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R F Gagel

Publications and source records attributed to R F Gagel.

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

Down-regulation of calcitonin gene transcription by vitamin D requires two widely separated enhancer sequences.

Transcription of the calcitonin (CT) gene is down-regulated by vitamin D in normal and transformed thyroid C cells. DNA transfer techniques have been previously used to map and characterize a cAMP-induced enhancer at nucleotides -255 to -129 and an enhancer of basal transcription at -1060 to -905 in the CT 5' flanking DNA. The same methods were used to identify a negative response element for vitamin D. Deletion mutants of a genomic fragment of CT extending from nucleotides -1460 to +90 were attached to a promoterless GH gene and transfected individually into the medullary thyroid carcinoma cell line TT. CT nucleotides -1460 to -129 induced significant basal transcription of the GH reporter gene in TT cells. Basal transcription was elevated 3-fold to 4-fold by treatment with cAMP analog. The biologically active metabolite of vitamin D3, 1,25-dihydroxyvitamin D3, had a minor (20%) inhibitory effect on basal transcription but inhibited more than 60% of the cAMP-induced transcription. We further investigated the cAMP-induced response and found that transcriptional activity of the downstream cAMP-induced enhancer was greatly synergized in the presence of the upstream enhancer of basal transcription. The latter enhancer contained three functional CANNTG sequences designated E1 (nucleotides -1060 to -1030), E2 (nucleotides -940 to -920), and E3 (nucleotides -920 to -900). E2 and E3 were essential for maximal cAMP-induced transcription. Detailed mapping of the vitamin D response showed that a minimum requirement for inhibition of the cAMP-induced enhancer by vitamin D was a sequence overlapping E3 (nucleotides -920 to -829). We conclude that a negative response element to vitamin D is located between nucleotides -920 and -829 in the CT 5' flanking DNA. It is possible that vitamin D inhibits transcription by interfering with the synergistic interaction between the cAMP-induced enhancer and the enhancer of basal transcription.

24,25-Dihydroxyvitamin D 3↗
PubMed1993

Nonfunctioning islet cell carcinoma of the pancreas.

BACKGROUND: Nonfunctioning islet cell carcinoma of the pancreas has a variable and often indolent natural history, which has resulted in a wide range of treatment recommendations. To more clearly define the natural history and appropriate treatment of this disease, we reviewed our institutional experience over the last 39 years. METHODS: The records of all patients confirmed to have a nonfunctioning islet cell carcinoma of the pancreas were retrospectively reviewed. Kaplan-Meier life tables were constructed and log-rank comparisons were performed. RESULTS: The 73 patients studied had an overall 5-year actuarial survival rate of 50%. Patients with localized disease at presentation (n = 39) had a significantly higher survival rate (p = 0.03) compared with patients with metastatic disease (n = 34). The 19 patients who underwent a potentially curative resection of the primary tumor had a significantly higher survival rate (p = 0.03) compared with the 20 patients with locally advanced, unresectable, nonmetastatic disease. Nine of these 20 patients died of complications of the primary tumor. In contrast, only 2 of 22 cancer-related deaths in the 34 patients with metastatic disease at diagnosis were due to the primary tumor. CONCLUSIONS: (1) Surgical resection should be performed in patients with resectable nonmetastatic disease. (2) Resection of the primary tumor in the presence of metastatic disease is rarely indicated. (3) Innovative treatment strategies are needed for patients with locally advanced, unresectable, nonmetastatic tumors of the pancreatic head.

Adolescent↗
PubMed1992

Identification of exon sequences and an exon binding protein involved in alternative RNA splicing of calcitonin/CGRP.

Transcripts derived from the 6 exon CALC I gene are differentially processed in a tissue-specific fashion to include or exclude a calcitonin-specific exon 4. All cell types which transcribe a second calcitonin/CGRP gene, CALC II, exclude exon 4. Substitution of the first 30 nucleotides of CALC I exon 4 with analogous CALC II sequence was sufficient to prevent recognition of exon 4 in in vitro or in vivo RNA splicing systems. UV crosslinking detected a approximately 66 kDa RNA-binding protein in HeLa nuclear extract which interacted with CALC I proximal exon sequence, but not CALC II or mutant sequences. UV crosslinking of this protein was inhibited by addition of nuclear extract from a cell type which normally causes exclusion of exon 4. These results identify an important regulatory element within exon 4 and support a model in which calcitonin production requires protein interaction with this sequence to facilitate exon recognition.

Amino Acid Sequence↗
PubMed1992

A preliminary analysis of consortium data for markers tightly linked to multiple endocrine neoplasia type 2A.

