ABSTRACT
BACKGROUND: Most of hematopoietic growth factor regulates proliferation and differentiation of blood cells through JAKs-STATs signal transduction pathway. Total saponins of Panax ginseng (TSPG) can promote in vitro differentiation of CD34+ hematopoietic progenitor cells into erythroid cells, with similar effectiveness of hematopoietic growth factor.Erythropoietin receptor (EpoR) expression on the cell membrane of progenitor cells is critical during the erythroid differentiation process.OBJECTIVE: To investigate the molecular mechanism of TSPG to induce erythroid cells through erythropoiesis and its receptor-mediated JAK2/STAT5 signal transduction.DESIGN, TIME AND SETTING: An in vitro cytological observation. The study was performed at the Department of Histology and Embryology, Institute of Basic Medicine, Chongqing Medical University from May 2006 to October 2008.MATERIALS: Umbilical cord blood of normal full-term pregnancy was provided by the First Hospital of Chongqing Medical University. TSPG, purity＞95%, provided by Chongqing Institute of Traditional Chinese Medicine, was diluted in RPMI-1640 for work concentration of 1 g/L and degermed by positive pressure filtration.in RPMI-1640 culture solution containing horse serum, with various dilutions of TSPG (0 as blank control, 10, 25, 50, 75,100 mg/L). The MNCs were cultured on 96-well culture plate, with 0.2 mL in each well. Early erythroid cells were counted on were harvested and cultured separately in RPMI-1640 culture solution containing 10% horse serum as control group and in TSPG (25 mg/L)- conditioned culture system as experimental group. 5 U/mL Epo was added for 0, 2, 5 and 30 minutes.Immunoprecipitation of JAK2/STAT5 was used for the effect of TSPG on Epo/EpoR-induced tyrosine phosphorylation of JAK2/STAT5.MAIN OUTCOME MEASURES: Effect of TSPG on proliferation of erythroid progenitor cells from human umbilical cord blood;Effect of Epo on the proliferation of hematopoietic cells; Effect of TSPG on EpoR expression of the umbilical blood cells; tyrosine phosphorylations of JAK2 and STAT5.RERULTS: TSPG (10-75 mg/L) promoted the colony formation of BEU-E, CFU-E, and the preferential differentiation into erythroid lineage cells was most induced from 25 mg/L of TSPG. Using the colorimetric MTT assay, MNCs exhibited proliferative responses to Epo (2-50 U/mL) reaching maximum at 5 U/mL Epo. The addition of TSPG did not increase the expression of EpoR after MNCs were incubated in the presence of with or without TSPG for 24 hours. The pretreatment with TSPG for 24 hours enhanced Epo-induced tyrosine phosphorylation of JAK2 and STAT5 (STAT5a and STAT5b).
ABSTRACT
Objective:To clarify further the mechanisms for total saponins of Panax ginseng(TSPG)in inducing differentiation of K562 cells,and to provide the theoretical basis and the experimental evidence of its clinical application.Methods:By using cell culture in vitro,morphological observation and flow cytometry,the effect of TSPG on K562 cells in expressing CD 15 ,HIR 2,EPO-R and GM-CSF-R were studied.Results:The results indicated that TSPG could induce the differentiation of K562 cells toward proerythroid cells and progranulocytic cells.It was also shown that after induction by TSPG,the ratio of positive K562 cells expressing CD 15 ,HIR 2,erythropoietin recepter(EPO-R) and granulocyte-macrophage colony stimulating facter recepter(GM-CSF-R) increased.Conclusion:K562 cells are abnormal cells whose differentiation are blocked,but its biological features are similar to the normal hematopoietic stem cells.The mechanism for TSPG inducing differentiation of K562 cells may be related to the higher expression of EPO-R and GM-CSF-R.
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Objective:Our purpose is to clarify the mechanisms for total saponins of Panax ginseng(TSPG) inducing K562 cells to apoptosis.Methods:By using morphological observation and TUNEL,the effect of TSPG on apoptosis of K562 cells were studied.The expression of Fas was studied by immunocytochemistry,and sfas was assayed by ELISA.Results:The results indicated that 50,100?g/ml TSPG could induce K562 cells to apoptosis.Our experiment also showed after induced by TSPG,the ratio of positive K562 cells of expression Fas is increased ( P
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Object To clarify the mechanism for total saponins of Panax ginseng C.A.Mey. (TSPG) inducing K562 cells to apoptosis, and to provide the theoretical basis and the experimental evidence of TSPG's clinical application. Methods By using in vitro cell culture, morphometry, flow cytometry, morphological investigation and immunocytochemistry, the effects of TSPG on apoptosis in K562 cells were studied. Results The results indicated that TSPG could inhibit the proliferation of K562 cells significantly, and induce K562 cells to apoptosis. It was also showed in the experiments that after induced by TSPG, the ratio of positive K562 cells expressing C-MYC and BCL-2 is decreased. Conclusion The mechanism for TSPG to induce K562 cells to apoptosis may be related to the expression of oncogene in K562 cells.
ABSTRACT
Object To study the effect of cryopreservation on the ability of hematopoietic stem cells from human bone marrow and the action of total saponins of Panax ginseng (TSPG) to reduce their freezing injury. Methods Gradual cooling methods were used to place mononuclear cells (MNC) in liquid nitrogen with 5% dimethylsulfoxide (DMSO) for 3—5 months. After thawed, the biological properties of MNC were monitored, which included the mean trypan blue exclusion rate and the mean recovery rate of MNC, CFU-Mix, CD34+ cell, and total cells; Thawed MNC were cultured with various concentrations of TSPG, the effect of TSPG on the recoverable ability from cryopreservation damage were detected by colony-forming assay, colorimetric MTT assay for cell proliferation and flow cytometry. Results A fraction of MNC lost their proliferatory ability after thawed, but the damage was not deteriorated with freezing time. TSPG (25 ?g/mL) could raise the colony production rate of thawed hematopoietic stem cells; TSPG (25—50 ?g/mL) could raise the colony production rate of thawed hematopoietic stem cells; TSPG (25—50 ?g/mL) could improve their proliferation; TSPG (25 ?g/mL) could also promote the entray of them into cell proliferatory cycle. Conclusion TSPG could induce the proliferation of thawed hemato- poietic stem cells and raise the post-freezing recoverable ability of hematopoietic stem cells.