Objective To investigate the roles of somatostatin(SS)positive intemeurons in the development and compensation of temporal lobe epilepsy.Methods Piloearpine-induced epilepsy rat model was established.Immunohistochemistry method was used to detect number changes and axonal sprouting of SS positive intemeurons in different domains of the hippocampus at difierent time points.Degeneration of SS positive interneurons and their neurophils were detected by the double immunofluorescence staining with SS and Fluoro-Jade B(FJB)at 7 and 60 days after status epilepticus (SE).Results In the exoerimental rat group,the number of SS positive neurons decreased in each hippocampal domain,and it reached the lowest at 7 days post-SE(There were 11.1±3.3 in hilus,2.8±0.9 in CA1region and 1.8±0.7 in CA1region,t=13.519,9.644 and 8.808,all P<0.01).In chronic phase,the number of SS neurons gradually recovered,and exceeded the control group in CA1 area at 60 days post-SE(12.8±1.5 vs 8.8±1.3,t=-4.506,P<0.01),however,the number of SS neurons in the hilus(25.5±4.6)and CA1 area(4.8±0.8)remained significantly less than normal levels(t value were 4.691 and 3.953.both P<0.01).Increased SS positive fibers were found in the lacunosum-molecular (1m)layer and outer molecular layer of dentate gyrus after 30 days post-SE,and numerous SS positive fibers were seen threnghout the layers of area CA1 at 60 days post-SE.Double immunofluuorescence revealed that a few SS positive interneurons and fibers were also labeled by FJB in area CA1 at 7 days post-SE and in CA domain/hilus at 60 days post-SE.Conclusions SS intemeurons loss plays an important role in the development of temporal lobe epilepsy.The loss is partially caIlsed by the degeneration and death of neurons;SS positive neurophils increase within area CA1 in chronic phase may play a significant role in the generation and compensation of temporal lobe epilepsy.

Adult ; Breast Neoplasms ; metabolism ; pathology ; secondary ; Carcinoid Tumor ; metabolism ; pathology ; surgery ; Carcinoma, Adenoid Cystic ; pathology ; Carcinoma, Lobular ; metabolism ; pathology ; secondary ; Cervical Intraepithelial Neoplasia ; metabolism ; pathology ; surgery ; Chromogranin A ; metabolism ; Diagnosis, Differential ; Female ; Humans ; Hysterectomy ; Keratins ; metabolism ; Neoplasms, Multiple Primary ; metabolism ; pathology ; surgery ; Ovarian Neoplasms ; metabolism ; pathology ; Sex Cord-Gonadal Stromal Tumors ; metabolism ; pathology ; Synaptophysin ; metabolism ; Uterine Cervical Neoplasms ; metabolism ; pathology ; surgery

Objective To observe the influence of recombinant human thyrotropin(rhTSH)on serum concentration of endogenous thyrotropin(TSH), free triiodothyronine(FT3), free thyroxine(FT4), thyroglobulin antibody(TGAb), and thyroglobulin(Tg). To evaluate the efficacy of rhTSH-aided radioiodine treatment in patients with differentiated thyroid carcinoma(DTC). Methods The study recruitment took place between November 2007 and March 2009. 62 patients(including 45 females)with biopsy confirmed DTC had undergone total or nearly total thyroidectomy, and received 131I treatment. 31 patients(including 22 females), median age of 45 years(23-72), received radioiodine treatment 4 weeks after L-thyroxine(T4)withdrawal. The other 31 patients(including 23 females), median age of 44 years(14-70), underwent rhTSH-aided radioiodine treatment. Before and after rhTSH injection, serum TSH, FT3, FT4, TGAb, and thyroglobulin were tested. Post-radiotherapy whole body scan was performed 5 to 7 days after radioiodine treatment and qualitatively and blindly evaluated by two nuclear medicine physicians. Follow-up took place 6 to 12 months after radioiodine treatment. The efficacy of rhTSH-aided radioiodine treatment was evaluated by whole body scan with diagnostic dose radioiodine. SPSS 13.0 statistical software was applied. Results (1)Before and after rhTSH-aided radioiodine treatment, the serum TSH was(1.08±4.01)vs(140.26±27.20)mIU/L(P＜0.05), thyroglobulin(23.75±132.92)vs(169.58±178.49)μg/L(P＜0.05), FT3(4.52±1.16)vs(4.42±1.11)pmol/L(P＞0.05), and FT4(15.09±5.83)vs(13.66±5.85)pmol/L(P＞0.05),respectively.(2)rhTSH-aided radioiodine ablation treatment had the same effect as L-T4withdrawal aided. The complete response ratio was 77.4% vs 71.0%(P＞0.05)by radioiodine whole body scan of diagnostic dose. Conclusion rhTSH-aided radioiodine treatment of DTC was effective and safe, and did at least at equivalent degree as did L-T4withdrawal. Furthermore, Serum thyroglobulin level could be effectively stimulated by rhTSH with tumor relapse or metastasis.

