BACKGROUND: The long-term cryopreservation of embryonic stem cells is an important link in its researches and application. Traditional protectants of cryopreservation often contain dimethyl sulfoxide, glycerine, animal serum, etc., but they cannot effectively ensure the survival rate of some cells, and even damage some functions of the cells.OBJECTIVE: To develop the suitable protectant for the cropreservation of mouse embryonic stem cells. DESIGN: An observational study.SETTINGS: Department of Physiology, Conghua College of Guangzhou Medical College; Sinocells Bio Technologies Co.,Ltd.MATERIALS: The experiment was carried out in the laboratory of Sinocells Bio Technologies Co.,Ltd. from October 2004 to September 2005. Mouse cell line mES-1 was given as a present by the Research Room of Cell Biology, Institute of Basic Medical Sciences, Academy of Military Medical Sciences of Chinese PLA.METHODS: The cells collected from cultured mES-1, a mouse embryonic stem cell line. Dimethyl sulfoxide was used as the main protectant for cryopreservation. There were four different groups according to different protectants. In the control group, the protectant for cryopreservation contained DMEM (high sugar), 10% dimethyl sulfoxide, 10% FBS,10 mg/mL ascorbicacid, 0.18 mg/mL inositol and 0.44 mg/mL folic acid; Besides, 7.56 mg/mL mannose in the mannose group, 34.23 mg/mL mycose in the mycose and 41.94 mg/mL saccharu in the saccharu group were supplemented respectively. The effects of different polyols on the survival rate and ability of multiple differentiation of mouse embryonic stem cells after cryopreservation were observed.MATN OUTCOME MEASURES: Formation rate of mouse embryonic stem cells colony; Formation rate of embryoid bodies.were all significantly decreased as compared with that before cryopreservation [control group: (24.0±8.8)%; mannose group: (42.0±10.1)%; mycose group: (84.0±8.2)%; saccharu group (70.0±14.2)%; before cryopreservation: (95.0±4.7)%,embryoid bodies in the mycose group was higher than those in the control group and mannose group [(90.0±5.2)%,(80.0±6.9)%, (82.0±9.6)%, P＜0.05], but had no significant difference as compared with that in the saccharu group.CONCLUSION: The protectant for cryopreservation by taking mycose as the main component can effectively maintain the abilities of self-renewing and multiple differentiation of mouse embryonic stem cells.

ObjectiveTo explore the effects of fisetin on the learning and memory abilities impairments induced by Aβ in mice.MethodsAlzheimer's disease (AD) animal model was made by single intracerebroventricular infusion of Aβ (1-42) through guide cannula.Fisetin was orally administered 7 days before Aβ infusion once a day,and continued throughout the experimental period.Water maze test began on day 3 after Aβ infusion.All mice were sacrificed and hippocampi were dissected immediately after behavioral test.The protein expression of hippocampal nuclear Nrf2 and the mRNA level of HO-1,GCLC and GCLM were examined by western blotting and RT-RCR techniques respectively.Results ( 1 ) Aβ (1-42) significantly increased escape latency in hidden platform test ( (25.4 ± 3.33 ) s ),and decreased the number of crossings in probe test ( 1.70 ± 0.25 ) compared with control ((9.05 ± 1.37 )s) for hidden plat form ;4.50 ± 0.41 for probe test) and Aβ (42-1)-treated group ( ( 10.80 ± 1.38)s) for hidden platform test; 4.10 ±0.39 for probe test; P＜0.01 ).The prolonged treatment with fisetin dose-dependently reversed the changes (10 mg/kg:17.54 ± 3.56s for hidden platform test;2.50 ± 0.40 for probe test,P＜0.05,20 mg/kg:( 13.04 ± 2.36) s for hidden platform test;3.60 ±0.36 for probe test,P＜0.01 ).(2) Aβ (1-42) significantly decreased the nuclear Nrf2 protein level (0.07 ±0.02),and mRNA level (0.45 ±0.04) of HO-1,GCLC (0.41 ± 0.04) and GCLM (0.26 ± 0.03 ) in the hippocampus of mice compared with control (0.18 ± 0.02 for Nrf2 ;0.83 ± 0.09 for HO-1 ; 1.01 ± 0.10 for GCLC; 0.65 ± 0.07 for GCLM) and Aβ (42-1 ) -treated group (0.21 ± 0.02 for Nrf2 ; 0.90 ± 0.08 for HO-1 ; 1.11 ± 0.11 for GCLC ; 0.72 ± 0.07 for GCLM) ( P ＜ 0.05 or P ＜ 0.01 ).However,fisetin administration significantly counteracted these changes ( 10 mg/kg:0.11± 0.01 for Nrf2 ; 0.56 ± 0.06 for HO-1 ; 0.61 ± 0.04 for GCLC ; 0.35 ± 0.04 for GCLM ; 20 mg/kg:0.16 ± 0.02for Nrf2;0.79±0.10 for HO-1;0.86±0.09 for GCLC;0.51±0.04 for GCLM;P＜0.05).ConclusionFisetin attenuates learning and memory impairments induced by Aβ (1-42) through activation of Nrf2 antioxidant signaling pathway.

