Effect of cryoprotectant removal by microfluidic chip on developmental capacity of oocytes / 生物医学工程学杂志

In order to reduce osmotic damage and chemical toxicity of cryoprotectants (CPA) to oocytes during unloading process, the microfluidic chip was used to remove CPA from porcine MⅡ oocytes in this study. Firstly, the effects of unloading time, composition and concentration of diluting solutions of microfluidic method on survival rate and developmental capacity of oocytes were studied, then microfluidic method was compared with traditional one-step and two-step CPA unloading protocols. The results showed that when the total time is 8 minutes, the survival rate and morula rate of oocytes treated with microfluidic method could achieve 95.99% ± 4.64% and 74.17% ± 1.18%, respectively, which were not significantly different from fresh control group (98.53% ± 2.94%; 78.22% ± 1.34%). In addition, 1 mol/L sucrose diluting solutions were more beneficial than other solutions, and it was also showed that microfluidic method achieved better survival, cleavage rate of oocytes than traditional methods. Microfluidic CPA removal protocol can reduce the damage to oocytes during unloading process, and may further improve the cryopreservation effect of oocytes.

Effect of different transfection reagents and injection methods in mice testicular injection on the expression of exogenous gene / 生物工程学报

The purpose of this study was to study the effect of three different transfection reagents (Lipofectamine™ LTX & PLUS™, Lipofectamine 2000 and Nano-PAMAM-D) and three different testicular injection methods (rete testicular injection, seminiferous tubules injection and testicular interstitial injection) on the efficiency of production transgenic mice. After the mixtures of plasmid DNA (pEFP-C1) and transfection reagent were injected with different testicular injection methods, the sperm density, vitality, positive sperm rates and PCR positive transgenic mice rate were examined 30 days after injection. The results showed that the damage degree from slight to serious of three transfection reagents was Lipofectamine™ LTX & PLUS™, Lipofectamine 2000, and PAMAM-D. The sperm positive rates with green fluorescence of these three groups were 35.65%±0.69%, 12.86%±0.35% and 10.04%±0.20%, respectively. The PCR positive rates of transgenic newborn mice were 29.17%, 13.70% and 5.88%, respectively. Among the groups of different testicular injection methods, the damage degree from slight to serious was rete testicular injection, seminiferous tubules injection, and testicular interstitial injection, whereas the sperm positive rates with green fluorescence were 35.13%, 15.13%, and 0%, respectively. The PCR positive rates of transgenic newborn mice among different testicular injection groups were 33.3%, 12.5%, and 0.0%. The combination of rete testicular injection and Lipofectamine™ LTX & PLUS™ had the lowest toxicity and highest transgenic efficiency in the production of transgenic mice.

Efficacy of rectally administered indomethacin for the prevention of post ERCP pancreatitis / 中华胰腺病杂志

Objectives To evaluate the efficacy of rectally administered indomethacin for the prevention of post-ERCP pancreatitis(PEP).Methods All eligible patients without high risk factors such as heart,lung,liver and kidney,coagulation dysfunction,without malignant disease and contraindication for NSAIDs,and pre-operative imaging study and lab test suggesting no pancreatitis,aged from 18 ～ 75 who underwent ERCP and EST were enrolled.In a randomized prospective trial,patients were randomized to receive a suppository containing indomethacin,100 mg,or an identical placebo 30 minutes after ERCP.PEP was diagnosed when there was pancreatitis related clinical symptoms,and serum amylase was higher than 3 times of the normal values,and when the patient needed more than 1 day hospitalization.Patients with PEP were evaluated with APACHE Ⅱ score 72 hours after ERCP.Results During 2004 ～ 2010,a total of 348 patients were enrolled,of which 182 received indomethacin and 166 received placebo.Six patients developed pancreatitis in the indomethacin group and 14 in the placebo group (3.3％ vs.8.4％,P ＜0.05),and the difference between the two group was statistically significant ( P ＜ 0.05 ).In those patients with PEP,the APACHE Ⅱ scores in indomethacin group (4.3 ± 1.3 ) were lower than that in the placebo group (7.4 ±1.7),and the difference between the two groups was statistically significant ( P ＜ 0.05 ).The incidence of hyperamylasemia in both groups was not statistically significant (9.3％ vs.10.8％,P ＞ 0.05 ).Conclusions This trial shows that rectally administered indomethacin after ERCP and EST can effectively reduce the incidence and severity of PEP.

Effect of trehalose on the freeze-dried boar spermatozoa / 生物工程学报

After freeze-drying, the ultrastructure of boar sperms was observed by optical and electron microscopy. The in vitro development ability of the sperm was also examined with intracytoplasmic sperm injection (ICSI). The rate of male pronuclear formation was (68.52%), for cleavage (59.17%) and for blastocyst formation (19.16%) of the trehalose group (0.2 mol/L), significantly higher than those of the 50 mmol/L EDTA group (64.59%, 56.26% and 15.62%) and the control group (35.36%, 52.33% and 8.60%) (P < 0.05). After storage for 60, 120 and 180 d at 4 degrees C, no significant difference in the in vitro development was observed (P > 0.05). The male pronuclear, cleavage and blastocyst formation after ICSI with freeze-dried spermatozoa incubated for 1 h was superior than those incubated for 2 h (P < 0.05). No significant differences in the structures after stored at 4 degrees C or -20 degrees C (P > 0.05) were observed between the trehalose group and EDTA group. The percent of B grade freeze-dried boar spermatozoa in the trehalose group was higher than that of the EDTA group (P < 0.05). Based on the ultrastructure observation, main cryogenic damage in freeze-dried boar spermatozoa was swelling, damage or rupture in the sperm acrosome, neck and tail.

