OBJECTIVETo investigate the effect of lead-exposed astrocyte conditioned medium (ACM) on the synaptic formation of neurons and to provide reference for the mechanism of lead neurotoxicity.

METHODSAstrocytes were cultured in the medium containing 50, 100, 200, 400, and 800 µmol/L lead acetate for 72 h. Alamar Blue was used to assess the cell viability of astrocytes, and then ACM was collected. Primarily cultured neurons were divided into six groups: pure culture group, non-glutamic acid (Glu)-induced ACM treatment group, Glu-induced lead-free ACM treatment group, and Glu-induced 50, 100, and 200 µmol/L lead acetate-exposed ACM treatment groups. Neurons were collected after being cultured in ACM for 24, 48, or 72 h. The content of synaptophysin (SYP) in neurons was determined by Western blot. The SYP expression in neurons was measured by immunofluorescence after being cultured in ACMfor 72 h.

RESULTSIn all lead-exposed groups, the cell viability of astrocytes declined with increasing concentration of lead (P < 0.05). The Western blot showed that compared with the pure culture group, the non-Glu-induced ACM treatment group and Glu-induced lead- free ACM treatment group had significantly increased content of SYP in neurons (P < 0.01); compared with the non-Glu-induced ACM treatment group, the Glu-induced ACM treatment groups had significantly reduced SYP expression in neurons (P < 0.05); compared with the Glu-induced lead-free ACM treatment group, all lead-exposed ACM treatment groups had the content of SYP in neurons significantly reduced with increasing concentration of lead after 72-h culture (P < 0.01), the 200 µmol/L lead-exposed ACM treatment group had significantly reduced content of SYP in neurons after 48-h culture (P < 0.01), and all lead-exposed ACM treatment groups showed no significant changes in the content of SYP in neurons after 24-h culture. Double-labeling immunofluorescence of SYP showed that all lead-exposed ACM treatment groups had a significant decrease in the number of SYP-fluorescent particles after 72-h culture (P < 0.05).

CONCLUSIONAstrocytes promote synaptic formation of neurons, which may be inhibited during lead exposure.

Astrocytes ; drug effects ; physiology ; Cell Survival ; drug effects ; Cells, Cultured ; Culture Media, Conditioned ; metabolism ; Glutamic Acid ; metabolism ; Lead ; toxicity ; Neurons ; drug effects ; Synapses ; drug effects ; physiology

Iron is an essential element for almost all living organisms. The possible role of iron for growth, adherence and cytotoxicity of Entamoeba histolytica was evaluated in this study. The absence of iron from TYI-S-33 medium stopped amebic growth in vitro. However, iron concentrations in the culture media of 21.4-285.6 microM did not affect the growth of the amebae. Although growth was not retarded at these concentrations, the adhesive abilities of E. histolytica and their cytotoxicities to CHO cell monolayer were correlated with iron concentration. Amebic adhesion to CHO cell monolayers was significantly reduced by low-iron (24.6 +/- 2.1%) compared with 62.7 +/- 2.8 and 63.1 +/- 1.4% of amebae grown in a normal-iron and high-iron media, respectively. E. histolytica cultured in the normal- and high-iron media destroyed 69.1 +/- 4.3% and 72.6 +/- 5.7% of cultured CHO cell monolayers, but amebae grown in the low-iron medium showed a significantly reduced level of cytotoxicity to CHO cells (2.8 +/- 0.2%). Addition of divalent cations other than iron to amebic trophozoites grown in the low-iron medium failed to restore levels of the cytotoxicity. However, when E. histolytica grown in low-iron medium were transferred to normal-iron medium, the amebae showed completely restored cytotoxicity within 7 days. The result suggests that iron is an important factor in the adherence and cytotoxicity of E. histolytica to CHO cell monolayer.
Animals ; CHO Cells ; Cell Adhesion/drug effects ; Cell Survival ; Cricetinae ; Cricetulus ; Entamoeba histolytica/*drug effects/*physiology ; Iron/*pharmacology

