This experiment was based on a full-length SAMDC sequence from the drought stress induced cDNA library of Cleistogenes songorica.Two methods,absolute quantification and relative quantification,were used to analyze the gene expression under different tissue and drought stress treatment by real-time quantitaive PCR experiments and were compared each other,while CsSAMDC gene was used as a case.Leaves and roots tissue were sampled from the plants at 0,4,6,8,and 10 d of drought stress and 1 and 4 d after rewatering.The 2-CT method was used to analyze the relative changes in gene expression from quantitative real-time PCR experiments.A standard curve was generated by the purified plasmid DNA.The primer concentration in the realtime PCR was optimized.All absolute quatification data were normalized by the normalization factor from the most stable house-keeping genes in C.songorica.Relative quantification results using 2-CT method showed that CsSAMDC transcripts increased 5.93?0.71 fold in roots,down regulated 0.62?0.13 fold in leaves,under drought stress.Absolute quantification results showed that CsSAMDC transcripts up-regulated in roots significantly under drought stress as 1ong as 8 to 1 0 days after drought stress,while down-regulated in both roots and leaves after drought stress within 6 days and 1 to 4 days after rewatering,compared with control.CsSAMDC transcipt showed similar tendency of gene expression compared by using relative and absolute quantification methods.

Objective To investigate the possible role of interstitial cell of Cajal (ICE)in the pathogenesis of diabetic gastroparesis and the protective effect of"Xiaopi prescription (消痞方)".Methods Fifty healthy male Spregue-Dawley (SD) rats were randomly divided into 5 groups (each n=10):normal, model,and 3 Xiaopi prescription groups:low,middle and high dosages.Diabetes was induced by intravenous injection of alloxan,and equal amount of normal saline was intravenously injected in the normal group.Gastric lavage method was used to administer the traditional Chinese medicine decoction of Xiaopi prescription in corresponding amount (5,10 and 20 g?kg~(-1)?d~(-1)) in respective low,middle and high dosage groups.In the normal control group and diabetic model group,only equal amount of normal saline was administered into the stomach.Gastric emptying rate was measured by method of nutritious semisolid paste;c-kit positive cells of ICC were quantitatively measured with immunohistochemistry assay and computer image analysis system;c-kit mRNA positive cells were quantitatively measured with in situ hybridization and computer image analysis system.Results①Gastric emptying rate:The rate was significantly lower in the model group than that in the normal control group (P0.05),but higher than those in the model group and the low dosage group (all P0.05).②c-kit immmunohistochemistry:c-kit positive cell presented yellow in color,and its membrane was stained yellow,this kind of cells primarily were distributed around the neural plexus in the inter-space between the circular and longitudinal muscular fibers, and around the ganglionic cells forming"sheath-like"structure.The results of numbers of c-kit positive cells in the various groups:the number of the cells in the model group was significantly lower than that in the normal group (P0.05),but the numbers in the former two dosage groups were obviously higher than those in the model group (all P0.05),being significantly lower than that in the normal group,middle and high dosage groups (all P0.05),but obviously higher than those in the model group (all P0.05),being significantly lower than that in the normal,middle and high dosage groups (all P

Objective To study the prevalence and trend of tuberculosis for related policy development in Shandong.Methods The population under the current study was randomly sampled,using both stratified clustering sampling and proportional population sampling methods,following the national survey protocol.A total of 35 clusters including about 1500 subjects per cluster were established,representing a population of 9.31 million.Questionnaire interview and chest X-ray exam were applied to all inhabitants above 15 years of age.Sputum microscopy and culture were given to all suspected cases with cough longer than 2 weeks or having abnormal X-ray results.Results In total,54 279 subjects were interviewed and examined,accounting for 95.78％ of eligible population.Of them,183 active pulmonary tuberculosis cases were identified,with 60.11％ asymptomatic.Two of the 35 (5.71％) clusters had no active tuberculosis cases found,and 24(68.57％) did not show smear positive results.The standardized prevalence rates of active,smear positive and bacteriologic positive pulmonary tuberculosis cases were 270.87/105,17.45/105 and 29.57/105,with the estimated case numbers as 211 900 (170 100-253 600),13 600 (5800-21 500) and 23 100 (13 200-33 000)respectively.Compared to the survey in 2000,the rates on smear positive and bacteriologic positive tuberculosis had decreased significantly,at a rate of 81.63％,and 75.56％ respectively.The rates in urban areas and in women decreased quickly than those in rural areas and in men.People living in the rural areas,being elderly or males,had significantly higher prevalence rates of tuberculosis.Conclusion Remarkable reduction of tuberculosis prevalence had been achieved despite the fact that tuberculosis remained a major public health problem in Shandong province.Symptomatic patients should be under more serious concern in order to improve the detection of early cases.More efforts should be given to rural population,especially elderly,male population.