We have analyzed DNA marker typing data contributed by six independent groups to estimate the pairwise genetic distances between these markers and the locus for multiple endocrine neoplasia type 2A (MEN 2A). We used LIPED to calculate these distances for female, male, and sex-average linkage maps and to determine the corresponding LOD scores. The preliminary analyses of this large data set (89 MEN 2A families and five non-MEN 2A references families, with 1,934 total individuals) are reported here. These refined estimates of the genetic map in this region will aid in the assignment of presymptomatic diagnoses. This study clearly points out the limitation of pairwise linkage analysis in further refining the position of MEN2A in this small region of chromosome 10. Further refinement of the genetic map position of MEN2A will be best accomplished by finding, verifying, and accurately mapping crossovers in specific families.

Chromosome Mapping↗
PubMed1991

Validation of an in vitro RNA processing system for CT/CGRP precursor mRNA.

The pre-mRNA encoding calcitonin (CT) and calcitonin gene-related peptide (CGRP) is differentially processed in a tissue-specific fashion to include or exclude the calcitonin-specific exon 4. A minigene containing a viral first exon and exons 4, 5, and 6 from the human CT/CGRP gene was correctly processed in transfected HeLa or F9 teratocarcinoma cells to produce mRNA that included or excluded exon 4, respectively. This processing decision could be reproduced in vitro using nuclear extracts from these two cell lines and an RNA precursor from a similar minigene. Supplementation of extract from HeLa cells with extract from F9 cells resulted in the F9 splicing pattern in which exon 4 was excluded. This model system may be useful for the purification of splicing factors important in the regulation of this splice choice.

Adenoviridae↗
PubMed1990

Transfection of calcitonin gene regulatory elements into a cell culture model of the C cell.

Calcitonin gene expression in the TT cell line can be regulated by phorbol esters, cAMP, glucocorticoids, and 1,25-dihydroxyvitamin D3. To further study the regulation of this gene we have sequenced 1460 bases 5' to the start of calcitonin gene transcription. This DNA sequence contains cis consensus elements for both phorbol ester- and cAMP-responsive elements. To study the role of these elements, calcitonin 5' flanking DNA was coupled to the human growth hormone gene as a reporter and transiently transfected into TT cells, a human thyroid C cell line. Treatment of transfected TT cells stimulated a two- to fivefold increase in reported gene product expression, confirming the existence of functional cAMP- and phorbol ester-dependent enhancers within the calcitonin 5' flanking sequence.

Animals↗
PubMed1990

Calcitonin exon sequences influence alternative RNA processing.

The pre-mRNA encoding calcitonin (CT) and CT gene-related peptide (CGRP) is differentially processed in a tissue-specific fashion to include exon 4 (which encodes CT) or exclude this exon and splice to exon 5 (which encodes CGRP). We have used a CT-specific in vitro RNA-processing system to identify cis-acting sequences required to prevent splicing to exon 5. Deletion mapping demonstrated the presence of an element within the first 45 nucleotides of the CT-specific exon 4 that was required to suppress splicing to the CGRP-specific exon 5. This element was able to function in a completely heterologous system to suppress splicing when the CGRP exon was replaced with a constitutive viral exon. The element was unable to suppress splicing in the absence of a proximal CT-specific 3' splice site. Our results suggest that CT-specific splicing requires assisted recognition of its 3' splice site.

Calcitonin↗
PubMed1990

Transcription of the human calcitonin gene is mediated by a C cell-specific enhancer containing E-box-like elements.

The calcitonin (CT) gene is expressed normally in thyroidal C-cells and in a restricted population of cells in the central and peripheral nerve system. To define the cis-elements within the 5'-flanking DNA of the human CT gene which mediate this cell-specific expression, we used DNA transfer techniques and a transient transfection approach. We found that a DNA sequence located between -1290 and -820 of the CT 5'-flanking DNA functioned as an enhancer of basal transcription in C-cells (from medullary thyroid carcinoma) but not in rat glioma (C6), hamster insulinoma (HIT), fibroblasts (3T3), or epithelial cells (HeLa and CV1). Further mapping revealed the presence of at least two elements within the enhancer region; an upstream element (USE, located between -1060 and -1030) which could not function independently but its removal caused 70-80% loss of enhancer activity and a downstream element (DSE, located at -1033 to -920) which functioned independently as a cell-specific enhancer but with reduced activity. The binding pattern of nuclear proteins from C-cells to the enhancer elements was studied by an electrophoretic mobility shift assay. A protein-DNA complex was formed with the USE which could be competed, specifically, by an oligonucleotide containing the microE2 motif of the immunoglobulin gene enhancer. A similar complex was formed with the DSE fragment. Nuclear proteins from HeLa cells failed to form complexes with USE. Moreover, the binding pattern of proteins derived from HeLa cells to DSE was different from that of C-cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Changing concepts in the management of hereditary and sporadic medullary thyroid carcinoma.

