TRAIL expression by activated human CD4(+)V alpha 24NKT cells induces in vitro and in vivo apoptosis of human acute myeloid leukemia cells

Human Valpha24NKT cells are activated by alpha-galactosylceramide (alpha-GalCer)-pulsed dendritic cells in a CD1d-dependent and a T-cell receptor-mediated manner. Here, we demonstrate that CD4(+)V alpha 24NKT cells derived from a patient with acute myeloid leukemia (AML) M4 are phenotypically similar to those of healthy donors and, in common with those derived from healthy donors, express tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) when the cells are activated by alpha-GalCer-pulsed dendritic cells but not prior to activation. We also show that myeloid leukemia cells from patients with AML M4, but not from patients with AML M0 or M1, undergo apoptosis following culture with TRAIL-expressing autologous or allogeneic healthy donor V alpha 24NKT cells. Apoptosis of AML M4 leukemia cells from patient peripheral blood was almost completely blocked by a neutralizing monoclonal antibody against TRAIL, indicating that TRAIL on V alpha 24NKT cells is essential for the induction of apoptosis in AML M4 leukemia cells. A nonobese diabetic-severe combined immunodeficient human leukemia (AML M4) model showed that human activated CD4(+)V alpha 24NKT cells induced apoptosis of human leukemia cells in vivo. This is the first evidence that activated V alpha 24NKT cells express TRAIL and that TRAIL causes apoptosis of monocytic leukemia cells from patients with AML M4 in vitro and in vivo. Adoptive immune therapy with activated V alpha 24NKT cells, or other strategies to increase activated V alpha 24NKT cells in vivo, may be of benefit to patients with AML M4. (Blood. 2001;97:2067-2074)     

Two distinct alpha beta T-cell lineages can be distinguished by the differential usage of T-cell receptor V beta gene segments.

Avian T cells can be divided into three subpopulations based on their expression of distinct T-cell receptors (TCR1, TCR2, and TCR3), ontogeny, and tissue distribution. The TCR1 cells appear to be the equivalent of mammalian gamma delta cells, but the derivation of cells expressing TCR2 and TCR3 has been unclear. Here we report that chickens contain two families of TCR beta variable (V) gene segments, V beta 1 and V beta 2. Furthermore, TCR2 and TCR3 represent subsets of alpha beta cells that are defined by mutually exclusive usage of these two families of V beta gene segments. Sequence comparisons of V beta 1 and V beta 2 with mammalian TCR beta V segments reveal that V beta 1 gene segments encode the conserved amino acids used to define the mammalian V beta consensus subgroup I, while V beta 2 encodes the amino acids used to define the mammalian V beta subgroup II. Although the beta chains of TCR2 and TCR3 cells are encoded by the same diversity (D), joining (J), and constant (C) region segments, V beta 1 gene segments undergo rearrangement before V beta 2 gene segments during T-cell development. This may result from the fact that TCR2 cells undergo V-DJ joining by deletional rearrangement, whereas TCR3 cells undergo V-DJ joining by inversional rearrangement. These data suggest that the TCR alpha beta cells can be divided into two distinct and evolutionarily conserved lineages based on V beta gene segment usage. The clear-cut separation of these lineages in the chicken may help to define their immunologic role.     

T cell receptor V beta repertoire of double-negative alpha/beta T cells in patients with systemic sclerosis.

OBJECTIVE. To analyze the T cell receptor V beta gene on double-negative (DN) alpha/beta T cells, which are increased in number, on peripheral blood lymphocytes (PBL) from patients with systemic sclerosis (SSc). METHODS. The DN alpha/beta T cells were sorted by flow cytometry from PBL obtained from 3 patients with SSc. The V beta repertoire was analyzed by polymerase chain reaction. RESULTS. Only 1 or 2 V beta genes (V beta 5/7, 5, or 17) were predominantly expressed on DN alpha/beta T cells from these 3 patients. CONCLUSION. The V beta repertoire on DN alpha/beta T cells in PBL from patients with SSc is rather restricted.     

