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1.
To examine the effects of co-culture with bone marrow mesenchymal stem cells on expansion of hematopoietic stem/progenitor cells and the capacities of rapid neutrophil engraftment and hematopoietic reconstitution of the expanded cells, we expanded mononuclear cells (MNCs) and CD34+/c-kit+ cells from mouse bone marrow and transplanted the expanded cells into the irradiated mice. MNCs were isolated from mouse bone marrow and CD34+/c-kit+ cells were selected from MNCs by using MoFlo Cell Sorter. MNCs and CD34+/c-kit+ cells were co-cultured with mouse bone marrow-derived mesenchymal stem cells (MSCs) under a two-step expansion. The expanded cells were then transplanted into sublethally irradiated BDF1 mice. Results showed that the co-culture with MSCs resulted in expansions of median total nucleated cells, CD34+ cells, GM-CFC and HPP-CFC respectively by 10.8-, 4.8-, 65.9- and 38.8-fold for the mononuclear cell culture, and respectively by 76.1-, 2.9-, 71.7- and 51.8-fold for the CD34+/c-kit+ cell culture. The expanded cells could rapidly engraft in the sublethally irradiated mice and reconstitute their hematopoiesis. Co-cultures with MSCs in conjunction with two-step expansion increased expansions of total nucleated cells, GM-CFC and HPP-CFC, which led us to conclude MSCs may create favorable environment for expansions of hematopoietic stem/progenitor cells. The availability of increased numbers of expanded cells by the co-culture with MSCs may result in more rapid engraftment of neutrophils following infusion to transplant recipients. Project supported by NIH-Blood, Heart & Lung (National Institute of Health, USA, IR 01 4L70593-01) and Zhejiang Provincial Science Foundation (No. 011103397), China  相似文献   

2.
In this paper,we investigate the effect and the possible mechanism of high glucose levels on the calcification of human aortic smooth muscle cells (HASMCs).HASMCs were divided into four groups: normal glucose group (NG),osmolality control group (OC),high glucose group (HG,HASMCs culture medium containing 30 mmol/L glucose),and high glucose plus recombinant human Noggin protein (bone morphogenetic protein-2 (BMP-2) antagonist) group (HN).The mRNA levels and the protein expressions of BMP-2 and core binding factor alpha-1 (Cbfα-1) were measured by real-time quantitative polymerase chain reaction (PCR) and Western blot.After induced by 10 mmol/L β-glycerol phosphoric acid,cells were harvested for assessments of alkaline phosphatase (ALP) activities at Days 1,2,and 3,and intracellular calcium contents at Days 7 and 14,respectively.High glucose levels increased the mRNA levels and the protein expressions of BMP-2 and Cbfα-1 (P0.05).The expression of Cbfα-1 was partially blocked by Noggin protein (P0.05),while BMP-2 was not (P0.05).After being induced by β-glycerol phosphoric acid,high glucose levels increased the ALP activity [(48.63±1.03) vs.(41.42±2.28) U/mg protein,Day 3;P0.05] and the intracellular calcium content [(2.76±0.09) vs.(1.75±0.07) μmol/mg protein,Day 14;P0.05] in a time-dependent manner when compared with the NG group,while the ALP activity could not be blocked by Noggin protein [(48.63±1.03) vs.(47.37±0.97) U/mg protein,Day 3;P0.05].These results show that high glucose levels can evoke the calcification of HASMCs by inducing osteoblastic trans-differentiation and intracellular calcium deposition via the BMP-2/Cbfα-1 pathway,which can be partially blocked by Noggin protein.  相似文献   

3.
Long-term preservation and easy transportation of human bone marrow-derived mesenchymal stem cells (hBM-MSCs) will facilitate their application in medical treatment and bioengineering. A pilot study on the freeze-drying of hBM-MSCs was carried out. hBM-MSCs were loaded with trehalose. The glass transition temperature of the freeze-drying suspension was measured to provide information for the cooling and primary drying experiment. After freeze-drying, various rehydration processes were tested. The highest recovery rate of hBM-MSCs was (69.33±13.08)%. Possible methods to improve freeze-drying outcomes are discussed. In conclusion, the present study has laid a foundation for the freeze-drying hBM-MSCs.  相似文献   

