Biomedical subjects

S Lohwasser

Publications and source records attributed to S Lohwasser.

10 recordsLinked to original sources

PubMed2001

Regulation of NKT cells by Ly49: analysis of primary NKT cells and generation of NKT cell line.

TCRalphabeta(+)NK1.1(+) (NKT) cells are known to express various NK cell-associated molecules including the Ly49 family of receptors for MHC class I, but its functional significance has been unclear. Here, we examined the expression of Ly49A, C/I and G2 on various NKT cell populations from normal and MHC class I-deficient C57BL/6 mice as well as their responsiveness to alpha-galactosylceramide (alpha-GalCer), a potent stimulator of CD1d-restricted NKT cells. The frequency and the level of Ly49 expression varied among NKT cells from different tissues, and were regulated by the expression of MHC class I and CD1d in the host. Stimulation of various NKT cells with alpha-GalCer suggested that Ly49 expression inversely correlates with the responsiveness of NKT cells to alpha-GalCer. Moreover, alpha-GalCer presented by normal dendritic cells stimulated purified Ly49(-), but not Ly49(+), splenic NKT cells, whereas MHC class I-deficient dendritic cells presented alpha-GalCer to both Ly49(+) and Ly49(-) NKT cells equally well. Therefore, MHC class I on APCs seems to inhibit activation of NKT cells expressing Ly49. To further characterize CD1d-restricted NKT cells, we generated an alpha-GalCer-responsive NKT cell line from thymocytes. The line could only be generated from Ly49(-)NK1.1(+)CD4(+) thymocytes but not from other NKT cell subsets, and it lost expression of NK1.1 and CD4 during culture. Together, these results indicate the functional significance of Ly49 expression on NKT cells.

Animals↗

PubMed2001

Ly49 and CD94/NKG2: developmentally regulated expression and evolution.

Murine natural killer (NK) cells express two families of MHC class I-specific receptors, namely the Ly49 family and CD94/NKG2 heterodimers. Stochastic co-expression of these receptors generates diverse receptor repertoires in adult NK-cell populations, whereas fetal NK cells have much more limited receptor diversity as they mostly express CD94/NKG2A but not Ly49. These receptors are also expressed on CD8-T cells and NK1.1+ T cells and regulate their functions, but their expression pattern on NK cells is significantly different from those on T cells. Thus, expression of Ly49 and CD94/NKG2 is developmentally regulated. NK cells acquire the Ly49 family of receptors in an orderly manner as they differentiate from bone marrow progenitors in vitro. Similarly, acquisition of CD94 and NKG2 by NK cells as they differentiate from embryonic stem cells is also orderly To gain insight into the mechanisms regulating Ly49 expression, potential regulatory regions of several Ly49 genes have been examined. Ly49 genes with different expression patterns have remarkably similar sequences in the putative regulatory regions. Finally, a functional Ly49 gene has been identified in baboon, and primate comparisons suggest that functional extinction of the Ly49 gene in the human lineage seems to have been a relatively recent event.

Amino Acid Sequence↗

PubMed2001

The non-classical MHC class I molecule Qa-1(b) inhibits classical MHC class I-restricted cytotoxicity of cytotoxic T lymphocytes.

The CD94/NKG2A heterodimer is an inhibitory receptor expressed on a subset of mouse NK cells. CD94/NKG2A recognizes the non-classical MHC class I (class Ib) molecule Qa-1(b) and inhibits NK cytotoxicity. Qa-1(b) presents a peptide derived from the leader sequence of classical MHC class I molecules. Here, we examined the role of CD94/NKG2A in T cell-mediated cytotoxicity. Soluble tetrameric Qa-1(b) bound to almost all CD8(+), but not CD4(+), T cells. This binding seems to be mediated by CD8, because COS cells transfected with CD8 also bound Qa-1(b) tetramer. Therefore, the expression of CD94/NKG2 in T cells was further examined by single-cell RT-PCR. Most murine CD8(+) T cells constitutively expressed CD94 and NKG2A transcripts, whereas they were not detected in CD4(+) T cells. Co-expression of Qa-1(b) and D(k) on target cells significantly inhibited cytotoxicity of D(k)-specific cytotoxic T lymphocytes generated by mixed lymphocyte reaction, indicating that Qa-1(b) on antigen-presenting cells interacts with CD94/NKG2A on CD8 T cells and regulates classical MHC class I-restricted cytotoxic T cells. These results suggest a significant role of CD94/NKG2A as an inhibitory receptor on CD8(+) T cells.

Animals↗

PubMed2000

The genomic organization of the mouse CD94 C-type lectin gene.

