The protoplasts of Micromonospora purpurea,the high yield strains of gentammicin producer were mutagenized by diethyl sulfate(DES)and ultraviolet radiation(UV)respectively,then fused,screened by gentamicin resistance and regenerated.The average fermentation unit 2200?U/ml could be achieved by shake flask for 10 batches.The average fermentation unit 1900?U/ml could be obtained by 5L fermentor for 7 batches.The quality of the end product conformed to CP2000,BP2000 and USP26 pharmacopoeia.

Currently, the randomized controlled trial (RCT) becomes the generally accepted golden standard for clinical trials. It can be classified into two types based on different design, that is, the explanatory randomized controlled trial (ERCT) and the pragmatic randomized controlled trial (PRCT). In designing a clinical trial, researcher may select either of them according to the goal of research. The design feature of PRCT and its application in designing clinical trials of acupuncture were introduced in this paper. Further attention should be paid on resolvent of how to apply PRCT in combining with other design methods organically to work out clinical trial scheme with high quality and consistent with the characteristics of acupuncture as possible, for evaluating the effect of acupuncture more accurately and objectively, so as to promote the international application and development of acupuncture therapy.
Acupuncture Therapy ; methods ; Biomedical Research ; Humans ; Randomized Controlled Trials as Topic ; methods

To explore the functions of human ribonuclease 9 (RNase 9), we constructed a mammalian fusion expression vector pcDNA-hRNase9, prepared recombinant human RNase 9-His fusion protein from HEK293T cells and determined its N-terminal amino acid sequences. According to the determined mature protein, recombinant human RNase 9 was prepared in E. coli. Ribonucleolytic activity and antibacterial activity of recombinant human RNase 9 were detected, and the distribution of human RNase 9 on tissues and ejaculated spermatozoa and in vitro capacitated spermatozoa were analyzed via indirect immunofluorescence assay. The results showed that recombinant human RNase 9 did not exhibit detectable ribonucleolytic activity against yeast tRNA, but exhibited antibacterial activity, in a concentration/time dependent manner, against E. coli. Immunofluorescent analyses showed that the predicted human RNase 9 was present throughout the epididymis, but not present in other tissues examined, and human RNase 9 was also present on the entire head and neck regions of human ejaculated spermatozoa and in vitro capacitated spermatozoa. These results suggest that human RNase 9 may play roles in host defense of male reproductive tract.
Adult ; Amino Acid Sequence ; Anti-Infective Agents ; metabolism ; Blotting, Western ; Cloning, Molecular ; Enzyme-Linked Immunosorbent Assay ; Epididymis ; enzymology ; Escherichia coli ; enzymology ; Genetic Vectors ; Humans ; Male ; Molecular Sequence Data ; Recombinant Fusion Proteins ; chemistry ; metabolism ; Ribonuclease, Pancreatic ; metabolism ; Ribonucleases ; chemistry ; metabolism ; Seminal Plasma Proteins ; chemistry ; metabolism ; Spermatozoa ; metabolism ; Testis ; enzymology ; Young Adult

OBJECTIVETo construct a method to culture pancreatic progenitor cells in hanging drop and on floating filter,and to examine if pancreatic progenitor cells can differentiate into mature endocrine cells with this method.

METHODSMurine embryos at day 12.5 were isolated and digested into single cells,which were then cultured in hanging drop for 24h and formed spheres.Spheres were cultured on the filter for 6 days,which floated in the dish containing medium.During culture,the expressions of pancreas duodenum homeobox-1(PDX-1)and neurogenin3(Ngn3)were determined.The expressions of endocrine and exocrine markers,insulin,glucagon,and carboxypeptidase(CPA)were determined on day 7 by immunohistochemistry.Insulin secretion of spheres stimulated by glucose was detected by ELISA.The changes of pancreatic marker expressions during culture were monitored by real-time polymerase chain reaction(PCR).

RESULTSOne day after the culture,there were still a large amount of PDX-1 positive cells in pancreatic spheres,and these cells proliferated.On day 3,high expression of Ngn3 was detected,and the Ngn3-positive cells did not proliferate.On day 7,The expressions of endocrine and exocrine markers in the differentiated pancreatic progenitor cells were detected,which were consistent with that in vivo.Insulin was secreted by spheres upon the stimulation of glucose.

CONCLUSIONIn hanging drop and on floating filter,pancreatic progenitor cells can differentiate into mature endocrine cells.