Anatomic localization of immature and mature dendritic cell subsets in dermatomyositis and polymyositis: Interaction with chemokines and Th1 cytokine-producing cells

OBJECTIVE: To clarify the involvement of dendritic cells (DCs), chemokines, and proinflammatory Th1 cytokines in the pathogenesis of the chronic muscle diseases dermatomyositis (DM) and polymyositis (PM). METHODS: We characterized by immunohistochemistry the DC subsets and their interaction with cells producing chemokines and the Th1 cytokines interleukin-17 (IL-17) and interferon-gamma (IFNgamma). Immature and mature DCs were defined by the expression of CD1a and DC-LAMP/CD83, respectively. RESULTS: Immature DCs were mainly detected in lymphocytic infiltrates in DM and PM muscle tissue samples. Mature DCs were detected in perivascular infiltrates and surrounded muscle fibers. IL-17-positive and IFNgamma-positive cells were also observed in perivascular infiltrates in both cases. We then focused on the expression of the CCL20/CCR6 chemokine/receptor complex, which controls immature DC migration, and on the expression of the CCL19/CCR7 and CCL21/CCR7 chemokine/receptor complexes, which control mature DC migration. CCL20 and CCR6 colocalized in lymphocytic infiltrates in DM and PM samples. CCL21 was rarely observed in DM samples and never observed in PM samples. CCL19- and CCR7-expressing cells were absent in both tissues. CONCLUSION: The close association between CCL20/CCR6 and immature DCs suggests the contribution of CCL20 to CCR6+ immature DC homing. Detection of mature DCs in DM and PM muscle tissue samples despite the lack of CCL19 and CCR7 is evidence for a local maturation of DCs in inflammatory muscle tissue without lymphoid organ organization.     

Estrogen receptor alpha and beta expression in theca and granulosa cells from women with polycystic ovary syndrome

A defining characteristic of dominant follicles is high estradiol concentrations. Abnormal expression of estrogen receptors (ERs) could contribute to poor follicular development and ovulatory failure in polycystic ovary syndrome (PCOS). The aim of this study was to determine whether there are differences in ERalpha and ERbeta expression in granulosa cells (GC) and theca cells (TC) from women with PCOS, compared with regularly cycling women. GC and TC were obtained by microdissection from 12 polycystic and 23 normal ovaries. ERalpha and ERbeta mRNA and protein expression were measured by semiquantitative RT-PCR and Western blot, respectively. In control ovaries, both GC and TC ERalpha mRNAs were higher in small antral (SA) than in dominant follicles. ERalpha mRNA was similar in PCOS and size-matched control follicles. In control follicles, ERalpha protein concentrations were higher in GC than in TC. In GC, the ERalpha concentrations were comparable among SA, dominant, and PCOS follicles. In TC, ERalpha concentrations were lower in dominant follicles but were markedly increased in PCOS. In control ovaries, GC and TC expression of ERbeta mRNA was higher in SA, compared with dominant follicles. In PCOS, ERbeta mRNA was intermediate between SA and dominant follicles in both GC and TC. In GC, the ERbeta protein concentrations followed the same pattern as mRNA expression; but in TC ERbeta, protein in PCOS was equivalent to that in dominant follicles. The results of this study demonstrate that there are significant alterations in the expression of ERalpha and ERbeta in PCOS that may be related to abnormal follicular development.     

Interferon-alpha and -beta differentially regulate osteoclastogenesis: role of differential induction of chemokine CXCL11 expression

In humans, type I interferon (IFN) is a family of 17 cytokines, among which the alpha subtypes and the beta subtype are differentially expressed. It has been suggested that IFN-beta activates a specific signaling cascade in addition to those activated by all type I IFNs. Nevertheless, no true biological relevance for a differential activity of alpha and beta IFN subtypes has been identified so far. Because type I IFNs are critical for the regulation of osteoclastogenesis in mice, we have compared the effect of IFN-alpha2 and IFN-beta on the differentiation of human monocytes into osteoclasts. Primary monocytes undergoing osteoclastic differentiation are highly and equally sensitive to both alpha2 and beta IFNs as determined by measuring the induction levels of several IFN-stimulated genes. However, IFN-beta was 100-fold more potent than the alpha2 subtype at inhibiting osteoclastogenesis. Expression profiling of the genes differentially regulated by IFN-alpha2 and IFN-beta in this cellular system revealed the chemokine CXCL11 as the only IFN-induced gene differentially up-regulated by IFN-beta. We show that recombinant CXCL11 by itself inhibits osteoclastic differentiation. These results indicate that autocrine-acting CXCL11 mediates, at least in part, the regulations of osteoclastogenesis by type I IFNs.     

