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Fang Chen,
Kang Zhou,
Lei Zhang,
Fengxia Ma,
Dandan Chen,
Junjie Cui,
Xiaoming Feng,
Shaoguang Yang,
Ying Chi,
Zhi-Bo Han,
Feng Xue,
Lijuan Rong,
Meili Ge,
Li Wan,
Shuxia Xu,
Wenjing Du, Shihong Lu,
Hongying Ren,
Zongchao Han
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ABSTRACT: All-trans retinoic acid (ATRA) induces clinical remission in most acute promyelocytic leukemia (APL) patients by inducing terminal differentiation of APL cells toward mature granulocytes. Here we report that human umbilical cord-derived mesenchymal stem cells (UC-MSCs) are capable of inducing granulocytic differentiation of APL-derived NB4 cell line as well as primary APL cells and also cooperate with ATRA in an additive manner. Transwell co-culture experiments revealed that UC-MSCs' differentiation-inducing effect was mediated through some soluble factors. Differentiation attenuation by IL-6Ra neutralization and induction by addition of exogenous IL-6 confirmed that IL-6 secreted by UC-MSCs was at least partially responsible for this differentiation induction process. Moreover, we found that UC-MSCs activated MEK/ERK signaling pathway in promyelocytic cells and pharmacological inhibition of MEK/ERK pathway reversed UC-MSCs-induced differentiation, indicating that UC-MSCs exerted effect through activation of MEK/ERK signaling pathway. These results demonstrate for the first time a stimulatory effect of MSCs on the differentiation of APL cells and bring a new insight into the interaction between MSCs and leukemic cells. Our data suggest that UC-MSCs/ATRA combination could be used as a novel therapeutic strategy for APL patients.
Stem cells and development 02/2013; · 4.15 Impact Factor
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Dandan Chen,
Fengxia Ma,
Shuxia Xu,
Shaoguang Yang,
Fang Chen,
Lijuan Rong,
Ying Chi,
Qinjun Zhao, Shihong Lu,
Zhibo Han,
Aiming Pang,
Zhongchao Han
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ABSTRACT: BACKGROUND AIMS: Toll-like receptors (TLRs) play an important role in innate and adaptive immunity by recognizing pathogen-associated molecular patterns (PAMPs). METHODS: In the present study, we investigated the expression and role of TLRs on human umbilical cord mesenchymal stem cells (UC-MSCs). The proliferation, differentiation and immunoregulatory activity of UC-MSCs primed with or without TLR ligands were determined. RESULTS: At the RNA level, the expression of TLR2, 4, 6 and 9 was relatively higher than that of other TLRs. However, TLR3 and TLR4 expression were relatively higher at the protein level. UC-MSCs expressed functional TLRs by nuclear factor-κB activation and cytokine expression assay. Poly-inosinic acid:cytidylic acid [Poly(I:C)] stimulation inhibited the proliferation of UC-MSCs, but the ligand of other TLRs had no significant effect. Poly(I:C) stimulation enhanced the adipogenic differentiation capability of UC-MSCs, but lipopolysaccharide inhibited the adipogenic differentiation. Poly(I:C) and CpG-oligonucleotide promoted the immunosuppressive potentiality of UC-MSCs, accompanied with the phosphorylation of interferon regulatory factor 3 (IRF3) and increased expression of indoleamine 2,3-dioxygenase and interferon β, whereas activation of other TLR ligands (synthetic analog fibroblast-stimulating lipopeptide-1 and lipopolysaccharide) failed to affect the immunoregulatory activity of UC-MSCs. CONCLUSIONS: Taken together, our data demonstrated that TLR activation influenced the function of UC-MSCs, which might have important implications in future efforts to explore the clinical potentials of UC-MSCs.
