Objective To summarize the experiences of the treatment of extrahepatic bile duct injury caused by cholecystectomy.Methods A retrospective analysis was made on the treatment of 11 cases of extrahepatic bile duct injuries in our hospital from 1989 to 2001.The position of injuries,types,the time from onset to diagnosis, operation methods and therapeutic effects were analyzed.Results Three of the 11 cases received an emergency operation,while 8 underwent selective operation.Seven cases (62.7%) of extrahepatic bile duct injury were diagnosed during the cholecystectomy,and the remaining 4 cases (37.3%) were diagnosed after operation.Seven cases of extrahepatic bile duct injury diagnosed during operation were treated immediately and 4 cases treated 3-4 days after the first operation.All the patients were cured and discharged.Conclusion The correct principles and methods of handling the cases of extrahepatic bile duct injury caused by cholecystectomy should be mastered;Roux-en-y cholangiojunostomy is the best option for the treatment of extrahepatic bile duct injury.

Objective To study the inhibitory effect of human antisense high mobility group box 1 (HMGB1) gene on the growth of human pancreatic adenocarcinoma cell line (PANC-1).Methods HMGB1 was cloned and reversely inserted into eukaryotic expression vector pcDNA3.1 to get pcDNA3.1/anti-HMGB1, which was then transfected by liposome method into PANC-1 cells.After G418 selection, HMGB1 expression of PANC-1 cells before and after transfection was detected by RT-PCR and Western blot, the cell proliferating activity in vitro was examined by MTT analysis, and cell cycle and apoptosis were assessed by flow cytometry.Results pcDNA3.1/anti-HMGB1 vector was obtained.After transfection with pcDNA3.1/anti-HMGB1, HMGB1 gene expression of PANC-1 cells was inhibited in both mRNA and protein level, and the proliferation of cultured pancreatic adenocarcinoma cells was suppressed with a suppression rate higher than 40 % on the sixth day.The cells of G1 phase increased obviously while the cell number of S phase decreased, the number of apoptosic cell increased.Conclusion Antisense HMGB1 gene can inhibit the proliferation of human ancreatic adenocarcinoma cells and increase the cell apoptosis rate.

Through ethical analysis of the conflicts appearing in four special cases in the four steps of assisted reproductive technology implementations, this paper pointed out the measures to solve them in the perspective of le-gal protection, standardized management, ethical supervision, and self-regulation for making the research and ap-plication of assisted reproductive technology serve patients better and promoting the healthy development of assisted reproductive technology.

Objective To investigate the possibility of transforming growth factor β (TGF-p) inducing chondrogenesis of precartilaginous stem cells (PSCs) and discuss expression mechanism of extracel-luar matrix. Methods PSCs were induced into a chondrogenic pathway in alginate bead culture in the absence of serum and in the presence of TGF-β1, β2, or-β3. The temporal pattern of expression of cartilage-specific extracellular matrix during chondrogenesis were analyzed by immunocytochemistry, immunoflu-orescence, RT-PCR, immunoprecipitation, Western blot and spectrophotometer. Results Type Ⅱ collagen staining was positive at days 7, 14 and 21 in alginate bead culture, showing most intense staining in the TGF-p3-treated culture. Expression of type Ⅱ collagen was increased in TGF-β3 group. Immunocytochemi-cal analysis of a number of other extracellular matrix components showed widespread expressions of aggre-can, fibromodulin and COMP in alginate bead culture that presented TGF-p3 for 21 days. The expressions of Aggrcan, fibromodulin, type Ⅰ and ⅹ collagen, and COMP were detected by RT-PCR in TGF-β3 group within 8 days, while type Ⅱ collagen began expression at days 8-21. COMP or type X collagen was present in TGF-β3 group at days 7, 14 and 21 by immunoprecipitation or Western blot analysis respectively. The extracted glycosaminoglycan content or the glycosaminoglycan/DNA rate in TGF-βl group was significantly lower than those in TGF-β2 group or TGF-β3 group at days 14 and 21 (P <0.01). Conclusions TGF-β can evocate chondrogenesis of PSCs, when rapid deposition of cartilage-specific extracellular matrix is involved. The sequential events in this pathway leading from the undifferentiated stem cells to mature chon-drocytes can be investigated by analysis of key matrix elements.

