Characteristics of gene sequence changes of hemorrhagic fever with renal syndrome virus in Heilongjiang Province / 中国实用内科杂志

OBJECTIVE: To study the gene sequence characteristics of hantavirus in Heilongjiang Province in order to find out the reasons for the changes of clinical characteristics of hemorrhagic fever with renal syndrome in Heilongjiang Province in recent years.METHODS: Totally 110 rat lung specimens,121 blood specimens from patients diagnosed with mild or atypical HFRS and 100 blood specimens from patients diagnosed with typical HFRS in the First Affiliated Hospital of Harbin Medical University and the Seventh Hospital of Qiqihar City from 2005 to 2015 were collected. The gene sequences were obtained by nucleic acid extraction, RT-NestPCR, and gene sequencing. Explore the possible reasons for the changes in clinical characteristics of HFRS by comparing the obtained sequences with previous strains, homology analysis, building phylogenetic trees of M gene, and finding out the law of nucleotide and amino acid loci changes. RESULTS: TM gene of twenty-six mild or atypical HFRS patients were successfully amplified, including 14 cases of HTN type and 12 cases of SEO type; M gene of twenty-two typical HFRS patients were amplified, including 19 cases of HTN type and 3 cases of SEO type. Compared with the standard strain 76-118, the nucleotide homology of hantavirus from mild or atypical HFRS patients, typical HFRS patients and mice was 74.4%-89.2%, 87.4%-90.3% and 88.1%-88.5%. Comparing hantavirus gene sequence from mice and from patients, the nucleotide homology was 79.7-99.1%. Hljh38 and Hljh39 from patients were significantly different from the other strains. They were the same subtype as Amur virus because they had high homology with Amur strains H5 and H8205(94.9%-97.6%). The deduced amino acids showed some variations compared with the standard strains, but no obvious variation law was observed. CONCLUSION: The reason for the changes of clinical characteristics of hemorrhagic fever with renal syndrome in Heilongjiang province is related to the change of viral type. There are also variations of hantavirus and amino acid, but the relationship between specific variation law and clinical manifestations needs to be further verified.

Immunotherapy with a chimeric AFP and HSP70 gene DNA vaccine targeting on a murine hepatocellular carcinoma / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To investigate immune responses and the anti-tumor effect of a constructed Chimeric AFP-Mt.HSP70 DNA vaccine in mice.</p><p><b>METHODS</b>Chimeric AFP-Mt.HSP70 was constructed by molecular clone techniques. Spleen cells derived from mice immunized twice were induced to secrete IFN gamma and were assayed using ELISA. The activity of the cytotoxic lymphocytes (CTL) derived from spleen cells was assayed using lactate dehydrogenase (LDH). 4 x 10(6) Hepa1-6 cells/200 microl were injected subcutaneously into the right axilla of each mouse bearing the tumor. The anti-tumor effect of the recombinant DNA vaccine was evaluated by measuring tumor sizes of the mice.</p><p><b>RESULTS</b>AFP-specific CTL reaction was induced by our chimeric DNA vaccine and Mt.HSP70 enhanced this effect (P < 0.05). The CTL activity was about 32% at E/T=50:1. The IFN gamma secreted by spleen cells of mice immunized with chimeric plasmids was about 200 pg/ml. It was higher than those in the other groups; Tumor sizes of mice immunized with fused plasmids were smaller than those in the other groups. Survival times of mice immunized with the fused plasmids were prolonged.</p><p><b>CONCLUSION</b>Chimeric DNA vaccine can induce AFP-specific CTL reaction and has an anti-tumor effect on transplanted tumors in our murine experiment.</p>

The role of dendritic cell and macrophage in hepatoma antigen-presenting / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To study the role of dendritic cells (DCs) and macrophages, differentiated from the same individual peripheral blood monocytes, in tumor antigen- presenting.</p><p><b>METHODS</b>DCs and macrophages were differentiated from human peripheral blood monocytes by adding both Granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4) or GM-CSF only. Then they were loaded with tumor antigen at different concentrations and cocultured with autologous T cells in 96-well flat-bottomed microtiter plates for five days at 37 degrees C, 5% CO(2). (3)H-thymine was added before the culture terminated, and twelve hours later, the cells were gathered to test the cpm value.</p><p><b>RESULTS</b>Both DCs and macrophages chased with tumor antigen could strongly stimulate the proliferation of autologous T cells, especially DCs. The stimulation effect with 20 microl/ml antigen was the most remarkable and the cmp values were 11,950.3 +/-1621.8, 8,708.5 +/-176.1, 402.5+/-43.1 in DCs group, Macrophages group, and lymphocytes group, respectively.</p><p><b>CONCLUSION</b>The antigen presenting role of DCs is stronger than that of macrophages from the same individual.</p>

Transient expression of fusion protein with a chimeric HBsAg-HSP70 construct in HepG2 cells / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate transient expression of fusion protein with a chimeric HBsAg-HSP70 construct in HepG2 cells.</p><p><b>METHODS</b>Enkaryotic expression plasmids inserted HBsAg gene or chimeric HBsAg-HSP70 gene were prepared and transfected into HepG2 cells by means of cationic liposome. mRNA were detected by RT-PCR and proteins expressed in the cells were detected by immunocytochemistry 48 hours later. HBsAg in cultured supernatants and cell lysates were assayed by ELISA.</p><p><b>RESULTS</b>Fusion protein (HBsAg-HSP70) transient expression in HepG2 cells were confirmed by RT-PCR, immunocytochemistry or ELISA, but fusion protein was not assayed in cell cultured supernatants by ELISA.</p><p><b>CONCLUSIONS</b>Transfection of HepG2 cells with a chimeric HBsAg-HSP70 construct leads to express fusion protein, but it does not secrete into cell cultured supernatants.</p>