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1.
Chinese Medical Journal ; (24): 4009-4013, 2012.
Article in English | WPRIM | ID: wpr-339909

ABSTRACT

<p><b>BACKGROUND</b>Calcium and phosphorus metabolic disturbance are common in dialysis patients and associated with increased morbidity and mortality. Therefore, maintaining the balance of calcium and phosphate metabolism and suitable intact parathyroid hormone (iPTH) level has become the focus of attention. We investigated the effects of different peritoneal dialysate calcium concentrations on calcium phosphate metabolism and iPTH in continuous ambulatory peritoneal dialysis (CAPD) patients.</p><p><b>METHODS</b>Forty stable CAPD patients with normal serum calcium were followed for six months of treatment with 1.25 mmol/L calcium dialysate (DCa1.25, PD4, 22 patients) or a combination of 1.75 mmol/L calcium dialysate (DCa1.75, PD2) and PD4 (18 patients) twice a day respectively. Total serum calcium (after albumin correction), serum phosphorus, iPTH, alkaline phosphatase (ALP) and blood pressure were recorded before and 1, 3 and 6 months after treatment commenced.</p><p><b>RESULTS</b>No significant difference was found in baseline serum calcium, phosphorus between the two patient groups, but the levels of iPTH were significantly different. No significant changes were found in the dosage of calcium carbonate and active vitamin D during 6 months. In the PD4 group, serum calcium level at the 1st, 3rd, 6th months were significantly lower than the baseline (P < 0.05). There was no significant difference in serum phosphorus after 6 months treatment. iPTH was significantly higher (P < 0.001) at the 1st, 3rd, and 6th months compared with the baseline. No differences were seen in ALP and blood pressure. In the PD4+PD2 group, no significant changes in serum calcium, phosphorus, iPTH, ALP and BP during the 6-month follow-up period.</p><p><b>CONCLUSIONS</b>Treatment with 1.25 mmol/L calcium dialysate for six months can decrease serum calcium, increase iPTH, without change in serum phosphorus, ALP, and BP. The combining of PD4 and PD2 can stabilize the serum calcium and avoid fluctuations in iPTH levels.</p>


Subject(s)
Aged , Alkaline Phosphatase , Metabolism , Blood Pressure , Physiology , Calcium , Metabolism , Chelating Agents , Female , Humans , Male , Middle Aged , Peritoneal Dialysis , Methods , Phosphorus , Metabolism , Retrospective Studies
2.
Article in Chinese | WPRIM | ID: wpr-282606

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the roles of human fetal liver stromal cells (hFLSCs) and human fetal bone marrow stromal cells (hFBMSCs) in the hematopoietic differentiation of human embryonic stem cells and analyze their gene expression profile changes.</p><p><b>METHODS</b>The embryonic bodies on day 4 were cocultured with hFLSCs or hFBMSC in the presence of cytokines. Flow cytometry was performed after 8 days of induction to detect the expressions of the hemangioblast markers KDR and CD34, and the differential gene expression profiles between hFBMSC and hFLSCs were examined by cDNA microarray analysis.</p><p><b>RESULTS</b>Eight days after the induction, (1.06-/+0.20)% of the hFLSCs and (8.8-/+1.49)% of the hFBMSCs were positive for KDR, with the positivity rates for CD34 of (1.25-/+0.16)% and (9.17-/+2.10)%, respectively. In hFLSCs and hFBMSCs cultures, 0.9-/+0.36 and 10.6-/+0.63 hemagioblast-like cell colonies were found, respectively. cDNA microarray analysis showed that 240 genes were highly expressed in hFBMSCs, and 21 genes related to secreted cytokines, cell adhesion molecules and extracellular matrix proteins were highly expressed.</p><p><b>CONCLUSION</b>The microenvironment including the cell matrix protein and cytokines secreted by the hFBMSCs might play an important role in hemangioblastic differentiation of human bone marrow stromal cells in vitro.</p>


Subject(s)
Antigens, CD34 , Genetics , Metabolism , Bone Marrow Cells , Cell Biology , Cell Differentiation , Physiology , Coculture Techniques , Embryonic Stem Cells , Cell Biology , Fetus , Gene Expression Profiling , Hematopoietic Stem Cells , Cell Biology , Humans , Liver , Cell Biology , Oligonucleotide Array Sequence Analysis , Stromal Cells , Physiology , Vascular Endothelial Growth Factor Receptor-2 , Genetics , Metabolism
3.
Article in Chinese | WPRIM | ID: wpr-233731

