Objective To clone and identify neurotrophic factor NT3 gene from mouse liver so as to establish a cell strain expressing high level of NT3 gene. Methods The target genes amplified by RT PCR were cloned into the shuttle vector pExchange 1neo, and then the DNA sequence was analyzed by enzyme digestion and sequencing. The recombinant expression vector pExchange NT3 1neo was employed to transfect the embryonic stem cell strain MESPU35, and then the transfected cells were selected by G418. The NT3 level in the transfected cells was detected by RT PCR. Results A gene fragment of 777 bp was obtained by RT PCR, and the DNA sequence was identical to mice NT 3 gene sequence of GenBank. The recombinant vector was constructed successfully and the constructed cell strain could express high level of NT 3 gene. Conclusion The successful cloning of NT3 gene from mouse liver, construction of pExchange NT3 1neo expression vector, and establishment of cell strain stably expressing high level of NT3 gene have laid a foundation for the further studies.

Objective To investigate the effectiveness and technical points of anatomical liver resection by trans-Glisson Sheath methylene blue staining in treatment of hepatolithiasis of right posterior lobe.Methods The clinical data of 12 cases of hepatolithiasis of right posterior lobe treated with anatomical liver resection by Glisson sheath methylene blue staining were retrospectively analyzed.Result 6 of 12 patients had undergone more than 2 previous biliary surgeries.All patients underwent contrast-enhanced CT scan and portography,hepatolithiasis of segment Ⅵ in 4 cases,right posterior lobe in 8 cases,accompanied by left lateral lobe bile duct stones in 2 cases,the right caudate lobe bile duct stones in 1 case.Methylene blue was injected into the portal vein,the methylene blue interface of segment Ⅵ or right posterior lobe displays well.Methylene blue interface was larger than the ischemia interface,which is in accordance with the anatomy.Along the methylene blue interface,hepatic resection was performed including right posterior lobe resection (n =9),segment Ⅵ resection (n =3),and combined with left lateral lobe resection (n =2) and the right caudate lobe resection (n =1).There was no postoperative mortality.Incision infection occurred in 5 cases,4 had right pleural effusion and 2 had a biliary fistula that were treated conservatively.With a mean follow-up period of 3.2 years,all patients are symptoms free and stone free.Conclusions Anatomical liver resection by methylene blue staining is a safe and effective treatment for hepatolithiasis of right posterior lobe.

Familial adenomatous polyposis is characterized by the multiple and adenomatous polyps in the colorectum combined with polyps in the stomach and duodenum, while it is rarely seen in the common bile duct (CBD).In July 2013, 1patient with FAP combined with adenomas in the CBD was admitted to the Fujian Provincial Hospital.The patient underwent laparoscopic CBD exploration and resection of masses due to acute pancreatitis 11 months ago, and was confirmed as with adenoma in the distal CBD by postoperative pathological examination.Multiple polyps were found in the stomach, duodenum,CBD, colorectum after admission to hospital, biopsy confirmed that polyps were tubular adenoma.The patient received pancreaticduodenectomy and was diagnosed as with duodenum-CBD tubular adenoma in postoperative pathological examination.The patient was followed up by telephone interview and outpatient examination and had a full recovery, in addition to the increasing of stools frequency and occasioned hematochezia, and then was treated by total colectomy at postoperative month 7.The tubular adenoma was confirmed by postoperative pathological examination.

Objective To provide anatomical information and clinical application of flaps based on the proximal ulnar artery perforators.Methods Ten fresh cadavers who underwent injected with artery imaging technology and dissected with layer by layer;Eighteen patients who sought surgical treatment with proximal ulnar artery perforator flap for soft tissue defects of the finger and dorsum wrist at our hospital between October 2011 and November 2012 were included in this study.Results The diameter and superficial length of the main perforator respectively were 0.5-0.9 mm and 33.0-47.0 mm in our dissection.There were 5-9 perforators given from the ulnar artery to supply skin over the medial side of the forearm.All of the 18 flaps survived after surgeries.The flap size ranged from 3.0 cm × 2.5 cm to 10.0 cm × 5.5 cm.All of the transplanted flaps presented favourable contours and good functions at 6 to 12 months' followed-up.Conclusion Proximal ulnar arter perforator flap has favourable appearance,constant vascular pedicle,reliable blood supply,and large diameter.The free transplantation of this flap offers a satisfactory alternative for repairing the small and medium-sized area of soft tissue defects of forearm and hand.

