Optically stimulated luminescence dating of cave deposits at the Xiaogushan prehistoric site, northeastern China.

The Xiaogushan cave site is one of the most important prehistoric sites in North China. The stone and bone artifacts found in the cave are similar to European contemporaneous artifacts. Cave deposits consist of five layers that have been dated from 46,353 ± 1179 to 4229 ± 135 cal. yr BP, using radiocarbon dating techniques on charcoal and bone samples collected from Layers 2-5. In this paper, optically stimulated luminescence (OSL) techniques were applied to date six samples taken from Layers 1-3. The luminescence properties of the fine-grained and coarse-grained quartz extracts indicate that the materials are suitable for OSL dating using a single-aliquot regeneration-dose (SAR) protocol. The OSL ages obtained are broadly consistent with the stratigraphy and the associated calibrated radiocarbon ages. The dating results show that the cave was first occupied by humans about 70 ka. The human occupation of the cave may be related to climate change. An occupation hiatus is inferred to between ∼ 17 to ∼ 10 ka. The stone and bone artifacts found in Layers 2 and 3 may indicate the Middle to Upper Paleolithic transitions in the region.

[Valproic acid induced intracellular GSH-redox imbalance and apoptosis of leukemic cells resistant to dexamethasone and doxorubicin].

OBJECTIVE: To investigate the anti-neoplastic effects of Valproic acid (VPA) on leukemic cells, especially drug-resistant lines, and to investigate whether modulation of GSH-redox status is involved in VPA-induced apoptosis. METHODS: After the treatment of VPA at various concentrations for indicated times, cellular proliferation of the Jurkat, CEM, HL-60, K562, K562/AO2 cells were evaluated via MTT assay; and the activities of Caspase-3, Caspase-8 and Caspase-9 were quantitatively analyzed by colorimetric assay. The morphological change and cell cycle distribution were also examined on Jurkat (Dexamethasone-resistant) and K562/AO2 (Doxorubicin-resistant) cell lines. The levels of intracellular glutathione/glutathione disulfide (GSH/GSSG) and the activities of the typical antioxidant enzymes, i.e., glutathione reductase (GSH-Rd) and glutathione peroxidase (GSH-Px), were measured on cell lysates of Jurkat and K562/AO2 cell lines prior to and after VPA treatment. Apoptosis rates of Jurkat and K562/AO2 cells treated with VPA along or in combination with N-acety-l-cysteine (NAC), catalase (CAT) or DL-buthionine-(S,R)-sulfoximine (BSO) were determined by Annexin V/propidium iodide (PI) staining with flow cytometry analysis. RESULTS: At concentrations comparable with that achieved at clinical settings, VPA inhibited cell proliferation, activated Caspase-3, 8, and 9, and induced cell cycle arrest in Jurkat and K562/AO2. A rapid decrease in GSH-Rd and GSH-Px activities and GSH content in Jurkat and K562/AO2 were detected after VPA treatment. Co-administration of NAC or CAT attenuated VPA-induced apoptosis. CONCLUSION: VPA inhibit cell proliferation, induce cell cycle arrest and apoptosis in drug-resistant leukemic cells. Apoptosis correlates with down-regulation of intracellular GSH and disruption of intracellular GSH-redox balance, possibly through inhibition of glutathione reductase and glutathione peroxidase.

[Isolation of multipotent mesenchymal stem cells from the tissue of umbilical cord for osteoblasts and adipocytes differentiation].

OBJECTIVE: To explore the methods and conditions for isolating and proliferating multipotent mesenchymal stem cells (MSCs) from the tissue of umbilical cord, with an aim to induce osteogenic and adipogenic differentiation in vitro. METHOD: The cord was dissected along the long axis, with vessels pulled away and then sutured into a "loop". Collagenase solution was filled into the loop. Suspended cells were collected from the loop suspension after 6-8 hours and centrifuged. The cells were finally cultured in polystyrene dishes. The single cell-derived colonies were obtained and tested for their immunophenotype and osteoblast and lipoblast differentiations. RESULT: Adherent cells were obtained from the tissue of umbilical cord, which proliferated and formed single cell-derived colonies. The colonies presented matrix cells immunophenotype and differentiated into osteoblasts that produced mineralized matrices, which were stained by alizarin red and alkaline phosphatase. The colonies also differentiated into adipocytes that accumulated lipid vacuoles, which were demonstrated by the morphology and oil red stains. CONCLUSION: MSCs can be isolate from the tissue of umbilical cords and proliferate in vitro. The proliferated colonies show matrix cell immunophenotypes and can differentiate into osteoblasts and adipocytes.

[Method and conditions of isolation and proliferation of multipotent mesenchymal stem cells].

OBJECTIVE: To investigate the method and conditions of isolation, proliferation of multipotent mesenchymal stem cells (MSCs) from human umbilical cord blood in vitro, and to induce osteogenic and adipogenic differentiation directly for identification. METHODS: Human umbilical cord blood was collected in asepsis condition, isolated by density gradient centrifugation, or sedimented red cell with methylcellulose, and then the same centrifugation was done, or obtained by negative immunodepletion of CD34+. These isolated mononuclear cells were used to carry on plastic adherent culture. To obtain single cell-derived colonies, these cells were proliferated clonally in medium which consists of L-DMEM or Mesencult medium and 10% fetal calf serum(FCS) respectively, then their differentiation potentiality to osteoblasts and lipoblasts was tested. RESULTS: The mononuclear cells isolated by sedimented and centrifugated way cultured in Mesencult medium and 10%FCS were most available. These adhesive cells could become obviously short rod-shape or shuttle-shape cells after 5-7 days. The colonies form well in 3rd-passage cells. The mononuclear cells obtained by only centrifugalized in density gradient were hard to form colony, isolated by immunomagnetic beads were hard to culture. The surface antigens of these colonies cells presented CD29, CD59, CD71 but not CD34, CD45 and HLA-DR etc. The colony cells differentiating into osteoblasts that produce mineralized matrices, stained by alizarin red, and differentiating into adipocytes that accumulate lipid vacuoles, stained by oil red. CONCLUSION: MSCs can be isolated from human umbilical cord blood and proliferate it in vitro. The way that mononuclear cells are sedimented red cell by methylcellulose and cultured by Mesencult medium and 10% FCS is the valid method of isolation. Proliferation colonies cells present matrix cell immunophenotypes, and can differentiate into osteoblasts and adipocytes.