Objective To study the potential effects of angiopoietin 1(Ang1) via adenovirus mediated gene transfer on ischemic heart disease in rabbits. Methods Forty-five male New Zealand white rabbits underwent high positioned double-ligation of the anterior descending left coronary artery, and were divided into three groups: Ang1 group (n=15) received direct myocardium injection of Ang1 recombinant adenovirus; DMEM group (n=15) and LacZ group (n=15) received only DMEM and LacZ recombinant adenovirus as controls respectively. The myocardial infarcted size was evaluated by N-BT macroscopic standing and the degree of angiogenesis was assessed by use of immunohistochemical analysis. The echocardiographic changes were measured on before operation and the 7 th, 14 th, and 28 th postoperative days. Results Animals in three groups had no significant difference in the percentage of infarction size of left ventricle at the 7 th, 14 th day and capillary density at the 7 th day. Animals in Ang1 group showed less infarcted size than DMEM group or LacZ group at the 28 th day and higher capillary density at the 14 th and the 28 th day. The results from echocardiographic measurements showed that animals in all three groups had no significant difference in the left ventricular systolic function before operation and at the 7 th , 14 th day, but the left ventricular systolic function in Ang1 group was better than that in DMEM group or LacZ group at the the 28 th day(P

Objective To explore the pathological changes by CT scan on localized thermochemotherapy dur- ing and after the operation of human gliomas.Methods Retrospective analysis was given to the CT scan of 37 pa- tients receiving thermochemotherapy during and after the operation,and the relation of the tumorous cells and mi- crovessels and CT density by EM were analyzed.Changes of tumorous cells and microvessels after localized ther- mochemotherapy on C_6 gliomas in rat were analyzed.Results When the tumor was low dense on CT pattern,less cellular number with increasing the amount of fluid between the cells was demonstrated pathologically.On EM,a lower cellular electron density was observed.The amount of fluid in cytoplasm was increased,the cytoplasm was porous,swelling denaturation was chiefly seen in organelle.If the tumor had mixed density on CT,cellular number was more,the amount of fluid was less.On EM,cellular electron density increased correspondingly,the fluid in cyto- plasm decreased,organdie was aggregated.After thermochemotherapy,the tumor reduced,liquefied,and vanished by CT scan.It could be observed that the tumorous cell become smaller,concentrated and cataclased,finally formed apoprotic bodies and separated from the cell in C_6 gliomas in EM.The tumorous vessels was less,smaller and thinker. Some vessels only could see the base membrane and no endothelioid cells.Conclusion The remaining tumors is van- ished by CT scan.The mechanisms of tumors disappearance proposes to explain that thermochemotherapy can dam- age C_6 glioma cells and microvessels,decrease microvessels density and induce tumor ceils apoptosis.That inhibits tu- morous angiogenesis and proliferation.

Animals ; Berberine/pharmacology* ; Blood Glucose ; Diabetes Mellitus, Experimental/drug therapy* ; Diabetes Mellitus, Type 2 ; Insulin ; Insulin Resistance ; Rats

OBJECTIVETo construct T vectors based on Xcm I recognition site and optimize the PCR fragments for its ligation.

METHODSWe firstly cloned the human histone H4 cDNA containing one Xcm I recognition site at both its 5' and 3' end into pCDNA 3.0 vector and then generated T vector with pCDNA 3.0 backbone by cutting the recombinant plasmid with Xcm I. To increase the ligation efficiency, the primers were firstly phosphorylated before DNA fragments amplification and then the PCR products were treated with Taq DNA polymerase and dATP after PCR amplification. Two DNA fragments with the length of 312 bp and 1 329 bp were ligated to it and the ligation mixture was transformed into E. coli DH5α competent cells and the positive rates of the transformants were evaluated by PCR and DNA agarose gel electrophoresis.

RESULTSOur results showed that the T vector produced by our method could ligate to the target DNA fragments with high efficiency. Besides, the phosphorylation state of the primers used for PCR amplification is also an important factor determining the cloning efficiency. What's more, as for longer DNA fragments, retreatment with Taq DNA polymerase and dATP after PCR amplification and purification could improve the ligation efficiency significantly.

CONCLUSIONOur protocol may overcome the dependence on blue/white screening to get positive clones and provide a potent way to generate T vectors and ligate them to the target PCR fragment.

