BACKGROUND:The hyperpoladzation-activated cyclic nucleotide-gated cation channel (HCN) gene is not increase the risk of induced cardiac arrhythmia,and can accept the modulation function of autonomic nervous system.Therefore,it is the first candidate gene for biological pacemakers.OBJECTIVE:To construct recombinant adenovirus vector containing human HCN4 gene and evaluate its transfection efficency in rat bone marrow mesenchymal stem cells (MSCs).DESIGN,TIME AND SETTING:The in vitro cytology-gene experiment was performed at the Lin Bai-xin Experimental Center,Second Hospital of Sun Yat-sen University from February to September 2008.MATERIALS:Ten SD rats were supplied by the experimental animal center of Sun Yat-sen University.Rlasmid pcDNA3.1-HCN4 containing target gene human HCN4,human embryo kidney 293 cells and Escherichia coll DH5α were preserved by our experimental center;Shuttle plasmid pShuttle-CMV containing green fluorescent protein gene and adenoviral backbone plasmid pAdxsi were bought from SinoGenoMax Co.,Ltd.METHODS:HCN4 cDNA segment was liberated from the cloning vector of pcDNA3.1-HCN4 via Hind Ⅲ+Xba Ⅰ,and subcloned into pShuttle-CMV,which was digested by I-Ceu Ⅰ +I-Sce Ⅰ double enzyme and subcloned into adenoviral plasmid to form recombinant adenovirus plasmid.Recombinant adenovirus plasmid was transfected into 293 cell lines by liposome,and the recombinant adenovirus AdHCN4 was packaged and transfected into rat MSCs.MAIN OUTCOME MEASURES:The identification of recombinant adenovirus plasmid vector,identification of recombinant adenovirus and its titration test;the transfection efficiency of recombinant adenovirus.RESULTS:Cloned sequence about 3.6kbp was obtained by Hind Ⅲ+Xho Ⅰ digestion after HCN4 cDNA segment was cloned into pShuttle-CMV.DNA sequencing results indicated that the clone location was correct.Recombinant adenovirus plasmid was cut into seven fragments while empty vector gained only six fragments after digested by Xho Ⅰ.The recombinant adenovirus was pathogenic to 293 cells after recombinant adenovirus plasmid was packaged in it.HCN4 cDNA (657bp) was amplified by PCR with virus titer of 2.5×10~(11) PFU/mL after transfected 293 cells with supematant.The efficiency of recombinant adenovirus infecting rat MSCs was about 90% when multiplicity of infection was 800.Rat MSCs expressed green fluorescence after transfection.CONCLUSION:The adenovirus vector with human HCN4 cDNA was established successfully,which can effectively transfect rat MSCs.

AIM:To research the characteristics of ventricular electrophysiology in right ventricular rapid pacing-induced congestive heart failure (CHF) dogs. METHODS:Dogs (n = 16) were randomly divided into 2 groups:the control (n = 7) and the CHF group (n = 9) induced by rapid right ventricular pacing at 240 pulse·min-1 for 4 to 5 weeks. The electrophysiologic parameters were evaluated by the technique of standard electric stimulation and monophasic action potential (MAP) recording. RESULTS:(1) Ventricular effective refractory period (VERP), ventricular MAP duration (MAPD90), ventricular late repolarization duration (VLRD) and intra- ventricular conduction time (IVCT) were prolonged by 26％ (P ＜ 0. 01), 43％ (P ＜ 0. 01), 318％ (P ＜ 0. 05), and 19％ (P ＜ 0. 01), respectively in CHF group. (2)The ratio of VERP to MAPD90(VERP/MAPD90) was decreased by 13％ (P＜0.05) in CHF group. (3) The dispersion of ventricular recovery time (VRT - D) was increased by 185％ (P ＜0. 01) in CHF group. (4) The ventricular fibrillation threshold (VFT) was decreased by 48％ (P ＜ 0. 01) in CHF group. CONCLUSION:The abnormal electrophysiological changes in the CHF condition may be contributing factors of lethal ventricular arrhythmias and sudden cardiac deaths in CHF.

Objective To learn the academic level and scientific impact of the Chinese Journal of Dermatology by using bihliometric methods. Methods Citation frequencies, first authors and their affiliation, citing journals of cited articles published in the Chinese Journal of Dermatology from 2002 to 2007 were statistically analyzed according to the database of Chinese Medical Citation Index (CMCI) by using bibliometric methods. Results From 2002 to 2007, a total of 2259 original papers were published in the Chinese Journal of Dermatology. Of them, 1224 were cited with a total citation frequency of 3963, citation rate of 54.1%, average citation frequency per paper of 3.2 and top citation frequency of 204. The number of first authors of cited papers amounted to 893, and 61 (6.63%) authors published 3 or more cited papers during this period. These authors were distributed in 30 provinces, municipalities, autonomous regions and 1 foreign country; Jiangsu, Beijing, Zhejiang, Guangdong, Shanghai and Shandong ranked in the top 6 regions with high total citation frequencies. There were 517 journals citing articles published in the Chinese Journal of Dermatology from 2002 to 2007;the top citing frequencies were observed in the Chinese Journal of Dermatology, China Journal of Leprosy and Skin Diseases, The Chinese Journal of Dermatovenereology, Journal of Clinical Dermatology and Southern China Journal of Dermato-Venereology, etc. Conclusions The articles in the Chinese Journal of Dermatology have a high academic quality and citation frequency. Chinese Journal of Dermatology is an important publication in the dermato- venereology field as well as a core periodical in medical field of China.