[Expression of HSP70 mRNA and MDR1 mRNA in K562 cells induced by heat shock and ADM].

This study was purpose to investigate the expression levels of HSP70 and MDR1 genes under heat shock and/or adriamycin (ADM) chemotherapy stimulation. The K562 cells were bathed in water at 43 degrees C for 1 hour, then the heat-treated K562 cells were collected and were cultured at 37 degrees C. The expression of HSP70 was assayed by immunocytochemistry, the growth suppression rate of K562 cells was detected by MTT assay, the function of P-gp and the expressions of HSP70 mRNA, MDR1 mRNA were detected by flow cytometry and real-time quantitative PCR (RT-PCR) respectively. The results showed that (1) the synthesis of HSP70 protein in K562 cells treated with high shock (43 degrees C) reached to high level after culture at 37 degrees C for 2 hours, and moved from cytoplasm to nucleolus, the expression of HSP70 began to decrease following 3 hours of culture at 37 degrees C, and gradually reached to normal level after culture at 37 degrees C for 5 hours, the location of HSP70 expression returned to cytoplasm; (2) the expressions of HSP70 mRNA and MDR1 mRNA increased following 43 degrees C heat shock, and were 4 and 5.8 times higher than that of control group at 37 degrees C culture for 2 hours respectively; (3) the expression of P-gp was higher in ADM group than that in control. The expressions of HSP mRNA and MDR1 mRNA increased significantly in heat shock plus ADM group and ADM group as compared with control (p<0.01). It is concluded that the heat shock and ADM chemotherapy both induce over expression of HSP70 and MDR1 which can maintain stability of K562 cells and may be related to formation of the MDR in leukemia.

[Antineoplastic effect of valproic acid and trichostatin on HL-60 and K562 cells].

The objective of this study was to investigate antineoplastic effects of valproic acid (VPA) and trichostatin (TSA) on HL-60 and K562 cells in vitro, and the synergic effects of VPA or TSA in combination with ATRA. The inhibitory effects of VPA, TSA and ATRA in various concentrations and different combinations on proliferation of HL-60 and K562 cells were observed by cell growth curves, 50% inhibitory concentration (IC(50)), as well as inhibition of leukemia colony growth at different time points. The characteristics of cell differentiation or apoptosis were analyzed by cytochemical staining, differentiation antigen detection, cell cycle assay and A(NBT)/A(MMT) value determination. The results showed that HL-60 cell had a lower IC(50) of VPA and TSA compared with K562 cells. ATRA could significantly enhance the inhibition of VPA, TSA on clonegenicity of HL-60 cells and inhibition of VPA on clonegenicity of K562 cells. HL-60 cells treated with VPA displayed the phenotype of neutrophilic like cells, and showed the increases of NBT reduction rate and CD11b expression. No evidence for K562 differentiation was found. It is concluded that both VPA and TSA inhibit HL-60 cells growth in vitro. VPA induces differentiation of HL-60 cells to granulocyte. VPA and TSA have a moderate anti-proliferative effect on K562 cells. None of these agents induces K562 cell differentiation.

[Erythropoietin increases transferrin receptor expression and the impact of erythropoietin on K562 leukemic cell cycle].

OBJECTIVE: Functionally, erythropoietin (EPO) can promote the proliferation and growth of erythroid progenitor cells, and it is widely used in the treatment of anemia in chronic diseases caused by tumor and inflammation. However, it is unclear whether EPO has any effect on tumor cell iron metabolism and tumor cell proliferation. The purpose of this study was to explore the effects of recombinant human EPO (rhEPO) on the expression of transferrin receptor (TfR, CD(71) antigen) of leukemic cell K562 and its relation to cell cycle. METHODS: In vitro culture of K562 cell was performed with additions of various concentrations of rhEPO and Fe. Treatments were terminated at 24 h and 72 h, respectively. Then each group of cells was incubated with FITC-IgG antibody to CD(71) or PI, a kind of DNA dye. And TfR expression and DNA synthesis status were analyzed by flow-cytometry. RESULTS: (1) The expression of TfR by K562 cells increased significantly when incubated for 72 h with different concentrations of rhEPO. The measurement values of 5 U/ml, 10 U/ml and 20 U/ml groups were 12.2 +/- 1.40, 10.7 +/- 0.99 and 11.1 +/- 0.90, respectively. They were markedly increased when compared with that of control group (6.27 +/- 0.11, P < 0.05). (2) When incubated with rhEPO (5 u/ml) alone or combined with FeCl(3) (100 micro mol/L), the percentages of cells in S phase were 51.1% and 59.6%, respectively. They significantly increased when compared with that of control group (42.9%, P < 0.05). CONCLUSIONS: Iron is very important for the proliferation of both normal cells and leukemic cells. It is essential to the activity of ribonucleotide reductase (RR). The authors hypothesized that rhEPO would increase the expression of TfR and intracellular iron content of leukemic cells, which would enhance the DNA synthesis and cell proliferation. Therefore, the clinical application of rhEPO to promote erythropoiesis of cancer patients should be cautious.