Objective To observe the effects of ( - )-epigallocatechin-3-gallate (EGCG) on human prostate cancer xenografted tumor growth and connexin43 expression in nude mice,and explore the mechanism of the EGCG on prevention for prostate cancer.Methods The methyl thiazolyl tetrazolium and annexin-V/PI double-labeled flow cytometry methods were used to observe the growth inhibiting rate (IR)and apoptosis rate (AR) of human prostate cancer cell line PC-3 which was treated by EGCG at different concentration (10,20 and 40 mg/L,respectively).The scrape-loading fluorescence dye transfer method was applied to assess the gap junction intercellular communication (GJIC) through fluorescence microscope.PC-3 cells were subcutaneously transplanted to establish tumor-bearing nude mice model.A total of 32 mice were randomly divided into four groups,both control group and three treatment groups were treated with different doses of EGCG ( 10,20 and 40 mg/kg,respectively).After two weeks,the mice prostate tumor tissues were taken out.The tumor wet weight was measured and tumor growth inhibiting rate was calculated.The tumor microvascular density (MVD) and apoptosis index (AI) were detected by the immunohistochemical techniques and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling techniques,respectively.Semi-quantitative reverse transcription polymerase chain reaction was used to examine the expression level of the Cx43 mRNA.Results EGCG at concentration ( 10 and 20 mg/L) could significantly inhibit the proliferation[(22.33 ±4.62)％,(38.67 ±5.67)％ vs (3.47 ±0.31 )％,P ＜0.01],induce the apoptosis [(7.84 ± 1.37 ) ％,( 24.53 ± 2.28 ) ％ vs ( 2.17 ± 0.70 ) ％,P ＜ 0.01] and enhance the GJIC of PC-3 cells.EGCG of different doses could inhibit prostate cancer xenografted tumor growth,induce tumor cells apoptosis and inhibit angiogenesis.EGCG ( 20 and 40 mg/kg) could effectively up-regulate Cx43 mRNA expression in xenografted tumor (0.58 ± 0.08,0.80 ± 0.07 vs 0.42 ± 0.04,P ＜ 0.0 ).The effects had significant correlation with the dose-dependent of EGCG ( P ＜ 0.05 ).Conclusions EGCG could up-regulate the Cx43 expression and enhance the gap junction intercellular communication mediated by Cx43 in the prostate tumor,which provide the experimental evidence for the mechanism of its effectively inhibiting the prostate cancer growth.

Objective To investigate kupffer cells (KCs) expressing indoleamine 2,3-dioxygenase(IDO)in the inhibition of allogeneic T-cell proliferation in vitro. Methods Real-time PCR was used to investigate the expression of IDO mRNA and FasL mRNA in KCs pretreated with or without IFNγ. High performance liquid chromatography was used to analyze the catabolism of tryptophan by IDO from KCs. Allogeneic T-cell response was used to confirm the inhibition of KCs in vitro. The proliferation of lymphocytes was detected using [3 H] thymidine incorporation. Cell cycle and lymphocyte apoptosis were evaluated by flow cytometric assay. Results Real-time PCR revealed IDO mRNA and FasL mRNA expression in KCs pretreated with IFN-γ. IDO catabolic effect was confirmed by a decrease in tryptophan and increase in kynurenine concentration. KCs expressing IDO and FasL from BABL/c mice acquire the ability to suppress the proliferation of T-cells from C57BL/6, which could be blocked by the addition of 1-methyl-tryptophan and anti-FasL antibody. The co-cultured T-cells with KCs expressing IDO and FasL could induce allogeneic T-cell apoptosis and exhibited cell-cycle arrest in G1. Conclusion In addition to the Fas/FasL pathway, IDO may also play an important role in KCs to inhibit allogeneic T-cell proliferation in vitro.

Objective To investigate the role of TLR4/NF-κB signal pathway in pathogenesis of brain inju-ry during deep hypothermia circulatory arrest(DHCA). Methods BV2 microglia cells were subjected to oxygen-glucose deprivation/reoxygenation(OGD/R),in vitro model for DHCA. The BV2 were randomly divided into the control group(C group)and the experimental group(O group). BV2 viability was determined by CCK-8 assay. TLR4 and its downstream signaling molecules ,MyD88 and phosphorylated NF-κB (p-p65) expressions were detected by Western blotting. TLR4 mRNA expression in BV2 microglial cells were determined by RT-PCR. Level of interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) in culture medium was detected by ELASA. Results Compared with the group C,BV2 microglia cell viability in experiment group was obviously weaker(P<0.05). Expressions of TLR4,MyD88 and phosphorylated NF-κB(p-p65)from the experiment group increased remarkedly than those from the group C (P < 0.05). TLR4 mRNA level was higher significantly in the group O than in the group C (P < 0.01). Production of IL-6 and TNF-α in the group O were up-regulated apparently compared to the group C(P<0.01). Conclusion TLR4/NF-κB signaling pathway contributed to activation of BV2 microglia cells treated by OGD/Reoxygenation ,which was probably the exactly way that involved in pathogenesis of brain injury during deep hypothermia circulatory arrest.