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R Ohlsson

Publications and source records attributed to R Ohlsson.

At least 37 records · Page 2Linked to original sources
PubMed1998

The paternal allele of the H19 gene is progressively silenced during early mouse development: the acetylation status of histones may be involved in the generation of variegated expression patterns.

Transcriptional silencing can reflect heritable, epigenetic inactivation of genes, either singly or in groups, during the life-time of an organism. This phenomenon is exemplified by parent-of-origin-specific inactivation events (genomic imprinting) for a subset of mammalian autosomal genes, such as H19. Very little is known, however, about the timing and mechanism(s) of silencing of the paternal H19 allele during mouse development. Using a novel in situ approach, we present evidence that the silencing of the paternal H19 allele is progressive in the trophectodermal lineage during early mouse development and generates variegated expression patterns. The silencing process apparently involves recruitment of histone deacetylases since the mosaic paternal-specific H19 expression reappears in trichostatin A-treated mouse conceptuses, undergoing in vitro organogenesis. Moreover, the paternal H19 alleles of PatDup.d7 placentas, in which a region encompassing the H19 locus of chromosome 7 is bipaternally derived, partially escape the silencing process and are expressed in a variegated manner. We suggest that allele-specific silencing of H19 share some common features with chromatin-mediated silencing in position-effect variegation.

Acetylation↗
PubMed1997

Inactivation of H19, an imprinted and putative tumor repressor gene, is a preneoplastic event during Wilms' tumorigenesis.

Genetic evidence shows that the parent of origin-dependent expression patterns of the Igf2 and H19 genes is coordinated in mouse, such that H19 controls the activity of Igf2 in cis. Equally compelling evidence for a similar situation in humans is absent, although the frequently observed activation of the maternal IGF2 allele (ie., loss of imprinting) in Wilms' tumors has been attributed to the silencing of the maternal H19 locus. We show here that loss of H19 activity is generally a preneoplastic event, which may be linked with an overgrowth lesion that has been proposed to be permissive for tumor formation. Although our results document one instance in which a postneoplastic loss of H19 activity correlates with loss of IGF2 imprinting at the cellular level, it appears that inactivation of H19 is more generally independent of loss of imprinting of IGF2, at least in our specimens. Our results imply that inactivation of H19 correlates with blastema overgrowth and can be independent of a regulatory role with respect to IGF2 imprinting status in cis.

Alleles↗
PubMed1996

Expression of a novel member of estrogen response element-binding nuclear receptors is restricted to the early stages of chorion formation during mouse embryogenesis.

Members of the nuclear hormone receptor gene family of transcription factors have been shown to be expressed in characteristic patterns during mouse organogenesis and postnatal development. Using an RT-PCR based screening assay, we have identified nuclear receptors expressed in embryonal carcinoma stem cells. One of the cDNAs characterized, mERR-2, was found to be expressed exclusively during a narrow developmental window in trophoblast progenitor cells between days 6.5 and 7.5 post coitum (p.c.). From 8.5 days p.c. and onwards, the mERR-2 gene activity evaded detection as analysed by in situ hybridization. We also show that the mERR-2 gene product and the estrogen receptor share a common target DNA-sequence recognition specificity unique among members of the gene family. Furthermore, efficient homodimerization and DNA-binding of the orphan receptor mERR-2 was found to be dependent on interaction with the heat shock protein 90, a molecular chaperone hitherto recognized to interact only with the steroid hormone receptor subgroup of nuclear receptors. Based on our results we suggest that the mouse orphan receptor mERR-2 has the potential to regulate overlapping gene networks with the estrogen receptor and may participate in signal transduction pathways during a short developmental period coinciding with the formation of the chorion.

Amino Acid Sequence↗
PubMed1996

Characterizing three-dimensional topography of engineering and biomaterial surfaces by confocal laser scanning and stylus techniques.

Three-dimensional measurements of surface topography were performed using a confocal laser scanner and a contact stylus instrument. Three surfaces known to be difficult to evaluate were chosen to be measured on the same area with the two instruments. The measurements from the optical and the contact stylus profilometer were compared with each other and with measurements obtained from high-resolution atomic force microscopy, which served as a reference instrument. Six implants manufactured from commonly used biomaterials were also measured on the same part of the implant, but not on the same area, with the optical and the contact profilometer in order to simulate the measurements that would be performed when different laboratories measure similarly treated surfaces. The numerical and visual differences achieved when measuring the same area with the two instruments investigated were compared. In general, we found an underestimation of the surface features with the contact stylus measurement and an overestimation with the confocal scanner. The stylus readings are mainly influenced by the radius of the stylus tip, the pressure of the stylus tip on the surface, and the hardness of the material. The optical profilometer has a tendency for creating spikes when surfaces with deep slopes are measured. For relatively soft metallic biomaterials, we found that using the optical instrument is the most appropriate method for surface roughness characterization, particularly when screw-shaped implants are analysed, whereas the stylus is preferred when larger areas with substantial slopes within the surface structure are to be evaluated.