4.
Retinoic acid level in the retina/choroid is altered in induced myopia models. All-trans-retinol dehydrogenase (RDH8) is an important enzyme of retinoic acid metabolism. This study aimed to investigate the association of the RDH8 gene with high myopia. Three single nucleotide polymorphisms (SNPs) [RDH851 (rs2233789), RDH8E5a (rs1644731), and RDH855b (rs3760753)] were selected, based on the linkage disequilibrium pattern of RDH8 from a previous study, and genotyped for 160 Han Chinese nuclear families with highly myopic (−10 diopters or worse) offspring as well as in an independent group with 166 highly myopic cases (−10 diopters or worse) and 211 controls. Family-based association analysis was performed using the family-based association test (FBAT) package, and genotype relative risk (GRR) was calculated using the GenAssoc program. Population-based association analysis was performed using Chi-square test. These SNPs were in linkage equilibrium with each other. SNPs RDH851 (rs2233789) and RDH8E5a (rs1644731) both did not show association with high myopia. SNP RDH855b (rs3760753) demonstrated significant association (P=0.0269) with a GRR of 0.543 (95% confidence interval=0.304–0.968, P=0.038). The association became statistically insignificant, however, after multiple comparison correction. Haplotype analysis did not show a significant association either. Population-based association analysis also showed no significant association (P>0.05). Our family- and population-based data both suggest that the RDH8 gene is unlikely to be associated with high myopia in Chinese.  相似文献   

5.
Naturally fermented pickles harbour many lactic acid bacteria (LAB). Forty-three LAB strains with conjugated linoleic acid (CLA)-producing ability were isolated from three naturally fermented pickle brines. Of these isolates, lp15 identified as Lactobacillus plantarum by API 50 CHL system and full-length 16S rDNA sequence analysis exhibited the highest CLA-producing ability (26.1% conversion) at 48 h in de Man Rogosa Sharpe (MRS) broth in the presence of 100 μg/ml of linoleic acid (LA). Compared to other strains, L. plantarum strain lp15 showed the highest tolerance upon increased levels of LA in the medium, i.e., up to 600 μg/ml. This strain converted about 25% of LA into CLA isomers [predominantly cis-9, trans-11 CLA (9-CLA) and trans-10, cis-12 CLA (10-CLA)], of which 75% was 9-CLA. Interestingly, though the conversion rate of LA into CLA by lp15 remained stable between 100 to 600 μg/ml LA levels in the medium, it dropped sharply at 1000 μg/ml. Taken together, the lp15 strain displayed relatively high LA tolerance with higher conversion rate, which implies that this strain is a valuable candidate for enhancing the CLA content in food-sources like pickles.  相似文献   

6.

Objective  

To study whether Tongxinluo (TXL) can induce angiogenesis in bone marrow mesenchymal stem cells (MSCs), and to investigate the underlying mechanism.  相似文献   

7.
INTRODUCTION Congestive heart failure is the end stage of manycardiovascular diseases. Myocardial infarction (MI)is a life-threatening event that may cause suddencardiac death and heart failure. Despite considerableadvances in diagnosis and treatment of heart disease,cardiac dysfunction after MI is still the majorworldwide cardiovascular disorder. Damaged myo-cardium after acute MI is gradually replaced by fi-brotic noncontractile cells to form scar tissue. Thedeveloping ventricul…  相似文献   