Gene expression of glucocorticoid receptor alpha and beta in giant cell tumour of bone: evidence of glucocorticoid-stimulated osteoclastogenesis by stromal-like tumour cells

Recent evidences point to a role of estrogens in males. We have earlier reported that tamoxifen, a synthetic non-steroidal antiestrogen, when administered to adult male rats, in the dose range of 0.04-0.4 mg/kg per day, reduced fertility. The reduced fertility was measured in terms of fertility index (a measure of the efficiency of the ovulated ovas to fertilise and implant), fecundity (siring ability) and litter size. The present study was done to investigate whether the reduction in fertility index was due to reduction in fertilising ability or increase in pre-implantation embryo loss. Also a dose related effect of tamoxifen from 0.02 mg to 2 mg/kg per day on the fertility of the male rats was studied. To study the fertilising ability, control and tamoxifen (0.4 mg/kg per day, the most effective dose) treated adult male rats were mated with normal cycling females and the females sacrificed at day 0-4 of gestation. Eggs fertilised/unfertilised were flushed from the oviduct/uterus and the number and types of eggs were noted. The index of fertilisation, a measure of the fertilising ability was determined. The studies demonstrate that the reduction in fertility is not due to decreased fertilising ability but because of the increased pre-implantation embryo loss as evident from an increase in number of abnormal eggs in the treated group with no change in index of fertilisation. A dose related decrease in fertility was observed. The present study suggests that tamoxifen at 0.02-2-mg dose is predominantly estrogenic in males and paternal factor/s sensitive to tamoxifen is involved in embryogenesis.     

Androgen receptor, estrogen receptor alpha, and estrogen receptor beta show distinct patterns of expression in forebrain song control nuclei of European starlings.

In songbirds, singing behavior is controlled by a discrete network of interconnected brain nuclei known collectively as the song control system. Both the development of this system and the expression of singing behavior in adulthood are strongly influenced by sex steroid hormones. Although both androgenic and estrogenic steroids have effects, androgen receptors (AR) are more abundantly and widely expressed in song nuclei than are estrogen receptors (ER alpha). The recent cloning of a second form of the estrogen receptor in mammals, ER beta, raises the possibility that a second receptor subtype is present in songbirds and that estrogenic effects in the song system may be mediated via ER beta. We therefore cloned the ER beta complementary DNA (cDNA) from a European starling preoptic area-hypothalamic cDNA library and used in situ hybridization histochemistry to examine its expression in forebrain song nuclei, relative to the expression of AR and ER alpha messenger RNA (mRNA), in the adjacent brain sections. The starling ER beta cDNA has an open reading frame of 1662-bp, predicted to encode a protein of 554 amino acids. This protein shares greater than 70% sequence identity with ER beta in other species. We report that starling ER beta is expressed in a variety of tissues, including brain, pituitary, skeletal muscle, liver, adrenal, kidney, intestine, and ovary. Similar to reports in other songbird species, we detected AR mRNA-containing cells in several song control nuclei, including the high vocal center (HVc), the medial and lateral portions of the magnocellular nucleus of the anterior neostriatum, and the robust nucleus of the archistriatum. We detected ER alpha expression in the medial portion of HVc (also called paraHVc) and along the medial border of the caudal neostriatum. ER beta was not expressed in HVc, in the medial and lateral portions of the magnocellular nucleus of the anterior neostriatum, in the robust nucleus of the archistriatum, or in area X. In contrast, ER beta mRNA-containing cells were detected in the caudomedial neostriatum and medial preoptic area in a pattern reminiscent of P450 aromatase expression in the same brain regions in other songbirds. These data suggest that estrogenic effects on the song system are not mediated via ER beta-producing cells within song nuclei. Nonetheless, the overlapping expression of ER beta- and aromatase-producing cells in the caudomedial neostriatum suggests that locally synthesized estrogens may act via ER beta, in addition to ER alpha, to mediate seasonal or developmental effects on nearby song nuclei (e.g. HVc).     

