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L Lefrancois

Publications and source records attributed to L Lefrancois.

At least 19 recordsLinked to original sources
PubMed2004

Ultrasound detection of spontaneous hepato-cellular carcinomas in X/myc bitransgenic mice.

AIMS: To evaluate trans-abdominal ultrasound for the detection of Hepatocellular carcinoma (HCC) in a bitrasgenic murine (X/myc) model using a commercially available high-frequency ultrasound unit. METHODS: Sixty-one female animals were included in this study. These animals were submitted to a single ultrasound examination of the liver under general anesthesia (isoflurane), and then euthanized. Results of ultrasound were compared with necropsy and histopathology. RESULTS: The lesions demonstrated a fairly consistent aspect (oval- or round-shaped, well-defined hypoechoic homogeneous lesions), and lesions as small as 2 mm were identified. For detection of hepatic nodules per mouse the sensitivity was 75%, the specificity was 100% and the accuracy was 88.5%. For detection of hepatic focal lesions per lesions the overall sensitivity was 60%, the specificity was 97%, and the accuracy was 75.9%. Contrast-enhanced harmonic ultrasound imaging did not improve the identification of the lesions in our experimental conditions. CONCLUSION: High-frequency ultrasound appears to be an efficient tool allowing new possibilities to use this animal model and evaluate new therapies in longitudinal studies, which are much more powerful.

Animals↗
PubMed1999

Expression of the transcription factor lung Krüppel-like factor is regulated by cytokines and correlates with survival of memory T cells in vitro and in vivo.

The transcription factor lung Krüppel-like factor (LKLF) is involved in naive T cell survival. Expression of LKLF is rapidly down-regulated upon T cell stimulation, raising the question of whether LKLF is reexpressed after activation, and what factors are required for such reexpression. Furthermore, the expression of LKLF in resting memory cells has not been determined. Here, we use the OT-I TCR transgenic mouse system to address these issues. LKLF was found to be reexpressed following culture of activated CD8 T cells in certain cytokines (IL-2, IL-7) but not others (IL-12) known to influence CTL development. Interestingly, induction of LKLF reexpression corresponded with long-term T cell survival and development of memory T cell phenotype. Furthermore, using OT-I cells stimulated in vivo, we demonstrated that Ag induced rapid LKLF down-regulation and that the factor is expressed by in vivo-derived memory T cells.

Adoptive Transfer↗
PubMed1999

Mucosal T lymphocyte numbers are selectively reduced in integrin alpha E (CD103)-deficient mice.

The mucosal lymphocyte integrin alpha E(CD103)beta 7 is thought to be important for intraepithelial lymphocyte (IEL) localization or function. We cloned the murine integrin gene encoding alpha E, localized it to chromosome 11, and generated integrin alpha E-deficient mice. In alpha E-/- mice, intestinal and vaginal IEL numbers were reduced, consistent with the known binding of alpha E beta 7 to E-cadherin expressed on epithelial cells. However, it was surprising that lamina propria T lymphocyte numbers were diminished, as E-cadherin is not expressed in the lamina propria. In contrast, peribronchial, intrapulmonary, Peyer's patch, and splenic T lymphocyte numbers were not reduced in alpha E-deficient mice. Thus, alpha E beta 7 was important for generating or maintaining the gut and vaginal T lymphocytes located diffusely within the epithelium or lamina propria but not for generating the gut-associated organized lymphoid tissues. Finally, the impact of alpha E deficiency upon intestinal IEL numbers was greater at 3-4 wk of life than in younger animals, and affected the TCR alpha beta+ CD8+ T cells more than the gamma delta T cells or the TCR alpha beta+ CD4+CD8- population. These findings suggest that alpha E beta 7 is involved in the expansion/recruitment of TCR alpha beta+ CD8+ IEL following microbial colonization. Integrin alpha E-deficient mice will provide an important tool for studying the role of alpha E beta 7 and of alpha E beta 7-expressing mucosal T lymphocytes in vivo.

Animals↗
PubMed1998

Differential requirements for ZAP-70 in TCR signaling and T cell development.