OBJECTIVETo compare the efficacy differences between acupoint catgut-embedding combined western medicine (ACCWM) and simple western medicine (WM) for treatment of epilepsy of generalized seizures type.

METHODSFifty-two patients with epilepsy of generalized seizures type were randomly divided into an ACCWM group (n=30) and a WM group (n=22). Dazhui (GV 14), Yaoshu (GV 2), Jiuwei (CV 15), Qihai (CV 6) were selected in the ACCWM group as the main acupoints combined with routine antiepileptic drugs, Valpromide was taken orally, 2.0 g/d for adult, 50 mg/kg x d for child. The WM group was treated with routine antiepileptic drugs. The epilepsy score and the attack frequency one year before the treatment and one year during treatment were observed and the therapeutic effects in the two groups were compared.

RESULTSThe epilepsy score and the attack frequency were both decreased in the two groups (P<0.01, P<0.05), and the epilepsy score in the ACCWM group decreased significantly (6.57 +/- 3.29 vs 10.73 +/- 2.54, P<0.05). The total effective rate of 90.0% (27/30) in the ACCWM group was superior to that of 68.2% (15/22) in the WM group (P<0.05).

CONCLUSIONThe therapeutic effect of ACCWM is superior to that of simple western medicine in treating epilepsy of generalized seizures type. It can improve significantly the attack symptoms so as to enhance life quality of the patients with epilepsy.

Acupuncture Points ; Acupuncture Therapy ; Adolescent ; Adult ; Anticonvulsants ; therapeutic use ; Catgut ; Child ; Child, Preschool ; Combined Modality Therapy ; Epilepsy ; drug therapy ; therapy ; Female ; Humans ; Male ; Middle Aged ; Seizures ; drug therapy ; therapy ; Valproic Acid ; analogs & derivatives ; therapeutic use ; Young Adult

An expression system is described for high-yield production of recombinant soluble human FasL (shFasL) in Dictyostelium discoideum cells. DNA encoding amino acids 141 - 281 of hFasL was PCR amplified from cDNA derived from activated human neutrophils. The resulting product was fused with a DNA fragment encoding hCG-beta signal peptide and cloned in the expression vector pMB12neo. Dictyostelium strain AX3 was transfected with this plasmid, yielding a recombinant strain called AX3-pCESFL95-H3. In order to improve the shFasL expression level, pMB12neo was optimized by replacing its transcriptional terminator/ polyadenylation segment of the 2H3 gene with an actin8 terminator/polyadenylation segment, yielding derived expression vector pMB74. The recombinant Dictyostelium strain called AX3-pLu8 was generated with this new plasmid. When the recombinant cells were cultivated in a complex HL-5C medium, a cell density of (1.5 - 2) x 10(7)/mL was reached, and the shFasL level expressed by strains AX3-pCESFL95-H3 and AX3-pLu8 was 23.5 microg/L and 206 microg/L, respectively. By using a newly developed synthetic medium called SIH as culture medium, higher cell density of (4 - 5) x 10(7)/mL was achieved. Correspondently, 111 microg/L and 420 microg/L shFasL were secreted by recombinant strains AX3-pCESFL95-H3 and AX3-pLu8, respectively.
Animals ; Chorionic Gonadotropin, beta Subunit, Human ; genetics ; Culture Media ; Dictyostelium ; genetics ; growth & development ; metabolism ; Fas Ligand Protein ; biosynthesis ; genetics ; Humans ; Neutrophils ; metabolism ; Recombinant Proteins ; biosynthesis ; genetics

In order to study the important factors involved in cationic liposome-mediated gene transfer, Lipofectamine 2000 or DOTAP was evaluated using three types of cells (Hep-2, MCF-7 and SW-480) in vitro transfection efficiencies. Different properties of the two reagents were analyzed and compared by DNA arrearage assay and MTT assay. Both Lipofectamine 2000 and DOTAP had strong capability to combine with DNA; Lipofectamine 2000 can get higher transfection efficiency of the three cells by using GFP as report gene, meanwhile, DOTAP can also get higher transfection efficiency against Hep-2 cell. However, DOTAP showed lower transfection efficiency against MCF-7 and SW-480 cell. On the other hand, the cytotoxicity assay showed that over 85% cell viability of MCF-7 cell could be achieved both by Lipofectamine 2000 and DOTAP under the optimal transfection condition. Relatively speaking, Lipofectamine 2000 has very high transfection efficiency in a broad range of cell lines, but because of the special selectivity of cell type on liposome, DOTAP also has a broad application prospect.
Cell Line, Tumor ; Cell Survival ; drug effects ; DNA ; genetics ; Fatty Acids, Monounsaturated ; chemistry ; toxicity ; Gene Transfer Techniques ; Genes, Reporter ; Genetic Vectors ; Green Fluorescent Proteins ; metabolism ; Humans ; Lipids ; chemistry ; toxicity ; Quaternary Ammonium Compounds ; chemistry ; toxicity ; Transfection

