Phycoerythrin （PE） is one of the most important proteins involved in light capturing during photosynthesis in red algae. Its potential biological activities had gained wide concerns. In the present study, tumor cyto...Phycoerythrin （PE） is one of the most important proteins involved in light capturing during photosynthesis in red algae. Its potential biological activities had gained wide concerns. In the present study, tumor cytotoxic and hydroxyl radical assay were preformed to detect the bioactivity of recombinant PE. Recombinant plasmids pGEX-PE and pBGL were transformed into E.coli BL21 to make two recombinant strains BEX （pGEX-PE） and BGL （pBGL）. PE expressing in BEX （pGEX-PE） was validated by SDS-PAGE and Western blotting analysis. SDS-PAGE analysis indicated that the PE-GST fusion protein was mostly inclusion bodies. Specific expression of PE was confirmed by Western blotting analysis. The recombinant E. coli BEX （pGEX-PE） cells were collected and sonicated. The supernatants were reserved for the tumor cytotoxic experiments. The result of tumor cytotoxic assay indicated that the supernatants containing PE had the activity of inhibiting the growth of Hela cells and with the increase of protein concentration, the inhibiting rate increased from 37.31% to 63.26%, which showed significant difference from the control. Hydroxyl radical scavenging effect was tested with supernatants of BEX （pGEX-PE） and BGL （pBGL） cell lysates treated with sonication and heating. For the sonication samples, the scavenging rates of the supernatants of BEX （pGEX-PE） and BGL （pBGL） cell lysates were significantly higher than the negative control BL2 I（pGEX-4T） （P〈0.02）, and the scavenging rates increased slowly following the increase of the protein content. For the heating samples, except for the 0.2 mgmL^-1 BGL （pBGL） products, the scavenging effects of the supernatants of BEX （pGEX-PE） and BGL （pBGL） cell lysates were stronger than that of negative control BL21（pGEX-4T）. However, the effect intensity was not positively correlated with the increase of the protein concentration. Though a partially decreased hydroxyl radical scavenging activity was led by heating, the biological activity was still retained and conspicuous. This research showed that phycoerythrin protein expressing in E. coli has the potential medical and sanitarian value.展开更多
We studied the rate of lead resoprtion in the gastrointestinal tract of the bird and accumulation of this element by organs and tissues different in their morphology and functions. The minimum lethal lead dosage makin...We studied the rate of lead resoprtion in the gastrointestinal tract of the bird and accumulation of this element by organs and tissues different in their morphology and functions. The minimum lethal lead dosage making 1.5 mg/kg of the live body weight leads to reduction of its mass recovering after its resoprtion in the stomach. Accumulation and elimination of lead from the body are related to dynamics of the body weight. The rate of these processes is in direct dependence and significantly different in various tissues and organs. Claws, kidneys, liver and feathers are characterized by very high rate of element accumulation and fat, heart and muscles - by the lowest rate. Intense elimination of lead from the body starts approximately in 2-2.5 months after the gastrointestinal tract is released from it.展开更多
To clone the gene coding the immunodominant region in the chlamydial protease-like activity factor （CPAF） from Chlaroydophila pneumoniae, to analyze immunoeoropetenee of the expressed protein, and to evaluate its va...To clone the gene coding the immunodominant region in the chlamydial protease-like activity factor （CPAF） from Chlaroydophila pneumoniae, to analyze immunoeoropetenee of the expressed protein, and to evaluate its value in serodiagnosis, the CPAF immunodominant region gene was amplified, ligated into a pGEX6p-2 vector, and then the expressed recombinant protein was purified with glutathione Stransferase （GST） agarose gel FF after renaturation, then identified by SDS-PAGE and Western blot. A new indirect ELISA was developed with the purified protein as coating antigen. The immunogenicity of the recombinant protein was evaluated by immunization to New Zealand rabbits, and its immunoreactivity was analyzed by reacting with anti-C, pneumoniae antibody. 