ABSTRACT

<p><b>OBJECTIVE</b>To characterize the time course of spontaneous differentiation of in vitro cultured human embryonic stem cells (hESCs) into hematopoietic cells to provide experimental evidence for induction of hematopoietic commitment of hESCs.</p><p><b>METHODS</b>In human embryoid bodies (hEBs) derived from spontaneous differentiation of chESC3, a hESC cell line we established previously, the expressions of such genes as KDR, Bmi1, Scl and gata2 were detected by RT-PCR every other day during the 12-day differentiation to monitor the process of the hematopoiesis. The hematopoietic stem cell marker CD34 was examined using flow cytometry to evaluate the efficiency of hematopoietic differentiation of the cells on days 6, 8, 10 and 12. The spontaneously differentiated hESCs were seeded in the hematopoietic colony culture system to study the hematopoietic colony forming ability. Immunocytochemical staining for CD45 was performed on the hEBs to examine the emergence of mature hematopoietic cells.</p><p><b>RESULTS</b>The expressions of the hematopoietic stem cell-related genes KDR and Bmi-1 were detected in the hESCs, and on days 4 to 6, the two genes were upregulated with prolonged cuture of the hEBs. Scl and gata2 gene expressions were detected since 6-8 days of culture and maintained high expressions till day 12. Flow cytometry revealed a gradual increase in CD34-positive cells in the culture, with positivity rates on days 6, 8, 10, and 12 of (1.4-/+0.4)%, (3.4-/+1.3)%, (5.5-/+2.2)%, and (5.1-/+1.7)%, respectively. The numbers of CD43-positive cell colonies on days 6, 8, 10, and 12 were 0, 7-/+2, 37-/+11, and 89-/+29 in each 10(5) cells, respectively. Immunocytochemical staining identified CD45-positive cells on days 10, 12, 15, and 18 in the cell colonies, with the positive cell numbers of 0, 40.5-/+15.09, 178.6-/+55.89, and 253.0-/+52.04, respectively.</p><p><b>CONCLUSION</b>The hESCs undergo spontaneous hematopoietic differentiation in 3 stages, including the differentiation into germ layer-specific cells (days 6-8), expansion period of the hematopoitic progenitors (days 8-12), and maturation of the hematopoietic cells (after day 15).</p>


Subject(s)
Animals , Antigens, CD34 , Metabolism , Basic Helix-Loop-Helix Transcription Factors , Genetics , Cell Culture Techniques , Cell Differentiation , Embryonic Stem Cells , Cell Biology , Metabolism , GATA2 Transcription Factor , Genetics , Gene Expression Regulation , Hematopoietic Stem Cells , Cell Biology , Metabolism , Humans , Mice , Nuclear Proteins , Genetics , Polycomb Repressive Complex 1 , Proto-Oncogene Proteins , Genetics , Repressor Proteins , Genetics , T-Cell Acute Lymphocytic Leukemia Protein 1 , Time Factors
4.
Article in Chinese | WPRIM | ID: wpr-814097

ABSTRACT

OBJECTIVE@#To observe the inductive efficiency of deriving hematopoietic colony-forming cells from murine embryonic stem (mES) cells co-cultured with bone marrow stromal cell-conditional medium (mBMEC-CM).@*METHODS@#After the day-4 embryoid bodies (4 dEBs) were derived from embryonic stem cell-D3 (ES-D3) cells, the cells of 4 dEBs were induced into hematopoietic colony-forming cells by co-culturing with mBMEC-CM. The numbers of 4 dEB-derived hematopoietic colonies (high proliferation potential-colony formation cells and burst forming unit-erythroid, HPP-CFC and BFU-E) were detected to explore the relation between the implanted 4 dEB-derived cell numbers and the colony numbers of BFU-E and HPP-CFC. The inducing effect of mBMEC-CM was observed according to the doses and days of induction.@*RESULTS@#The number of 4 dEB-derived cells within 1 x 10(7)-4 x 10(7)/L was positively related to the colony numbers of HPP-CFC and BFU-E (HPP-CFC, r=0.916,P< 0.05; BFU-E, r=0.927, P<0.05). The inducing doses of mBMEC-CM within 0-20% were positively related to the colony numbers of HPP-CFC and BFU-E (HPP-CFC, r=0.909, P<0.05; BFU-E, r=0.927, P<0.01). The colony numbers of HPP-CFC and BFU-E derived from the 4 dEB-derived cells were the highest after 3 days of induction, followed by those of 6 days and 9 days.@*CONCLUSION@#Bone marrow endothelial cell-conditional medium can promote the generation of HPP-CFC and BFU-E from murine embryonic stem cells.


