Murine Intestinal Intraepithelial Lymphocytes Research Articles

High-throughput Raman spectroscopy allows ex vivo characterization of murine small intestinal intra-epithelial lymphocytes (IEL).

T cells are considered to be critical drivers of intestinal inflammation in mice and people. The so called intra-epithelial lymphocyte (IEL) compartment largely consist of T cells. Interestingly, the specific regulation and contribution of IELs in the context of inflammatory bowel disease remains poorly understood, in part due to the lack of appropriate analysis tools. Powerful, label-free methods could ultimately provide access to this cell population and hence give valuable insight into IEL biology and even more to their disease-related functionalities. Raman spectroscopy has demonstrated over the last few years its potential for reliable cell characterization and differentiation, but its utility in regard to IEL exploration remains unknown. To address this question experimentally, we utilized a murine, T cell-driven experimental model system which is accepted to model human gut inflammation. Here, we repopulated the small intestinal IEL compartment (SI IELs) of Rag1-deficient mice endogenously lacking T cells by transferring naïve CD4+ T helper cells intraperitoneally. Using multivariate statistical analysis, high-throughput Raman spectroscopy managed to define a cell subpopulation ex vivo within the SI IEL pool of mice previously receiving T cells in vivo that displayed characteristic spectral features of lymphocytes. Raman data sets matched flow cytometry analyses with the latter identifying T cell receptor (TCR)αβ+ CD4+ T cell population in SI IELs from T cell-transferred mice, but not from control mice, in an abundance comparable to the one detected by Raman spectroscopy. Hence, in this study, we provide experimental evidence for high-throughput Raman spectroscopy to be a novel, future tool to reliably identify and potentially further characterize the T cell pool of small intestinal IELs ex vivo.

Statins directly suppress cytokine production in murine intraepithelial lymphocytes

Statins, inhibitors of the enzyme 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, are known not only as cholesterol-lowering agents but also as anti-inflammatory mediators. However, their regulatory effect on intestinal mucosal immunity remains unclear. The present study examined the possible direct effects of statin on intestinal intraepithelial lymphocytes (IELs), the front line cells of the intestinal mucosal immune system. Murine IELs were isolated from the small intestines of C57BL/6 mice. IELs activated with anti-CD3/CD28 monoclonal antibodies produced interferon (IFN)-γ, tumor necrosis factor (TNF)-α, interleukin (IL)-2, and IL-4 in significant numbers; however, they did not produce IL-5. Both simvastatin and lovastatin suppressed IEL production of IFN-γ, TNF-α, IL-2, and IL-4 in a dose-dependent manner, whereas 48-h treatment with high concentrations (5×10−5M) of simvastatin and lovastatin did not affect the number of IELs. The suppressive effect of the simvastatin was significantly restored by the addition of mevalonate, farnesyl pyrophosphate ammonium salt, and geranylgeranyl pyrophosphate ammonium salt, which are downstream metabolites of HMG-CoA. These findings suggest that statins have direct suppressive effects on the production of T helper 1-cytokines and IL-4 in IELs; these effects are associated with inhibition of the mevalonate pathway to some extent.

Norepinephrine suppresses IFN-γ and TNF-α production by murine intestinal intraepithelial lymphocytes via the β1 adrenoceptor

We examined whether norepinephrine (NE) had direct effects on cytokine production by murine intestinal intraepithelial lymphocytes (IELs), compared with splenocytes. CD3+ IELs and CD3+ splenocytes expressed α1B, α1D, α2C, β1, β2, and β3 adrenoceptors (ARs). NE significantly suppressed IFN-γ and TNF-α production by IELs and splenocytes ex vivo. The suppressive effects of NE in IELs were reversed by β1 AR antagonist CGP-20712A, whereas those in splenocytes were reversed by β2 AR antagonist ICI118,551. In IELs, β1 AR agonist xamoterol mimicked the suppressive effects of NE. These results indicated NE regulates intestinal mucosal immune responses mediated by IELs via β1 AR.

