Donor MHC gene to mitigate rejection of transplantation in recipient mice / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Donor organ rejection continues to be a significant problem for patients receiving transplants. We therefore tested whether transferring a donor's major histocompatibility complex (MHC) gene to the recipient would mitigate the rejection of transplanted hearts in mice.</p><p><b>METHODS</b>H-2K(k) gene from donor mice was amplified using nested polymerase chain reaction (PCR) and ligated into a mammalian expression vector, which was then transfected into thymus ground mass cells collected from the recipients. Clones stably expressing the transgene were then injected into the recipients' thymus visualized using ultrasound. Control mice were administered cells previously transfected with empty vector. Following heart transplantation, cardiac activity was monitored electrocardiographically. Recipient thymus cells were tested for MHC antigenicity using flow cytometry and spleen cells were subjected to mixed lymphocyte culture tests. Finally, the transplanted hearts were sectioned, stained and examined under light microscopy.</p><p><b>RESULTS</b>Southern analysis following nested PCR revealed clear expression of H-2K(k) gene. Following transplantation, electrocardiosignals were detectable highly significantly longer in recipients administered thymal cells expressing donor H-2K(k) than in those receiving control cells. Flow cytometric analysis using an anti-H-2K(k) antibody confirmed its expression in H-2K(k) treated recipients but not in control mice. Mixed lymphocyte cultures containing H-2K(k) treated cells showed significantly less proliferation than those containing control cells. Hearts from control mice showed substantially greater lymphocyte infiltration than those from H-2K(k) treated mice and large areas of necrosis.</p><p><b>CONCLUSION</b>Rejection of transplanted hearts can be mitigated substantially by introducing the donor's MHC into the recipient.</p>

Expression and Biological Characterization of Human Osteoprotegerin Fused with Mycobacteria Heat Shock Protein 65 / 中国生物工程杂志

A fused functional gene of human OPG and Mhsp65 was amplified by PCR,and cloned into the prokaryotic expression vector pET-28a.The BL21(DE3) strain of E.coli was transformed using the recombinant plasmid pET-28a-OPG-HSP65 and the expected protein was expressed by induction with IPTG.Result of SDS-PAGE indicated that the expected recombinant protein of 23 kDa was expressed with high yield as inclusion body.The fusion protein could be specifically recognized by both the anti-His antibody and anti-human OPG monoclonal antibody in Western blot analysis.The purified and refolded fusion protein could inhibit osteoclast proliferation and bone absorption in vitro.The results of mouse ear swelling assay and expressions of TNF-?,IFN-? and IL-17 mRNAs detected by real-time quantitative PCR demonstrated that the fusion protein had an anti-inflammation activity.The results suggest that the fusion protein of human OPG and Mhsp65 may act as a potential therapeutics for rheumatoid arthritis.