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

Objective To investigate the photo-toxic effects of indocyanine green (ICG) based on photodynamic therapy (PDT) on vascular endothelial cells in vitro. Methods Human umbilical vein endothelial cells were incubated with ICG with different concentrations in 96-well plate. According to the dose of ICG , they were divided into 6 groups as 0, 50, 100, 150, 200 and 250 μmol/L group. Each group consists of 8 wells, 4 of them were irradiated with a 810 nm laser of total power 24.0 J/cm2. Cell viability was measured by MTT assay 24 h after light irradiation. Morphological changes in cells were observed by light microscope. Results With the irradiation of 810 nm light , there was a significant decrease in relative cell viability in all groups except 0 μmol/L group (all P < 0). With the increase of concentration of ICG, the cell viability dropped rapidly (P < 0). Apoptotic changes can be seen in all groups except 0 μmol/L group and necrosis was the main change in cells of 200 μmol/L and 250 μmol/L group. Conclusion PDT with appropriate concentration of ICG has a strong inhibition to vascular endothelial cells in vitro and ICG-PDT may be a good new therapy for nevascularization.

OBJECTIVETo study immunomodulating activity of Lonicera Japonica flavone by investigating immune enzymatic activity of serum and antoxidized activity of lymphoid organs in mice.

METHODSFifty KM mice were randomly divided into control group, model group, low dose group, middle dose group and high dose group(n = 10), respectively. And low dose group, middle dose group and high dose group were given Lonicera Japonica flavone with 100 mg/kg, 200 mg/kg and 400 mg/kg every day, respectively, while control group and model group were administered with NS. After continuously giving drug 7 weeks, other groups were injected with Dexamethasome (Dex: 25 mg /kg) for 3 days by subcutaneous injection, but the control group were treated with NS. And after giving Lonicera Japonica flavone 1 week simultaneously, organ indexes , the activity of acid phosphatase (ACP), alkaline phosphatase (AKP) and lysozyme (LSZ) in serum , and the content of monoamine oxidase (MAO), total antioxidant capacity (T-AOC), total superoxide dismutase (SOD) and malondialdehyde (MDA) in lymphoid organs in mice were tested, respectively.

RESULTSLonicera Japonica flavone could significantly improve the organ indexes, and significantly improve the activity of ACP, AKP and LSZ in serum, and significantly improve the contents of T-AOC and SOD, but reduce that of MAO and MDA in lymphoid organs in immunosuppressed mice.

CONCLUSIONIonicera Japonica flavone can significantly improve the activity of immune enzyme in serum and the antioxidized activity of lymphoid organs in mice. It suggests that Ionicera Japonica flavone has a good immunomodulatory effects.

Acid Phosphatase ; blood ; Alkaline Phosphatase ; blood ; Animals ; Antioxidants ; metabolism ; Flavones ; pharmacology ; Immunomodulation ; Lonicera ; chemistry ; Malondialdehyde ; metabolism ; Mice ; Monoamine Oxidase ; metabolism ; Muramidase ; blood ; Superoxide Dismutase ; metabolism

ObjectiveTo study the effect of behavior training on the expression of NR2B around the infarcted focus and in the cortex of temporal lobe in rats with bilateral hippocampal infarction.Methods54 SD rats were randomized into pre-training, 7 d, 14 d, 21 d after training groups, pre-immobilization, 7 d, 14 d, 21 d immobilization groups and control group. Behavior training and immobilization were performed on the 3rd day after the infarction. Immunohistochemistry was used to detect the expression of NR2B around the infarcted focus and in the cortex of temporal lobe at different points of time.ResultsThe expression of NR2B was abundant in the normal hippocampus and cortex of temporal lobe of rats. The expression of NR2B decreased after infarction and increased after behavior training. However, the expression of NR2B had little increased in immobilization groups, and showed significant difference compared with that in behavior training groups (P<0.01).ConclusionBehavior training can accelerate the expression of NR2B around the infarcted focus and in the cortex of temporal lobe in rats with bilateral hippocampal infarction.

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.

OBJECTIVETo study the curative effect of Zhiyang Pingfu Liquid (ZPL) in treating epidermal growth factor receptor inhibitors (EGFRIs) associated adverse reactions of the skin.

METHODSAll 54 patients with pathologically confirmed malignant tumor had EGFRIs induced adverse reactions of the skin to various degrees. ZPL was externally applied for them all, once or twice per day, 14 days consisting of one therapeutic course. Changes of adverse skin reactions, time for symptoms relief, adverse skin reaction types suitable for ZPL were observed before and after treatment.

RESULTSEGFRIs associated skin adverse reactions were improved to various degrees after they used ZPL. The shortest symptoms relief time was 1 day while the longest was 12 days, with an average of 6.93 days and the median time 7 days. Compared with before treatment, itching, rash/scaling, acne/acneform eruptions were obviously improved (P < 0.05).

CONCLUSIONZPL could alleviate EGFRls associated adverse skin reactions, especially showed better effect on itching, rash/scaling, acne/acneform eruptions.