Glutamine supplementation increases Th1-cytokine responses in murine intestinal intraepithelial lymphocytes

Intestinal intraepithelial lymphocytes (IELs) are major effector cells in the gut mucosal immune system, and are phenotypically distinct from thymic and peripheral T cells. Although nutritional supplementation with glutamine affects the intestinal immune response, it remains unclear whether this is a direct effect via the IEL-derived cytokines. This study examined changes in IEL-derived cytokine production following treatment with glutamine in vitro. Murine IELs were purified and activated with PMA plus ionomycin, and then cultured in the presence of various glutamine concentrations. IEL-derived cytokines were measured using a cytometric bead array (CBA) system, and IEL subsets were analyzed by flow cytometry. Treatment with glutamine increased the production of IL-2 and IFN-γ from IELs in the presence of PMA plus ionomycin, but had no effect on TNFα, IL-4, or IL-5 production. Treatment with alanine or glucose had no regulatory effect on IEL-derived cytokines. Glutamine therefore had a direct effect on the production of selected IEL-derived Th1-cytokines, and enteral supplementation with glutamine may influence the intestinal immune responses mediated by IELs.

Stimulatory and costimulatory effects of IL-18 directed to different small intestinal CD43 T cell subsets

This study has examined the stimulatory and costimulatory effects of IL-18 on two subsets of murine small intestinal intraepithelial lymphocytes (IELs) defined by the expression of the CD43 S7 glycoform. Data from gene array studies and real-time PCR indicated that S7(+) IELs had significantly higher levels of gene expression for the IL-18 receptor and the IL-18R accessory protein than S7(-) IELs. IL-18 costimulation of IELs in conjunction with CD3-induced activation resulted in significantly greater proliferation than CD3 stimulation alone. In CFSE dilution experiments, IL-18 costimulation favored the S7(+) IEL population. IL-18 costimulation did not affect apoptosis of either S7(-) or S7(+) IELs compared with CD3 stimulation alone. Although IL-18 costimulation did not alter the total number of IFN-gamma-producing cells relative to CD3 stimulation alone, twice as many S7(+) IELs were IFN-gamma -secreting cells than S7(-) IELs in both CD3-stimulated and IL-18-costimulated cultures. Notably, direct IL-18 stimulation in the absence of CD3 activation induced an IFN-gamma response that was predominantly directed to the S7(+) population, indicating that IL-18 is itself an IFN-gamma activational signal for intestinal T cells. In contrast, direct IL-18 stimulation of IELs did not generate TNF-alpha-producing cells, indicating a differential response in the activation of proinflammatory cytokines following IL-18 exposure. These findings point to distinctly different activational effects of IL-18 on IELs, both with regard to the type of functional responses elicited and with respect to the IEL subsets affected.

Upregulation of ICOS on CD43+ CD4+ murine small intestinal intraepithelial lymphocytes during acute reovirus infection

Murine intestinal intraepithelial lymphocytes (IELs) can be classified according to expression of a CD43 glycoform recognized by the S7 monoclonal antibody. In this study, we examined the response of S7+ and S7− IELs in mice during acute reovirus serotype 3 (Dearing strain) infection, which was confirmed by virus-specific real-time PCR. In vivo proliferation increased significantly for both S7− and S7+ IELs on day 4 post-infection as determined by BrdU incorporation; however, expression of the inducible costimulatory (ICOS) molecule, which peaked on day 7 post-infection, was upregulated on S7+ CD4+ T cells, most of which were CD4+8− IELs. In vitro ICOS stimulation by syngeneic peritoneal macrophages induced IFN-γ secretion from IELs from day 7 infected mice, and was suppressed by treatment with anti-ICOS mAb. Additionally, IFN-γ mRNA increased in CD4+ IELs on day 6 post-infection. These findings indicate that S7− and S7+ IELs are differentially mobilized during the immune response to reovirus infection; that the regulated expression of ICOS is associated with S7+ IELs; and that stimulation of IELs through ICOS enhances IFN-γ synthesis during infection.

