CELLS AND THE LUNG

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cells share with B cells and T cells the property of using somatic gene rearrangement to generate diverse surface antigen receptors. Nonetheless, while we are confident of the contributions made to protective immunity by B cells and by T cells, the case remains enigmatic for cells (1).

By analysis of cell dynamics, and studies of cell-deficient mice, cells have been implicated in immune responses to numerous infectious and noninfectious challenges. Thus, in the lung, interleukin 4 (IL-4) production by cells was claimed to be essential in a murine model of airway hyperresponsiveness (2). Paradoxically, cells have also been reported to downregulate airway hyperresponsiveness, independent of T cell-mediated events (3). Strikingly, cell-deficient mice are fatally defective in their immune response to Nocardia, a gram-positive bacterium delivered to the lung by aerosol. The failure appeared to involve an insufficient mobilization of neutrophils (4). Neutrophils also respond to ozone-treatment of animals. cell-deficient animals likewise succumbed to exaggerated damage induced by ozone (4).

These several reports indicate that cells are relevant to the biology of the lung, at least as explored in mouse models. Moreover, the results reported can be reconciled to some extent with documented specificities and effector functions of cells.

	CELL SPECIFICITIES AND ANTIGEN SAMPLING

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The T cell receptor (TCR) resembles immunoglobulin and TCR in presenting a series of solvent-exposed loops supported by two  sheets (5). The variability in putative CDR3 is greater than that in TCR (6) and CDR1/2 are strikingly diverged among different TCRs within a species (1, 6). This is consistent with the evidence that cells, unlike T cells, are not "locked in" to recognition of peptide plus MHC (7). In support of this are additional data that cells are usually CD4CD8, and hence unable to exploit the capacity of CD8 and CD4 to coengage (with the TCR) MHC class I and class II antigens, respectively.

Some antigen specificities of some subsets of cells have been elucidated. Thus, human peripheral blood cells recognize low molecular mass mycobacterial products, for example, ethylamine, and isopentenyl pyrophosphate, which is also produced by actively growing mammalian cells (8). The mechanism by which cells recognize these products is unelucidated. Indeed, although there can be high sequence diversity in TCR, it is not evident how antigens of any great diversity can be sampled. This is because cells are seldom found in the conventional T/B zones of the lymph nodes and spleen, respectively, which are the known anatomical sites where diverse antigen can be presented, and the responding, cognate lymphocytes rapidly expanded. Instead, cells are commonly found within epithelia, or broadly spread throughout secondary lymphoid organs such as the spleen. This is true even in animals, such as young cattle, where > 75% of CD3⁺ cells may comprise CD4CD8 cells (1).

Because of the potential difficulty in sampling antigen, and based on experimental data that cells could respond to MHC class I structures (11), it was proposed that the cells might recognize class Ib MHC molecules, expressed as sentinels of cell stress or infection (12). Data supporting this come from evidence that human gut cells recognize MICA (MHC class I chain-related protein A) (13), and that a significant subset of murine splenic cells recognizes T10/T22 (14). Both products are expressed by activated cells, epithelial cells, and lymphoid cells (14, 15). These data establish that cells have the potential to recognize and effect function toward activated epithelial cells and lymphoid cells. This has been confirmed by tissue culture experiments (13, 16).

	CELL EFFECTOR FUNCTIONS

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Diverse effector functions have been attributed to cells. For example, helper T cell type 1 (Th1) and Th2 clones have been characterized, and shown to support the production of the respective classes of immunoglobulin (Ig) in vivo (18). This activity is heightened in T cell-deficient mice, with high titers of IgE supported by cell activity. This appears to be exaggerated on particular genetic backgrounds (M. Girardi, A. C. Hayday, and R. E. Tigelaar, unpublished data). Mucosal IgA synthesis is low in -deficient mice (19). In short, the localization of cells to tissues appears to be paralleled by their capacity to support the production of "body surface" Ig. These data demonstrate the potential of cells to be major contributors to mucosal B cell responses.

In contrast to CD4⁺ T cells, cells are usually biased away from Th2 and toward Th1 and the production of interferon  (IFN-). In addition, intraepithelial lymphocytes (IELs) are reported to produce active fibroblast growth factor VII (FGFVII), which can promote the growth and differentiation of epithelial cells (20). Paradoxically, they can also be cytolytic toward epithelial cells (13) and may express granulysin. In summary, cells display varied effector functions and the phenotype manifested by cell activity (or deficiency) will most likely vary according to the nature of the stimulus, the genetics of the animal, and the stage of the animal's development.

	CELLS REGULATE IMMUNOPATHOLOGY AND INFLAMMATION

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To assess the contribution of cells to mucosal immune responses, our laboratory has focused on the use of Eimeria vermiformis, a natural coccidian pathogen of the mouse gut (21). Such parasites are abundant, infecting mucosal epithelial cells in a wide range of vertebrates. The strength of an infection is readily measured by counting oocysts released into fecal pellets over a period of from 1 to 2 wk postinfection. On rechallenge, immunocompetent animals are highly resistant, even to significantly higher inocula.

Eimeria infection of T cell-deficient mice revealed a marked susceptibility to primary infection, and a failure to develop immunity to rechallenge (22). The effector role of T cells is probably mediated by IFN--producing, Th1 cells. In contrast, TCR^/ mice showed no defects in either parameter (22). Such negligible contribution of cells to protective immunity has been observed in numerous infections. Nonetheless, TCR^/ mice show an exaggerated immunopathology, attributable to the action of T cells (22). Whether this reflects a capacity of cells directly to regulate T cell activity or whether, via the action of FGFVII, cells ordinarily protect the epithelium, is not currently resolved. It has been considered that in the absence of FGFVII from cells, epithelial homeostasis is dysregulated in health and disease (1).

