A strain of Mucor named M2,which could produce protease,was isolated from a traditional fermented soybean product.Culture conditions of proteases produced by M2 were studied therefore.The results showed that nitrogen source and carbon source preferred for protease production were soybean protein isolate and glucose,while inorganic salts preferred were KH2PO4,CaCl2 and MgCl2.The suitable culture conditions for protease production were as follows:culture temperature was 28℃,inoculation volume was 2%,liquid level was 100 mL in 300 mL triangle bottle at pH 5,rotating speed was 150 r/min and culture time was 48 h.The obtained protease activity in culture was about 4.35 U/mL.The protease produced by Mucor was analyzed with SDS-PAGE.The protease had a molecular weight of 36.4 kD.

Objective:To improve the primary culture method of rat pulmonary microvascular endothelial cells(PMVECs) and obtain purified PMVECs.Methods:The modified tissue block pasted culture method was used to isolate and culture Wistar rat PMVECs. The morphous of cultured cells were observed by microscopy. The cultured cells were identified by detecting factor Ⅷ related antigen and binding isolectin B4. Results and Conclusion:The morphous of cultured primary PMVECs in vitro showed short fusiform shape or polygon, and the monolayer of cultured cells displayed the shape of pavingstone. But the morphous changed followed the transfer of culture and the change of culture condition. The cultured cells had characterization of binding isolectin B4 and negative immunocytochemical staining for factor Ⅷ related antigen. The cultured PMVECs have good growth state and purity,and can be subcultringed stably.The observation of cell morphous integrating with immunocytochemical staining is a reasonable identification method for PMVECs.

Objective To establish a method of isolation, purification, primary culture and identification of alveolar type Ⅱ epithelial cells(AT-Ⅱ).Methods The AT-Ⅱs were isolated from Wistar rats by trypsin,purified by differential centrifugation, erythrocyte spallation, differential adherence and immune adherence, and identified by observing the morphology of cultured cells under the inverted phase and tannic acid staining. Results and Conclusion The cultured primary AT-Ⅱs in vitro presented single or island form growth, and their shapes were round or elliptical. A great deal of fine particles showed sharp contrast, and were observed in intracytoplasm. The cell nuclei were clear. They were positive for tannic acid staining.The primary culture AT-Ⅱs obtained from improved isolation and purification have good growth state and purity, and are suitable for research in vitro.

Objective To explore whether the degree of 99Tcm-methoxyisobutylisonitrile (MIBI) uptake in non-small cell lung cancer (NSCLC) could be correlated with the treatment response to three-dimensional conformal radiotherapy.Methods A total of 102 patients with NSCLC were studied with 99Tcm-MIBI SPECT before radiotherapy.The patients were classified by a follow-up CT as responders (complete or partial remission) and non-responders (stable or progressive disease).After intravenous administration of 740 MBq 99Tcm-MIBI, SPECT imaging at 10-30 min (early) and 2-3 h (delayed) were performed.Region of interest (ROI) was placed over the tumors and contralateral normal lung tissue.The uptake ratio of tumor to contralateral normal lung (T/N) was obtained from both early (ER) and delayed (DR) SPECT images.The retention index (RI) was measured as:RI = (DR - ER)/ER×100%.Statistical analysis was performed by two independent-sample t-test and Mann-Whitney U test using software SPSS 13.0.Results 99Tcm-MIBI uptake was significantly higher in responders than in non-responders:2.36 ±0.17 vs 1.82 ±0.14 (ER) and 2.48 ± 0.20 vs 1.94 ± 0.16 (DR), respectively (t = - 13.1,- 12.7, both P＜ 0.05).The median RI in the responders group was also significantly higher than that in the non-responders group (6.60% vs 5.13%, z = - 6.83, P ＜ 0.05).Conclusion ER, DR and RI of 99Tcm-MIBI SPECT might be useful to estimate the treatment response to three-dimensional conformal radiotherapy in patients with NSCLC.