Effect of foot shock stress on the interferon-γ production of murine intestinal intraepithelial lymphocytes

To investigate the effect of stress on interferon (IFN)-γ production by intestinal intraepithelial lymphocytes (IEL), we exposed male C3H/HeN mice to electric foot shock for 30 min a day for 5 consecutive days. Immediately after the final foot shock stress, IEL from small intestine were isolated by Percoll density gradient. The stress induced a marked suppression of IFN-γ production by IEL stimulated with immobilized anti-CD3 mAb and a marked decrease in the proportion of IFN-γ-producing CD3 + IEL or αβTCR + IEL stimulated with PMA + ionomycin. The αβTCR + subset was the major cause of stress-induced suppression of IFN-γ production by IEL. Glucocorticoid induced the suppression of IFN-γ production by IEL in vitro, which was reversed by mifepristone (RU486), a glucocorticoid receptor antagonist. In vivo administration of RU486 reversed the stress-induced suppression of IFN-γ production by IEL. In conclusion, repeated foot shock stress suppressed IFN-γ production of IEL by stress-induced elevation of endogenous glucocorticoid. Substantial suppression of the αβTCR + subset was the major cause of the suppression.

Repeated Stress Suppresses Interferon-.GAMMA. Production by Murine Intestinal Intraepithelial Lymphocytes

Intestinal intraepithelial lymphocytes (IEL), one of the major effector components in the mucosal immune system, are phenotypically and functionally distinct from thymic and peripheral T cells. To investigate the effect of repeated stress on the number and function of IEL, we exposed male C3H/HeN mice to mild electric foot shock for 30 min/day for 5 consecutive days. Immediately after the final foot shock stress, the blood, spleen, thymus and small intestine of each of the mice were obtained. As a functional measure, we evaluated interferon (IFN)- gamma production by IEL, since IFN-gamma is a key immunomodulating cytokine in mucosal immune responses. Serum corticosterone level was elevated immediately after foot shock stress. There were no significant changes in the number of whole IEL and CD3+ IEL subsets after the stress. In contrast, the stress led to a significant decrease in the total number of thymocytes, particularly the reduction in the number of CD4+CD8+ thymocytes. Thymocytes expressed the highest level of intracellular glucocorticoid receptor (GR), followed by splenocytes and IEL. The foot shock stress induced a marked suppression of IFN-gamma production by IEL, when stimulated with immobilized anti-CD3 monoclonal antibody. Furthermore, corticosterone suppressed the IFN-gamma production by cultured IEL, which was prevented by Mifepristone (RU486), a GR antagonist. In summary, repeated foot shock stress did not alter the numbers of IEL and CD3+ IEL subsets, but suppressed IFN-gamma production by IEL, which was probably mediated by the elevated corticosterone. We therefore propose that stress influences host defense by suppressing the production of IFN-gamma in IEL.

Maximum Immunobioactivity of Murine Small Intestinal Intraepithelial Lymphocytes Resides in a Subpopulation of CD43+ T Cells

CD43 has been linked to many function-associated T cell activities. Using mAbs that recognize two different CD43 determinants, we show that, although mouse small intestinal intraepithelial lymphocytes (IELs) expressed the CD43 core molecule reactive with mAb R2/60, only about one-half of the total IELs-including some but not all of the TCRalphabeta and TCRgammadelta cells-expressed the CD43 S7(-) reactive determinant. CD43 S7(+) IELs secreted more IL-2, IL-4, IL-10, IL-17, and IFN-gamma following anti-CD3 stimulation, and were >4-fold more cytotoxic in fresh isolates and >16-fold more cytotoxic after anti-CD3 stimulation, than S7(-) IELs. S7(+) but not S7(-) IELs from the ileum of IL-10(-/-) mice spontaneously produced IFN-gamma. In vivo BrdU uptake by IELs in non-Ag-primed mice was greatest in the S7(+) population, indicating that significantly more S7(+) IELs than S7(-) IELs undergo cell expansion under normal homeostatic conditions. DNA microarray analyses showed that S7(+) IELs expressed higher levels of genes associated with activated T cells, whereas S7(-) IELs expressed genes used in the regulation of NK cells. These findings define two functionally distinct populations of IELs based on CD43 expression independent of TCR class, and they identify a subset of IELs that may serve as a target to better control intestinal inflammation.

