A double targets of high throughput screening model for xanthine oxidase inhibitors and superoxide anion scavengers was established. In the reaction system of xanthine oxidase, WST-1 works as the probe for the ultra oxygen anion generation, and product uric acid works as xanthine oxidase activity indicator. By using SpectraMax M5 continuous spectrum enzyme sign reflectoscope reflector, the changes of these indicators' concentration were observed and the influence factors of this reaction system to establish the high throughput screening model were studied. And the model is confirmed by positive drugs. In the reaction system, the final volume of reaction system is 50 μL and the concentrations of xanthine oxidase is 4 mU x mL(-1), xanthine 250 μmol x L(-1) and WST-1 100 μmol x L(-1), separately. The Z'-factor of model for xanthine oxidase inhibitors is 0.537 4, S/N is 47.519 9; the Z'-factor of model for superoxide anion scavengers is 0.507 4, S/N is 5.388 9. This model for xanthine oxidase inhibitors and superoxide anion scavengers has more common characteristics of the good stability, the fewer reagent types and quantity, the good repeatability, and so on. And it can be widely applied in high-throughput screening research.

A double targets of high throughput screening model for xanthine oxidase inhibitors and superoxide anion scavengers was established. In the reaction system of xanthine oxidase, WST-1 works as the probe for the ultra oxygen anion generation, and product uric acid works as xanthine oxidase activity indicator. By using SpectraMax M5 continuous spectrum enzyme sign reflectoscope reflector, the changes of these indicators' concentration were observed and the influence factors of this reaction system to establish the high throughput screening model were studied. And the model is confirmed by positive drugs. In the reaction system, the final volume of reaction system is 50 μL and the concentrations of xanthine oxidase is 4 mU x mL(-1), xanthine 250 μmol x L(-1) and WST-1 100 μmol x L(-1), separately. The Z'-factor of model for xanthine oxidase inhibitors is 0.537 4, S/N is 47.519 9; the Z'-factor of model for superoxide anion scavengers is 0.507 4, S/N is 5.388 9. This model for xanthine oxidase inhibitors and superoxide anion scavengers has more common characteristics of the good stability, the fewer reagent types and quantity, the good repeatability, and so on. And it can be widely applied in high-throughput screening research.
Enzyme Inhibitors ; pharmacology ; Free Radical Scavengers ; pharmacology ; High-Throughput Screening Assays ; Superoxides ; Uric Acid ; Xanthine ; Xanthine Oxidase ; antagonists & inhibitors

To control the quality of Humulus scandens, the quality standard was established in this study. According to the method recorded in the Appendix of Chinese Pharmacopoeia (2010 Edition) , the water and ash inspections were carried out. The component luteoloside and cosmosiin in Humulus scandens were identified and assayed by TLC and HPLC. The results showed a strong characteristics microscopic of Humulus scandens, and trichoromethane-methanol-formic acid (10: 3: 0. 3) as the mobile phase of TLC, the spots at 365 nm with a UV lamp was clear. The 16 batches of samples were analyzed by HPLC with a gradient elution of acetonitrile and phosphate solution (0.2%) at a flow rate of 1.0 mL · min(-1) and detected at 350 nm. The content of luteoloside was 0.015%- 0.651% (average 0.148%); the content of cosmosiin was 0.003%-0.118% (average 0.036%). The linear calibration curve of luteoloside and cosmosiin was acquired in the ranges of 0.011-0.364 g · L(-1) (r = 1.000 0) and 0.003-0.096 g · L(-1) (r = 1.000 0), respectively. The average recovery was 100.5% and 98.5%, respectively. The methods are convenient and reliable, which can be ap- plied for quality assessment of Humulus scandens.
China ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; analysis ; standards ; Humulus ; anatomy & histology ; chemistry ; Quality Control

To promote bilingual teaching reform and explore a proper bilingual teaching mode in China,we studied the bilingual teaching of the course “Biochemistry on special subjects”.The present paper mainly designs and discusses the object of the course,teaching materials,contents and methods as well as the building of feedback and evaluation system of the course.