BACKGROUNDResults of perimembranous ventricular septal defects (pmVSD) transcatheter closure have been reported in the literature mostly using a Amplatzer VSD device. However, the data of percutaneous closure of pmVSD with VSD occluder (VSD-O) made in China are still limited. We sought to analyze safety, efficacy, and follow-up results of percutaneous closure of pmVSD with VSD-O made in China.

METHODSBetween February 2005 and June 2009, 78 patients underwent percutaneous closure of pmVSD at our institution. A VSD device made in China was used for all subjects. The safety and the efficacy of the VSD-O were investigated.

RESULTSThe average age at closure was 11 years (range 2.5 to 44 years). The attempt to place device was successful in 74 patients (94.9%). The average device size used was 8 mm (range 5 to 16 mm). No deaths occurred. Total occlusion rate was 62.8% at completion of the procedure, rising up to 87.2% at discharge and 99.0% during follow-up. A total of eight early complications occurred (10.3%), but in all subjects these were transient. The average follow-up period was 40.5 months. The most significant complication was complete atrioventricular block (cAVB) in the early phase (five subjects, 6.4%) and during the follow-up (1 subject, 1.3%), which saw no need for pacemaker implantation in six subjects. Cox proportional hazards regression analysis showed that the age was only the variable significantly associated with the occurrence of this complication during the procedure (P = 0.025; relative risk 0.22). The subjects who experienced this complication were less than five years old.

CONCLUSIONSPercutaneous pmVSD closure used VSD-O made in China is associated with excellent success and closure rates, no mortality, and low morbidity. Longer follow-up data and improvements in device characteristics are needed to reduce the risk of cAVB.

Adolescent ; Adult ; Cardiac Catheterization ; adverse effects ; methods ; Child ; Child, Preschool ; China ; Female ; Heart Septal Defects, Ventricular ; therapy ; Humans ; Infant ; Male ; Prospective Studies ; Treatment Outcome ; Young Adult

AIMTo develop a sensitive and specific HPLC method for determination of trigonelline in rabbit plasma, and study the pharmacokinetics in rabbit.

METHODSAfter ig of fenugreek extract and i.v. of trigonelline in rabbit, the biological samples could be well purified after precipitation of protein with methanol and acetonitrile. Asahipak NH2P-50 column was used, the mobile phase consisted of acetonitrile-water (90:10) at a flow-rate of 1.2 mL.min-1, and detection wavelength was set at UV 265 nm. The column temperature is 30 degrees C.

RESULTSThe calibration curve was linear in the range from 0.98 mg.L-1 to 31.28 mg.L-1, with r = 0.9986, the detection limit of this method was 50 micrograms.L-1. The concentration-time curves of trigonelline in rabbits after ig and i.v. administration were shown to fit one-compartment and two-compartment open model, respectively. The main parameters after ig of fenugreek extract were as follow: T1/2(Ka) was 0.9 h, T1/2(Ke) was 2.2 h, V was 0.64 L.kg-1, AUC was 1.93 mg.min.L-1. The main parameters after i.v. of trigonelline were as follows: T1/2 alpha was 10.8 min, T1/2 beta was 44.0 min, K21 was 0.044 min-1, K10 was 0.026 min-1, K12 was 0.017 min-1, AUC was 931.0 mg.min.L-1.

CONCLUSIONTrigonelline showed a middle rate of absorption and fast rate of elimination in rabbit. Meanwhile, the method is simple, accurate, with a good reproducibility, and it provide a basic method for the investigation of trigonelline and fenugreek pharmacokinetics.

Alkaloids ; blood ; isolation & purification ; pharmacokinetics ; Animals ; Antineoplastic Agents, Phytogenic ; blood ; isolation & purification ; pharmacokinetics ; Area Under Curve ; Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; isolation & purification ; pharmacokinetics ; Female ; Male ; Plants, Medicinal ; chemistry ; Rabbits ; Seeds ; chemistry ; Trigonella ; chemistry

OBJECTIVETo study the clinic application of compound flap pedicled with arterial arch of palpebral margin in repairing severe full defect of eyelid.