Objective To investigate the diagnosis and therapeutic methods of primary tumors of duodenum(PTD).Methods Clinical data of 86 patients with primary duodenal tumors who were hospitalized in the First Affiliated Hospital of China Medical University during 26 years were analyzed retrospectively.Of all the tumors,18 cases were benign tumors and 68 cases were malignant tumors.Results The clinical(manifestations) were symptoms of bile duct obstruction,GI and nonspecific symptoms,and signs included(fever),anemia and weight loss.The correct preoperative diagnostic rate by auxiliary examinations were:(duodenal) endoscopy 92%,air barium double radiography 89%,MRCP 85%,BUS 33%,CT 51%,MRI 75%,and duodenal endoscopy combined with air barium double radiography were 95%.Tumors were mostly located in the descending portion of duodenum(71 cases).Eighteen cases of benign PTDs were resected(completely) with a 5-year survival of 92.3%.Among the 68 maligant PTD cases,35 cases underwent(pancreatoduodenectomy),and 8 cases received segmental duodenectomy,with a resection rate of 63.2% and the 5-year survival rate of 31.0%.The remaining 25 cases,in which the tumors were unresectable,were treated by bypass operations,the 5-year survival rate was 0%.Conclusions Patients with PTD usually lack specific symptoms and signs,but combination of endoscopy,GI air barium double radiography and other(examinations) can improve the preoperative positive diagnostic rate.Segmental duodenectomy and simple tumor resection are curable for primary benign tumors of duodenum;while for primary malignant tumors of(duodenum),the therapy of choice should be pancreatoduodenectomy.

Objective To investigate the energy metabolism state and the posttransplantive function under different warm ischemia time (WIT). Methods The left lobe of pancreas was resected, and according to WIT (30, 60, 90, 120 min), the canines were randomized into 4 groups. The graft was perfused and preserved by the UW solution (0-4 ℃) cold storage method for 24 h. And then the heterotopic segmental autotransplantation of pancreas was performed, while the normal control group and no warm ischemia(NWI)group were also set up. Blood sugar (BG) and intravenous glucose tolerance test (IVGTT) K value and the plasma insulin level were examined in the first week after pancreas transplantation. The amylase level of pancreatic juice 24 h and one week after transplantation were detected either. The tissue concentrations of adenosine triphosphate (ATP) and total adenine nucleotides (TAN) were determined using high performance liquid chromatography (HPLC). The pancreas cortex was taken for morphological analysis and ultrastructure study 1 h after transplantation. Results Plasma glucose in groups of 30 min and 60 min was returned to normal 3-5 days after transplantation, while groups of 90 min and 120 min could not one week after transplantation. IVGTT K values in groups of NWI, 30 min and 60 min were more than 1, while those in groups of 90 min and 120 min were less than 1 one week after transplantation. The plasma insulin levels in groups of 90 min and 120 min were much lower than those in groups of NWI, 30 min and 60 min (P

