1.Effects of human ERMAP-siRNA on erythroid differentiation of K562 cells induced by Ara-C.
Jie-Fang LIANG ; Ying CHEN ; Tie-Zhen YE ; Ying-Yi HE ; Xin-Rong HE ; Li-Dan LIN ; Sai-Jun GAO
Journal of Experimental Hematology 2009;17(1):49-53
In order to investigate the potential role of human ERMAP gene in erythropoiesis, the ERMAP-dsDNA was designed, ERMAP-shRNA expressing plasmids was constructed, and ERMAP-shRNA/K562 cell was established. Cell morphology, biphenylamine staining, expression of cell surface antigens as well as quantitative level of human ERMAP gene were observed during K562 cells differentiating toward erythroid lineage induced by Ara-C. The results showed that at 72 hours after Ara-C treatment, ERMAP-shRNA/K562 cell size became large with increasing cytoplasm content. The percentage of biphenylamine positive cells increased from 1.17% to 2.04% (p < 0.05), but still lower than that in group K562 + Ara-C. The percentage of CD36(-)/CD235a(+) increased from 8.83% to 11.28%, CD36(+)/CD235a(+) increased from 1.23% to 2.64%, and CD36(+)/CD235a(-) increased from 0.59% to 1.47% respectively, which were all lower than that in group K562 + Ara-C at either time point. At the same time, the level of ERMAP expression increased slowly from 2.52 x 10(-3) to 4.53 x 10(-3), which was also significantly lower than that of group K562 + Ara-C. It is concluded that the ERMAP-shRNA inhibits the Ara-C-induced erythroid differentiation of K562 cells, which further suggests that there is relationship between hERMAP and erythroid differentiation and development.
Blood Group Antigens
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genetics
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Butyrophilins
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Cell Differentiation
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drug effects
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Cytarabine
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pharmacology
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Erythropoiesis
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drug effects
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Humans
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K562 Cells
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RNA, Messenger
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RNA, Small Interfering
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pharmacology
2.Effect of zishen shengxue recipe on endogenous erythropoietin in patients with renal anemia undergoing long-term hemodialysis.
Jin-chuan TAN ; Jin-huan HU ; Jing-wen SU
Chinese Journal of Integrated Traditional and Western Medicine 2010;30(1):42-44
OBJECTIVETo investigate the mechanism of Zishen Shengxue Recipe (ZSR) in treating renal anemia by observing its effect on serum level of endogenous erythropoietin in patients undergoing long-term hemodialysis.
METHODSSixty renal anemia patients undergoing long-term hemodialysis were randomly and equally assigned to two groups. The treated group was treated with subcutaneous injection of erythropoiesis stimulating factor (rHuEpo) combined with oral intake of ZSR, and the control group treated with rHuEpo alone. They were observed for eight weeks, and the blood levels of endogenous human erythropoietin (Epo), hemoglobin (Hgb), hematocrit (Hct), as well as the residual renal function (RRF) in the two groups were compared.
RESULTSSerum Epo level in the control group was unchanged after treatment (P>0.05), while that in the treated group increased significantly, and showed significant difference in comparing with that in the control group (P<0.05). Levels of Hgb and Hct increased and RRF decreased in both groups (P<0.01), but the treated group showed higher increments and lesser decrement than those in the control group (P<0.05).
CONCLUSIONSZSR can enhance the blood levels of Hgb, Hct and Epo, postpone the descent of RRF, and correct the anemic status in patients. Its mechanism of action is possibly through alleviating the inhibition of uremic toxin on erythropoiesis, in the meanwhile of promoting the secretion of Epo.
Adult ; Anemia ; blood ; therapy ; Drugs, Chinese Herbal ; therapeutic use ; Erythropoiesis ; drug effects ; Erythropoietin ; blood ; Female ; Hematinics ; therapeutic use ; Hematocrit ; Hemoglobins ; analysis ; Humans ; Male ; Middle Aged ; Renal Dialysis
3.Effect of nerve growth factor on erythropoiesis in mice and its underlying mechanism.
