CD8 lymphocyte population heterogeneity has been examined by using reovirus serotype 1, strain Lang (reovirus 1/L) as a model infection. We have previously reported that gut mucosal infection with reovirus stimulates the appearance of virus-specific cytotoxic T cell precursors (pCTL) in Peyer's patches (PP). The effectors that mediate reovirus-specific cytotoxicity were found to express both the Thy-1 and CD8 Ag and were MHC-restricted in their recognition of reovirus Ag. To further characterize the virus-specific precursor and effector cells we have analyzed PP cells for the expression of a novel surface Ag (termed germinal center and T cell Ag (GCT)) found on germinal center B cells and a subpopulation of CD8+ T cells. Gut mucosal infection with reovirus 1/L is capable of increasing the proportion of GCT+ CD8+ T cells in PP. Positive selection as well as depletion of GCT+ cells has demonstrated that pCTL can express this Ag, and depletion experiments have demonstrated that effector CTL express the GCT Ag. Thus, a subpopulation of GCT+ cells have been identified as Ag-specific precursor and effector CTL. These observations indicate that the expression of the GCT Ag may provide a means to identify recently stimulated pCTL or effector CTL in gut mucosal tissues.

Reovirus, serotype 1, causes a transient, asymptomatic infection of the murine intestine when given intraduodenally or orally. However, this infection markedly perturbs both B- and T- cell populations in Peyer's patches (PP) resulting in: 1) a rapid and persistent increase in specific precursors for cytotoxic T cells (pCTL) and a gradient of frequencies highest in PP and lowest in distal lymphoid tissue; 2) a similar increase in memory B cells committed to IgA; 3) the transient appearance of a subset of germinal center B cells identified by MAb, GC-T; 4) the appearance of pCTL among intraepithelial lymphocytes; and 5) the antigen non-specific alteration in Ig isotype potential of B cells previously primed and found in PP. The pCTL appearing upon acute gut mucosal infection with reovirus are Thyl+, Lyt2+, virus-specific, viral serotype non-specific, class I MHC haplotype restricted and occur within the subset of T cells which newly appears also identified by MAb GC-T. Infections of both neonatal and severe-combined immunodeficient mice indicate that the elements of the immune system may operate at many levels to resist, limit, contain, and resolve viral infection.

A monoclonal anti-idiotope termed 87.92.6 mimics the neutralization/cell-attachment site of the reovirus type 3 hemagglutinin (HA3). The second complementarity determining regions of the VH and VL of 87.92.6 share sequence similarity with a determinant on the HA3. We have used synthetic peptides (termed VH, VL, and Reo peptides, respectively) to probe the immunologic significance of this sequence similarity. Antibodies specific for Reo peptide or VL peptide neutralized reovirus type 3 infectivity. Although Reo peptide was an effective immunogen by itself, free VL peptide or VH peptide were unable to elicit antibodies unless they were linked to each other (VH-VL peptide). Immunization with reo peptide, 87.92.6, or the HA3 elicited a specific lymphocyte proliferative response to VH peptide, indicating that VH peptide may bear an important TH determinant. As found previously for 87.92.6, VL peptide elicited a delayed-type hypersensitivity response specific for reovirus type 3. Reovirus type 3 specific cytolytic lymphocytes specifically lysed targets coated with VH-VL peptide, but not VH or VL peptide alone. These results suggest that immune cross-reactivity between an external Ag and an internal image antibody can be understood at the primary structural level. These observations may have important implications for understanding the development of autoantibodies, network interactions, and the regulation of immune responses.

Reovirus 1 has been intraduodenally administered to mice to determine whether virus-specific cytotoxic cells can be elicited within the intestinal epithelium. We have found that reovirus-stimulated, but not control mice, generate virus-specific cytotoxic effector cells when isolated intraepithelial lymphocytes are restimulated in vitro. These effector cells are restricted by major histocompatibility complex (MHC) class I determinants and thus appear to utilize the T cell antigen receptor and to be of the T cell lineage. These results raise the possibility that cytotoxic T lymphocytes (CTL) may be an important component of the host immune response in the intestinal epithelium.

