Si-qi Lu

Capital Medical University, Peping, Beijing, China

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Publications (10)2.28 Total impact

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    ABSTRACT: To inhibit the expression of pyruvate kinase (PK) mRNA in Giardia lamblia by specific hammerhead ribozyme. The constructed hammerhead-GCV vector (pGCV-PKH) which aims to PK mRNA was electroporated into G. lamnblia trophozoites (group A). Electroporated trophozoites (group B) and normal trophozoites (group C) served as control Trophozoites in each group were collected at 24, 48, 72 and 96 h post-electroporation, respectively. The concentrations of trophozoites were calculated and the growth curves were constructed. At 24, 48, 72 and 96 h post-electroporation, mRNA of each group was detected by RT-PCR and real-time PCR, respectively. The PK activity was tested by ultraviolet spectrophotometry. The growth curve showed that the growth of trophozoites was considerably depressed after 96 h post-electroporation. RT-PCR result displayed that the specific ribozyme mRNA was detected in group A from 24 h to 96 h post-electroporation. At 24 and 48 h after transfection, the PK mRNA level of group A decreased to 5% (5 +/- 0.17) and 8% (8 +/- 0.19) of the level in group C, respectively; and the PK activity of group A decreased to 32% (32 +/- 0.64) and 38% (38 +/- 0.65) of the level in group C. PK mRNA expression in G. lamblia has been inhibited by specific hammerhead ribozyme.
    Zhongguo ji sheng chong xue yu ji sheng chong bing za zhi = Chinese journal of parasitology & parasitic diseases 08/2010; 28(4):257-60.
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    ABSTRACT: To construct a recombinant vector of Giardia canis virus (GCV)-specific hammerhead ribozyme of pyruvate kinase (PK) in Giardia lamblia. Using RNA draw software, the secondary structure of PK gene in Giardia was predicted and the cleavage site of ribozyme was selected. The antisense-specific hammerhead ribozyme (PKH) targeting this site was designed. The DNA encoding the ribozyme was synthesized and the recombinant vector pGCV-PKH was constructed. The extracellular and intracellular cleavage of PK mRNA was carried out with the linear recombinant vector. Trophozoites in each group were observed with fluorescent microscopy at 24th hour after transfection. Real-time PCR was used for relative quantitative analysis of cleavage products. The recombinant vector of GCV-specific hammerhead ribozyme of pyruvate kinase in Giardia lamblia (pGCV-PKH) was constructed. Intracellular cleavage assays showed that the green fluorescence could only be seen in pGCV-GFP transfected trophozoites. The relative content of PK mRNA in pGCV-PKH transfected group was 33.14% of that in normal control group. It could cleave PK mRNA extracellularly and the efficiency was 58.5%. The recombinant vector can transfect Giardia trophozoites, and cleave mRNA of PK intracellularly and extracellularly.
    Zhongguo ji sheng chong xue yu ji sheng chong bing za zhi = Chinese journal of parasitology & parasitic diseases 04/2010; 28(2):98-102, 138.
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    International Journal of Infectious Diseases - INT J INFECT DIS. 01/2009; 13.
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    ABSTRACT: Giardia lamblia is an early branching eukaryotic microorganism that derives its metabolic energy primarily from anaerobic glycolysis. In most organisms, glycolysis is catalyzed by pyruvate kinase (PK), allowing the generation of two ATP molecules from one molecule of pyruvate. Giardia has both PK and pyrophosphate-dependent pyruvate phosphate dikinase (PPDK), which catalyzes the generation of five ATP molecules from pyruvate by pyrophosphate-dependent glycolysis and offers a potential selective advantage. In order to evaluate the importance of pyrophosphate-dependent glycolysis, we used ribozyme-mediated cleavage of the PPDK transcript to decrease PPDK transcript levels to 20% of normal. The accompanying decrease in PPDK enzyme activity decreased ATP levels to 3% of normal and increased glycogen deposition, confirming the importance pyrophosphate-mediated glycolysis that was previously suggested by cell lysate studies. PPDK is not found in vertebrates, so specific inhibitors may be useful for treatment of infections caused by anaerobic protists that depend on pyrophosphate-dependent glycolysis.