Biocompatible Materials↗
PubMed1996

Genomic imprinting and mammalian development.

Genomic imprinting, which results in the mono-allelic expression of certain genes in a parent of origin-dependent manner, represents a specialized form of gene regulation which may be vitally important for mammalian development. The mechanisms which underlie imprinting and the molecular nature of the imprint itself remain elusive but most likely include epigenetic modifications of DNA, such as methylation and chromatin structure changes. It is clear, however, that many of the known imprinted genes play important developmental roles and that changes in the functional imprinting of some of these genes may have important pathological consequences, including placental abnormalities.

Animals↗
PubMed1996

Allele-specific in situ hybridization (ASISH) analysis: a novel technique which resolves differential allelic usage of H19 within the same cell lineage during human placental development.

Precursory studies of H19 transcription during human foetal development have demonstrated maternally derived monoallelic expression. Analyses in extra-embryonic tissues, however, have been more equivocal, with discernible levels of expression of the paternal allele of H19 documented in the first trimester placenta. By refining the in situ hybridization technique we have developed an assay to enable the functional imprinting status of H19 to be determined at the cellular level. This assay involves the use of oligonucleotide DNA probes that are able to discriminate between allelic RNA transcripts containing sequence polymorphisms. Biallelic expression of H19 is confined to a subpopulation of cells of the trophoblast lineage, the extravillous cytotrophoblast, while the mesenchymal stroma cells maintain the imprinted pattern of monoallelic expression of H19 throughout placental development. This data demonstrates that the low level of paternal H19 expression previously detected in normal human placenta is not due to a random loss of functional imprinting, but appears to result from a developmentally regulated cell type-specific activation of the paternal allele. In addition, biallelic expression of H19 does not seem to affect the functional imprinting of the insulin-like growth factor II gene, which is monoallelically expressed at relatively high levels in the extra-villous cytotrophoblasts. These results imply that the allelic usage of these two genes in normal human placental development may not be directly analogous to the situation previously documented in the mouse embryo.

Alleles↗
PubMed1995

An Inr-containing sequence flanking the TATA box of the human c-sis (PDGF-B) proto-oncogene promoter functions in cis as a co-activator for its intronic enhancer.

High-level activity of the human PDGF-B promoter in choriocarcinoma cell lines depends upon an atypical, intronic enhancer-like element which does not function with heterologous promoters tested. An extensive series of mutant PDGF-B promoter-driven constructs identified a sequence flanking the TATA box which is required specifically for enhancer-mediated transcription in human choriocarcinoma cell lines. This element, which we here term an enhancer-dependent cis co-activator (EDC) contains an Inr (initiator) consensus sequence upstream of the TATA box which is required, but not sufficient for its function. Requirement for the EDC is cell type-specific, since it was dispensable for enhancer-mediated transcription in a human breast cancer cell line. Although it lies within the region defined, the TATA box itself is not required for EDC function, or for basal promoter function which may derive from a second Inr-like sequence situated at the transcriptional start site. These observations indicate that interactions between some promoter and enhancer elements may be more complex than that generally described for 'classical' enhancer systems and may suggest an additional function for the initiator motif.

Base Sequence↗
PubMed1995

Expression, promoter usage and parental imprinting status of insulin-like growth factor II (IGF2) in human hepatoblastoma: uncoupling of IGF2 and H19 imprinting.

We have studied the promoter utilization and parental imprinting status of human IGF2 in three genetically informative hepatoblastomas from patients ranging in age from 9 months to 3 years. In all three cases, there is a downregulation of promoter P1 in the tumor tissues while the P2 and P3 promoters are upregulated compared to the normal liver. One of three patients displayed loss of imprinting (LOI) of IGF2 in the tumor tissue. We also investigated the expression of the H19 gene in all three cases and the methylation pattern in H19 from the patient with LOI of IGF2. The expression of H19 was greatly reduced in all tumors. Monoallelic H19 expression however, was retained even in the case which showed LOI of IGF2. Unlike the situation in Wilms' tumor, no differences in the methylation pattern between the normal liver and tumor tissues were observed in the H19 promoter or 3' region, using HpII analysis. We show here, that in contrast to the situation in Wilms' tumor, H19 expression is not a prerequisite for maintaining a monoallelic IGF2 expression.

