Objective To construct a prokaryotic expression vector in BL21 to secretorily expressα-Cyclodextrin Glycosyltransferase(α-CGTase). Methods α-CGT gene was amplified from Bacillus macerens genome by PCR.pET26b and α-CGT gene were connected after digested with Nco I, Xho I respectivly, and then transformed into Escherichia coli BL21 strain.α-CGTase was expressed in fermentation culture medium and AA-2G was prepared by using α-CGTase, VC and starch.Results α-CGTase was expressed secretorily and the enzyme activity was up to 120 U/mL.AA-2G was prepared by the biotransformation of VC and starch using α-CGTase which proved to be correct by HPLC.Conclusion AA-2G was prepared by using self-madeα-CGTase, after optimized the preparation conditions the yield of AA-2G was 17.46 g/L, and the conversion rate reached 58.2%(mg/mg).

Objective To investigate the inhibition by Guangxi Selenocosmia huwena toxins on human glioma cell strain U251 and to interpret its possible mechanisms, with regard to initial clarify its mechanism. Methods Culture medium of Containing 10% fetal bovine serum wet RPMI1640 of U251 cells incubated with culture, MTT was involved indetecting the cytotoxic effects. HE staining and inverted phase contrast microscope was used to detected the change of cell shape and cellular nucleus shape. TUNEL was used to detected apoptosis ofhuman glioma cell strain U251. Results The proliferation of U251 was obviously inhibited by Guangxi Selenocosmia huwena toxins in dose- and time- dependent manner. The inhibitive effect of Guangxi Selenocosmia huwena toxins on U251 was obvious, with the Two half-inhibitory concentration of 82.60 μg/mL and 71.12 μg/mL at 48 h and 72 h respectively, This depressanteffct offer conspicuous dose-effect relationship. We could observesome cell nucleus became to pycnosis and nuclear fragmentation (cell apoptosis). We also could dye apoptosis cell nucleus in brown. Conclusion Guangxi Selenocosmia huwena toxins can inhibit theproliferation of glioma U251 which was implemented through the inducement ofglioma apoptosis.

To study in vitro anti-influenza viral activities of Chinese traditional patent medicines for influenza prevention and treatment, neuraminidase (NA) activity assay was used to examine NA inhibitory activity of 33 Chinese traditional patent medicines through fluorimetric assay, and influenza virus induced cytopathic effect (CPE) inhibition assay was used to verify their anti-influenza viral activities in vitro. The assay results showed that most liquid preparations displayed relatively high NA inhibitory activities, such as Shuanghuanglian oral liquid, Qingkailing oral liquid, Qingre Jiedu oral liquid, and Reduning injection. Among liquid preparations, Shuanghuanglian oral liquid not only displayed the highest NA inhibitory effect, but also exhibited obvious in vitro anti-viral activity in CPE experiment. Among solid preparations, Shuanghuanglian powder for injection showed the highest activity on NA inhibition, and Fufang Yuxingcao tablet showed relatively strong anti-influenza viral activity in CPE cells. From the results, it can be concluded that most Chinese traditional patent medicines possessed NA inhibitory activity, but only a few of them displayed significant in vitro anti-influenza viral activities. These results will provide important information for the isolation of active constituents, and for the clinical uses of Chinese traditional patent medicines for influenza treatment and prevention.
Animals ; Antiviral Agents ; pharmacology ; Cell Line ; Cytopathogenic Effect, Viral ; drug effects ; Dogs ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; administration & dosage ; isolation & purification ; pharmacology ; Influenza A Virus, H1N1 Subtype ; drug effects ; enzymology ; Influenza A Virus, H3N2 Subtype ; drug effects ; enzymology ; Influenza B virus ; drug effects ; enzymology ; Medicine, Chinese Traditional ; Neuraminidase ; antagonists & inhibitors ; metabolism ; Plants, Medicinal ; chemistry

OBJECTIVETo compare the antioxidant active components from two species of chamomile-matricaria and Roman chamomile produced in Xinjiang.

METHODThe TLC-bioautography was used, with 1,1-Diphenyl-2-picrylhydrazyl (DPPH) radical as the experimental model. The peak areas of various antioxidant components were obtained by TLC-scanning for analyzing antioxidant active components contained in volatile oil extracts and flavone extracts from the two species of chamomiles. The total peak area was taken as the indicator for comparing the antioxidant capacities of the two types of extracts, and comparing them with the total antioxidant activity of flavone extracts of the two species of chamomiles.