Journal of Experimental Hematology 2008;16(6):1365-1371
This study was aimed to explore the effect of nerve growth factor (NGF) on erythropoiesis and to elucidate the underlying mechanism. Using flow cytometry, colony forming assay, blood cell counter, fluorescent real-time quantitation PCR, and enzyme-linked immunosorbent assay (ELISA), the changes in the bone marrow cells (BMCs) proliferation cycle, CFU-E and BFU-E counts, the peripheral blood erythroid related parameters, kidney EPO, BMC GM-CSF, spleen EPO receptor (EPOR) mRNA expression, and serum EPO, GM-CSF, and IL-1 concentrations were all determined after NGF was injected intramuscularly into the thigh of mice, meanwhile the change of BFU-E and CFU-E counts and its relationship with EPO, IL-3 were investigated. The results indicated that the cell proportion in S+G2/M phase, the CFU-E and BFU-E counts of BMCs and the spleen EPOR mRNA expression in injection of NGF (7.5 microg/kg) for 7 days were significantly higher than that in injection of physiological saline for 13-19 days; red blood cell, hemoglobin, and reticulocyte counts increased as well. In vitro, NGF stimulated a dose-dependent increase of CFU-E colonies formation in the semisolid culture system with or without exogenous EPO; the colony counts in the system with NGF alone were significantly higher than that in the system with exogenous EPO alone. The BFU-E counts in the system with exogenous NGF and IL-3 were significantly higher than that in the system with exogenous EPO and IL-3. It is concluded that the NGF promotes the responsibility of hematopoietic cells to EPO and activates the same signal transduction pathway as EPO in hematopoietic cells, and then accelerates the BMCs into mitosis, the HSCs differentiating into erythroid cells, and CFU-E and BFU-E formation.
Animals
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Bone Marrow Cells
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cytology
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drug effects
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Cell Proliferation
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drug effects
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Cells, Cultured
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Erythrocytes
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cytology
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metabolism
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Erythropoiesis
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Granulocyte-Macrophage Colony-Stimulating Factor
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blood
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Hematopoietic Stem Cells
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cytology
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drug effects
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Male
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Mice
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Mice, Inbred BALB C
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Nerve Growth Factor
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pharmacology
4.Effect of low-dose hydroxyurea with sodium butyrate on globin gene expression in human erythroid progenitor cells.
Qian-qian WU ; Xin-hua QIAN ; Mei-jia XU
Journal of Southern Medical University 2009;29(10):2073-2081
OBJECTIVETo investigate the effects of combined use of low-dose hydroxyurea (HU) and sodium butyrate (NaB) on the expression of 7 globin genes (zeta, alpha, epsilon, Ggamma, Agamma, delta, and beta) in human erythroid progenitor cells.
METHODSHuman erythroid progenitor cells were cultured using a two-step liquid culture system and treated with HU and NaB either alone or in combination. The inhibitory effects of the agents on the cell growth were monitored with trypan blue exclusion assay, and the changes in the mRNA of the 7 globin genes were detected using RT-PCR.
RESULTSLow-dose HU combined with NaB resulted in significantly lower inhibition rate of the erythroid progenitor cells than routine dose HU and NaB used alone (28.56% and 38.80%, respectively, P<0.05). Compared with untreated cells (0.653-/+0.092 and 0.515-/+0.048), HU combined with NaB significantly increased the expression of Ggamma-and Agamma- mRNA (1.203-/+0.018 and 0.915-/+0.088, respectively, P<0.05), and HU and NaB used alone produced similar effects (1.305-/+0.016 and 0.956-/+0.029 for HU, and 1.193-/+0.070 and 0.883-/+0.012 for NaB, P>0.05). HU and NaB, either used alone or in combination or at different doses, caused no significant changes in the other globin genes (zeta, alpha, epsilon, delta and beta) (P>0.05).
CONCLUSIONLow-dose HU combined with NaB can up-regulate gamma globin gene expression, especially Ggamma-mRNA expression, to decrease the growth inhibition on human erythroid progenitor cells in vitro, but produces no significant effect on the expressions of zeta, alpha, epsilon, delta and beta genes.