Hormonal regulation of estrogen receptor alpha and beta gene expression in human granulosa-luteal cells in vitro

Estrogen is one of the major sex steroid hormones that is produced from the human ovary, and its actions are established to be a receptor-mediated process. Despite the demonstration of estrogen receptor (ER) expression, little is known regarding the regulation of ER in the human ovary. In the present study we investigated the expression and hormonal regulation of ERalpha and ERbeta in human granulosa-luteal cells (hGLCs). Using RT-PCR amplification, both ERalpha and ERbeta messenger ribonucleic acid (mRNA) were detected from hGLCs. Northern blot analysis revealed that ERalpha is expressed at a relatively lower level than ERbeta. Basal expression studies indicated that ERalpha mRNA levels remain unchanged, whereas ERbeta mRNA levels increased with time in culture in vitro, suggesting that ERbeta is likely to play a dynamic role in mediating estrogen action in hGLCs. The regulation of ERalpha and ERbeta expression by hCG was examined. hCG treatment (10 IU/mL) significantly attenuated the ERalpha (45%; P < 0.01) and ERbeta (40%; P < 0.01) mRNA levels. The hCG-induced decrease in ERalpha and ERbeta expression was mimicked by 8-bromo-cAMP (1 mmol/L) and forskolin (10 micromol/L) treatment. Additional studies using a specific protein kinase A (PKA) inhibitor (adenosine 3',5'-cyclic monophosphorothioate, Rp-isomer, triethylammonium salt) and an adenylate cyclase inhibitor (SQ 22536) further implicated the involvement of the cAMP/PKA signaling pathway in hCG action in these cells. The hCG-induced decrease in ERalpha and ERbeta mRNA levels was prevented in the presence of these inhibitors. Next, the effect of GnRH on ER expression was studied. Sixty-eight percent (P < 0.001) and 60% (P < 0.001) decreases in ERalpha and ERbeta mRNA levels, respectively, were observed after treatment with 0.1 micromol/L GnRH agonist (GnRHa). Pretreatment of the cells with a protein kinase C (PKC) inhibitor (GF109203X) completely reversed the GnRHa-induced down-regulation of ERalpha and ERbeta expression, suggesting the involvement of PKC in GnRH signal transduction in hGLCs. In agreement with the semiquantitative RT-PCR results, Western blot analysis detected a decrease in ERalpha and ERbeta proteins levels in hGLCs after treatment with hCG (10 IU/mL), GnRH (0.1 micromol/L), 8-bromo-cAMP (1 mmol/L), forskolin (10 micromol/L), or phorbol 12-myristate 13 acetate (10 micromol/L). Functionally, we demonstrated an inhibition of progesterone production in hGLCs in vitro by 17beta-estradiol, and this inhibitory effect was eliminated by pretreatment of 10 IU/mL hCG or 0.1 micromol/L GnRHa for 24 h before 17beta-estradiol administration. In summary, we observed a differential expression of ERalpha and ERbeta mRNA in hGLCs in vitro. The demonstration of hCG- and GnRHa-induced down-regulation of ERalpha and ERbeta gene expression suggests that hCG and GnRH may contribute to the control of granulosa-luteal cell function. Furthermore, our data suggest that the effects of hCG and GnRH on ERalpha and ERbeta expression in hGLCs are mediated in part by activation of PKA and PKC signaling pathways, respectively.     