Reovirus, a virus that contains neither an envelope nor glycosylated polypeptides, has been found to induce virus-specific, major histocompatibility complex (MHC) class I antigen restricted, cytotoxic T lymphocyte (CTL) responses. The cytotoxic T cells require in vitro stimulation in the presence of virus to phenotypically express cytotoxic activity. Utilizing reovirus types 1 and 3, the CTLs derived from mice infected with one serotype can lyse target cells infected with a second serotype of reovirus. In addition, lymphocytes primed in vivo with one serotype develop into fully functional CTLs during in vitro stimulation with the other serotype of reovirus. Therefore, these results suggest that reovirus induced CTLs are virus, but not serotype specific. Common determinants shared by reovirus polypeptides from reovirus types 1 and 3 are most likely the stimuli for the majority of CTLs responses to reovirus.

Many years after Johann Peyer described localized accumulations of lymphoid cells in the mammalian intestinal mucosa (Peyer’s patches, PP) (1) we proposed a special role for them in the development of secretory IgA responses (2). Initially we found that rabbit PP cells were enriched sources of precursors for IgA-plasma cells and could repopulate the spleen and intestinal lamina propria of irradiated recipients with such plasma cells much more effectively than could lymphoid cells from other sources (2). This was a somewhat accidental finding made while trying to elucidate the mechanism for transporting IgA from lamina propria plasma cells into the gut lumen. Jose O’Daly had been following up the detection of secretory component (SC) in glandular epithelial cells (3) and observed that many of the apparently nonspecialized epithelial cells lining the Lieberkühn crypts contained numerous cytoplasmic granules that stained positively for both IgA and SC (4). The issue became the direction of flow — were the granules the result of endocytosis of sIgA from the intestinal fluid in the crypt lumen or were they formed at the plasma membrane on the basal side of the cells from IgA dimer secreted by the proximal plasma cells?

In this report we have shown that reovirus 1/L is an effective mucosal immunogen capable of generating a cytotoxic T cell (CTL) and associated helper T cell response to the nominal antigens associated with reovirus 1/L. The effectors that mediate reovirus-specific cytotoxicity are Thy-1+, Lyt-2+, and major histocompatibility complex (MHC)-restricted in their recognition of reovirus antigens, and can therefore be classified as CTLs. Frequency analysis of precursor CTLs occurring in Peyer's patches (PP) and peripheral lymph nodes (PLN) 6 d and 6 mo after intraduodenal stimulation have demonstrated that a persistent gradient of precursors is established, with higher frequencies present in PP. The generation of a CTL response in PP may be important in preferentially repopulating mucosal tissues with effector CTLs that could result in the local containment of infections in the gut. We also found that reovirus 1/L generates a virus-specific B cell response that is dominated by IgA memory cells after intraduodenal immunization. We hypothesize that the efficacy of reovirus 1/L at stimulating T and B cells in the gut mucosa is related to its ability to selectively enter PP via microfold (M) cells after enteric application. In this study we have also demonstrated that PP cells, upon in vitro culture and unrelated to prior reovirus priming, can generate natural killer-like (NK) cytotoxic activity. This may be an in vitro correlate of the in vivo generation of effectors that may populate mucosal tissues (i.e., the intestinal epithelium) with NK-like effector cells.

The experiments described in this report were designed to determine whether murine Peyer’s patches are capable of generating a cytotoxic T cell (CTL) response to nominal (non-MHC encoded) antigens. To date, Kagnoff (1) is the only researcher who has been able to directly demonstrate CTL activity in Peyer’s patches. He demonstrated that a cytotoxic T cell response could be detected in Peyer’s patches from mice chronically fed tumor cells that were mismatched at the major histocompatibility locus. However, he was unable to demonstrate a similar response in Peyer’s patches when mice were chronically fed tumor cells bearing minor histocompatibility differences. While his study demonstrated that a CTL response could occur in Peyer’s patches, the generation of allospecific CTL’s, whose precursors are present at high frequency among normal T cells (2), may not be representative of the potential of Peyer’s patches to generate CTL activity directed to nominal antigens. We have utilized reovirus serotype 1 (reo 1) as a model nominal antigen to determine whether intraduodenally applied virus induces a detectable CTL response in murine Peyer’s patches.

The ray method is used to study the free oscillations of an incompressible, inviscid, perfectly conducting fluid of constant density contained in a rotating spherical shell, in the presence of a constant toroidal magnetic field. For sufficiently large values of the rate of rotation and sufficiently small values of the azimuthal wave number, the oscillations are confined to seven bands of latitude for a range of frequencies, which are more bands than have been found in the non-hydromagnetic case. The phase velocity is shown to have no preference for either eastward or westward drift.