Functionally active CD8alphabeta+ TCRgammadelta intestinal intraepithelial lymphocytes in athymic nu/nu mice.

Murine intestinal intraepithelial lymphocytes (IEL) encompass a high proportion of TCRgammadelta cells. A vast majority of these TCRgammadelta IEL express CD8alpha, but not CD8beta (CD8alphaalpha homodimer), and are considered to develop in intestinal epithelial layers independently of a functional thymus. Here we show that TCRgammadelta cells expressing both CD8alpha and CD8beta (CD8alphabeta heterodimer) appear in athymic nu/nu mice, although their appearance is random. The IEL comprising CD8alphabeta(+) TCRgammadelta cells expressed pronounced cytolytic and IFN-gamma-producing activities after TCRgammadelta ligation, which were markedly stronger than activities of IEL lacking CD8alphabeta(+) TCRgammadelta cells. Purified CD8alphabeta(+) TCRgammadelta cells expressed strong cytolytic activities and produced large quantities of IFN-gamma after TCR engagement. CD8alphabeta(+) TCRgammadelta cells were also identified among IEL from euthymic C57BL/6 mice, although their abundance varied among individual animals. However, cytolytic and IFN-gamma-producing activities in euthymic C57BL/6 mice were markedly lower than those in athymic nu/nu mice. Our findings suggest that CD8alphabeta(+) TCRgammadelta cells can develop in the intestine independently of a functional thymus/thymic epithelial cells and that they perform biological functions in situ.

CD43 potentiates CD3-induced proliferation of murine intestinal intraepithelial lymphocytes.

The involvement of CD43 in cell proliferation of murine intestinal intraepithelial lymphocytes (IEL) has been studied in in vitro CD3-stimulated cell cultures. In the presence of either IL-2 or IL-15, CD3 stimulation of IEL resulted in low levels of proliferation as measured by thymidine incorporation, whereas no proliferation occurred upon CD3 stimulation in the absence of cytokines. The combination of both cytokines to IEL cultures synergistically enhanced CD3-induced proliferation by approximately threefold that of cultures supplemented with either cytokine alone. Most importantly, however, proliferation of IEL was significantly greater when CD3 stimulation occurred in conjunction with CD43 triggering, indicating that CD43 functions as a coactivational signal for murine IEL. These findings indicate that a spectrum of potential proliferative responses exist among murine IEL depending on the types and combinations of signals received, and that because under normal conditions murine IEL are largely devoid of CD28 expression, a classical T-cell coactivational molecule, the capacity for high-level IEL proliferation may reside with CD43.

Qa-2–Dependent Selection of Cd8α/α T Cell Receptor α/β1 Cells in Murine Intestinal Intraepithelial Lymphocytes

Qa-2–Dependent Selection of Cd8<b>α/α</b> T Cell Receptor <b>α/β</b>1 Cells in Murine Intestinal Intraepithelial Lymphocytes

Origins of intestinal intraepithelial lymphocytes: direct evidence for a thymus-derived γδ T cell component