Iron is an essential element for almost all living organisms. The possible role of iron for growth, adherence and cytotoxicity of Entamoeba histolytica was evaluated in this study. The absence of iron from TYI-S-33 medium stopped amebic growth in vitro. However, iron concentrations in the culture media of 21.4-285.6 microM did not affect the growth of the amebae. Although growth was not retarded at these concentrations, the adhesive abilities of E. histolytica and their cytotoxicities to CHO cell monolayer were correlated with iron concentration. Amebic adhesion to CHO cell monolayers was significantly reduced by low-iron (24.6 +/- 2.1%) compared with 62.7 +/- 2.8 and 63.1 +/- 1.4% of amebae grown in a normal-iron and high-iron media, respectively. E. histolytica cultured in the normal- and high-iron media destroyed 69.1 +/- 4.3% and 72.6 +/- 5.7% of cultured CHO cell monolayers, but amebae grown in the low-iron medium showed a significantly reduced level of cytotoxicity to CHO cells (2.8 +/- 0.2%). Addition of divalent cations other than iron to amebic trophozoites grown in the low-iron medium failed to restore levels of the cytotoxicity. However, when E. histolytica grown in low-iron medium were transferred to normal-iron medium, the amebae showed completely restored cytotoxicity within 7 days. The result suggests that iron is an important factor in the adherence and cytotoxicity of E. histolytica to CHO cell monolayer.
Animals ; CHO Cells ; Cell Adhesion/drug effects ; Cell Survival ; Cricetinae ; Cricetulus ; Entamoeba histolytica/*drug effects/*physiology ; Iron/*pharmacology

OBJECTIVE: To investigate the effects of autophagy on osteogenic differentiation of stem cells from the apical papilla (SCAPs) in the presence of tumor necrosis factor- (TNF-) stimulation . METHODS: SCAPs treated with TNF- (0, 5, and 10 ng/mL) with or without 5 mmol/L 3-MA were examined for the expression of autophagy marker LC3-Ⅱ using Western blotting. The cells were transfected with GFP-LC3 plasmid and fluorescence microscopy was used for quantitative analysis of intracellular GFP-LC3; AO staining was used to detect the acidic vesicles in the cells. The cell viability was assessed with CCK-8 assays and the cell apoptosis rate was analyzed using flow cytometry. The cells treated with TNF- or with TNF- and 3-MA were cultured in osteogenic differentiation medium for 3 to 14 days, and real- time PCR was used to detect the mRNA expressions of osteogenesis-related genes (ALP, BSP, and OCN) for evaluating the cell differentiation. RESULTS: TNF- induced activation of autophagy in cultured SCAPs. Pharmacological inhibition of TNF--induced autophagy by 3-MA significantly decreased the cell viability and increased the apoptosis rate of SCAPs ( < 0.05). Compared with the cells treated with TNF- alone, the cells treated with both TNF- and 3-MA exhibited decreased expressions of the ALP and BSP mRNA on days 3, 7 and 14 during osteogenic induction ( < 0.05) and decreased expression of OCN mRNA on days 3 and 7 during the induction ( < 0.05). CONCLUSIONS Autophagy may play an important role during the osteogenic differentiation of SCAPs in the presence of TNF- stimulation.
Autophagy ; drug effects ; physiology ; Cell Differentiation ; drug effects ; physiology ; Cell Survival ; drug effects ; Cells, Cultured ; Dental Papilla ; cytology ; Green Fluorescent Proteins ; Humans ; Osteogenesis ; physiology ; Stem Cells ; drug effects ; physiology ; Transfection ; Tumor Necrosis Factor-alpha ; administration & dosage ; antagonists & inhibitors ; pharmacology

OBJECTIVETo determine the optimal concentration and processing time of EMS mutation for suspension cells from Pinellia ternata.

METHODUnder four EMS concentration gradients (0.1% , 0.2%, 0.4%, 0.6%) and three processing time gradients (0.5, 1.0, 2.0 h), the suspension cells of P. ternata were mutagenized.

RESULT AND CONCLUSIONThe results showed that the survival rate was significantly different under the different concentrations of EMS and the different processing time. In the same processing time, the EMS concentrations were increased, but the suspension cells survival rate decreased gradually. The optimum EMS concentration for the mutagenesis was 0.4% and the best processing time was 1 hour.

Cell Survival ; drug effects ; genetics ; Dose-Response Relationship, Drug ; Ethyl Methanesulfonate ; pharmacology ; Mutagenesis ; drug effects ; Mutation ; drug effects ; Pinellia ; cytology ; drug effects ; genetics ; physiology ; Suspensions ; Temperature ; Time Factors

This study determined the effects of selenium on the growth of Fusarium strains and the effects of products extracted from the fungal cultures on relevant indicators of chondrocytes injury. The results showed that selenium supplementation resulted in differential effects on the mycelial growth of the strains. Levels of the chondrocyte injury indicators, including cell viability, proteoglycan and type II collagen contents and their mRNA expressions, were all reduced to varying degrees when the chondrocytes were incubated with fermentation extracts, the inhibitory effect varied depending on selenium content supplemented to fungal culture media. The results indicated that certain chain relations existed between the content of selenium in the environment, the production of some metabolites by fungi, and the occurrence of chondrocyte damage. The extent of this relationship and the role it plays in Kaschin-Beck disease pathogenesis merit further study.
Animals ; Cell Survival ; Cells, Cultured ; Chondrocytes ; pathology ; Fermentation ; Fusarium ; drug effects ; physiology ; Rabbits ; Selenium ; pharmacology

