Spontaneous Expression of Interleukin-2 In Vivo in Specific Tissues of Young Mice
Spontaneous Expression of Interleukin-2 In Vivo in Specific Tissues of Young Mice
JULIA A.YANG-SNYDER 0
ELLEN V.ROTHENBERG 0
0 aHoward Hughes Medical Institute and Department of Pharmacology K516, University of Washington , Box 357750, Seattle, Washington 98195-7750 , USA
In situ hybridization and immunohistochemistry were used to determine the spectrum of tissues in which interleukin-2 (IL-2) mRNA and protein are found in healthy, normal young mice. In neonatal animals, IL-2 is expressed specifically by distinct, isolated cells at three major sites: the thymus, skin, and gut. Based on morphology and distribution, the IL-2-expressing cells resemble CD3e T cells that are also present in all these locations. Within the thymus of postweanling animals, both TcRafl and TcR),6 lineage cells secrete "haloes" of the cytokine that diffuse over many cell diameters. Within the skin, isolated cells expressing IL-2 are seen at birth in the mesenchyme, and large numbers of IL-2-expressing cells are localized around hair follicles in the epidermis in 3-week-old animals. At this age, a substantial subset of CD3 cells is similarly localized in the skin. Significantly, by 5 weeks of age and later when the CD3e cells are evenly distributed throughout the epidermis, IL-2 RNA and protein expression are no longer detectable. Finally, within the intestine, IL-2 protein is first detected in association with a few discrete, isolated cells at day 16 of gestation and the number of IL-2 reactive cells increases in frequency through El9 and remains abundant in adult life. In postnatal animals, the frequency of IL2-positive cells in villi exceeds by greater than fivefold that found in mesenteric lymph node or Peyer's patches. Overall, these temporal and spatial patterns of expression provide insight into the regulation of IL-2 in vivo and suggest a role for IL-2 expression distinct from immunological responses to antigen.
Dendritic epidermal T cells; gut-associated lymphocytes; TCR-yt3 thymocytes; TCR-a/3 thymocytes; in situ hybridization; immunohistochemistry; developmental regulation; interleukin-2
INTRODUCTION
Interleukin-2 (IL-2) has been best studied as a growth
factor secreted by activated T cells in the course of the
immune respohse. Its expression is strictly dependent
on inducing stimuli, with requirements forseparate
signals from Ca+2-dependent, ras-dependent, and
1995; Serfling et al., 1995). These requirements are
met in mature T cells when they encounter antigen on
antigen-presenting cells, thereby delivering
coordinated signals triggered by the crosslinking of TcR/
CD3 complexes, CD4 (or CD8 in some cases), and
CD28. Accordingly, analysis of the cis and trans
regulatory elements critical for IL-2 expression has
shown several discrete protein-DNA interactions that
require activation by TcR/CD3- and/or
CD28dependent signaling mediators
(Jain et al., 1995;
Serfling et al., 1995)
. The observation that IL-2 is also
expressed primarily or exclusively by T cells has led
to a general assumption that the primary function of
IL-2 expression in vivo is to regulate T-cell clonal
expansion as triggered by T-cell recognition of
foreign antigen.
This assumption has been challenged in recent
years by the phenotype of IL-2 "knockout" mice.
Animals homozygous for targeted disruption of the
IL-2 gene show initially normal numbers of T cells of
various subsets and detectable but relatively subtle
defects in their immediate proliferative responses to
antigen
(Schorle et al., 1991; Ktindig et al., 1993;
Krimer et al., 1994; Cousens et al., 1995; Kneitz et
al., 1995)
. By contrast, multiple aspects of their
longterm hematopoietic regulation and lymphocyte
homeostasis are disrupted
(Sadlack et al., 1993, 1995;
Krimer et al., 1995; Reya, 1996)
.
These findings raise several questions about the
role of IL-2 expression in vivo. First, is IL-2
expressed in homeostatic contexts as well as in acute
immune responses? Second, does IL-2 expression
regulate hematopoietic populations in nonlymphoid
tissues? Lastly, is mature cell behavior affected by
prior exposure or lack of exposure to IL-2 during
development? In order to address these questions, we
systematically examined various organs of normal
young mice to determine where IL-2 is detectably
expressed. Our studies show that IL-2 mRNA and
protein are associated with specific tissues in situ.
These tissues invariably contain T cells, suggesting
that IL-2 may be involved in regulating lymphocytic
and/or hematopoietic function(s) at these sites.
RESULTS
Overall Distribution of IL-2-Expressing Cells in
Neonatal Mice
To survey the range of tissues that might express the
IL-2 gene in mice, neonatal mouse body sections were
analyzed by in situ hybridization with 35S-labeled
probes. The results of these analyses showed that
most tissues of the body were devoid of hybridization
above background, as established with sense-strand
control probes. Brain, heart, lung, liver, stomach,
bladder, and oth (...truncated)