Cloning of transcription factor PcFBA-1 in Pogostemon cabin and its interaction with FPPS promoter / 中国中药杂志

Farnesyl diphosphate synthase(FPPS) is a key enzyme at the branch point of the sesquiterpene biosynthetic pathway, but there are no reports on the transcriptional regulation of FPPS promoter in Pogostemon cabin. In the early stage of this study, we obtained the binding protein PcFBA-1 of FPPS gene promoter in P. cabin. In order to explore the possible mechanism of PcFBA-1 involved in the regulation of patchouli alcohol biosynthesis, this study performed PCR-based cloning and sequencing analysis of PcFBA-1, analyzed the expression patterns of PcFBA-1 in different tissues by fluorescence quantitative PCR and its subcellular localization using the protoplast transformation system, detected the binding of PcFBA-1 protein to the FPPS promoter in vitro with the yeast one-hybrid system, and verified its transcriptional regulatory function by dual-luciferase reporter gene assay. The findings demonstrated that the cloned PcFBA-1 had an open reading frame(ORF) of 1 131 bp, encoding a protein of 376 amino acids, containing two conserved domains named F-box-like superfamily and FBA-1 superfamily, and belonging to the F-box family. Moreover, neither signal peptide nor transmembrane domain was contained, implying that it was an unstable hydrophilic protein. In addition, as revealed by fluorescence quantitative PCR results, PcFBA-1 had the highest expression in leaves, and there was no significant difference in expression in roots or stems. PcFBA-1 protein was proved mainly located in the cytoplasm. Furthermore, yeast one-hybrid screening and dual-luciferase reporter gene assay showed that PcFBA-1 was able to bind to FPPS promoter both in vitro and in vivo to enhance the activity of FPPS promoter. In summary, this study identifies a new transcription factor PcFBA-1 in P. cabin, which directly binds to the FPPS gene promoter to enhance the promoter activity. This had laid a foundation for the biosynthesis of patchouli alcohol and other active ingre-dients and provided a basis for metabolic engineering and genetic improvement of P. cabin.

Effects of aqueous extracts of gecko on stemness of hepatocellular carcinoma / 中国中药杂志

To explore the effects and mechanism of aqueous extracts of gecko on cancer stem cells properties of hepatocellular carcinoma. In vitro, MTT assay was used to detect the cells growth in Huh7 and Hep3B. Spheroid-forming assay and flow cytometry were performed to observe the the stemness of Huh7 and Hep3B cells. The protein expressions of β-catenin, CD44, c-Myc, CCND1, Sox2, Oct4, Nanog and ABCG2 were detected by Western blot. Interacting proteins were detected by co-immunoprecipitation; and a subcutaneous xenograft model was used to detect the stemness of hepatoma carcinoma cells. The results indicated that aqueous extracts of gecko induced cell growth inhibition in a dose- and time-dependent manner, with the IC₅₀ of (0.750±0.112) g•mL⁻¹ for Huh7 and (0.454±0.039) g•mL⁻¹ for Hep3B, respectively. The number and size of tumor spheres formed by hepatoma carcinoma cells were decreased after treatment by aqueous extracts of gecko(P<0.05); the proportions of cells staining with putative markers for cancer stem cells, such as CD133 and CD44, were decreased(P<0.05). After treatment with aqueous extracts of gecko, the expression levels of β-catenin, CD44, c-Myc, CCND1, Sox2, Oct4, Nanog and ABCG2 were decreased. Co-immunoprecipitation results showed that the aqueous extracts of gecko could inhibit the interaction between LRP6 and Frizzled6, indicating that the aqueous extracts of gecko could inhibit the proliferation of hepatoma cells, the formation of tumor spheres and the proportion of tumor stem cells, and inhibit the Wnt signaling pathway by targeting LRP6 to prevent the formation of LRP6 and Frizzled6 complexes.