The mouse natural killer (NK) gene complex is located on chromosome 6 and contains a number of genes encoding C-type lectin receptors which have been found to regulate NK cell function. Among these are CD94 and four NKG2 genes. Like its human counterpart, the mouse CD94 protein associates with different NKG2 isoforms and recognizes the atypical MHC class I molecule Qa-1b. Here, the genomic organization of the mouse CD94 gene was determined by analysing a BAC clone containing the CD94 gene. The mouse CD94 gene contains six exons separated by five introns. Exons I and II encode the 5' untranslated region (UTR) and the transmembrane domain. Exon III encodes the stalk region and exons IV-VI encode the carbohydrate recognition domain (CRD). Furthermore, we cloned and sequenced the CD94 promoter region, and putative regulatory DNA elements were identified. Further studies on the CD94 promoter region may help to elucidate the restricted expression pattern of CD94 in NK cells and a subpopulation of T cells.

Alternative Splicing↗

PubMed1999

IFN-gamma production and cytotoxicity of IL-2-activated murine NK cells are differentially regulated by MHC class I molecules.

Activation of NK cells by target cells leads to cytotoxicity as well as production of various cytokines including IFN-gamma. MHC class I molecules on target cells regulate NK cytotoxicity. However, little is known about the regulation of IFN-gamma production by NK cells. We examined the production of IFN-gamma in individual murine NK cells stimulated with tumor cell lines by flow cytometric analysis of intracellular IFN-gamma. Among several tumor lines tested, the rat basophilic leukemia line RBL-1 induced particularly high level of IFN-gamma production in IL-2-activated NK cells, whereas other lines, including the prototypic NK target YAC-1, induced very low or no IFN-gamma production. Transfection of murine classical MHC class I molecules into RBL-1 cells substantially inhibited IFN-gamma production. This inhibition of IFN-gamma production by MHC class I was independent of Ly-49 or CD94/NKG2A expression on NK cells. These results indicate that some target cells directly stimulate IL-2-activated NK cells and induce IFN-gamma production, but the requirements for the induction of IFN-gamma production seem different from those for NK cytotoxicity. Furthermore, similar to NK cytotoxicity, induction of IFN-gamma production is inhibited by MHC class I on stimulating cells. However, the MHC class I-specific receptors inhibiting IFN-gamma production are different from those for NK cytotoxicity.

Animals↗

PubMed1999

Diversity of NK cell receptor repertoire in adult and neonatal mice.

Murine NK cytotoxicity is regulated by two families of MHC class I-specific receptors, namely Ly49 and CD94/NKG2. We developed a single-cell RT-PCR method to analyze expression of all known Ly49 and NKG2A genes in individual NK cells and determined the receptor repertoires of NK cells from adult and neonatal (1-wk-old) C57BL/6 mice. In adult mouse NK cells, up to six different receptors were coexpressed in random combinations. Of 62 NK cells examined, 42 different patterns of receptor expression were observed. Most of them expressed at least one Ly49, whereas NKG2A was detected in 32% of the cells. Over 75% of them expressed Ly49C, I, or NKG2A, which are thought to recognize self-class I MHC (H-2b). Coexpression of multiple Ly49 receptors and NKG2A was stochastic. In contrast, very few neonatal NK cells expressed any Ly49, but almost 60% of them expressed NKG2A. These results demonstrate that adult NK cells are quite heterogeneous and have diverse receptor repertoires. They also suggest that the expression of NKG2A precedes Ly49 expression in NK cell ontogeny, and NKG2A is a major inhibitory receptor in neonatal NK cells.

Aging↗

PubMed1999

Cloning of murine NKG2A, B and C: second family of C-type lectin receptors on murine NK cells.

Multiple NK cell receptors for MHC class I have been identified. They include killer inhibitory receptors and CD94/NKG2 heterodimers in humans and the Ly49 family in mice. Here we report the cloning of murine NKG2A, B and C. The deduced amino acid sequence of mouse NKG2A contains only one consensus cytoplasmic immunoreceptor tyrosine-based inhibitory motif (ITIM). NKG2A from B6 and BALB/c mice differ by six amino acid residues in the extracellular domain. Murine NKG2B, like its human conterpart, appears to be a splice variant of NKG2A and lacks a large portion of the stalk region. Murine NKG2C lacks an ITIM in its cytoplasmic domain, a feature shared by human and rat NKG2C. However, unlike the human counterpart, the transmembrane domain of mouse NKG2C does not contain a charged amino acid residue. Mouse NKG2A mRNA was detected in IL-2-activated NK cells and spleen cells but not in other tissues. The NKG2A gene was localized on the distal portion of chromosome 6 where the NK complex has been located. These results further extend the repertoire of C-type lectin receptors on murine NK cells.