Objectiye To optimize the depth of the microchannel and the time point for sperm collection,and improve the efficiency of sperm screening on a microfluidic device. Methods Microchannels with four different depths of 25, 50, 100 and 200 μm were tested. Mice sperm were added to the inlet of the microchannel. The relative quantity and motility of sperm in the outlet were recorded at different collection times, i.e. ,5, 15, 30 and 60 min. Statistical method one-way ANOVA and appropriate post-hoc testing were applied to analyze differences between different groups, and further to select the best-fit depth of the microchannel and the time point for collection. Results In microchannels with depths of 25, 50, 100 and 200 μm, the sperm motilities measured in each outlet were (85.4 ± 2.3)%, (85.8 ± 5.8)%,( 87. 2 ± 2. 8 ) %, (76. 5 ± 2. 8 ) % respectively with statistical significance ( F = 5.8, P ＜ 0. 05 ). No obvious differences were found among 25-100 μm channels, however the motility dramatically decreased in the 200 μm group. The relative sperm quantities were (5.2 ±2.0)%, (7.2 ±2.5)%,(12.3 ±2.0)%,(7. 7 ± 1.1 ) % respectively with statistical significance ( F = 6. 9, P ＜ 0. 05), which increased with channel depth from 25 to 100 μm,while it decreased in the 200 μm channel Taking 2 indexes into account, 100 μmwas the most fit channel depth for sperm motility screening. The sperm motility in the outlet gradually decreased with time. At the time points of 5, 15, 30 and 60 min after adding sperm, the sperm motilities were (99. 6 ±0. 7)%, (87.2 ±2. 8)%, (79. 3 ±2. 2)% and (62. 6 ±8.0)% respectively with statistical significance ( F = 37. 3, P ＜ 0. 01 ). Yet the relative quantities of sperm in the outlet increased almost three times in this process. At the time points mentioned above, the relative quantities of sperm were (5.8±1.1)%, (10.6 ± 0.9)%, (12.1 ± 1.7)%, (17.9 ± 3.4)% respectively with statistical significance ( F = 17.8, P ＜ 0. 01 ). Thus 15-30 min was the ideal screening time. Conclusion An effective microdevice for sperm screening with optimized depth and collection time period is developed,which may contribute significantly for the screening of healthy sperm on microfluidic chips.

OBJECTIVETo observe the pathological changes of renal tissue in the rats with paraquat (PQ) poisoning as well as the serum creatinine (SCr) levels and expression levels of hypoxia-inducible factor-1α (HIF-1α) and transforming growth factor-β (TGF-bgr;) in renal tissue at different time points after PQ poisoning, and to investigate the association of HIF-1α with renal injury after PQ poisoning.

METHODSForty-eight healthy male Sprague-Dawley rats were randomly divided into control group (n = 6) and PQ group (n = 42). The control group was given a single dose of 1 ml saline by gavage; the PQ group was given a single dose of 1 ml PQ (50 mg/kg), which was prepared by diluting 20% raw liquid of PQ with saline, by gavage. The PQ group was further divided into 2, 6, 12, 24, 48, 72 and 120 h PQ subgroups (n = 6 for each subgroup) to be examined at 2, 6, 12, 24, 48, 72, or 120 h after gavage. Their arterial blood was collected for blood gas analysis as well as blood lactic acid (BLA) and SCr measurement; renal sections were subjected to HE staining; the protein expression of HIF-1α and TGF-β in renal tissue was measured by Western blot.

RESULTSThe BLA level and SCr level began to rise at 6h after poisoning. Compared with the control group, the 6, 12, 24, 48, 72 and 120 h PQ subgroups had significantly increased BLA and SCr levels (P < 0.05); the 72 and 120 h PQ subgroup showed hypoxemia (P < 0.05). The protein expression of HIF-1α in PQ group increased significantly at 6h and reached the peak level at 72 h, with a significant difference from that in the control group at 6, 12, 24, 48, 72, and 120 h (P < 0.05). The protein expression of TGF-β in PQ group began to rise at 24 h, reached the peak level at 72 h, and declined at 120 h, with a significant difference from that in the control group at 24, 48, and 72 h (P < 0.05). The protein expression of HIF-1α was positively correlated with SCr level (r = 0.9308, P = 0.0008), uncorrelated with arterial partial pressure of oxygen (r = -0.6996, P = 0.0534), and positively correlated with BLA level (r = 0.9483, P = 0.0003). The pathological changes of renal tissue mainly included the degeneration and necrosis of renal tubular epithelial cells, which worsened as the time went on and appeared less severe at 120 h.