RESULTSAccording to the result of TLC-bioautography in volatile oil extracts from the two species of chamomiles, volatile oil extracts from chamomile showed four white antioxidant spots, including en-yne-dicycloether, and volatile oil extracts from Roman chamomile showed only one white antioxidant spot. The TLC-scanning result showed that the peak area of antioxidant spots of volatile oil extracts from chamomile was significantly larger than that of volatile oil extracts from Roman chamomile. According to the test on the antioxidant activity of the two species of chamomiles with ultraviolet-visible spectrophotometry, the concentration of chamomile after scavenging 50% of DPPH radicals was 0.66 g x L(-1), whereas the figure for Roman chamomile was 0.33 g x L(-1). According to the result of TLC-bioautography in flavone extracts from the two species of chamomiles, flavone extracts from chamomile showed seven yellowish antioxidant spots, including apigenin and apigenin-7-glucoside, and flavone extracts of Roman chamomile showed eight yellowish antioxidant spots, including apigenin and apigenin-7-glucoside. The TLC-scanning results showed that the peak area of antioxidant spots of flavone extracts from Roman chamomile was significantly larger than that of flavone extracts from chamomile.

CONCLUSIONVolatile oil extracts from the two species of chamomiles have significant difference in the antioxidant activity in TLC-bioautography. Specifically, the antioxidant activity of volatile oil extracts from chamomile is stronger than volatile oil extracts from Roman chamomile; the known antioxidant active components in volatile oil extracts from chamomile is en-yne-dicycloether, while all of the other three antioxidant active components as well as antioxidant active components in volatile oil extracts from Roman chamomile are unknown components and remain to be further determined. Considering the significant difference in the number of antioxidant active spots in volatile oil extracts from the two species of chamomiles, the result can be applied to distinguish the two species of chamomiles. The antioxidant activity determination result for flavone extracts from two species of chamomiles was consistent with the result of TLC-bioautography, showing that flavone extracts from chamomile and Roman chamomile are more antioxidant active, while that of Roman chamomile is stronger than chamomile. Flavone extracts from both of the two species of chamomiles contain apigenin and pigenin-7-glucoside, which are known, while all of the other five antioxidant active components contained in flavone extracts from chamomile and the other six antioxidant active components contained in flavone extracts from Roman chamomile are unknown and remain to be further identified. The method lays a foundation for further identification of antioxidant active components contained in chamomile.

Antioxidants ; chemistry ; isolation & purification ; Apigenin ; chemistry ; isolation & purification ; Biphenyl Compounds ; metabolism ; Chamaemelum ; chemistry ; Chromatography, Thin Layer ; methods ; Flavones ; chemistry ; isolation & purification ; Free Radical Scavengers ; chemistry ; isolation & purification ; Glucosides ; chemistry ; isolation & purification ; Matricaria ; chemistry ; Oils, Volatile ; chemistry ; isolation & purification ; Picrates ; metabolism ; Plant Extracts ; chemistry ; isolation & purification ; Plant Oils ; chemistry ; isolation & purification

Objective To study the prevalence and grade of cerebral microbleeds (CMB) among patients with different subtypes of acute ischemic cerebrovascular diseases, and investigate the clinical significance of CMB.Methods Consecutive 259 patients with acute ischemic cerebrovascular diseases, admitted to our hospitals from September 2009 to July 2010, were included; according to the stroke subtypes, these patients were classified into groups of atherothrombotic infarction (n=146),cardioembolic infarction (n=28), small artery infarction (n=50), infarction of unknown origin (n=19) and transient ischemia attack (TIA, n=16). The patients without cerebral vascular diseases were served as controls (n=96). The baseline data were registered and all patients were performed gradient echo-T2*weighted imaging (GRE-T2*WI); the prevalence and grade of CMB between each 2 different subtypes of acute ischemic cerebrovascular diseases were compared; the prevalence of CMB in patients with acute ischemic infarction for the first time and patients with recurrent cerebral infarction was compared.Results The prevalence and grade of CMB between each 2 different infarction subtypes varied with a statistical difference (P＜0.05). Apart from that of TIA group, the prevalence of all infarction groups was statistically higher than that of the controls (P＜0.05) with small artery infarction group being the highest (68.0%). The prevalence of CMB in patients with recurrent infarction was statistically higher than that in patients with primary infarction (P＜0.05).Conclusion The prevalence of CMB among different subtypes of infarction is high with the subtype of small artery infarction enjoying the highest rate; the prevalence of CMB in recurrent infarction goes higher as compared with that in primary infarction; the relapse of the cerebral infarction is possiblely related to the presence of CBMs.

