An evaluation of the mitogenic reactivity of intestinal intraepithelial lymphocytes of chickens.
A colorimetric assay employing MTT (3-[4,5-dimethylthiazole-2-yl], 2-5-diphenyltetrazolium bromide) was used to determine the mitogenic response of intestinal intraepithelial lymphocytes (i-IELs) of chickens to T- and B-cell mitogens. Comparisons between mitogenic responses of i-IELs and peripheral blood lymphocytes (PBLs) were made to examine potential relationships. The results from this study indicated that T-cell mitogens, concanavalin A (Con A), and phytohemagglutinin-P (PHA-P) induced mitogenic stimulation in i-IELs. Although stimulation indexes of both i-IELs and PBLs were similar, the optical densities (ODs) of i-IEL cultures containing Con A or PHA-P were 20- to 50-fold lower than the ODs of PBL cultures containing the same mitogen. The lower conversion of MTT to formazan resulting in lower ODs in i-IEL cultures indicated a lower level of cellular activity in the i-IELs than in the PBLs. The mitogenic responses of both i-IELs and PBLs to Con A and PHA-P were dose dependent. The responsive concentration of Con A for i-IELs was within the range of 25-50 micrograms/ml, whereas the responsive concentration of PHA-P for i-IELs was 50 micrograms/ml. Three days of incubation was found to be adequate to induce a significant (P < 0.05) mitogenic response for both T-cell mitogens. Lipopolysaccharide was unable to induce a mitogenic response in i-IELs, which was attributed to the lack of B cells in the i-IEL population. This technique may prove useful in evaluating and studying the role of i-IELs in local cell-mediated immune responses of the gastrointestinal tract.
Agrawal
P K
PK
Department of Veterinary Microbiology and Preventive Medicine, College of Veterinary Medicine, Iowa State University, Ames 50011, USA.
Reynolds
D L
DL
D000818:Animals; D001402:B-Lymphocytes / Q000187:drug effects; D002478:Cells, Cultured; D002645:Chickens / Q000276:immunology*; D003124:Colorimetry / Q000379:methods / Q000662:veterinary; D003208:Concanavalin A / Q000494:pharmacology; D007413:Intestinal Mucosa / Q000166:cytology / Q000276:immunology*; D008213:Lymphocyte Activation; D008214:Lymphocytes / Q000276:immunology*; D008934:Mitogens / Q000494:pharmacology*; D010835:Phytohemagglutinins / Q000494:pharmacology; D013601:T-Lymphocytes / Q000187:drug effects
D008934:Mitogens; D010835:Phytohemagglutinins; C028784:phytohemagglutinin-P; D003208:Concanavalin A