Recent linkage of the gene for multiple endocrine neoplasia type 2A and 2B to the centromeric region of chromosome 10 has provided new insight into the causes of medullary thyroid carcinoma and has provided tools to diagnose gene carriers status for this syndrome with greater than 90% certainty. This review focuses on how these advances influence the clinical management of both sporadic and hereditary medullary thyroid carcinoma and discusses how tests based on the genetic linkage studies will aid in the early diagnosis and treatment of this syndrome. In addition, the authors have focused on several controversial management questions regarding the type and extent of surgery for this thyroid tumor, the management of the patient with metastatic disease, and the approach to management of other manifestations of multiple endocrine neoplasia types 2A and 2B. This review attempts to provide a balanced overview of these complex issues.

Adrenal Gland Neoplasms↗
PubMed1989

Multiple endocrine neoplasia type 2a associated with cutaneous lichen amyloidosis.

PURPOSE: To describe and characterize the association of hereditary cutaneous lichen amyloidosis with multiple endocrine neoplasia type 2a (MEN 2a). DESIGN: Survey of a family for two diseases. SETTING: Evaluation of patients at a clinical research center. PATIENTS: Nineteen family members with MEN 2a. MEASUREMENTS AND MAIN RESULTS: In this family cutaneous lichen amyloidosis presented as multiple infiltrated papules overlying a well-demarcated plaque in the scapular area of the back (right or left). Immunohistochemical studies showed amyloid that stained for keratin but not calcitonin. Three family members had the characteristic skin lesion and also carried the gene for MEN 2a; two additional members carried the gene for MEN 2a, but did not manifest the observable skin changes associated with lichen amyloidosis. CONCLUSIONS: From the findings in this kindred and in another recently reported but unrelated family with an identical type of pruritic skin rash and MEN 2a, several conclusions can be drawn. First, the syndrome of cutaneous amyloidosis and MEN 2a is a clearly defined autosomal dominant hereditary syndrome. Second, the dermal amyloid appears to be caused by deposition of keratin-like peptides rather than by calcitonin-like peptides. Third, known families with hereditary lichen amyloidosis should be screened to determine the true frequency of this syndrome.

Adrenal Gland Neoplasms↗
PubMed1989

1,25-Dihydroxyvitamin D3 silences 3',5'-cyclic adenosine monophosphate enhancement of somatostatin gene transcription in human thyroid C cells.

Treatment of the somatostatin (SRIF)-producing TT cell line with 1,25 dihydroxyvitamin D3 (1,25 D3) lowered intracellular SRIF mRNA and peptide concentration. In separate experiments, the cAMP analog 8-(4-chlorophenylthio)-cAMP stimulated a rapid increase in SRIF mRNA content of the TT cells and SRIF peptide secretion. To determine whether 1,25 D3 could inhibit either the transcriptional or secretory effects of cAMP, TT cells were pretreated with 1,25 D3 for 4 days followed by treatment with the cAMP analog. Pretreatment with 1,25 D3 inhibited the cAMP-mediated increase of SRIF mRNA, but had no effect on the secretory response. We conclude that the ability of 1,25 D3 to silence SRIF gene expression is more potent than cAMP enhancer activity but that 1,25 D3 has no effect on that portion of the cAMP-dependent pathway which regulates peptide secretion.

Calcitriol↗
PubMed1989

The clinical and screening age-at-onset distribution for the MEN-2 syndrome.

The decision to screen for multiple endocrine neoplasia type 2 (MEN-2) is generally based on family history, the rationale for this approach being the presumed 100% penetrance of the disease. To determine the validity of this presumption we have estimated--by applying modifications of the life-table method--the clinical and screening age-at-diagnosis distributions for MEN-2, using families from the Cancer Research Campaign Medullary Thyroid Cancer Register and one large American family. The clinical penetrance of MEN-2 is shown to be incomplete, an estimated 41% of gene carriers not presenting with symptoms by age 70 on the basis of clinical history. Screening by the standard tests for detecting the earliest manifestations of the syndrome increases the penetrance to an estimated 93% by age 31. There is no evidence of a difference in the age-at-diagnosis distributions between maternal and paternal transmission, or among different families, but there is some suggestion of an earlier onset of medullary thyroid cancer in female gene carriers, and of a tendency of pheochromocytoma to cluster in families. These results can be used to calculate risks to relatives of affected individuals, which in turn can be used to guide decisions on which individuals to screen.

Adrenal Gland Neoplasms↗
PubMed1989

Familial cutaneous lichen amyloidosis in association with multiple endocrine neoplasia type 2A: a new variant.