Estrogen receptors alpha and beta in rat decidua cells: cell-specific expression and differential regulation by steroid hormones and prolactin

Estradiol is known to play an important role in the growth and differentiation of rat uterine stromal cells into decidual cells. In particular, this hormone with progesterone is necessary for blastocyst implantation and subsequent decidualization in the rat. Although binding experiments have demonstrated the presence of estrogen-binding sites, no evidence exists as to whether the rat decidua expresses both isoforms of the estrogen receptor (ER), alpha and beta. In this investigation, we analyzed the expression of decidual ERalpha and ERbeta, studied their regulation by PRL and steroid hormones and examined the ability of decidual ERp to transduce the estradiol signal to the progesterone receptor. Immunocytochemistry, RT-PCR, and Northern blot analysis showed that both ER species are coexpressed in the decidua during pseudopregnancy. Interestingly, these genes were preferentially found in a cell population localized in the antimesometrial site of the uterus where blastocyst implantation takes place. Using decidual cells in primary culture obtained from pseudopregnant rats and a decidua-derived cell line (GG-AD), we show a differential regulation of ERalpha and ERbeta by PRL and ovarian steroid hormones. Whereas PRL, estradiol, and progesterone all increased ERbeta messenger RNA (mRNA) expression in a dose-dependent manner, only PRL up-regulated the mRNA levels of ERa. Estradiol had no effect on ERalpha expression, whereas progesterone markedly decreased its mRNA levels. Interestingly, progesterone, which up-regulates the ability of PRL to signal to a PRL-regulated gene in mammary-gland derived cells, prevented PRL stimulation of decidual ERalpha and had no synergistic effect on ERbeta expression. The use of GG-AD cells, which express only ERbeta, allowed us to demonstrate that this receptor subtype is functional and transduces estradiol signal to the progesterone receptor. In summary, the results of this investigation have revealed that ERbeta is expressed in addition to ERalpha in the rat decidua, and that the expression of both ERs are cell specific and differentially regulated by PRL and steroids. One salient finding of this investigation is that progesterone down-regulates ERalpha, but concomitantly increases the expression of a functional ERbeta that mediates estradiol up-regulation of the decidual progesterone receptor.     

Selective expression of the V beta 14 T cell receptor on Leishmania guyanensis--specific CD8+ T cells during human infection

Peripheral blood mononuclear cells from subjects never exposed to Leishmania were stimulated with Leishmania guyanensis. We demonstrated that L. guyanensis-stimulated CD8(+) T cells produced interferon (IFN)- gamma and preferentially expressed the V beta 14 T cell receptor (TCR) gene family. In addition, these cells expressed cutaneous lymphocyte antigen and CCR4 surface molecules, suggesting that they could migrate to the skin. Results obtained from the lesions of patients with localized cutaneous leishmaniaisis (LCL) showed that V beta 14 TCR expression was increased in most lesions (63.5%) and that expression of only a small number of V beta gene families (V beta 1, V beta 6, V beta 9, V beta 14, and V beta 24) was increased. The presence of V beta 14 T cells in tissue confirmed the migration of these cells to the lesion site. Thus, we propose the following sequence of events during infection with L. guyanensis. After initial exposure to L. guyanensis, CD8(+) T cells preferentially expressing the V beta 14 TCR and secreting IFN- gamma develop and circulate in the periphery. During the infection, these cells migrate to the skin at the site of the parasitic infection. The role of these V beta 14 CD8(+) T cells in resistance to infection remains to be determined conclusively.     

Selective expression of IL-7 receptor on memory T cells identifies early CD40L-dependent generation of distinct CD8+ memory T cell subsets

Several recent studies have demonstrated that T-helper cell-dependent events during the initial priming period are required for the generation of CD8(+) T cell-mediated protective immunity. The underlying mechanisms of this phenomenon have not yet been determined, mostly because of difficulties in studying memory T cells or their precursor populations at early stages during immune responses. We identified IL-7 receptor (CD127) surface expression as a marker for long-living memory T cells, most importantly allowing the distinction between memory and effector T cells early after in vivo priming. The combination of surface staining for CD127 and CD62L further separates between two functionally distinct memory cell subsets, which are similar (if not identical) to cell subsets recently described as central memory T cells (CD127(high) and CD62L(high)) and peripheral effector memory T cells (CD127(high) and CD62L(low)). Using this new tool of memory T cell analysis, we demonstrate that CD8(+) T cell priming in the absence of T cell help or CD40L specifically alters the generation of the effector memory T cell subset, which appears to be crucial for immediate memory responses and long-term maintenance of effective protective immunity. Our data reveal a unique strategy to obtain information about the quality of long-term protective immunity early during an immune response, a finding that may be applied in a variety of clinical settings, including the rapid monitoring of vaccination success.     

