Autophagy Of Peripheral Blood CD4～+ T Lymphocyte In Asthma

Mature T lymphocytes were divided into subset of CD4~+ T and CD8~+T lymphocyte, and CD4~+ T lymphocyte could differentiate into Th1, Th2 cells. Activated CD4~+ T lymphocyte induced recruitment of multiple leucocyte into airway from blood, and provoke mediators of inflammation releasion of eosinophils, mast cells and lymphocytes. Furthermore, activated Th2 cells could show Th2 dominance response by releasing IL-4, 5, 13, which concerning with asthma, the immunity disbalance disease. Regulation of Th1/Th2 balance could make help in asthma prevention and treatment.Spinozzi reported that reduction of lymphocyte apoptosis was one reason of inflammation persistence in asthma airway. Therefore, CD4~+ T lymphocyte anergy and apoptosis induction was believed kind of progress on regulation of Th1/Th2 balance.Now, it's proved that programmed cell death (PCD) included not only apoptosis but also autophagy, oncosis and autoschizis, and regulation network may be hidden among them. Therefore, further study on PCD of CD4~+ T lymphocyte could make progress on Th1/Th2 disbalance proceed, and make further step in new treatment.Autophagy, the so called"PCD type II", was named for the widespread existing degradation/recycle system in eukaryocyte, in which macromolecule and organells degradation were carried out by regulation of amino acid and hormone, and degradation product turned into raw material of de novo synthesis of protein and organells. Sometimes, autophagy could protecte the cell from apoptosis and necrosis. Many irritations, such as oxidation, ischemia/reperfusion, TNF-α,FasL, calcium overload and so on, could lead to apoptosis agent releasing after osmosis transport tunnel damage of mitochondria. Autophagy was then initiated to remove the damaged mitochondria to protect the cell.Now, apoptosis induction of glucocorticoid in CD4~+ T lymphocyte was proved but not autophagy induction. Therefore, we observed autophagy in peripheral blood T lymphocyte and it's subset in asthma to make further understanding of PCD in them and effect of glucocorticoid.Part one: Autophagy in human peripheral blood T lymphocyte MethodsPeripheral blood T lymphocyte of healthy adult was separated with Percoll (1.073 g/mL) and harvested by filtration through nylon column. Cell morphology was observed by electron microscope, and fluorescent microscope at 0 and 72h. And incidence rate of autophagy was analyzed by flow cytometry after MDC stain.Results1. T lymphocyte showed typical morphology of autophagy under electron microscope at 72h.2. T lymphocyte showed typical autophagic vacuoles under fluorescence microscope at 72 h.3. There was significant statistic difference between 0 and 72h autophagy incidence rate of T lymphocyte.Part two: Autophagy in peripheral blood CD4~+ T lymphocyte of asthmaMethodsPeripheral blood CD4~+ T lymphocyte of asthma patient was harvested after separation with Percoll (1.073 g/mL), nylon column filtration and negative magnetic separation. The cultured cells were divided into control and DXM group, and incidence rate of autophagy and apoptosis was analyzed by flow cytometry.Results1. There was significant statistic difference in autophagiy incidence rate of CD4~+ T lymphocyte between control and DXM group in asthma at 72h. 2. There was significant statistic difference in apoptosis incidence rate of CD4~+ T lymphocyte between control and DXM group in asthma at 72h.Conclusions1. T lymphocyte showed typical morphology of autophagy at 72h in heathy control group.2. There was significant statistic difference between 0 and 72h autophagy incidence rate of T lymphocyte in heathy control group.3. There was significant statistic difference in autophagiy incidence rate of CD4~+ T lymphocyte between control and DXM group in asthma at 72h.