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E Simpson

Publications and source records attributed to E Simpson.

At least 91 records · Page 5Linked to original sources
PubMed1993

Exon-specific northern analysis and rapid amplification of cDNA ends (RACE) reveal that the proximal promoter II (PII) is responsible for aromatase cytochrome P450 (CYP19) expression in human ovary.

Estrogens are synthesized from C19 steroids by a unique form of cytochrome P450, aromatase cytochrome P-450 (P-450AROM; the product of the CYP19 gene). We have shown that tissue-specific expression of human P-450AROM is determined, in part, by the use of alternative promoters. Previous methods of analysis for determining the specific 5'-termini of the different transcripts included S1 nuclease protection, primer extension, and Northern analysis. In the present study we have used the RACE procedure (rapid amplification of cDNA ends) to amplify and clone the 5' termini of P-450AROM transcripts expressed in human corpus luteum (CL). Sequencing of the resulting clones supports the results of the previously performed studies. Specifically, the proximal promoter, PII, is the predominant promoter utilized in CL, such that the start of transcription occurs 26 bp downstream of the putative TATA sequence. A minority of the clones possess an alternative 5'-end, namely I.3. Exon-specific Northern analysis confirms that the majority of the P-450AROM transcripts in CL tissue contain sequence specific for promoter II. Similarly, exon-specific Northern analysis indicates that transcripts in human follicles, as well as granulosa cells in culture, contain primarily sequence specific for promoter II.

Aromatase↗
PubMed1993

Deletion of Y chromosome sequences located outside the testis determining region can cause XY female sex reversal.

An approach designed to map and generate mutations in the region of the short arm of the mouse Y chromosome, known to be involved in sex determination and spermatogenesis, is described. This relies on homologous Yp-Sxra pairing and asymmetrical exchange which can occur at meiosis in XY males carrying Sxra on their X chromosome. Such exchange potentially generates deficiencies and duplications of Yp or Sxra. Three fertile XY females were found out of about 450 XY offspring from XSxra/Y x XX crosses. In all three, despite evidence for deletion of Y chromosomal material, the Sry locus was intact. Each deletion involved a repeat sequence, Sx1, located at a distance from Sry. Since expression of Sry was affected these results suggest that long range position effects have disrupted Sry action.

Animals↗
PubMed1993

Loss of the 'azoospermia factor' (AZF) on Yq in man is not associated with loss of HYA.

We have typed 9 EBV cell lines from azoospermic or severely oligospermic patients for the expression of H-Y antigen, in order to test the hypothesis of the coincidence of AZF and HYA genes. Of nine patients with cytogenetically normal Y chromosomes, 7 could be tested for HYA expression and of these 6 were H-Y positive. Of the three patients showing Yq structural abnormalities, two could be tested for H-Y expression and one was negative, the other positive. These results therefore show no correlation between spermatogenic failure and the absence of HYA, thus separating the AZF locus from HYA.

Adult↗
PubMed1993

T cell deletion follows chronic antigen specific T cell activation in vivo.

Exposure of mice transgenic for a TCR (F5) to cognate peptide antigen results in thymic depletion of CD4+CD8+ cells and expansion and activation of peripheral CD8+ TCR(tg)+ T cells. In the thymus apoptotic DNA ladder is evident as early as 3 h after peptide injection. Long exposure of intact or thymectomized F5 TCR transgenic mice to peptide antigen leads to depletion of most of the peripheral CD8+ T cells bearing the F5 receptor, with the remaining cells having lower levels of transgenic TCR compared with non-treated animals. In the thymus of intact F5 TCR transgenic mice such continuous exposure to antigen results in the reappearance of CD4+CD8+ with lower levels of the transgenic receptor.

Animals↗
PubMed1993

T cell repertoire selection by mouse mammary tumour viruses.