8.
Objective: To investigate the directed transplantation of allograftic bone marrow-derived mesenchymal stem cells (MSCs) in myocardial infarcted (MI) model rabbits. Materials and Methods: Rabbits were divided into 3 groups, heart infarcted model with MSCs transplanted treatment (MSCs group, n=12), heart infarcted model with PBS injection (control group, n=20), sham operation with PBS injection (sham group, n=l 7). MSCs labelled by BrdUrd were injected into the MI area of the MSCs group. The same volume of PBS was injected into the MI area of the control group and sham group. The mortality, LVIDd, LVIDs and LVEF Of the two groups were compared 4 weeks later. Tropomyosin inhibitory component (Tn I) and BrdUrd immunohistochemistry identified the engrafted cells 4 weeks after transplantation. Result: The mortality of the MSCs group was 16.7% (2/12), and remarkably lower than the control group's mortality [35% (7/20) (P＜0.05)].Among the animals that survived for 4 weeks, the LVIDd and LVIDs of the MSCs group after operation were 1.17±0.21 cm and 0.74±0.13 cm, and remarkably lower than those of the model group, which were 1.64±0.14 cm and 1.19±0.12 cm (P＜0.05); the LVEF of the MSCs group after operation was 63±6%, and remarkably higher than that of the model group,which was 53±6% (P＜0.05). Among the 10 cases of animals that survived for 4 weeks in the MSCs group, in 8 cases (80%),the transplanted cells survived in the non MI, MI region and its periphery, and even farther away; part of them differentiated into cardiomyocytes; in 7 cases (70%), the transplanted cells participated in the formation of blood vessel tissue in the MI region. Conclusion: Transplanted allograftic MSCs can survive and differentiate into cardiomyocytes, form the blood vessels in the MI region. MSCs transplantation could improve the heart function after MI.  相似文献   

9.

Objective  

To explore the effects of insulin-like growth factor-1 (IGF-1) on migration, proliferation and differentiation of mesenchymal stem cells (MSCs).  相似文献   

10.
Gene and drug therapies are being developed to alleviate vision loss in patients with Stargardt’s disease and age-related macular degeneration (AMD). To evaluate the therapeutic effects of these treatments, organic solvents are routinely used to extract and quantify bisretinoid lipofuscin constituents, such as N-retinylidene-N-retinyl-ethanolamine (A2E) and all-trans-retinal dimer (ATR-dimer). By high-performance liquid chromatography (HPLC), we found that A2E and ATR-dimer were both altered by tetrahydrofuran (THF) and chloroform, but were stable in dimethyl sulfoxide (DMSO) or methanol (MeOH). In addition, cyclohexane and ethanol (EtOH) did not alter ATR-dimer, whereas an alteration of A2E occurred in EtOH. On the basis of these findings, we designed processes II–IV, generated by modifications of process I, a routine method to measure bisretinoid compounds in vivo. Extra amounts of either ATR-dimer or A2E in mouse eyecups were released by processes II–IV versus process I. Efforts to clarify the effects of organic solvents on lipofuscin pigments are important because such studies can guide the handling of these fluorophores in related experiments.  相似文献   

11.
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12.
Type Ⅰ,Ⅲ and Ⅴ collagens were extracted from bovine dermis and cornea by using pepsin treatment in acetic acid solution,followed by salt precipitation and dialysis,to purify and isolate each type of collagens.The preparation process was analyzed by using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE).A reducing agent,2-mercaptoethanol,was used to remove disulfide bonds and analyze the structure of the bonds involved between α chains in some types of collagens.The use of delayed reducing methods resulted in the difference between α1(Ⅲ) and α1(Ⅰ) chains in a mixture containing type Ⅰ and Ⅲ collagens.The structure of disulfide bonds among α chains exists potentially in type Ⅴ collagen prepared from the pepsin-treatment extraction at 4℃,which differs from type Ⅲ collagen in relation to the locations of disulfide bonds.Compared with pepsin-treated collagen at 4℃,the relative molecular weights of α1(Ⅴ) and α2(Ⅴ) chains treated at room temperature decrease by 4.6% and 6.0%,respectively.It is concluded that type Ⅰ,Ⅲ and Ⅴ collagens can be prepared from bovine dermis and cornea by the use of pepsin treatment,salt precipitation and dialysis.The interchain disulfide bonds lie potentially near the edges of termini of type Ⅴ collagen molecules in extracellular matrix,and a small number of interchain crosslinks exist in type Ⅴ collagen.  相似文献   

13.

