IFN-γ could inhibit the proliferation and induce the differentiation of MR2 cells to some extent. The effects were more obvious after both interferons in combination with ATRA respectively （P〈0.05）. Moreover, the maturation of MR2 cells induced by IFN-γ＋ATRA group was more higher than that by IFN-α＋ATRA group （P〈0. 05）. Both interferons could induce the expressions of PML protein. Conclusion：Both interferons can inhibit MR2 cells proliferation, which may be related to the expression of PML protein induced by both interferons. The inducing differentiation effects of IFN-γ＋ATRA group on MR2 cells are more powerful than those of IFN-aq-ATRA group, which may be related to the different signal transduction pathway of both interferons.     

Growth and differentiation of neural stem cells with superparamagnetic iron oxides labeling in vitro

Objective： To study the growth and differentiation of superparamagnetie iron oxides（SPIOs） labeled neural stem cells （NSCs）. Methods： After NSCs were cultured and subcuhured from newborn rat brain, they were magnetically labeled with ferumoxides （a kind of SPIOs ）. Growth, differentiation and other biology properties of the cells were investigated with immunocytochemistry, transmission electron microscopy （TEM） and Prussian blue staining. Results： Nestin positive cells were found in the culture and offspring clones. NSCs could be differentiated into positive GFAP and NF200 cells in serum culture. When NSCs incubated with ferumoxides, the iron particles were seen in intracellular as well as in offspring clones. With the increase in concentration of ferumoxides （5.6-11.2/μg/ml）, ferumoxides showed no significant difference effects on the growth and differentiation of NSCs. When the concentration of ferumoxides exceeded 22.4μg/ml ,there was significant difference（P〈0.05）. Conclusion： We successfully label NSCs with ferumoxides,it is useful for tracking of magnetic labeled NSCs in vivo with MRI.     

Forskolin augments the effects of glucocorticoids on proliferation and differentiation of a human osteosarcoma cell line by u

Ｆｏｒｓｋｏｌｉｎａｕｇｍｅｎｔｓｔｈｅｅｆｆｅｃｔｓｏｆｇｌｕｃｏｃｏｒｔｉｃｏｉｄｓｏｎｐｒｏｌｉｆｅｒａｔｉｏｎａｎｄｄｉｆｆｅｒｅｎｔｉａｔｉｏｎｏｆａｈｕｍａｎｏｓｔｅｏｓａｒｃｏｍａｃｅｌｌｌｉｎｅｂｙｕｐ－ｒｅｇｕｌａｔｉｏｎｏｆｇｌｕ...     

Clinical outcomes after autologous haematopoietic stem cell transplantation in patients with progressive multiple sclerosis

Background Multiple sclerosis （MS） is a continuously disabling disease and it is unresponsive to high dose steroid and immunomodulation with disease progression. The autologous haematopoietic stem cell transplantation （ASCT） has been introduced in the treatment of refractory forms of multiple sclerosis. In this study, the clinical outcomes followed by ASCT were evaluated for patients with progressive MS. Methods Twenty-two patients with secondary progressive MS were treated with ASCT. Peripheral blood stem cells were obtained by leukapheresis after mobilization with granulocyte colony stimulating factor. Etoposide, melphalan, carmustin and cytosine arabinoside were administered as conditioning regimen. Outcomes were evaluated by the expanded disability status scale and progression free survival. No maintenance treatment was administered during a median follow-up of 39 months （range, 6 to 59 months）. Results No death occurred following the treatment. The overall confirmed progression free survival rate was 77% up to 59 months after transplantation which was significantly higher compared with pre-transplantation （P=0.000）. Thirteen patients （59%） had remarkable improvement in neurological manifestations, four （18%） stabilized their disability status and five （23%） showed clinical recurrence of active symptoms. Conclusions ASCT as a therapy is safe and available. It can improve or stabilize neurological manifestations in most patients with progressive MS following failure of conventional therapy.     

