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S Melmed

Publications and source records attributed to S Melmed.

At least 127 records · Page 7Linked to original sources
PubMed1994

Dominant negative inhibition of tumorigenesis in vivo by human insulin-like growth factor I receptor mutant.

Although insulin-like growth factor I (IGF-I) is a mitogenic growth factor, its role in tumorigenesis is unclear. We therefore transfected wild-type and truncated beta-subunit mutant (952STOP) human IGF-I receptor cDNAs into Rat-1 fibroblasts. Rat-1 transfectants expressed 2.5- to 7-fold increased IGF-I receptor mass, while the Kd for IGF-I binding was unchanged. The Rat-1 cells transfected with wild-type receptor cDNA responded to in vitro IGF-I treatment by increased proliferation and DNA synthesis. Cells overexpressing wild-type receptors were also transformed as evidenced by ligand-dependent colony proliferation in soft agar. After injection into athymic nude mice, all wild-type transfectants formed solid sarcomas within 3 weeks, and ex vivo tumor cell assays confirmed continued overexpression of human IGF-I receptors. In contrast, both DNA synthesis and proliferation of 952STOP-transfected cells were attenuated below that of untransfected cells. 952STOP cells were nonresponsive to IGF-I in vitro and were unable to sustain anchorage-independent growth. No tumors were induced for up to 8 weeks after injection of 952STOP transfectants into athymic mice, despite the presence of demonstrable endogenous IGF-I receptors on the 952STOP-transfected cells. Therefore, 952STOP behaves as a dominant negative inhibitor of endogenous IGF-I receptor function, probably by assembling nonfunctional hybrid rat/mutant human receptor tetramers.

Animals↗
PubMed1994

Heterogeneous expression of two somatostatin receptor subtypes in pituitary tumors.

The expression of two somatostatin receptor subtypes, SSTR1 and SSTR2, was studied in 27 pituitary tumors and 1 chronic lymphocytic leukemia pituitary infiltrate. Solution hybridization techniques were used for RNA analysis. SSTR1 and SSTR2 were, respectively, expressed in 3 of 7 and 9 of 10 GH-secreting tumors, 1 of 9 and 5 of 9 nonfunctioning tumors, 4 of 5 and 0 of 5 of prolactinomas, and 1 of 3 and 0 of 3 ACTH-secreting tumors. The chronic lymphocytic leukemia infiltrate expressed the SSTR2 subtype. No correlation was detected among tumor size, level of hormonal hypersecretion, and somatostatin receptor expression status. Two acromegalic patients who responded to octreotide therapy exclusively expressed the SSTR2 subtype in their tumors. The results indicate that two SSTR subtypes are heterogeneously expressed in different pituitary adenoma cell types.

Adenoma↗
PubMed1994

H-ras mutations in human pituitary carcinoma metastases.

Molecular mechanisms of pituitary tumorigenesis were studied using Polymerase chain reaction-single stranded conformational polymorphism with DNA sequencing to identify potential mutations in the ras protooncogenes and the tumor suppressor gene p53 in invasive pituitary adenomas and carcinomas. Sequencing of exons 5 through 8 of the p53 gene revealed no mutations, nor were mutations detected in the N- or K-ras protooncogenes in four of the carcinomas and their respective metastatic deposits. Point mutations of H-ras however, were identified in three distant metastatic pituitary tumor secondaries, but not in their respective primary pituitary carcinomas, or in six invasive adenomas. Two of the mutations included a G to C substitution at codon 12, and a G to A substitution at codon 18, resulting in a glycine to arginine, and an alanine to threonine change at these amino acids, respectively. A third mutation involved a single base pair (adenine) deletion in codon 3 of H-ras which causes a frame shift, resulting in a termination signal at codon 19. These results suggest that point mutations in p53 and ras are not associated with pituitary tumorigenesis, however, point mutations of the H-ras gene may be important in the formation and or growth of pituitary metastases. This observed genomic instability will be of value in predicting the potential metastatic behavior of these aggressive pituitary tumors.

Adenoma↗
PubMed1994

Expression of three somatostatin receptor subtypes in pituitary adenomas: evidence for preferential SSTR5 expression in the mammosomatotroph lineage.

The expression of three somatostatin receptor subtypes, SSTR3, SSTR4, and SSTR5, was evaluated in 33 pituitary tumor specimens. SSTR3 expression was studied by reverse transcription coupled to polymerase chain reaction, whereas SSTR4 and SSTR5 expression was determined by ribonuclease protection assay. SSTR3 was expressed in 6 of 7 GH-secreting tumors, all 8 clinically nonfunctioning tumors, all 3 prolactinomas, and 1 of 2 ACTH-secreting tumors tested. Eight nonfunctioning adenomas had undetectable messenger ribonucleic acid levels of SSTR4, and only 1 of them expressed SSTR5. SSTR4 expression was also undetectable in 11 GH-secreting tumors, 3 prolactinomas, and 1 ACTH-secreting tumor tested. In contrast, SSTR5 was highly expressed in 10 of 11 GH-secreting adenomas and 1 prolactinoma. Two prolactinomas and 1 ACTH-secreting tumor had low levels of expression of SSTR5. The widespread pituitary adenoma expression of SSTR3, regardless of hormonal secretory type, suggests that SSTR3 might be involved in a somatostatin action(s) other than GH or TSH regulation. SSTR5 is expressed predominantly in mammosomatotroph-derived tumors, suggesting that this receptor subtype may be an important determinant of GH secretion in acromegaly.