The Syk/ZAP-70 family of protein tyrosine kinases is indispensable for normal lymphoid development. Syk is necessary for the development of B cells and epithelial gammadelta T cells, whereas ZAP-70 is essential for the normal development of T cells and TCR signaling. In this study, we show that although development of the alphabeta lineage was arrested in the thymus, CD3-positive T cells, primarily of the gammadelta lineage, were present in the lymph nodes of mice lacking ZAP-70. Moreover, in the absence of ZAP-70, dendritic epidermal T cells were fewer in number and of abnormal morphology, and intestinal intraepithelial lymphocytes, normally containing a large proportion of gammadelta T cells, were markedly reduced. These data suggest that gammadelta T cells show a variable dependence upon ZAP-70 for their development. Biochemical analyses of thymocytes revealed a lack of basal zeta-chain tyrosine phosphorylation. However, several other substrates were inducibly tyrosine phosphorylated following TCR stimulation. Thus, TCR-mediated signaling in ZAP-70-deficient thymocytes is only partially impaired. These studies suggest that Syk compensates only partially for the loss of ZAP-70, and that there is an absolute requirement of ZAP-70 for alphabeta T cells and epithelial gammadelta T cells, but not for some gammadelta T cells in peripheral lymphoid tissues.

Animals↗
PubMed1994

Gamma delta T cells contribute to immunity against the liver stages of malaria in alpha beta T-cell-deficient mice.

The functional role of gamma delta T cells (expressing the gamma delta heterodimeric T-cell receptor for antigen) in infectious diseases remains largely unknown. We have therefore attempted to define the possible role of these T cells in the immune response against the various developmental stages of malaria parasites. For this purpose, we monitored the immune response and the development of liver and blood stages of Plasmodium yoelii, a rodent malaria parasite, in immunized and nonimmunized alpha beta T-cell-deficient and gamma delta T-cell-deficient mice. Immunization of alpha beta T-cell-deficient mice with irradiated sporozoites induced an immune response that significantly inhibited the development of the parasite's liver stages. This inhibitory immune response was abolished by an antibody-mediated transient in vivo depletion of gamma delta T cells. Two gamma delta T-cell clones were derived from malaria-immunized alpha beta T-cell-deficient mice. The adoptive transfer of one of these gamma delta T-cell clones to normal mice inhibited the development of liver stages, following sporozoite inoculation. These results provide evidence for gamma delta T-cell-mediated protective immunity against parasites, in the absence of alpha beta T cells. As for the blood phase of the infection, both normal mice and gamma delta T-cell-deficient mice cleared the blood stages of the nonlethal strain of P. yoelii, while alpha beta T-cell-deficient mice failed to control the parasitemia.

Animals↗
PubMed1993

Nonimmunogenic tumor cells may efficiently restimulate tumor antigen-specific cytotoxic T cells.

Induction of immunity to a viral protein that had been transfected into a tumor cell line was studied. The nucleoprotein (NP) of vesicular stomatitis virus (VSV) was used as a model tumor-associated Ag after transfection into EL-4, and H-2b thymoma originating from C57BL/6 mice. The NP-transfected cell line (EL-4NP) was lysed by NP-specific CTL and was found to restimulate NP-specific CTL in vitro as efficiently as did VSV-infected macrophages. Despite both of these in vitro characteristics, C57BL/6 mice inoculated with EL-4NP did not mount a measurable NP-specific CTL response and developed a lethal tumor as rapidly as did mice given control EL-4. This lack of immunogenicity could not be explained by down-regulation of MHC class I molecules or by loss of NP; even EL-4NP cells metastasizing to the spleen kept their high restimulatory capacity and excellent target characteristics. However, once mice were immunized with VSV or with a vaccinia-VSV-NP recombinant virus they were protected against tumor growth of EL-4NP by CD8+ CTL but not by CD4+ T cells. Taken together, the failure of the tumor-associated Ag to induce a protective T cell response in vivo despite its excellent capacity to restimulate CTL in vitro may encourage adjuvant immunotherapy in cancer; even the effects of weakly immunizing tumor vaccines, e.g., recombinant viruses, may be efficiently amplified by tumor cells.

Animals↗
PubMed1993

Emigration of selected subsets of gamma delta + T cells from the adult murine thymus.

Cells bearing the gamma delta form of the TCR make up only 1-3% of T cells in the adult murine thymus and peripheral lymphoid organs. Evidence from studies of nude mice suggests that the development of at least some gamma delta T cells is thymus dependent; however, until now it has not been directly demonstrated that gamma delta cells are exported from the thymus. In this paper we have used the technique of labelling thymocytes in vivo with FITC, followed by flow cytometric analysis to trace cells emigrating from the thymus to the spleen. Using this approach we have been able to demonstrate for the first time that gamma delta T cells are exported from the adult murine thymus to the spleen. We also demonstrate that the cells emigrating to the spleen are a selected subset of gamma delta thymocytes being heat stable antigen positive, Thy-1+, and expressing low levels of CD44 (Pgp-1). In addition, investigation of TCR V gamma gene usage among adult gamma delta + thymocytes, recent emigrants, and spleen cells, indicated a selective emigration of gamma delta cells expressing certain V gamma genes.