The contribution of T cell precursors from the thymus and the bone marrow to the pool of intestinal intraepithelial lymphocytes (IELs) has been studied in a system using donor cells from enhanced-green fluorescent protein (EGFP +) transgenic mice adoptively transferred into EGFP − recipient mice. Consistent with previous studies, regeneration of γδ and αβ T cell populations in the intestinal epithelium occurred within 2–3 weeks of bone marrow transfer into irradiatiated EGFP − animals and prior to T cell repopulation of the spleen, of interest, however, although transfer of whole adult EGFP + thymocytes to non-irradiated EGFP − congenitally-athymic nude mice produced αβ T cells in both the spleen and intestine, γδ T cells in significant numbers were detected only in the intestine of recipient mice. In contrast, transfer of CD3 −, CD4 −, CD8 − immature thymocytes resulted in no detectable T cells in either the intestine or the spleen of nude mice up to twelve weeks post-cell transfer, suggesting that intestinal IELs generated from thymocytes arose from differentiated lineage-committed cells rather than from immature thymocytes. These findings provide direct evidence for both thymus-independent and thymus-dependent sources of intestinal γδ T cells, and they suggest that murine IELs consist of diverse groups of T cells with distinct developmental origins.

Qa-2-dependent selection of CD8alpha/alpha T cell receptor alpha/beta(+) cells in murine intestinal intraepithelial lymphocytes.

Murine intestinal intraepithelial lymphocytes (iIELs) are made up of a heterogeneous mix of T cells with unique phenotypes. Whereas CD8(+) T cells in peripheral lymphoid organs use CD8alpha/beta and are selected on MHC class Ia molecules, a majority of iIELs use CD8alpha/alpha. Here, we report that the presence of CD8alpha/alpha TCR-alpha/beta cells in iIELs is independent of classical MHC class I molecules K(b) and D(b), as illustrated by their presence in K(b)/D(b) double-knockout mice and in mice lacking a nonclassical MHC class I molecule, CD1d. Most strikingly, their presence is decreased by approximately 70% in mice lacking transporter associated with antigen processing (TAP). The TAP-dependent nonclassical MHC class I molecule Qa-2 is strongly implicated in the presence of these cells, as inferred from the low numbers of CD8alpha/alpha TCR-alpha/beta T cells in mice deficient in Qa-2 genes. Second, a Qa-2-transgenic mouse made in a Qa-2(-) strain showed an increase in the numbers of CD8alpha/alpha cells among its iIELs. Thus, the presence of CD8alpha/alpha TCR-alpha/beta cells in iIELs is mainly dependent on the nonclassical MHC class I molecule Qa-2.

Functional role of CD95 ligand in concanavalin A-induced intestinal intraepithelial lymphocyte cytotoxicity.

Freshly isolated murine intestinal intraepithelial lymphocytes (IEL) express CD95 ligand (CD95L), as shown by reverse transcription-polymerase chain reaction (RT-PCR) and fluorescence-activated cell sorter (FACS) analysis. Between 15 and 25% of IEL could be stained with an antibody to CD95L. Therefore it was investigated whether the CD95L/CD95 pathway was effective in IEL cytotoxicity. Stimulation of IEL in vitro with concanavalin A (Con A) induced a strong cytotoxic response, which was much higher when using CD95-expressing target cells. This effect was most evident when comparing the specific lysis of CD95-transfected target cells of the leukaemia cell line L1210 with that of the untransfected parental cell line. In addition, an antibody to CD95 was able to dramatically reduce the specific lysis of CD95-expressing target cells. After stimulation with Con A, which is able to bind to CD95L, the effects were more obvious compared with the triggering of the T-cell receptor (TCR)-alphabeta or gamma delta. On the other hand, EGTA reduced the Con A-induced cytotoxicity. Together these findings support a role of the CD95L/CD95 pathway in IEL cytotoxicity, even though the reaction was Ca2+ sensitive. As a function, CD95L-expressing IEL should be able to contribute to the elimination of CD95-expressing target cells in the intestine.