Mouse mammary tumour viruses (Mtv) are B-type retroviruses. These can be exogenous, transmitted via maternal milk, or endogenous, as proviral integrations into the mouse genome, transmitted vertically in a Mendelian fashion. A number of different sites of integration of endogenous Mtvs have been reported in various inbred mouse strains. An open reading frame (ORF), within the long terminal repeat (LTR) of Mtv, encodes a type 2 integral membrane glycoprotein. The ORF products are expressed in association with MHC class II molecules at the cell surface and have an affinity for certain T cell receptor (TCR) V beta chains such that CD4+8+ TCR+ double positive thymocytes expressing these V beta chains undergo programmed cell death in mice carrying the appropriate endogenous or exogenous Mtvs. This constitutes a measurable part of negative repertoire selection of the T cell repertoire. Some positive selection of the T cell repertoire also appears to be TCR V beta-specific, although the involvement of polymorphic ligands other than MHC molecules is not apparent. This minireview summarizes the published work on the TCR V beta specificity and chromosomal localization of the various mouse mammary tumour proviral integrations leading to negative selection, and discusses the nature of TCR V beta-specific positive selection.

Animals↗
PubMed1993

Positive and negative selection in transgenic mice expressing a T-cell receptor specific for influenza nucleoprotein and endogenous superantigen.

A transgenic mouse was generated expressing on most (> 80%) of thymocytes and peripheral T cells a T-cell receptor isolated from a cytotoxic T-cell clone (F5). This clone is CD8+ and recognizes alpha alpha 366-374 of the nucleoprotein (NP 366-374) of influenza virus (A/NT/60/68), in the context of Class I MHC Db (Townsend et al., 1986). The receptor utilizes the V beta 11 and V alpha 4 gene segments for the beta chain and alpha chain, respectively (Palmer et al., 1989). The usage of V beta 11 makes this TcR reactive to Class II IE molecules and an endogenous ligand recently identified as a product of the endogenous mammary tumour viruses (Mtv) 8, 9, and 11 (Dyson et al., 1991). Here we report the development of F5 transgenic T cells and their function in mice of the appropriate MHC (C57BL/10 H-2b, IE-) or in mice expressing Class II MHC IE (e.g., CBA/Ca H-2k and BALB/c H-2d) and the endogenous Mtv ligands. Positive selection of CD8+ T cells expressing the V beta 11 is seen in C57BL/10 transgenic mice (H-2b). Peripheral T cells from these mice are capable of killing target cells in an antigen-dependent manner after a period of in vitro culture with IL-2. In the presence of Class II MHC IE molecules and the endogenous Mtv ligand, most of the single-positive cells carrying the transgenic T-cell receptor are absent in the thymus. Unexpectedly, CD8+ peripheral T-cells in these (H-2k or H-2d) F5 mice are predominantly V beta 11 positive and also have the capacity to kill targets in an antigen-dependent manner. This is true even following backcrossing of the F5 TcR transgene to H-2d scid/scid mice, in which functional rearrangement of endogenous TcR alpha- and beta-chain genes is impaired.

Animals↗
PubMed1993

A comparative study of T-cell receptor V beta usage in non-obese diabetic (NOD) and I-E transgenic NOD mice.

The non-obese diabetic (NOD) mouse is a model for the study of insulin-dependent diabetes mellitus (IDDM). Recently transgenic NOD mice have been derived (NOD-E) that express the major histocompatibility complex (MHC) class II I-E molecule. NOD-E do not become diabetic and show negligible pancreatic insulitis. The possibility pertained that NOD-E mice are protected from disease by a process of T-cell deletion or anergy. This paper describes our attempts to discover whether this was so, by comparing NOD and NOD-E mouse T-cell receptor V beta usage. Splenocytes and lymph node cells were therefore tested for their ability to proliferate in response to monoclonal anti-V beta antibodies. We were unable to show any consistent differences between NOD and NOD-E responses to the panel of antibodies used. Previously proposed V beta were shown to be unlikely candidates for deletion or anergy. T cells present at low frequency (V beta 5+) in both NOD and NOD-E mice were shown to be as capable of expansion in response to antigenic stimulation as were more frequently expressed V beta. Our data therefore do not support deletion or anergy as mechanisms which could account for the observed disease protection in NOD-E mice.