CONCLUSIONThe HIF-1α expression in renal tissue increases significantly in the early stage of PQ poisoning, which is associated with increased BLA and SCr levels and causes upregulated expression of TGF-β that promotes renal fibrosis.

Animals ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Kidney ; metabolism ; pathology ; Male ; Paraquat ; toxicity ; Rats ; Rats, Sprague-Dawley ; Transforming Growth Factor beta ; metabolism

Ischemic stroke is a primary cause of death and long-term disability all over the world. This disease is resulted from ischemia and hypoxia in brain tissues because of insufficient blood supply and causes a series of physiochemical metabolism disorders and physiological dysfunction. Its high disability ratio has bright huge burdens to society, governments and families. However, there is not efficacious medicine to treat it. In this study, a right middle cerebral artery occlusion was established in rats to observe the multi-path and multi-aspect intervention effects of Tibetan patent medicine Ruyi Zhenbao pills in reducing injuries to Nissl bodies, cerebral edema and inflammatory reactions and preventing cellular apoptosis, in order to lay a foundation for defining its therapeutic mechanism in acute ischemic stroke.
Animals ; Brain Ischemia ; drug therapy ; Male ; Medicine, Chinese Traditional ; Medicine, Tibetan Traditional ; NF-kappa B ; physiology ; Patents as Topic ; Rats ; Rats, Sprague-Dawley ; Stroke ; drug therapy

Objective To detect the expressions of CD19 and CD20 of B-lymphocytes (CD19+B, CD20+B) and natural killer (NK) cells in the peripheral blood of epileptic children and explore their significances. Methods Four hundred and fifty-eight epileptic children, admitted to our hospital from January 2008 to December 2010, were chosen as patient group; another 52 healthy subjects were chosen as controls. The expressions of CD19+B and CD20+B and NK cells were detected by flow cytometry; their results were compared. Ninety-two epileptic children were treated with intravenous immunoglobulin (IVIG) and the effect of IVIG therapy was studied. Results The CD19+B level ([22.35±6.54]％) and CD20+B level ([21.50±8.41]％) in the epileptic children were obviously higher than those in the healthy controls ([16.86±4.02)％,[16.13±4.19]％, P＜0.05); and the level of NK cells ([9.1 1±4.90]％) in the epileptic children was significantly decreased as compared with that in the healthy controls ([14.72±4.15]％, P＜0.05). The CD19+B level ([18.26±5.03]％) and CD20+B level ([16.74±5.12]％) 6months after MG treatment were decreased significantly as compared with those before treatment ([22.74±6.25]％,[21.61±8.03]％, P＜0.05]; while the level of NK cells ([14.65±4.58]％) 6 months after IVIG treatment was increased significantly as compared with that before treatment ([9.07±4.76]％,P＜0.05). Among the 92 patients treated with IVIG, 70 enjoyed good results and 22 had non-effective resutls; The changesofCD19+Blevel ([7.99±5.34]％) and CD20+B ([8.21±5.21]％) before and after treatment in effectively treated patients by IVIG were significantly different as compared with those in ineffectively treated patients by MG ([3.78±2.76]％,[3.66±2.48]％, P＜0.05); however, the changes of level of NK cells before and after treatment showed no significant difference between effectively treated patients and ineffectively treated patients ([5.28±4.55]％,[4.53％±4.43]％, P＞0.05). Conclusion Dysfunctions of B-lymphocytes and NK cells exist in epileptic children, and MG treatment shows good effect on immune dysfunction of them. The levels of CD19-B and CD20-B can be used as monitoring indicators in the IVIG treatment of epileptic children.

OBJECTIVETo investigate the effects of three kinds of artificial dermal scaffolds on vascularization and scar formation of wounds in pigs with full-thickness burn.

METHODSEighteen Bama miniature pigs were divided into chitosan scaffold (CS) group, sulfonated carboxymethyl chitosan scaffold (SCCS) group, and acellular dermal matrix (ADM) scaffold group according to the random number table, with 6 pigs in each group. Every pig in all groups was inflicted with 4 or 8 full-thickness scald wounds on the back (totally 96 wounds). Forty-eight hours after injury, eschars of all wounds were excised. Twenty-four wounds in CS group were transplanted with double-layer artificial dermis of collagen-chitosan and silicone rubber, those in SCCS group with double-layer artificial dermis of collagen-sulfonated carboxymethyl chitosan and silicone rubber, and those in ADM scaffold group with ADM. The rest 24 wounds in the three groups were dressed with vaseline gauze as control group. After 2 weeks of treatment, all wounds of every group were covered with skin. In post treatment (scaffold transplantation or gauze covering) week (PTW) 1, 2, 3, and 4, gross condition of wound was observed, and specimens from central parts of wounds were harvested for observation and assessment of vessels or cells with positive expression of CD31, α smooth muscle actin (α-SMA), TGF-β(1) and TGF-β(3) with SP staining. Data were processed with one-way analysis of variance and LSD test.