Differential Requirements of CD45 Protein Tyrosine Phosphatase for Cytolytic Activities and Intrathymic and Extrathymic Development of Intestinal Intraepithelial Lymphocytes

AbstractCD45 is a transmembrane protein tyrosine phosphatase essential for Ag receptor-mediated signaling in both T and B cells. In this study we investigated roles of CD45 in development and cytolytic activities of murine intestinal intraepithelial lymphocytes (i-IEL) using CD45 exon 6 knockout (CD45−/−) mice. Interestingly, the total cell number of i-IEL was significantly reduced in CD45−/− mice during aging (10–20 wk of age), whereas the i-IEL number was normally increased in the wild-type littermates. Especially, the number of γδTCR+ i-IEL decreased markedly in CD45−/− mice during aging. The i-IEL in CD45−/− mice were more susceptible to in vitro spontaneous apoptosis than the normal i-IEL, implying that CD45 is required for maintenance of the cellularity of i-IEL. Results from in vivo analyses of the extrathymic and intrathymic development of i-IEL suggested that CD45-mediated signaling is required for the intrathymic, but not the extrathymic, development of i-IEL. Moreover, the whole i-IEL from CD45−/− mice showed a significantly reduced cytolytic activity, and the residual cytolytic activity was completely diminished by depleting CD45+ i-IEL, suggesting that CD45 is indispensable for the TCR-mediated cytolytic activity of i-IEL. Furthermore, we found differential contributions of CD45 and p56lck to development and induction of cytolytic activities of i-IEL.

Expansion of a novel pulmonary CD3(-) CD4(+) CD8(+) cell population in mice during Chlamydia pneumoniae infection.

A new pulmonary T-cell-like lymphocyte population with the phenotype CD3(-) CD4(+) CD8(+) was discovered in mice. CD4(+) CD8(+) but CD3(+) cells among murine intestinal intraepithelial lymphocytes have previously been described. We describe herein a dramatic expansion of the CD3(-) CD4(+) CD8(+) cell population in response to experimental respiratory infection. After intranasal Chlamydia pneumoniae infection, CD4(+) CD8(+) cells became transiently the dominant lymphocyte type (maximum of 87% of all lymphocytes) in the lungs of NIH/S mice but remained virtually undetectable in spleen and blood. The enrichment of these cells was not a C. pneumoniae-specific event, since infection of NIH/S mice with influenza A virus also resulted in an increase in the number of CD4(+) CD8(+) cells (maximum of 42% of all lymphocytes). In addition to outbred NIH/S mice, two other mouse strains were studied: BALB/c (H-2(d)) and C57BL/6 (H-2(b)). C. pneumoniae-infected BALB/c mice responded with an intermediate increase in the number of CD4(+) CD8(+) cells in lungs, whereas C57BL/6 mice did not respond. The double-positive CD4(+) CD8(+) cells lacked a major part of the T-cell receptor complex, being both CD3(-) and TCR alpha beta-. However, when they were stimulated in vitro with a T-cell mitogen, they responded by proliferation but did not secrete gamma interferon. The dramatic expansion of this cell population at the infection site suggests an active role for them in respiratory infection, but the specification of this requires further study.

A step-wise expansion of intestinal intraepithelial T lymphocytes in association with microbial colonization is defined by sensitivity to cyclosporin A.

Murine intestinal intraepithelial lymphocytes (IELs) consist of T cells bearing alpha beta-antigen receptor (alpha beta-IELs) and those bearing gamma delta-IELs). Although gamma delta-IELs outnumber alpha beta-IELs in germ-free (GF) mice, oral inoculation of fecal suspension from conventional (CV) mice into GF mice induced the increase in number of alpha beta-IELs, leaving the number of gamma delta-IELs unchanged, and the number of alpha beta-IELs reached the level of CV mice by 3 weeks after conventionalization. Expansion of alpha beta-IELs and increase in their CD44+ subset in conventionalized mice were not affected until 2 weeks after beginning of daily injection of cyclosporin A (CsA). However, further expansion of alpha beta-IELs during 2-3 weeks after conventionalization was blocked by injection of CsA. Although the relative constitution of CD4- 8-, CD4+ 8-, CD4- 8 alpha alpha+, CD4- 8 alpha beta+ and CD4+ 8+ subsets among alpha beta-IELs was comparable between control and CsA-treated groups, CsA injection resulted in the decrease in ratio of high-density fraction cells to low density fraction cells in IELs. CsA completely abrogated the expansion of T cells in peripheral lymph nodes stimulated by alloantigens in vivo, and proliferation of IELs from GF mice induced by immobilized anti-alpha beta-T-cell receptor (TCR) monoclonal antibodies (mAb) in vitro was also eliminated by CsA. These results indicate that microbial colonization-induced expansion of alpha beta-IELs is subdivided into two steps: the early phase of expansion takes place via TCR-non-mediated pathway and the late phase of expansion requires TCR-mediated signal transduction.