Animals↗
PubMed1992

Thymic depletion and peripheral activation of class I major histocompatibility complex-restricted T cells by soluble peptide in T-cell receptor transgenic mice.

Injection of mice transgenic for a class I major histocompatibility complex-restricted T-cell receptor with a soluble peptide antigen from influenza virus nucleoprotein results in clonal depletion of double-positive immature thymocytes in the thymus and activation of mature T cells in the periphery, accompanied by a transient up-regulation of the T-cell receptor and CD3 and CD8 coreceptor molecules.

Animals↗
PubMed1992

Expression of major histocompatibility complex class I antigens at low levels in the thymus induces T cell tolerance via a non-deletional mechanism.

Transgenic CBA (H-2k haplotype) mice expressing the H-2 Kb major histocompatibility complex (MHC) class I gene under control of transcriptional promoter elements from a milk protein gene display high-level H-2 Kb transcription in lactating mammary glands and low-level transcription in skin and thymus of male and virgin female transgenic mice. However, H-2 Kb antigen could be detected only in lactating mammary gland epithelial cells by immunohistological methods. All transgenic mice are tolerant of H-2 Kb since they fail to reject skin grafts from mice expressing H-2 Kb molecules. Furthermore, anti-H-2 Kb cytotoxic responses could not be generated using responder T cells from transgenic mice but T cells from the same mice proliferated, in the presence of interleukin-2, in response to stimulator cells expressing H-2 Kb. Tolerance to H-2 Kb is induced in the thymus since CBA mice grafted with thymus tissue from transgenic mice fail to reject H-2 Kb disparate skin grafts. However, experiments with double-transgenic mice also expressing a T cell receptor with anti-H-2 Kb specificity reveal that tolerance induction is not brought about by elimination of thymocytes bearing H-2 Kb-reactive receptors. Instead, a non-deletional mechanism which results in down-modulation of both CD8 and T cell receptor expression in peripheral T cells correlates with the induction of tolerance in these mice. These data reveal that extremely low levels of self-antigen expression in the thymus are sufficient to induce tolerance via non-deletional mechanisms.

Animals↗
PubMed1992

Mechanisms of transplantation immunity.

In summary, this chapter describes the biology and genetics of the major and minor histocompatibility antigens and the nature of in vitro and in vivo immune responses to them and to tissue-specific antigens. It reviews the nature and action of immune response genes. It gives an account of how tolerance to histocompatibility antigens was originally defined and the prospects of intervention aimed at establishing tolerance to these and tissue-specific antigens in adult animals, including man.

Animals↗
PubMed1992

A new approach to the cloning of genes encoding T-cell epitopes.

The molecular structure of antigens recognized exclusively by T cells, such as minor histocompatibility antigens and some antigens that provoke autoimmune responses, has proved difficult to determine. Recently, several antigens induced on tumor cells by mutagen treatment have been cloned by transfection of genomic DNA libraries into P1.HTR cells, screening for antigen expression using T-cell clones, and subsequent recovery of the integrated DNA by cosmid rescue. We have modified this technique and have stably transfected P1.HTR cell lines with polyoma T antigen, which allows episomal replication of the shuttle vector, pCDM8. Using pCDM8-CAT constructs, we have determined the frequency of transfection and plasmid copies taken up per cell under optimal transfection conditions. Using a pCDM8 construct which expresses the tumor-specific antigen, P91A (pCDM8-tum-), that is recognized by a T-cell clone, we have found that cells transfected with this antigen can be recognized by the T-cell clone when they are present at only 1%-3% of a mixed population. Progeny of a single cell transfected with pCDM8-tum-: pCDM8-CAT at proportions of 1:10, 1:25, and 1:50 are recognized by the T-cell clone. Furthermore, Hirt extracted plasmid DNA from transfectants expressing the tum- antigen can be amplified in bacteria, transfected back into P1.HTR recipients, and recognized by the T-cell clone. This approach should enable reasonably rapid screening of cDNA libraries for even relatively low abundance messages encoding, for example, minor histocompatibility and alloantigens, and allow their subsequent cloning.