HMBA-induced differentiation of myeloid leukemic cell lines is assoc iated with altered G1 cell cycle regulators and related genes

Theproliferationanddifferentiationofhematopoieticcellscanberegulatedbyanumberofphysiologicalagentsincludinghexamethylenebisacetamide (HMBA) Clinically ,HMBAhasbeenusedforthetreatmentofacutemyeloidleukemiaandmyelodysplasticsyndrome 1 However,themechanismoftheeffectofHMBAonthedifferentiationofmyeloidleukemiccellsislargelyunkown Uptonow ,relatedreportshavenotbeenfound WeusedHL 6 0andU937celllinestostudytheeffectofHMBAonthedifferentiationofmyeloidleukemiccellsandtoexplorethepossiblemechan…     

Effects of different subtypes of histamine receptors on proliferation and differentiation of murine colony forming unit granulocyte-macrophage and colony forming unit megakaryocyte.

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Angiogenesis and neural progenitor proliferation induced by intracarotid administration of mesenchymal stem cells in rats with cerebral ischemia

1Introduction Strokeisoneoftheleadingcausesofdisability worldwide.Effectivetherapeuticsfortherehabilitationof strokesurvivorshaveyettobedeveloped.Bonemarrow stromalcellsormesenchymalstemcells(MSCs)havethe potentialforself renewalanddifferentiationintodiverse celltypes.Anumberofrecentpublicationshavereported thatMSCscannotonlygiverisetobone,cartilage,but alsodifferentiateintomyocytes,hepatocytes,glialcells,andneurons[1,2].Itisindicatedbysomestudiesthat MSCscanpassthroughtheblood brainbarrie…     

Molecular tissue engineering: Applications for modulation of mesenchymal stem cells proliferation by transforming growth factor β1 gene transfer

Summary The effect of transforming growth factor β₁ (TGF-β₁) gene transfection on the proliferation of bone marrow-derived mesenchymal stem cells (MSCs) and the mechanism was investigated to provide basis for accelerating articular cartilage repairing using molecular tissue engineering technology. TGF-β₁ gene at different doses was transduced into the rat bone marrow-derived MSCs to examine the effects of TGF-β₁ gene transfection on MSCs DNA synthesis, cell cycle kinetics and the expression of proliferating cell nuclear antigen (PCNA). The results showed that 3 μl lipofectaminemediated 1 μg TGF-β₁ gene transfection could effectively promote the proliferation of MSCs best; Under this condition (DNA/Lipofectamine= 1μg/3μl) flow cytometry and immunohistochemical analyses revealed a significant increase in the³H incorporation, DNA content in S phase and the expression of PCNA. Transfection of gene encoding TGF-β₁ could induce the cells at G0/G1 phase to S1 phase, modulate the replication of DNA through the enhancement of the PCNA expression, increase the content of DNA at S1 phase and promote the proliferation of MSCs. This new molecular tissue engineering approach could be of potential benefit to enhance the repair of damaged articular cartilage, especially those caused by degenerative joint diseases. This project was supported by a grant from National Natural Science Foundation of China (No. 30170270).     

Distribution ofintimal proliferation in long lesions covered with multiplestents:insightfrom an intravascular ultrasound study

Distribution ofintimal proliferation in long lesions covered with multiplestents:insightfrom an intravascular ultrasound study@F.Liu@JGe@D.Welge@M.Hauda@D.BaumgartandR.Erbel     

Dexamethasone impairs the differentiation and maturation of murine dendritic cells by Toll-like receptor 4-nuclear factor-κB pathway

Background Recent studies have demonstrated that dexamethasone （DEX） interferes with immune responses by targeting key functions of dendritic cells （DCs） at the earliest stage. However, the cellular and molecular mechanisms are still incompletely understood. This study aimed to explore the possible mechanisms by investigating the roles of DEX on differentiation, maturation ＆ function of murine DCs and the effects of DEX on DCs via Toll-like receptor 4 （TLR4）-nuclear factor （NF）-KB mediated signal pathway. Methods Immature DCs （imDCs） were cultured from murine bone marrow （BM） cells. We added DEX into culture medium at different time. The expression of CD11c, CD86 and I-Ab （mouse MHC class II molecule） was determined by flow cytometry. We determined the expression of NF-κB and its inhibitory protein I-κBα by electrophoretic mobility shift assay （EMSA） and Western blotting, respectively. The productions of interleukin （IL）-12p70 and IL-10 in cell culture supernatants were determined by enzyme-linked immunosorbent assay （ELISA）. Results DEX impaired differentiation of DCs from murine bone marrow progenitors, and inhibited lipopolysaccharide （LPS） induced maturation of DCs. DEX significantly inhibited NF-κB expression of normal DCs, the higher the DEX concentration or the longer the DEX treatment time, the more obvious the effect. However, DEX had little effect on LPS-induced NF-KB activation, and partially impaired LPS-induced I-κBα degradation. DEX significantly decreased LPS induced IL-12p70 production by DCs. Interestingly, our results showed a synergistic effect between DEX and LPS on the production of IL-10 by DCs. Conclusions DEX inhibits the differentiation and maturation of murine DCs involved in TLR4-I-κB-NF-κB pathway, and also indirectly impairs Thl development and interferes with the Thl-Th2 balance through IL-12 and/or IL-10 secretion by DCs.     