The effect of n-hexane on the gonad toxicity of female mice / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To investigate the toxic effects of n-hexane on the Ganod of female mice.</p><p><b>METHODS</b>n-Hexane was administered to four groups of mice by inhalation at doses of 0, 3.0, 15.1, and 75.8 mL/m3 respectivelyfor five weeks. Each group consisted of 10 mice, of which half were injected in first with 10 IU of pregnant mare serum gonadotrophin (PMSG) on the 33rd days, and then with 10 IU of human chorionic gonadotrophin (HCG) 48 hrs later. After the treatment, mouse sera were sampled and ovulating hormone (LH), follicle-stimulating hormone (FSH), estradiol (E2), and progesterone (P4) levels were measured by electrochemiluminescence immunoassays (ECLIA). In each group, the right ovaries of the non-super-ovulated mice were stained with hematoxylin and eosin while ovaries on the left side were prepared with the TUNEL method in order to detect apoptotic cells.</p><p><b>RESULTS</b>The duration of the diestrus stage decreased significantly (P < 0.05) in the 75.8 mL/m3 group. All super-ovulated mice in each treatment group produced fewer eggs than those in the control group (P < 0.05). The number of follicles in ovaries in the 75.8 mL/m3 group was smaller compared with the control group (P < 0.05).The serum P4 levels in each treatment group were lower than those in the control group (F = 6.196, P < 0.01). The cell apoptotic rate in the 75.8 mL/m3 group was higher (P < 0.05).</p><p><b>CONCLUSION</b>n-Hexane may have directly mediated via alterations hormone secretion and promoted granulosal cell apoptotic, which may be one of the important mechanisms for n-hexane induced mouse ovary impairment.</p>

Proteomic analysis of human fetal and adult brain cortex / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To study the differences of protein expression levels in the brain cortex of human fetus and adult with proteomics technique, and provide preliminary data on the change of proteins during brain development.</p><p><b>METHODS</b>Proteins extracted from human temporal lobes in fetal (3 month and 5 month respectively) and adult (30 years old) brain were separated by two-dimensional gel electrophoresis (2DE). The proteins were then stained with colloidal Coomassie blue to produce a high-resolution map of the proteiome. The differential protein spots were analyzed by PDQuest 7.0 software and 8 spots, which were gradually reduced or gradually increased in brain development process and the protein spots of difference over two-fold in the brain, were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF/TOF).</p><p><b>RESULTS</b>(1) On average, 642, 511 and 527 protein spots could be obtained in the temporal lobes of adult, 3 month and 5 month fetus. The matching rate of images was 87%. The basic proteins in adult brain were obviously much more than that in the fetus; (2) There were 172, 171 and 152 singular protein spots in temporal lobes of adult, 3 month and 5 month fetus respectively.(3) Compared with adult, there were 131 and 115 different protein spots in the 3 month and 5 month fetus respectively. There were 60 and 40 protein spots with more than 2 fold difference, among which 24 and 17 were down-regulated, and 36 and 23 were up-regulated respectively. (4) There was different expression in proteins such as serum albumin, triosephosphate isomerase, etc. in the 3 groups. Fatty acid binding protein 7 and unnamed proteins were only highly expressed in the 3 month brain; ribulose-1, 5-bisphosphate carboxylase/oxygenase large subunit and transducin beta-1 subunit were up-regulated in adult brain. Serum albumin decreases gradually with brain development. However, ATP synthase, mitochondrial F0 complex, and triosephosphate isomerase increase gradually with brain development.</p><p><b>CONCLUSION</b>The proteins of human brain cortex were obviously changed from embryonic stage to adult. The differentially displayed proteins may provide further insight into the understanding of development of human brain.</p>

Principles of ethics review on traditional medicine and the practice of institute review board in China / 中国结合医学杂志

As one of the significant parts of medical science research in China, the research on Chinese medicine (CM) reflects the essence of healthcare tradition in the country both theoretically and clinically, and embodies the values of Chinese culture. Therefore, in the practice of ethics review on CM research protocols, besides abiding by the contemporary prevalent international principles and guidelines on bioethics, which emphasizes the scientific and bioethical value of the study, we should also stress the CM theoretical background and relevant clinical experience in the framework of Chinese culture and values. In this paper, we went over the traits of CM clinical research and the experience from the practice of ethics review by the institution review board for bioethics, and then attempted to summarize the key points for the bioethics review to CM researches in China, so as to serve as reference for the bioethics review to traditional and alternative medicine researches.

