Objective To prepare the recombinant epitopes of HSV-gB and HSV-gD protein and provides a new antigen protein for the development of herpes simplex virus(HSV)vaccine.Methods The epitopes of HSV-gB and HSV-gD protein were analyzed by epitope prediction software.A novel gene named X which encoded 9 predicted epitopes of HSV-Gb and HSV-gD protein was designed and synthesized using chemical method.X gene was cloned into vector PET-28a(+),expressed in Escherichia cob' BL21(DE3),and analyzed by Western blot.Results X gene was successfully designed and expressed in Escherichia coli BL21(DE3).Western blot analysis showed that recombinant X protein,which was with His marker,can be detected by anti-His antibody.Conclusion In this study we establish a newmethod to express recombinant epitope protein,which may be a new protein for developing vaccine against HSV infection.

Objective To study the expression of mammalian target of rapamycin(mTOR)in ischemic postconditioning(I-postC)-induced attenuation of ischemia/reperfusion(I/R)injury in rat skeletal muscle.Methods A total of 48 healthy male Wistar rats were randomly divided into 3 groups(n=16 each group):I/R group(4-hour ischemia followed by 12-or 24-hour reperfusion),ischemic preconditioning (IPC)group(3 cycles of 5-minute ischemia followed by 5-minute reperfusion),and I-postC group(3 cycles of 1-minute reperfusion followed by 1-minute ischemia).The rat model of I/R injury in right hind limb model was established by clamping the right femoral artery.The changes in the morphology,wet-to-dry weight ratio(W/D),malondialdehyde(MDA),and myeloperoxidase(MPO)in skeletal muscle were compared.The expression of mTOR was detected by Western blot and immunohistochemistry.Results In I-postC and IPC groups,the skeletal muscle edema was less severe,the levels of MDA and MPO significantly decreased,and the expression of mTOR significantly in creased,compared with I/R group(all P＜0.03).There was no significant difference between I-postC and IPC groups.Conclusion Ipostc may attenuate I/R injury in rat hind limbs by activating mTOR signal pathway,which is similar to the mechanism of IPC.

Objective To analyze the relationship between the expression of MHC class I chain-related gene A/B(MICA/B) on different human cancer cells and their sensitivity to NK cell cytotoxicity.Methods Hie expression MICA/B in K562,Bcap37,769P and A549 cells was measured by FACS.Isolation of NK cells were obtained by the modified RosetteSep~((R)) procedure.Cytotoxicity of NK cells at different ef0.50)%,(90.72±0.64)%,(55.59±0.55)%,and(3.84±0.10)% respectively.The purity of NK cells obtained by the modified RosetteSep~((R)) procedure was(96.52±2.42)%,whereas the cytotoxicity of NK cells against K562,Bcap39 and 769P was much higher than that of A549(P<0.01).The expression of MICA/B was significantly correlated with the cytotoxicity of NK cells at E:T ratios of 5:1,10:1 and 20:1 respectively(P<0.01).Conclusion The MICA/B expression on cancer cell lines was correlated with NK cell-mediated cytolysis and influenced the cytotoxicity of NK cells.

Objective To explore the relationship between KISS-1 gene and metastasis of bladder carcinoma,and to study the effect of the stable expression of KISS-1 gene on the invasion of bladder carcinoma cell line T24.Methods Fluorescent quantitative PCR was used to detect the expression of KISS-1 mRNA in primary bladder carcinoma without metastasis and primary bladder carcinoma with metastasis.Recombinant vector pIRES2-AKS-1 was constructed and transfected into T24 cells.Single clone of stably transfected cells was screened,and the changes in the invasive ability of T24 cells was detected after transfection.Results The expression level of KISS-1 mRNA in primary bladder carcinoma with metastasis was significantly lower than that in primary bladder carcinoma without metastasis(P＜0.05).The expression of KISS-1 protein in the single clone of stably transfected cells increased significantly,and the invasive ability significantly decreased(P＜0.05).Conclusion KISS-1 gene is correlated with the metastasis of bladder carcinoma,and the up-regulated expression of KISS-1 gene can inhibite the invasiveness of T24 cell line.

Objective To explore the expression of 10 kDa interferon-gamma-induced protein(IP-10)in pathological scar and its role in the pathogenesis of pathological scar.Methods Imrnunohistochemistry was performed to detect the expression and distribution of IP-10 in 28 patients with keloid(group K),34 patients with hypertrophic scar(group HS),and 20 normal controls(group N).The data were collected and analyzed statistically.Results The expression of IP-10 was significantly higher in groups K and HS than in group N(P＜0.01),but no significant difference in the expression of IP-10 was found between groups K and HS.Conclusion IP-10 may enhance the formation of pathological scar by attracting T lymphocytes and inducing immune/inflammatory response.

