Currently,there are a lot of targets for the treatment of HBV infection,both for the host,and for the virus itself.However,existed clinical drugs can only control HBV infection,and can not remove the HBV,especially cccDNA.Therefore,the chronic persistent infection caused by HBV related diseases is still seriously threat to human health.People are still impatient for the development of new effective anti-HBV drugs.In this paper,we review the recent research of the anti-hepatitis B virus based on the host as the target.

Aim To investigate the method of cell culture for smooth muscle cells from rat cerebral basilar artery and understand cells growth and biological characteristics.Methods The explant attached method was applied for cell culture of rat basilar artery smooth muscle cells(BASMCs).The cultured BASMCs were identified by immunocytochemical staining.The activities of cells were indicated by the dynamic changes of intracellular calcium concentration observed by RF-5 000 fluorospectro-photometer.Results BASMCs grew out of tissue blocks by 5 days,reached confluency could be subcultured after 2 weeks.Cultured cells were identified by intensely positive immunocytochemical staining to smooth muscle actin-specific.Introduction of calcium channel agonists induced significant increase in Fura-2 fluorescence ratio(F340/F380)and cells were in good condiction.Conclusion Explant attached method is simple,efficient and economic.It provides an ideal cell model for the study of pathogenesis of the cerebral vascular diseases.

In the present study, the total RNA was extracted from three instar larvae of Musca domestica, the cDNA sequence encoding the ORF of cecropins was amplified by RT-PCR, and the target fragment was further sequenced after being cloned into T vector pUCm-T. Then, the cDNA sequence of the mature cecropins was amplified by PCR with recombinant plasmid pUCm-T/cecropin as template, the N-terminal rare codon GGA of E. coli was changed to the favorable codon GGC,and a Asn codon AAC was added in front of the stop coden TAA in the C- terminus. This mutant gene designated as mCecropin was then ligated with the fusion expression vector pGEX-4T-1. After restriction analysis and DNA sequencing, the positive recombinant plasmid pGEX-4T-1/mCecropin was transformed to different strains of E. coli cells and the fusion protein was expressed after IPTG induction. The fusion protein was assayed by SDS-PAGE and the E. coli BL21(DE3) cell was chosen as the host cell for the expression of the fusion protein. The expressed fusion protein GST-mCecropin was purified by GSTrap affinity coloum and the GST marker was then cleaved by thrombin. In this way, the fusion protein mCecropin with antibacterial activity was obtained after purification with HiTrap benzamidine column.

Objective Using shotgun mass spectrometry to detect proteins probably contained in bone marrow stromal cells(BMSCs) conditioned medium. MethodsMixed with BMSCs conditioned medium was divided into two parts which is(>5kD and <5kD) by means of ultrafiltration. The two parts were used to culture neural stem cells(NSCs) separately, and the proportions of neurons, astrocytes and oligodendrocytes in the offsprings of NSCs were calculated, then the effective part that could regulate the differentiation of NSCs was detected by Shotgun mass spectrometry. Results The BMSCs conditioned medium which is >5kD could promote the NSCs differentiate into more neurons and oligodendrocytes. The SDS-PAGE of this part showed that the most proteins were above 14kD, then the protein bands were enzymed. In total, 456 proteins were identified by Shotgun mass spectrometry after all the protein bands were enzymed, there were 154 similar proteins, 17 hypothetical proteins and 56 unknown proteins. And in the rest of 229 proteins, most of them were cytoskeletal proteins, secreted proteins, signal transduction proteins, enzymes, transporter and so on. Conclusion Many proteins secreted by BMSCs could regulate the differentiation of NSCs so as to prove the protein components probably existed in the BMSCs conditioned medium.

Objective To investigate the diagnostic value of matrix gamma carboxy glutamic acid protein(MGP) for coronary heart disease(CHD) .Methods Enzyme linked immunosorbent assay was performed for the detection of serum MGP level in health‐y subjects and CHD patients with different coronary artery calcium score(CACS) .Receiver operating characteristic(ROC) curve was used to analyze the diagnostic value of MGP for CHD .Results Between CHD patients and healthy subjects ,and CHD patients with different CACS ,the difference of serum MGP level was significant ,and serum MGP level was positively correlated with CACS (P<0 .05) .ROC curve of showed that the area under ROC curve was 0 .667 ,the diagnostic threshold was 70 .69 pg/mL ,the diag‐nostic sensitivity was 58 .80% ,the specificity was 83 .70% and the Youden index was 0 .425 .Conclusion CHD patients might be with abnormal serum MGP level ,which could be positively correlated with CACS .MGP might be with significant value for the diag‐nosis of CHD ,could be useful for the clinical prevention and early diagnosis of CHD .