Effects of a Nonapeptide Thymic Hormone on Intestinal Intraepithelial Lymphocytes in Mice Following Administration of 5-Fluorouracil

A significant fraction of murine small intestinal intraepithelial lymphocytes (i-IELs) maturate in local sites outside the thymus. However, there is evidence suggesting that extrathymic differentiation of i-IELs is still influenced by the thymus or thymus-derived factors. Facteur thymique serique (FTS), a nonapeptide thymic hormone, is involved in several aspects of intra- and extrathymic T cell differentiationin vivo.In this study, we investigated the effects of FTS on the kinetics of i-IELs in mice following a single administration of 5-fluorouracil (5-FU). FTS treatment significantly accelerated the recovery in cell number of i-IELs after administration of 5-FU. Flow cytometric analysis revealed that this accelerated recovery was mainly due to a rapid increase in CD8αα+i-IELs. Similar findings were also evident in adult thymectomized (ATX) mice, indicating that FTS treatment caused a rapid recovery of CD8αα+i-IELs following 5-FU administration in the absence of a functional thymus. Furthermore, expression levels of the mRNAs for interleukin-2, interferon-γ, and transforming growth factor β1in the i-IELs were augmented by FTS treatment. Notably, FTS treatment protected mice from 5-FU-induced lethal toxicity, accompanied with an inhibition of the translocation ofEnterobactericeae.These results suggest that FTS has an important function in the extrathymic maturation and activation of i-IELs in the small intestine following 5-FU administration, which may contribute at least partly to the protection against 5-FU-induced lethal toxicity.

Interleukin 2 (IL-2) and interleukin 7 (IL-7) reciprocally induce IL-7 and IL-2 receptors on gamma delta T-cell receptor-positive intraepithelial lymphocytes.

In this study, we describe the interaction between cytokine and cytokine receptor (R) for the activation and proliferation of gamma delta T-cell receptor-positive T cells (gamma delta T cells). gamma delta T cells isolated from murine intestinal intraepithelial lymphocytes (IELs) were separated into gamma delta (Dim) and gamma delta (Bright) fractions according to the intensity of gamma delta T-cell receptor expression. The gamma delta T cells express low levels of IL-2R and IL-7R as shown by flow cytometry and reverse transcriptase-PCR analysis, whereas gamma delta (Bright) T cells did not express either receptor. Our study also revealed that recombinant marine (rm)IL-2 and rmIL-7 reciprocally induced high expressions of IL-7R and IL-2R, respectively, on gamma delta (Dim) T cells but not on gamma delta (Bright) cells. Thus, treatment of gamma delta (Dim) T cells with rmIL-2 and rmIL-7 resulted in high proliferative responses, whereas gamma delta (Bright) T cells did not respond to these two cytokines. The sources of these two cytokines for gamma delta T cells were neighboring epithelial cells (IL-7) and alpha beta T cells (IL-2 and IL-7). Cytokine signaling by IL-2 and IL-7 from alpha beta T cells and epithelial cells was necessary for the expression of IL-7R and IL-2R, respectively, on a subset of gamma delta T cells (e.g., gamma delta (Dim) T cells) in mucosa-associated tissue for subsequent activation and cell division.