Objective To understand the treatment circumstance of ST-elevation myocardial Infarction (STEMI) patients at public hospitals in Shenzhen.Methods Directed by Public Hospital Administration at Shenzhen Municipality (PHASM) and led by Chest Pain Treatment Quality Control Center at Shenzhen People's Hospital (CPTQCC-SZ),25 public hospitals in Shenzhen, including 15 PCI-capable hospitals and 10 non-PCI-capable hospitals,we investigated on the overall treatment conditions and the STEMI patient treatment situations from October to December 2015 in these hospitals. A regression analysis was performed between a few factors and the success rate of STEMI treatment was reviewed. Results 383 STEMI cases twere registered between October to December 2015 in the 25 public hospitals in Shenzhen,with 324 case treated in PCI-capable hospitals and 59 cases in non-PCI-capable hospitals. There were statistical differences between the PCI-capable hospitals and non-PCI-capable hospital in fields of total number of senior cardiologists (work year ≥ 3 year),total number of beds in general cardiology beds and number of beds in cccu(all P<0.01). There was no difference in the time of obtaining the first ECG at patient arrival between hospitals(P=0.052).Time for laboratory results availability for troporin was significantly shorter in PCI-capable hospital[(25.0±4.2)min vs.(58.0±2.8)min,P=0.002] .Among the PCI-capable hospitals,the mean D-to-B time was 320 minutes, and mean F-to-B time was 380 minutes. In non-PCI-capable hospitals,D-to-N time ranged from 20 to 350 minutes and F-to-N time ranged from 25 to 380 minutes. Conclusions There are gaps among the overall conditions of the public hospitals in Shenzhen. The overall conditions and chest pain treatment conditions of non-PCI-capable hospitals had bigger gaps with PCI-capable hospitals.

BACKGROUNDMicroRNAs (miRNAs) are highly conserved small non-coding RNAs of 18 - 25 nucleotides (nt) that mediate post-transcriptional gene regulation. Hepatitis B virus (HBV) can cause either acute or chronic hepatitis B, and is a high risk factor for liver cirrhosis and hepatocellular carcinoma. Some mammalian viruses have been shown to modulate the expression of host cellular miRNAs. However, interactions between the HBV and the host cellular miRNAs are largely unknown.

METHODSmiRNA microarray and Northern blotting analysis were used to compare the expression profile of cellular miRNAs of a stable HBV-expressing cell line HepG2.2.15 and its parent cell line HepG2. mRNA microarray assay and the miRanda program were used to predict the miRNA targets. A flow cytometric assay was further used to investigate the expression of human leukocyte antigen (HLA)-A.

RESULTSEighteen miRNAs were differentially expressed between the two cell lines. Among them, eleven were up-regulated and seven were down-regulated in HepG2.2.15 cells. Northern blotting analysis confirmed that the expression of miR-181a, miR-181b, miR-200b and miR-146a were up-regulated and the expression of miR-15a was down-regulated, which was in consistent with the results of the microarray analysis. Furthermore, some putative miRNA targets were predicted and verified to be linked with mRNA expression. The 3'-UTR of HLA-A gene had one partially complementary site for miR-181a and miR-181a might down-regulate the expression of HLA-A.

CONCLUSIONHBV replication modulates the expression of host cellular miRNAs, which may play a role in the pathogenesis of HBV-related liver diseases.

Blotting, Northern ; Cell Line, Tumor ; metabolism ; virology ; Flow Cytometry ; Gene Expression Profiling ; Gene Expression Regulation ; HLA-A Antigens ; metabolism ; Hepatitis B virus ; growth & development ; physiology ; Humans ; MicroRNAs ; genetics ; Oligonucleotide Array Sequence Analysis