Animals↗
PubMed1992

PCR-analyzed microsatellites of the mouse genome--additional polymorphisms among ten inbred mouse strains.

Eighty sequences from the mouse genome database containing microsatellites (simple sequence repeats) have been analyzed for size variation among ten different inbred strains of mice; 62/80 (77.5%) showed polymorphism of at least three alleles. We have been able to detect all the polymorphisms by agarose gel electrophoresis, often running the gels for up to 3 h. Between individual pairs of mouse strains to be used in chromosomal mapping studies in our laboratory, 35-60% polymorphism occurred. There are potentially enough microsatellites within the mouse and human genome to have a marker at every 1-cM distance. This simple approach will, therefore, continue to be useful in genome mapping studies, leading eventually to high-resolution maps of both the mouse and human genomes; this should allow for physical mapping and cloning of specific genes.

Animals↗
PubMed1992

T-cell-targeted immunotherapy.

The pace of T-cell research is matched only by the speed with which fundamental advances are being developed as new therapies. This report from a recent meeting updates developments in a number of immunointervention strategies.

AIDS Vaccines↗
PubMed1992

Deletion mapping of H-Y antigen to the long arm of the human Y chromosome.

A gene encoding or controlling the expression of the H-Y transplantation antigen was previously mapped to the human Y chromosome. We now report the sublocalization of this gene on the long arm of the human Y chromosome. Eight patients with Y-chromosomal abnormalities were examined with a series of existing and new DNA markers for the Y chromosome. The resulting deletion map was correlated with H-Y antigen expression. We conclude that the H-Y antigen gene maps to a portion of deletion interval 6 that is identified by specific DNA markers.

Adolescent↗
PubMed1992

Recombination between the X and Y chromosomes and the Sxr region of the mouse.

The Sxr (sex-reversed) region that carries a copy of the mouse Y chromosomal testis-determining gene can be attached to the distal end of either the Y or the X chromosome. During male meiosis, Sxr recombined freely between the X and Y chromosomes, with an estimated recombination frequency not significantly different from 50% in either direction. During female meiosis, Sxr recombined freely between the X chromosome to which it was attached and an X-autosome translocation. A male mouse carrying the original Sxra region on its Y chromosome, and the shorter Sxrb variant on the X, also showed 50% recombination between the sex chromosomes. Evidence of unequal crossing-over between the two Sxr regions was obtained: using five markers deleted from Sxrb, 3 variant Sxr regions were detected in 159 progeny (1.9%). Four other variants (one from the original cross and three from later generations) were presumed to have been derived from illegitimate pairing and crossing-over between Sxrb and the homologous region on the short arm of the Y chromosome. The generation of new variants throws light on the arrangement of gene loci and other markers within the short arm of the mouse Y chromosome.

Animals↗
PubMed1992

A molecular deletion map of the Y chromosome long arm defining X and autosomal homologous regions and the localisation of the HYA locus to the proximal region of the Yq euchromatin.

41 Y-linked DNA probes that detect sequences on the Y chromosome long arm have been used to analyse genomic DNA from a series of 23 patients with deletions of Yq. Southern blot analysis has differentiated 15 distinct breakpoints, which divide Yq into 14 mapping intervals. From the pattern of DNA sequences present in each patient, it has been possible to produce a congruent deletion map, with the exception of two cases which are not compatible with the consensus order. These patients can be explained by the presence of inversion polymorphisms on Yq in the general population or by complex rearrangements induced during the formation of the deleted chromosomes. The distribution of sequences on the Y long arm has defined distinct regions of homology with autosomes, the Y short arm and the long and short arms of the X. A number of the patients have been typed for the presence or absence of H-Y antigen (as determined by the cytotoxic T-cell assay) and it has been possible, from analysis of informative cases, to assign the locus to the proximal region of the Yq euchromatin.

Cell Line↗