300 clinical sera samples were respectively detected by mieroimmunofluorescenee （MIF） as reference method and the indirect ELISA, and the difference between the two methods was analyzed. Cross-reactivity against Chlamydia trachomatis was investigated with the indirect ELISA to detect anti-C, trachomatis positive antisera. The results indicated that a 51.3 kDa recombinant protein was obtained. Western blot assay proved that the recombinant protein could merely specifically react with human anti- C. pneurnoniae antisera. The titers of the specific IgG antibodies in the immunized New Zealand rabbits were above 1 ： 16 000. Anti- C. pneumoniae IgG positive and negative reference sera were detected with the indirect ELISA, and the concordance rate of negative and positive results were both 100% （40/40）. The sensitivity and specificity of the indirect ELISA in comparison with MIF were 93.8% （45/48） and 100% （252/252） separately by detecting 300 clinical sera samples, and the concordance rate between the two methods was 99.0%. No cross reaction against C. trachomatis was found with the indirect ELISA to detect anti-C, trachomatis positive antisera. In conclusion, the prepared recombinant protein of the CPAF immunodominant region shows excellent immunocompetence and can be used to develop a new indirect ELISA as a method to detect anti-C, pneumoniae antibody for diagnosis of C. pneumoniae infection.展开更多
The soluble HLA-G1 (sHLA-G1) isoform was found to be secreted by trophoblast cells at the matemo-fetal interface, which suggests that it may act as an immunomodulator during pregnancy. In this paper, we reported that ...The soluble HLA-G1 (sHLA-G1) isoform was found to be secreted by trophoblast cells at the matemo-fetal interface, which suggests that it may act as an immunomodulator during pregnancy. In this paper, we reported that GST-sHLA-G1α chain could bind to its receptor ILT-2 on NK92 cells and then the latter recruited Src homology 2 domaincontaining tyrosine phosphatase-1 (SHP-1), which consequently dephosphorylated some important protein tyrosine kinases and blocked the activation of downstream molecules such as MEK and ERK so that the cytotoxicity of natural killer (NK) cells was inhibited. These results indicated that GST-sHLA-G1α chain might be exploited in new immunotherapy strategies aiming at inducing immunotolerance during allograft, xenograft and autoimmune situations. In addition, we found that modification of O-linked β-N-acetylglucosamine (O-GlcNAc) was involved in NK cells' activating and inhibitory signals. This may provide a novel molecular target for inducing immunotolerance but needs further study.展开更多
文摘Phycoerythrin （PE） is one of the most important proteins involved in light capturing during photosynthesis in red algae. Its potential biological activities had gained wide concerns. In the present study, tumor cytotoxic and hydroxyl radical assay were preformed to detect the bioactivity of recombinant PE. Recombinant plasmids pGEX-PE and pBGL were transformed into E.coli BL21 to make two recombinant strains BEX （pGEX-PE） and BGL （pBGL）. PE expressing in BEX （pGEX-PE） was validated by SDS-PAGE and Western blotting analysis. SDS-PAGE analysis indicated that the PE-GST fusion protein was mostly inclusion bodies. Specific expression of PE was confirmed by Western blotting analysis. The recombinant E. coli BEX （pGEX-PE） cells were collected and sonicated. The supernatants were reserved for the tumor cytotoxic experiments. The result of tumor cytotoxic assay indicated that the supernatants containing PE had the activity of inhibiting the growth of Hela cells and with the increase of protein concentration, the inhibiting rate increased from 37.31% to 63.26%, which showed significant difference from the control. Hydroxyl radical scavenging effect was tested with supernatants of BEX （pGEX-PE） and BGL （pBGL） cell lysates treated with sonication and heating. For the sonication samples, the scavenging rates of the supernatants of BEX （pGEX-PE） and BGL （pBGL） cell lysates were significantly higher than the negative control BL2 I（pGEX-4T） （P〈0.02）, and the scavenging rates increased slowly following the increase of the protein content. For the heating samples, except for the 0.2 mgmL^-1 BGL （pBGL） products, the scavenging effects of the supernatants of BEX （pGEX-PE） and BGL （pBGL） cell lysates were stronger than that of negative control BL21（pGEX-4T）. However, the effect intensity was not positively correlated with the increase of the protein concentration. Though a partially decreased hydroxyl radical scavenging activity was led by heating, the biological activity was still retained and conspicuous. This research showed that phycoerythrin protein expressing in E. coli has the potential medical and sanitarian value.
