Objective To construct directional expressing cDNA library from normal testes in Chinese males for further studying the structure and functions of spermatogenesis related genes. Methods mRNA of normal testes from a Chinese male was isolated and the cDNA synthesized by reverse transcription.After being digested by SalI and NotI ,dscDNA was subcloned into expressing plasmid pSPORT1 directionally.Finally,the directional expressing cDNA library from normal testes in Chinese males was constructed. Results The transformation rate of the library was 7?10 5.Ten clones were chosen and analysed. Digested by SalI and NotI ,the fragments of 0.5～2.0 kb were obtained with cDNA inserted. Conclusions The present study indicates that the directional expressing cDNA library from normal testes in Chinese males is qualified to screen genes and is considered to be helpful for studying the structure and functions of spermatogenesis related genes.

Multilevel models are applicable to both the quantitative data and categorical variables. We used the methods, including the multilevel models, analysis of covariance and CMH chi-square test, to analyse different types of data, to explore the application of multilevel models in the analysis of the multicenter clinical trial center effect. The results showed that the analysis of covariance is more sensitive to find the center effect for quantitative data, while multilevel models are more sensitive to categorical variables. It can be seen that results with different analytical methods for center effect are not the same, and the most appropriate method should be selected in accordance with the characteristics of data, the objective of research, and the applicable conditions of the various methods in practical use.
Clinical Trials as Topic ; Humans ; Models, Theoretical ; Research Design

OBJECTIVETo observe the effect of Draconis Sanguis-containing serum on the expressions of NGF, BDNF, CNTF, LNG-FR, TrkA, GDNF, GAP-43 and NF-H in Schwann cells, and investigate the possible mechanism of Draconis Sanguis to promote peripheral nerve regeneration.

METHODSD rats were randomly divided into 2 groups: the Draconis Sanguis group (orally administered with Draconis Sanguis-containing balm solution) and the blank group (equivoluminal balm) to prepare Draconis Sanguis-containing serum and blank control serum. Schwann cells were extracted from double sciatic nerves of three-day-old SD rats, divided into 2 groups: the Draconis Sanguis group and the blank control group, and respectively cultured with 10% Draconis Sanguis-containing serum or blank control serum. The mRNA expressions of NGF, BDNF, CNTF and other genes in Schwann cells were measured by RT-PCR analysis 48 hours later.

RESULTMost of the Schwann cells were bipolar spindle and arranged shoulder to shoulder or end to end under the microscope and identified to be positive with the immunocytochemical method. To compare with the blank group, mRNA expressions of NGF, LNGFR, GDNF and GAP-43 significantly increased (P < 0.01). Whereas that of BDNF decreased significantly (P < 0.05), and so did that of TrkA, CNTF (P < 0.01), with no remarkable difference in NF-H-mRNA.

CONCLUSIONTraditional Chinese medicine Draconis Sanguis may show effect in nerve regeneration by up-regulating mRNA expressions of NGF, LNGFR, GDNF and GAP-43 and down-regulating mRNA expressions of TrkA, BDNF and CNTF.

Animals ; Arecaceae ; chemistry ; Brain-Derived Neurotrophic Factor ; genetics ; metabolism ; Cells, Cultured ; Ciliary Neurotrophic Factor ; genetics ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; GAP-43 Protein ; genetics ; metabolism ; Gene Expression ; drug effects ; Glial Cell Line-Derived Neurotrophic Factor ; genetics ; metabolism ; Male ; Nerve Growth Factor ; genetics ; metabolism ; Nerve Regeneration ; drug effects ; Neurofilament Proteins ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptor, trkA ; genetics ; metabolism ; Schwann Cells ; drug effects ; physiology ; Serum ; chemistry

Object To determine the content of glycyrrihizic acid obtained when each individual ingredient in MAHUANG DECOCTION was decocted separately and then mixing the extracts with boiling water in comparison with that obtained by decocting the total composition together as a whole in the traditional way. Methods The contents of glycyrrhizic acid was determined by HPLC. Hypersil C 18 column was used, with acetonitrile∶0.1% acetic acid (33∶67) as the mobile phase and detected at the wavelength of 254 nm. Results The average recovery of glycyrrhizic acid when separately decocted was 102.43%, RSD=2.65%, while that of decoction in whole was 99.41%, RSD=3.11%. Conclusion The method was simple and accurate and was not interfered by other constituents in the prescription.