Cytotherapy 01/2013; · 3.63 Impact Factor
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ABSTRACT: To isolate pancreatic progenitor cells with the surface markers of hematopoietic stem cells, the expression of stem cell antigen (Sca-1) and c-Kit and the coexpression of them with pancreatic duodenal homeobox-1 (PDX-1), neurogenin 3 (Ngn3), and insulin were examined in murine embryonic pancreas. Then different pancreatic cell subpopulations were isolated by magnet-activated cell sorting. Isolated cells were cultured overnight in hanging drops. When cells formed spheres, they were laid on floating filters at the air/medium interface. With this new culture system, pancreatic progenitor cells were induced to differentiate to endocrine and exocrine cells. It was shown that c-Kit and Sca-1 were expressed differently in embryonic pancreas at 12.5, 15.5, and 17.5 days of gestation. The expression of c-Kit and Sca-1 was the highest at 15.5 days of gestation. c-Kit rather than Sca-1 coexpressed with PDX-1, Ngn3, and insulin. Cells differentiated from c-Kit-positive cells contained more insulin-producing cells and secreted more insulin in response to glucose stimulation than that from c-Kit-negative cells. These results suggested that c-Kit could be used to isolate pancreatic progenitor cells and our new culture system permitted pancreatic progenitor cells to differentiate to mature endocrine cells.
International Journal of Endocrinology 01/2012; 2012:948683. · 1.87 Impact Factor
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Jianping Li,
Shaoguang Yang, Shihong Lu,
Hui Zhao,
Jianming Feng,
Wenqian Li,
Fengxia Ma,
Qian Ren,
Bin Liu,
Lei Zhang,
Yizhou Zheng,
Zhong Chao Han
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ABSTRACT: Aplastic anemia (AA) is generally considered as an immune-mediated bone marrow failure syndrome with defective hematopoietic stem cells (HSCs) and marrow microenvironment. Previous studies have demonstrated the defective HSCs and aberrant T cellular-immunity in AA using a microarray approach. However, little is known about the overall specialty of bone marrow mesenchymal stem cells (BM-MSCs). In the present study, we comprehensively compared the biological features and gene expression profile of BM-MSCs between AA patients and healthy volunteers. In comparison with healthy controls, BM-MSCs from AA patients showed aberrant morphology, decreased proliferation and clonogenic potential and increased apoptosis. BM-MSCs from AA patients were susceptible to be induced to differentiate into adipocytes but more difficult to differentiate into osteoblasts. Consistent with abnormal biological features, a large number of genes implicated in cell cycle, cell division, proliferation, chemotaxis and hematopoietic cell lineage showed markedly decreased expression in BM-MSCs from AA patients. Conversely, more related genes with apoptosis, adipogenesis and immune response showed increased expression in BM-MSCs from AA patients. The gene expression profile of BM-MSCs further confirmed the abnormal biological properties and provided significant evidence for the possible mechanism of the destruction of the bone marrow microenvironment in AA.
PLoS ONE 01/2012; 7(11):e47764. · 4.09 Impact Factor
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ABSTRACT: Heart failure is a leading cause of morbidity and mortality worldwide. The current strategies for treatment are limited, and new therapeutic approaches are needed. Experimental studies and clinical trials suggest that stem cell transplantation may improve cardiac function and prevent cardiac remodeling of the injured heart. Although the results of the studies were exciting, many problems remain to be resolved such as the best method of delivering the targeted cells. Direct injection into the myocardium and intracoronary artery infusion are the two most used methods of delivery in clinical settings. However, in a portion of patients with occluded coronary arteries and poor collaterals, transplanted cells may not reach the target ischemic lesion. To resolve this problem, we hypothesize that retrograde coronary venous delivery of stem cells may be a promising therapeutic strategy for the patients with occluded coronary arteries and poor collaterals.
Clinical Transplantation 09/2011; 25(6):830-3. · 1.67 Impact Factor
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Hongying Ren,
Qinjun Zhao,
Tao Cheng, Shihong Lu,
Zhong Chen,
Lei Meng,
Xiaofan Zhu,
Shaoguang Yang,
Wen Xing,
Yongdi Xiao,
Qian Ren,
Ying Chi,
Dongsheng Gu,
Renchi Yang,
Zhong Chao Han
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ABSTRACT: The acceleration of capillarization and venularization of hepatic sinusoids after cell therapy would not be beneficial to restoration after liver disease. The goal was to observe the effects of umbilical cord (UC)-derived mesenchymal stromal cells (MSC) on liver microcirculation and their therapeutic potential in liver fibrosis.