Subject(s)
Animals , Bone Marrow Cells , Cell Biology , Cell Differentiation , Cells, Cultured , Coculture Techniques , Culture Media, Conditioned , Embryonic Stem Cells , Cell Biology , Endothelial Cells , Cell Biology , Hematopoietic Stem Cells , Cell Biology , Mice , Stromal Cells , Cell Biology
5.
Article in Chinese | WPRIM | ID: wpr-813962

ABSTRACT

OBJECTIVE@#To observe the inductive efficiency of deriving hematopoietic cells from human embryonic stem (hES) cells co-cultured with human yolk sac stromal cells, fetal liver stromal cells or fetal bone marrow stromal cells,in order to discuss the effect of the different hemopoietic microenvironment on hemopoietic cytogenesis.@*METHODS@#We used two-step method to induce the hES cells into the hematopoietic cells. In the first step the hES cells were co-cultured with cytokines by formation of the day 5 embryoid bodies (5d EBs). In the second step the 5d EB cells were induced into the hematopoietic cells by co-culturing with human yolk sac stromal cells, fetal liver stromal cells or fetal bone marrow stromal cells for 10 days. The inductive efficiencies of deriving hematopoietic cells from hES cells co-cultured with the different hemopoietic microenvironment were reflected by the expression levels of flk, CD34 and CD45 antigen.@*RESULTS@#Flow cytometry analysis demonstrated that the population of the cells co-cultured with human yolk sac stromal cells contained flk (1.80%+/-0.56%), CD34 (1.30%+/-0.14%) or CD45 (1.05%+/-0.63%) positive cells; the population of the cells co-cultured with human fetal liver stromal cells contained flk (34.00%+/-25.45%), CD34 (38.40%+/-24.80%) or CD45 (72.60%+/-25.70%) positive cells; the population of the cells co-cultured with human fetal bone marrow stromal cells contained flk (2.50%+/-1.48%), CD34 (3.20%+/-0.56%) or CD45 (1.65%+/-0.21%) positive cells. Compared with spontaneous differentiation of EBs, all of the three stromal cells could induce EBs into the hematopoietic cells (P<0.05).@*CONCLUSION@#The inductive efficiency of deriving hematopoietic cells from EBs co-cultured with human fetal liver stromal cells was higher than EBs co-cultured with human yolk sac stromal cells and fetal bone marrow stromal cells.


Subject(s)
Antigens, CD34 , Cell Differentiation , Cells, Cultured , Cellular Microenvironment , Coculture Techniques , Embryonic Stem Cells , Cell Biology , Fetus , Cell Biology , Hematopoietic Stem Cells , Cell Biology , Humans , Leukocyte Common Antigens , Mesenchymal Stem Cells , Cell Biology , Stromal Cells , Cell Biology , Yolk Sac , Cell Biology
6.
Article in Chinese | WPRIM | ID: wpr-356588

ABSTRACT

Embryonic stem cells are pluripotent and their differentiation in vitro can serve as an experimental model to explore the molecular mechanisms of early embryonic development. To investigate the effect of stromal cell conditioned medium combined with cytokines (sccm + cys) on the differentiation from human embryonic stem cells to hematopoietic cells and endothelial cells, the mouse fibroblast feeder cells to make human embryonic stem cells grown into embryonic bodies (EBs) were initially deleted. After culture for 3 days, EB cells were trypsinized into single cells and induced for 8 days by sccm + cys. Then, the differentiated cells were cultured in the semisolid medium containing 0.9% methylcellulose and cytokines to study the colony forming and self-renewal ability of cells. Immunocytochemical staining was used to check the surface markers of the colony cells. During the induction, mRNA expression of flk-1, BMI-1, scl, and Zeta-globin genes was tested by RT-PCR. Surface markers, such as flk-1, CD34 were tested by the flow cytometry. The results demonstrated that: (1) cell clusters containing 20-30 cells were formed after culture for 8 - 14 days in the semisolid medium, replanting these cells resulted in similar cell cluster forming. In addition, CD45 positive in big cell colonies were also found in the semisolid medium; (2) attached cell colonies appeared after culture for 8 days in the semisolid medium and VIII factor, UEA and KDR could be detected as negative by immunocytochemical staining; (3) on the 4(th) day of induction, mRNAs of flk-1, BMI-1, scl and Zeta-globin were all expressed. On the 8(th) day of induction, all of the above genes except Zeta-globin were expressed, while ES cell and EB cells which served as controls did not express scl and Zeta-globin genes; (4) on the 8(th) day of induction, the proportions of flk-1(+) cells and CD34(+) cells among all the inducing population were 9.8% and 16.8%, respectively, while the corresponding positive populations were 0.36% and 1.16% in spontaneously differentiated 11(th) day's EB, and 0.04% and 0.16%, respectively, in ES cells. If is concluded that embryonic stem cells can differentiate into hematopoietic cells and endothelial cells in combinant culture system of this study.


