Objective To study the influence of HBV replication regulator,enhancer I and Pre- S2,on the immune response of HBV DNA vaccine.Methods DNA fragments of HBsAg,PreS2 HBsAg,HBsAg-enhancer I and PreS2-HBsAg-enhancer I region of HBV were amplified by PCR using the complete genome DNA of HBV adr subtype,and inserted into VR1012 vectors,respective- ly.The recombinant plasmids were transfected into HepG2 cells,and injected into Balb/C mice.The expression of HepG2 cells and the cellular and humoral immune response of mice were tested by cell immuno-chemistry,ELISA and ELISPOT.Results The target protein were expressed by transfected HepG2 cells,enhancer I and Pre-S2 can promote the expression of HBsAg in transfected cells.The HBsAb and the HBsAg specific CTL in inoculated mice were found in the second week after injection, PreS2 but not enhancer I can promote the immune response in inoculated mice.Conclusions When inserted into HBV DNA vaccine,enhancer I and PreS2 can promote the expression of HBsAg in transfected HepG2 cells,PreS2 can promote the immune response in inoculated Balb/C mice.

OBJECTIVETo observe anti-atherosclerotic effect of Xuefu Zhuyu Granule (XZU) and Danlou Tablet (DT) on blood lipids, platelet derived growth factor (PDGF), vascular smooth muscle cells (VSMCs) proliferation, extracellular signal-regulated kinase (ERK) signal pathway in atherosclerosis (AS) model rats, and to explore their potential mechanisms.

METHODSForty male Wistar rats were randomly divided into five groups, i.e., the normal control group, the model group, the Atorvastatin group, the DT group, the XZG group, 8 in each group. Rats in the normal control group were fed with basic forage for 12 weeks, while rats in the other four groups were fed with high fat forage plus intraperitoneal injection of vitamin D3 to build AS model. Then rats in the model control group, the Atorvastatin group, the DT group, the XZG group were administered with normal saline, Atorvastatin suspension (0.18 mg/mL), DT suspension (45 mg/mL), and XZG (1 g/mL) by gastrogavage for 8 successive weeks, respectively. After intervention serum levels of TC, TG, LDL-C, HDL-C, and PDGF were detected by ELISA. Pathological changes in thoracic aorta were observed by HE staining. Protein expression levels of ERK1/2 and pERK1/2 in thoracic aorta were measured by Western blot.

RESULTSCompared with the normal group, serum TC, TG, LDL-C, PDGF levels, and expression levels of ERK1/2 and pERK1/2 significantly increased (P <0. 05) in the model control group. HE staining showed irregular intimal thickness, accumulated endothelial foam cells, lipids deposited, disarranged media VSMCs, forming typical AS plaque. Compared with the model group, TC and PDGF levels decreased in all medicated groups (P < 0.05, P < 0.01). Serum levels of TG and LDL-C significantly decreased in the Atorvastatin group and the DT group (P < 0.01, P < 0.05). Expressions of ERK1/2 and pERK1/2 significantly decreased in the Atorvastatin group, the DT group, and the XZG group (P < 0.01). HE staining also showed typical AS plaque in three medicated groups, but with reduced pathological degree of endometrial hyperplasia and plaque area.

CONCLUSIONSXZG and DT could reduce the plaque area and attenuate pathological degree of AS in model rats, thereby postponing the progress of AS. Its mechanism might be achieved through reducing serum lipids and release of PDGF, inhibiting ERK signal pathway activation and VSMC proliferation.

Animals ; Aorta, Thoracic ; Atherosclerosis ; drug therapy ; Drugs, Chinese Herbal ; administration & dosage ; pharmacology ; therapeutic use ; Extracellular Signal-Regulated MAP Kinases ; Lipids ; Male ; Plaque, Atherosclerotic ; Rats ; Rats, Wistar ; Tablets

Adenomatoid Tumor ; metabolism ; pathology ; surgery ; Adult ; Biomarkers, Tumor ; metabolism ; Female ; Humans ; Hysterectomy ; methods ; Keratins ; metabolism ; Lymphangioma ; pathology ; Uterine Neoplasms ; metabolism ; pathology ; surgery ; Vimentin ; metabolism

In vivo Mammalian Bone Marrow Micronucleus Test is included in the standard battery genotoxicity testing,with great application prospects in medicine,public health,food and drug safety evaluation fields.Establishing standardized experimental methods and conditions in GLP condition and accumulating a certain range of background data could effectively ensure the reliability of the test system,and also provide strong basis to support the experimental data.We herein summarized the background data of mouse and rat bone marrow micronucleus tests performed from 2007 to 2015,to expound the standardized data collection method for rodent animal bone marrow micronucleus test.

