Objective To construct and identify of a plasmid mediated high efficiency of RNA interference(RNAi) against Nogo-66 receptor(NgR).Methods After cloning of NgR by RT-PCR,the fragments were inserted into pcDNA3.1/CT-GFP-TOPO to produce plasmid expressing NgR-GFP fusion protein.Four pairs of oligonucleotide were designed according to NgR sequence and annealed.The resulting fragments were ligated into short hairpin RNA(shRNA) expressing plasmid.The plasmids expressing NgR-GFP fusion protein and shRNA were co-transfected into AAV-293 cells.RNAi efficiency against NgR was observed under fluorescent microscope and calculated by Western blotting.Results The interference efficiency of one sequence was above 90%.Conclusion A plasmid mediated high RNAi efficiency against NgR is constructed successfully.

AIM: This study was designed to investigate the protective role of hypobaric hypoxic pretreatment (HHPT) on hippocampal neurons in Babl/c inbred mice. METHODS: HHPT was produced by simulating a 7 000 m high altitude 2.5 h/d for 3 d. At 36 h after last time decompression, three subgroups consisted of both the control and pretreatment mice were subjected to the 12 000 m high altitude hypobaric hypoxia for 4 h, the severe ischemia produced by bilateral common carotid artery occlusion for 18 min, and the severe ischemia/hypoxia produced by permanently ligation of right lateral common carotid artery followed by a 8 000 m high altitude hypobaric hypoxia for 4 h, respectively. The extents of protection to hippocampal CA1 neurons by HHPT were evaluated by accounting the number of intact neurons between HHPT and control group. RESULTS: The results indicated that HHPT protected hippocampal neurons against severe hypobaric hypoxia, severe ischemia, and ischemia combined with hypobaric hypoxia. CONCLUSION: Hypobaric hypoxic pretreatment induces a delayed protection to hippocampal neurons against hypoxic and ischemic injuries.

AIM: This study was designed to explore the differentially expressed genes between hypobaric hypoxic delayed preconditioning (HHDP) and normal mouse hippocampus. METHODS: HHDP was produced by treating the animals at a 7 000 m high altitude for 2.5 h/d for 3 d. At 36 h after last time decompression, total RNA was isolated from hippocampus. cDNA was synthesized and amplified by SMART PCR. cDNA libraries of differentially expressed gene between HHDP and control hippocampus were constructed. 452 clones from forward (subtracted control from preconditioning) cDNA library and 74 clones from backward (subtracted preconditioning from control) one were screened by reverse Northern hybridization. RESULTS: Screening with subtracted probes, hybridization signal of 85 gene fractions decreased and that of 217 gene fractions increased by more than 2 times in HHDP hippocampus compared with control. Screening with unsubtracted probe, hybridization signal of 44 gene fractions decreased and that of 135 gene fractions increased by more than 2 times in HHDP hippocampus compared with control. Some of the clones had been sequenced. Analysis and comparison with the data of GenBank were performed. The results showed that mouse cytochrome C oxidase subunit 1, NADH dehydrogenase subunit 1 and 6, deleted in split-hand/split-foot 1 region (DSS1) and cDNA corresponding to clone IMAGE: 5251089 of mice cDNA library were increased in hippocampus of HHDP mice. cDNA corresponding to clone IMAGE: 3593193, mus musculus adult male olfactory brain cDNA and mus musculus bladder RCB-0544 MBT-2 cDAN were decreased in hippocampus of HHDP mice. CONCLUSION: Many genes expresses differentially in hippocampus of mice during HHDP. This may be one of the molecular mechanisms of HHDP.

AIM: This study was designed to explore the effect of hypoxia on expression of P450 2C11 mRNA in cultured rat astrocytes. METHODS: Cultured rat astrocytes were randomly divided into 4 groups: glutamate group (G), hypoxic group (H), hypoxia + glutamate group (H+G) and control (C). Cells of control group were exposed to nomorxic (95% air, 5%CO 2) condition,that of G and H+G were incubated with 100 ?mol/L L-glutamate ,then cells of H and H+G exposed to hypoxic conditions (5%CO 2, 95%N 2) at 37℃?Each group had five timepoints which included 0 h, 3 h, 6 h, 12 h, 24 h, respectively. Expression of mRNAs of P450 2C11 were detected with reverse transcription ploymerase chain reaction (RT-PCR). RESULTS: Expression of P450 2C11 mRNA did not change in H and C at all the above mentioned timepoints. While it showed marked and continuous increase from 6 h to 24 h in G and from 3 h to 24 h in G+H. P450 2C11 mRNA expression was significantly higher in G than both in H and C, and it was the highest of all groups in G+H. CONCLUSIONS: Glutamate and hypoxia+glutamate increased the expression of P450 2C11 mRNA in cultured rat astrocytes continuously in 24 hours. Hypoxia alone did not increase the expression of P450 2C11 mRNA but enhanced the effect of glutamate on expression of P450 2C11 mRNA in astrocytes, which is maybe one of the mechanisms of hypoxic-induced cerebral dilation.

Objective To discuss the efficacy of retroperitoneal laparoscopic dismembered pyeloplasty for pediatric ureteropelvic junction obstruction(UPJO). Methods Retroperitoneal laparoscopic dismembered pyeloplasty was performed on 24 patients with UPJO.The age of the patients ranged from 3 to 14 years(mean,7 years).The diagnosis was set up by ultrasonography,IVU,CT or magnetic resonance urography.All the patients had hydronephrosis,with 4 cases mild,10 cases moderate and 10 severe ones. Results All operations were completed laparoscopieally,without conversions to open surgery during operation.The mean operative time was 180 min(range 150 to 200).The mean blood loss was 60 ml(range 40 to 100)and the mean postoperative hospital stay was 7 d (range 5 to 9).All 24 patients were followed-up for 6 to 24 months(mean,1 4 months).There was no stricture at UPJ.Eighteen patients had complete resolution of hydronephrosis and 6 patients showed obvious alleviation in hydronephrosis. Conclusion Retroperitoneal laparoseopie dismembered pyeloplasty could be a mini-invasive,safe,and effective procedure for the treatment of UPJO in pediatric patients.

