[Morphology and differentiation pathway of superantigen SEB-activated NKT subsets].
OBJECTIVE
This study is to explore the biological characteristics of NKT subsets through investigating their morphology and differentiation pathways.


METHODS
Splenic lymphocytes from C57BL/J mice were treated with either Staphylococcal entertoxin B (SEB) or ConA and then cultured in vitro. On day 5 and day 10, the lymphocytes were harvested. Normal adherent macrophages and normal lymphocytes were harvested on day 0. Expression of CD69 molecule on the cell surfaces of the lymphocytes and large lymphocytes by SEB-activated was measured using staining of fluorescent antibody and flow cytometry (FACS). And the percent (%) of NKT subsets was determined and analyzed the differentiation pathways of the NKT subsets. Appearance of subsets was observed and compared under a light microscope at x400 magnification.


RESULTS
Expression levels of CD69 molecule on the SEB-activated lymphocytes and the giant lymphocytes were significantly increased to 55% and 68.95% respectively as compared to the level (0.11%) detected on the normal lymphocytes (both P<0.01; n=3). The percent (%) of CD8(+) NK1.1(+) and TcRVbeta8(+) NK1.1(+) NKT subsets in the giant lymphocytes was significantly enhanced to 30.29% and 31.48% (84.0 or 38.9 folds) originally from 0.36% and 0.81%, respectively. Based on the cell distribution shown in the upper part of the FACS picture, they should be large-scale selection. The SEB-activated lymphocytes in size were larger than not only the ConA-activated cells but also the adherent macrophages with an increase of 5 fold observed under a microscope. They are no-adherent cells. There were a few granule seen in cytoplasm . The value of cytoplasm vs nuclei was less than 1.0. The SEB-activating CD8(+) and TcRVbeta8(+) NKT subsets were not relative to a NK source. They were produced directly from T cells differentiation.


CONCLUSIONS
The SEB-activating NKT cells are giant lymphocytes in size, consisting predominantly of CD8(+) NK1.1(+) NKT cells. They are considered as a subsets of T lymphocytes.
Chen
Yu
Y
Department of Immunology, General Hospital of PLA, Beijing 100853, China. leoyu0323@yahoo.com.cn
Guo
Ye-lei
YL
Zhong
Jiang
J
D000818:Animals; D015703:Antigens, CD / Q000032:analysis; D000945:Antigens, Differentiation, T-Lymphocyte / Q000032:analysis; D002454:Cell Differentiation; D004768:Enterotoxins / Q000276:immunology*; D005260:Female; D037181:Lectins, C-Type / Q000032:analysis; D008213:Lymphocyte Activation; D051379:Mice; D008810:Mice, Inbred C57BL; D055611:Natural Killer T-Cells / Q000166:cytology* / Q000276:immunology; D018089:Superantigens / Q000276:immunology*; D000099192:CD69 Antigens
D015703:Antigens, CD; D000945:Antigens, Differentiation, T-Lymphocyte; D004768:Enterotoxins; D037181:Lectins, C-Type; D018089:Superantigens; D000099192:CD69 Antigens; C031818:enterotoxin B, staphylococcal