Objective  

Human embryonic stem cells (hESCs) have recently been reported as an unlimited source of mesenchymal stem cells (MSCs). The present study not only provides an identical and clinically compliant MSC source derived from hESCs (hESC-MSCs), but also describes the immunomodulative effects of hESC-MSCs in vitro and in vivo for a carbon tetrachloride (CCl4)-induced liver inflammation model.  相似文献   

14.
Surface display is effectively utilized to construct a whole-cell biocatalyst. Codon optimization has been proven to be effective in maximizing production of heterologous proteins in yeast. Here, the cDNA sequence of Rhizopus oryzae lipase (ROL) was optimized and synthesized according to the codon bias of Saccharomyces cerevisiae, and based on the Saccharomyces cerevisiae cell surface display system with α-agglutinin as an anchor, recombinant yeast displaying fully codon-optimized ROL with high activity was successfully constructed. Compared with the wild-type ROL-displaying yeast, the activity of the codon-optimized ROL yeast whole-cell biocatalyst (25 U/g dried cells) was 12.8-fold higher in a hydrolysis reaction using p-nitrophenyl palmitate (pNPP) as the substrate. To our knowledge, this was the first attempt to combine the techniques of yeast surface display and codon optimization for whole-cell biocatalyst construction. Consequently, the yeast whole-cell ROL biocatalyst was constructed with high activity. The optimum pH and temperature for the yeast whole-cell ROL biocatalyst were pH 7.0 and 40 °C. Furthermore, this whole-cell biocatalyst was applied to the hydrolysis of tributyrin and the resulted conversion of butyric acid reached 96.91% after 144 h.  相似文献   

15.
Background： Endothelial and smooth muscle cells were used as seeding cells and heterogeneous acellularized matrix was used as scaffold to construct the tissue-engineered graft. Methods： A 2 weeks piglet was selected as a donor of seeding cells. Two-centimetre length of common carotid artery was dissected. Endothelial cells and smooth muscle cells were harvested by trypsin and collagenase digestion respectively. The isolated cells were cultured and expanded using routine cell culture technique. An adult sheep was used as a donor of acellularized matrix. The thoracic aorta was harvested and processed by a multi-step decellularizing technique to remove the original cells and preserve the elastic and collagen fibers. The cultured smooth muscle cells and endothelial cells were then seeded to the acellularized matrix and incubated in vitro for another 2 weeks. The cell seeded graft was then transplanted to the cell-donated piglet to substitute part of the native pulmonary artery. Results： The cultured cells from piglet were characterized as endothelial cells by the presence of specific antigens vWF and CD31, and smooth muscle cells by the presence of specific antigen a-actin on the cell surface respectively with immunohistochemical technique. After decellularizing processing for the thoracic aorta from sheep, all the cellular components were extracted and elastic and collagen fibers kept their original morphology and structure. The maximal load of acellular matrix was decreased and 20% lower than that of untreated thoracic aorta, but the maximal tensions between them were not different statistically and they had similar load-tension curves. Three months after transplantation, the animal was sacrificed and the graft was removed for observation. The results showed that the inner surfaces of the graft were smooth, without thrombosis and calcification. Under microscopy, a great number of growing cells could be seen and elastic and collagen fibers were abundant. Conclusion： Cultured self-derived endothelial and smooth muscle cells could be used as seeding cells and heterogeneous acellularized matrix could be used as scaffold in constructing tissue-engineered graft.  相似文献   

