Document 0862
 DOCN  M9650862
 TI    Rhodamine 123: a useful probe for monitoring T cell activation.
 DT    9605
 AU    Ferlini C; Biselli R; Nisini R; Fattorossi A; Laboratory of Immunology,
       D.A.S.R.S., Pomezia, Rome.
 SO    Cytometry. 1995 Nov 1;21(3):284-93. Unique Identifier : AIDSLINE
       MED/96139620
 AB    The T cell activation pathway involves an increase in mitochondrial
       activity. This can be evaluated in individual cells using the
       fluorescent probe rhodamine 123 (Rh123) and flow cytometry. Peripheral
       blood mononuclear cells (PBMC) were stimulated with optimal
       concentrations of phytohaemagglutinin (PHA), superantigens (Sag) SEA and
       SEC2, and allogeneic cells. Activation kinetics were followed at days 1,
       2, 4 and 7. In all activation conditions, Rh123 uptake was augmented
       with the CD25 expression, cell size, and DNA synthesis. Rh123 uptake
       reflected an increase in mitochondrial activity and mass, as assessed by
       experiments in which Rh123 was substituted for by the 10-nonyl acridine
       orange, which stains mitochondria in an energy-independent manner. The
       spectral characteristics of Rh123 allowed us to double stain cells with
       Rh123 and phycoerythrin-conjugated monoclonal antibodies. In
       PHA-activated cultures, CD4+ and CD8+ cells incorporated essentially the
       same amount of Rh123 at all time points, suggesting that the two subsets
       did not differ in their activation kinetics. Accordingly, after 1 week
       of culture, no significant modification in the CD4/CD8 ratio was
       observed. Sag-activated CD4+ cells incorporated a higher amount of Rh123
       than did CD8+ cells and preferentially expanded after 1 week of culture
       as indicated by the increase in the CD4/CD8 ratio. The different
       behavior of the CD4 and CD8 subsets observed by dual color flow
       cytometry in the PHA and Sag models was confirmed using purified CD4+
       and CD8+ cell preparations obtained by immunomagnetic sorting. CD4+
       cells were also the preferential target in the allogeneic model,
       although the magnitude of the phenomenon was lower than in the Sag
       model. Present data indicate that Rh123 is a reliable marker for
       monitoring the mitochondrial compartment during T cell activation. The
       possibility of phenotyping Rh123-stained cells adds to the applicability
       of the probe.
 DE    CD4-Positive T-Lymphocytes/CYTOLOGY/*IMMUNOLOGY  CD8-Positive
       T-Lymphocytes/CYTOLOGY/*IMMUNOLOGY  Enterotoxins/IMMUNOLOGY  Flow
       Cytometry/METHODS  *Fluorescent Dyes  Human  In Vitro  Leukocytes,
       Mononuclear/CYTOLOGY/IMMUNOLOGY  *Lymphocyte Transformation
       Phytohemagglutinins/IMMUNOLOGY  *Rhodamines  JOURNAL ARTICLE

       SOURCE: National Library of Medicine.  NOTICE: This material may be
       protected by Copyright Law (Title 17, U.S.Code).