Extracellular glycerol in patients with severe traumatic brain injury / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To study the factors affecting extracellular glycerol (Gly) in patients with severe traumatic brain injury (STBI).</p><p><b>METHODS</b>Perilesional extracellular Gly and cerebral blood flow (CBF) in 53 patients with STBI were consecutively monitored. Simultaneously, the intracranial pressure (ICP) and cerebral perfusion pressure (CCP) were monitored. The hourly minimum of CCP and CBF and the hourly maximum of ICP levels were matched with the hourly Gly. Gly values were divided into several groups according to regional ICP (less than 15 mm Hg or larger than 15 mm Hg), CCP (less than 70 mm Hg or larger than 70 mm Hg), CBF (less than 50 AU or 50-150 AU) and the outcomes (death or persistent vegetative state group, severe or moderate disability group, and good recovery group).</p><p><b>RESULTS</b>In comparison with the severe or moderate disability group, the Gly concentration of the death or persistent vegetative state group increased significantly, but CBF and CCP decreased significantly. In comparison with the good recovery group, the Gly concentration of the severe or moderate disability group increased significantly, but CBF and CCP decreased significantly. The Gly concentrations in patients with ICP larger than 15 mm Hg, CCP less than 70 mm Hg and CBF less than 50 AU were respectively higher than those of patients with ICP less than 15 mm Hg, CCP larger than 70 mm Hg and 50 AU less than CBF less than 150 AU. In patients with diffuse axial injury, the mean Gly concentration was (201.17+/-55.00) micromol/L, which was significantly higher than that of the patients with epidural hematoma (n equal to 7, 73.26+/-8.37, P less than 0.05) or subdural hematoma (n equal to 9, 114.67+/-62.88, P less than 0.05), but it did not increase significantly when compared with those in patients with contusion(n equal to 24, 167.48+/-52.63).</p><p><b>CONCLUSION</b>Gly can be taken as a marker for degradation of membrane phospholipids and ischemia, which reflects the severity of primary or secondary insult.</p>

Effect of mild hypothermia on glucose metabolism and glycerol of brain tissue in patients with severe traumatic brain injury / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To study the effect of mild hypothermia on glucose metabolism and glycerol of brain tissue in patients with severe traumatic brain injury (STBI) using clinical microdialysis.</p><p><b>METHODS</b>Thirty-one patients with STBI(GCS less than or equal to 8) were randomly divided into hypothermic group(Group A) and control group(Group B). Microdialysis catheters were inserted into the cerebral cortex of perilesional and normal brain tissue. All samples were analyzed using CMA microdialysis analyzer.</p><p><b>RESULTS</b>In comparison with the control group, lactate/glucose ratio(L/G), lactate/pyruvate ratio(L/P) and glycerol(Gly) in perilensional tissue were significantly decreased; L/P in normal brain tissue was significantly decreased. In control group, L/G, L/P and Gly in perilensional tissue were higher than that in normal brain tissue. In the hypothermic group, L/P in perilensional tissue was higher than that in relative normal brain.</p><p><b>CONCLUSIONS</b>Mild hypothermia protects brain tissues by decreasing L/G, L/P and Gly in perilensional tissue and L/P in "normal brain" tissues. The energy crisis and membrane phospholipid degradation in perilensional tissue are easier to happen after traumatic brain injury, and mild hypothermia protects brain better in perilensional tissue than in normal brain tissue.</p>

High-sensitive C-reactive protein and Insulin Resistance in Elderly Hypertension Patients / 中华高血压杂志

