Biomedical subjects

F Hedborg

Publications and source records attributed to F Hedborg.

At least 19 recordsLinked to original sources

PubMed2001

Evidence of chromaffin oxygen sensing in neuroblastoma.

With the aid of IGF2 and VEGF in situ hybridization; tyrosine hydroxylase, chromogranin A, and Ki67 immunohistochemistry; and TUNEL staining applied to a large series of clinical neuroblastomas and to an animal model, we show here that stroma-poor neuroblastomas show evidence of chromaffin differentiation similar to that of type 1 small intensely fluorescent (SIF) cells and that this occurs in a vascular-dependent fashion, indicating a role for local tumor hypoxia in the differentiation process.

Animals↗

PubMed1999

Mosaic allelic insulin-like growth factor 2 expression patterns reveal a link between Wilms' tumorigenesis and epigenetic heterogeneity.

Numerous observations link the loss of imprinting of insulin-like growth factor 2 (IGF2) and an overdosage of this growth factor gene with cancer, in general, and with Wilms' tumorigenesis, in particular. It is not known, however, if loss of imprinting correlates with specific stages of neoplasia or if allelic expression patterns vary within the tumor. By applying an allele-specific in situ hybridization technique to formalin-fixed thin sections, we show that the parental IGF2 alleles can be differentially expressed, not only in Wilms' tumors, but also in nephrogenic rests (which represent premalignant lesions) of Wilms' tumor patients. Moreover, a subpopulation of mesenchymal cells, which surrounds tumor nodules, expresses IGF2 biallelically irrespective of the imprinted state of IGF2 within the tumor. These data show that Wilms' tumorigenesis involves epigenetic heterogeneity as visualized by variable allelic IGF2 expression patterns.

Alleles↗

PubMed1999

Inhibition of angiogenesis induces chromaffin differentiation and apoptosis in neuroblastoma.

Inhibition of angiogenesis has been shown to reduce tumor growth, metastasis, and tumor microvascular density in experimental models. To these effects we would now like to add induction of differentiation, based on biological analysis of xenografted human neuroblastoma (SH-SY5Y, WAG rnu/rnu) treated with the angiogenesis inhibitor TNP-470. Treatment with TNP-470 (10 mg/kg s.c., n = 15) reduced the tumor growth by 66% and stereological vascular parameters (Lv, Vv, Sv) by 36-45%. The tumor cell apoptotic fraction increased more than threefold, resulting in a decrease in viable tumor cells by 33%. In contrast, the mean vascular diameter (29 microm) and the mean tumor cell proliferative index (49%) were unaffected. TNP-470-treated tumors exhibited striking chromaffin differentiation of neuroblastoma cells, observed as increased expression of insulin-like growth factor II gene (+88%), tyrosine hydroxylase (+96%), chromogranin A, and cellular processes. Statistical analysis revealed an inverse correlation between differentiation and angiogenesis. It is suggested that by inhibiting angiogenesis, TNP-470 induces metabolic stress, resulting in chromaffin differentiation and apoptosis in neuroblastoma. Such agonal differentiation may be the link between angiostatic therapy and tumor cell apoptosis.

Animals↗

PubMed1998

Developmental gene expression of sympathetic nervous system tumors reflects their histogenesis.