    Biochemical and Biophysical Research Communications 04/2008; 367(2):394-8. · 2.28 Impact Factor
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    ABSTRACT: To analyze the genetic characteristics of two rabies virus isolates from Henan province and to compare their relations hip with known rabies virus isolates and vaccine strains. Rabies viral antigens were detected in 100 canine brains by immunofluorescence assay method. Rabies virus was isolated through inoculating the suspensions of positive brains into suckling mice. N gene and G were amplified by RT-PCR and sequenced. Phylogenetic trees were constructed for the analysis on genetic characteristics of rabies virus. Two rabies virus strains were isolated (Henan Hb1 and Henan Sq1). Data from sequential comparison revealed that the nucleotide and amino acid identities of N and G gene between the two isolates were 99.3% and 98.9%, and 98.7% and 98.4% respectively. The two isolates were more closely related to CTN, with the homogeny of N gene and G gene as 89.1% and 85.6%-85.7% at the nucleotide level, but 97.6%-98.0% and 92.3% at the amino acid level respectively, than to other vaccine strains. When comparing with other known viruses including Chinese isolates, the two stains shared closer identity with the isolates from Indonesia, and the rates of homogeny of N and G gene were 92.1%-93.2% and 91.9%-92.1% at the nucleotide level, 97.5%-98.6% and 96.0%-96.2% at the amino acid level, respectively. Data from the deduced amino acid sequences revealed that some amino acid residues including the residues in the N and G antigenic sites were substituted in the two isolates. Furthermore, phylogenetic analysis showed that the two isolates were also more closely related to the strains from Indonesia and vaccine strain CTN than to any other known street viruses and vaccine strains. Both Henan Hbl and Henan Sql belonged to genotype 1. However, the N and G gene diverged from known street viruses and vaccine strains at either nucleotide level or amino acid level.
    Zhonghua liu xing bing xue za zhi = Zhonghua liuxingbingxue zazhi 04/2007; 28(3):261-7.
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    ABSTRACT: To establish genetic method in detecting Cryptosporidium parvum and Giardia lamblia which often coinfected with AIDS patients. Cryptosporidium oocysts and Giardia cysts were isolated and purified from fecal samples of the individuals infected with C. parvum and G. lamblia, respectively. Genomic DNAs were extracted. Two pairs of specific primers were designed or synthesized according to the 18S rRNA gene from C. parvum or the triose phosphate isomerase (tim ) gene from G. lamblia. Polymerase chain reaction(PCR) technique was used to amplify the DNA samples from the oocysts and the cysts, and those from the 6 control samples, including Schitosoma japonicum, Toxoplasma gondii , Entamoeba histolytica, Trichinella spiralis, Trichomonas vaginalis and human blood cells. DNA samples from 30 fecal samples of AIDS patients were detected with the same method. One fragment of 500 bp was amplified with the primer of C. parvum, and the other one of 683 bp was amplified with the primer of G. lamblia. Twenty pg and 0.4 pg DNA of C. parvum and G. lamblia could be detected separately. The specificity of these two pairs of PCR primers was confirmed by the failure in the amplification of the control DNA samples. Out of 30 cases of AIDS patients, 7 showed C. parvum positive, while non Giardia was detected. Genetic detection method for C. parvum and G. lamblia detection was established which was more sensitive and specific.
    Zhonghua liu xing bing xue za zhi = Zhonghua liuxingbingxue zazhi 11/2006; 27(10):884-8.
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    ABSTRACT: Trophozoites of Giardia lamblia were axenically cultivated with modified TYI-S-33 medium contained 500 microg/ml metronidazole (12h LC50). The morphology of drug-treated trophozoites was observed with light and electron micro-scopes at 2, 4, 8, 12 h respectively. The light microscopy revealed that the trophozoites treated with MTZ showed swollen, detached from the wall of the culture tube, and were with vacuoles in the cytoplasm. Movement of the flagella become slowly or stopped. Electronic microscopy showed that the trophozoites were swollen and deformed; lots of vacuoles were seen in the cytoplasm; the contents of cytoplasm were depleted and the nuclei deformed. This study indicated that MTZ has injured the morphology of G. lamblia.