Objective To investigate the relationship between high-sensitive C-reactive protein(hsCRP)and insulin resistance(IR)in elderly hypertension patients.Methods The levels of ambulatory blood pressure mo- nitoring(ABPM),fasting plasma glucose(FPG),fasting insulin(FINS),von Willebrand factor(vWF),hsCRP were measured in elderly patients with hypertension alone(n=260),hypertension coexit with diabetes(n=230), and healthy matched subjects(n=250).Results The levels of FINS,FPG,vWF,hsCRP,systolic blood pres- sure(SBP),diastolic blood pressure(DBP)in the HT and HT+DM group were significantly higher than that in the NC group(P

Influence of temperature on extracellular glucose and lactate after traumatic brain injury / 中国医学科学院学报

<p><b>OBJECTIVE</b>To study the changes of extracellular glucose (Glu), lactate (Lac), and the ratio of lactate/pyruvate (L/P) in patients with traumatic brain injury under different body temperatures.</p><p><b>METHODS</b>Catheters for microdialysis were punctured into the penumbra zone of injured brain tissue (INJ), relatively normal brain tissue (NOR), and abdominal subcutaneous tissue (ABD) respectively in 51 patients to collect the fluid. The perfusion rate was 0.3 microl/min and one tube of fluid was collected for each hour. The average collection time was (67.10 +/- 18.27) hours. Concentrations of Glu, Lac, and pyruvate (Pyru) in the fluid were analyzed using CMA microdialysis analyzer. Patients were divided into 7 groups according to their rectal temperature (RT) values, which were RT < 33.0 degrees C, 33.0-33.9 degrees C, 34.0-34.9 degrees C, 35.0-35.9 degrees C, 36.0-36.9 degrees C, 37.0-37.9 degrees C, and > or = 38.0 degrees C.</p><p><b>RESULTS</b>The concentration of Glu in ABD was significantly higher than that in the brain tissue (P < 0.05). The Glu in NOR were significantly higher and the highest in 33.0 degrees C compared with that in the INJ when RT < 36.0 degrees C (P < 0.05). The concentration of Lac in ABD was significantly lower than that in brain (P < 0.05). The Lac in NOR were much higher than that in INJ when RT < 35.0 degrees C or > or = 37.0 degrees C (P < 0.05). The ratio of L/P decreased along with the increase of body temperature (P < 0.001). The ratio of L/P significantly decreased in an order of INJ > ABD > NOR when RT was lower than 33.0 degrees C, which was changed to the order of NOR > INJ > ABD when RT was higher than 34.0 degrees C.</p><p><b>CONCLUSION</b>Treatment of hypothermia may play more protective role when RT were between 33-34 degrees C or 36-37 degrees C.</p>

Expression vector for the inhibitor of growth-1 gene is constructed and the NLS-GFP fusion protein expresses in MRC-5 cells / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To construct the NLS(ING1)-GFP vector, transfer it into MRC-5 cells and establish a cell model expressing NLS (ING1)-GFP fusion protein.</p><p><b>METHODS</b>Firstly, cDNA fragment of nuclear locating sequence (NLS) of inhibitor of growth-1 gene (ING1) was gained by RT-PCR and inserted into multi-clone site of pEGFP-C1 to construct the NLS (ING1)-GFP expression vector. Then the vector was used to transfect the MRC-5 cells to observe the subcellular signal localization of green fluorescence protein (GFP).</p><p><b>RESULTS</b>We successfully constructed the expressing vector of NLS (ING1)-GFP fusion protein. After transferring the fusion expressing vector into MRC-5 cells, we observed that green fluorescence signal located in the cell nucleus. However, the green fluorescence signal located in the cytoplasm in MRC-5 cells transfected with pEGFP-C1 control only expressing GFP.</p><p><b>CONCLUSION</b>In living cells, physiologically p33 ING1b locates absolutely in nucleus. The p33(ING1b) NLS plays a decisive role in the transporting process of subcellular localization.</p>