CD4^ T CELL HETEROGENEITY 
Kim Bottomly, Ph.D., Associate Investigator 
CD4^ T cells have been shown to be functionally 
heterogeneous, with CD4^ T cells directing both 
humoral and cell-mediated immunity. Because of 
the diverse capabilities of CD4-bearing T cells, Dr. 
Bottomly became interested in determining if a sin- 
gle cell could mediate all functions characteristic of 
CD4^ T cells or if specialized subsets existed within 
the CD4^ T cell population. Two approaches were 
used in the initial studies. First, monoclonal an- 
tibodies were shown to identify phenotypic dif- 
ferences between CD4-bearing T cells, and these 
phenotypic differences correlated with functional 
differences. Studies in both the rat and human sys- 
tems showed that CD4"'" T cells could be divided 
into naive and memory cells on this basis. Second, 
mouse monoclonal T cells showed differences in 
their functional capabilities, and this correlated 
with the cytokines they released. Thl cells are 
cloned T cells that release interleukin-2 (IL-2), in- 
terferon-7 (IFN-7), and lymphotoxin (LT); these 
cells also mediate delayed hypersensitivity re- 
sponses, activate macrophages, and kill class II 
major histocompatibility complex (MHC) -bearing B 
cells. Th2 cells are cloned T cells that release inter- 
leukin-4 (IL-4) and interleukin-5 (IL-5) and help B 
cells, eosinophils, and mast cells. Further studies 
comparing these two approaches have pointed to 
inconsistencies or differences in the defined CD4''^ 
subsets. To resolve these differences and to investi- 
gate functional equivalents of Thl and Th2 cells in 
vivo, this laboratory has produced a monoclonal 
antibody that distinguishes between cloned lines of 
Thl and Th2 cells. This antibody has been used to 
separate normal CD4 T cells to study their function 
and activation requirements. 
I. Characteristics of Subsets of CD4"'' T Cells. 
Initial studies using the monoclonal antibody 
16a showed that CD4 T cells from normal mice 
could be accurately subdivided into stable popula- 
tions. When the density of the determinant was 
used as a marker, 16A high-density cells produced 
IL-2 and IFN-7 upon activation with T cell mitogens 
in short-term culture; whereas the I6A low-density 
cells produced IL-4 and IL-5 and provided excellent 
helper T cell function. Thus it appears that I6A 
monoclonal antibody divides normal CD4 T cells 
into Thl-like and Th2-like T cell subsets. By im- 
munoprecipitation, I6A was shown to bind to com- 
mon leukocyte antigen or CD45. This is of particu- 
lar interest in that the majority of other antibodies 
that subset CD4 T cells in other species are also di- 
rected against CD45. Dr. Bottomly and her col- 
leagues compared the subsets defined by I6A in the 
three species with subsets defined by other anti- 
CD45 antibodies. 
In the rat and human, CD4 T cells are subdivided 
by anti-CD45 antibodies into naive and memory T 
cells. To examine whether 16A separates compara- 
ble populations. Dr. Bottomly and her colleagues 
tested I6A high- and low-density cells for their abil- 
ity to make memory or recall responses to a specific 
antigen. Recall T cell activation, as measured by 
cytokine release, indicated that both subsets con- 
tain memory T cells. Thus subsets defined by I6A 
are not equivalent to those subsets defined in the 
rat and human. A second possibility suggested by 
the data is that I6A subdivides memory T cells into 
Thl-like and Th2-like cells. Either their immediate 
precursor is uncommitted functionally and be- 
comes committed upon contact with antigen/anti- 
gen presenting cells or their immediate precursor is 
precommitted and upon differentiation becomes a 
Thl or Th2 cell. The latter possibility seems un- 
likely, in that cloned T cell lines derived from I6A- 
high cells that produce IL-2, switch to the produc- 
tion of IL-4. Thus it would appear that l6A-high 
cells can give rise to I6A-I0W cells, phenotypically 
and functionally. Finally, a study of the phenotype 
and function of l6A-high and I6A-I0W cells over 
time after activation with antigen suggests that the 
main activity of l6A-high cells is to release IL-2, 
clonally expand, and give rise to I6A-I0W cells. By 
contrast, the main activity of I6A-I0W cells is to re- 
lease IL-4, IL-5, and IFN-7 upon stimulation. Be- 
cause most helper activity and cytolytic activity is 
found in this population, they may be considered T 
effector cells. Thus the data suggest that I6A sepa- 
rates CD4 T cells into a T proliferator population 
and a T effector population. 
II. I6A Defines Two Subsets of Memory T Cells. 
Analysis of the proliferative capabilities of l6A-de- 
fined subsets of CD4 indicates that l6A-high cells 
give the best recall T cell proliferative response, 
which decreases over time. This proliferative re- 
sponse is blocked by anti-IL-2 antibodies. I6A-I0W 
cells have a limited proliferative potential early after 
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