Objective To construct a novel tissue?engineered bone matrix gelatin (BMG)/poly[butylene succinate?co?tere?phthalate] (PBST) biphasic scaffold for annulus fibrosus. Methods The PBST spinning fibers were prepared by electrospinning and the porosity and water absorption rate were tested. Rabbit annulus fibrosus cells were isolated, cultured and identified through SafraninOstaining, and collagenⅡimmunohistochemical staining in vitro. And then annulus fibrosus cells were implanted on the PBST fiber, whose growth situation was observed by scanning electron microscope (SEM). Then the BMG/PBST biphasic scaf?fold was constructed by BMG as the outer annular fibrosus and PBST fiber as the inner annular fibrosus. The annulus fibrosus cells were implanted on the biphasic scafflod and cultured for 3, 7 and 21 days in vitro. The biomechanical and biological property was observed at the predetermined time point. Results The porosity of the fiber was 61.83%±7.33%and its water absorption rate was 297.34%± 57.13%. The identified result of annulus fibrosus cells were positive, suggesting that the cells have still kept their annulus fibrosus cells characteristics. The cells growth could be observed through SEM at 3rd and 7th day after implanted on the fi?bers. After cultured on the BMG/PBST scaffold, HE staining proved that the cells could ingress into the inner of fiber with time. SafraninOstaining and collagenⅡimmunohistochemical staining proved that the cells can secreted abundant proteoglycan and collagenⅡ, the special annulus fibrosus cell extracellular matrix. Compared with the BMG/PBST scaffold without cells, the elastic modulus of biphasic scaffold was increased from 14.83±1.02 MPa to 17.56±1.47 MPa after cultured with cells for 21 days in vitro. Conclusion The novel tissue?engineered biphasic scaffold for annulus fibrosus constructed with BMG and PBST fiber spinning has good cytocompatibility and biomechanical characteristics, which provide a basis for the complete tissue engineered interverte?bral disc.

BACKGROUND:Tissue-engineered intervertebral disc has provided a new biological therapeutic approach for intervertebral disc degeneration. Tissue-engineered annulus fibrosus is one key step of constructing a complete tissue-engineered intervertebral disc. OBJECTIVE:To sum up the research progress of tissue-engineered annulus fibrosus from the folowing aspects: structural features, scaffold materials, seed cels. METHODS:PubMed database and Wanfang database (2000-2015) were retrieved by the key words of “tissue engineering, intervertebral disc, annulus fibrosus, seed cel, scaffold, construction” in Chinese and in English, respectively. According to inclusion and exclusion criteria, 48 literatures were involved for summarization. RESULTS AND CONCLUSION: Previous studies about tissue-engineered annulus fibrosus only focused on cel adhesion, proliferation and extracelular matrix secretion on the scaffold. Currently, tissue-engineered annulus fibrosus exhibit similar features to the natural annulus fibrosus in the folowing aspects: cel function, tissue structure and mechanical features, and relevant animal experiments have achieved certain results in animal experiments. However, it is stil difficult to build a tissue-engineered annulus fibrosus entirely similar to the natural one, and we need to further improve scaffold materials, culture conditions, colection of seed cels. The current strategies of annulus fibrosus construction stil focus on single phase of scaffold, and the biphasic scaffold and complete intervertebral disc scaffold wil be the trend of the researches. Technology of induced differentiation of stem cels provides a broach source of seed cels for tissue-engineered annulus fibrosus.

OBJECTIVE:To establish the quality standards of processed Sophora japonica.METHODS:Different batches of processed S. japonica products were detected in respect of property,microstructure,TLC,moisture,total ash,acid insoluble ash,extract inspection,content of total flavonoids and rutin. RESULTS:The limit value of test term for S. japonica,stir-baked S. japonica,S. japonica carbon were confirmed. CONCLUSION:Established standard is applicable for the quality control of processed S. japonica.