Anemia, Sickle Cell ; genetics ; Butyrates ; administration & dosage ; pharmacology ; therapeutic use ; Cells, Cultured ; Drug Therapy, Combination ; Erythroid Precursor Cells ; cytology ; drug effects ; physiology ; Erythropoiesis ; drug effects ; Humans ; Hydroxyurea ; administration & dosage ; pharmacology ; therapeutic use ; RNA, Messenger ; genetics ; metabolism ; gamma-Globins ; genetics ; metabolism
5.Effect of combination of bushen jianpi recipe and erythropoietin on serum tumor necrosis factor alpha in patients with anemia.
Qiu-fen LI ; Qiong-ying MA ; Cai-feng ZHU
Chinese Journal of Integrated Traditional and Western Medicine 2004;24(2):106-108
OBJECTIVETo study the effect of treatment of renal anemia by combination of Bushen Jianpi Recipe (BJR, a Chinese experience recipe for supplementing Shen and supporting Pi) and low dosage erythropoietin (EPO), and the influence of treatment on change of serum tumor necrosis factor alpha (TNF-alpha) so as to explore the possible mechanism of integrative Chinese and western medicine (ICWM) in treating renal anemia.
METHODSPatients with renal anemia were randomly divided into two groups, the ICWM group and the control group, symptomatic and supporting treatment such as dialysis, supplementing of ferrous, foliac acid and vitamin B12, was given to both groups. Additionally, to the ICWM group, 50 IU/kg of EPO subcutaneous injection for twice every week, and oral intake of BJR, one dose per day taken in two parts, were given, and to the control group, EPO alone, 50 IU/kg by subcutaneous injecting, 3 times per week, was given. The therapeutic course for two groups was 3 months. Blood levels of hemoglobin (Hb), hematocrit (Hct), TNF-alpha were measured before treatment and the therapeutic effect was observed.
RESULTSAfter treatment, the levels of Hb and Hct increased significantly in both groups (P < 0.01), comparison between the two groups in Hb and Hct after being treated for 3 months showed significant difference (P < 0.05). The serum level of TNF-alpha was markedly higher than normal range in both groups before treatment, it significantly lowered after treatment in the ICWM group (P < 0.05), but unchanged in the control group.
CONCLUSIONCombination of BJR and EPO could significantly inhibit the production of TNF-alpha, this may be an important factor for ICWM in effectively improving sensitivity to EPO.
Adult ; Anemia ; blood ; drug therapy ; etiology ; Chronic Disease ; Drug Therapy, Combination ; Drugs, Chinese Herbal ; therapeutic use ; Erythropoiesis ; drug effects ; Erythropoietin ; therapeutic use ; Female ; Glomerulonephritis ; blood ; complications ; drug therapy ; Humans ; Male ; Middle Aged ; Phytotherapy ; Recombinant Proteins ; Tumor Necrosis Factor-alpha ; metabolism
6.Role of Ras/ERK-dependent pathway in the erythroid differentiation of K562 cells.
Chi Dug KANG ; In Rok DO ; Kwang Woon KIM ; Byung Kwon AHN ; Sun Hee KIM ; Byung Seon CHUNG ; Byung Hak JHUN ; Mi Ae YOO
Experimental & Molecular Medicine 1999;31(2):76-82
The chronic myelogenous leukemic K562 cell line carrying Bcr-Abl tyrosine kinase is considered as pluripotent hematopoietic progenitor cells expressing markers for erythroid, granulocytic, monocytic, and megakaryocytic lineages. Here we investigated the signaling modulations required for induction of erythroid differentiation of K562 cells. When the K562 cells were treated with herbimycin A (an inhibitor of protein tyrosine kinase), ras antisense oligonucleotide, and PD98059 (a specific inhibitor of MEK), inhibition of ERK/MAPK activity and cell growth, and induction of erythroid differentiation were observed. The ras mutant, pZIPRas61leu-transfected cells, K562-Ras61leu, have shown a markedly decreased cell proliferation rate with approximately 2-fold doubling time, compared with the parental K562 cells, and about 60% of these cells have shown the phenotype of erythroid differentiation. In addition, herbimycin A inhibited the growth rate and increased the erythroid differentiation, but did not affect the elevated activity of ERK/MAPK in the K562-Ras61leu cells. On the other hand, effects of PD98059 on the growth and differentiation of K562-Ras61leu cells were biphasic. At low concentration of PD98059, which inhibited the elevated activity of ERK/MAPK to the level of parental cells, the growth rate increased and the erythroid differentiation decreased slightly, and at high concentration of PD98059, which inhibited the elevated activity of ERK/MAPK below that of the parental cells, the growth rate turned down and the erythroid differentiation was restored to the untreated control level. Taken together, these results suggest that an appropriate activity of ERK/MAPK is required to maintain the rapid growth and transformed phenotype of K562 cells.