Animals ; Cell Culture Techniques ; Cell Differentiation ; Cells, Cultured ; Endocrine Cells ; cytology ; Homeodomain Proteins ; metabolism ; Insulin ; metabolism ; Mice ; Pancreas ; cytology ; Stem Cells ; cytology ; Trans-Activators ; metabolism

OBJECTIVETo find out how to prepare high-density dental ceramics through isostatic pressing so that sintering shrinkage will be reduced.

METHODSTo prepare Al2O3/ZrO2 composite powder first, then to mold through dry-pressing, and to shape the green-body through isostatic pressing. The green-bodies were sintered at the temperature of 1 400 degrees C and kept at the temperature for different period of time (2 h, 3 h, 4 h). After that, the density and fracture strength were measured and the microstructure observed by scanning electron microscope (SEM).

RESULTSThe sample product's density, line-shrinkage, and fracture strength of ceramics was rising with the sintering time lengthened. The sample product kept under the temperature of 1 400 degrees C for 4 hours, the fracture strength was (497.27 +/- 78.45) MPa and glass phase distributed evenly in the ceramics and the grains were integrated owing to the glass phase. The longer the sintering time, the more even the microstructure was.

CONCLUSIONThe sintering quality and the efficiency were improved through isostatic pressing.

Ceramics ; Dental Materials ; Glass ; Temperature

OBJECTIVETo study the differential gene expression profiles related to toxic effects in rats exposed to silica.

METHODSWistar rats exposed to SiO2 (50 mg/ml) and 1 ml normal saline by intratracheal injection served as the exposure and control groups, on the 14th day after exposure all rats were executed and the rat lung tissues were obtained. The differential gene expression profiles in the lung tissues of rats exposed to silica were detected using confocal fiber beads gene chip technique, and the differential expression profiling data were analyzed using the database for annotation, visualization and integrated discovery (DAVID) bioinformation analysis tool.

RESULTSThe results of present study indicated that 1567 genes with differential expression were identified in 22107 genes of rat lung tissues in exposure group, including 765 up-regulated genes and 802 down-regulated genes as compared to control group. In the 461 genes related to toxic effects, 285 genes were up-regulated and 176 genes were down-regulated in exposure group. The trends of up-regulation of HMOX1 and SOD2 genes in RT-PCR assay were similar to those in gene chip technique.

CONCLUSIONA large number of genes related to toxic effects in the rats with silica-induced pulmonary fibrosis appeared up-regulation or down-regulation. There may be a complex gene regulation network in the pulmonary fibrosis induced by SiO2, and the toxicological mechanism is an important part in the development of pulmonary fibrosis.

Animals ; Lung ; drug effects ; metabolism ; Male ; Oligonucleotide Array Sequence Analysis ; Pulmonary Fibrosis ; chemically induced ; genetics ; metabolism ; Rats ; Rats, Wistar ; Silicon Dioxide ; toxicity ; Transcriptome

OBJECTIVETo transfer the effective elements of Bupleurum scorzonerifolium into carrot, and provide theoretical data for the exploitation, improvement and selection of the germplasm of Chinese medicinal plants.

METHODThe protoplasta of Bupleurum scorzonerifolium irradiated by ultraviolet light (UV) at an intensity of 300 microW.(cm2)-1 for 0, 1, 2 min respectively were fused with those of carrot Fisch by PEG method. The regenerated clones, derived form a single fused cell, were examined for their hybrid nature by phenotype and Esterase isoenzyme analysis.

RESULTNine clones were identified as the somatic hybrids between B. scorzonerifolium and carrot.

CONCLUSIONThis provides a firm foundation for the further analysis of the main active components saikosaponin of somatic hybrids and the screening out of high-medicine-content hybrid cell lines.

Bupleurum ; cytology ; genetics ; growth & development ; Cell Fusion ; Culture Techniques ; Daucus carota ; cytology ; genetics ; growth & development ; Esterases ; analysis ; Hybrid Cells ; enzymology ; Hybridization, Genetic ; Isoenzymes ; analysis ; Plants, Medicinal ; cytology ; genetics ; growth & development ; Protoplasts ; cytology

OBJECTIVETo investigate the protective effect of suppressive oligodeoxynucleotides (Sup ODN) on interferon-γ (IFN-γ) and signal transducers and activators of transcription (pSTAT4) expression of Silica-induced pulmonary inflammation in Mice.