BACKGROUND: Deletion of glial cell-derived neurotrophic factor and endothelin receptor type B gene will induce abnormal development of enteric nervous system. Neural stem cell transplantation can repair nervous system from anatomy and function,and be considered as a vector of gene transfection.OBJECTIVE: To transfect recombinant adenovirus carrying glial cell-derived neurotrophic factor and endothelin receptor type B gene into mouse neural stem cells, and to observe expression of target gene.DESIGN: A cell-gene study.MATERIALS: New-born Kunming mice were provided by the Animal Center of Tongji Medical College, Huazhong University of Science and Technology, China. jetPEI reagent was purchased from PolyPlus Co, France. The pAdTrack-CMV-GE with green fluorescent protein (GFP) was gifted by Doctor Sun Nianfeng and Zhang Jinghui in our laboratory.METHODS: Neonatal mouse brain tissues were sterilely obtained to prepare monoplast suspension. Adenovirus expressing glial cell-derived neurotrophic factor and endothelin receptor type B gene with GFP was dissolved in NaCI to prepare JetPEI/DNA complex. Subcultured neural stem cells in DMEM/F12 were regulated to 5×10~8/L, and 400 μL cell suspension and 100 μL JetPEI/DNA complex were seeded on a 24-well plate at 37 ℃ in 5% CO_2 incubator. Neural stem cells were harvested at 24, 48 and 72 hours following transfection.MAIN OUTCOME MEASURES: The efficiency of transfection was detected using fluorescence microscope and flow cytometry.Target gene expression in neural stem cells was determined using RT-PCR.RESULTS: Bright green fluorescence of the transfected cells could be observed under fluorescence microscope after 24 hours of transfection. The positive rate of GFP was 15.36%, 24.67%, 25.73% at 24, 48 and 72 hours following transfection respectively.Neural stem cells expressed glial cell-derived neurotrophic factor and endothelin receptor type B gene at various time points.Strap brightness was low at 24 hours, and exogenous gene expression was great at 72 hours.CONCLUSION: The target genes were successfully transfected into neural stem cells by using jetPEI reagent. Moreover, glial cell-derived neurotrophic factor and endothelin receptor type B gene effectively transcribed and expressed in target cells.

BACKGROUND:Magnetic resonance diffusion tensor imaging can display the dispersion changes of peripheral nerve injury and be used to conduct quantitative research, so it has good application prospects in displaying the nerve injury and regeneration. OBJECTIVE:To investigate the possibility of magnetic resonance diffusion tensor imaging of rabbit acute sciatic nerve traction injury, and to figure out the value of diffusion tensor parameters in the diagnosis of peripheral nerve injuries and to reveal the pathologic basis. METHODS:The right hind limb sciatic nerves of 32 New Zealand white rabbits were selected to make the regeneration and repair models, the left hind limb nerves as the sham-operation side. Diffusion tensor imaging examination of sciatic nerves were performed at 1 and 3 days, 1, 2, 3, 4, 6 and 8 weeks after operation with 1.5 T MRI. Fractional anisotropy and apparent diffusion coefficient were measured through diffusion tensor tracing
reconstruction, and then the pathological examination was performed. RESULTS AND CONCLUSION:Diffusion tensor imaging revealed only the proximal nerve, injured nerve as wel as the middle of the distal nerve at 1 day after traction injury. At 1 week, the nerve of distal portion appeared thinner and shorter fiber bundle. At 2-6 weeks after operation, the fiber bundle was increased and thickened. At 8 weeks after operation, the distal nerve fibers had nearly restored to the level before injury. There was significant difference in the fractional anisotropy value of traction portion and distal portions between traction injury and sham-operation group at 1 day-8 weeks after operation (P<0.05). While there was significant difference in the fractional anisotropy value of proximal traction portion between traction injury and sham-operation group 1 day-1 week after operation (P<0.05). There were no significant differences in the apparent diffusion coefficient values between traction injury and sham-operation group at 1 day-8 weeks after operation. Fal of fractional anisotropy value in the early stage of nerve traction injury was the result of myelin sheath broke down and axonal disintegrated;recovery of fractional anisotropy value resulted from myelin sheath proliferated and myelin sheath grew slowly to mature. Diffusion tensor tracing can show the abnormal change of the sciatic nerve with traction injury in rabbit clearly and early, and the measurement of fractional anisotropy value can be used as the sensitive method to monitor the degeneration and regeneration after nerve traction injury.

