Objective To explore the change of degrade of clonazepam in serum with single and repeated - dose administration in children with seizure, and find a reasonable method for using the clonazepam. Methods Children with seizures were divided into single - dose paradigam, repeated - dose paradigam, and decreased - dose paradigam. The concentration of CZP in serum was determined by high performance liquid chromatography( HPLC). Results The serum concentrations of clanazepain in single - dose paradigam were (101.9?12.1),(76.9 ? 5.8),(50.7?2.9),(30.9?5.4),(21.5?6.8)?g/L,the time point that the blood samples collected were 15,30,60,120 and 480 min. The serum concentrations in repeated - dose paradigam were (97. 2 ? 6. 1),(130.4? 13. 4), (99. 4 ? 9.8),(79.6?2.4)?g/L,in decreased-dose paradigam were( 101.1 ?13.1),(123.1 ?6. 6), (99.4 ?9. 8), (79. 3 ? 2. 2)?g/L,in these two groups,the time point were 15,45,60 and 120 min. Conclusion Repeated administration of CZP with decreased dose may increase its effectiveness in treatment without substantially increasing toxicity.

In order to improve the thermostability of the Penicillium expansum Lipase (PEL), the lipase encoding genes was mutated by site-directed mutagenesis. A recombinant vector pAO815-ep8-K55R which contain double mutant genes was constructed by overlap extension PCR using the cDNA of a random-mutant lipase ep8 (a single site mutant) as the template and two special primers were used to generate another mutation site K55R. The recombinant vector was transformed into Pichia pastoris GS115 by electroporation and the recombinant mutant GS-pAO815-ep8- K55R can secret double-mutant lipase PEL-ep8-K55R-GS into the medium when it was induced by Methanol. The yield of the double-mutant lipase is 508 u/mL, which is 81% that of the wild type lipase PEL-GS (627 u/mL) and 55% that of random-mutant PEL-ep8-GS (924 u/mL). The specific activity of double-mutant lipase is 2309.1 u/mg, which is similar to random-mutant lipase PEL-ep8-GS and the wild type lipase PEL-GS. The optimum temperature of the double-mutant lipase is same with the wild type lipase PEL-GS and random-mutant lipase PEL-ep8-GS. While the Tm of the double-mutant lipase is 41.0 degrees C, 2.3 degrees C higher than the wild type lipase PEL-GS and 0.8% higher than the random-mutant lipase PEL-ep8-GS, indicating that the double-mutant lipase PEL-ep8-K55R-GS has higher thermostability.
Electroporation ; Enzyme Stability ; Hot Temperature ; Lipase ; genetics ; metabolism ; Mutagenesis, Site-Directed ; Mutant Proteins ; metabolism ; Penicillium ; enzymology ; Pichia ; genetics ; metabolism ; Protein Engineering ; methods ; Recombinant Proteins ; biosynthesis ; genetics

OBJECTIVETo investigate the role of chemokine receptor CXCR3 in patients with atopic dermatitis.

METHODSThe expression of CXCR3 mRNA was measured by fluorescent quantitative polymerase chain reaction, and the relationship between CXCR3 mRNA expression and the disease severity (graded according to SCORAD index system) was assessed by correlation analysis.

RESULTSCXCR3 mRNA expression was significantly higher in patients with atopic dermatitis than in healthy control subjects (P7lt;0.01), and showed obvious positive correlation with SCORAD index system.

CONCLUSIONThese data suggest an important role of CXCR3 in the development and progression of atopic dermatitis.

Adolescent ; Case-Control Studies ; Child ; Dermatitis, Atopic ; genetics ; Female ; Gene Expression Regulation ; Humans ; Male ; RNA, Messenger ; genetics ; metabolism ; Receptors, CXCR3 ; genetics