OBJECTIVE: To evaluate the value of the nano-hydroxyapatite/ collagen (nHAC) as a scaffold for cartilage tissue engineering. METHODS: The normal cartilage from patients after total hip arthroplasty of osteoarthritis was selected, and then chondrocytes were digested, separated, and amplificate in vitro. The chondrocytes were seeded onto the nHAC scaffold and were cultured in a 3-dimensional environment for 5 and 10 days. The effects of the composite scaffold on the cell adhesion, proliferation, morphological changes, and synthesis of the extracellular matrix were observed by scanning electronic microscopy and immunohistochemistry. RESULTS: The chondrocytes could adhere to the surface of the scaffolds, proliferate, and migrate into the scaffolds. They maintained round and could secrete extracellular matrix on the porous scaffold. CONCLUSION The nHAC can be used as the cartilage cell carrier.
Cells, Cultured ; Chondrocytes ; cytology ; Collagen ; Durapatite ; Humans ; Prostheses and Implants ; Tissue Engineering ; methods ; Tissue Scaffolds

Objective To evaluate and study percutaneous nephrolithotomy (PCNL) related factors in the treatment of renal calculi caused postoperative fever and its prevention measures. Methods Making a retrospective analysis of the clinical records of 150 patients who underwent PCNL, including age, gender, diabetes history and previous ipsilateral renal surgery, stone type, stone size, whether the complication of upper ureteral stones, preoperative urinary tract infection, hydronephrosis and pyonephrosis, preoperative renal fistula, postoperative centering vein pressure, intraoperative perfusion and the operation time from January 2015 to June 2017. After the operation, the patients were divided into two groups: fever group and non-fever group, and analyzed the related factors of the fever. Results Among the 150 cases, fever occurred in 27 cases after PCNL, taking up 18%. Gender, history of diabetes, staghorn calculi or staghorn stone, stone size, with ureteral calculi, preoperative urine leukocyte count, renal abscess, preoperative renal fistula, postoperative central venous pressure, intraoperative perfusion and operation time between the two groups, the differences that were statistically significant (P < 0.05). Logistic that multivariate analysis showed that female patients with upper ureteral calculi, perfusion, intraoperative volume,preoperative pyonephrosis, long operation time are independent risk factors of fever after operation (OR^>1, P<0.05). Conclusion Routine bacterial culture of urine should be performed before percutaneous nephrolithotomy, urinary tract infection and kidney empyema should be treated thoroughly. The reasonable shorter operative duration and perfusion fluid volume could significantly reduce the incidence of fever after PCNL.

OBJECTIVE: To explore the effect of insulin-like growth factor (IGF-1) on the concentration of NO and PGE(2) in the supernatant of rabbit articular chondrocytes induced by IL-1, and to explore the mechanism of IGF-1 in the development of osteoarthritis (OA). METHODS: The samples were divided into 7 groups: IL-1beta 10 microg/L group, IL-1beta 10 microg/L+IGF-1 1 microg/L group, IL-1beta 10 microg/L+IGF-1 10 microg/L group, IL-1beta 10 microg/L+IGF-1 50 microg/L group, IL-1beta 10 microg/L+IGF-1 100 microg/L group, IGF-1 50 microg/L group, and a blank control group. The chondrocytes from the articular cartilage of 2 month old rabbits were cultivated and identified, and then co-cultured in the second filial generation chondrocytes on plates with or without recombinant human IGF-1 or IL-1. The concentration of NO was detected by nitrate reductase kit, and that of PGE(2) by enzyme-linked immunosorbent assay (ELISA). The results were analyzed by statistical method. RESULTS: The average value of NO and PGE(2) was (89.971+/-10.224) micromol/L and (22.028+/-8.731) micromol/L in the IL-1beta 10 microg/L group, and (12.404+/-8.809) micromol/L and (1.900+/-0.227) ng/L in the blank control group. The concentration of NO and PGE(2) in IL-1beta 10 microg/L group was significantly higher than that in the blank control group (P<0.05). At the same concentration of 10 microg/L, IGF-1 could dose-dependently decrease the increase of NO and PGE(2) concentration induced by IL-1beta in the chondrocytes supernatant in vitro, and the optimum concentration of IGF-1 was 50 microg/L. CONCLUSION IL-1 can significantly increase the concentration of NO and PGE(2), and IGF-1 can dose-dependently decrease the concentration of NO and PGE(2) in the chondrocytes supernatant in vitro. The optimum concentration of IGF-1 was 50 microg/L.
Animals ; Cartilage, Articular ; cytology ; metabolism ; Cells, Cultured ; Chondrocytes ; drug effects ; metabolism ; Dinoprostone ; metabolism ; Insulin-Like Growth Factor I ; pharmacology ; Interleukin-1 ; pharmacology ; Nitric Oxide ; metabolism ; Osteoarthritis ; metabolism ; Rabbits