Subject(s)
Cell Differentiation , Cells, Cultured , Embryonic Stem Cells , Cell Biology , Endothelial Cells , Cell Biology , Flow Cytometry , Globins , Genetics , Metabolism , Hematopoietic Stem Cells , Cell Biology , Humans , Immunohistochemistry , Nuclear Proteins , Genetics , Metabolism , Polycomb Repressive Complex 1 , Proto-Oncogene Proteins , Genetics , Metabolism , Repressor Proteins , Genetics , Metabolism , Reverse Transcriptase Polymerase Chain Reaction , Time Factors , Vascular Endothelial Growth Factor Receptor-2 , Genetics , Metabolism
7.
Article in Chinese | WPRIM | ID: wpr-266802

ABSTRACT

<p><b>OBJECTIVE</b>To establish the determination method of 2,3,5,4'-tetrahydroxystilbene-2-O-beta-D-glucoside in Yangan Oral Liquid.</p><p><b>METHOD</b>2,3,5,4'-tetrahydroxystilbene-2-O-beta-D-glucoside was determined by HPLC on C18 column, in acetonitrile-water (14:86) solution as a mobile phase, detection wavelength as 320 nm.</p><p><b>RESULT</b>This method showed a good linear relationship. The average recovery was 99.05% and RSD was 1.31%.</p><p><b>CONCLUSION</b>The method was simple with styong specificity and good repriducibility and can be used for quality control for Yangan Oral Liquid.</p>


Subject(s)
Chromatography, High Pressure Liquid , Methods , Drug Combinations , Drugs, Chinese Herbal , Chemistry , Glucosides , Plants, Medicinal , Chemistry , Polygonum , Chemistry , Quality Control , Stilbenes
8.
Article in Chinese | WPRIM | ID: wpr-355704

ABSTRACT

The purpose of this study was to observe the bone marrow endothelial cell-conditioned medium (BECM) and cytokines, i.e. vascular endothelial growth factor (VEGF), stem cell factor (SCF) and EPO promoting the generation of hematopoietic precursor cells from mouse embryonic stem cells (ESC) in vitro. Day 4 embryoid body (4dEB) cells were derived from ESC-D3 cell line, a murine ESC line, and then induced with BECM and/or cytokines. Four groups, i.e. BECM, BECM + VEGF + SCF + EPO, VEGF + SCF + EPO and control (spontaneous differentiation), were designed. Immunochemistry staining and flow cytometry were adopted to observe the antigen expression, RT-PCR to detect hematopoietic transcription factors, and hematopoietic progenitor assay to examine hematopoietic differentiation. The results showed that the cells induced from ESC expressed hematopoietic precursor cell antigens (c-kit, Sca-1, Thy-1 and CD34), transcription factors (c-myb, SCL and beta-H1) and generated HPP-CFC and BFU-E. The effect of BECM + VEGF + SCF + EPO was the most potent in the inducing groups according to the numbers of hematopoietic precursor cells and colonies. It is concluded that BECM promotes the differentiation of ESC into hematopoietic precursor cells in vitro, and this effect is the strongest when BECM combining with VEGF + SCF + EPO.


Subject(s)
Animals , Cell Differentiation , Culture Media, Conditioned , Embryo, Mammalian , Cell Biology , Endothelial Cells , Physiology , Erythropoietin , Pharmacology , Female , Hematopoietic Stem Cells , Cell Biology , Mice , Stem Cell Factor , Pharmacology , Stem Cells , Cell Biology , Vascular Endothelial Growth Factor A , Pharmacology
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