Antineoplastic Agents ; adverse effects ; Biomedical Research ; Drugs, Chinese Herbal ; administration & dosage ; therapeutic use ; Exanthema ; chemically induced ; Humans ; Neoplasms ; drug therapy ; Pruritus ; Receptor, Epidermal Growth Factor ; antagonists & inhibitors ; Skin ; drug effects ; Skin Diseases ; drug therapy

The clinical efficacy and safety of auricular acupuncture (AA) for treatment of primary insomnia was evaluated. After a comprehensive retrieval in domestic and foreign databases, literatures were strictly screened and Revman 5.2 software was applied to perform a Meta-analysis on eligible randomized controlled trials (RCTs). The evidence quality was assessed with GRADE profiler 3.6 software. As a result, 8 articles were included involving 894 patients. Compared among AA and sham AA, placebo AA, blank control, there was significant difference in Pittsburgh sleep quality index (PSQI) [WMD = -3.48, 95% CI (-3.96, -3.00)], sleep latency LWMD = -10.14, 95% CI (-17.16, -3.12)] and sleep awakening times [WMD = -9.98, 95% CI (-1.10,-0.48)]. Compared between AA and western medication, there was significant difference in PSQI [WMD = -3.62, 95% CI (-4.59, -2.65)]. The evidence quality was moderate in AA vs. sham AA, placebo AA or blank control, while that of the rest was extremely low. No reports of adverse events were described in all studies. In conclusion, for the treatment of primary insomnia, AA could effectively improve sleep quality, but due to the low evidence quality, cautious attitude should be taken on this conclusion, and clinical trials with large sample and high quality were needed in the further.
Acupuncture, Ear ; Humans ; Randomized Controlled Trials as Topic ; Sleep ; Sleep Initiation and Maintenance Disorders ; physiopathology ; therapy

OBJECTIVETo explore the attention characteristics of children with different clinical subtypes of attention deficit hyperactivity disorder (ADHD) and to provide a basis for clinical intervention.

METHODSA total of 345 children diagnosed with ADHD were selected and the subtypes were identified. Attention assessment was performed by the intermediate visual and auditory continuous performance test at diagnosis, and the visual and auditory attention characteristics were compared between children with different subtypes. A total of 122 normal children were recruited in the control group and their attention characteristics were compared with those of children with ADHD.

RESULTSThe scores of full scale attention quotient (AQ) and full scale response control quotient (RCQ) of children with all three subtypes of ADHD were significantly lower than those of normal children (P<0.01). The score of auditory RCQ was significantly lower than that of visual RCQ in children with ADHD-hyperactive/impulsive subtype (P<0.05). The scores of auditory AQ and speed quotient (SQ) were significantly higher than those of visual AQ and SQ in three subtypes of ADHD children (P<0.01), while the score of visual precaution quotient (PQ) was significantly higher than that of auditory PQ (P<0.01). No significant differences in auditory or visual AQ were observed between the three subtypes of ADHD.

CONCLUSIONSThe attention function of children with ADHD is worse than that of normal children, and the impairment of visual attention function is severer than that of auditory attention function. The degree of functional impairment of visual or auditory attention shows no significant differences between three subtypes of ADHD.

Attention ; Attention Deficit Disorder with Hyperactivity ; psychology ; Child ; Female ; Humans ; Male

OBJECTIVETo evaluate the efficacy and toxicity of taxol-based chemotherapy in the treatment of advanced gastric cancer (AGC).

METHODSTwenty-nine patients with AGC treated with taxol-based protocols: taxol plus 5-fluouracil 17 patients, taxol plus cisplatin 10 patients, taxol plus epirubicin 2 patients.

RESULTSTwenty-six patients were evaluated for clinical response. There was no complete response, PR 10 (34.5%), SD 12 (41.4%), PD 4 (13.8%). The total response rate was 34.5%. The clinical beneficial response rate was 72.4% (21/29). Median time to progression (mTTP) was 5.8 months and median overall survival time was 9.3 months. The main side effects were: suppression of bone marrow in 26 patients (89.7%), alopecia in 25 (86.2%), myodynia and arthrodynia in 23 (79.3%). There was no death during the treatment.

CONCLUSIONTaxol-based chemotherapy is an effective and well tolerated regimen in the treatment of AUC, which can relieve suffering and improve quality of life of the patients. It can be used as the second-line therapy for relapsed advanced gastric cancer.

Adult ; Aged ; Alopecia ; chemically induced ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Cisplatin ; administration & dosage ; Drug Administration Schedule ; Epirubicin ; administration & dosage ; Female ; Fluorouracil ; administration & dosage ; Humans ; Lymph Nodes ; pathology ; Lymphatic Metastasis ; Male ; Middle Aged ; Neoplasm Staging ; Paclitaxel ; administration & dosage ; adverse effects ; Quality of Life ; Remission Induction ; Stomach Neoplasms ; drug therapy ; pathology

OBJECTIVETo identify the localization of hair follicles stem cell (HFSC) in different stages of hair and explore the differentiating capacity of HFSC into epidermis in vitro.

METHODSHFSC were detected by K19 immunostaining in normal human skin. Then, the isolated HFSC through enzyme digestion were seeded on dermal equivalent (DE) and cultured between the air-liquid interfaces for 14 days. Skin-equivalents was harvested and used for evaluation.

RESULTSHFSC mainly located in outer root sheet in hair follicle and human anagen hair follicles containing two distinct reservoirs for K19-positive cells located in the bulge and bulb of the follicle. These two reservoirs fused in line of outer root sheets during the catagen-telogen transition phase and individualized again in the newly forming anagen hair follicle. Based on DE, growing HFSC built a multilayered and confined epidermis.

CONCLUSIONHFSC located in outer root sheets can promote hair cycle and differentiate into epidermis in vitro.

Cell Differentiation ; physiology ; Cells, Cultured ; Epidermis ; cytology ; Hair Follicle ; cytology ; Humans ; Stem Cells ; cytology