Establishment Of CD3 ~ + CD4 ~ + T Lymphocyte Sorting And The Significance Of Integrin α4β7 Expression In Intestinal Tract

Objective：To establish a method to sort CD3+CD4+T lymphocytes in theperipheral blood quickly and accurately with less disturbing cell activity.Methods： First of all,peripheral blood mononuclear cells (PBMC) wereprepared by using continuous density Percoll gradient centrifugation technique,and then CD3+CD4+T lymphocytes were sorted by flow cytometry (FCM).Themethod was evaluated by the purity, shape and the survival rate of the sortedcells, respectively. Results：Continuous density Percoll gradient centrifugationcould well prepare PBMC.The purity of CD3+CD4+T lymphocytes was(51.9±9.6)%before sorting, while the purity of CD3+CD4+T lymphocytes was(95.9±0.8)%after sorting，it was significantly different between before and aftersorting (P＜0.01). The sorted CD3+CD4+T lymphocytes survival rate was(95.8±1.2)%and had intact cell shape. Conclusion：FCM not only produceshighly pure CD3+CD4+T lymphocytes with little influence on activity and shape but aslo may be carried out under sterile conditions, which insures the cells to besatisfactorily used in functional research. Objective：To establish an accurate method to sort human intestinalCD3+CD4+T lymphocytes with less disturbing cell activity. Methods：First ofall,digestive enzymes decomposed human intestinal tissue,and then whichpassed through a40μm cell strainer into cell suspension.Mononuclear cellswere collected by using continuous density Percoll gradient centrifugationtechnique,then flow cytometry sorted them to get CD3+CD4+T lymphocytes.The method was evaluated by the purity, shape and the survival rate of thesorted cells, respectively.Results：Digestive enzymes well decomposed humanintestinal tissue into cell suspension, by using continuous density Percollgradient centrifugation the purity of CD3+CD4+T lymphocytes was（43.9±7.3）%,while the purity of CD3+CD4+T lymphocytes was（96.9±1.2）%afterfluorescence-activated cell sorting，it was significantly different between beforeand after fluorescence-activated cell sorting (P＜0.01). The sortedCD3+CD4+T lymphocytes survival rate was (97.8±1.6)%and had intact cellshape. Conclusion：Combination of Percoll gradient centrifugation techniqueand fluorescence-activated cell sorting not only produce highly pure humanintestinal CD3+CD4+T lymphocytes with little influence on activity and shape but aslo may be carried out under sterile conditions, which insures the cells to besatisfactorily used in functional research. Objective：To provide evidence of molecular level of intestine as majorsite of HIV infection,we detected α4β7+CD3+CD4+T lymphocytes expressionrate in peripheral blood and intestinal tissue.Methods：The expression ofintegrin α4β7+,CD3+and CD4+in T lymphocytes were tested by flowcytometry,then data were analyzed statistically.Results：The expression rate ofα4β7+CD3+CD4+T lymphocytes wa（s30.2±5.1）%in peripheral blood and（45.2±6.2）%in intestinal tissue respectively,it was significantly different betweenperipheral blood and intestinal tissue (P＜0.01).Conclusion：Integrin α4β7makes CD3+CD4+T lymphocytes to the gut-associated lymphoid tissue,theexpression rate of α4β7+CD3+CD4+T lymphocytes is higher in intestinal tissuethan in peripheral blood.The high expression of integrin α4β7in CD3+CD4+Tlymphocytes makes intestinal tissue become major site of HIV infectionpossibly.