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Objective To investigate the expression of serum amyloid A (SAA) in patients adipose tissue with gestational diabetes mellitus (GDM) and the correlations between SAA and insulin resistance (IR) and body mass index (BMI).Methods A total of 60 single full-term pregnant women underwent cesarean section from June 2013 to December 2013 was enrolled in this study (GDM group,n =30;control group,n =30);serum SAA level was detected with Enzyme-Linked Immunosorbent Assay (ELISA);and mRNA expression of SAA1 in adipose tissue was determined by reverse transcription PCR (RT-PCR);SPSS software was used to compare these markers,and the correlations between SAA and HOMA-IR,BMI were analyzed with Pearson correlation method.Results SAA,mRNA expressions in omental and subcutaneous fat in GDM group (0.447 ± 0.069,0.291 ± 0.067) were significantly higher than those in control group (0.194 ± 0.070,0.231 ± 0.068,P ＜ 0.01).Serum SAA levels [(21.038 ± 6.648) mg/L] and homeostasis model assessment of insulin resistance(HOMA-IR) (4.168± 2.416) in GDM group were significantly higher than those in control group [(14.384 ± 12.770) mg/L,2.045 ± 1.008,P ＜ 0.05];SAA1 mRNA expression levels in omental and subcutaneous fat were positively correlated with serum SAA (r =0.353,0.342,P ＜ 0.01).SAA1 mRNA expression levels in omental were positively correlated to pregestational BMI,late gestational BMI,weight gain in pregnancy and HOMA-IR (r =0.543,0.644,0.340,0.473,P ＜ 0.01),and SAA1 mRNA expression levels in subcutaneous fat were positively correlated to pregestational BMI,late gestational BMI,and HOMA-IR (r =0.788,0.693,0.504,P ＜ 0.01),but was no correlation with weight gain in pregnancy(r =0.013,P ＞ 0.05).Conclusions SAA mRNA expressions in omental and subcutaneous fat in GDM group and serum SAA levels increase,which is positively correlated with BMI and the degree of insulin resistance,SAA may participate in the formation of GDM by increasing insulin resistance.SAA may be used as a new monitor of GDM.
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Objective To construct the pcDNA3. 1-Brugia malayi (Bm) glyceraldehyde-3-phosphate dehydrogenase (GAPDH) eukaryotic recombinant plasmid and to study its effect on mouse cellular immunity response. Methods Total RNA was prepared from periodic Bm. The target gene fragments were amplified by reverse transcription-polymerase chain reaction (RT-PCR) technique and then were inserted into the cloning vector. pGEM-T Easy, and sub-cloned into pcDNA3. 1. Purified pcDNA 3. 1-BmGAPDH recombinant plasmid and CpG were injected into the anterior tibial muscle of BALB/c mice in order to induce host immunity response. Mice injected with PBS and mice injected with blank plasmid were prepared as controls. The mouse models were immunized for 3 times with an interval of 2 weeks. RT-PCR was utilized to detect target gene expression in the muscle tissue. MTT method was used to measure the immunized mice T lymphocytes stimulation index, while enzyme-linked immunoassay (ELISA) was used to determine the serum interleukin (IL)-4 and interferon (IFN)-γ level. Means were compared using t test with SPSS software. Results The recombinant plasmid pcDNA3. 1-BmGAPDH was constructed sucessfully. The target gene was 1020 bp long and its homology with known gene sequence in database was 99%. BmGAPDH gene in the injected muscle of the immunized mice was detected by PCR. The proliferation of spleen T lymphocytes was higher in pcDNA3-BmGAPDH group than in the 2 control groups which were 1. 398, 1. 006 and 1. 017,respectively (P< 0. 05). The levels of IFN-γ and IL-4 in serums from the immunized mice were significantly higher than those of the PBS control group and blank plasmid control group which were 163.905, 58.589, 51. 317 and 107. 906, 27.111, 34.627, respectively (P<0. 05). Immune adjuvant CpG could accelerate and boost antigen-specific immune responses induced by vaccine, which presented as significant increase of IFN-γ and lymphocyte proliferation at 4 weeks after immunization.Conclusion The recombinant eukaryotic plasmid pcDNA3. 1-BmGAPDH is constructed and could elicit cellular immune responses in immunized mice.
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Objective To study the mechanism of killing microfilariae in host by detecting the change of amino acids in Brugia malayi microfilariae. Methods The composition and contents of amino acid in microfilariae of periodic Brugia malayi before and after the effect of cytotoxicity were analyzed with automatic amino acid analyzer. Results The result showed that microfilariae contained 17 kinds of amino acids, but lacked Tryptophan. The total amount of amino acids of the microfilariae after cytotoxicity was lower than that before cytotoxicity( P
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Inthis study, the DNA fragment encoding the regions Ⅰ to Ⅱ of CSP gene from Plasmodium falciparm isolate FCC1/HN was cloned into an. expression vector pcDNA3 contained cytomegalovius (CMV) and transformed into human Hela cell line. The expressed protein PfCSP (condidate vaccine) was used to immunize BALB/c mice by subcutaneous、 intravenous or intraperitoneal administration respectively. The splenocyte of BALB/c mice immumized with the condidate vaccine released significantly IL-2 and IFN-γ following stimulation with this vaccine. It is associated with increase of the splenic T lymphocyte proliferation stimulated by this vaccine and enhancement of NK cell killing activity in the former studies. These results suggested that the vaccine could stimulate T cell response and enhance the cell-mediated immunity.
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The determination of human growth hormor.e (hGH) in urine by means of LAB-ELISA technique was described. The major factors affecting the Sensitivity of LAB-ELISA were studied systematically. The optimized conditions have been established. The sensitivity of LAB-ELISA is at the level of pg/ml. There were little cross binding with human prolactin, thyroid-stimulating hormone, follicular-stimulating hormone, and luteinizing hormone. Human growth hormone levels in overnight urine in 50 normal children and 11 cases of GH-deficiency (GHD) were measurejl by the LAB-ELISA technique. Results of recovery test (recovery rate 100 ?1.4%) and precision test (intra-assay CV 4.72-12.00%, inter-assay CV 4.89-11.17%, and bias 0.20-12.00%)indicate that the method is highly specific,precise and stable.