Comparisons of the developing human sympathetic nervous system (SNS) to tumors presumed to derive from these cells may suggest tumor progenitors and predict tumor biologic behavior. Classic neuroblastoma (NB) and its more highly differentiated stroma-rich subtypes, extra-adrenal sympathetic paraganglioma, and pheochromocytoma were examined for the presence of the developmentally characterized gene products NSE, S-100, CD44, Bcl-2, HNK-1, PNMT, TrkA, IGF2, and tyrosine hydroxylase. The marker gene expression profiles of these tumors were compared with those similarly determined for a number of normal prenatal and postnatal human SNS cell types. Sympathetic paraganglioma, pheochromocytoma, and stroma-rich NB display marker expression profiles mimicking those of childhood sympathetic paraganglia, adrenal chromaffin cells, and sympathetic neurons, respectively. A selection of differentiating, extra-adrenal NB tumors with prognostically favorable features possess marker gene expression profiles paralleling that observed for fetal extra-adrenal sympathetic paraganglia/small intensely fluorescent cells. In contrast, undifferentiated, clinically aggressive NB tumors manifest characteristics mirroring that of embryonic/early fetal sympathetic neuroblasts of sympathetic ganglia and of the adrenal gland. These findings suggest that clinical features, such as primary tumor location and age at diagnosis, provide prognostic information for NB patients by virtue of the existence and biology of the presumed tumor progenitor cell type.

Adrenal Gland Neoplasms↗

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↗

PubMed1997

Homozygous deletion of the neurofibromatosis-1 gene in the tumor of a patient with neuroblastoma.

The neurofibromatosis type 1 (von Recklinghausen, NF1) gene has been proposed as a suppressor gene in tumors associated with neurofibromatosis. Recent publications have indicated that the NF1 gene can be rearranged in neuroblastoma cell lines. We analyzed DNA from a neuroblastoma patient with NF1 inherited as a familial trait on the paternal side. Using PCR and Southern techniques we showed that the patient had a constitutional deletion of several exons of the paternally derived NF1 gene and that the maternal copy of the gene had been deleted in the tumor of the patient. This is the first instance of a homozygous deletion reported in a primary neuroblastoma tumor. This suggests that NF1 inactivation in involved in the development or progression of some neuroblastomas in agreement with the hypothesized two hit model of inactivation for a tumor suppressor. These results are concordant with other groups that have detected unbalanced translocations t(1;17) in neuroblastoma tumors, with a breakpoint in chromosome 17 that may coincide with the location of the NF1 gene.

Blotting, Southern↗

PubMed1997

Delimitation of a critical tumour suppressor region at distal 1p in neuroblastoma tumours.

We analysed DNA from 68 neuroblastoma tumours for loss of heterozygosity (LOH) on the distal chromosome 1p (1p-LOH) using PCR-based DNA polymorphisms. Fifteen tumours (22%) displayed 1p-LOH. The shortest region of overlap (SRO) for the deletions was defined proximally by marker D1S244 and distally by marker D1S80. The CDC2L1 locus, located on chromosome 1p36, has been put forward as a neuroblastoma tumour suppressor. We analysed coding regions of the CDC2L1 gene in a subset of aggressive neuroblastoma tumours with known allelic loss for different 1p-markers. Single-stranded conformation polymorphism, heteroduplex and sequencing analysis of tumour DNA did not reveal any significant changes in the coding region. Using a DNA sequence polymorphism, we showed, in a primary tumour with an interstitial allelic deletion, that this tumour had both alleles of the CDC2L1 locus retained in the tumour. Thus, we showed that the neuroblastoma tumour suppressor critical region on 1p in our material is defined by loci D1S244 and D1S80 and that the CDC2L1 locus is distal to the critical region.

Child↗

PubMed1997

In vivo spontaneous neuronal to neuroendocrine lineage conversion in a subset of neuroblastomas.

Neuroblastoma is an embryonal tumor derived from the sympathetic nervous system. Although all neuroblastomas have a neuronal character, a subset of tumors also show evidence of extra-adrenal neuroendocrine differentiation in discrete cell layers. A characterization of the cells of the developing human sympathetic nervous system was performed, identifying growth-associated protein-43, neuropeptide tyrosine, and Bcl-2 as marker genes for sympathetic neurons. Whereas all neuroblastomas express growth-associated protein-43, neuropeptide tyrosine, and Bcl-2, tumors with differentiating cells with neuroendocrine features expressed these genes only in the morphologically immature, proliferating cells. Thus, with neuroendocrine tumor cell differentiation, neuronal marker gene expression vanished and proliferation ceased and was succeeded by expression of chromogranin A/B and insulin-like growth factor-2, markers of neuroendocrine chromaffin differentiation. These tumors appear to provide examples of spontaneous lineage conversion from a neuronal to a neuroendocrine phenotype.

