(1/894) Immunohistochemical localization of CD1a-positive putative dendritic cells in human breast tumours.
The presence of a high number of infiltrating CD1a+ cells in malignant neoplasms has been reported to be associated with an improved prognosis, reduced tumour recurrence and fewer metastases. This study identified a population of CD1a+ cells within the lymphoid cell infiltrate in human ductal breast carcinoma (n = 52), which was significantly different from normal breast tissue, in which only two out of nine cases expressed CD1a+ cells (P = 0.0192). In the majority of cases, the infiltrate was low compared with the number of macrophages and T cells present (results not shown). There was no correlation between the number of CD1a+ cells and tumour grade, with all tumour grades expressing similar numbers of infiltrating CD1a+ cells. There was clear evidence, however, that the CD1a+ cells were closely associated with tumour cells. It is likely that CD1a+ cells have a role in antigen capture and presentation in human tumours, and this study documents the density of CD1a+ cells in a large sample of all histological grades of human breast carcinomas. (+info)
(2/894) Long-term culture of human CD34(+) progenitors with FLT3-ligand, thrombopoietin, and stem cell factor induces extensive amplification of a CD34(-)CD14(-) and a CD34(-)CD14(+) dendritic cell precursor.
Current in vitro culture systems allow the generation of human dendritic cells (DCs), but the output of mature cells remains modest. This contrasts with the extensive amplification of hematopoietic progenitors achieved when culturing CD34(+) cells with FLT3-ligand and thrombopoietin. To test whether such cultures contained DC precursors, CD34(+) cord blood cells were incubated with the above cytokines, inducing on the mean a 250-fold and a 16,600-fold increase in total cell number after 4 and 8 weeks, respectively. The addition of stem cell factor induced a further fivefold increase in proliferation. The majority of the cells produced were CD34(-)CD1a- CD14(+) (p14(+)) and CD34(-)CD1a-CD14(-) (p14(-)) and did not display the morphology, surface markers, or allostimulatory capacity of DC. When cultured with granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4), both subsets differentiated without further proliferation into immature (CD1a+, CD14(-), CD83(-)) macropinocytic DC. Mature (CD1a+, CD14(-), CD83(+)) DCs with high allostimulatory activity were generated if such cultures were supplemented with tumor necrosis factor-alpha (TNF). In addition, p14(-) cells generated CD14(+) cells with GM-CSF and TNF, which in turn, differentiated into DC when exposed to GM-CSF and IL-4. Similar results were obtained with frozen DC precursors and also when using pooled human serum AB+ instead of bovine serum, emphasizing that this system using CD34(+) cells may improve future prospects for immunotherapy. (+info)
(3/894) Biochemical characterization of CD1d expression in the absence of beta2-microglobulin.
CD1d is a major histocompatibility complex class I-like molecule that exhibits a distinct antigen processing pathway that functions in the presentation of hydrophobic antigens to T cells. CD1d has been previously shown to be expressed on the cell surface of human intestinal epithelial cell lines in vivo and a transfected cell line in vitro independently of beta2-microglobulin (beta2m). To define the relationship between CD1d and beta2m and characterize the biochemical structure of CD1d in the absence of beta2m, we have used a newly generated series of CD1d transfectants and CD1d-specific antibodies. These studies show that in the absence of beta2m, CD1d is expressed on the cell surface as a 45-kDa glycoprotein that is sensitive to endoglycosidase-H and is reduced to 37-kDa after N-glycanase digestion. In contrast, in the presence of beta2m, CD1d is expressed on the cell surface as a 48-kDa endoglycosidase-H-resistant glycoprotein. Pulse-chase metabolic labeling studies demonstrate that acquisition of endoglycosidase-H resistance of CD1d is observed in the presence of beta2m but not in the absence of beta2m even after a 24-h chase period. Thus, CD1d is able to be transported to the cell surface independently of beta2m; however, in the absence of beta2m, the glycosylation pattern of CD1d is altered and consistent with an immature glycoprotein. (+info)
(4/894) T cell-tropic simian immunodeficiency virus (SIV) and simian-human immunodeficiency viruses are readily transmitted by vaginal inoculation of rhesus macaques, and Langerhans' cells of the female genital tract are infected with SIV.
Intravaginal inoculation with T cell-tropic molecular clones of simian immunodeficiency virus (SIV) or simian-human immunodeficiency virus (SHIV) or some dual-tropic strains of SIV or SHIV produced systemic infection in rhesus macaques. Vaginal inoculation with other dual-tropic molecular clones of SIV or SHIV did not infect rhesus macaques even after multiple inoculations. While in vitro measures of macrophage tropism do not predict which primate lentiviruses will produce systemic infection after intravaginal inoculation, the level to which a virus replicates in vivo after intravenous inoculation does predict the outcome of intravaginal inoculation. Another series of studies, using combined in situ hybridization and immunolabeling to simultaneously detect SIV RNA and identify the immunophenotype of infected cells, demonstrated that a large proportion (approximately 40% in some animals) of the SIV-infected cells in the vagina and cervix were Langerhans' cells. This is the first in vivo demonstration that Langerhans' cells in the genital tract are infected with SIV and that dendritic cells are significant reservoirs for lentiviruses. (+info)
(5/894) Expression of the nlsLacz gene in dendritic cells derived from retrovirally transduced peripheral blood CD34+ cells.