Objective To investigate the best digestion time for rabbit rib cartilage cells separation by collagenase Ⅱ diges‐tion method .Methods Rabbit cartilage tissue was taken from 3‐month‐old New Zealand white rabbits ,and then digested in 0 .2%collagenase Ⅱ solution for 4 ,8 ,12 ,16 and 20 h under vibration at 37 ℃ ,respectively .The corresponding experiments were named as group A ,B ,C ,D ,and E .The obtained primary cells were cultured and passaged after counting .The cell morphology ,proliferation rate ,secretion of extracellular matrix of passage 2 cells were compared to chose the best digestion time .Results The primary chon‐drocytes obtained in group C ,D and E were more than the other two group A and B .The chondrocytes cytoactive in group C and D were better than the other three group A ,B and E .The passage 2 chondrocytes in group C ,D had better secretion of extracellular matrix capability than the other three groups .Conclusion The best collagenase Ⅱ digestion time is 12 to 16 h ,at this digestion time period ,more primary chondrocytes with high cytoactive and strong proliferation capability can be obtained ,and the phenotype of their passage 2 cells is quite good .

Objective To discuss interventional treatment technique for benign ureteral strictures. Methods A total of 19 cases with 20 benign ureteral strictures were treated with balloon dilatation and double-pigtail ureteral stent placement. Results Among 19 caces,16 were succeeded in a rate of 84.2%.The three failure cases included one of compression by aberrant vessels,one by compression of fibrous-strings and the last one due to tortuosity of the ureter.Re-examinations,by intravenous urography and B ultrasound were carried out in 16 successors,showed gradually release of hydro-nephrosis and renal function improvement.Iatrogenic foreign body ocurred in 2 cases,caused by the protruding tail of stent which had been taken out under ureteroscopy. Conclusions Interventional technique for benign ureteral strictures possesses adventages with mild injury,simple operation,and a new effective method.

A fusion protein of glucagons-like peptide-1 and human serum albumin (GLP-1/HSA) was expressed and secreted into the fermentation broth with recombinant Pichia pastoris. The productivity of expressed GLP-1/HSA could reach 63.6mg/L in 10L fermentor. After concentrated with hollow-fiber ultrafiltration membrane, GLP-1/HSA was purified from fermentation broth by hydrophobic chromatography, negative ion exchange chromatography and gel filtration chromatography in turn. The HPLC analysis showed that the purified GLP-1/HSA had an overall purity of 95.8%. Furthermore, the analysis of in vivo activity indicated that GLP-1/HSA had the bioactivity of native GLP-1, and could significantly reduce blood glucose level 4h after intraperitoneal administration. It was concluded that a great deal of GLP-1/HSA with higher purity could be harvested by Pichia pastoris expression system and the established purification methods. Preliminary studies show a new potential for developing the long-acting GLP-1 analogs for clinical applications.

Objective To isolate and culture sweat gland epithelial cells in vitro,and to study the effects of acetylcholine (ACh) on intracellular flee calcium concentration ([Ca~(2+)]i) of cultured sweat gland epithelial cells.Methods Sweat glands epithelial cells were collected by enzymatic digestion.After ACh was added to the primary and first passage cells,[Ca~(2+)]i was examined using confocal laser scanning microscopy (CLSM) and the Ca~(2+) sensitive dye Fura 3/AM.Results The primary and first passage epithe- lial cells grew well.After ACh was added,opening of the calcium channel and significant [Ca~(2+)]i increase were observed when the primary and first passage cells were incubated with high concentration of calcium (2 mmol/L);no significant [Ca~(2+)]i increase was observed in those cultured without calcium.Conclusion Upon stimulation with ACh,calcium channels of cultured primary and first passage sweat gland epithelial cells would open,influx of extracellular Ca~(2+) occurred,which resulted in an increase of [Ca~(2+)]i.Extracellular bound calcium was therefore converted into intracellular free calcium.