Background Even though the change in wavefront aberrations with correction modality is well documented in the literature,little is known about the underlying mechanism.Complete understanding of the causes responsible for the wavefront change in the combined lens-eye system is important since it provides basic knowledge for further improving the technique to correct refractive error by correcting lenses.Objective The aim of this study was to investigate the influence of refractive correction lens on optical property of the eye by analyzing Zernike aberrations in myopic eyes with contact lens correction.Methods This study was approved by the Ethic Committee of Wenzhou Medical College.Written informed consent was obtained from each subject before entering this study.Zernike aberrations of 52 myopic eyes of 26 subjects with the spherical equivalent-1.75 to-8.50 D were measured using a Hartmann-Shock wavefront sensor.The human eye aberrations were examined at the uncorrected condition,rigid-gas-permeable contact lens (RGP-CL) corrected condition and soft contact lens (Soft-CL) corrected condition.The differences of wavefront aberrations and Zernike coefficients were compared by repeated measurement of single factor variance analysis,and correlation of the aberration changes between uncorrected condition and RGP-CL corrected condition or Sofi-CL corrected condition,between the right eyes and left eyes in different conditions were analyzed by Pearson linear correlation.Results Mean total root-mean-square (tRMS) was (0.71 ± 0.30)μm,(0.54±0.19)μm and (0.74±0.32)μm in the uncorrected condition,RGP-CL corrected condition and Soft-CL corrected condition,with a significant difference (F =8.758,P＜0.001),and tRMS was significant declined under the RGP-CL corrected condition compared with uncorrected condition (t =2.746,P =0.008),and tRMS in RGP-CL corrected condition was significantly lower than that in Soft-CL corrected condition (t =3.428,P =0.001).The high RMS (hRMS) was (0.34±0.12) μm,(0.28 ±0.12) μm,(0.40±0.14) μm in the uncorrected condition,RGP-CL corrected condition and Soft-CL corrected condition,with a significant difference among them (F =10.681,P＜0.001).An insignificant decrease of hRMS was seen in the RGP-CL corrected condition compared with uncorrected condition (t =1.987,P=0.053),but hRMS value was significant higher in the Soft-CL corrected condition than that in the uncorrected condition (t=2.101,P=0.041) and RGP-CL corrected condition (t=4.266,P＜0.001).Compared with uncorrected condition,the axis astigmatism (C5) and spherical aberration (C12) in the RGP-CL corrected condition and spherical aberration (C12) in the Soft-CL corrected condition were significantly reduced (P＜0.05),and the absolute values of trefoil (C6),vertical coma (C7) and tetrafoil (C10) in the RGP-CL corrected condition were lower than those of the uncorrected condition,but vertical coma (C7) absolute value in the Soft-CL corrected condition was increased (P＜0.05).A significantly positive correlation was seen in the spherical aberration (C12) between the RGP-CL corrected condition and uncorrected condition (r =0.763,P＜0.001),and less significant correlation was in the secondary astigmatism (C11) between the Soft-CL corrected condition and uncorrected condition(r=0.469,P＜0.001).Conclusions Different contact lens corrected conditions exert their effects on ocular wavefront structure due to its unique interaction with the eye.RGP-CL wearing has strong modification on wavefront aberrations probably due to its molding effect on corneal surface,which reduces the bilateral symmetry.High order wavefront aberration can be modified by Soft-CL wearing.

