1.Isolation and specific detection of two major schistosoma gut-associated circulating antigens
Zongli QIAN ; Ping LU ; Zhaojun WANG ; J GOVERT ; Dam VAN ; M.Deelder ANDRÉ
Chinese Medical Journal 2001;114(6):614-617
Objectives To investigate the nature of the common epitopes of Schistosoma japonicum (S. japonicum) circulating anodic (CAA) and circulating cathodic antigen (CCA) and to try to obtain sufficient purified material to set up a standard series for quantitative determinations. Methods Isolation of the two worm fractions from a trichloroacetic acid (TCA) soluble preparation of S. japonicum adult worm antigen (AWAj-TCA) via Mono-Q anion exchange chromatography was performed and analysis of specific reactivity of the eluted fractions was done by antigen-capture Enzyme Linked Immuno Sorbent Assay (ELISA) specific for CAA or CCA with reference to affinity purified preparations of S. mansoni CAA and CCA. Results When an ionic strength gradient was used, CCA was eluted in two major peaks, an unbound fraction CCA-1, and a major bound fraction, CCA-2. Two additional minor peaks, CCA-3 and CCA-4, were eluted at higher ionic strengths. CAA was only detected in the bound fraction, partly overlapping with CCA-3. In the CCA-1 and CCA-2 fractions, reactivity was only found in the antigen-capture ELISA using anti-CCA McAbs both for capture and detection. The CAA fraction was predominantly found to be positive in the antigen-capture ELISA using anti-CAA McAbs both for capture and detection. However, in ELISA using combined anti-CCA and anti-CAA McAbs for capture and detection, this fraction showed some reactivity. Conclusion The two CCA fractions contain molecules which bear at least two CCA-epitopes; the CAA fraction contains molecules which contain at least two CAA-epitopes, and one CCA-epitope.
2.Effect of oxLDL on the uptake and clearance rate of cholesterol in v-SMC originated from human apoA1 transgenic mice.
Ying ZENG ; Gaofeng ZHAO ; Jun QIAN ; Zongli WANG ; Mingpeng SHE
Chinese Medical Journal 2002;115(4):584-588
OBJECTIVETo study the inhibition effect of oxLDL on the uptake and clearance of intra-cellular (3)H-cholesterol in v-SMC from the human-apoA1 transgenic mice (C57BL/6) and the changes in human-apoA1 mRNA expression in v-SMC from human apoA1 transgenic mice after oxLDL stimulation.
METHODSv-SMC originally isolated from human apoA1 transgenic mice connected with a recombined mouse metallothionein-I (MT-I) promoter was used, and the effect of oxLDL on the uptake and clearance of intracellular (3)H-cholesterol was studied in v-SMC of the transgenic and control mice respectively, the study of h-apoA I mRNA expression from v-SMC of the transgenic mice were done by RT-PCR and Northern blot.
RESULTSoxLDL (30 microg/ml) strongly promoted the SMC proliferation. No difference was found in (3)H-cholesterol uptake between nSMC and trSMC, and the uptake rates of both kinds of SMC rose 100% after oxLDL stimulation. The efflux rates of (3)H-cholesterol in trSMC were much higher than those of nSMC (40% - 50%). After oxLDL stimulation, the clearance rates fell by 28% and 10%, respectively, for nSMC and trSMC. The result of RT-PCR and Northern blot showed that h-apoA1 gene expression was markedly increased by the stimulation of oxLDL.
CONCLUSIONExpression of the h-apoA1 gene in C57BL/6 mice enables them to reduce the accumulation of cholesterol in v-SMC. The trSMC can alleviate the harmful effect of oxLDL due to the increase of h-apoA1 expression.
Animals ; Apolipoprotein A-I ; genetics ; metabolism ; Blotting, Northern ; Cell Division ; drug effects ; Cells, Cultured ; Cholesterol ; pharmacokinetics ; Dose-Response Relationship, Drug ; Gene Expression ; Humans ; Lipoproteins, LDL ; pharmacology ; Mice ; Mice, Inbred C57BL ; Mice, Transgenic ; Muscle, Smooth, Vascular ; cytology ; drug effects ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Time Factors