Cell Differentiation↗

PubMed1996

A developmental model of neuroblastoma: differentiating stroma-poor tumors' progress along an extra-adrenal chromaffin lineage.

The prognosis of children with neuroblastoma (NB) is dependent upon the patient's age at diagnosis, the location of the primary tumor, and histologic tumor cell differentiation. These characteristics, as well as the presumption that NB results from clonal expansion of primitive cells involved in sympathetic nervous system (SNS) development, predict that a model of tumorigenesis based upon normal fetal SNS histogenesis might indicate tumor progenitor status and define biologic and clinical behavior. Immunohistochemistry and in situ hybridization were used to examine a panel of marker gene products predicted or shown to be expressed during SNS development in the normal human fetal SNS from 8 to 24 weeks' gestational age. A similar analysis was performed in a selection of clinical NB tumors, and the results were compared. In a subset of differentiating, often extra-adrenal NB tumors in patients who frequently had a favorable outcome; advancing morphologic tumor cell differentiation spatially paralleled an advancing fetal extra-adrenal chromaffin marker gene expression phenotype (ie, increasing TrkA, TrkC, TH, IGF-2, and neuron-specific enolase expression but a lack of phenylethanolamine N-methyltransferase expression). In these tumors, expression of gene products associated with normal fetal sympathetic ganglionic differentiation (ie, Bcl-2, HNK-1, and neuropeptide Y) was lost with morphologic tumor cell differentiation. In contrast, undifferentiated tumors, the majority of which were high stage, adrenal in origin, and prognostically unfavorable, displayed marker expression characteristics mirroring that of an early fetal ganglionic lineage. Thus, we show that morphologic differentiation in stroma-poor NB tumors, long held as an important prognostic feature in tumor grading systems, often corresponds to an extra-adrenal chromaffin rather than a ganglion cell or adrenal medullary chromaffin phenotype. Understanding the biology of extra-adrenal chromaffin tissues may provide an explanation for the clinically less aggressive nature of differentiating NB tumors and suggest potential mechanisms for spontaneous regression and/or treatment response.

Biomarkers, Tumor↗

PubMed1995

Deletion of chromosome 1p loci and microsatellite instability in neuroblastomas analyzed with short-tandem repeat polymorphisms.

We have analyzed DNA from 46 neuroblastoma tumors of all clinical stages and five ganglioneuroma tumors together with corresponding control DNA for loss of heterozygosity (LOH) on the distal 1p chromosomal region (1p-LOH). The markers used for the analyses were genetically mapped DNA polymorphisms detectable with PCR analysis. In general, there was concordance among aggressive tumor stage, 1p deletion, and N-myc amplification, although exceptions were found. Twelve (26%) of the 46 neuroblastoma tumors displayed 1p-LOH, 11 being stage 4 and 1 stage 2 (which progressed subsequently to stage 4), whereas 10 stage 4 tumors showed no 1p-LOH. Of 12 neuroblastomas shown to have N-myc amplification, 10 had 1p-LOH. In 8 cases it was possible to test for parental origin of the chromosome involved in 1p-LOH. No significant correlation between LOH and paternal or maternal allele was found. Commonly deleted loci in the distal 1p region in the neuroblastoma tumors indicated that the region for a tentative neuroblastoma tumor suppressor gene is defined proximally by marker D1S244 and distally by marker D1S80. One striking feature of three stage 2 neuroblastomas and one of the stage 3 tumors was the presence in the tumor DNA of alleles not present in the constitutional DNA of the patients, i.e., microsatellite instability. The significance of this phenomenon in localized neuroblastoma tumors remains to be clarified. Aggressive neuroblastoma in young children (younger than 2 years of age) seems to be a homogenous disorder consistently showing concomitant 1p-LOH and N-myc amplification. In the majority of unfavorable neuroblastoma in older children, however, neither 1p-LOH nor N-myc amplification could be detected. This indicates that neuroblastoma in older children is a biologically more heterogenous disorder in which genetic alterations other than deletions of chromosome 1p and amplification of N-myc also may contribute to tumorigenesis.