AIM: To determine the contents of ephedrine、laetrile、glycyrrhizic acid and liquiritin in Huagai Powder traditional decoction and granule decoction(Herba Ephedrae,Semen Armeniacae Amarum,Radix et Rhizoma Glycyrrhizae,etc.) by HPLC. METHODS: An Agilent SB-C_(18) column was used for ephedrine,the mobile phase was acetonitrile-0.1% phosphoric acid(4∶96),detection wavelength at 207 nm;An Agilent XDB-C_(18) column was used for laetrile,the mobile phase was methanol-water(23∶77),detection wavelength at 215 nm;An Agilent XDB-C_(18) column was used for glycyrrhizic acid,the mobile phase was methanol-0.2 mol/L ammonium-acid(67∶33∶1),detection wavelength at 250 nm; An Agilent XDB-C_(18) column was used for liquiritin,the mobile phase was acetonitrile-0.5% acetic acid(20∶80),detection wavelength at 276 nm. RESULTS: There were discrepancies between traditional decoction and granules in the contents of four ingredients,the average contents in granules were more than those in the traditional decoction. CONCLUSION: The peaks of the four ingredients are(segregated)(effectively),and the method is simple,conveient,exact and has good reproducibility,the other ingredients do not have interferences for the determination.

OBJECTIVE: To study the mechanism of herbal application along meridians for treatment and prevention of asthma by using serum pharmacological test to observe the effects of serum containing herbs against the constriction of tracheal spiral strips induced by acetylcholine chloride (Ach). METHODS: Guinea pigs were randomly divided into normal control group, normal saline (NS) application group, aminophylline application group, aminophylline injection group, 1-day herb application group, 7-day herb application group and 14-day herb application group. Asthma was induced by Hutson's method in guinea pigs except the normal control group. Guinea pigs in herb application groups were treated by external application of a compound herbal medicine 60 min once every day. Guinea pigs in NS application group were treated by external application of NS. Guinea pigs in the two aminophylline-treated groups were treated by external application and intraperitoneal injection of aminophylline at a dose of 400 mg/kg, respectively. The guinea pigs were killed and the sera were obtained after 1-day, 7-day and 14-day treatment in the three herb application groups, 7-day treatment in the NS application group, the aminophylline application and injection groups, respectively. Serum pharmacological method was used to do the experiment, the effects of different sera on the constriction of tracheal strips were observed, and the constriction rates were calculated. RESULT: The serum containing herbs had an effect in reducing the constriction of tracheal spiral strips induced by Ach, and the effect was similar to that of the serum obtained from the aminophylline injection group. The constriction rate of the tracheal spiral strips was decreased when herbal application treatment was prolonged within a period of time, and it became stable when herbal application treatment was between 7-14 days. The constriction of tracheal spiral strips induced by Ach could be reduced remarkably when it was previously treated by serum containing herbs. CONCLUSION: The anti-acetylcholine function with a time-dependent effect is one of the mechanisms of herbal application treatment along meridians for asthma, and furthermore, herbal application treatment along meridians might be useful for preventing asthma.

BACKGROUND: Previously, there are some researches on studying the effects of hypoxia or hypothermia singly. Nowadays, study on the synthetic action of hypoxia and hypothermia has been attached widely importance.OBJECTIVE: To explore the combination effects of acute hypothermia and hypoxia on superoxide dismutase(SOD) and malondialdehyde in normal rabbits and rabbits with myocardial ischemia.DESIGN: Randomized controlled trial.SETTING, MATERIALS and INTERVENTIONS: Experimental site:Physiological Laboratory of Youjiang Ethnic Medical College. A total of 32 healthy rabbits weighed between 1.8to 2.5 kg were chosen. They were randomly divided into 4 groups each with 8 rabbits: normal group, hypothermia and hypoxia group, ischemic group with normal temperature and oxygen,ischemic group with hypothermia and hypoxia. Normal rabbits and ischemic rabbits were put in fridge at low temperature( -10 ± 2) ℃ and low oxygen (oxygen content of 8.5% ) for one hour and tested activity of SOD and concentration of malondialdehyde in serum.MAIN OUTCOME MEASURES: Comparison of serum SOD and malondialdehyde between normal rabbits and myocardial ischemic rabbits under the acute hypothermia and hypoxia conditions.RESULTS: The SOD level of rabbits in hypothermia and hypoxia group [(371.04 ± 29.96) kNU/L] was much lower than that of normal temperature and oxygen group[ (424.09 ± 22.59) kNU/L] ( t = 5.21, P < 0.01) while the content of malondialdehyde[ (5.58 ± 0. 44) μ mol/L] was greater than that of normal group[ (4.44 ± 1.11) μ mol/L] ( t = 3.21, P < 0.05) . In myocardial ischemia group with hypothermia and hypoxia, SOD was greatly lower than that of myocardial ischemia group with normal temperature and oxygen(t=4.37, P < 0.01) while the malondialdehyde was greatly higher than that of ischemic group with normal temperature and oxygen group(t=2.94, P <0.05).CONCLUSION: The combined effects of acute hypothermia and hypoxia will aggravate the damage to body.