Two fragments G1 and G2 of the glycoprotein G gene of bovine respiratory syncytial virus(BRSV) were selected for expression in Escherichia coli based on the analysis of glycoprotein G by DNA Star software.Then the two fragments of glycoprotein G were amplified by PCR with synthesized G gene of BRSV as the template.The amplified fragments G1 and G2 are 570bp and 308bp in length,respectively.The PCR products were cloned into pET30a vector and expressed in soluble form in E.coli after induction of cultured E.coli with IPTG.Both of the recombinant proteins G1 and G2 were purified by immobilized Ni ion affinity chromatography under native conditions.Then the purified proteins were analysed by Western blotting.The results showed that the purified recombinant protein G1 retained good antigenicity and specificity.But the purified recombinant protein G2 didn't possess biological activity.Antibodies against BRSV were detected in suspected bovine serum samples in China by using indirect ELISA and Western blotting with the purified recombinant protein G1.The purified recombinant protein G1 might be used as antigen for establishing serological methods for diagnosis of BRSV infection.And the purified recombinant protein G1 might also be used for preparing polyclonal and monoclonal antibodies for research on biological functions of glycoprotein G of BRSV.

Objective:To identify the effect of ?-synuclein overexpression on mitochondrial membrane structure with atomic force microscopy. Methods:?-syn expression was mediated by AAV (adeno-associated viral vector) and Recombinant AAV/?-syn and AAV/LacZ viral particles were stereotaxically injected in the left side of rat substantia nigra (SN) for rat model of ?-synuclein overexpression. Mitochondria were isolated from rats SN of Brain. Mitochondria were analysis with JC-1 staining,atomic force microscopy and Western blot. Results:By 16 weeks post-infection of AAV-?-syn,the level of ?-syn increased about 2 times in mitochondrial fraction with Western blot and mitochondrial membrane potential (??) decreased with JC-1 staining. Furthermore,mitochondria swelling and porous like structure formed on the mitochondrial membrane with atomic force microscopy. Conclusion:The data suggested that ?-syn could accumulate in mitochondria,might form mitochondrial membrane pores and lead to ?? decreases. ?-syn might lead to mitochondrial dysfunction in Parkinson's disease.

Objective To investigate the clinical outcomes derived from Ningmitai combined with tamsulosin to prevent double-J stent syndrome after laser lithotripsy with ureteroscope. Methods 117 patients underwent laser lithotripsy with ureteroscope and then placed a double-J stent for draining were collected from January 2010 to January 2013. Patients with double-J stent placement were divided into four groups determined by dosage regimen. Tamsulosin group (30 cases) was treated with tamsulosin (0.4 mg once daily) lonely, Ningmitai group (29 cases) was treated with Ningmitai (1.52 g, trice time a day) lonely, tamsulosin combined Ningmitai group (30 cases) was treated with tamsulosin and Ningmitai at the same time, operation control group (28 cases) was neither tamsulosin nor Ningmitai. The catheter was removed on the 3rd day post-lithotripsy and then remained double-J stent for 1 month. The scores of urinary tract, pain and the incidence of gross hematuria were assessed. Results The significant differences in the improvement of symptom score (χ2=22.038, P=0.000), pain score (χ2=9.876, P=0.020) and hematuria (χ2=8.000, P=0.046) were found among tamsulosin group, Ningmitai group, and tamsulosin combined Ningmitai group. The number of patients with symptomless, slight symptom in tamsulosin combined Ningmitai group were higher than those of tamsulosin group, Ningmitai group, operation control group (symptomeless:14 vs. 6, 3 and 2 cases;slight symptom:13 vs. 9, 5, 4 cases). The number of patients with>Ⅱpain score (7 vs. 9, 14, 17 cases) and incidence of hematuriag [26.6%(8/30) vs. 56.7%(17/30), 58.6% (17/29), 53.6% (15/28)] were lower in tamsulosin combined Ningmitai group than those of tamsulosin group, Ningmitai group, operation control group. The drug combination of Ningmitai with tamsulosin had the synergism to relived symptom and pain, and showed the more obviousthan lonely use. Conclusion The drug combination of Ningmitai with tamsulosin can be used in clinic for prophylactic purpose to prevent double-J syndrome.