Objective To observe the immune effect of DNA vaccines encoding mutated HBV pre-c/c gene(VE2,VFA) in mice. Methods Three kinds of plasmid VEC(DNA vaccines encoding HBV pre-c/c gene),VE2 and VE4 were injected into the thigh muscles of different group of BALB/c mice. Blood and splenocytes from mice were isolated at 4 weeks after immunization. We also have mouse groups immunized with three of these plasmid combined with IFN-γ gene plasmids. The anti-HBc and anti-HBe antibody in peripheral blood in mice were detected by enzyme linked immunusorbent assay( ELISA),antigen-specific cell immune responses were detected by CTL test and enzyme linked immunospot assay( ELISPOt). Results We found that anti-HBe titers of VE2 and VE4 immunizing groups are higher than VEC group( P < 0.05). We also observed that VE2 and VE4 could induce stronger antigen-specific immune responses than VEC and when combined with IFN-γ plasmid,the antigen-specific immune responses are stronger than those without combination immunization in mice (P < 0.05). Conclusions The DNA vaccine VE2 and VE4 could induces stronger antigen-specific immune responses than VEC,and when combined with IFN-γ plasmid,the antigen-specific immune responses are improved in mice.

OBJECTIVETo prepare cytochrome (CYP)2E1-silenced hepatocytes by lentivirus-mediated RNA interference technology and to investigate the hepatotoxicity of trichloroethylene (TCE) in CYP2E1-silenced hepatocytes.

METHODSShort hairpin RNA fragments were designed and synthesized and were then ligated into the lentiviral vector; single colonies were screened; the plasmid was extracted after PCR and sequence identification and then transferred into L02 hepatocytes; the CYP2E1-silenced hepatocytes were selected; real-time quantitative PCR and Western blot were used to evaluate the interference effects. The obtained CYP2E1-silenced hepatocytes, as well as normal L02 hepatocytes, were treated with TCE (0, 0.25, 0.50, 1.00, 2.00, and 4.00 mmol/L). The cell viability and half maximal inhibitory concentration (IC50) of TCE were measured; the apoptotic rate of cells was measured by flow cytometry; the mRNA expression levels of apoptosis genes and oncogenes were measured by real-time quantitative PCR.

RESULTSThe IC50s of TCE for L02 hepatocytes and CYP2E1-silenced hepatocytes were 15.1 mmol/L and 23.6 mmol/L, respectively. The apoptotic rate increased as the dose of TCE rose in the two types of cells; the CYP2E1-silenced hepatocytes hada significantly lower apoptotic rate than L02 hepatocytes when they were exposed to 2.0 and 4.0 mmol/L TCE (P < 0.05 or P < 0.01). The mRNA expression level of bcl-2 (anti-apoptosis gene) in CYP2E1-silenced hepatocytes was 15% ∼ 60% higher than that in L02 hepatocytes (P < 0.01), while the mRNA expression levels of caspase-3 and caspase-9 (apoptosis genes) in CYP2E1-silenced hepatocytes were 30% ∼ 60% lower than those in L02 hepatocytes (P < 0.01). The mRNA expression level of p53 (cancer suppressor gene) in CYP2E1-silenced hepatocytes was 81 - 278% higher than that in L02 hepatocytes (P < 0.01), while the mRNA expression levels of c-fos and k-ras (oncogenes) in CYP2E1-silenced hepatocytes were 20-68% lower than those in L02 hepatocytes (P < 0.01).

CONCLUSIONCYP2E1-silenced cells can be successfully prepared by lentivirus-mediated RNA interference technology. Silencing CYP2E1 gene can reduce the hepatotoxicity of TCE and inhibit the expression of some apoptosis genes and oncogenes, suggesting that CYP2E1 gene plays an important role in TCE metabolism and is related to the hepatotoxicity of TCE.

Apoptosis ; drug effects ; genetics ; Cell Line ; Cell Survival ; drug effects ; genetics ; Cytochrome P-450 CYP2E1 ; genetics ; metabolism ; Genetic Vectors ; Hepatocytes ; drug effects ; metabolism ; Humans ; Lentivirus ; genetics ; RNA Interference ; Trichloroethylene ; toxicity

OBJECTIVETo improve the diagnosis and treatment of primitive neuroectodermal tumors (PNET) of the pancreas.

METHODSOne patient with PNET of the pancreas was reported in this article. The corresponding literatures on the diagnosis and treatment was reviewed.

RESULTSThe patient was diagnosed as pancreatic PNET by her clinical, microscopic, and immunohistochemical features as well as cytogenetic analysis after the resection of the tumor located in the uncinate process in PUMC Hospital. Radiochemotherapy was given after the operation for 8 months and no recurrence was observed. Since PNET of pancreas have no specific clinical symptoms and most patients have jaundice and/or abdominal pain, the diagnosis depended on the immunohistochemical features of positive P30/32(MIC2) and at least two of the neural markers. The cytogenetic analysis showed translocation mainly harbored the characteristic t (11; 22) (q24; q12). Since pancreatic PNET were highly aggressive, early chemotherapy, close follow-up, and immediate surgical interventions were required as early as possible.