RESULTS(1) Degree of vascularization in SCCS group was better than that in the other three groups. (2) The number of vessels with positive expression of CD31 in CS, SCCS, ADM scaffold, and control groups increased gradually from PTW 1 to PTW 3, and decreased in PTW 4. There were statistical differences among 4 groups from PTW 1 to PTW 4 (with F value respectively 24.005, 38.822, 25.274, 3.856, P < 0.05 or P < 0.01). The numbers of vessels that expressed CD31 in SCCS group from PTW 1 to PTW 3 were more than those in the other three groups (with P values all below 0.05). (3) The numbers of vessels that expressed α-SMA in CS, SCCS, and ADM scaffold groups from PTW 1 to PTW 3 showed the similar trend of change to those of vessels that expressed CD31, which increased gradually in control group from PTW 1 to PTW 4. There were obvious differences among 4 groups from PTW 1 to PTW 4 (with F value respectively 22.637, 28.087, 62.651, 18.055, P values all below 0.01). The number of vessels that expressed α-SMA in SCCS group from PTW 1 to PTW 4 was more than that in the other three groups (with P values all below 0.05). (4) From PTW 1 to PTW 4, the number of cells with expression of TGF-β(1) in CS group was respectively (127 ± 8), (167 ± 19), (170 ± 18), (144 ± 10) per 400 times visual field, that in SCCS group was respectively (171 ± 17), (207 ± 25), (130 ± 30), (69 ± 16) per 400 times visual field, that in ADM scaffold group was respectively (106 ± 8), (159 ± 17), (171 ± 11), (145 ± 11) per 400 times visual field, and that in control group was respectively (100 ± 20), (150 ± 18), (200 ± 14), (172 ± 20) per 400 times visual field. There were statistical differences among 4 groups from PTW 1 to PTW 4 (with F value respectively 29.675, 9.503, 13.107, 54.515, P values all below 0.01). Compared with those in SCCS group, the number of cells that expressed TGF-β(1) in the other three groups was decreased in PTW 1, 2 but increased in PTW 3, 4 (with P values all below 0.05). (5) The number of cells that expressed TGF-β(3) in 4 groups increased gradually from PTW 1 to PTW 3, and decreased or increased continually in PTW 4. There were statistical differences among 4 groups from PTW 1 to PTW 4 (with F value respectively 140.612, 945.850, 714.037, 119.147, P values all below 0.01). The number of cells with positive expression of TGF-β(3) in SCCS group from PTW 1 to PTW 4 was more than that in the other three groups (with P values all below 0.05).

CONCLUSIONSThe collagen-sulfonated carboxymethyl chitosan dermal scaffold can rapidly induce growth and maturation of blood vessels during wound healing after burn. It is beneficial for wound repair at early stage with inhibition of scar proliferation.

Acellular Dermis ; Animals ; Burns ; surgery ; Chitosan ; analogs & derivatives ; Cicatrix ; pathology ; Collagen ; Dermis ; transplantation ; Female ; Neovascularization, Physiologic ; Skin Transplantation ; Skin, Artificial ; Swine ; Tissue Scaffolds ; Wound Healing

To explore the possible linkage between telomerase and acute leukemia, we detected telomerase activity expressed in 3 leukemia cell lines, 22 acute leukemia bone marrow and 6 normal bone marrow with PCR ELISA assay. Results showed that telomerase activities of three leukemia cell lines were positive with the average (1.57 +/- 0.056) U, normal bone marrow samples average was (0.085 +/- 0.081) U, telomerase value from 22 acute leukemia patients was (0.512 +/- 0.294) U. Telomerase activity is higher expressed in acute leukemia than normal samples and decreased significantly after chemotherapy (P < 0.01). The results suggested that telomerase activity was related to some malignant diseases and might be used as a marker for tumor diagnosis and therapy.