Amino Acid Sequence↗

PubMed1999

Expression analysis of new Ly49 genes: most transcripts of Ly49j lack the transmembrane domain.

Five new Ly49 genes, named Ly49j-n, have recently been identified in C57BL/6 mice. This study examined the expression of three of these new genes, Ly49j, k, and n. To determine whether the Ly49j, k, and n genes were transcribed, gene-specific primers were used to amplify cDNA clones for each gene from C57BL/6 interleukin-2-activated natural killer (NK) cell cDNA. A full-length cDNA for Ly49j was detected which encodes a 267 amino acid protein and shares approximately 96% nucleotide identity with Ly49c and i. COS cells transfected with the Ly49j cDNA were shown to react with the monoclonal antibody 8H7, suggesting that the gene likely encodes a functional protein. Many different sized Ly49k and n transcripts were observed, although it is likely that they do not encode functional proteins due to missing exons or severe truncations in the open reading frames. Interestingly, the most abundant Ly49j transcript detected was shown to lack exon 3, which encodes the transmembrane domain. Similar studies performed on the same source of NK cell cDNA using Ly49c- and i-specific primers revealed the presence of transmembrane-less Ly49i transcripts, although at a much lower frequency than observed for Ly49j. We also detected Ly49g and h transcripts lacking the transmembrane domain. Despite the absence of the transmembrane region, the resulting Ly49 transcripts maintain their open reading frames, and therefore could potentially encode cytoplasmic proteins with a role in NK cell function.

Alternative Splicing↗

PubMed1997

[Space occupying lesion in the anterior mediastinum in a patient with Basedow disease and previously diagnosed osteosarcoma].

HISTORY AND CLINICAL FINDINGS: A 24-year-old woman with osteosarcoma of the right thigh, diagnosed 12 years ago, complained at a follow-up examination of decreased exercise tolerance, increased nervous tension, heat intolerance, weight loss, hair loss and irregular stools. Examination revealed tachycardia (100/min), mild exophthalmus and a small goitre. INVESTIGATIONS: A decreased basal TSH level (0.002 mU/ml), raised peripheral thyroid hormone (fT4 6.7 ng/dl, total T3 7.8 micrograms/l) and a TSH receptor antibody titre of 33.4 U/l) were compatible with immune type Graves' disease. Radiology revealed an upper mediastinal space-occupying lesion which scintigraphically was separate from thyroid tissue. A metastasis of the osteosarcoma or thymus hyperplasia were considered the most likely cause. TREATMENT AND COURSE: The mediastinal lesion regressed under thyrostatic treatment with carbimazole (20 mg daily by mouth). But the clinical picture, localization and negative scintigraphy provided the diagnosis of transitory thymus hyperplasia in the course of Graves' disease. CONCLUSION: In immune type Graves' disease with a mediastinal space-occupying lesion, not only intrathoracic goitre but also thymus hyperplasia should be considered in the differential diagnosis.

Adult↗

PubMed1997

[Morphological changes in human colon carcinoma after chemotherapy with 5-fluorouracil--a study in the nude mouse model].

Colorectal cancer represents one of the most important challenges in the field of cancer chemotherapy. 5-fluorouracil (5-FU) is used as the first-choice chemotherapeutic agent in the treatment of advanced gastrointestinal cancer. Despite this fact very little is known about the morphological changes of colon carcinoma after treatment with 5-FU. Thymusaplastic nude mice (n = 59) with subcutaneously heterotransplanted human coloncarcinoma (tubulo-papillary adenocarcinoma, grade II-III) were treated with 5-FU (40 mg/kg/body weight i. p. for five days). The animals were sacrificed, tumors were removed at 0 h, 24 h, 36 h, 48 h, 54 h, 60 h, 66 h, 72 h, 94 h, 118 h, 240 h after chemotherapy and the tumor tissue was embedded for light-microscopic and ultrastructural examination. Hydropic mitochondria and cytoplasmatic vacuoles were observed in tumor cells as early as 0 h after chemotherapy. These changes displayed a focal pattern. Damaged tumor tissue was surrounded by tumor cells with intact ultrastructure. In some regions tumor cells were separated from the basal membrane and showed signs of necrosis. These focal changes within the tumor tissue were observed from 0-240 h after treatment. Tumor capillaries did not show any damage. Treatment with 5-FU led to focal cytotoxic effects in the tumor. The remaining intact vascular system of the tumor may be a target for new therapy modalities.

Adenocarcinoma, Papillary↗