| Na+/H+ exchanger(NHE) is widely present in the plasma transmembrane proteins, with pHi set point, determine intracellular pHi levels of intracellular homeostasis, regulating cell behavior and development as well as the occurrence of tissue injury. There are nine isoforms of NHE protein with varying tissues and cellular distribution. The most clearest is NHE1,NHE1 not only have the functions but also plays an important role in tissue injury, especially in the myocardium.In 2006, chicken NHE1(chNHE1) proved to be ALV-J receptor.Subgroup J avian leukemia virus(ALV-J) is an RNA virus, and it can cause tissue injury, growth inhibition, performance degradation or tumors. However, due to its hypermutation and the study of its infection mechanism is not clearly now,so it lead to widely complex disease and brings serious harm to our poultry industry. ALV-J infect host cells through specfic interactions between viral Env and cell receptor of chNHE1, and then causing tissue damage and reducing produce performance or other conditions. The Env directly bonded the first extracellular loop1 of chNHE1, then Env transmembrane domain(TM) has change conformationa to promote fusion of the virus and cells, this is the key step to virus enters the cell.chNHE1 first extracellular loop 1(ECL1) is the maximum outer membrane, and therefore ECL1 is a key domain between ALV-J and chNHE1. The viral infection can be blocked by blocking cell receptor.Thus the chance of viral infection are likely to be reduced.Thereby reduce the extent of tissue injury. Here is a question, whether chNHE1 play a decisive role in ALV-J-induced tissue damage? This issue deserves further study.This study was prepared antibody of chNHE1 first outer membrane to detect the blocking effect of ALV-J infection and tissues protection, so as to clarify the decisive role chNHE1 in ALV-J-induced tissue injury.To explore this question, we obtained ECL1 gene from normal SPF chicken kidney and prepared receptor antibody(anti-ECL1). We set up four groups: A(Normal group), B(Anti-ECL1 group), C(ALV-J group), D(ALV-J + Anti-ECL1 group); the Anti-ECL1 antibody by cell toxicity testing to ensure that no damage to the cell case, and madeanti-ECL1 antibody and ALV-J incubating at the same time on the DF-1 cells. We were detected the blocking results by quantitative PCR and ELISA. The test results show that in48 h, ALV-J transcription level of Group D was significantly lower than group C, the level of ALV-J transcription of Group D had no significant difference with group C as the time goes on; ELISA detection of ALV-J antigen p27 expression level within 72 h was significantly lower than group C, proofed Anti-ECL1 antibody can effectively block the ALV-J infection for a limited time. we injected anti-ECL1 antibody to ALV-J-infected chickens to detection whether anti-ECL1 antibody play a crucial role to ALV-J infected chicken tissues and organs.relative quantitative PCR results showed that compared with group C ALV-J transcription level had a significant reduction in group D, ALV-J antigen expression level reducing in group D, ALV-J transcription level significantly lower in group D in the liver, but ALV-J antigen expression had no significant change, the results show that anti-ECL1 antibody has a decisive effect in ALV-J infected chicken heart and liver, but other tissues or organs has no significant protective impact. Furthermore, in order to detect chNHE1 expression in immune organs of ALV-J infections chicken, we were used immunohistochemistry to detect chNHE1 expression in bone marrow, thymus, spleen and bursa of Fabricius of ALV-J infected chicken, and we found that chNHE1 up-regulation, chNHE1 expression showed significantly in the bone marrow myeloid cell lines.Currently in the field of human medical research shows, NHE1 is a contributing factor to ischemia / reperfusion or myocardium injury or heart failure. Many scholars to developed NHE1 inhibitor or chemical drugs to reduce the degree of myocardial injury, to ensure personal safety.This study also provide some reference for this work.Furthermore ECL1 is the key domain of ALV-J receptor, but whether ALV-J has a co-receptor or other cofactors are still in the study.In summary, this study shows that anti-ECL1 polyclonal antibody can effectively block ALV-J infection in a limited time; anti-ECL1 antibody have a significant protective effect in myocardium and liver, it also proved ECL1 domain is not the only binding site to ALV-J infected cells; the anti-ECL1 antibody has protection to organizations is also based on the ALV-J targeted level infifferent organization; chNHE1 expression were up-regulated in infected ALV-J chicken immunohistochemistry organ, especially on myeloid cell lines inmyeloid in bone marrow; chNHE1 play a decisive role in ALV-J-induced tissue injury; thisdiscovery for the present study human medicine myocardial damage caused by various diseases and laid part of the reference; also is provides a new way of thinking and methods to the control ALV-J virus and other related virus. |
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