CONCLUSIONPNET can occur in pancreas, and diagnosis and treatment should be made as early as possible to improve the outcome.

Child ; Combined Modality Therapy ; Female ; Follow-Up Studies ; Humans ; Neuroectodermal Tumors, Primitive ; diagnosis ; therapy ; Pancreatic Neoplasms ; diagnosis ; therapy

OBJECTIVETo develop an HPLC method for determination of fingerprints and evaluate green tangerine peel (GTP) processed with vinegar.

METHODThe samples were extracted with 50% ethanol using ultrasonic bath and analyzed on a Chromasil C18 column eluted with methanol and water containing 0.2% acetic acid in a linear gradient program. The flow rate was 1.0 mL x min(-1), and detection wavelength was set at 280 nm.

RESULTThe RSD values of relative retention times and peak areas of main peaks were all less than 5%. The fingerprints of Geqingpi and Sihuaqingpi were different. The fingerprints of the processed and the unprocessed were almost same, and the fingerprint differences among the different batches processed with the same technology from the same unprocessed GTP were not obvious.

CONCLUSIONThe influence of processing with vinegar on 50% ethanol extracts were not evident, and the technological pross was stable. It was found that the fingerprints of Geqingpi and Sihuaqingpi were different, but it should be study further.

Acetic Acid ; Chromatography, High Pressure Liquid ; methods ; Citrus ; chemistry ; Drug Stability ; Fruit ; chemistry ; Plants, Medicinal ; chemistry ; Technology, Pharmaceutical ; methods

OBJECTIVETo clone tpn17 and tpn47 genes of Treponema pallidum and then construct their prokaryotic expression systems,to establish ELISAs based on rTpN17 and rTpN47 as antigens and to evaluate the sensitivity and specificity of the ELISAs for detection of serological diagnosis of syphilis.

METHODSThe whole length of tpn17 and tpn47 genes was amplified by PCR and then their prokaryotic expression systems were constructed. SDS-PAGE was used to measure the expression of the target recombinant proteins rTpN17 and rTpN47. Ni-NTA affinity chromatography was applied to extract rTpN17 and rTpN47, while Western blot was performed to determine the specific immunoreactivity of rTpN17 and rTpN47. By using rTpN17 and rTpN47 as the coated antigen, respectively, ELISAs (rTpN17-ELISA and rTpN47-ELISA) were established to detect serum samples from 200 healthy individuals, 25 RA patients, 17 SLE patients and 211 syphilis patients. The detection effects of the ELISAs were compared to those of TRUST and TPHA.

RESULTThe sequence similarity of the cloned tpn17 and tpn47 genes was 100 % compared with the corresponding sequences in GenBank. The expression outputs of rTpN17 and rTpN47 were approximately 37.2 % and 26.8 % of the total bacterial proteins, respectively. Both the extracted rTpN17 and rTpN47 could take place remarkable conjugation reactions to the sera with positive antibody against Treponema pallidum.The positive detection rate of TPHA (99.1%) was the highest (P<0.001). The positive detection rates of rTpN17-ELISA (85.3 %) and rTpN47-ELISA (84.3 %) were similar (P>0.05). The positive detection rates of TRUST (72.5 %) was lower than that of rTpN17-ELISA (P=0.001) but similar to that of rTpN47-ELISA (P=0.014). The detection results of all the serum samples from healthy individuals, RA patients and SLE patients were negative, whereas 7.1 % (3/42) of the samples from RA or SLE patients were positive.

CONCLUSIONrTpN17 and rTpN47 are still maintaining their original immunoreactivity. The ELISAs using rTpN17 or rTpN47 as the antigen are rapid, simple and convenient, higher sensitivity and specificity methods for serological screening and detection of syphilis.

Antibodies, Bacterial ; Antigens, Bacterial ; Blotting, Western ; Enzyme-Linked Immunosorbent Assay ; methods ; Female ; Humans ; Male ; Syphilis ; diagnosis ; Syphilis Serodiagnosis ; Treponema pallidum ; chemistry ; immunology ; isolation & purification