AIM: To compare the effects of hypoxia on expression of vascular endothelial growth factor (VEGF), iNOS and eNOS mRNA in cultured umbilical vein endothelial cells (UVECs) obtained from Tibetan and Han. METHODS: UVECs were obtained from native Tibetan and immigrant Han, respectively and cultured under hypoxia conditions (0.5% oxygen) for 2 h, 4 h, 12 h, and 24 h and normoxic conditions. VEGF, iNOS and eNOS mRNAs were detected with methods of RT-PCT. RESULTS: VEGF and iNOS mRNAs were up-regulated while eNOS mRNA depressed by hypoxia similarly in Tibetan and Han UVECs. CONCLUSION: Our results suggest that the changes of VEGF, iNOS and eNOS mRNA expression are common pathways in the mechanisms of hypoxic responses.

BACKGROUND:Studies have shown that human telomerase reverse transcriptase (hTERT) has the ability to enhance cel proliferation, maintain telomere length, prolonged cel life cultured in vitro. OBJECTIVE:To observe the effect of hTERT gene-modified bone marrow mesechymal stem cel transplantation on neural function recovery of rats with cerebral infarction. METHODS:Rat models of middle cerebral artery occlusion were established and randomized into model group, cel transplantation group and hTERT-modified cel transplantation group, with 20 rats in each group. Rats in the three groups were respectively injected via tail vein with 1 mL PBS, passage 9 bone marrow mesenchymal stem cel suspension (2.5×107/L) and hTERT-modified passage 9 bone marrow mesenchymal stem cel suspension (2.5×107/L), respectively. Modified neurological severity scores were determined before and after transplantation; RT-PCR and western blot assay were used to measure hTERT expression at gene and protein levels; TUNEL method was adopted to detect cel apoptosis in the brain. RESULTS AND CONCLUSION:hTERT-modified bone marrow mesenchymal stem cels had prolonged cel cycle, and with the increase in passage number, the cels showed good growth with no changes in morphology. The expressions of hTERT mRNA and protein were superior in the hTERT-modified cel transplantation group than the cel transplantation group, and there was a significant difference (P < 0.05). Modified neurological severity scores and number of apoptotic cels were decreased significantly in the hTERT-modified cel transplantation group compared with the other two groups (P < 0.05). These findings indicate that hTERT-modified bone marrow mesenchymal stem cels can promote neural functional recovery of rats with cerebral infarction.

BACKGROUND:Numerous clinical studies have confirmed that the microenvironment at a spinal cord injury site can be obviously improved through hyperbaric oxygen therapy; however, what effect does hyperbaric oxygen have on the microenvironment of the injured brain? OBJECTIVE:To investigate the effect of hyperbaric oxygen therapy on nerve regeneration microenvironment and the recovery of rat nerve function after focal cerebral infarction. METHODS:Rat models of focal cerebral infarction were established by occlusion of the middle cerebral artery and subjected to hyperbaric oxygen therapy. Sham group and model group were established as comparison. In the sham group, rat models of focal cerebral infarction were established but did not receive any treatment. Rats in the model group were placed in a hyperbaric oxygen therapy chamber but the pressure and oxygen concentration were not administered. RESULTS AND CONCLUSION:Compared with the model group, the score of rat limb function at 16 days after treatment and the expression of growth associated protein 43 in the rat cerebral infarcted area at postoperative 14 days were significantly increased , but infarct volume at postoperative 24 hours was al significantly decreased in the hyperbaric oxygen therapy group (alP < 0.05). These results confirmed that hyperbaric oxygen therapy can improve nerve regeneration microenvironment and promote the recovery of rat nerve function after focal cerebral infarction.

To increase the utilization rate of expanded criteria donor (ECD) kidney, the kidney preservation methods have been ever advancing in recent years. The application of normothermic machine perfusion (NMP) promotes the preservation, evaluation and repair of ex vivo donor kidneys and accelerates the innovation of surgical approaches of kidney transplantation. Ischemia-free kidney transplantation (IFKT), which initiated by Organ Transplantation Center of the First Affiliated Hospital of Sun Yat-sen University, keeps the blood flow and oxygen supply of the donor kidney with NMP machine during the entire process of acquisition, preservation and transplantation, thereby fundamentally avoiding ischemia-reperfusion injury (IRI) of the donor kidney and reducing the risk of delayed graft function (DGF) and acute rejection after surgery. In this article, recent progresses upon the kidney NMP, surgical procedures and short-term outcomes of IFKT were reviewed, aiming to provide reference for enhancing the utilization rate of ECD donor kidney and resolving the issue of organ shortage.