Preliminary study on the mechanism of halofuginone inhibiting the activity of HepG2 cells / 药学学报

This study aimed to investigate halofuginone's inhibitory effect and mechanism on the activity of hepatocellular carcinoma cells. HepG2 cells were used to detect the effects of halofuginone. After treatment, cell activity, cell migration, cell cycle, and cell apoptosis were detected by CCK-8, transwell, and flow cytometry, respectively. The expression levels of growth and metabolism-related factors such as citrate synthase (CS), ketoglutarate dehydrogenase (OGDH), and isocitrate deoxygenase (IDH) were detected by real-time quantitative PCR and Western blot. Compared with the control group, the activity of HepG2 cells was significantly inhibited by halofuginone (P < 0.01), the migration rate of HepG2 cells was decreased (P < 0.01), the apoptosis of HepG2 cells was induced (P < 0.01), and the cell cycle was arrested in S phase (P < 0.01). The expression levels of tricarboxylic acid key enzymes CS, IDH3, and OGDH were up-regulated, the expression level of isocitrate dehydrogenase isoenzymes IDH1 and IDH2 were down-regulation. In conclusion, halofuginone can inhibit the proliferation and migration of HepG2 cells and promote apoptosis in a dose-dependent manner, which may be due to the promotion of the aerobic metabolism of cells.

[Analysis of DC-SIGN and DC-SIGNR genetic polymorphism in Chinese Han population].

OBJECTIVE: To understand the genetic polymorphism of DC-SIGN's and DC-SIGNR's neck regions in normal Chinese Han population, and to obtain the genetic data of the two loci in Chinese Han population. METHODS: The genotypes and alleles of repeat sequences of DC-SIGN and DC-SIGNR neck region were typed by PCR, agarose gel electrophoresis and sequencing. Polymorphism information content (PIC) of DC-SIGNR was calculated. RESULTS: DC-SIGN genetic polymorphism was rare. Allele 7 was most and its frequency was 0.9808. 4-, 5-, 6- and 8- alleles were also found, although their frequencies were very low. Caucasians had only 6- and 8- allele mutants; DC-SIGNR genetic polymorphism was high, its PIC was 0.5312, 4-,5-,6-,7-,8-,9- alleles and 16 genotypes were found in normal Chinese Han population. The differences of 6/5,7/4,7/5,7/6,7/7,9/5,9/7,9/9 genotypes distribution and 5-,6-,7-,9- alleles frequency between normal Chinese Han population and Caucasian population were all extremely distinct (P<0.01). The inserted mutation seemed more in Chinese Hans than Caucasian population. CONCLUSION: DC-SIGN and DC-SIGNR genotypes and alleles distribution in Chinese Han population are significantly different from Caucasian population and with Chinese own population genetic characteristics, compared with Caucasians.

[High-flux detection of HBV lamivudine resistant strains by genechip and analysis of mutant hotspots in those strains].

OBJECTIVE: Lamivudine resistant HBV strains in Shenzhen were detected at multiple sites and in large amounts to understand further the distribution of lamivudine resistant mutants. METHODS: 552 Hepatitis B patients's sera were examined using genechip method. Among them, 192 samples of lamivudine resistant mutant were further analyzed. RESULTS: In those 192 lamivudine resistant samples, 191 were YMDD mutants, 124 mutants of codon 528 and 9 mutants of codon 555. 88% YMDD mutants were multi-mutants of YVDD and codon 528; single mutants of YIDD; multi-mutants of YIDD and codon 528. 91% codon of YMDD mutants were GTG, ATT; the other 9% were ATA, ATC. CONCLUSIONS: These results suggest that mutants of codon 552 (YMDD) are core mutants. Mutants of codon 528 and 555 are incidental mutants, YVDD mutants always emerge with mutants of codon 528, but YIDD mutants appear differently. 9% YMDD mutants's codons are ATA or ATC. This may be the reason for the low positive rate shown by using the conventional PCR methods.

[Early indexes to predict the therapeutic effect of interferon on chronic hepatitis B].