AIM:To study the effects of FK506 binding protein 12.6(FKBP12.6) overexpression on the performance of sarcoplasmic reticulum(SR) in ventricular myocytes of rats with heart failure.METHODS:The adenovirus(Ad)-mediated gene transfer was used to overexpress FKBP12.6 in ventricular myocytes of rats with heart failure.Western blotting and reverse transcriptase-polymerase chain reaction(RT-PCR) analysis were used to reveale specific overexpression of FKBP12.6.X-rhod-1-AM was used as the Ca2+ indicator,and cells were viewed under a confocal microscope.RESULTS:Adenovirus mediated overexpression of FKBP12.6 resulted in a 5-fold increase in relative FKBP12.6 mRNA levels and a 4-fold increase in relative FKBP12.6 protein levels at 48 h after transfection compared with control.The amplitude of the fluorescence [Ca2+]i transient was significantly increased in Ad-FKBP12.6-GFP cardiomyocytes compared with Ad-GFP myocytes(peak F/F0,3.16?0.42 vs 1.43?0.38,P

To study the bioequivalence of a kind of progesterone sustained release suppository, a randomized cross-over study was conducted in 12 rabbits. A single rectal dose of 2.75 mg/kg progesterone sustained released suppository (tested formulation, T) and progesterone suppository (reference formulation, R) was administered; a multiple dose of 2.75 mg/kg was given up to seven times with an interval of 8 h. Concentrations in serum were determined by a competitive enzyme immunoassay. The main parameters of T were: for single and multiple doses, Cmax was 48.8 +/- 11.8 ng/mL and 43.5 +/- 9.4 ng/mL, Tmax was 0.5 +/- 0.3 h and 0.4 +/- 0.3 h, AUC(0-24 h) was 362.4 +/- 143 ng x h x mL(-1) and 310.6 +/- 70.3 ng x h x mL(-1), respectively. The relative bioavailability of T to R were (104.2 +/- 13.4)% and (111.4 +/- 19.1)%, respectively. Statistical analysis showed that the two formulations were bioequivalent and T had sustained released feature.
Administration, Rectal ; Biological Availability ; Cross-Over Studies ; Progesterone/administration & dosage ; Progesterone/*pharmacokinetics ; Random Allocation ; Suppositories

Objective To identify the imbalance of T cell-specific transcription factors T-bet/GATA-3,and to explore the modulation with dexamethasone and imiquimod in CD4+T cells from ovalbumin (OVA)sensitized mice.Methods CD4+T cells were obtained fromsingled-cell suspension of spleen(after lysis of RBC).ELISA assay was used to detect the concentrations of IL-4,IL-5 and IFN-γin superna tants and cell pellets,and the expression of T-bet and GATA-3 was detected by Western blot.Resuits In the control group,tIle low levels of IFN-γ were detected in the supernatants during 24 h.In OVA treatment group,the concentrations of IL-4,IL-5 were increased significantly,and the concentrations of IFN-γ were always low in the supernatants.In the dexamethasone treatment group,the concentrations of IFN-γ,IL-4 and IL-5 were all low in the supernatants during 24 h.In the imiquimod treatment group,the concentrations of IFN-γ were increased significantly,and the concentrations of IL-4 and IL-5 were decreased in the super natants.It worked at 6 h,and achieved the peak at 12 h,lasted over 24 h.In the control group,the expres sions of T-bet and GATA-3 were detected in CD4+T cells during 24 h.In OVA treatment group,the expressions of T-bet were decreased,and that of GATA-3 were increased rapidly in CD4+T cells.In dexam ethasone treatment group,the expressions of T-bet were always low in CD4+T cells,and that ofGATA-3 were no change during 24 h.In imiquimod treatment group,the expressions of T-bet were increased,andthat of GATA-3 were decreased in CD4+T cells.The protein expressions worked at 6 h.and achieved the peak at 12 h,lasted over 24 h.Conclusion The imbalance T cell-specific transcription factors T-bet/GA-TA-3 contributes to both high expression of GATA-3 and low expression of T-bet in CD4+T cells from OVA sensitized mice.Dexamethasone treatment inhibits the expression of T-bet in CD4+T cells and has no func tion in GATA-3.Imiquimod treatment modulates key master switches GATA-3 and T-bet that results in com mitting T helper cell to a TH 1 phenotype and imiquimod may play a key role in the regulation of TH2 cytokine responses in asthma.