Such dysregulated immunopathology has been reported for several other infections of TCR^/ mice (1). Interestingly, cells can also respond to and limit inflammatory damage elicited by noninfectious agents (23). Indeed, the spontaneous, autoreactive hyperresponsiveness of T cells that leads to lupus- and nephritis-type symptoms in MRL/lpr mice is exaggerated when crossed onto the TCR^/ background (24). By 24 wk, mortality rates for ^/ MRL/lpr mice were ~ 67% compared with ~ 25% for ⁺ MRL/lpr mice (24). These data demonstrate incontrovertibly that a defect in cell immunoregulatory function can reduce life span, contingent on an appropriate genetic background.

	AN AGE-DEPENDENT ROLE FOR CELLS: A HYPOTHESIS

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Nonetheless, the general importance of an immunoregulatory role for cells in adult animals is as yet unclear. In addition, such immunoregulation may not represent the primary role of cells, just as immunoregulation is not the primary role of T cells. Indeed, in preadolescent members of several species, cells outnumber T cells, the putative targets of regulation. Hence, we further examined the Eimeria system for more overt immunoprotective cell function.

First, we compared the susceptibility to infection of adult TCR^/ mice with that of TCR( × )^/ mice that lack all T cells. The latter were even more susceptible than the former, indicating that cells respond to the infection, and can contribute to pathogen clearance, even though they are not required in T cell-competent animals (22). cells appear biased toward Th1 activity and IFN- production (1). Interestingly, IFN-^/ mice were as susceptible as TCR( × )^/ mice, supporting the hypothesis that cells contribute an antipathogenic effect via production of IFN-. The question, therefore, was whether there might be a set of conditions under which this or a related activity of cells might be more obviously critical to the health of the host.

cells are disproportionately abundant in young animals, between birth and adolescence. Remarkably, studies of humans and mice indicate that during this ontogenetic period, T cells are extremely Th2 biased. To test whether this context placed greater reliance on cells for immunoprotection, Eimeria infections were repeated in young mice, before, at, or shortly after weaning. In complete contrast to adult mice, TCR^/ mice now showed little or no increased susceptibility to primary infection. Rather, an increased susceptibility was commonly shown by TCR^/ mice (E. Ramsburg, S. J. Roberts, and A. C. Hayday, unpublished data). We therefore believe that the responsiveness and immunoprotective potential of cells may be especially important in young animals. This may be particularly true for animals, such as chickens, that do not receive transplacental maternal immunoglobulin, and in which cell numbers are particularly high.

Interestingly, when young mice were rechallenged with Eimeria only the T cell-deficient animals failed to show immunity. Hence, T cells are not "ignorant" of the initial challenge, and although they fail to make an obvious contribution to the primary response, they clearly establish a memory pool. Possibly these events accelerate the ontogenetic development of Th1 activity in the T cell pool (Figure 1).

View larger version (12K): [in this window] [in a new window]   Figure 1.   Hypothesis for involvement of cells in "infection-based immunoregulation" in young animals.

	SUMMARY

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cells have been retained as a component of the tripartite lymphocyte system ( T cells, B cells, and cells) for more than 500 million years. This suggests a strong selective pressure for their retention. At the same time, cells are represented to different degrees in different species: high in ungulates, in chickens, and in intestinal repertoires of mice; low in humans and in the system circulation of mice. Nonetheless, in all vertebrates in which cells have been studied, the cells are disproportionately abundant in young animals, and at body surfaces. In seeking a property of cells that might satisfy all these points, we have found evidence that cells may be important for controlling primary infections in young animals, prior to T cells assuming this role.

We do not yet know the breadth of pathogens to which this hypothesis may apply. But somewhat similar data were obtained in studies of Cryptosporidium infection of young versus adult mice (25). In that case, T cell-deficient animals showed a defect in immunoprotection only during the early phases of infections of very young mice. This is entirely consistent with our findings. Nonetheless, T cell-deficient mice also showed defects in immunoprotection when infected as young mice, and it is also true that the generally accepted period of susceptibility of mice to Cryptosporidium infection is during the first 4 wk of life. How can this be if cells are effective at providing immunoprotection during this period? Possibly the key is in the nature of the infecting organism. Cryptosporidium, unlike Eimeria, is not self-clearing in immunodeficient animals. Therefore, defects in host responsiveness seen in T cell-deficient mice after 3 to 4 wk possibly represent a failure of a robust T cell memory response to develop. This is consistent with our findings that a long-lasting immunoprotective memory response to Eimeria requires T cells, irrespective of the age of infection.

The mechanism of action of cells is now under investigation. At least three nonmutually exclusive mechanisms can be considered, based on the effector functions outlined above. The cells may be directly antipathogenic through cytolytic and/or Th1 activities; they may activate antipathogenic activities in other cells, for example, macrophages or neutrophils, via the action of cytokines; and they may protect the survival and growth of healthy epithelial cells, via the production of epithelial growth factors. The antipathogenic role for cells in young animals could be of great significance when exposure to the external environment is at its most precipitous and potentially catastrophic. In addition, it is possible that an early skewing toward Th1 responses, driven by cells, might accelerate the bias away from Th2 responses and thereby reduce susceptibility to allergy.