Biomedical subjects

Zhonghua Jiang

Publications and source records attributed to Zhonghua Jiang.

6 recordsLinked to original sources

PubMed2006

Construction of eukaryotic expressing plasmids encoding HA and HA1 of influenza A virus and their transient expression in HEK293 cells.

In order to explore the feasibility and protective efficiency of influenza DNA vaccine, we constructed eukaryotic expressing plasmids encoding HA and HA1 of influenza A virus (A/PR/8/34) and studied their expression in HEK293 cells. HA and HA1 genes were amplified by RT-PCR and cloned into pcDNA3.1(+) to generate pcDNA3.1(+)/HA and pcDNA3.1(+)/HA1, respectively. After verification of the cloning fidelity by restriction endonuclease digestion, PCR, and sequencing, pcDNA3.1(+)/HA and pcDNA3.1(+)/HA1 were transfected into HEK293 cells using PolyFect Transfection Reagent. Immunofluorescence assay was used to detect the transient expressing cells. Fluorescence microscopy revealed strong expression of target gene in HEK293 cells transiently transfected with either pcDNA3.1(+)/HA or pcDNA3.1(+)/HA1. Therefore, the results confirm the successful construction of eukaryotic expressing plasmids capable of driving the eukaryotic expression of influenza virus antigen HA and HA1, which is likely to provide a basis for both further investigation of the mechanism of influenza viral infection and the development of influenza DNA vaccine.

Cell Line↗

PubMed2006

[Suppression of telomerase activity by plasmid-mediated RNA interference].

This study was aimed to construct a plasmid expressing siRNA specific for the human telomerase reverse transcriptase (hTERT) gene and to evaluate the ability of small interference RNA(siRNA) for inhibiting telomerase activity in HeLa cells. 64 nucleotides, in which 19 nt were homologous with hTERT gene, were chemically synthesized, annealed and linked into pSUPER to get pSUP-hTE. Then pSUP-hTE was digested with enzyme. We obtained its fragmant concluding promoter and 64nt. So we cloned it into pEGFP-C1 for constructing pEGFP-hTE which contains neo gene and the enhanced green fluorescent protein (EGFP). Recombinant pEGFP-hTE was transfected to HeLa cells. These cells were screened with medium containing G418. When stable colonies appeared, G418-resistant cells were harvested and propagated. At the different cell generations, hTERT mRNA and protein expression, telomerase activity and cell growth activity were analyzed. Compared with control cells, HeLa cells transfected with pEGFP-hTE showed that hTERT mRNA level and hTERT protein expression decreased and telomerase activity reduced by 38%, but the cells growth activity displayed no changes. So pEGFP-hTE could specifically inhibit expression of hTERT and telomerase activity. These results suggested that siRNA targeting hTERT gene might provide a new strategy for cancer biotherapy.

Base Sequence↗

PubMed2003

[Apoptosis of cancer cells induced by influenza virus and its elementary mechanism].

OBJECTIVE: To study the apotosis-inducing effects of influenza virus on cancer cells and the role of FasL. METHODS: Four strains of cancer cells (Hela, Raji, SMMC-7721 and SPC-A-1) were infected with 20 m. o. i influenza virus and the effects of apoptosis were detected at different time points post infection by electron microscopy, DNA agrose gel electrophoresis, PI stained flow cytometry, and Annexin-V FITC/PI stained flow cytometry. And the concentration of FasL in the culture supernatant was measured with the use of ELISA at the same time. RESULTS: All the four strains of cancer cells showed apoptotic morphological changes. DNA agarose electrophoresis also showed "ladder" in a time dependent manner after virus infection. During this course, there was an obvious increase of FasL concentration in the cell culture supernatant. CONCLUSION: Influenza virus can induce tumour cells apoptosis in vitro, and it is probably related to the increased expression of FasL.

Adenocarcinoma↗

PubMed2003

Apoptosis in Raji cell line induced by influenza A virus.