Alleles↗
PubMed1995

Paternally derived H19 is differentially expressed in malignant and nonmalignant trophoblast.

The paternal allele of the H19 gene has been shown to be transcriptionally inactive in the developing human embryo. Using reverse transcription PCR and RNase protection assays, we demonstrate that expression of H19 is predominantly, but not exclusively, from the maternal allele in the human placenta. In situ hybridization analysis shows strong expression of the H19 gene in eight complete hydatidiform moles, hyperplastic tissues consisting of trophoblasts which contain only paternally derived genetic material, indicating that H19 is not functionally imprinted in this tissue. H19, a putative growth suppressor, is oppositely imprinted to the neighboring insulin-like growth factor II (IGF2) gene and an up-regulation of IGF2 expression has been linked previously to a down-regulation of H19 expression in the progression to Wilms' tumor. Two cases of complete hydatidiform mole which progressed to choriocarcinoma show high levels of expression of both H19 and IGF2. The choriocarcinomas which developed from these complete hydatidiform moles showed similar expression of IGF2 but a decreased number of H19-positive cells, which may reflect selection for cells expressing IGF2 and against those expressing H19 in this tissue.

Alleles↗
PubMed1995

Monoallelic expression of IGF2 at the human fetal/maternal boundary.

IGF2 is expressed in both placental and decidual tissues, enabling an analysis of the parental imprinting over the fetomaternal boundary. Evidence is provided that IGF2 is monoallelically expressed in both placenta and pregnant, as well as nonpregnant, endometrium. These observations suggest that the maternally derived IGF2 allele is inactivated during germline transmission. Comparison of promoter usage in decidua and placental samples shows that the P3 promoter appears to regulated independently of the others. These observations are discussed with respect to current models of IGF2 imprinting and the hypothesized conflict of parental reproductive interests which bears on the phenomenon of parental imprinting.

Alleles↗
PubMed1995

H19 is imprinted in the choroid plexus and leptomeninges of the mouse foetus.

It has been proposed that either the Igf-2 gene or the H19 gene--but not both--can be expressed from a given chromosome. Igf-2 is known to be biallelically expressed in the choroid plexus and leptomeninges, however, raising the question of whether H19 is down-regulated or absent there. We found by in situ hybridization that H19 is indeed expressed in the choroid plexus and leptomeninges of the developing mouse foetus. Comparison with the expression pattern of Igf-2 showed that the genes are coexpressed in all areas, with the exception of the choroid plexus epithelium. To evaluate whether H19 is also biallelically expressed in these tissues, we microdissected embryos from interspecific crosses and performed RNAse protection analysis on the isolated RNA. This revealed that H19 maintains its imprint in the choroid plexus/leptomeninges, being transcribed from the maternal allele at a level comparable to that in normal liver. We discuss the significance of these results for current models of Igf-2 and H19 imprinting.

Alleles↗
PubMed1995

Should health care be rationed by age?

Can rationing of health care by chronological age be justified on ethical grounds? The principles of equality, solidarity, liberty and efficiency are used as value premises in the discussion. Health care rationing by age is inconsistent with the principles of equality and liberty. But in some situations such rationing has support from the principles of solidarity and efficiency. The compromise suggested is that, as a rule, rationing by age should not be permitted, except in situations with intrinsic scarcity (as is the case with transplantation) and in situations with temporary extrinsic scarcity (as was the case with dialysis for a period of time). However, the main purpose is not to defend this position, but to identify and analyze the ethical conflicts such rationing gives rise to.

Age Factors↗
PubMed1995

Promoter-specific IGF2 imprinting status and its plasticity during human liver development.

IGF2 has been shown to be expressed preferentially from the paternally derived allele, although the maternal allele can be found active during both prenatal and postnatal development as well as in neoplastic tumours in humans. We addressed here whether or not the biallelic expression patterns that can be seen during postnatal human liver development reflected a coordinated change in the activities of the four promoters of human IGF2. We show here that the P2, P3 and P4 promoters, but not the P1 promoter, display monoallelic activity in embryonic, neonatal and younger infant liver specimens. The P2, P3 and P4 promoters can, however, be found active either monoallelically or biallelically or even monoallelically on opposite parental alleles in older infant and adult liver specimens. In contrast, H19, which is closely linked to IGF2, is monoallelically expressed in all postnatal liver samples analysed. We conclude that the functional imprinting status of IGF2 during postnatal liver development appears to be promoter/enhancer-specific and either partly or completely independent of H19.