Objective To harvest pancreatic tissues from rats , prepare decellularized bio-derived pancreatic scaffolds ( DBPS) , and to examine the integrity and biocompatibility of the scaffolds .Methods Normal pancreases were harvested from healthy adult SD rats .DBPS was prepared by perfusing SDS and Triton X-100 through bile duct and the portal vein, respectively.After decellularization, normal pancreatic tissue and DBPS were compared via HE staining , and transmission electron microscopy ( TEM ) . Abdominal wall and subcutaneous implantations were used to compare biocompatibility , and the remain quantity of residual protein and growth factors were determined via enzyme linked immunosorbent assay(ELISA).MTT assay was used to test the scaffolds’ cytotoxicity.The scaffolds were co-cultured with endotheliocyte .Results HE staining and TEM study indicated no residual cells in the DBPS as well as preservation of the complete extracellular matrix .The remain quantity of residual protein and growth factors in ECM was high .The abdominal wall and subcutaneous implantation revealed that DBPS triggered a lower immune response as compared to the control group.MTT assay showed little cytotoxicity .Endotheliocyte assembled and growed with the scaffolds together .Conclusion DBPS are completely decellularized , and exhibit a higher level of biocompatibility in vivo.Using the way of vessels can make the integrity of extracellular matrix to be fully preserves and contain more growth factors .So using vessels way is better than bile duct .

A series of cycloberberine derivatives were designed, synthesized and evaluated for their anti-cancer activities in vitro. Among these analogs, compounds 6c, 6e and 6g showed strong inhibition on human HepG2 cells. They afforded a potent effect against DOX-resistant MCF-7 breast cells as well. The primary mechanism showed that cell cycle was blocked at G2/M phase of HepG2 cells treated with 6g using flow cytometry assay. It significantly inhibited the activity of DNA Top I at the concentration of 0.1 mg mL-1. Our results provided a basis for the development of this kind of compounds as novel anti-cancer agents.

Background To optimize the culture method of human retinal microvascular endothelial cells is very important for the study of retinal angiogenesis disease.Human retinal microvascular endothelial cells have been successfully cultured in previous studies,but further improvement of the culture method to harvest higher yields and purity cells is still needed.Objective This study was to design a modified method to isolate and purify human retinal microvascular endothelial cells much easily and quickly,and to compare the expression of specific markers of vascular endothelial cells,factor Ⅷ and CD31/CD34 in the cells.Methods The use of human donor eyeballs was approved by the Ethic Commission of Zhongshan Ophthalmic Center of Sun Yat-sen University.The retina tissue from healthy donor was isolated and digested by the two-step digestion method with 2％ trypsin and 0.133％ collagenase Ⅳ.Human retinal microvascular endothelial cells were collected and plated in 60 mm dishes coated by 0.1％ fibronectin and cultured in endothelial cell-specialized medium supplemented with 10％ fetal bovine serum,0.3 mg/L β-endothelial cell growth factor (ECGF) and 100 ng/L sodium heparin.During the culturing,the growth situation of the cells was monitored by morphological observation,and immunohistochemical staining was performed to probe vascular endothelial cell-specific membrane protein CD31,CD34 and factor Ⅷ for identification of the cell purity.Results Human retinal microvascular endothelial cells were isolated successfully from the retina by the twostep digestion method.The primary cultured cells adhered to well 72 hours later and achieved confluence with the typical cobblestone appearance 9 to 10 days after cultured.The cells exhibited the blue nuclei and reddish cytoplasm by regular haematoxylin and eosin stain and showed a strong positive response for CD31,CD34 and factor Ⅷ by immunohistochemistry.The positive dye of CD31 and CD34 was lower than Ⅷ factor in both endothelial cells.Conclusions Modified culture method of human retinal microvascular endothelial cells can improve cell culture result and purify target cells.

ObjectiveTo explore the effect of musicotherapy on agitation behaviors in patients with aged dementia (AD).Methods22 AD patients were treated with different musicotherapies, and examined with Mini-Mental State Examination (MMSE), Cohen-Mansfield Agitation Inventory (CMAI) and self made sociality psychological inventory before and after treatment.ResultsAfter musicotherapy, the incidence rate of agitation behaviors of AD patients decreased significantly ( P<0.001). The incidence rates of agitation behaviors among patients with low, middle and high MMSE scores were significantly different ( P<0.001). After musicotherapy, both depressions of patients' cognitive function and scores of agitation behaviors were significant different compared with that before treatment (P<0.01). Effectual rates of musicotherapy on sociality psychological emotion, communication, language and intercourse of this group patients were 86.36%, 90.91%, 72.73% and 77.27%, respectively.ConclusionMusicotherapy can effectively alleviate the symptoms of agitation behaviors in AD patients.