METHODSSixty Balb/c mice were randomly divided into 4 groups, normal control group, silicious group, suppressive oligodeoxynucleotides (Sup ODN) group, control oligodeoxynucleotides (Con ODN) group. Except the normal control group injected normal saline, the rest groups were induced by the intratracheal instillation of 0.1 ml (5 g/L) of sterilized silica suspension. Sup ODN group and Con ODN group were treated by i.p. injection of 0.3 ml (1mg/mL) of suppressive or control ODN 3 h before silica administration. After 7 days, the animals were killed and levels of IFN-γ were detected by ELISA. The pathologic changes in lung tissues of mice were observed with HE staining. Expressions of IFN-γ and pSTAT4 in lung tissue were detected with immunohistochemistry and quantified by Image-Pro Plus 7.0.

RESULTSHE staining showed that the lung tissue of silicious group were damaged seriously than Sup ODN group. Compared with the normal control group (serum: (280.1±41.3) pg/ml, lung tissue: (0.249±0.373), IFN-γ increased in silicious group (serum: (886.3±81.7) pg/ml, lung tissue: (0.270±0.300) (P < 0.05). Compared with the normal control group and Con ODN group [(894.5±91.6) pg/ml], IFN-γ in the serum of Sup ODN group decreased significantly (P < 0.01). Compared with the silicious group , IFN-γ in lung tissue decreased in Sup ODN group (0.241±0.250) (P < 0.05). Compared with the normal control group (0.279±0.353), pSTAT4 in lung tissue increased significantly in silicious group (0.313±0.231) (P < 0.01). Compared with the silicious group, pSTAT4 in lung tissue decreased significantly in Sup ODN group (0.269±0.523) (P < 0.01).

CONCLUSIONSup ODN attained protective effect on Silica treated mice by suppressing expression of IFN-γ and pSTAT4.

Animals ; Female ; Inflammation ; metabolism ; Interferon-gamma ; metabolism ; Lung ; drug effects ; metabolism ; pathology ; Mice ; Mice, Inbred BALB C ; Oligodeoxyribonucleotides ; pharmacology ; Phosphorylation ; STAT4 Transcription Factor ; metabolism ; Silicon Dioxide ; toxicity

OBJECTIVETo explore the relationship between the change in neuron specific enolase (NSE) and brain malfunction in burned patients.

METHODSThe serum samples of 11 burned patients with brain dysfunction were collected for the development of the serum level of neuron specific enolase with radioimmunoassay, and the correlation between condition of systemic inflammation and the levels of neuron specific enolase was assessed.

RESULTSThe level of NSE in burn patients with cerebral malfunction was obviously higher than that in control, and the level was correlated with the systemic inflammation.

CONCLUSIONThe change in the level of serum NSE could reflect the damage degree of central nervous system to some extent.

Adult ; Brain Diseases ; enzymology ; Burns ; enzymology ; Humans ; Male ; Middle Aged ; Phosphopyruvate Hydratase ; blood ; Systemic Inflammatory Response Syndrome ; enzymology

OBJECTIVETo establish a new culture method to induce the differentiation of embryonic pancreatic cells into mature endocrine cells.

METHODSMouse embryos at day 12.5 were used and embryonic pancreata were isolated. The isolated embryonic pancreata were cultured on the filter for 7 days, which floated in the dish containing medium. During culture, the expression of pancreas duodenum homeobox-1 (PDX-1), a pancreatic stem cell marker, was examined at day 1. The expression of neurogenin 3 (Ngn3), a pancreatic progenitor cell marker, was examined at day 3. The expressions of endocrine and exocrine markers, insulin, glucagon, and carboxypeptidase (CPA) were examined at day 7 by immunohistochemistry. The kinetics of pancreatic marker expression during culture was assayed by real-time PCR.

RESULTSMany pancreatic stem cells still existed in embryonic pancreata cultured for 1 day; meanwhile, these pancreatic stem cells proliferated in high rate. A large amount of pancreatic progenitor cells were found in embryonic pancreata cultured for 3 days.Pancreatic stem/progenitor cells differentiated into mature endocrine and exocrine cells in embryonic pancreata after having been cultured for 7 days. Furthermore, the expression pattern of pancreatic marker is consistent with that in vivo.

CONCLUSIONWe successfully established a new culture method, with which embryonic pancreatic cells can efficiently differentiate into mature endocrine cell.

Animals ; Basic Helix-Loop-Helix Transcription Factors ; metabolism ; Cell Culture Techniques ; Cell Differentiation ; Cells, Cultured ; Embryo, Mammalian ; Endocrine Cells ; cytology ; Female ; Homeodomain Proteins ; metabolism ; Male ; Mice ; Nerve Tissue Proteins ; metabolism ; Pancreas ; cytology ; Trans-Activators ; metabolism