16.
Several different pharmacological effects have been described for Nigella sativa (Siah-Daneh), including an anti-inflammatory effect. In the present study, the effect of the extract of N. sativa on lung pathology and blood interleukin-4 (IL-4) and interferon-γ (IFN-γ) of sensitized guinea pigs was examined. Three groups (n=8 for each group) of guinea pigs sensitized to ovalbumin (OA) were given drinking water alone, and drinking water containing low and high concentrations of the plant extract, respectively. The animals of the control group (n=8) were treated with saline instead of OA and were given drinking water. The pathological changes of the lung, including infiltration of eosinophils and lymphocytes, local epithelial necrosis, the presence of oedema, thickening of the basement membrane, smooth muscle layer hypertrophy, mucosal secretion, and the presence of mucosal plug, and blood IL-4 and IFN-γ of sensitized guinea pigs were evaluated. The lungs of the sensitized group showed significant pathological changes (P<0.001). Blood IL-4 and IFN-γ were increased in sensitized animals compared to the controls (P<0.01 and P<0.001, respectively). Treatment of sensitized animals with the extract led to a significant decrease in pathological changes of the lung (P<0.01 to P<0.001), except for the oedema in the sensitized group treated with low concentration of the extract, but an increased IFN-γ. These results confirm a preventive effect of N. sativa extract on lung inflammation of sensitized guinea pigs.  相似文献   

17.
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18.
Objective: Large segmental bone defect repair remains a clinical and scientific challenge with increasing interest focusing on combining gene transfection with tissue engineering techniques. The aim of this study is to investigate the effect of connective tissue growth factor (CTGF) on the proliferation and osteogenic differentiation of the bone marrow mesenchymal stem cells (MSCs). Methods: A CTGF-expressing plasmid (pCTGF) was constructed and transfected into MSCs. Then expressions of bone morphogenesis-related genes, proliferation rate, alkaline phosphatase activity, and mineralization were examined to evaluate the osteogenic potential of the CTGF gene-modified MSCs. Results: Overexpression of CTGF was confirmed in pCTGF-MSCs. pCTGF transfection significantly enhanced the proliferation rates of pCTGF-MSCs (P<0.05). CTGF induced a 7.5-fold increase in cell migration over control (P<0.05). pCTGF transfection enhanced the expression of bone matrix proteins, such as bone sialo-protein, osteocalcin, and collagen type I in MSCs. The levels of alkaline phosphatase (ALP) activities of pCTGF-MSCs at the 1st and 2nd weeks were 4.0- and 3.0-fold higher than those of MSCs cultured in OS-medium, significantly higher than those of mock-MSCs and normal control MSCs (P<0.05). Overexpression of CTGF in MSCs enhanced the capability to form mineralized nodules. Conclusion: Overexpression of CTGF could improve the osteogenic differentiation ability of MSCs, and the CTGF gene-modified MSCs are potential as novel cell resources of bone tissue engineering.  相似文献   

19.
目的:探讨外源性EGFP基因转染修饰兔骨髓间充质干细胞(m esenchymal stem cells,MSCs)的可行性。方法:用增强型荧光绿色蛋白(enhanced green fluorescent protein,EGFP)作为报告基因,以脂质体法转染骨髓间充质干细胞,观察转染结果、表达情况及对靶细胞活力的影响。结果:经荧光显微镜下观察证实:成功地对骨髓间充质干细胞实现了基因转染。结论:兔骨髓间充质干细胞能够在体外适宜的条件下进行长期培养;骨髓间充质干细胞可直接作为基因靶细胞,建立稳定表达EGFP的骨髓间充质干细胞系具有可行性,为进一步做标记的MSCs细胞的移植研究奠定基础。  相似文献   

20.
Therapeutically delivered mesenchymal stem cells (MSCs) improve ventricular remodeling. However,the mechanism underlying MSC cardiac remodeling has not been clearly determined. Congestive heart failure (CHF) was induced in rats by cauterization of the left ventricular free wall. MSCs were cultured from autologous bone marrow and injected into the border zone and the remote myocardium 5 d after injury. Ten weeks later,when compared with sham operation,CHF significantly increased nucleus mitotic index,capilla...  相似文献   

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