BACKGROUND AND OBJECTIVE: Gene transfer and expression of exogenous genetic information coding for an immunogenic protein in antigen presenting cells (APCs) can promote an immune response. This was investigated by retroviral transfer of a marker gene into CD34+ derived APCs. DESIGN AND METHODS: To achieve long term expression of a specific transgene in APCs, G-CSF mobilized peripheral blood CD34+ cell populations were retrovirally transduced with the bacterial nlsLacZ, a marker gene used here as a model, in the presence of IL-3, IL-6, GM-CSF and SCF prior to being induced to differentiate into dendritic and macrophage cells by GM-CSF and TNF-a. RESULTS: Addition of IL-4 was found to induce dendritic differentiation preferentially by inhibiting proliferation and differentiation of the macrophage lineage. As assessed by X-Gal staining, LacZ gene expression was observed in cells from both the dendritic lineage (CD1a+/CD14-) which still exhibits the highest immunostimulatory activity in mixed lymphocyte reaction and from the macrophage lineage (CD1a-/ CD14+). INTERPRETATION AND CONCLUSIONS: This study sets out the possibility of transducing dendritic and macrophage progenitors present in the CD34+ cell population and in using a marker gene such as nlsLacZ to study gene expression in antigen presenting cell compartments. (+info)
(6/894) Expression of CD1d2 on thymocytes is not sufficient for the development of NK T cells in CD1d1-deficient mice.
CD1 is an MHC class I-like molecule that has been conserved throughout mammalian evolution. Unlike MHC class I molecules, CD1 can present unique nonprotein antigens to T cells. The murine CD1 locus contains two highly homologous genes, CD1d1 and CD1d2. CD1d1 is essential for the development of a major subset of NK T cells that promptly secrete IL-4 following activation. However, the function of CD1d2 has not yet been demonstrated. In the present study, we examined the expression of CD1d2 in CD1d1-deficient (CD1d1 degrees) mice with the anti-CD1 Ab 3H3. Unlike CD1d1, which is expressed by all lymphocytes, CD1d2 can be detected only on the surface of thymocytes. To determine whether CD1d2 can select a unique subset of NK T cells, we compared the remnant population of NK T cells in CD1d1 degrees and CD1d1, CD1d2-double deficient (CD1d1 degrees CD1d2 degrees) mice. No significant difference in the number of NK T cells and cytokine secretion capacity can be detected between CD1d1 degrees and CD1d1 degrees CD1d2 degrees mice, indicating that CD1d2 cannot substitute for CD1d1 in NK T cell development. The inability of CD1d2 to select NK T cells is not due to the structural constraints of CD1d2 since CD1d2-transfected cells can be recognized by both NK T cell hybridomas and freshly isolated NK T cells. Given the structural similarities, it is possible that the low levels of surface expression and limited tissue distribution of CD1d2 may prevent it from functioning in the selection and expansion of NK T cells. (+info)
(7/894) Juvenile hemochromatosis locus maps to chromosome 1q.
Juvenile hemochromatosis (JH) is an autosomal recessive disorder that leads to severe iron loading in the 2d to 3d decade of life. Affected members in families with JH do not show linkage to chromosome 6p and do not have mutations in the HFE gene that lead to the common hereditary hemochromatosis. In this study we performed a genomewide search to map the JH locus in nine families: six consanguineous and three with multiple affected patients. This strategy allowed us to identify the JH locus on the long arm of chromosome 1. A maximum LOD score of 5.75 at a recombination fraction of 0 was detected with marker D1S498, and a LOD score of 5. 16 at a recombination fraction of 0 was detected for marker D1S2344. Homozygosity mapping in consanguineous families defined the limits of the candidate region in an approximately 4-cM interval between markers D1S442 and D1S2347. Analysis of genes mapped in this interval excluded obvious candidates. The JH locus does not correspond to the chromosomal localization of any known gene involved in iron metabolism. These findings provide a means to recognize, at an early age, patients in affected families. They also provide a starting point for the identification of the affected gene by positional cloning. (+info)
(8/894) Immunolocalization of CD1d in human intestinal epithelial cells and identification of a beta2-microglobulin-associated form.
In order to better understand the role of intestinal CD1d, we sought to define the cellular localization and further characterize the biochemical structure of CD1d in human intestinal epithelial cells (IEC). Using a CD1d-specific rabbit anti-gst-CD1d antibody, immunoprecipitation of radiolabeled cell surface proteins detected a previously identified 37 kDa protein as well as a 48-50 kDa protein which were confirmed by Western blotting with a CD1d-specific mAb, D5. Immunoprecipitation of protein lysates with the CD1d-specific mAb, D5 and 51.1.3, and the beta2-microglobulin (beta2m)-specific mAb, BBM.1, followed by N-glycanase digestion and Western blotting with the D5 mAb showed that the 48-50 kDa protein was a beta2m-associated, CD1d glycoprotein. CD1d was immunolocalized to the apical and lateral regions of native small and large intestinal IEC as defined by confocal laser microscopy using the D5 mAb and the rabbit anti-gst-CD1d antibody. In addition, a large apical intracellular pool of CD1d was identified. Identical observations were made with polarized T84 cells. Selective biotin labeling of apical and basolateral cell surfaces followed by immunoprecipitation with the D5 mAb, N-glycanase digestion and avidin blotting confirmed the presence of glycosylated CD1d on both cell surfaces and immunolocalization of the 37 kDa non-glycosylated form of CD1d to the apical cell surface. These studies show that CD1d is located in an ideal position for luminal antigen sampling and presentation to subjacent intraepithelial lymphocytes. (+info)