Objective To construct 2 recombinant plasmids carrying cassette chromosome recombinase C(ccrC)and ccrAB respectively and introduce the plasmids into methicillin-resistant Staphlococcus aureus(MRSA)strain 81/0432,and to observe the precise excision of Staphylococcal cassette chromosome mec(SCCmec)island from bacterial chromosome.Methods ccrC and ccrAB genes were amplified with chromosomal DNAs isolated from MRSA strains 81/0342 and N31S as PCR templates.We constructed recombinant plasmids pSR5C and pSR2AB by cloning ccrC and ccrAB genes into temperature-sensitive plasmid pYI3,after introducing them into MRSA strains 81/0432 and N315 by electroporation.PCR was performed to identify SCCmec excision from the bacterial chromosome.The transformants were serial passaged for 10 days,and then the drug resistance of these rransformants was detected by replica experiment.Results The fragment length of ccrC gene was 1.9 kb,smaller than the fragment length of ccrAB from N315.The recombinant plasmids of pSR5C and pSR2AB were successfully constructed.After these 2 recombinant plasmids were introduced into MRSA strain 81/0342,type-V SCCmec island was excised from the chromosome and formed a closed circular structure in the bacteria.However,type-Ⅱ SCCmec island could be excised only in N31S strain after the expression of pSR2AB.Replica experiment verified that transformed strains of 0342(pSR2AB),0342(pSR5C),and N315 (pSR2AB)were mostly susceptible to ceftizoxim or tobramycin after the excision of SCCmec island.Conclusion cciC could serve as a recombinase as ccrAB,which could mediate precise excision of SCCmec island from the chromosome of type-V MRSA strain.This study shows that type-V SCCmec island is widely disseminated between Staphylococcus aweus strains in community setting.

Objective To compare the two ways of twice irradiation on the rabbit liver VX2 tumor using high intensity focused ultrasound (HIFU).Methods Totally 45 tumor-bearing rabbits were randomly divided into 3 groups:group A(one-time irradiation group) received a one-time ablation;group B and C(twice irradiation group) firstly received a low-dose irradiation(without ablation),then group B received ablation on the next day while group C received it 2 days later.Results Compared with one-time irradiation group,the total treatment time of twice irradiation group was not significantly different,but the time of each irradiation,the incidence of skin erythema and the energy efficiency factor(EEF)were less,and that of group B were the least.After ablation,the typical coagulation necrosis in tumor tissues occurred,and the recurrence and metastasis were effectively controlled in all the three groups.Conclusion The total treatment time and efficacy of twice irradiation were same as one-time ablation,but the time and dose of each irradiation significantly decreased,the damage efficiency was enhanced and the complications were reduced.The way of continuous two-days twice irradiation was the most effective,which would be a safe and effective method of HIFU treatment.

Objective To prepare the long-circulating liposomes of paraoxonase(PON).Methods The long-circulating liposomes of paraoxonase were prepared by film dispersion method.The encapsulation efficiency was determined by gel column.The effects of the factors on the encapsulation efficiency,such as the weight ratio of paraoxonase to phospholipid,cholesterol(Choi) to phospholipid,PEG-cholesterol (PEG-Chol) and the iron strength of water phase,were investigated respectively.Then the formulation was optimized by orthogonal design.Results The encapsulation efficiency of the paraoxonase liposomes was 87.66±3.46%,and the average diameter of the liposomes was about 126 nm.There was no significant change on encapsulation efficiency on 15 d at 4 ℃,and the activity of paraoxonase was maintained basically stable.Conclusion The preparation of PEG-modified paraoxonase liposomes was easy and practicable,and the property investigation in vitro showed that the paraoxonase liposomes were stable.

Objective To observe the effects of nonylphenol(NP) on L-type Ca~(2+) currents(I_(Ca-L)) in isolated guinea-pig ventricular myocytes.Methods The ventricular myocytes were isolated from guinea-pig hearts.The L-type Ca~(2+) currents in the ventricular myocytes treated with NP were measured with whole cell patch-clamp technique.Results Nonylphenol inhibited lew,at different potentials.Nonylphenol (10~(-6) mol·L~(-1),10~(-5) mol·L~(-1)) reduced the peak amplitude of I_(Ca-L) from-3.2±1.5 pA·pF~(-1) to-1.6±0.8 pA·pF~(-1)(P<0.01) and-1.4±0.7 pA·pF~(-1)(P<0.01),respectively.Nonylphenol did not influence the activation curve of I_(Ca-L) significantly.Conclusion Nonylphenol could in hibit the L-type calcium currents in isolated guinea-pig ventricular myocytes with a concentration-dependent manner.Objective To observe the effects of nonylphenol(NP) on L-type Ca~(2+) currents(I_(Ca-L))in isolated guinea-pig ventricular myocytes.Methods The ventricular myocytes were isolated from guinea-pig hearts.The L-type Ca~(2+) currents in the ventricular myocytes treated with NP were measured with whole cell patch-clamp technique.Results Nonylphenol inhibited lew,at different potentials.Nonylphenol hibit the L-type calcium currents in isolated guinea-pig ventricular myocytes with a concentration-dependent manner.

Objective To explore the changes of the amount and function of Vα24~+Vβ11~+,CD161~+Vα24~+NKT cells in the peripheral blood of systemic lupus erythematosus (SLE) patients. Methods The amount of Vα24~+Vβ~+,CD16~+Va24~+NKT cells in the peripheral blood of 30 SLE cases and 30 healthy persons were detected by flow cytometry. Results Vα24~+Vβ11~+,CD161~+Vα24~+NKT cells in the peripheral blood of SLE patients were significantly lower than that of normal control (P < 0.05). Vα24~+Vβll~+ and CD161~+Va24~+NKT cells in the pe-ripheral blood of SLE patients positively correlated with the level of the complement (C3,C4) and negatively correlated with SLE disease ac-tivity index (SLEDIA) and ESR. Conclusion Activated NKT cells decreased in the peripheral blood of SLE patients. Vα24~+Vβ11~+ and CD161~+Vα24~+NKT cells might be involved in the pathogenesis and development of SLE.