Multiple endocrine neoplasia type 2A (MEN 2A) is a rare hereditary disease transmitted in families as an autosomal dominant trait. We have identified a family in which the expression of a rare autosomal dominant form of cutaneous lichen amyloidosis appears to cosegregate with MEN 2A. In this family the skin lesion presented as multiple infiltrated papules overlying well demarcated plaques over the scapular area (right or left). Immunohistochemical studies demonstrated amyloid which stained for keratin but not calcitonin. A total of 19 members were screened. Three members of the family have the characteristic skin lesion and MEN 2A; two additional members have MEN 2A but have not manifested observable skin changes of lichen amyloidosis. Another unrelated Italian family with a similar type of pruritic skin rash and MEN 2A has been reported recently. Although the initial skin biopsies were negative for amyloidosis, subsequent biopsy established the association of MEN 2A with amyloidosis in this family also. When these kindreds are combined, several conclusions can be drawn. First, the syndrome of cutaneous amyloidosis and MEN 2A appears to be a clearly defined autosomal dominant hereditary syndrome. Whether this syndrome can be linked to chromosome 10 is not yet known. Second, the dermal amyloid appears to be caused by deposition of keratin-like peptides rather than calcitonin-like peptides. Third, we believe that patients with the hereditary form of cutaneous amyloid should be screened for medullary thyroid carcinoma to determine the true frequency of this syndrome.

Amyloidosis↗
PubMed1989

Transcriptional regulation of the human calcitonin gene: a progress report.

We have applied DNA transfer techniques to study the transcriptional regulation of the calcitonin (CT) gene in a C-cell line (TT) derived from a human medullary thyroid carcinoma. TT cells were transfected with a fusion gene containing the CT gene promoter and 5'-flanking DNA attached to the promoter-less growth hormone gene (reporter). We quantitated the reporter gene product to monitor transcriptional activation by the CT promoter and deletion mutants of the 5'-flanking DNA. We found that the proximal CT promoter which includes the DNA sequence from +1 to -129 bp upstream from the CT transcription start site did not induce transcription in C-cells or in NIH 3T3 cells. The attachment of additional 5'-flanking DNA, extending up to -1460 bp enhanced transcription up to twelvefold in TT cells but had no effect on transcription in 3T3 cells. Deletion of a sequence located at -1290 to -820 bp on the CT 5'-flanking DNA abolished the transcription of the reporter gene. Attachment of the DNA sequence located between -1333 to -731 to the fusion gene, containing the CT promoter (+1 to -129) and the reporter gene, restored transcription of the reporter gene in TT cells. We conclude that an enhancer of CT transcription, which is active in C-cells but not in 3T3 cells, is located between -1290 and -820 of the CT 5'-flanking DNA.

Calcitonin↗
PubMed1989

Calcitonin production: an examination of the regulatory effects of extracellular calcium and 1,25-dihydroxyvitamin D3.

Acute elevations of the serum calcium concentration stimulate release of preformed calcitonin. Whether chronic alterations of the serum calcium of 1,25 dihydroxyvitamin D3 concentrations cause an effect on synthesis or secretion of this major calcium-regulating hormone has not been determined with certainty. Recent studies, including those presented in this manuscript, indicate that 1,25 dihydroxyvitamin D3 and, perhaps, increases of the serum calcium concentration inhibit transcription of the calcitonin gene resulting in decreased production of calcitonin. Inhibition of calcitonin production in the hypercalcemic or vitamin D toxic patient may play a role in the pathophysiology of these conditions.

Calcitonin↗
PubMed1988

The clinical outcome of prospective screening for multiple endocrine neoplasia type 2a. An 18-year experience.

An important question facing physicians who care for families with multiple endocrine neoplasia type 2a is whether prospective screening to detect early abnormalities of the thyroid, parathyroid, or adrenal glands favorably influences the ultimate course of the disease. An 18-year study of a large family has allowed us to examine the effect of early treatment on the clinical course of the disease. Of 22 patients who underwent thyroidectomy for early C-cell abnormalities, 19 remained free of detectable medullary thyroid carcinoma according to all criteria, at a mean of 11 years after thyroidectomy. None of the 22 patients had evidence of parathyroid disease either at the time of surgery or after a mean follow-up of 10 years. Prospective screening for adrenal medullary abnormalities by means of measurement of 24-hour urinary epinephrine excretion and the ratio of urinary epinephrine to norepinephrine was predictive of pheochromocytoma in 10 of 11 patients (with a false negative result in one patient) but was not useful in diagnosing adrenal medullary hyperplasia. We conclude that regular, prospective screening and early treatment of the manifestations of multiple endocrine neoplasia can prevent metastasis of medullary thyroid carcinoma and the morbidity and mortality caused by pheochromocytoma.

Adrenal Gland Neoplasms↗