Essential requirement of an invariant V alpha 14 T cell antigen receptor expression in the development of natural killer T cells.

NK1.1+ T [natural killer (NK) T] cells express an invariant T cell antigen receptor alpha chain (TCR alpha) encoded by V alpha 14 and J alpha 281 segments in association with a limited number of V betas, predominantly V beta 8.2. Expression of the invariant V alpha 14/J alpha 281, but not V alpha 1, TCR in transgenic mice lacking endogenous TCR alpha expression blocks the development of conventional T alpha beta cells and leads to the preferential development of V alpha 14 NK T cells, suggesting a prerequisite role of invariant V alpha 14 TCR in NK T cell development. In V beta 8.2 but not B beta 3 transgenic mice, two NK T cells with different CD3 epsilon expressions, CD3 epsilon(dim) and CD3 epsilon(high), can be identified. CD3 epsilon(high) NK T cells express surface V alpha 14/V beta 8 TCR, indicating a mature cell type, whereas CD3 epsilon(dim) NK T cells express V beta 8 without V alpha 14 TCR and no significant CD3 epsilon expression (CD3 epsilon(dim)) on the cell surface. However, the latter are positive for recombination activating gene (RAG-1 and RAG-2) mRNA, which are only expressed in the precursor or immature T cell lineage, and also possess CD3 epsilon mRNA in their cytoplasm, suggesting that CD3 epsilon(dim) NK T cells are the precursor of V alpha 14 NK T cells.     

Oligoclonal expansion of T cell receptor V beta 2 and 3 cells in the livers of mice with graft-versus-host disease.

The nonsuppurrative destructive cholangitis lesions in the B10.D2 (donor) into BALB/c (host) mouse graft-versus-host disease (GVHD) model are dependent on CD4 T cells that use a T cell receptor-beta chain variable region (Vbeta) repertoire, which is heavily biased toward Vbeta2 and Vbeta3 usage. We hypothesized that liver Vbeta2(+) and Vbeta3(+) CD4 T cells originate from donor mice and recognize BALB/c minor histocompatibility alloantigens and BALB/c endogenous retroviral superantigen-6, respectively. To test this hypothesis, we determined the donor:host chimera status of infiltrating liver lymphocytes and the clonal states of liver Vbeta2(+) and liver Vbeta3(+) CD4 cells isolated from GVHD mice. A limited donor TCR Vbeta repertoire composed of Vbeta1(+), 2(+), 3(+), 4(+), 6(+), and 8(+) cells infiltrated the livers of GVHD mice on day 3. Consistent with a response to immunodominant host minor histocompatibility antigens, we detected oligoclonal liver Vbeta2(+) T cells in 40% of GVHD mice studied on day 3 and in 100% of GVHD mice studied on day 14. Typical of superantigen stimulation, extremely polyclonal liver Vbeta3(+) T cells were detected in 100% of GVHD mice studied on day 3 and 40% of GVHD mice studied on day 14. Yet, the liver Vbeta3(+) T cells in 60% of the day 14 GVHD mice were oligoclonal, pointing to a response to minor histocompatibility antigens.     