Association of β-adrenergic receptor genes polymorphisms with incidence of subsequent cardiovascular events in Han Chinese patients with coronary artery disease

Background Sequence variants in the 13-adrenergic receptor （ADRB） genes have a close relationship with the development of coronary artery disease （CAD） and the patient＇s prognosis. However, there is a lack of data on the role of the variants in ADRBs genes in Han Chinese patients with CAD. We aimed to investigate the association of genetic variants in the ADRB1 and ADRB2 genes with the incidence of major adverse cardiac event （MACE） in Han Chinese patients with CAD. Methods A total of 545 Han Chinese patients with CAD undergoing percutaneous coronary intervention （PCI） were recruited to the study and followed for one year. Three variant sites in ADRB1 （rs1801253） and ADRB2 （rs1042713 and rs1042714） were genotyped. The effect of the ADRB1 and ADRB2 genotypes on MACE within one year was assessed. Results There were 47 cases of MACE during follow-up. There was no significant difference in the incidence of MACE among patients carrying different genotypes of the three variants in ADRB1 and ADRB2 （Log-rank, all P 〉0.05）. Cox regression analysis showed no association between three variants in ADRB1 and ADRB2 genes and the incidence of MACE during one-year follow-up, the adjusted hazard ratios （95% confidence interval） for rs1801253, rs1042713 and rs1042714 were 1.05 （0.54-2.02）, 1.24 （0.58-2.64）and 1.66 （0.81-3.42）, respectively. Conclusion Our data did not support a relationship between the three polymorphisms of ADRB1 （rs1801253） and ADRB2 （rs1042713 and rs1042714） genes and risk of subsequent cardiovascular events after PCI in Han Chinese patients with CAD.     

Rapid identification of the origin of hematopoietic cells after allogeneic hematopoietic stem cell transplantation for leukemia with DNA-fingerprinting

Allogeneichematopoieticstemcelltransplan-tation(AHCT)isusedtotreatleukemia,severeaplasticanaemiaandsoon.OneoftheimportantindexesofsuccessfulAHCTisthedetectionofdonorcellsafterreconstitutionofhematopoieticfunctionintherecipient.AvarietyoftechniqueshavebeenusedtoconfirmthesurvivalofthegraftafterAHCT,whichincludeRBCphenotyping,RBCandWBCisoenzymeanalysis,immunoglobu-linallotyping,andsoon,However,thesemarkersmaybeeasilyaffectedbymanyadditionalfactorssuchasmultipletransfusions,samesex,samebl…     

Corelation of cell proliferation in hibition and apoptosis induction with expression of human β5 integrin on hematopoietic cells

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Effects of Kangquan Recipe (康泉方) on sex steroids and cell proliferation in rats with benign prostatic hyperplasia

Objective:To investigate the effects of Kangquan Recipe(康泉方,KQR)on sex steroids and cell proliferation in an experimental benign prostatic hyperplasia(BPH)model in rats.Methods:Seventy-two SD rats were randomly divided into six groups:the normal group,the model group,the finasteride group,and the low-, middle-,and high-dose KQR groups,12 in each group.Except those in the normal group,the rats were injected with testosterone after castration for the establishment of BPH model and then given respectively w...     

Sertoli cell-mediated differentiation of male germ cell-like cells from human umbilical cord Wharton’s jelly-derived mesenchymal stem cells in an in vitro co-culture system

Background

Microenvironment signals play a critical role in directing the differentiation of stem cells. Sertoli cells (SCs) provide a unique microenvironment that is essential for germ cell differentiation.

Methods

Our previous study has demonstrated that human umbilical cord Wharton’s jelly-derived mesenchymal stem cells (HUMSCs) could differentiate towards male germ cells in vitro, but HUMSC-derived germ-like cells expressed only few germ cell markers. The aim of this study was to investigate the effect of SCs on the differentiation of HUMSCs towards male germ cells using a co-culture system that mimicked the in vivo male germ cell microenvironment.