Human UC MSC labeled with or without CM-DIL were transplanted into NOD/SCID mice with carbon tetrachloride (CCl4)-induced chronic liver fibrosis models. Because of the high autofluorescence on the injured liver sections, we used immunohistochemistry, Western blot and reverse transcriptase-polymerase chain reaction (RT-PCR), but not immunofluorescence, in order to avoid false images under a confocal fluorescence microscope.
Human-specific alpha-fetoprotein and albumin mRNA and proteins were detected in CCl4-treated mouse livers receiving human UC MSC transplants. We only observed the gene expression of human-specific endothelial-like cells markers CD31 and KDR by RT-PCR, but not protein expression by immunohistochemistry, in UC MSC-transplanted mouse livers. Vascular endothelial growth factor (VEGF) expression in injured livers 4 weeks after UC MSC transplantation was higher than in normal livers. However, UC MSC injection did not increase significantly the vascular density labeled by CD31 and (vWF) in the injured livers of UC MSC-transplanted mice compared with non-transplanted mice after CCl4 treatment. In addition, liver function was partly improved after UC MSC transplantation.
Human UC MSC can differentiate into hepatocyte-like cells but do not accelerate the capillarization and venularization of hepatic sinusoids, finally leading to the partial improvement of liver function in mice with CCl4-mediated chronic liver fibrosis.
Cytotherapy 02/2010; 12(3):371-83. · 3.63 Impact Factor
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Dongsheng Gu,
Zhenping Chen,
Haifeng Zhao,
Weiting Du,
Feng Xue,
Jing Ge,
Tao Sui,
Hao Wu,
Bin Liu, Shihong Lu,
Lei Zhang,
Renchi Yang
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ABSTRACT: Immune thrombocytopenia (ITP) is an acquired organ-specific autoimmune disease with a polarization of T(h)1. Both the T(h)1 chemokine CXCL10 and T(h)2 chemokine CCL2 have been studied in several autoimmune diseases, but the status of these chemokines in ITP is still unknown. The aims of this study were to determine the expression of CXCL10 and CCL2 and their receptors, CXCR3 and CCR2, in ITP patients, and to conduct a preliminary study of the pathogenic roles of these factors in ITP. Plasma samples from 49 patients with ITP and 24 normal healthy subjects were assayed for CXCL10 and CCL2 plasma concentration by enzyme-linked immunosorbent assay. Real-time quantitative polymerase chain reaction was performed to determine the mRNA expression of these chemokines and their receptors in the PBMNC of 24 normal controls and 28 active ITP patients as well as splenocytes of nine ITP patients. The CXCL10 levels in the plasma samples from patients with active ITP were significantly higher than those from healthy controls (p = 0.007) and decreased to normal levels in patients with remission ITP. In contrast, CCL2 levels were similar in patients with active disease, patients in remission, and control subjects. PBMNC of patients with active disease expressed more CXCL10 mRNA (p = 0.031) but less CCR2 mRNA (p = 0.005). Lower peripheral platelet count correlated with higher CXCL10 levels and CXCL10/CCL2 ratios. Our study demonstrated that plasma levels of CXCL10 and CXC10/CCL2 ratio were higher in patients with active ITP than in healthy donors, and had an association with platelet counts of the patients. CXCL10 might be a pathogenic factor of this disorder.
Human immunology 02/2010; 71(6):586-91. · 2.55 Impact Factor
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ABSTRACT: Mesenchymal stem cells (MSCs) have been implicated in antitumor therapy for hematopoietic and non-hematopoietic tumors. Cell-contact and soluble factors are demonstrated to play a role in the growth inhibition of tumor cells mediated by MSCs in vitro, while there is little clue about signaling pathways involved in the process. P38 MAPK has been implicated as a suppressor of cell proliferation and tumorigenesis. We here investigate whether p38 MAPK is involved in MSC-induced growth inhibition of leukemic tumor cells. Methods: We characterized the effect of human umbilical cord mesenchymal stem cells (UC-MSCs) on proliferation, cell cycle and phosphorylation pattern of p38 MAPK in HL60 and K562 cells. SB203580, a specific inhibitor of p38 MAPK, or p38 MAPK-small interfering RNA (siRNA), were used to identify the role of p38 in growth suppression by UC-MSCs. We also investigated the expression of cell cycle regulators.
