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Objective: To investigate the effect of SIRT1/NF-κB signal axis on delaying hematopoietic stem cell and progenitor cell senescence with ginsenoside Rg1 in ageing model rat induced by D-galactose. Methods: Male SD rats (n = 40) aging from 6 to 8 weeks old were randomly divided into control group, aging model group, positive control group, Rg1 treated group, and Rg1 prevented group (n = 10). The aging rat model was prepared by sc D-galactose for continuous 42 d, then ginsenoside Rg1 was given in different time. After 2 d of the treatment, the Sca-1+ HSC/HPC was isolated by magnetic cell sorting (MACS). The changes of cells observed by senescence-associated β-galactosidase (SA-β-Gal) staining, cell cycle analysis and culture of mixed hematopoietic progenitor cell were used to investigate the treated aging effect of ginsenoside Rg1.The expression of senescence associated SIRT1, NF-κB mRNA and protein was examined by real time fluorescence quantitative PCR (FQ-PCR) and Western blotting. Results: In ginsenoside Rg1 treated group and ginsenoside Rg1 prevented group, the percentage of positive cells expressed SA-β-Gal and the number of cells entering G0/G1 phase were lower than that of aging model group, but the number of CFU-Mix was increased than aging model group. Compared with aging model group, the expression of SIRT1 mRNA and protein was upregulated and the expression of NF-κB mRNA and protein was downregulated in Rg1 treated group and prevented group. Changes in Rg1 prevented group were more than those in Rg1 treated group. Conclusion: SIRT1/NF-κB signal axis may play a key role in the anti-aging effect of Rg1 to Sca-1+ HSC/HPC senescence in ageing model rat induced by D-galactose.
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0.05).Conclusion:The results support our hypothesis that APS can be used to treat the hematopoietic type of acute radiation sickness.APS pretreated groups had better results than APS treated groups in initial observation.
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Objective To study the effect of Sca1+ cells on the HSCs`(hemopoietic stem cells) differentiation to dendritic cells(DCs),and identify the morphology,function and surface markers of the cells.Methods CD117+ HSCs were isolated and purified from the bone marrow of healthy Balb/c mice by magnetic affinity cell sorting.After cell expansion by treatment with the support of Sca1+ cells,the HSCs were induced for directional differentiation into DC-like cells.Studied the surface markers by FACS and function via LSCM and animal experiments. Results After 10 days of culture,we demonstrated that Sca1+ cells induced HSCs to differentiate into a distinct regulatory DC subset with high expression of CD11b but low expression of Ia.They had phagocytotic activity,and suppressed the GVHD(graft versus host disease) reaction.Conclusion HSCs can differente into dendritic cells with the support of Sca1+ cells.
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Aim:To investigate the effect of Hemangiopoietin (HAPO) on the hematopoiesis reconstitution in sub-lethally irradiated Balb/c mice.Methods: Balb/c mice were underwent total body irradiation at 700 cGy 137Cs ? radiation and were treated with HAPO or recombinant human granulocyte colony stimulating factor (rhG-CSF) after irradiation. The hematopoiesis reconstitution of mice were detected. Cells from bone marrow of Balb/c mice were cultured with HAPO or rhG-CSF for 24 hours or 72 hours before or after the cells were irradiated. The viability of cells were assessed and the ability of in vitro hematopoiesis reconstitution were also detected. Result: rhG-CSF and HAPO treated mice both showed increased survival rate and increased colony forming units. The peripheral WBC number increased greatly. The HAPO group was most quickest compared with rhG-CSF group and PBS control group. The number of bone marrow cells at day 14 of rhG-CSF group was higher than that in HAPO group, but the number of bone marrow cells at day 32 of rhG-CSF was lower than that in HAPO group. The number of bone marrow cells at day 42 of rhG-CSF was below normal. The number of bone marrow cells at day 42 of HAPO group was nearly normal. The number of CFU-GEMM in HAPO group was most compared with that in rhG-CSF group and PBS control group at day 7, 14 and 21 after radiation. The survival rate of cells after radiation in HAPO group was markedly higher than that in PBS control group, but the survival rate of cells after radiation in rhG-CSF group was no notable difference compared with that in PBS control group. In MTT assay, both HAPO and rhG-CSF incubation stimulated proliferation of bone marrow cell at 72 hours after radiation. Bone marrow cells formed Hematopoietic islands in HAPO group after radiation and were positive for sca-1 and CD31. CD31 positive endothelial cells increased around the Hematopoietic islands. There was no Hematopoietic islands formation, few CD31 positive endothelial cells and no sca-1 positive cells in PBS control group. Conclusion: HAPO can promote hematopoiesis reconstitution in sub-lethally irradiated Balb/c mice. It can increase the survival rate of mice and stimulate the proliferation of hematopoietic stem cells.
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AIM: To explore transdifferentiation potential of Sca-1 + cells from murine fetal liver. METHODS: 2?10 3 of Sca-1 + cells from male murine fetal liver were transfused into female mouse irradiated lethally with ? ray from 60 Co source (10 Gy) via tail vein. Two months later, FISH and immunohistochemistry were used to detect the situation for transdifferentiating of the donor cells (male cells) in tissues of female recipient mouse. RESULTS: The renal tubular epitheliocyte-like and neurocyte-like cells with Y chromosome were found on the sections of renal and brain tissues from female recipient mice. These cells have phenotype characteristics of RCA+/CD - 45 F - 4/80 and NueN +/CD -45 F - 4/80, respectively. CONCLUSION: The evidence is provided for Sca-1 + cells from murine fetal liver to transdifferentiate into both renal and brain tissue cells.