Background: Previous studies showed that combining apparent diffusion coefficient (ADC) value with the Spondyloarthritis Research Consortium of Canada (SPARCC) index value might provide a reliable evaluation of the activity of ankylosing spondylitis (AS), and that contrast?enhanced (CE) magnetic resonance imaging (MRI) is unnecessary. However, the results were based on confirming only a small random sample.This study aimed to assess the role of CE?MRI in differentiating the disease activity ofAS by comparingADC value with a large sample. Methods: A total of 115 patients with AS were enrolled in accordance with Bath AS Disease Activity Index and laboratory indices, and 115 patients were divided into two groups, including active group (n = 69) and inactive group (n = 46). SPARCC, ΔSI, and ADC values were obtained from the short tau inversion recovery (STIR), diffusion?weighted imaging (DWI), and CE?MRI, respectively. One?way analysis of variance and receiver operating characteristic analysis were performed for all parameters. Results: The optimal cutoff values (with sensitivity, specificity, respective area under the curve, positive likelihood ratio, and negative likelihood ratio) for the differentiation between active and inactive groups are as follows: SPARCC = 6 (72.06%, 82.61%, 0.836, 4.14, 0.34); ΔSI (%) = 153 (80.6%, 84.78%, 0.819, 5.3, 0.23); ADC value = 1.15 × 10?3 mm2/s (72.73%, 81.82%, 0.786, 4, 0.33). No statistical differences were found among the predictive values of SPARCC, ΔSI, and ADC. Multivariate analysis showed no significant difference between the combination of SPARCC and ADC values with and without ΔSI. Conclusions: Using large sample, we concluded that the combination of STIR and DWI would play significant roles in assessing the disease activity, and CE?MRI sequence is not routinely used in imaging of AS to avoid renal fibrosis and aggravation of kidney disease.

A series of cycloberberine derivatives were designed, synthesized and evaluated for their anti-cancer activities in vitro. Among these analogs, compounds 6c, 6e and 6g showed strong inhibition on human HepG2 cells. They afforded a potent effect against DOX-resistant MCF-7 breast cells as well. The primary mechanism showed that cell cycle was blocked at G2/M phase of HepG2 cells treated with 6g using flow cytometry assay. It significantly inhibited the activity of DNA Top I at the concentration of 0.1 mg mL-1. Our results provided a basis for the development of this kind of compounds as novel anti-cancer agents.
Antineoplastic Agents ; chemical synthesis ; chemistry ; pharmacology ; Berberine ; analogs & derivatives ; chemical synthesis ; chemistry ; pharmacology ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; DNA Topoisomerases, Type I ; metabolism ; Doxorubicin ; pharmacology ; Drug Resistance, Neoplasm ; Hep G2 Cells ; Humans ; MCF-7 Cells ; Molecular Structure ; Structure-Activity Relationship

The total RNA was extracted from ginseng leaves of Panax ginseng. The Cu/Zn-SOD gene was amplified via RT-PCR and the pET-28(a)-Cu/Zn-SOD expression vector was constructed. The pET-28 (a)-Cu/Zn-SOD recombinant plasmid was transformed into Escherichia coli BL21 (DE3) competent cells and was induced by IPTG in order to select optimal induction of expression conditions. The target protein was purified by the nickel ions (Ni ) affinity chromatography and the target protein enzyme activity was determinated by the xanthine oxidase method. The similarity of the Cu/Zn-SOD gene sequences and the Cu/Zn-SOD gene sequences of Korean ginseng in NCBI was 99. 00%. The target protein expression level was about 44.42%, and the molecular weight was 16.30 kDa after the pET-28(a)-Cu/Zn-SOD recombinants were induced by IPTG. The purified Cu/Zn-SOD protease activity reached 10,596.69 U x mg(-1). The P. ginseng pET-28(a)-Cu/Zn-SOD prokaryotic expression vector was built by the method of molecular biology, which provided the foundation for studying the Cu/Zn-SOD biology function.
Cloning, Molecular ; Escherichia coli ; genetics ; Gene Expression ; Genetic Engineering ; methods ; Genetic Vectors ; genetics ; Panax ; enzymology ; genetics ; Sequence Analysis ; Superoxide Dismutase ; genetics ; isolation & purification ; metabolism