Objective To explore the roles of follicular helper T cells (Tfh) in the pathogenesis of pemphigus and bullous pemphigoid (BP).Methods Venous blood was collected from 20 patients (including 10 cases of pemphigus and 10 cases of BP) and 20 age- and sex-matched normal human controls.Enzyme linked immunosorbent assay (ELISA) was performed to measure the levels of serum interleukin (IL)-21 and anti-BP180-NC16A antibody,and flow cytometry to detect the percentage of Tfh in peripheral blood mononuclear cells (PBMCs).The relationship of anti-BP180-NC16A antibody titer to Tfh and IL-21 levels was assessed by linear correlation analysis.Results The serum IL-21 level and Tfh percentage in PBMCs were significantly higher in patients with pemphigus than in the normal controls (95.33 ± 33.69 vs.54.50 ± 18.13 pg/ml,t =3.38,P＜ 0.01;12.08％ ± 4.74％ vs.6.15％ ± 1.62％,t =3.74,P ＜ 0.01),and higher in patients with BP than in the normal controls ( 106.70 ± 44.91 vs.55.37 ± 15.89 pg/ml,t =3.41,P ＜ 0.01; 11.85％ ± 3.14％ vs.6.03％ ± 1.74％,t =5.13,P ＜ 0.01 ).The titer of anti-BP180-NC 16A antibody was positively correlated with the percentage of Tfh (r =0.67,P＜ 0.05) and serum level of IL-21 (r=0.77,P＜ 0.01) in patients with BP.Conclusions There is a significant increase in the percentage of Tfh and serum level of IL-21 in patients with pemphigus and BP,hinting a certain role of Tfh and IL-21 in the pathogenesis of pemphigus and BP.

AIMTo investigate the bio-affinities of ligustilide and butylidenephthalide to rat aortic smooth muscle cells and the inhibitory effects of them on bFGF-stimulated proliferation of rat vascular smooth muscle cell (VSMC).

METHODSVSMCs were cultured from rat aorta pectoralis and identified by an immunohistochemical method. The bio-affinities between solute (ligustilide or butylidenephthalide) and cell membrane were measured by rat aortic cell membrane chromatography (CMC). The inhibitory effects of ligustilide and butylidenephthalide on bFGF-stimulated VSMC proliferation were evaluated by MIT colorimetric method.

RESULTSBoth ligustilide and butylidenephthalide had selective affinities to rat aortic smooth muscle cell as the same as verapamil, one of the calcium ion antagonists. They could potently inhibit the bFGF-stimulated VSMC proliferation at the concentrations of 5.5 and 11.1 micromol x L(-1), separately (P < 0.05), but had no effects on the normal VSMC growth.

CONCLUSIONBoth ligustilide and butylidenephthalide can inhibit the abnormal proliferation of VSMC induced by bFGF.

4-Butyrolactone ; analogs & derivatives ; isolation & purification ; pharmacology ; Animals ; Aorta, Thoracic ; cytology ; Cell Proliferation ; drug effects ; Cells, Cultured ; Female ; Fibroblast Growth Factor 2 ; antagonists & inhibitors ; Ligusticum ; chemistry ; Male ; Muscle, Smooth, Vascular ; cytology ; drug effects ; Myocytes, Smooth Muscle ; cytology ; drug effects ; Phthalic Anhydrides ; isolation & purification ; pharmacology ; Plants, Medicinal ; chemistry ; Rats ; Rats, Sprague-Dawley

Objective To investigate the correlation between the expression of HIF-1a (hypoxia inducible factor 1,HIF-1a) in perihematomal brain issue and formation of brain edema in patients with hypertensive cerebral hemorrhage.Method Perihematomal brain issue was collected in the course of hematoma elimination in 32 patients with hypertemive intraeerebral hemorrhage.Expressions of HIF-1a and vascular endothelial growth factor (VEGF) were observed by immunohistochemistry.The volume of perihematomal brain edema on computed tomographie scan was determined by computed tomographic scan before surgery.The results of staining and the volume of perihematomal brain edema were analyzed in double blind fashion.Results HIF-1a protein immunohistochemical staining positive cells (2.8?0.8/HP) were identified dispersedly from 4 hours after acute hemorrhagic stroke in perihematomal brain issue,and reached the peak at 24～48 hours (12.5?3.9/HP).High expression of HIF-1a progressed at 48～72 hours (12.2?1.8/HP) after acute hemorrhagic stroke.There was a positive correlation between the expression of HIF-1a and VEGF (r=0.76,t=6.37,P