Child↗

PubMed1995

Cellular death in neuroblastoma: in situ correlation of apoptosis and bcl-2 expression.

Apoptosis is the selective physiologic deletion of cells that are no longer required. Over-expression of the bcl-2 proto-oncogene extends survival of neurons otherwise destined for apoptosis. The unique capacity of neuroblastoma (NB) to undergo spontaneous regression and the prognostic dichotomy of children with this malignancy led us to evaluate bcl-2 expression and apoptosis in NB. An in situ DNA nick-labeling technique to detect apoptotic cells, as well as immunohistochemistry and morphology, were utilized in a selection of NB tumor specimens and in the human fetal sympathetic nervous system. bcl-2 expression was present in all 28 NB tumors examined and in sympathetic ganglia of the human fetus. Measurement of overall bcl-2 expression and of extent of apoptosis correlated with favorable prognosis. In low-stage tumors, bcl-2 expression was most intense in poorly differentiated tumor cells adjacent to fibrovascular stroma. Cells distant from the stroma exhibited increasing degrees of chromaffin differentiation, with apoptosis most evident in bcl-2-negative neuroblasts adjacent to well-differentiated NB cells. The spatial distribution of bcl-2 expression, apoptosis and chromaffin differentiation in favorable-prognosis NB may provide insight into mechanisms of persistent tumor existence or regression.

Apoptosis↗

PubMed1995

The expression profile of alternatively spliced neuronal c-src RNA distinguishes between human tumours of the sympatho-adrenal lineage.

Human neuronal and neuroendocrine tumour specimens and cell lines were analysed regarding proteins and transcripts coded by the proto-oncogene c-src. At the protein level, most of the neuroblastomas and phaeochromocytomas expressed the neuronal c-src form, pp60c-srcN. None of the other neuroendocrine tumours, i.e. paragangliomas, neuroendocrine pancreatic tumours, or carcinoid tumours and small-cell lung carcinomas of different types, appeared to express the neuronal form. In the brain, c-src is transcribed into 3 differently spliced mRNA variants, c-src, c-srcNI, and c-srcNI+NII. The expression of these transcripts was analysed by PCR amplification of fragments covering the mini-exons I and NII of the corresponding cDNAs. The PCR products were analysed by Southern hybridization and characterized by determination of their sequences. Neuroblastomas, paragangliomas, retinoblastomas and the phaeochromocytomas expressed neuronal c-src splice variants. However, whereas neuroblastomas and retinoblastomas contained all 3 transcripts, the phaeochromocytomas and paragangliomas expressed, with 2 exceptions, only the c-src and the c-srcNI+NII mRNA species. To assess whether neuroblastomas display adrenal chromaffin characteristics, they were analysed regarding expression of the chromaffin marker enzyme, phenylethanolamine-N-methyl transferase. Whereas phaeochromocytomas were positive, all neuroblastomas were immuno-chemically negative for this enzyme. These results and the c-src expression profile suggest that neuroblastomas, including those with an adrenal location, do not originate from the adrenal chromaffin differentiation lineage. The data further suggest neuronal c-srcNI mRNA as a marker for sympathetic neuronal cells of the sympatho-adrenal lineage.

Adrenal Gland Neoplasms↗

PubMed1995

Biochemical evidence for a mature phenotype in morphologically poorly differentiated neuroblastomas with a favourable outcome.