T cell receptor (alpha, beta, gamma) gene rearrangements and expression in normal and leukemic large granular lymphocytes/natural killer cells

The large granular lymphocyte (LGL) population, which effects a natural killer (NK) function, consists of cells whose lineage derivation has not been clearly established on the basis of phenotypic and functional properties. To clarify the relationship of LGL/NK cells to T cells we studied patterns of rearrangement and expression of the T cell receptor (Ti) genes alpha, beta, and gamma in normal human LGLs; in CD8+, CD8-, Mol+, and Mol- LGL subsets; and in 17 cases of leukemic LGL proliferations (T gamma LPD). T alpha, T beta, and T gamma genes were not expressed, nor were T beta and T gamma genes rearranged in normal LGLs or LGL subsets. The T gamma LPD were divided into two groups. One group (15/17 cases) was characterized as CD3+ and displayed Ti gene rearrangements. Seven of these cases were reactive with monoclonal antibody WT31, which suggested expression of an alpha/beta heterodimer on the cell surface. The other group (2/17 cases) was CD3- with unrearranged Ti genes. These results indicate that the normal LGL/NK population is homogeneous and distinct from the normal T cell population because it does not express, and as a result, cannot effect its immune function through the T cell receptor molecules. Conversely, T gamma LPDs represent a heterogeneous group of lymphoproliferative diseases within which the CD3-, Ti- cases most likely represent the neoplastic counterpart of normal LGL cells. The more frequent CD3+ cases may be related to recently described NK-like T cells. The observations that normal LGLs maintain germline T gamma genes and that many CD3+ T gamma LPD display an alpha/beta heterodimer suggest that a T gamma-containing receptor may not be necessary for NK or NK-like cytotoxicity.     

T cell receptor V beta expression in human intestine: regional variation in postnatal intestine and biased usage in fetal gut.

A panel of T cell receptor V beta specific monoclonal antibodies was used to analyse V beta gene usage at different sites in human postnatal and fetal intestine. In normal small intestine, at a single site, different patients showed expansion of T cells expressing individual V beta s. Lamina propria and epithelial T cells from the same patient showed overlapping but not identical V beta dominance. V beta dominance was also shown in the T cells of the colonic lamina propria. Analysis of two separate regions of intestine from the same patient (5-100 cm apart) showed that T cells expressing a dominant V beta region were often present at both sites. In most patients, however, major biases in T cell V beta usage (two to 12-fold variation) were also apparent between the two sites. Analysis of V beta expression in human fetal intestine also showed considerable skewing, although the most common dominant V beta in postnatal intestine (V beta 22) was never predominant in fetal intestine. Patchy local variation in the expression of individual V beta s therefore occurs against a background of V beta dominance over large regions of the human gut. Furthermore the results from fetal gut show that factors other than luminal antigen control V beta expression in the gut.     

Regulation of CCR6 chemokine receptor expression and responsiveness to macrophage inflammatory protein-3alpha/CCL20 in human B cells

The regulation of CCR6 (chemokine receptor 6) expression during B-cell ontogeny and antigen-driven B-cell differentiation was analyzed. None of the CD34(+)Lin(-) hematopoietic stem cell progenitors or the CD34(+)CD19(+) (pro-B) or the CD19(+)CD10(+) (pre-B/immature B cells) B-cell progenitors expressed CCR6. CCR6 is acquired when CD10 is lost and B-cell progeny matures, entering into the surface immunoglobulin D(+) (sIgD(+)) mature B-cell pool. CCR6 is expressed by all bone marrow-, umbilical cord blood-, and peripheral blood-derived naive and/or memory B cells but is absent from germinal center (GC) B cells of secondary lymphoid organs. CCR6 is down-regulated after B-cell antigen receptor triggering and remains absent during differentiation into immunoglobulin-secreting plasma cells, whereas it is reacquired at the stage of post-GC memory B cells. Thus, within the B-cell compartment, CCR6 expression is restricted to functionally mature cells capable of responding to antigen challenge. In transmigration chemotactic assays, macrophage inflammatory protein (MIP)-3alpha/CC chemokine ligand 20 (CCL20) induced vigorous migration of B cells with differential chemotactic preference toward sIgD(-) memory B cells. These data suggest that restricted patterns of CCR6 expression and MIP-3alpha/CCL20 responsiveness are integral parts of the process of B-lineage maturation and antigen-driven B-cell differentiation.