To establish induction and liquid culture system for hairy roots of Danshen (Salvia miltiorrhiza), Agrobacterium rhizogenes A4, LBA9402, 15834 as test bacterium were used to infect aseptic leaves of Danshen. The hairy roots were induced and positive transgenic hairy roots were selected with PCR using rolB and rolC as the target gene. Then hairy roots of S. miltiorrhiza were harvested and salvianolic acids were extracted with 70% methanol containing 1% formic acid. The content of salvianolic acid B (SalB) and rosmarinic acid (RA) were determined by HPLC. According to the above research results, the Danshen hairy roots induced by A. rhizogenes LBA9402 were inoculated into the following group of culture media: MSOH, MS, B5, and 6,7-V liquid media. Then the same methods of extraction and determination for the content of Danshen hairy roots were adopted. Last, the hairy roots of S. miltiorrhiza induced by A. rhizogenes LBA9402 were inoculated into the MSOH liquid media with different pH values. The content of salvianolic acid were extracted with 70% methanol containing 1% formic acid and determined by HPLC. As a result, three kinds of A. rhizogenes A4, LBA9402, 15834 could induce hairy roots and Ri plasmids were integrated into the genome of S. miltiorrhiza by PCR. Danshen hairy roots induced by A. rhizogenes LBA9402 and A4 produced much more salvianolic acid, which were (3.27 ± 0.37)% [including (1.04 ±0.36)% of RA and (2.22 ± 0.29)% of SalB] and (3.17 ± 0.20)% [including (0.92 ± 0.31)% of RA and (2.25 ± 0.26)% of SalB], respectively. Hairy roots induced by A. rhizogenes LBA9402 when they were cultured in MSOH liquid media produced much more salvianolic acid, which was (4.56 ± 0.36)%, including (1.12 ± 0.26)% of RA and (3.44 ± 0.23)% of SalB. Hairy roots induced by A. rhizogenes LBA9402 produced the most salvianolic acid when they were cultured in MSOH liquid media with the pH value 4.81, which was 4.85%, including 1.16% of RA and 3.69% of SalB. So Danshen hairy roots induced by A. rhizogenes LBA9402 and A4 produced much more salvianolic acid when they were cultured in MSOH liquid media with the pH value 4.81. The research had established the foundation on genetic engineering to improve the quality of S. miltiorrhiza.
Agrobacterium ; physiology ; Benzofurans ; analysis ; metabolism ; Cell Culture Techniques ; instrumentation ; methods ; Cinnamates ; analysis ; metabolism ; Culture Media ; chemistry ; metabolism ; Depsides ; analysis ; metabolism ; Drugs, Chinese Herbal ; analysis ; metabolism ; Plant Roots ; chemistry ; growth & development ; metabolism ; microbiology ; Salvia miltiorrhiza ; chemistry ; growth & development ; metabolism ; microbiology

AIM: To define the indications of C2 pedicle lag screw or C2 pedicle screw associated with C3 lateral mass screw to treat Hangman fracture, and evaluate the clinical applications. METHODS: Thirty-four patients with Hangman fracture were selected from Changzheng Hospital between February 2000 and December 2004, including 10 patients of type Ⅰ, 12 patients of type Ⅱ, and 8 patients of type Ⅱa, 4 patients of type Ⅲ. Levine-Edwards classification and graded: 2 patients of grade C spinal injury, 12 patients of grade Ds and 20 patients of grade E according to Frankel scale. Patients with typeⅠHangman's fracture were fixed by C2 pedicle lag screw, and patients with typeⅡ, Ⅱa and Ⅲ Hangman fracture were fixed by C2 pedicle screw associate C3 lateral mass screw. Postoperative neurofunctional recovery and fracture healing were studied. RESULTS: A total of 34 subjects were involved in the analysis of results. ① Frankle classification of postoperative neurofunctional recovery: There were 2 cases of grade C, 1 case recovered to grade E, and one case recovered to grade D. There were 12 cases of grade D, including 11 cases of recovered to grade E and one without change. ② In 3-6 months follow-up, X-ray of anterior-posterior and lateral position showed that all fractures healed without loose screw, vertebra artery injury or other complications. CONCLUSION: C2 pedicle lag-screw, C2 pedicle screw associated with C3 lateral mass screw are safe, effective in the treatment of Hangman fracture, which can to retain the function of upper cervical spine as great as possible.