Treatment with UC-MSCs led to potent proliferation-inhibition of HL60 and K562 cells without inducing apoptosis. Growth inhibition by UC-MSCs was due to G0/G1 arrest. UC-MSCs increased phosphorylation of p38 MAPK in HL60 and K562 cells. Pharmacological inhibition or genetic silencing (through siRNA) of p38 MAPK partially abrogated the proliferation-suppression and cell cycle arrest caused by UC-MSCs. UC-MSCs also modulated the expression of cell cycle regulatory proteins in HL60 and K562 cells while SB203580 reversed the effect.
Taken together, our findings indicate that p38 MAPK is critical for the growth inhibitory effect of UC-MSCs on leukemic tumor cells.
Cellular Physiology and Biochemistry 01/2010; 26(6):799-808. · 2.86 Impact Factor
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ABSTRACT: The regulation of growth and apoptosis in K562 cells by human bone marrow mesenchymal stem cells (MSCs) from leukemia patients was investigated.
K562 cells were cocultured with leukemic MSCs under serum deprivation. Cell Counting Kit-8 (CCK-8), PI staining, Annexin V/PI binding and FACS assays were used to investigate cell proliferation, cell cycle status, and apoptosis of K562 cells cultures in the presence or absence of 10% serum. Western blotting was used to determine the levels of Akt, phosphorylated Akt (p-Akt), the BCL-2 family member Bad, and phosphorylated Bad (p-Bad) proteins in K562 cells after coculturing with MSCs. The effects of LY294002 (a specific inhibitor of PI3K) on protein expression were also determined.
K562 cell proliferation was inhibited by coculture with MSCs and the dominant cell cycle was the G0-G1 phase. The proportion of apoptotic K562 cells was decreased and the levels of p-Akt and p-Bad were upregulated after exposing K562 cells to MSCs. However, when LY294002 was used, p-Akt and p-Bad proteins inK562 cells showed a significant reduction, while no distinct variation was seen in the nonphosphorylated Akt and Bad protein levels.
Leukemic MSCs can inhibit K562 cell expansion and modulate the cell cycle to a state of relative quiescence. This allows the K562 cells to endure adverse conditions such as serum starvation. The PI3K-Akt-Bad signaling pathway may be involved in this antiapoptotic process via phosphorylation of the Akt and Bad proteins. Blocking MSC-induced transduction of the PI3K-Akt-Bad pathway may be a potential strategy for a targeted therapy to combat leukemia.
Journal of Experimental & Clinical Cancer Research 11/2009; 28:141. · 2.15 Impact Factor
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ABSTRACT: The ATP-binding cassette efflux transporter, ABCG2, is widely expressed in a variety of normal tissues, stem cells, as well as cancer cells. Existing data suggest that ABCG2 plays an important role in the maintenance of the stem cell phenotype and multidrug resistance of cancer cells. However, the potential role of ABCG2 in other cellular processes remains speculative and poorly understood. Here, we demonstrated that ABCG2 is involved in the proliferation of cancer cells. We used RNA interference approach to efficiently and specifically down-regulate ABCG2 protein levels in MCF-7/MX and A549 cells. We showed that knockdown of ABCG2 significantly inhibited the proliferation of these cells. Suppression of ABCG2 reduced the percentage of cells in the S phase of the cell cycle and enhanced G0/G1 accumulation. The G0/G1 growth arrest was associated with down-regulation of cyclin D3 and up-regulation of p21. Furthermore, blocking of ABCG2 function by chemical inhibitor fumitremorgin C also inhibited cell proliferation via the prolonged G0/G1 interval. Taken together, these findings suggest that ABCG2 correlates with cell cycle progression, highlighting a novel function of ABCG2 in cancer cell proliferation.