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AIM: To study whether Sca-1 + cells from fetal liver can be induced to differentiate into neuronal cells in vitro. METHODS: Sca-1 + cells from 14 5-days-old murine fetal liver were isolated with a magnetic cell sorting kit, and were cultured in Dulbecco's modified Eagle's medium (DMEM)/F12 supplemented with 10% fetal bovine serum (FBS), and passaged at a ratio of 1∶3 when cells reached more than 80% confluence. The 5 passage cells were induced by 10 -3 mol/L ?-mercaptoethanol (?-ME) and 5 ?10 -7 mol/L all-trans-retinoic acid (RA) for 24 hours, and then incubated in serum-free medium for 5 hours to 5 days. The characteristics of treated cells were assayed by immunocytochemistry staining analysis at 5 hours, or 5 days.RESULTS: Cells treated with ?-ME and RA exhibited neuronal phenotype and expressed neuron-specific protein such as neuron-specific nuclear protein (NeuN), neuronfilament-M, and neuron-specific tubulin-1 (TuJ-1) but not tau, MAP-2, or the astrocyte-specific marker glial fibrillary acidic protein (GFAP).CONCLUSION: Sca-1 + cells from fetal liver, of which most are regarded as hematopoietic stem cells, could differentiate into early immature neuronal cells in vitro . These findings suggest that Sca-1 + cells from fetal liver may be an alternative source in cell therapy and gene therapy of neural dysfunction.
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liver.Combined informational analysis showed that the presenting frequency of the cells containing Y chromosome was consistent with the irradiative sensitivity of the organ.CONCLUSION: These findings suggest that the capability of differentiation of Sca-1 positive cells from murine fetal liver was potentially connected to the extent of damage in these organs when transferred in vivo.
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AIM: To study the effect of acute renal failure (ARF) on the differentiated frequency of Sca-1+ cells from murine fetal liver in irradiated mice. METHODS: The Sca-1+ cells from murine fetal liver were isolated with magnetic cell sorting (MACS) technique, the sex of which was identified by PCR. The 2?104 Sca-1+ cells were transplanted into a lethally irradiated ([ 60Co], 8 Gy) inbred female mouse. After 8 weeks, these recipient mice were divided to A, B, and C groups at random (A group: irradiated; B group: ARF; C group: ARF and Sca-1+). The mice in B and C groups were induced to ARF with 50% (V/V) glycerin (11.6 mL/kg). 72 hours later, the mice in C group were injected with the fresh prepared Sca-1+ cells again. 8 weeks later, mice were sacrificed, and their kidneys were taken out, fixed and slices were prepared. Fluorescence in situ hybridization (FISH) of renal slices was performed and the pictures of them were taken and analyzed. RESULTS: The cells containing Y chromosome were found in renal slices from the mice in A, B and C groups, which located in epithelial cells of renal tubules, interstitium, glomeruli, and glomerular margin and increased gradually. The double and encircle zone of Y chromosome cells were found in the slices from the mice in B and C groups separately, which was consist of new renal tubules. The differentiation frequency of Sca-1+ cells in kidney in A, B and C groups were (1.65?0.18)%, (8.58?1.34)% and (18.13?1.91)%, respectively, which showed significant difference between former group and later group (P
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AIM: To determine whether Sca-1~+ cells from fetal liver can differentiate into neural cells. METHODS: The sex of 14.5-day-old murine fetuses was determined by PCR analysis of sry gene, and Sca-1~+ cells from male fetal liver were isolated with a magnetic cell sorting kit, 2?10~3 of which were then transplanted into lethally irradiated female mice. The donor cells and their characteristics in recipient brains were identified and detected by FISH and immunohistochemistry double-staining analysis at 60, 120, 180 days after transplantation. RESULTS: There existed many male cells in brains of female recipients, some of them express neuron-specific nuclear protein (NeuN), and some of them express the astrocyte-specific marker, i.e. glial fibrillary acidic protein (GFAP). CONCLUSION: Sca-1~+ cells from fetal liver, which contain hematopoietic stem cells, can differentiate into neuronal cells and astrocytes in the brains of adult mice.
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Objective:To explore potential differentiation from antigen-1 positive stem cells (Sca-1 +cells) in murine fetal liver tissue into cardiomyocytes in vivo. Methods: Sca-1 + cells from the murine fetal liver of 14.5 d were separated by Magnetic cell sorting (MACS).The sequence of the sex determination region of the Y chromosome was determined by PCR.Sca-1 + cells (2?10 3) of male mice were transfused to female mice receiving 8-12 weeks irradiation at a lethal dose of ?-ray(10 Gy) from a 60Co source.Two months later,the mice were killed and the hearts were removed to make slices. Fluorescence in situ hybridisation (FISH) tests with a Y chromosome probe were used to detect sex of the cells.Immunohistochemistry with antibodies of desmin,Flt-1,white blood cell common antigen CD45 and macrophage F 4/80 were used to detect tissue characteristics of cardiomyocytes. Results: Cells with Y chromosome were found in the cardiomyocytes of female mice transplanted with Sca-1 + cells,and showed phenotypic characteristics of Desmin +/Flt-1-/CD45-/F- 4/80 at the same time.Conclusion: Sca-1 + cells from the murine fetal liver are mostly hematopoietic stem cells,having the potential of differentiating into cardiomyocytes.