italic>Salvia miltiorrhiza Bunge is a traditional Chinese medicinal herb widely used to treat cardiovascular and cerebrovascular diseases at clinic. Its main water-soluble components are rosmarinic acid (RA) and salvianolic acid B (SAB), which are produced by phenylpropanoid pathway. 4-Hydroxyphenylpyruvate reductase (HPPR) is a key enzyme in phenylpropanoid metabolism pathway. SmHPPR1 was cloned from S. miltiorrhiza and was constructed into plant expression vector pJR-SmHPPR1. On this basis, SmHPPR1 transgenic Arabidopsis plants were induced and the content of 4-hydroxyphenyllactic acid (pHPL) was determined. SmHPPR1-overexpressing (SmHPPR1-OE) hairy roots of S. miltiorrhiza were obtained and the concentration of active components and transcriptome analysis were performed. The results showed that the concentration of pHPL in SmHPPR1 transgenic Arabidopsis T1 was 0.594 mg·g^-1 dry weight. The concentration of RA, SAB and total salvianolic acid in SmHPPR1-OE-3 hairy roots were 1.09, 1.29, 1.15 times of that in control-3, respectively, and the content of Danshensu was 36.26% of that in control-3. Transcriptomic analysis revealed that overexpression of SmHPPR1 caused the upregulation of other phenylpropanoid pathway genes like SmTAT2. Protein-protein interaction indicated CYT (TR74706_c0_g1), NADP⁺ (TR26565_c0_g1) and NADP⁺ (TR68771_c0_g1) is the central node of the network and participated in metabolic process and cellular process. The tracking work in this study proved that SmHPPR1 could catalyze the reduction of 4-hydroxyphenylpyruvic acid to 4-hydroxyphenyllactic acid in SmHPPR1 transgenic Arabidopsis, and SmHPPR1-overexpressing in hairy roots of S. miltiorrhiza could increase the concentration of salvianolic acids through synergistically regulating other pathway genes.

Objective To investigate the geographical distribution of excessive iodine in drinking water and to demarc aI|e the excessive iodine regions in Hebei Province.Methods In 173 counties of Hebei Province, town was surveyed as an elementary unit.Five villages were sampled according to the direction of east,west, south,north,center in every town.If the water sources were more than 5 in a village,water was sampled according to the direction ofeast,west, south,north, center, respectively;If the water sources were less than 5 in a village, all were sampled:If the village used water of cental supply,only one sample wag collected.Arsenic-cerium catalysis was used to determine the iodine concentration of drinking water.Results Two thousands and forty-nine towns were investigated and 19 352 water samples were determined in Hebei Province.One hundred and seventy- two towns were confirmed to be excessive iodine areas,where the range of water iodine Wag 0.1～2840.4μg/L The water 8amDle with 18 358 had a water iodine median less than 150.0μg/L The areas with water iodine median less than 150.0μg/L,in between 150.0μg/L and less than 300.0μg/L,and equal to or more than 300.0 μg/L accounted for 94.86%,2.92%and 2.22%,respectively.There were 110 towns with water iodine median being between 150.0μg/L and iess than 300.0 μg/L and 62 towns with water iodine median equal to or more than 300 gμ/L.Exeessive iodine towns distributed in 33 counties of 6 cities involving 5 854 960 residents in'Hebei Provmce. MOBt of the water sources with excessive iodine Cangzhou were deep wells and there was a positive correlation between we depth and water iodine concentration (r=0.430,P<0.01), while they were shallow wells in Handan, Xingtai.Hengshui,and no correlation was found between well depth and water iodine concentration(r=-0.060,-0.119.-0.121,P>0.05).Conclusions Six cities have excessive iodine water resotlrees in Hebei Province, mostly in Handan.xingtai,Hengshui and Cangzhou Cities,which all are low-lying land.The water with excessive iodine is hypogene,and the towns with excessive iodine water distributein patchy or spot pattern.

Ischemic stroke is one of the most important causes of death and disability in humans,but the effective methods for treating brain injury after stroke are quite limited.Sphingosine 1-phosphate (S1P) is a pleiotropic lipid.There is certain interdependence between its metabolism and regulation and the molecular mechanisms involved in important biological events following cerebral ischemia.Membrane lipid therapy with S1P as the core may be an effective neuroprotective strategy of ischemic stroke.

Objective To explore knowledge, attitude, practice (KAP) and related determinants on nutrition among caregivers of those rural stranded children under 7 years of age in China and to provide evidence for setting up relevant health education program. Methods 1691 caregivers of the stranded children randomly selected were surveyed by a standard questionnaire. Logistic regression models were used to screen the determinants on KAP regarding nutrition. Results Rates on awareness, positive attitude and approprite behavior were lower in caregivers of children whose parents both left (47.8%, 55.4%, 41.8%, respectively) the countryside, when compared with those only one parent was away from home (59.9%, 59.5%, 38.0%, respectively). Data from multivariable logistic regression models showed that caregivers' KAP on nutrition was related to age, educational background, average family income, and willingness on the job as well as the age of the child. Conclusion Improving caregivers' KAP on nutrition and setting up appropriate health education program were in urgent need.

OBJECTIVETo investigate the role of c-Jun N-terminal kinase (JNK) signal transduction pathway in the rats of nonalcoholic fatty liver disease (NAFLD).