Neuroblastoma is an embryonal tumour of the sympathetic nervous system with marked heterogeneity in terms of histological maturity and clinical course. A previous study revealed that high tumour levels of the csrc protein, particularly its neuronal isoform (pp60csrcN), correlated with favourable outcome. To test whether this feature reflects a higher degree of neuronal maturation in these tumours, an extended series (47 consecutive neuroblastomas and 10 ganglioneuromas) were analysed for levels of csrc protein isoforms, neuron-specific enolase, and synaptophysin. Immunoblotting and radioimmunoassay techniques were employed. The results were compared with conventional histological signs of neuronal maturation. High pp60csrcN levels were specific for prognostically favourable neuroblastomas and correlated with high neuronal marker levels. However, signs of histological maturation correlated poorly with these parameters. It is therefore concluded that low stage tumours are highly differentiated in biochemical terms despite their frequently immature histology. Furthermore, the clinical usefulness of these biochemical parameters as prognostic markers was compared with established parameters in a multivariate analysis. Stage 4 disease, MYCN amplification, and age above 18 months at diagnosis was the most powerful combination of variables found for predicting a poor outcome. As expected, none of the neuronal differentiation markers investigated could add to the prediction of aggressive disease when compared with this model. However, high expression of pp60csrcN appeared to be useful in predicting long-term survival in high stage infant neuroblastoma.

Age Factors↗

PubMed1995

Differentiation and survival influences of growth factors in human neuroblastoma.

Human neuroblastoma cell lines are established from high-stage, highly malignant tumours. Despite this and the fact that these tumours are arrested at an early, immature stage, many cell lines have the capacity to undergo neuronal differentiation under proper growth conditions. One such cell line is the noradrenergic SH-SY5Y cell line. These cells can be induced to mature by a variety of modalities, resulting in different mature phenotypes. The use of this cell system as a model to study the stem cell character of neuroblastoma is reviewed and discussed. In particular, we focus on growth factor dependencies in the SH-SY5Y system, and compare that to the normal situation, i.e. growth factor control of sympathetic neuronal and neuroendocrine differentiation during human and rat embryogenesis.

Cell Differentiation↗

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↗

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↗

PubMed1994

Overlapping patterns of IGF2 and H19 expression during human development: biallelic IGF2 expression correlates with a lack of H19 expression.

The spatial patterns of IGF2 and H19 gene expression are strikingly similar during parts of human embryonic/fetal and early postnatal development. Notable exceptions were found with the ciliary anlage of the embryonic retina and the choroid plexus/leptomeninges, where transcripts from the IGF2 but not the H19 locus could be detected. Moreover, in contrast to the other tissue samples examined, the choroid plexus/leptomeninges expressed both parental IGF2 alleles. Whilst RNase protection analysis revealed a weak activity of the P1 promoter in the choroid plexus/leptomeninges, the P2, P3 and P4 promoters were all active wherever IGF2 was expressed. We discuss these observations with respect to a hypothesized coordinated control of the reciprocally imprinted and closely linked IGF2 and H19 loci.

Alleles↗

PubMed1994

The cell type-specific IGF2 expression during early human development correlates to the pattern of overgrowth and neoplasia in the Beckwith-Wiedemann syndrome.

Overstimulation by insulin-like growth factor II is implied in several overgrowth conditions and childhood cancers. We have therefore studied spatial and temporal expression patterns of the insulin-like growth factor II gene (IGF2) and the insulin-like growth factor type 1 receptor gene during normal human development (5.5 to 23.0 weeks postfertilization). The set of cell types with the most abundant IGF2 expression correlated strikingly to the organomegaly and tumor predisposition of the Beckwith-Wiedemann syndrome. Intrauterine growth and postnatal organ weights of a prematurely born child with a full-blown syndrome are presented. The cell type-specific IGF2 expression of these organs and of multifocal Wilms' tumors from two other children affected by the Beckwith-Wiedemann syndrome were also studied. The results clarify and extend previous findings concerning human prenatal IGF2 expression and are consistent with a short range overstimulatory role of locally produced IGF II ensuing after the first trimester in the Beckwith-Wiedemann syndrome.

Beckwith-Wiedemann Syndrome↗