OBJECTIVE: To summarize the clinical changing characters of the clinical markers after interferon treatment in chronic hepatitis B (CHB) and make out practical indexes to predict the effect. METHODS: 150 CHB patients were randomly divided into two groups: therapeutic group (90) and control group (60) in the prospective controlled trial. The levels of endogenous interferon before treatment, interferon antibody at the end of the second month and fourth month after treatment, alanine aminotransferase (ALT) and HBV DNA in the serum were detected. Then the data was analysed to find out indexes for predicting the effect. RESULTS: (1) The clearance rate of HBeAg had no significant difference in age except for 20 - 30 and 30 - 40 (t > 2.331 2, P < 0.01). (2) It was more effective if ALT level was higher than 400 U/L before treatment and it decreased more than 50% two months after treatment. (3) The patients whose HBV DNA was negative (dot hybridization) or less than 10(6) copies/ml before treatment had higher rate of HBeAg clearance. (4) There was no effect on patients whose interferon antibody turned positive at the end of the second month. (5)A predictive method of comprehensive factors was made out, whose sensitivity, specificity, and accuracy were 80%, 100% and 90%, respectively. CONCLUSION: The clinical characters of these Chinese patients are different from those of the westerners and the effects of interferon have close relation to the levels of ALT, HBV DNA and interferon antibody.

[Molecular cloning and expression of the severe acute respiratory syndrome-associated coronavirus nucleocapsid protein and its clinical application].

OBJECTIVE: To clone and express nucleocapsid (N) protein of the severe acute respiratory syndrome (SARS)-associated coronavirus, and to evaluate its antigenicity and application value in the development of serological diagnostic test for SARS. METHODS: SARS-associated coronavirus N protein gene was amplified from its genomic RNA by reverse transcript nested polymerase chain reaction (RT-nested-PCR) and cloned into pBAD/Thio-TOPO prokaryotic expression vector. The recombinant N fusion protein was expressed and purified, and its antigenicity and specificity was analyzed by Western Blot, to establish the recombinant N protein-based ELISA for detection of IgG antibodies to SARS-associated coronavirus, and SARS-associated coronavirus lysates-based ELISA was compared parallelly. RESULTS: The recombinant expression vector produced high level of the N fusion protein after induction, and that protein was purified successfully by affinity chromatography and displayed higher antigenicity and specificity as compared with whole virus lysates. CONCLUSION: The recombinant SARS-associated coronavirus N protein possessed better antigenicity and specificity and could be employed to establish a new, sensitive, and specific ELISA for SARS diagnosis.

[Construction of a genechip method for detection of hepatitis B virus DNA, basal core promotor and Pre-C mutants].

BACKGROUND: To establish a genechip method for detection of hepatitis B virus (HBV) DNA, basal core promotor (BCP), and Pre-C mutants. METHODS: This study used two kinds of technology (PCR, oligochip), which can detect five mutant hotspots including nt 1 896, nt 1 899, nt 1 862, nt 1 764 and nt 1 762. The results of genechip method was verified by DNA sequencing. RESULTS: In detecting HBV DNA, the results of genechip were 100% consistent with those of DNA sequencing. In detecting HBV BCP and Pre-C mutants, 146 codons showed the same results using both methods, except for only 4 codons (P greater than 0.05). CONCLUSION: This convenient high throughput genechip method could detect several BCP and Pre-C mutant codons at the same time. These results suggest that genechip method has the same positive rate and specificity with DNA sequencing method. It has more advantages than the latter in detecting mixed mutants and therefore may be used in clinical practice.

Computational Identification of microRNAs and Their Targets / 中国生物工程杂志

The discovery of microRNAs (miRNAs) has introduced a new paradigm into gene regulatory systems. Since inception, computational methods have been an invaluable tool complementing experimental approaches, and many discoveries have been obtained through combination of experimental and computational approaches. The knowledge that has been accumulated about the principles of miRNAs and target recognition were reviewed. The currently available computational methodologies and software for prediction of miRNA and their target genes also have been discussed.

Computational Identification of microRNAs and Their Targets / 中国生物工程杂志

The discovery of microRNAs (miRNAs) has introduced a new paradigm into gene regulatory systems. Since inception, computational methods have been an invaluable tool complementing experimental approaches, and many discoveries have been obtained through combination of experimental and computational approaches. The knowledge that has been accumulated about the principles of miRNAs and target recognition were reviewed. The currently available computational methodologies and software for prediction of miRNA and their target genes also have been discussed.