Objective To investigate the association between the single nucleotide polymorphisms (SNP)in-308 loci of tumor necrosis factor-α(TNF-α)gene promoter region and chronic hepatitis B (CHB).Methods Genotypes of-308 loci of the TNF-αpromoter were examined by the polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP)in 142 patients with chronic hepatitis B (CHB group)and 150 healthy controls (HC group).The indexes for evalua-ting the curative effect were the ALT,AST,HBeAg,HBV-DNA and the viral load weight,HBV-LP and HBV-PreS1,mean-while,the correlations between related indexes and SNP in TNF-α308 loci were explored as well.Results There was no sta-tistical significance in frequency distribution difference of the genotypes and alleles of-308 loci between CHC and HC groups (P>0.05),the protective factors of TNF-α308 allele A may be not associated with CHB (OR=1.529,OR95%CI:0.872~2.684).There was no association between TNF-αgene promoter-308G/A polymorphism and the positive rates of AST, ALT,HBV-LP and HBV-PreS1 (P>0.05),however,TNF-α-308G/A polymorphism associates with the positive rates of HBeAg and HBV-DNA,and A-allele of 308 loci may increase the risk of HBeAg and HBV-DNA positive expression (HBeAg:OR=3.256,OR95%CI=1.105~9.594;HBV-DNA:OR=2.847,OR95%CI=1.059~7.655).Furthermore,A-allele compared with Gallele,statistically significant differences were observed in the certain HBVDNA viral load range of104~107 copies/ml (P <005).Conclusion TNFαgene promoter308G/A polymorphism would not be associated withCHB,but the TNFα308 gene G mutation of Aallele,which was associated with HBVDNA viral load,may be the susceptible factors of HBV infection.

BACKGROUND:Stromal cel derived factor-1 is a smal molecular protein with a wide range of biological activity that can cause immune cel chemotaxis, and it also has a chemotactic effect on bone marrow stem cels and periodontal ligament cels.
OBJECTIVE:To investigate the effect of stromal cel derived factor-1 with different concentrations on the proliferation of bone marrow stem cels and to probe the best concentration.
METHODS:Bone marrow stem cels from beagle dogs were culturedin vitro and stimulated by different concentrations of stromal cel derived factor-1 (100, 200, 300 μg/L). MTT was used to detect the influence of stromal cel derived factor-1 on the proliferation of bone marrow stem cels so as to screen the best concentration of stromal cel derived factor-1. Then, stromal cel derived factor-1 at the best concentrations was used to intervene the bone marrow stem cels, and MTT was used again to detect the proliferation of bone marrow stem cels.
RESULTS AND CONCLUSION:Stromal cel derived factor-1 at concentrations of 100, 200, 300 μg/L could promote the proliferation of bone marrow stem cels, and the effect was more notable at 200 and 300 μg/Lbut withno significant difference. Therefore, 200 μg/L was considered to be the best concentration of stromal cel derived factor-1 for intervention of bone marrow stem cels. Compared with the blank control group, 200 μg/L stromal cel derived factor-1 could significantly promote the proliferation of bone marrow stem cels. Taken together, stromal cel derived factor-1 can promote the proliferation of bone marrow stem cels, and its best concentration is 200 μg/L.