Androstadienes/pharmacology
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Ca(2+)-Calmodulin Dependent Protein Kinase
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Cell Differentiation/drug effects
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Enzyme Inhibitors/pharmacology
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Erythroid Progenitor Cells/physiology*
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Erythroid Progenitor Cells/cytology
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Erythropoiesis*
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Flavones/pharmacology
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Human
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K562 Cells
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Leukemia, Myeloid/pathology
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Oligonucleotides, Antisense/pharmacology
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Quinones/pharmacology
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ras Proteins/metabolism*
7.Effects of Adjuvant Androgen on Anemia and Nutritional Parameters in Chronic Hemodialysis Patients Using Low-dose Recombinant Human Erythropoietin.
Myeung Su LEE ; Seon Ho AHN ; Ju Hung SONG
The Korean Journal of Internal Medicine 2002;17(3):167-173
BACKGROUND: Recombiant human erythropoietin (epoetin) has greatly contributed to improvement of the anemia of chronic renal failure patients on hemodialysis. However, the reduced erythropoietic effect to epoetin and its high cost have induced lots of supplementary treatments. Therefore, we performed a prospective study to evaluate the effects of adjuvant low-dose androgen therapy in patients using a lower-dose of epoetin than the commonly recommended dose on anemia and the nutritional parameters. METHODS: 17 patients of hemoglobin (Hgb) less than 9 g/dL even after being treated with 1,000 U epoetin subcutaneously (s.c.) 3 times per week on a stable status for more than 6 months, who were on hemodialysis at our institution were examined. They were injected with the same dose of epoetin s.c. and nandrolone decanoate 100 mg intramuscularly (i.m) weekly for another 6 months. Blood test was performed every month before therapy for 6 months and after therapy for 6 months and the mean values were reviewed for comparison. RESULTS: Hgb (7.75+/-0.9 vs 8.99+/-1.39 g/dL, p < 0.01) and hematocrit (Hct) (23.68+/-2.85 vs 26.66+/-3.91%, p < 0.01) were apparently changed before and after adjuvant therapy. Hgb and Hct, weekly dose of epoetin were not statistically different in 9 male patients before and after adjuvant therapy. The weekly dose of epoetin was not statistically different in 8 female patients, but Hgb and Hct (8.02+/-0.6 vs 9.72+/-1.31 g/dL, 24.54+/-1.7 vs 28.74+/-3.06%, p < 0.01) were statistically different before and after adjuvant therapy. In comparison between male and female groups, weekly doses of epoetin and nandrolone decanoate were significantly greater in the female group than the male group (epoetin; 50.66+/-6.23 vs 61.18+/-8.76 U/kg/week, nandrolone decanoate; 1.69+/-0.2 vs 2.04+/-0.29 mg/kg/week, p < 0.05). CONCLUSION: Our data show that the adjuvant androgen therapy is effective for the anemia of hemodialysis patients who did not recover from anemia even after being continuously treated with low-dose epoetin.
Adult
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Anabolic Steroids/*administration & dosage/blood
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Anemia, Aplastic/blood/*drug therapy
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Chemotherapy, Adjuvant
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Erythropoiesis/drug effects
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Erythropoietin, Recombinant/*administration & dosage/blood
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Female
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Human
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Kidney Failure, Chronic/blood/drug therapy
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Male
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Middle Age
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Nandrolone/*administration & dosage/blood
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Nutritional Status
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Prospective Studies
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*Renal Dialysis