METHODSAccording to eyelid structure and the defect size, the two compound flaps were designed beside the defect based on the arterial arch of the palpebral margin. If the defective area was too large, the lateral compound flap may be extended to lower or upper eyelid 0.5 cm away from the outer canthus, then cut and propelled the two compound flaps to repair the full eyelid defect.

RESULTS20 cases had been cured with this method since 1998. In this cases, 4 cases were basal cell carcinoma of eyelid, 2 cases were squamous carcinoma, 3 angiomas, 6 chromatophore nexuses, 3 traumatic defects, 2 congenital defects. The largest length of eyelid full defect was 1.7 cm and the smallest was 0.8 cm. 6 cases were upper eyelid defect and 14 cases were lower eyelid defect. All the compound flaps survived completely without any complications. All cases obtained satisfactory results functionally and esthetically.

CONCLUSIONSRepairing full eyelid defect with the compound eyelid flap is the same kind tissue repairing. It can not only provide enough tissues to primary repair large full defect of the upper or lower eyelid to restore normal anatomical structure and appearance of the eyelid, but also is easy to be operated without severe secondary deformities. The arterial arch of the palpebral margin is constant and the blood supply of the compound flap is reliable. It is an ideal method of repairing the eyelid defect.

Adult ; Aged ; Child ; Child, Preschool ; Eyelids ; blood supply ; transplantation ; Female ; Humans ; Infant ; Male ; Middle Aged ; Ophthalmic Artery ; transplantation ; Reconstructive Surgical Procedures ; methods ; Surgical Flaps

Objective To detect the resistance index and esterase activity of each generation of DDVP-resistant Culex mosquitoes and analyze the relationship between insecticide resistance and esterase. Methods WHO bioassay and micro-plate measurement were used for the detection. Results The resistance index increased to 12.17 after 43 generations' insecticide selection compared to 1.00 as sensitive isolate. The nonspecific esterase(NSE) activity of the mosquitoes became strengthened with the extension of the generations, and the individual frequency of those with OD values no less than 0.9 increased gradually, consistent basically to the bioassay. The AChE average inhibition rate decreased with the extended generation and increased resistance, and the individual frequency of those with inhibition rate less than 30% became strengthened with the extension of generations, showing a positive correlation. Conclusion The activity of NSE and AChE shows a correlation with DDVP resistance.

Objective To investigate the effect of methylprednisolone sodium succinate on oxygen indexes (PaO2/FiO2) and serum biochemical indexes in patients with respiratory distress syndrome.Methods Seventy-two patients with respiratory distress syndrome were randomly divided into treatment group and control group,36 cases in each group.Control group was given routine treatment.Treatment group was given methylprednisolone sodium succinate 120 mg · d-1,by intravenous drip on the basis of control group.All the patients were treated continuously for two weeks.The PaO2/FiO2 and acute physiology and chronic health evaluation Ⅱ (APACHE Ⅱ) scores before and after treatment were observed.The levels of nitric oxide (NO),interleukin-6 (IL-6),tumor necrosis factor-α (TNF-α),C-reactive protein (CRP) and adverse drug reactions in two groups were observed.Results After treatment,the PaO2/FiO2 levels in treatment group and control were (287.62 ±24.57),(213.21 ±25.67)mmHg,the APACHE Ⅱ scores were 11.23 ± 1.66,15.59 ± 2.21,all with significant difference (all P ＜ 0.05).The levels of NO were (58.88±32.76) and (74.28 ±32.94) μ mol· L-1,the levels of IL-6 were (167.94 ±25.29) and (213.11±21.50) pg· mL-1,the levels of TNF-α were (22.35 ±8.93) and (36.30±9.88) pg·mL-1,the levels of CRP were (6.43 ± 3.02) and (9.25 ± 2.94) mg · L-1,all with significant differences (all P ＜ 0.05).There were 2 cases of short-term elevated blood pressure in treatment group,and there was no adverse drug reaction in control group,without significant difference (P ＞ O.05).Conclusion Methylprednisolone sodium succinate in the treatment of respiratory distress syndrome can significantly improve the PaO2/FiO2,reduce the APACHE Ⅱ score and serum biochemical indicators,and can improve the clinical effect with high safety.