International Journal of Cancer 08/2009; 126(4):841-51. · 5.44 Impact Factor
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ABSTRACT: This study was designed to investigate the dynamics of transmigration and engraftment of hematopoietic stem/progenitor cells (HS/PCs) from umbilical cord blood (UCB) introduced via intra-bone marrow transplantation (IBMT), which is reserved as a novel strategy for possible clinical transplantation.
The early distribution pattern and engraftment level of human HS/PCs introduced via traditional intravenous transplantation (IVT) and IBMT routes were compared in the xenotransplanted nonobese diabetic/severe combined immunodeficient mouse model by means of flow cytometric analysis and an optical imaging system.
It was obvious that a good deal of IVT-introduced donor cells were entrapped in the liver and lung, 0.06% +/- 0.01% and 0.07% +/- 0.02%, respectively. Meanwhile three to six times fewer IBMT-introduced donor cells were entrapped in recipients' liver and lung (p<0.05 and p<0.05, respectively). Superior 8-week engraftment of human cells was observed in IBMT recipients (54.019% +/- 31.338%) than in IVT recipients (12.197% +/- 10.350%) when given transplants of 1.0 x 10(4) UCB CD34(+) cells and, furthermore, human hematopoietic cell engraftment was observed in IBMT, but not in IVT recipients when given transplants of 1.0 x 10(3) UCB CD34(+) cells.
Our results demonstrated that higher levels of human hematopoietic cell engraftment in nonobese diabetic/severe combined immunodeficient recipients achieved by IBMT might be due to the superior in vivo motility potential of IBMT-introduced HS/PCs. Clinical transplantation using transplants of UCB containing limited numbers of HS/PCs might benefit from the efficient IBMT strategy.
Experimental hematology 05/2009; 37(8):990-7. · 3.11 Impact Factor
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Jie Xu,
WenBin Liao,
DongSheng Gu,
Lu Liang,
Meng Liu,
WeiTing Du,
PengXia Liu,
Lei Zhang, ShiHong Lu,
ChunLan Dong,
Bin Zhou,
Zhongchao Han
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ABSTRACT: In contrast to hematopoietic stem cells, there is still a lack of definitive cell markers for specific isolation and identification of mesenchymal stem cells (MSCs). Thus a homogenous population of MSCs is only obtained after several passages, when multilineage potential or other distinctive features of very early progenitors may be already somewhat compromised. Recently a novel surface marker the neural ganglioside GD2 has been reported to distinguish MSCs from all other cells within marrow. Here, we found that MSCs derived from umbilical cord (UC-MSCs) also expressed this marker at early-passages. More importantly, UC-MSCs were the only cells within umbilical cord expressing this marker. Compared to unsorted cells, GD2(+)-sorted cells not only possessed much higher clonogenicity and proliferation capacity but also had significantly stronger multi-differentiation potentials. Flow cytometric analysis revealed that GD2(+)-sorted cells showed increased expression of SSEA-4, Oct-4, Sox-2 and Nanog, the typical markers expressed in embryonic stem cells, in comparison to unsorted or GD2-negative MSCs. Take together, our data demonstrate that the cells selected by GD2 are a subpopulation of MSCs with feature of primitive precursor cells and provide evidence that GD2 can be a cell surface marker suitable for the isolation and purification of UC-MSCs in early-passage culture.
Cellular Physiology and Biochemistry 02/2009; 23(4-6):415-24. · 2.86 Impact Factor
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ABSTRACT: Nanog plays an important role in embryonic stem (ES) cells pluripotency and self-renewal, yet the precise mechanism through which Nanog accomplishes this important function remains unclear. To understand comprehensive molecular mechanism by which Nanog mediates, we identified genome-wide molecular changes upon silencing Nanog in ES cells by using microarray technology. In order to downregulate Nanog expression efficiently, four siRNAs were designed on the basis of the conserved Nanog sequence and their effects on the Nanog expression were tested. Among these four siRNAs, Nanog-siRNA-P1 was found to be most effective. Once Nanog was downregulated, ES cells underwent differentiation by showing morphological change and decreased proliferation rate. Microarray analysis was then used to identify the altered gene expression after Nanog was silenced. A series of differentially expressed genes due to reduced expression of Nanog was identified as Nanog-related genes. These genes identified here could provide insights into the roles of Nanog in ES cells self-renewal and early differentiation.