Animals↗
PubMed1992

Mechanism of self-tolerance of gamma/delta T cells in epithelial tissue.

The present study examined mechanisms of tolerance for T cell receptor gamma/delta (TCR-gamma/delta) cells. Using a transgenic (Tg) model, we demonstrate that although alloantigen (Ag)-specific TCR-gamma/delta cells are deleted in the thymus and spleen of Ag-bearing mice, intraepithelial lymphocytes (IELs) expressing normal levels of the Tg TCR were present. However, Tg+ IELs from Ag-bearing mice were unresponsive to activation. Furthermore, self-reactive Tg+ IELs decreased in number over time. Thus, in epithelial tissue, Tg TCR-gamma/delta cells are eliminated subsequent to and most likely as a result of the induction of clonal anergy.

Animals↗
PubMed1992

A program to facilitate cytotoxicity assay data reduction.

A report of a program which performs the initial computational reduction of the raw data from a cytotoxicity assay and outputs the reduced data as an image of the arrangement of the assay(s) upon the microwell plate. The program accepts the raw data either as manual or diskette file input.

Algorithms↗
PubMed1991

Phenotypic complexity of intraepithelial lymphocytes of the small intestine.

A detailed phenotypic analysis of intraepithelial lymphocytes (IEL) of the small intestine was performed using multicolor fluorescence flow cytometry. CD4+8+ IEL (double positives; DP) could be detected in significant numbers in preparations from several mouse strains. DP IEL expressed Tcr alpha, beta and Thy1. Comparison of Tcr alpha, beta levels of thymocytes and IEL revealed that whereas the majority of DP thymocytes expressed low Tcr levels, DP IEL expressed high, mature T cell levels of Tcr. In addition, DP IEL were generally Ly3- (CD8 beta), unlike their thymic counterparts, which are Ly3+. Ly3 was not present on Tcr gamma, delta IEL, whereas CD4-8+ Tcr alpha, beta IEL contained Ly3- and Ly3+ subsets. The Ly3- population in either Tcr-bearing subset could be further subdivided by Thy1 expression. Ly1 (CD5) expression was also examined, and none of the Tcr gamma, delta IEL were Ly1+. Based on Thy1, Ly3, and Ly1 expression, four CD4-8+ Tcr alpha, beta IEL subsets were detected. The results indicate the cellular complexity of the IEL compartment rivals that found in the thymus. These findings are discussed in light of recent data suggesting an extrathymic origin of some IEL.

Animals↗
PubMed1991

Developmental expression of CD45 alternate exons in murine T cells. Evidence of additional alternate exon use.

The CD45 glycoprotein family exhibits cell line-age-associated structural heterogeneity arising in part from alternate 5' exon shuffling. Previous studies of exons involved in the final glycoprotein structure have provided evidence of alternate exon use for only three exons (Ex-4, 5 and 6). However, our prior data using reverse transcription-polymerase chain reaction (RT-PCR) suggested the presence of at least one additional CD45 alternate exon. By using RT-PCR, Southern blotting with exon-specific or exon splice junction-specific oligonucleotide probes and direct DNA sequencing of RT-PCR products, we demonstrated additional alternate use involving Ex-7. PCR analysis of stage I thymocytes (CD4-CD8-) revealed only faintly detectable bands for two isoforms: one lacking Ex-4, 5, 6 and 7 (a "minus-one" [Ex(-1)] isoform), and a smaller isoform preliminarily characterized as also lacking Ex-8. Stage II thymocytes (CD4+CD8+) prominently expressed both Ex(-1) and zero alternate exon (Ex(0] isoforms, with one exon (Ex(1] and two exon (Ex(2] isoforms also present. Among stage III thymocytes, both CD4+CD8- and CD4-CD8+ cells expressed only Ex(-1) and Ex(0) isoforms. CD45 alternate exon use in resting CD4+ and CD8+ lymph node T cells was divergent, with CD8+ cells additionally expressing an Ex(2) isoform. Among alloreactive T cell clones, band intensity for the Ex(1) isoform in CD4+ BC-3 cells was much less than for resting CD4+ T cells, while the CD8+ CTL clone 8.2.2 exhibited production of the higher alternate exon isoforms, Ex(2) and Ex(3). We conclude that at least four and possibly five alternate exons exist in the CD45 glycoprotein family, with a previously unrecognized isoform lacking Ex-4, 5, 6 and 7 prominently expressed in T cells. Shuffling of CD45 alternate exons appears to occur in an organized and predictable sequence during cellular maturation and activation.

Animals↗
PubMed1990

Characterization of the CD45 molecule on murine intestinal intraepithelial lymphocytes.

