ABSTRACT
Objective] To explore the humoral and cellular immune responses in mice to eukaryotic expression recombinant plasmid encoding histidine rich protein 2 (HRP\|Ⅱ) of Plasmodium falciparum. [Methods] The start and stop codes were introduced into HRP\|Ⅱ gene fragment, the reading frame and the position of start and stop codes in HRP\|Ⅱ were identified by sequencing. HRP\|Ⅱ fragment containing the start and stop codes was cloned into pcDNA3 1(\|) to form pcDNA3 1(\|)/HRP\|Ⅱ. The BALB/c mice were immunized i.m. with the plasmids for 3 times in 3 weeks intervals. Two weeks after the last immunization, the sera and splenocytes were collected to investigate anti\|HRP\|Ⅱ antibodies by ELISA and the splenocytes proliferation response to HRP\|Ⅱ. [Results] Sequence data show that the reading frame and the position of start and stop codes are correct. Restriction enzyme digestion indicated that the HRP\|Ⅱ gene fragment containing start and stop codes was successfully cloned into pcDNA3 1(\|). Mice raised significant anti\|HRP\|Ⅱ antibodies after pcDNA3 1(\|)/HRP\|Ⅱ immunization, and the splenocytes proliferated prominently when stimulated with HRP\|Ⅱ protein. [Conclusion] Eukaryotic expression recombinant plasmid \{encoding\} HRP\|Ⅱ gene can induce significantly humoral and cellular immune response in mice. HRP\|Ⅱ gene may be a good candidate for P.falciparum blood\|stage multiple DNA vaccine.
ABSTRACT
AIM:To construct hybrid nucleic acid vaccines which contain Plasmodium falciparum merozoite surface protein 1 (MSP1 ) block 1 7gene and gene fragment coding for several T cell epitopes. METHODS:MSP1 block 1 7gene of FUP strain was connected with a polyvalent gene fragment which contains several T cell epitopes from MSA1 ,MSA2 ,RESA,IL- 1 and Tetanus toxin,the connected gene fragment (hybrid gene,HG) was cloned into eukaryotic expression plasmid p CDNA3.1 (- ) ,VR1 0 1 2 for intracellular expression,VR1 0 1 2 /TPA for secretion,the recombinants were identified by PCR and restriction enzyme digestion. RE- SUL TS &CONCLUSION:The hybrid nucieic acid vaccines:VR1 0 1 2 /HG,VR1 0 1 2 /TPA/ HG and p CDNA3.1 /HG were successfully constructed.
ABSTRACT
Two pairs of primers specific to small subunit ribosomal DNA of Plasmodium falciparum were designed and the expected SSUrDNA fragment was amplified for detecting P.falciparum infection with double-ternperature-nested polymerase chain reaction using DNA prepared by boiling method. The results showed that the nested PCR could amplify a constant size of desired SSUrDNA fragment of P. falciparum which was further confirmed by digestion of restrlction endonuclease and could detect parasitemia level of 0. 8 ? 10-6. It has great potentials for identifying Plasmodium species in ring form of erythrocytic stage and detecting mixed Plasmodium infections. Therefore, it is suggested that this method is sensitive, accurate, simple and rapid in detecting Plasmodium falciparum in blood samples for malaria diagnosis.
ABSTRACT
Aim To investigate the anti-arrhythmic effect and mechanism of ?-opioid receptor during myocardial ischemia and reperfusion in rats,and to initially determine the regulation of U50488H(U50,a selective ?-opioid receptor agonist) to angiotensinⅡ(AngⅡ),endothelin(ET) and nitric oxide(NO) in rats.Methods Rats were randomly divided into 7 groups,i.e.,control group,ischemia/reperfusion group(I/R),U50488H+I/R group,PTX group(PTX,a Gi/o proteininhibitor),Glib group(glibenclamide,a K_(ATP) channel blocker),Che group(chelerythrine,a selective PKC inhibitor),and Gen group(Genistein,a Tyrosine kinase inhibitor) respectively.The arrhythmia occurrence and score in different groups were observed and counted.The contents of AngⅡ,ET and NO in plasma of rats were also examined.Results ① Compared with I/R group,the arrhythmia score of U50+I/R group was significantly decreased.The effect of pared with I/R group,the arrhythmia score of U50+I/R group was significantly decreased.The effect of glibenclamide and chelerythrine respectively,the anti-arrhythmic effects induced by U50488H in the rats during myocardial ischemia and reperfusion were significantly attenuated or even completely blocked.③ The anti-arrhythmic effects of U50488H were not significantly affected by pretreatment with genistein.④ In comparison with normal rats,the contents of AngⅡand ET in plasma of I/R group were significantly increased,but the content of NO was decreased.With the administration of U50488H,the contents of AngⅡand ET in plasma of rats in U50488H+I/R group were significantly decreased.Meantime the content of NO was increased.Conclusions ① U50488H-induced anti-arrhythmic effects in the rats with myocardial ischemia and reperfusion are mediated by ?-opioid receptor.The signaling pathway may be related with(Gi/o,) PKC,and K_(ATP) channel.② The activation of ?-opioid receptor may elicit anti-arrhythmic effect through the down-regulations of AngⅡ or ET and up-regulation of NO in plasma of rats.