Journal of Cellular Biochemistry 05/2008; 104(6):2348-62. · 2.87 Impact Factor
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ABSTRACT: Neuropilin-1 (NRP-1) is a novel receptor of vascular endothelial growth factor (VEGF) and expressed in endothelial cells and tumor cells. The role of NRP-1 in the growth and progression of leukemia is unknown. Here we studied the mRNA expression and effect of NRP-1 in leukemic cells. Our results showed that NRP-1 mRNA was expressed in six of seven leukemic cell lines and primary leukemias derived from all 24 patients with acute myeloid leukemia (AML). Reduced NRP-1 expression by RNA interference led to a decrease of VEGF-mediated mitogenic and migration responses in acute myeloid leukemic cell line HEL. Increased NRP-1 expression was directly correlated with the blast percentage in both peripheral blood and bone marrow of AML patients. Our data demonstrated that a higher level of NRP-1 mRNA was expressed in leukemias and NRP-1 promoted proliferation and chemotaxis of leukemic cells in response to VEGF. Inhibition of NRP-1 functions may provide a new therapeutic strategy for treatment of AML.
Leukemia & lymphoma 03/2008; 49(2):331-8. · 2.40 Impact Factor
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ABSTRACT: Hemangiopoietin (HAPO) is a novel human growth factor acting on the primitive cells of both hematopoietic and endothelial cell lineages. Our previous study has shown that HAPO exerts a proliferative effect on endothelial cells. However, the mechanism of this action remains unclear. Thus, we studied the signal transduction pathway whereby HAPO promotes cell proliferation in human umbilical vein endothelial cells (HUVECs). In this study, recombinant human HAPO (rhHAPO) stimulated the proliferation of HUVECs in a dose-dependent manner. The transient phosphorylation of Akt occurred after addition of rhHAPO to HUVECs. LY294002, a specific inhibitor of PI-3K, significantly inhibited Akt phosphorylation and completely abrogated HAPO-stimulated proliferation of HUVECs. rhHAPO enhanced the expression of cyclin D1, where as LY294002 inhibited the up-regulation of cyclin D1. Moreover, rhHAPO is able to selectively enhance the mitogenic activity of VEGF for vascular endothelial cells. Overall, these findings demonstrate that HAPO induces endothelial cell proliferation through the PI-3K/Akt pathway.
Cellular Physiology and Biochemistry 02/2008; 22(1-4):307-14. · 2.86 Impact Factor
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ABSTRACT: Idiopathic thrombocytopenic purpura (ITP) is a disease putatively relating to abnormal immune function and auto-antiplatelet immunoglobulin. We examined whether polymorphism of CD72, an inhibitory receptor of B cells, affect the susceptibility to ITP, or associated with the clinical characteristics of ITP. A case-control study was carried out in 206 Chinese ITP patients and 169 healthy controls. The detection of variable number of tandem repeats in CD72 intron 8 was performed by polymerase chain reaction and subsequent analysis with polyacrylamide gel electrophoresis. We did not find direct association between CD72 genotypes and susceptibility to ITP. The haplotype that contained one repeat of 13 nucleotides in intron 8 (designated as *1, and haplotype containing two repeat of 13 nucleotides in intron 8 is designated as *2) was significantly associated with early first onset age (< or = 14) in ITP patients (P = 0.03). ITP patients with CD72*1\*1 and *1\*2 genotype had a 3.09-fold [95% confidence interval (CI), 1.32-7.25] and 1.98-fold (95% CI, 0.92-4.25) increased risk of appearing ITP manifestation at their childhood respectively. The haplotype CD72*1 is apparently a risk allele, whereas CD72*2 a protective allele for child-onset of ITP disease.