    Zhongguo ji sheng chong xue yu ji sheng chong bing za zhi = Chinese journal of parasitology & parasitic diseases 11/2006; 24(5):387-8.
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    ABSTRACT: To study the in vitro effect of dihydroartemisinin (DHA) on Giardia lamblia. Trophozoites of G. lamblia were cultivated with modified TYI-S-33 medium that contains dihydroartemisinin (DHA). The trophozoites were morphologically observed respectively with light and electron microscopes after treated with the drug. The mortality increased with the prolongation of the time of the drug action and the increase of drug concentration (P<0.01). While at the same concentration of 100 microg/ml, the mortality increased from 46.6% for 12 h to 100% for 24 h (P<0.05). For 12 h, the mortality of G. lamblia was from 46.6% at concentration of 100 microg/ml to 100% at 200microg/ml. Under optical microscope, deformation and swelling of the parasites were observed when treated with DHA for 12, 24 and 48 h. Movement of the flagella became slow or stopped. Under electron microscope, the trophozoites were swollen and deformed, vacuoles were seen in the cytoplasm, and the cell membrane ruptured and fell off. The cytoplasm protrusions appeared on the surface of the plasma membrane. The adhesive disc changed into large bubbles and the perinuclear space became wider and the deformed nucleus was seen. DHA shows a strong impairment on the plasma membrane and cytoskeleton of Giardia lamblia.
    Zhongguo ji sheng chong xue yu ji sheng chong bing za zhi = Chinese journal of parasitology & parasitic diseases 10/2005; 23(5):292-5.
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    ABSTRACT: To evaluate the protective efficacy against a lethal dose of E. coli O157:H7 after intranasal, oral and subcutaneous immunization with the outer membrane protein (OMP) extracted from the whole cell of E. coli O157:H7. Female BALB/C mice were immunized three times (on days 0, 7 and 14) with OMP and CT or Complete Frund adjuvant. On the 21st day after the last immunization, serum, fecal extracts and vaginal washes were collected for the detection of antigen-specific antibody responses by ELISA before the oral challenge with E. coli O157:H7 933. And the antigens that induced the specific antibody responses were analysed by Western-blotting. Then, the mice were orally challenged with 0.3 ml 4.25 x 10(10)/ml live E. coli O157:H7, and the mortality was recorded. On the 7th day after the challenge, the mice were sacrificed and the heart, liver, lung, kidney, small intestine and colon were collected. Then the histology lesions were observed by light microscopy. In ELISA, both intranasal and oral immunization, with CT as a mucosal adjuvant, induced strong anti-OMP IgA responses in serum, fecal extract and vaginal washes, and anti-OMP IgG responses in serum. However, the intranasal immunization was much more effective to induce specific IgA and IgG responses than the oral immunization. In contrast to mucosal immunization, subcutaneous immunization only induced high levels of specific IgG antibodies in serum, and did not effectively promote IgA immune response. The results of the protective efficacy after challenge showed that both intranasal and oral immunizations with OMP provided significant protection (86.7% to 40%, P < 0.01 and 73.3% to 40%, P < 0.05) against a lethal dose of E. coli O157:H7 challenge, and intranasal immunization possessed a better protective ability (86.7% to 73.3%, P < 0.05). In contrast, the mice immunized subcutaneously were not protected. They were died more early after oral challenge. Furthermore, the histopathological features of the kidney, colon and other organs also appeared to be well correlated with immunization route. In comparison with the mice immunized subcutaneously and those in control group, the severity of the histopathological lesions in the tissues of the mice immunized intranasally was minimal. These results suggested that the intranasal immunization should be the best choice of vaccine development against E. coli O157:H7.
    Zhonghua yi xue za zhi 01/2004; 84(1):58-62.
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    ABSTRACT: AIM: T o d e t e c t a n d o b s e r v e t h e e f f e c t o f dihydroartemisinin on the cytoskeleton of Giardia lamblia in vitro.