OBJECTIVE: To study the apoptotic effects of influenza A virus on the Raji cell line. METHODS: Cultured Raji cells were infected with influenza A virus at a multiplicity of infection (m.o.i) of 20 and the effects of apoptosis were detected at different time points post infection using the following methods: electron microscope, DNA agarose gel electrophoresis, PI stained flow cytometry (FCM) and Annexin-V FITC/PI stained FCM. RESULTS: Raji cells infected with influenza A virus showed changes of morphology apoptosis, DNA agarose electrophoresis also demonstrated a ladder-like pattern of DNA fragments in a time-dependent manner. PI stained FCM showed "apoptosis peak" and FITC/PI stained FCM showed apoptotic cells. Quantitative analysis indicated that the percentage of apoptotic Raji cells increased after infection, and cycloheximide (CHX), an eukaryotic transcription inhibitor, could effectively inhibit the apoptotic effects of influenza A virus in vitro. CONCLUSIONS: Influenza A virus can induce apoptosis in Raji cell line suggesting that it may lead to a potential method for tumor therapy.

Apoptosis↗

PubMed2003

[Construction and screening of TsF cDNA library specific for AchR].

OBJECTIVE: To construct the T suppress factor (TsF) cDNA library specific for AchR and screen out the positive clones of TsF. METHODS: The full-length cDNA was synthesized by the reverse transcripting from the TsF PolyA+mRNA, which had been extracted from the AchR specific suppressor lines (ARSL). The cDNA library was obtained using Lambda gt11 vector. Finally, this cDNA library was screened by Western blotting using Analysis of this cDNA library showed that titer of the monoclonal antibody specific for TCR-achain. RESULTS: recombinant bacteriophage was 1.4 x 10(7) pfu/ml, the rate of positive recombinant was 86.9%. The recombinant bacteriophage DNA were digested by EcoRI and electrophoresis analysis found that the main size of insertion was 0.8 to approximately 4.0 kb . As a result of screening, 27 positive clones were obtained. The positive recombinant bacteriophage DNA were digested by EcoRI, and electrophoresis analysis found that the size of insertion was 1 kb. Conclusion The results suggested that this cDNA library is available, the recombination phages can express TsF specific for AchR effectively and could be used to prepare the products of gene engineering of TsF.

Animals↗

PubMed2002

[The changes of thromboxane B2 (TXB2) and 6-keto-prostaglandin F1 alpha(6-keto-PGF1 alpha) in the lungs of rats drowned in hypothermic-sea-water].

OBJECTIVE: To observe changes of thromboxane B2 (TXB2), 6-keto-prostaglandin F1 alpha(6-Keto-PGF1 alpha) and TXB2/6-Keto-PGF1 alpha (T/P) in lungs of rats drowned in hypothermic sea water and to assess their influence on the blood-gas. METHODS: Rats of different groups were drowned nearly to death in hypothermic sea water and then taken out of the water rapidly, observed at room temperature, after that the following steps were taken in 5, 15, 30, 60, 240 min and 360 min groups, that were 1 ml arterial blood taken from left heart for blood-gas analysis including pH, PaO2 and PaCO2, rectal temperature observed; at last, the ratio of left dry lungs with left wet lungs was assessed, TXB2 and 6-Keto-PGF1 alpha in right lungs were examined in all above groups and dead group(14 rats dead, only 4 examined). RESULTS: The rectal temperature[(20.13 +/- 0.48) degree C], pH(6.68 +/- 0.03), PaO2[(45.00 +/- 6.30) mm Hg)], TXB2[(97.46 +/- 17.46) ng/L] and 6-Keto-PGF1 alpha[(25.59 +/- 8.12) ng/L] dropped to the lowest point in the 5 minutes group(P < 0.01), while PaCO2[(89.18 +/- 5.10) mm Hg] reached the highest point(P < 0.01), all above items from 5 minutes group then showed a recovering tendency, but only the pH in 240 minutes and 360 minutes groups as well as TXB2 in 360 minutes group and dead group reached near the level of normal control groups (P > 0.05); T/P had a rising tendency and reached the highest point in the 360 minutes group. CONCLUSIONS: The production and secretion of TXB2 and 6-Keto-PGF1 alpha were influenced by hypothermia, hypoxemia and acidosis, the imbalance of T/P could be one of factors influencing the improvement of blood gas index.

6-Ketoprostaglandin F1 alpha↗