ABSTRACT
Objective:To evaluate the immune response efficiency of the recombinant abrin B chain protein (rATB) in BALB/c mice.Methods:BALB/c mice were vaccinated intraperitoneally (i.p.) with the purified rATB protein.ELISA was used to detect the IgG,IgG1 and IgG2a.Meanwhile,the expression of IL-4 and IFN-γwere detected by flow cytometry.Then the protein neutralization assay and efficacy of passive protection were done .Results:The anti-rATB antibody was detected by ELISA after vaccinated the rATB in the mice and results showed that the highest titer reached 106 ,which was acquired after the last vaccination .Meanwhile,a strong secondary response was triggered in mice when challenged with toxin .In contrast ,the antibody response of the PBS-vaccinated mice was less than 1:10 and P<0.05 indicated obvious difference between test group and control group .The result of IgG 1 was the same as the IgG ,while the result of IgG2a had not changed.There was significant difference for IL-4 between two groups (P<0.05),while no significant difference for IFN-γwas observed.All BALB/c mice survived after challenged with 5×LD50 of AT.Transfusion of sera from immunized mice provided passive protection in naive mice.Furthermore, the rATB could induce specific neutralizing antibodies against abrin toxin.Conclusion: The recombinant protein can induce the Th 2-type immune response and trigger a good immune response and protective efficacy,which means it may be a promising vaccine candidate against human exposure to AT .
ABSTRACT
Objective:To explore the immunity provided to BALB/c mice by immunization with the recombinant ricin B chain protein (rRTB). Methods:Female BALB/c mice were randomly selected into rRTB-vaccinated group and PBS group. Mice were subcu-taneously (s. c. ) injected four times with 14 days immunization time interval. Changes of antibodies (IgG,IgG1and IgG2a) in serum were detected by ELISA. Meanwhile,the expression of IL-4 and IFN-γ were detected by flow cytometry. Results:The mean IgG titers reached 106 after the fourth immunization and a strong secondary response was induced in vaccinated mice when challenged with toxin. There was significant difference between rRTB-vaccinated group and PBS group ( P<0. 05 ) . The same result was shown in IgG1. However,no changed was detected in IgG2a. Meanwhile,there was significant difference for IL-4 between two groups (P<0. 05), while no significant difference for IFN-γwas observed. Conclusion:rRTB can produce higher levels of antigen-specific antibodies ( IgG and IgG1),and cytokine (IL-4) of splenocytes,which means the recombinant protein can induce the Th2-type immune response and trigger a good immune response. rRTB may be a potentially valuable vaccine candidate against human exposure to AT.
ABSTRACT
αB-crystallin is the structural protein of vertebrate lens, w hich is w idely expressed in non-lens tissue. As one of the heat shock protein fam ily m em bers,αB-crystallin possesses biological proper-ties of m olecular chaperones and anti-apoptotic effects. Multi-factor injuries, such as retinopathy, inflam-m ation and nervous system diseases, have a closely relationship w ith αB-crystallin. T his paper review s the research progress of the expression and m echanism ofαB-crystallin in retina and extraocular tissues and organs.
ABSTRACT
Background: Transgenic plants inhabiting single Bt gene are prone to develop insect resistance and this resistance has been reported in case of some important yield-devastating insect larvae of commercial crops, such as cotton and rice. Therefore, it has become essential to adapt new strategies to overcome the problem of insect resistance and these new strategies should be sophisticated enough to target such resistant larvae in broad spectrum. Among these, plants may be transformed with Bt gene tagged with some fusion-protein gene that possesses lectin-binding capability to boost the binding sites for crystal protein gene within insect mid-gut in order to overcome any chances of insect tolerance against Bt toxin. Enhanced chloroplast-targeted Bt gene expression can also help in the reduction of insect resistance. Results: In the present investigation, a combined effect of both these strategies was successfully used in cotton (G. hirsutum). For this purpose, plant expression vector pKian-1 was created, after a series of cloning steps, carrying Cry1Ac gene ligated with chloroplast transit peptide towards N-terminal and Ricin B-Chain towards C-terminal, generating TP-Cry1Ac-RB construct. Conclusions: Efficacy of pKian-1 plasmid vector was confirmed by in-planta Agrobacterium-mediated leaf GUS assay in tobacco. Cotton (G. hirsutum) local variety MNH-786 was transformed with pKian-1 and the stable integration of TP-Cry1Ac-RB construct in putative transgenic plants was confirmed by PCR; while fusion-protein expression in cytosol as well as chloroplast was substantiated by Western blot analysis. Whereas, confocal microscopy of leaf-sections of transgenic plants exposed that hybrid-Bt protein was expressing inside chloroplasts.
Subject(s)
Chloroplasts/genetics , Chloroplasts/metabolism , Plants, Genetically Modified , Chloroplast Proteins/isolation & purification , Ricin/analysis , Protein Sorting Signals , Blotting, Western , Cloning, Molecular , Microscopy, Confocal , Agrobacterium , Chloroplast Proteins/genetics , InsectaABSTRACT
AIM: To determine whether the high mobility group protein I (HMGI) is able to bind to the upstream sequence of platelet-derived growth factor B-chain gene and to characterize the HMGI-binding AT-rich regions. METHODS: Recombinant human HMGI (rhHMGI) protein was prepared and electrophoresis mobility shift assay (EMSA) was used. RESULTS: The binding of rhHMGI to PDGF-B (-1 758 / +43 bp) was observed in vitro. Two major HMGI-binding fragments -1 392 / -1 180 bp and -188 / +43 bp were identified, which contained the same AT-rich sequence TTTATAAA (-1 333 / -1 326 bp, -1 314 / -1 307 bp and -30 / -23 bp). An oligonucleotide bound to the TTTATAAA and the GAGACC, the core sequence of the shear stress response element of the PDGF-B, could also bind to the HMGI. Furthermore, HMGI facilitated the binding of NF-?B to the GAGACC in the oligonucleotide. CONCLUSION: The HMGI could bind to the upstream sequence of the PDGF-B gene via the AT-rich sequence TTTATAAA, which may play a role in the transcriptional regulation of the PDGF-B gene.