Proceeding from the requirements of teaching target for postgraduate students,the course of progress in pathophysiology was constructed and administrated.The course objective was defined which combined teaching knowledge with fostering students' ability together,especially the ability to think new ideas and to do scientific research.Aiming at this teaching target,the teaching contents which combined with the direction of scientific research of the department was growing together with scientific development,especially in new knowledge and new technique.Multiple teaching means and several mode of examine were adopted during the process of teaching practice.

Objective:To observe the effect of astragaloside Ⅳ on lysophosphatidic acid(LPA)- induced neurite retraction of N1E-115 cells and its potential mechanism.Methods:N1E-115 cells were divided into blank group, model group, the low, medium and high dose groups of astragaloside Ⅳ. The blank group and model group was not intervened by astragaloside; while the low, medium and high dose groups were treated with 20,40 and 80 μg/ml astragaloside Ⅳ for 24 h. Each group was cultured with serum-free medium for 12 h. The model group and astragaloside Ⅳ groups were intervened by 40 μmol/L LPA for 10 min. Each group was observed and photographed with the inverted microscope, and the number of neurites in N1E-115 cells was counted by Image J software. The fluorescence expression of recombinant ras homolog gene family member A (RhoA), rho associated coiledcoil protein kinase 2 (ROCK2), phospho-rho associated coiledcoil protein kinase 2 (p-ROCK2) and phospho-myosin light chain 2 (p-MLC2) proteins was detected by immunohistochemistry. Real-time fluorescent quantitative polymerase chain reaction was used to detect the mRNA expression levels of RhoA and ROCK2 ; the protein expression levels of RhoA, ROCK2, p-MLC2 and myosin light chain 2 (MLC2) were detected by Western blotting.Results:Compared with 20 μg/ml astragaloside Ⅳ group, the inhibition rate of neurite retraction in 40 and 80 μg/ml astragalosideⅣ groups increased ( P<0.05). Compared with model group, the average fluorescence intensity of RhoA, p-ROCK2, p-MLC2 in 20, 40, 80 μg/ml astragaloside Ⅳ groups and the ROCK2 average fluorescence intensity in 40 μg/ml astragaloside Ⅳ group were decreased ( P<0.05, P<0.01); the expression of RhoA mRNA (0.89±0.09, 0.41±0.01, 0.09±0.03 vs. 1.50±0.01) and ROCK2 mRNA (0.89±0.09, 0.14±0.01, 0.20±0.01 vs. 1.62±0.17) decreased in 20, 40, 80 μg/ml astragaloside Ⅳ groups ( P<0.05, P<0.01); the ROCK2 protein (0.75±0.06, 0.57±0.02, 0.66±0.01 vs. 1.08±0.02), p-MLC2 protein (1.72±0.03, 1.40±0.04, 1.29±0.03 vs. 2.19±0.11), MLC2 protein (1.13±0.02, 0.68±0.03, 0.75±0.03 vs. 1.60±0.03) in 20, 40, 80 μg/ml astragaloside Ⅳ groups and the RhoA protein (0.35±0.01, 0.40±0.03 vs. 0.57±0.08) in 20, 40 μg/ml astragaloside Ⅳ groups were decreased ( P<0.05, P<0.01). Conclusion:Astragaloside Ⅳ can prevent LPA-induced neurite retraction and promote damaged nerve regeneration. The mechanism may down-regulae the protein expression levels of RhoA, ROCK2, p-ROCK2, p-MLC2 and MLC2 in RhoA-ROCK2 signaling pathway, and inhibite nerve growth cone collapse.

To summarized the experiences from our basic experimental and clinical research on peritoneal dialysis. In the past 16 years, peritoneal fibrosis rat models and rabbit models of peritonitis were first established successfully in our laboratory in China. Peritoneal mesothelial cells were also separated and identificated. Besides, we assessed the biocompatibility of peritoneal dialysis fluid and analyzed the molecular mechanism of peritoneal mesothelial cell injury. We demonstrated the key role of transforming growth factor-beta1 (TGF-beta1), connective tissue growth factor (CTGF) and peroxisome proliferative activated receptor-gamma (PPAR-gamma) in the pathogenesis of peritoneal fibrosis, as well as their regulation of molecular mechanism. Furthermore, we transfected the plasmids encoding TGF-beta1-shRNA or pCTGF-shRNA into peritoneal cells and tissues by nanocarrier technologies. In clinical research, the positioning of peritoneal dialysis catheters, peritoneal dialysis treatment modalities and the prevention and treatment of its complications were studied. The characteristics and mechanism of solute transport in peritoneal dialysis was also explored.
Animals ; Connective Tissue Growth Factor ; metabolism ; Fibrosis ; physiopathology ; prevention & control ; Humans ; Kidney Failure, Chronic ; metabolism ; therapy ; Peritoneal Dialysis ; methods ; Peritoneal Dialysis, Continuous Ambulatory ; adverse effects ; Peritoneum ; pathology ; Rabbits ; Rats ; Retrospective Studies ; Tissue Adhesions ; physiopathology ; prevention & control ; Transforming Growth Factor beta ; metabolism