The CD45 molecule was analyzed from murine intestinal intraepithelial lymphocytes (IEL). Immunofluorescent staining of CD8+ IEL revealed varying degrees of reactivity with mAb specific for CD45-restricted determinants, some which are typically expressed only by B cells. Immunoprecipitation of CD45 molecules from IEL yielded an array of proteins with apparent (m.w.) ranging from 180,000 to 260,000. The m.w. 260,000 form was restricted to IEL, was distinct from the B220 molecule, and was the only CD45 isoform that expressed the CD45-associated carbohydrate differentiation Ag CT1. Moreover, the CT1 determinant was present on cells of the Thy-1- but not the Thy-1+ IEL subset. Sequential immunoprecipitation studies indicated that expression of the m.w. 260,000 protein was not restricted to CT1+ cells. The protein composition of the m.w. 260,000 CD45 isoform was examined by using the polymerase chain reaction for analysis of CD45 variable exon usage. In contrast to B cells in which the major CD45 mRNA contained all three variable exons (exons 4, 5, and 6), IEL CD45 mRNA contained significant amounts of two-exon, single exon, and zero variable exon forms. Restriction enzyme analysis identified the single exon form as exon 5 and the two-exon form as a mixture of exons 4 and 5 and exons 5 and 6. Metabolic labeling of CD45 in pulse-chase experiments suggested that the generation of this high m.w. protein was caused by post-translational modifications, perhaps glycosylation. Overall, the results indicated that the high m.w. form of CD45 and the addition of the CT1 determinant were generated via IEL-specific post-translational modifications and not by novel alternate exon usage.

Animals↗
PubMed1990

Extrathymic selection of TCR gamma delta + T cells by class II major histocompatibility complex molecules.

The influence of MHC antigens on TCR gamma delta usage in CD8+ intraepithelial lymphocytes (IELs) was examined using a pan-reactive and V delta 4 region-specific MAb. While an average of 30% of IELs from the majority of mice of various MHC haplotypes were V delta 4+, a 2-fold or greater percentage of IELs from H-2k mice were V delta 4+. Analysis of IELs from F1 mice indicated that the increase in TCRs using V delta 4 was likely to be the result of positive selection. The V delta 4 usage patterns of IELs from recombinant inbred strains and from mice recombinant within H-2 revealed that the increase in V delta 4 usage mapped to H-2 and required I-E expression. Moreover, selection of TCRs using V delta 4 occurred in chimeric mice in the absence of a thymus. The results demonstrate an extrathymic selective mechanism for gamma delta TCRs of CD8+ IELs and suggest that these cells may exhibit MHC class II-restricted antigen recognition.

Animals↗
PubMed1990

Expression of gamma/delta T cell receptors on lymphocytes from the lactating mammary gland.

gamma/delta cells were at least four times more frequent in lactating mouse mammary glands than among T cells of the most proximal lymph nodes. Two-color staining of freshly isolated T cells and a study of clonally expressed gamma/delta receptors on hybridomas further revealed that the mammary gamma/delta population is heterogeneous, including at least three different subsets, among them cells expressing V gamma 5, V gamma 4 together with V delta 4, or none of these V regions.

Animals↗
PubMed1989

Intraepithelial lymphocytes. Anatomical site, not T cell receptor form, dictates phenotype and function.

The function and structure of the TCR proteins of intraepithelial lymphocytes (IEL) were examined using a panel of mAbs specific for TCR-gamma/delta. Three subsets of TCR-gamma/delta+ IEL could be detected with five mAbs, termed GL1-GL5. The mAbs were able to trigger lysis via crosslinking of the IEL TCR and all of the subsets were constitutively cytolytic. Immunoprecipitation of IEL TCR proteins revealed that the GL2 mAb reacted only with gamma, delta heterodimers containing high Mr delta chains, while the other mAbs precipitated all of the observed gamma and delta proteins. Two-color fluorescence analysis showed that the GL2+ subset was contained within the larger GL1+ subset. The GL3 and GL4 mAbs appear to be specific for all TCR-gamma/delta while GL2 was V delta 4 specific. Analysis of IEL for TCR-alpha/beta expression demonstrated that approximately 20% of B6 IEL were TCR-alpha/beta+. Interestingly, this population of IEL contained Thy-1- and CT1+ cells, indicating that the unique phenotype of IEL was not restricted to TCR-gamma/delta+ cells. Moreover, the TCR-alpha/beta+ IEL were also constitutively cytolytic, suggesting that the intestinal milieu was controlling the functional programming of IEL regardless of TCR type. The mAbs reported here as well as the ability to exploit the distinct phenotype of IEL should prove useful in determining the function of IEL and the TCR-gamma/delta.

Animals↗