A study of the proliferative response of rabbit T cells using the BrdU-Hoechst method

Con-A- and PHA-induced proliferation of cells from rabbit thymus, spleen and mesenteric lymph node was studied with the DNA-fluorescent probe 33258 Hoechst. The fluorescence of this probe is quenched when 5-bromo-2'-deoxy-uridine is incorporated into nascent DNA during the S phase. Fluorescence decreased with increasing content of newly formed DNA per cell. Proliferation kinetics and the number of Con-A- and PHA-reactive cells (C+ and P+ cells) were determined cytofluorometrically . Lymphocytes from control and dexamethasone (DX)-treated animals start their proliferation early: after 42 hr about 25% of the control and the majority of the DX-resistant cells finished their second cell division. Small numbers of C+ (12.0%) and P+ (3.5%) cells were found in control thymus, while these percentages were enhanced in DX thymus: 32.5 and 27.0% respectively; 50% of the spleen T cells in control and DX animals are C+ or P+ and 75% of the lymph-node T cells are C+ (after DX 45%) and 50% are P+ (after DX also 50%). It is concluded that in thymus and lymph nodes, a steroid sensitive (Ss) C+P-, and in lymph nodes a Ss C+P+ cell pool is present. A mitogen non-proliferative cell pool (C-P-) is present in control and DX thymus.