Osteosarcoma is the most common primary malignant bone cancer in children and adolescents. Emerging evidence has suggested that the capability of a tumor to grow is driven by a small subset of cells within a tumor, termed cancer stem cells (CSCs). Although several methods have been explored to identify or enrich CSCs in osteosarcoma, these methods sometimes seem impractical, and chemotherapy enrichment for CSCs in osteosarcoma is rarely investigated. In the present study, we found that short exposure to chemotherapy could change the morphology of osteosarcoma cells and increase sarcosphere formation in vitro, as well as increase tumor formation in vivo. Furthermore, methotrexate (MTX)-resistant U2OS/MTX300 osteosarcoma cells were larger in size and grew much more tightly than parental U2OS cells. More importantly, U2OS/MTX300 cells possessed a higher potential to generate sarcospheres in serum-free conditions compared to parental U2OS cells. Also, U2OS/MTX300 cells exhibited the side population (SP) phenotype and expressed CSC surface markers CD117 and Stro-1. Notably, U2OS/MTX300 cells showed a substantially higher tumorigenicity in nude mice relative to U2OS cells. Therefore, we conclude that chemotherapy enrichment is a feasible and practical way to enrich osteosarcoma stem cells.
Animals ; Antigens, Surface ; metabolism ; Antimetabolites, Antineoplastic ; pharmacology ; Bone Neoplasms ; metabolism ; pathology ; Cell Line, Tumor ; Cell Proliferation ; Drug Resistance, Neoplasm ; Humans ; Methotrexate ; pharmacology ; Mice ; Mice, Nude ; Neoplasm Transplantation ; Neoplastic Stem Cells ; drug effects ; pathology ; Osteosarcoma ; metabolism ; pathology ; Phenotype ; Proto-Oncogene Proteins c-kit ; metabolism

OBJECTIVETo explore the effect of the improved plastic and reconstruction of the anus in situ.

METHODSImproved plastic and reconstruction of anus in situ was performed in 38 cases of low rectal cancers operated while Miles radical operation. Improvement includes: (1) The internal sphincter was rebuilt with 4 layers of muscle layer of the endmost of colon. (2) The last of gracilis was divided into 2 parts to reconstruct the superficial part and deep part of external sphincter muscle. (3) The rectum cape improvement is to firmly stitch the levator ani outside the external sphincter muscle in front of the colon. (4) The rectum valve is improved into three artificial rectum valves.

RESULTSThe form and function and their long term survival rate were good, the rate of superior anus function was 94.73%.

CONCLUSIONIt mains the results of improved plastic and reconstruction of anus in situ is near that of normal persons.

Adult ; Aged ; Anal Canal ; surgery ; Female ; Humans ; Male ; Middle Aged ; Reconstructive Surgical Procedures ; methods ; Rectum ; surgery

OBJECTIVETo observe the dynamic changes of CD3+, CD4+, and CD8+ T lymphocytes in the peripheral blood of patients with sepsis and discuss the clinical significance.

METHODSSixty-four patients admitted in the Emergency Center and Emergency Intensive Care Unit of the Second Hospital of Wenzhou Medical University between August, 2007 and July, 2009 were enrolled in this study. CD3+, CD4+, and CD8+ T lymphocytes in the peripheral blood were detected by flow cytometry on days 1, 7 and 14 after admission, and the results were compared between the patients with improvement of the condition and those without improvement, with 20 healthy subjects as the control group.

RESULTSOn day 1 after admission, CD3+ and CD4+ T lymphocytes and CD4+/CD8+ T cell ratio were obviously lower in the 2 groups of patients with sepsis than in the control group (P<0.05), but no significant difference was found in CD8+ T lymphocytes. The sepsis patients with clinical improvement showed significant higher CD3+ and CD4+ T lymphocyte percentages and CD4+/CD8+ T cell ratio than those without improvement on day 1. In the patients with clinical improvement, CD3+ and CD4+T lymphocytes and CD4+/CD8+ T cell ratio increased gradually with time and till day 14, they were comparable with the control levels; in the patients without improvement, CD3+ and CD4+ T lymphocytes and CD4+/CD8+ T cell ratio showed no obvious alterations in the course of observation.

CONCLUSIONImmune imbalance occurs in patients with sepsis represented by lowered CD3+ and CD4+T lymphocyte percentages and CD4+/CD8+ T cell ratio in relation to the severity of the condition. CD3+ and CD4+ T lymphocytes and CD4+/CD8+ T cell ratio can be used as the indicators for assessing the severity of sepsis.

Adult ; Aged ; Aged, 80 and over ; CD4-CD8 Ratio ; Case-Control Studies ; Female ; Flow Cytometry ; Humans ; Male ; Middle Aged ; Sepsis ; blood ; immunology ; T-Lymphocyte Subsets ; cytology

Cholesterol Ester Storage Disease/genetics* ; Humans ; Mutation ; Sterol Esterase/genetics*