Objective To evaluate the effect of preoperative transarterial chemoembolization (TACE) for resectable hepatocellular carcinoma (HCC).Methods HCC patients undergoing up-front hepatectomy (group A) were compared with those receiving TACE before hepatectomy (group B).Results Tumor size decreased significantly after TACE (t =3.3 1,P =0.021).The rates of tumor encapsulation and liver adhesions were significantly more often seen in group B.There were fewer tumor-residual and more frequent necrosis in group B.Operative time in group B was longer (t =2.71 ,P =0.046).The average blood loss and complication rate were of no difference between the two groups.The occurrence of pleural effusion and intrahepatic recurrence rate in group A was higher than group B (x2 =3.85 ,P =0.031) (x2 =2.76,P =0.046).The overall survival rate from the second year postoperative in group B was higher than group A (x2 =3.37, P =0.043).Conclusions TACE could diminish tumor, advance encapsulation and reduce tumor-residual.Preoperative TACE does not improve 1-, 2-, and 3-year tumor-free survival rates but improve 1-, 2-, and 3-year overall survival rates.

Objective To evaluate the short-term and long-term elinical effect of hepatectomy using Tissue-Link & Cusa,compared to the Pringle maneuver.Methods Clinical data of 87 HCC patients who had received hepatectomy by the Pringle's Maneuver (group A) or Tissue-Link & Cusa (group B) were retrospectively analyzed.Results The average amount of bleeding in Group A was more than group B (t =2.030,P =0.023).The time of operation in group A was shorter than group B (t =-2.896,P =0.006).The postoperative supplement of albumin in group A was more than group B,the level of serum total bilirubin on 7th day after operation was higher than group B (P ＜ 0.05).There was no significant difference in postoperation complications and the time in hospital (P ＞0.05).The rate of incisional recurrence and the rate of metastasis in or out of the liver in group A were higher than group B (P =0.029,0.021,0.016).The 2-and 3-year tumor-free survival rates and the 3-year overall survival rates in group A were lower than that in group B (P =0.047,0.036,0.042).Conclusions Hepatectomy using Tissue-Link & Cusa is superior to the Pringle's maneuver for the treatment of primary hepatocellular carcinoma.It has a clear operative field,less operative bleeding,less damage to liver function and a lower relapse rate of incisional margin.

Objective To explore the protective effect of folic acid supplementation in different ways on lipopolysaccharide-induced external malformations and skeletal malformations in mice.Methods The pregnant mice were divided into six groups randomly,including saline control group,FA control group,LPS group,LPS+FAig group,LPS+FAip group,and LPS+FAdw group.All dams were medicated on gestational day 8～12,and then sacrificed on gestational day 18,for each litter,the number of live fetuses,dead fetuses,resorption fetuses and external malformations of live fetuses were all counted.All fetuses were subsequently evaluated the skeletal malformations.Results A low dose of LPS injection during the second trimester resulted in external and skeletal malformations.Exencephaly and encephalomeningocele were two of the most common external malformations,skeletal malformations consisted mainly of the incompletion of supraoccipital ossification,sternal malformation and rib malformation.FA supplementation by three ways all attenuated the external and skeletal malformations.And the best protective effect was by oral administration.The incidence of external,sternal and rib malformation was all decreased.Conclusions Three ways can prevent the mice during the second trimester from the external and skeletal malformations caused by LPS injection,and the best protective effect was by intragastric administration.

Objective To evaluate the effect of chronic low potassium on K+uptake rate in the my?ocardium and skeletal muscle of rabbits. Methods Thirty?two adult male rabbits, aged 12-14 weeks, weighing 2?0-2?7 kg, were randomly divided into 4 groups ( n=8 each) using a random number table:normal feeding group ( group N) , low potassium feeding group ( group L) , potassium supplementation con?trol group ( group SC ) and potassium supplementation experimental group ( group SE ) . N and SC groups were given a normal diet only, and L and SE groups were fed with a low potassium diet for 15 days. Potassi?um chloride ( KCl) 0?5 mol∕L was then infused intravenously at the initial rate of 60 μmol·kg-1 ·min-1 in SE and SC groups. Blood samples were obtained from the central artery of the left ear every 5 min for meas?urement of plasma K+ concentrations. The infusion rat of KCl was then adjusted until the plasma K+concen?tration reached 5?5 mmol∕L and maintained at this level for 1 h, and then infusion was stopped. The total volume of KCl infused was recorded. The hearts and soleus muscle of animals were excised for determination of K+content. K+uptake and uptake rate were calculated. Results Compared with N group, the plasma K+concentration, and K+content in the myocardium and soleus muscle were significantly decreased in group L ( P<0?05) . Compared with SC group, the total volume of KCl infused, and K+uptake and uptake rate in the myocardium and soleus muscle were significantly increased in group SE ( P<0?05) . Conclusion Chro?nic hypokalaemia can increase K+ uptake rate in the myocardium and skeletal muscle of rabbits.