Alleles↗
PubMed1995

IGF2 expression is a marker for paraganglionic/SIF cell differentiation in neuroblastoma.

Neuroblastoma is a childhood tumor of the sympathetic nervous system. Observations in the Beckwith-Wiedemann syndrome suggest that sympathetic embryonal cells with an abundant expression of the insulin-like growth factor 2 gene (IGF2) may be involved in the genesis of low-malignant infant neuroblastomas. We have therefore compared the cell type-specific IGF2 expression of the human sympathetic nervous system during early development with that of neuroblastoma. An abundant expression in normal sympathetic tissue was specific to extra-adrenal chromaffin cells, ie, paraganglia and small intensely fluorescent (SIF) cells, whereas sympathetic neuronal cells were IGF2-negative. A subpopulation of neuroblastomas expressed IGF2, which correlated with an early age at diagnosis, an extra-adrenal tumor origin, and severe hemodynamic signs of catecholamine secretion. Histologically IGF2-expressing tumors displayed a lobular growth pattern, and expression was restricted to the most mature and least proliferative cells. Typically, these cells were morphologically and histochemically similar to paraganglia/SIF cells and formed distinct ring-like zones in the center of the lobules around a core of apoptosis-like tumor cells. The similarities found between IGF2-expressing neuroblastoma cells and paraganglia/SIF cells in terms of histological features, anatomical origin, and age-dependent growth suggest a paraganglionic/SIF cell lineage of most infant tumors and also of extra-adrenal tumors diagnosed after infancy. Furthermore, since paraganglia/SIF cells undergo postnatal involution, the same cellular mechanism may be responsible for spontaneous regression in infant neuroblastoma.

Biomarkers, Tumor↗
PubMed1995

Normal development and neoplasia: the imprinting connection.

The observation that a number of autosomal genes are expressed in a parent of origin-dependent monoallelic manner has fuelled a frantic research effort into the underlying mechanisms and biological functions of this phenomenon, termed genomic or parental imprinting. The level of intrigue associated with this subject has been heightened by the discovery that the "transcriptional phenotype" of some imprinted genes shows developmental and tissue-specific variation, and that some imprinted genes are expressed biallelically in tumors. Here we describe some further examples of variation in the allele-specific transcription of an imprinted gene, human IGF2. Analysis of different sub-clones of an established tumor cell line (Jeg-3) revealed examples of both a switch from monoallelic to biallelic expression, as well as monoallelic expression from the opposite parental allele. Examination of IGF2 expression in adult human liver clearly demonstrated that the functional imprinting is manifested in a promoter-specific manner. The P1 promoter produced biallelically derived transcripts, whereas the remaining three promoters were utilized in a complex pattern of mono- and biallelic expression which varied from sample to sample. These observations emphasize the need to re-examine the imprinting phenomenon and its plasticity in terms of the cis elements and trans-acting factors involved in the transcriptional regulation of these genes both in the normal and pathological contexts.

Adult↗
PubMed1994

Insulin-like growth factor 2 cannot be linked to a familial form of Beckwith-Wiedemann syndrome.

The Beckwith-Wiedemann syndrome (BWS) is characterised by congenital malformations and organomegaly associated with an increased risk for development of childhood neoplasms. Both a sporadic and a familial form have been described in the literature. It has been suggested that duplications or rearrangements of the short arm of chromosome 11 (11p15.5) underlie the aetiology of the disease. This region of chromosome 11 contains the insulin-like growth factor 2 (IGF2) gene, which has been shown to be parentally imprinted in the sporadic form of the BWS with only the active, paternally-derived allele being duplicated. The familial form of BWS, which exhibits a predominantly maternal inheritance, has been suggested to result from a relaxation of IGF2 imprinting. This could render both parental IGF2 alleles active, thereby generating a similar gene dosage as in the sporadic from of the BWS. To address this issue, we used an RNase protection assay based upon a polymorphic region within exon nine of IGF2. We show here that only the paternally-inherited IGF2 allele is transcriptionally active in the index patient of one family with inherited BWS. In addition, highly informative IGF2 DNA markers were used to perform linkage analysis. Since these data ruled out a common maternally-transmitted IGF2 allele in the affected patients, we argue that IGF2 cannot be linked to the hereditary form to the disease.

Alleles↗