Cloning, Molecular ; DNA, Complementary ; genetics ; Escherichia coli ; genetics ; Genetic Vectors ; Histones ; genetics ; Humans ; Polymerase Chain Reaction ; methods

OBJECTIVETo analyze the expression of Cyc1 in nasopharyngeal carcinoma (NPC) and evaluate the interfering efficiency of a lentivirus interfering vector targeting Cyc1 in NPC cells.

METHODSMicroarray technique was used to examine the expression of Cyc1 in NPC tissues. Real-time PCR was utilized to confirm the high expression of Cyc1 in NPC tissues and NPC cell lines. The recombinant Cyc1 shRNA-expressing plasmid (pLentiU6/Cyc1-shRNA) was stably transfected into NPC cells, and the interfering efficiency against Cyc1 was evaluated by quantitative RT-PCR.

RESULTSThe result of microarray showed that Cyc1 was highly expressed in NPC tissues compared to noncancerous nasopharyngeal tissues, as confirmed by Real-time PCR. All of the 8 NPC cells showed a high expression of Cyc1, among which 5-8F cells showed the highest expression. Sequence analysis indicated that the recombinant plasmid pLentiU6/Cyc1-shRNA was successfully constructed and could significantly and stably suppress the expression of Cyc1 in NPC cells.

CONCLUSIONCyc1 is highly expressed in NPC cells. The lentivirus vector constructed can markedly inhibit the expression of Cyc1 in NPC cells, which provides assistance in the investigation of the function and molecular mechanism of Cyc1 in NPC.

Carcinoma ; Cell Line, Tumor ; Cytochromes c1 ; genetics ; metabolism ; Genetic Vectors ; Humans ; Lentivirus ; genetics ; Nasopharyngeal Neoplasms ; metabolism ; Oligonucleotide Array Sequence Analysis ; Plasmids ; RNA, Small Interfering ; genetics

OBJECTIVETo establish a reliable model for drug screening and therapy by culturing rat femoral head and inducing cartilage degeneration quickly in vitro.

METHODSThe femoral heads from the same SD rats of two-month old were divided into control group and experimental group respectively. They were cultured with DMEM medium plus 10% fetal bovine serum or DMEM medium plus 10% fetal bovine serum plus 50 ng/ml IL-1β for three days. Femoral heads were fixed in 4% paraformaldehyde, decalcified, dehydrated, embedded in paraffin and cut into slices. Specimens were stained with Toluidine blue and Safranine O-Fast Green FCF. The protein expression levels of type II collagen, MMP13, Sox9 and ADAMTS5 were analyzed by immunofluorescence.

RESULTSBoth the Toluidine blue and Safranine O staining were pale in the margin of femoral heads which were stimulated with IL-1β for three days compared to that in control group. The Fast Green FCF staining was positive at the edge of the femoral head in experimental group, which indicated that cartilage became degenerated. The expression levels of both type H collagen and Sox9 were decreased significantly while the expression levels of MMP13 and ADAMTS5 were increased in experimental group.

CONCLUSIONThe model of cartilage degeneration is established by culturing and inducing the degeneration of the femoral heads quickly in vitro.

Animals ; Cartilage Diseases ; genetics ; metabolism ; Collagen Type II ; genetics ; metabolism ; Disease Models, Animal ; Femur Head ; metabolism ; Humans ; In Vitro Techniques ; Interleukin-1beta ; genetics ; metabolism ; Male ; Matrix Metalloproteinase 13 ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; SOX9 Transcription Factor ; genetics ; metabolism

The influenza A virus matrix protein2 gene(M2)which deleted transmembrane region was amplified by overlap extending PCR,and the multiepitope gene of hemagglutinin(HA)was PCR amplified with seven continuous synthesized segments by designing primer.The two gene segments were separately cloned into pMD18T vector to sequence analysis and prokarytic expression vector pET28a+ to construct the recombinant plasmid pET28a+M2dHA.The recombinant plasmid was transformed into E.coli BL21(DE3),and the high expression strain was obtained by screening monoclones.The recombinant protein existed as inclusion bodies,which accounted about 45% of the total cellular protein.The inclusion bodies were washed with 1% Triton X100 solution twice,and dissolved in 8 mol/L urea solution.The solution protein was purified by Ni+2 affinity chromatography,and refolded by dilution renaturation,then purified by Q Sepharose FF cation exchange column.The purity of the protein was over 90% by HPLC analysis.The result of Western blot showed it has good antigenicity and specificity.These results strongly supported for the further study of the broadspectrum influenza virus subunit vaccine.