BACKGROUND: Although there are many studies on acute patellar dislocation in adolescents, the optimal treatment strategy is under discussion.OBJECTIVE: To summarize the clinical efficacy of reconstruction of the medial patellofemoral ligament (MPFL) in the treatment of adolescent acute patellar dislocation using double-pulley technique.METHODS: Thirty cases of acute patellar dislocation were enrolled and received reconstruction of the MPFL using double-pulley technique. We proceeded to prepare the bone bed for anchor placement at the medial point of the superior edge of the patella and at the upper 1/3 patella along the long axis of the patella using arthroscopic grinding. Two double-loaded 5.0 anchors were respectively placed at the medial point of patella and at the upper 1/3 patella parallel to the patellar articular surface, and then tied using the double-pulley technique. Long longitudinal incisions were performed at the medial condyle and at the adductor tubercle to expose these two structures, the deep fascias tunnel between two incisions were penetrated to pull the tendon weave end through the tunnel until the lateral side along a femoral tunnel; finally the tendon was fixed using screws in appropriate tightness. All patients were followed for more than 10 months, and the patellar-related parameters and knee function parameters were compared before and after operation. RESULTS AND CONCLUSION: There were 12 cases of intra-articular cartilage injury and 4 cases of meniscus injury. The patella title angle, Q-angle and outward shift distance at the last follow-up were less than those before operation. Lysholm and International Knee Documentation Committee scores at the last follow-up were higher than those before operation. The mean operation time was (90±10.5) minutes. There was one patient with knee movement angle from 0° to 30° and one patient with a patellar fracture caused by slipping and treated with internal fixation. None of patients appeared with patellar dislocation, positive apprehension test infection or poor wound healing. These results indicate that the simple and mini-invasive double-pulley technique is a good treatment strategy for acute patellar dislocation, because it is consistent with patellar anatomical stability, holds patellar trajectory visibility, quick recovery and good curative effect. Additionally, osteotomy is recommended when the simple efficacy is not satisfactory.

Objective To study the effects of L-arginine on the reperfusion injury of hypothermic perfusion and on preservation of isolated pancreas(ISP) in canines . Methods After segmental pancreatic transplantation(SPTS) model were set up,the animals were randomly divided into 3 groups:(1)ISP irregulated and preserved with Euro Colln Solution(ECS)plus L-arginine(L-arg group);(2)IPS irregulated and preserved with ECS plus N-nitro-L-arginine methy ester(L-NAME group);and(3)IPS irregulated and preserved with ECS plus normal saline(Control group).The IPS were irregulated and preserved at 0-4℃ for 24h.Serum concentration of amylase and lipase were detected. NO,NOS and myeloperoxidase (MPO) and expression of iNOS mRNA in the pancreas graft were measured. Histological examination was performed. Results The level of NO in the L-arg group was significantly higher than that in the control group(P

Objective To study the effects of cholecystokinin (CCK) on canine Oddi sphincter(SO)function after pancreas transplantation with bladder drainage. Methods Normal canine and transplant canine SO manometry before and after CCK administration was carried out. Results SO basal pressure in control group was(18 5?2 8) mm Hg, frequency was(9 7?1 5)per min, amplitude was (47?6) mm Hg, motility index was(236?56). After CCK administration, basal pressure, frequency, amplitude and motility index decreased significantly to(10 2?2 2) mm Hg ,(5 0?1 2)per minute,(19?5) mm Hg and(50?17), all P

Objective To explore the clinical application of laparoscopic splenectomy in treatment of spleen disease at our hospital.Methods We reviewed laparoscopic splenectomy carried out at our hospital since 1995,patients were grouped by date.63 laparoscopic splenectomies were divided into six groups.Operation time,intraoperative blood loss,postoperative hospital stay,time to feeding,days of drainage,amount of drainage,postoperative complications and indications for surgery were compared.Results Patient's age averaged at 44.19 years,body mass index averaged at 23.75,3 patients were converted to open surgery.Mean operating time,blood loss,postoperative hospital stay,time to feeding,converting rate in the 53 cases which had the surgery after 2003 were much better than the 10 cases before 2003.Surgical indications for laparoscopic splenectomy were limited to hematopoietic disease related splemegaly before 2003,the indication range significantly expanded after 2003,during which laparoscopic splenectomy were mainly applied to treat spleen tumors.Conclusions There is a marked learning curve in laparoscopic splenectomy after 10 surgeries before 2003 we have achieved the level.With the suitable approach,apparatus and skillful technique,laparoscopic splenectomy is safe and feasible to treat tumors of the spleen.