To study the role of hematopoietic microenvironment abnormality in development of minimal residual disease and its mechanism, the viability of HL-60 cells was investigated by means of bone marrow stromal cell culture system or co-culture system of bone marrow stromal cell with HL-60 cells and idarubicin (IDA), flow cytometry and ELISA. The results showed that viability of HL-60 cells gradually decreased along with the increase of IDA dose and prolongation of culture time. Amount of HL-60 cells co-cultured with leukemia bone marrow stramal cells was significantly increased as compared with that of the control (P < 0.05). Bone marrow stromal cells or stromal cell conditioned medium reduced the effect of IDA on HL-60 cells in culture. In conclusion, leukemia bone marrow stromal cells contribute to increasing resistance of HL-60 cells to chemotherapeutic agents, and play some role in developing minimal residual disease.
Bone Marrow Cells ; physiology ; Cell Survival ; drug effects ; Coculture Techniques ; Dose-Response Relationship, Drug ; Drug Resistance, Neoplasm ; HL-60 Cells ; drug effects ; Humans ; Idarubicin ; pharmacology ; Stromal Cells ; physiology

The change of endothelial cell viability due to corticosteroid treatment in stored rabbit corneas was investigated. Hydrocortisone was injected into the anterior chamber of enucleated eyeballs which were stored in a moist chamber. After 24,48, or 72 hours of storage, the cornea was removed and stained with trypan blue. The unstained endothelial cells were counted under the light microscope in order to determine the density of viable endothelial cells. The same procedures were done on the contralateral eye with normal saline injected into the anterior chamber instead of hydrocortisone as a control. The density of viable endothelial cells in the corticosteroid-treated group was higher than that of the control group by 1.75%,14.39%, and 27.40% in 24,45, and 72 hour-stored corneas, respectively.
Animals ; Cell Survival/drug effects ; Endothelium, Corneal/*drug effects ; Female ; Hydrocortisone/*pharmacology/physiology ; Male ; Rabbits ; Time Factors ; Tissue Preservation/*methods

OBJECTIVEWe investigated the role of endoplasmic reticulum stress (ERS) in silica-induced apoptosis in alveolar macrophages in vitro.

METHODSRAW264.7 cells were incubated with 200 μg/mL silica for different time periods. Cell viability was assayed by the MTT assay. Cell apoptosis was evaluated by DAPI staining, flow cytometry analysis, and Western blot analysis of caspase-3. Morphological changes in the endoplasmic reticulum were observed by transmission electron microscopy. The expression of ERS markers binding protein (BiP) and CCAAT-enhancer-binding protein homologous protein (CHOP) was examined by Western blotting and real-time PCR. As an inhibitor of ERS, 4-phenylbutyric acid (4-PBA) was used in the experiments.

RESULTSSilica exposure induced nuclear condensation and caspase-3 expression in RAW264.7 cells. The number of apoptotic cells increased after silica exposure in a time-dependent manner. Silica treatment induced expansion of the endoplasmic reticulum. In addition, the expression of BiP and CHOP increased in silica-stimulated cells. Furthermore, 4-PBA treatment inhibited silica-induced endoplasmic reticulum expansion and the expression of BiP and CHOP. Moreover, 4-PBA treatment attenuated nuclear condensation, reduced apoptotic cells, and downregulated caspase-3 expression in silica-stimulated cells.

CONCLUSIONSilica-induced ERS is involved in the apoptosis of alveolar macrophages.

Animals ; Apoptosis ; drug effects ; Butylamines ; Cell Survival ; drug effects ; Endoplasmic Reticulum Stress ; physiology ; Mice ; RAW 264.7 Cells ; Silicon Dioxide ; toxicity

Phagocytosis or endocytosis by macrophages is critical to the uptake of fine particles, including nanoparticles, in order to initiate toxic effects in cells. Here, our data enhance the understanding of the process of internalization of silver nanoparticles by macrophages. When macrophages were pre-treated with inhibitors to phagocytosis, caveolin-mediated endocytosis, or clathrin-mediated endocytosis, prior to exposure to silver nanoparticles, Interleukin-8 (IL-8) production was inhibited. Although cell death was not reduced, the inflammatory response by macrophages was compromised by phagocytosis and endocytosis inhibitors.
Cell Line ; Cell Survival/drug effects ; Endocytosis/*physiology ; Humans ; Interleukin-8/*metabolism ; Macrophages/drug effects/*metabolism ; Metal Nanoparticles/*chemistry ; Phagocytosis/*physiology ; Silver/*chemistry/pharmacology