Effects of angiotensin Ⅱ and captopril on outward potassium channel currents in canine atrial myocytes / 中华急诊医学杂志

Objective To observe effects of angieminⅡ(AngⅡ)and captopril on outward potassium channel currents in canine atrial myoeytes,and to study mechanisnof Ang II and capupril on atrial arrhythmia.Method Ten healthy adult mongrel dogs(general class),weighing 15 to 20 kg,male and female informality,were provided bythe service centre of Tianjin Li-qun experimental animals.Single canine atrial myotcyte was acutely isolated and whole-cell configtmtion of the patch-clamp tchnique was used to detec trapidly activating delayed reefifier outward K+ current(Ikr),slowly activating delayed recti fier outward K+ current(Iks),ultra-rapidly aetivatin delayed rectifier outward K+ current(Ikur)and transient outward potassium current(Ito)before and after An II and captopril peffion.Software of pClamp 7.0 for windows and pClampfit 7.0 Was used to measure current and data were expressed as mean±standard deviation(x±s).SPSS 10.0 statistical was used for statistical analysis.The paired t test was useel for comparison betwn before and after treatment.P<0.05 was comidered as statistical significance.Results AngII(0.5/mol/L)increased Ikr and Iks,ilfibited Ito[(19.54±2.41)pA/pF vs.(24.83±2.52)pA/pF,P=0.001;(20.69±2.29)pA/pF Vfl.(25.59±3.42)pA/pF,P:0.0003;(6.34±1.93)pA/pF vs.(3.71±1.50)pA/pF,P=0.001)],and had no effect on k[(19.78±1.22 pA/pr Vs.(20.39±1.50)pA/pF,P=0.258)].Captopril(5tot/L)had no significant effect on Ikr.,b.k and[(19.11 4-4.91)pA/pF vs.(18.99 4-4.04)-∥pF,P=0.808;(20.76 4-2.89)pA/pF vs.(20.27 a-3.46)pA/pF,P=0.305;(18.50 4-3.78)pA/pF vs.(18.25 4-4.02)pA/pF,P=0.704;(7.31±1.99)pA/pF vs.(6.89±2.12)pA/pF,P=0.136)].Conclusioas AngⅡmay promote atrial electrical remocof atrial fibrillation through outward potassium currents.As angiotemin-eonverting enzy/ne inhibitor.captioruk can prevent atrial electrical rodding of atrial fibrillation by inhibiting renin-angiotensin-system.

Optimization of porcine embryo vitrification / 生物工程学报

The purpose was to optimize the vitrification for porcine embryos cryopreservation. Blastocyst/Morula (5-6th day-embryos) were collected from superovulated Bama mini-pigs (sows/gilts). We compared different cryopreservation methods, cryopreservation tools, thining of zona pellucida (ZP) and recipient breeds on the efficiency of porcine embryo cryopreservation. The results showed that: in embryo survival rate and blastocyst cell number, there were no significant differences between cryopreservation method I [embryos were vitrified by two step method with open pulled straw (OPS) and glass micropipette (GMP) in solution 1 (TCM199 + 20% FBS + 10% EG + 10% DMSO) for 3 min, and solution 2 (TCM199 + 20% FBS + 20% EG + 20% DMSO + 0.4 mol/L SUC) for 1 min, stored in liquid nitrogen] and method II[Blastocysts were cultured for 25 min in NCSU23 + 7.5 microg/mL cytochalasin B, centrifuged at approximately 13 000 xg for 12-13 min, and recovered back into pNCSU23. They were then equilibrated for 5 min in 2 mol/L ethylene glycol in pNCSU23, washed quickly in the vitrification medium, 8 mol/L ethylene glycol, 7% polyvinylpyrrolidone (PVP) in pNCSU23, loaded into OPS/GMP, and plunged into liquid nitrogen]. GMP vitrification method was more suitable and efficient than OPS method (P < 0.05) in embryo survival rate (83.8% vs 77.6%) and blastocyst cell number (53.1 vs 47.5) after thawing. Thining of ZP did not increase the survival rate, but significantly improved blastocyst cell number in the survival blastcysts (60.1 and 46, P < 0.01). Local pig breeds (Fengjing sows) were more suitable as recipients for embryo transfer of vitrified/warmed blastcysts, which can improve pregnant rate and embryo efficiency.

Relationship between Serum Von Willebrand Factor and Diabetic Nephropathy in Type 2 Diabetic Patients / 中国医师杂志

Objective To investigate the change of serum von willebrand factor(vWF)level in the different stages of diabetic nephropathy and its significance. Methods 50 healthy subjects and 150 patients with type 2 diabetes mellitus were enrolled in this study. Diabetic patients were further divided into three subgroups according to their urinary albumin excretion rate(UAER): 56 patients with normal UAER,49 patients with microalbuminuria and 45 patients with clinical proteinuria. Serum vWF concentration was measured with emzyme linked immunosorbent assay (ELISA) method. Results vWF concentration was significantly higher in patients with type 2 diabetes mellitus than that in normal controls. Serum vWF concentration increased with the elevation of UAER. Serum vWF level was positively related with UAER, blood creatine and course of disease independently. Conclusion Serum vWF concentration increased in the patients with type 2 diabetes mellitus and the increased extent of vWF was consistent with the severe degree of diabetic nephropathy.