METHODSSixty four Sprague-Dawley rats were randomly divided into four groups: 8-week control group (NG8w), 12-week control group (NG12 w), 8-week high-fat diet (HG8w), and 12-week high-fat diet group (HG12w), with 16 rats in each group. Glucose infusion rate (GIR) was tested by euglycemic hyperinsulinemic clamp; aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG), total cholesterol (TC), free fatty acid (FFAs), fast insulin (FIns), tumor necrosis factor alpha (TNF alpha), superoxide dismutase (SOD) and malondialdehyde (MDA) were tested by biochemistry automatic analyzer or RIA; The expression of JNK1, insulin receptor substrate-1 (IRS-1), phospho-IRS-1 Ser307 (p-IRS-1 Ser307), Protein kinase B (PKB) and phospho-PKB Ser473 (p- PKB Ser473) were detected by Western blot.

RESULTSCompared to control group, body weight, liver index, serum levels of ALT, AST, TG, TC, FIns, FFAs, TNF alpha, and TC, TG FFAs, MDA in liver homogenates were increased, while the level of SOD, and GIR were decreased. The expression of JNK1 protein and p-IRS-1 Ser307 in liver tissue was up-regulated, while expression of p-PKB Ser473 was decreased (P < 0.05). A positive correlation was found between the expression intensity of JNK1 and IR (Pearson correlation: 0.718, P < 0.01).

CONCLUSIONThe high-fat could induce the expression of JNK1, which in turn modulates the phosphorylation of proteins in the insulin signaling pathway, and induces insulin resistant.

Alanine Transaminase ; blood ; Animals ; Biomarkers ; blood ; Dietary Fats ; administration & dosage ; Disease Models, Animal ; Fatty Liver ; etiology ; metabolism ; pathology ; Insulin Receptor Substrate Proteins ; metabolism ; Insulin Resistance ; JNK Mitogen-Activated Protein Kinases ; metabolism ; Lipids ; blood ; Liver ; metabolism ; pathology ; Male ; Proto-Oncogene Proteins c-akt ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Signal Transduction

OBJECTIVETo set up the HPLC fingerprint of Glycyrrhiza uralensis in decoctions prepared from various combinations of the recipe named Mahuang decoctions and study the influence of compatibility on fingerprint of G. wralensis in decoctions.

METHODThe RP-HPLC method was used, chromatography conditions were Zorbax SB-C18 column (4.6 mm x 250 mm, 5 microm), the acetonitrile-1.2% acetic acid (gradient elution) as mobile phase and detective wavelength at 254 nm.

RESULTThe method is stable and reliable with a good reproducibility and provides a reference standard for the quality control of G. uralensis, it can be used to determine the fingerprint of G. aralensis in Mahuang decoctions. The result showed that 8 peaks were common in the fingerprint of G. uralensis in Mahauang decoctions. The area changes of 8 common peaks in decoctions prepared from various combinations of Mahuang decoctions were studied.

CONCLUSIONThe fingerprint of G. uralensis in Mahuang decoctions is markedly influenced by herba ephedrae and ramulus cinnamomi and semen armeniacae amarum. The areas of 7 common peaks were reduced obviously, while the relative areas of 6 common peaks were increased remarkably.

Chromatography, High Pressure Liquid ; methods ; Cinnamomum ; chemistry ; Drug Combinations ; Drug Interactions ; Drugs, Chinese Herbal ; chemistry ; Ephedra sinica ; chemistry ; Glycyrrhiza uralensis ; chemistry ; Plants, Medicinal ; chemistry ; Prunus ; chemistry ; Quality Control ; Reproducibility of Results

OBJECTIVETo explore curative effects of advanced elastic intramedullary nail technology in treating children with femoral subtrochanteric fracture.

METHODSForm March 2009 to December 2010, 16 children with femoral subtrochanteric fracture were treated with advanced elastic intramedullary nail technology. There were 12 males and 4 females ranging in age from 7 to 15 years old with a mean of 9.8 years old. According to fracture classification, 9 cases were stable fractures (traverse or short oblique type) and 7 cases were unstable fractures (comminuted or long oblique type). Radiographs and complications were retrospectively reviewed. Fracture healing were evaluated according to Flynn criteria.

RESULTSAll children were followed up from 11 to 32 months with an average of 21 months. No incision infecton,bone nonunion and breakage of screw occurred. The fracture healing time was from 8 to 16 weeks with an average of 10.1 weeks. According to Flynn criteria, 14 cases got excellent results, 1 moderate and 1 poor.

CONCLUSIONAdvanced elastic intramedullary nail technology for the treatment of children with femoral subtrochanteric fracture is a safe and effective fixation, which is good for early mobilization, shorter the duration of hospital stays and reduce complications.

Adolescent ; Child ; Female ; Follow-Up Studies ; Fracture Fixation, Intramedullary ; methods ; Hip Fractures ; surgery ; Humans ; Male