Journal of Clinical Immunology 01/2008; 28(3):214-9. · 3.08 Impact Factor
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ABSTRACT: To determine whether live attenuated Salmonella carrying platelet factor 4 cDNAs can protect mice from radiation damage, the attenuated Salmonella SL3261 was used as oral vector for targeted gene delivery. The recovery of mice receiving sublethal total-body irradiation (TBI) was investigated after the oral administration of attenuated Salmonella carrying cDNA for platelet factor 4 (PF4) or truncated PF4. This oral gene therapy protected mice from radiation damage after TBI. The number of bone marrow cells and high proliferative potential colony-forming cells (HPP-CFCs) increased significantly at day 7. Similarly, the administration of PF4 or PF4(17-70) protein also improved the survival of mice after TBI. Both PF4 gene therapy and protein administration accelerated hematopoietic recovery in vivo in mice after irradiation. In vitro, PF4 also promoted survival and proliferation of 5-fluorouracil-resistant hematopoietic stem/progenitor cells after irradiation. These data demonstrate a novel biological function of PF4 as a protector against radiation injury and suggest that attenuated Salmonella could be used in vivo as a PF4 DNA delivery vector in the management of radiation injury.
Radiation Research 09/2006; 166(2):352-9. · 2.68 Impact Factor
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ABSTRACT: The multidrug resistance (MDR) mediated by P-glycoprotein (P-gp), the MDR1 gene product, is one of the major obstacles in leukemia treatment. The present study was designed to explore a MDR1-targeted small interfering RNA (si-MDR1) approach for reversal of P-gp-mediated MDR in the MDR human leukemia cell line k562/A02. It was found that si-MDR1 significantly inhibited MDR1 expression at both mRNA and protein levels. Depletion of MDR1 by si-MDR1 correlated with the increased sensitivity of the cells to cytotoxic agents and with the enhanced intracellular retention of daunorubicin (DNR). One base-pair mutated control (si-MDR1-Mut) lost the effect of si-MDR1 on both the degradation of mdr1 mRNA and the reduction of P-gp expression. These findings indicate that siRNA specifically and efficiently interferes with the expression of mdr1 and could be used as a molecularly defined therapeutic approach for MDR in the treatment of leukemia.
Cancer Gene Therapy 12/2004; 11(11):707-12. · 2.80 Impact Factor
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ABSTRACT: Essential thrombocythemia (ET) is a chronic myeloproliferative disorder with abnormal megakaryocyte/platelet production. Recent studies have found that Bcl-xL, as a member of the bcl-2 family of proteins that inhibit apoptosis, is essential in megakaryocytic differentiation. In this study the expression of Bcl-xL was evaluated during megakaryocytic differentiation in ET patients.
To study the role of Bcl-xL in megakaryocyte differentiation, we evaluated the effect of small interfering RNA (siRNA) on the expression of Bcl-xL. CD34+ cells from patients with ET, chronic myeloid leukemia (CML), polycythemia vera (PV) and normal individuals were cultured in serum-free medium supplemented with thrombopoietin (TPO). Immunocytochemical staining and flow cytometric analysis were used to evaluate the Bcl-xL expression during megakaryocytic differentiation of CD34+ cells.
When exposured to si-Bcl-xL, the percentage of K562 cells induced into megakaryocytes in 72 hours was lower than the corresponding percentage of control cells. CD41a+ cells from the three groups of patients and the control group were cultured. At day 10, the percentage of Bcl-xL- cells in CD41a+ cells from ET patients was 61.0+/-28.1%, which was significantly higher than that from patients with CML (2.5+/-20.9%), PV (33.6+/-10.0%) or control subjects (15.1+/-13.0%).]
These results demonstrate that Bcl-xL is down-regulated early during in vitro differentiation of megakaryocytes from ET patients; this might reflect an early entry of megakaryocytes into a degenerating mature stage.
Haematologica 11/2004; 89(10):1199-206. · 6.42 Impact Factor