Results

HUMSCs formed clump-like features on SC monolayers after seeding for 3 weeks. Differentiated cells formed round colonies that share the morphological features of spermatogonial colonies. RT-PCR, immunofluorescence, confocal microscopy, and Western blot analyses revealed the expression of early germ cell markers STELLA and VASA and male germ cell-specific marker DAZL in differentiated HUMSCs, confirming the presence of cells with characteristics of male germ cells.

Conclusion

The HUMSC-SC co-culture system mimics a native microenvironment for germ cell colonization without any in vitro artificial manipulation and can be used to explore the mechanisms controlling the differentiation of male germ cells from HUMSCs. Male germ cells derived from HUMSCs may be used in the therapy for male infertility.     

Differential thymosin β10 expression levels and actin filament organization in tumor cell lines with different metastatic potential

Background To investigate the differential expression levels of thymosin [β10 (Tβ10) and the corresponding changes of actin filament organization in human tumor cell lines with different metastatic potential.Methods Four groups of nine human tumor cell lines with different metastatic potential were analyzed for the amount of Tβ10 mRNAs by Northern blot and for their peptide expression levels by immunohistochemistry. The filamentous actin (F-actin) was observed by staining of TRITC-phalloidin to detect changes in actin organization.Results In comparison with non-/weakly metastatic counterparts, Tβ10 was upregulated in highly metastatic human lung cancer, malignant melanoma and breast cancer cell lines. Staining of TRITC-phalloidin revealed less actin bundles, a fuzzy network of shorter filaments and some F-actin aggregates in the highly metastatic tumor cells. Meanwhile, the actin filaments were robust and orderly arranged in the non-/weakly metastatic cancer cell lines.Conclusion Tβ10 levels correlate positively with the metastatic capacity in human tumors currently examined. The increasing metastatic potential of tumor cells is accompanied by a loss of F-actin,poorly arranged actin skeleton organizations and presence of F-actin aggregates. There is a consistent correlation between the elevated Tβ10 expression and the disrupted actin skeleton.     

Effects of anti-CXCR4 monoclonal antibody 12G5 on proliferation and apoptosis of human acute myelocytic leukemia cell line HL-60

Objective:To investigate the expression of CXCR, on HL-60 cell line and the proliferation, apoptosis of HL-60 cell line cocultured with bone marrow stromal cells, so as to assess the possibility of 12G5. an anti-CXCR4 monoclonal antibody, in eradicating the minimal residual disease. Methods:The activity of SDF-1 was inhibited by 10μg/ml 12G5. After treatment with 12G5. the status of adhesion was observed, and the adhesion rates, apoptosis and cell cycles were detected after 24 h of treatment. Cell growth rates were measured by trypan blue exclusion. Cell growth curve was plotted, and the expression of PCNA and apoptosis related protein including PCNA, Bcl-2 and Fas were detected with immunohis-tochemical technique. Results :(1) There was middling degree expression of CXCR4 on HL-60 membrane. From 0 h to 6 h, as the time of 12G5 incubation along, the expression of CXCR4 decreased gradually. (2) After treatment for 24 h, the adhesion rates in the experiment group and the control were (39. 4±7. 9)% and (51. 4±5. 9)%, respectively. (3)After treatment for 24 h, the percentage of HL-60 cells in G0/G1 phase were (55. 21±4. 9)%, and that in S phase and G2/M phase were (30. 40±4. 1)% and (14. 39±5.2)%, respectively, with the corresponding proportions being (44. 67±2. 2) % , (45. 30±3. 7)% . and (10. 03±2. 6)% in the control. (4) The percentage of apoptotic HL-60 cells was (8. 95±1. 7)% in the experiment group, compared to (3. 97±2. 4)% in the control. (5)The survival rates of HL-60 cells decreased markedly at 48 h to 96 h, and the proliferation slowed down at this time duration. (6)The expression of PCNA and Bcl-2 down-regulated significantly, but the Fas protein expression was up-regulated. Conclusion :12G5 could inhibit the capability of adhesion and proliferation of HL-60 cells and it can induce more cells to enter G0/G1 phase and promote apoptosis. It may be helpful by inhibiting the bioactivity of SDF-1 with 12G5 in the therapy of marrow residual disease.