OBJECTIVETo construct a lentiviral vector carrying the γ-synuclein(SNCG) gene and establish a human colorectal carcinoma cell line SW1116 stably expressing this gene, and then investigate the inhibition of the growth and invasion capacity of SW1116 cells.

METHODSRNA interference fragment was designed according to the SNCG sequence (GenBank: No.NM003087.2), and then SNCG RNAi effective target genes were screened. After the Oligo DNA of target sequences was synthesized, the lentiviral vectors carrying LV-SNCG-RNAi-EGFP (RNAi group) and LV-SNCG-NC-EGFP (NC group) were constructed and packaged to produce lentivirus venom. The supernatants of different virus-producing cells were used to transfect SW1116 cells respectively. Wild SW1116 cells were used as blank control (CON group) EGFP fluorescence was detected by fluorescent microscopy and the differential expression of SNCG mRNA and protein was detected by real-time PCR and Western blot. CCK-8, soft agar assay and Transwell chamber were employed to estimate the inhibiting effect on growth and invasion of SW1116 respectively.

RESULTSRecombinant lentiviral vectors respectively carrying the SNCG-RNAi-EGFP and SNCG-NC-EGFP were successfully constructed and the supernatants of lentivirus could effectively infect SW1116 cells. The titer of the virus carrying LV-SNCG-RNAi-EGFP or LV-SNCG-NC-EGFP was 8×10(8) TU/ml. Real-time PCR and Western blot confirmed that compared with the NC group, SNCG-RNAi group had lower SNCG expression (1.009±0.161 vs. 0.114±0.030, P=0.009), and showed tremendous silencing effect as 76.8%(P<0.05). SNCG protein expression was also significantly reduced (RNAi:12.001±2.884, NC:32.443±4.731, CON:34.308±6.920, P<0.05). After SNCG knockdown, the number of proliferation cells was obviously reduced at 48, 72, 96 and 120 hours respectively(P=0.036). In soft agar assay, clones in RNAi group were smaller[RNAi:(0.582±0.103) mm, NC:(1.863±0.316) mm, CON:(1.749±0.525) mm]. Colony formation rate of RNAi group was down to (17.1±3.5)%, which was significantly lower than (36.5±4.3)% in NC group and (33.8±3.9)% in CON group. In migration test, the number of invasion cell was 37.4±9.3 in RNAi group, which was significantly less than 112.3±8.6 in NC group and 100±0.0 in CON group.

CONCLUSIONExpression of SNCG mRNA and protein plays an important role in the growth and the invasion capacity of SW1116 cells.

Cell Line, Tumor ; Cell Proliferation ; Colorectal Neoplasms ; pathology ; Genetic Vectors ; Humans ; Lentivirus ; RNA Interference ; RNA, Messenger ; RNA, Small Interfering ; genetics ; Real-Time Polymerase Chain Reaction ; Transfection ; gamma-Synuclein ; genetics

Objective : To investigate the effect and mechanism of ARRB1 on Armillariella tabescens polysaccharides reversal of 5-fluorouracil ( 5-FU ) -induced chemotherapeutic intestinal mucosal injury． Methods : Twelve ARRB1 knockout ( ARRB1 －/ － ) and wild-type ( WT) C57BL /6 mice were randomly divided into Control，Model and ATPS groups (200 mg / kg) ，respectively．5-FU (50 mg / kg) was injected intraperitoneally for 7 days to establish a model of chemotherapeutic intestinal mucosal injury．The histopathological damage of jejunum was evaluated by HE staining ; the activity of serum superoxide dismutase (SOD) and diamine oxidase (DAO) was measured by kits ; the expression of tight junction protein (TJ) markers ZO-1，Occludin，Claudin-1 and proliferation-associated protein Ki-67 was detected by immunohistochemistry．Crypt isolation and organoid culture were used to detect the growth status of small intestinal organoids. Results : 5-FU chemotherapy reduced body weight，aggravated histopathological damage in small intestine，decreased SOD level，TJ protein and Ki-67 protein expression，increased serum DAO level，decreased spherical structure formation rate and organoid formation rate ; compared with the model group，after ATPS treatment，WT mice recovered body weight，decreased pathological damage，increased serum SOD level，TJ protein and Ki-67 protein expression，DAO levels decreased，and the rates of spherical structure for- mation and organoid formation were significantly higher．However，ARRB1 －/ － mice failed to reverse the effect of 5- FU after ATPS treatment． Conclusion ATPS reverses 5-FU-induced intestinal mucositis through the protective effects of ARRB1 on intestinal barrier and organoid growth.