文摘We studied the rate of lead resoprtion in the gastrointestinal tract of the bird and accumulation of this element by organs and tissues different in their morphology and functions. The minimum lethal lead dosage making 1.5 mg/kg of the live body weight leads to reduction of its mass recovering after its resoprtion in the stomach. Accumulation and elimination of lead from the body are related to dynamics of the body weight. The rate of these processes is in direct dependence and significantly different in various tissues and organs. Claws, kidneys, liver and feathers are characterized by very high rate of element accumulation and fat, heart and muscles - by the lowest rate. Intense elimination of lead from the body starts approximately in 2-2.5 months after the gastrointestinal tract is released from it.
文摘To clone the gene coding the immunodominant region in the chlamydial protease-like activity factor （CPAF） from Chlaroydophila pneumoniae, to analyze immunoeoropetenee of the expressed protein, and to evaluate its value in serodiagnosis, the CPAF immunodominant region gene was amplified, ligated into a pGEX6p-2 vector, and then the expressed recombinant protein was purified with glutathione Stransferase （GST） agarose gel FF after renaturation, then identified by SDS-PAGE and Western blot. A new indirect ELISA was developed with the purified protein as coating antigen. The immunogenicity of the recombinant protein was evaluated by immunization to New Zealand rabbits, and its immunoreactivity was analyzed by reacting with anti-C, pneumoniae antibody. 300 clinical sera samples were respectively detected by mieroimmunofluorescenee （MIF） as reference method and the indirect ELISA, and the difference between the two methods was analyzed. Cross-reactivity against Chlamydia trachomatis was investigated with the indirect ELISA to detect anti-C, trachomatis positive antisera. The results indicated that a 51.3 kDa recombinant protein was obtained. Western blot assay proved that the recombinant protein could merely specifically react with human anti- C. pneurnoniae antisera. The titers of the specific IgG antibodies in the immunized New Zealand rabbits were above 1 ： 16 000. Anti- C. pneumoniae IgG positive and negative reference sera were detected with the indirect ELISA, and the concordance rate of negative and positive results were both 100% （40/40）. The sensitivity and specificity of the indirect ELISA in comparison with MIF were 93.8% （45/48） and 100% （252/252） separately by detecting 300 clinical sera samples, and the concordance rate between the two methods was 99.0%. No cross reaction against C. trachomatis was found with the indirect ELISA to detect anti-C, trachomatis positive antisera. In conclusion, the prepared recombinant protein of the CPAF immunodominant region shows excellent immunocompetence and can be used to develop a new indirect ELISA as a method to detect anti-C, pneumoniae antibody for diagnosis of C. pneumoniae infection.
文摘The soluble HLA-G1 (sHLA-G1) isoform was found to be secreted by trophoblast cells at the matemo-fetal interface, which suggests that it may act as an immunomodulator during pregnancy. In this paper, we reported that GST-sHLA-G1α chain could bind to its receptor ILT-2 on NK92 cells and then the latter recruited Src homology 2 domaincontaining tyrosine phosphatase-1 (SHP-1), which consequently dephosphorylated some important protein tyrosine kinases and blocked the activation of downstream molecules such as MEK and ERK so that the cytotoxicity of natural killer (NK) cells was inhibited. These results indicated that GST-sHLA-G1α chain might be exploited in new immunotherapy strategies aiming at inducing immunotolerance during allograft, xenograft and autoimmune situations. In addition, we found that modification of O-linked β-N-acetylglucosamine (O-GlcNAc) was involved in NK cells' activating and inhibitory signals. This may provide a novel molecular target for inducing immunotolerance but needs further study.