Adenoma↗
PubMed1994

Insulin-like growth factor-1 activation of extracellular signal-related kinase-1 and -2 in growth hormone-secreting cells.

Intracellular pathways mediating feedback regulation by insulin-like growth factor-1 (IGF-1) of pituitary GH gene expression remain incompletely understood. Extracellular signal-related kinases (ERKs), a family of serine/threonine kinases, are activated by tyrosine kinase-associated growth factor receptors. To further define the IGF-1 postreceptor events occurring in GH-secreting cells, we investigated the activity of ERKs in response to IGF-1 in GC cells following stable transfection with either wild type human IGF-1 receptor cDNA (WT cells) or a mutant cDNA encoding a truncated, kinase-defective IGF-1 receptor with a dominant negative effect on endogenous receptor function (952STOP cells). Zymography of immunoprecipitated ERKs in myelin basic protein (MBP)-containing polyacrylamide gels demonstrated dose-dependent induction of ERK-1 and -2 activity by IGF-1 in GC cells with maximal activity occurring at 6 min. IGF-1-induced ERK activity in WT-transfected cells was up to 80-fold basal and 4-fold that observed in GC cells. 952STOP cells expressing the tyrosine kinase-deficient receptor were refractory to IGF-1 action, demonstrating minimal ERK induction. In contrast, 12-O-tetradecanoylphorbol 13-acetate stimulated ERK activity to the same degree in all three cell types regardless of their IGF-I receptor status. Forskolin (50 microM), isobutylmethylxanthine (0.5 mM), and forskolin/isobutylmethylxanthine in combination attenuated IGF-1-induced ERK activity in WT cells by 54, 55, and 75% respectively. The rapid, dose-dependent, and IGF-1 receptor-dependent activation of ERKs and the attenuation of this effect by cAMP suggest an interrelated role for both molecules in IGF-1 signal transduction in GH-secreting cells.

Animals↗
PubMed1994

Pituitary neoplasia.

Anterior pituitary tumors may secrete trophic hormones with resultant peripheral clinical disorders, or they may be clinically silent. Hypothalamic hormones control anterior pituitary hormone secretion and cell proliferation and also have been implicated in tumor promotion in animal models. Pituitary tumors are, however, monoclonal in origin. Several activating oncogene mutations as well as allelic loss of tumor suppressor genes have been described. Interestingly, apparently sporadic pituitary tumors appear to exhibit genetic defects similar to those for MEN 1-associated tumors. The multistep process of pituitary tumorigenesis is becoming increasingly apparent, and its understanding will provide novel prognostic and therapeutic options for these patients.

Adenoma↗
PubMed1993

Recurrent acromegaly resulting from ectopic growth hormone gene expression by a metastatic pancreatic tumor.

BACKGROUND: Acromegaly is usually the result of a pituitary growth hormone (GH)-cell adenoma or is more rarely due to ectopic secretion of GH-releasing hormone (GHRH). The authors previously described a more unusual form of acromegaly secondary to ectopic GH synthesis by a pancreatic islet cell tumor. METHODS: One year after tumor resection and transient disease remission, multiple abdominal metastases were identified with accompanying elevated levels of circulating GH and insulin-like growth factor-1 (IGF-1). Serial 24-hour GH sampling was performed before and after intravenous GHRH or thyrotropin releasing hormone (TRH) administration during treatment with bromocriptine; treatment with the somatostatin (SRIF) analogue octreotide; or no treatment. RNA from abdominal tumor tissue was extracted and subjected to Northern gel electrophoresis and GH hybridization analysis. RESULTS: Neither GHRH nor TRH resulted in stimulation of the elevated GH levels. Bromocriptine and octreotide did not suppress GH secretion but attenuated the thyroid stimulating hormone (TSH) response to TRH administration. Octreotide (as much as 1500 micrograms/d) was clinically, biochemically, and radiographically ineffective. GH-secreting abdominal tumor tissue expressed a 0.9-kb mRNA transcript consistent with the size of authentic human GH mRNA. CONCLUSION: The natural history and ectopic nature of a GH-producing pancreatic carcinoma has been documented, with biochemical remission occurring after initial tumor resection, with autonomous GH hypersecretion after tumor recurrence, and with RNA analysis demonstrating ectopic activation of GH gene transcription.

Acromegaly↗
PubMed1993

Recombinant growth hormone treatment of amyotrophic lateral sclerosis.

Based on the known trophic effects of growth hormone (GH) on nerve and muscle 75 patients with ALS were treated for up to 18 months with synthetic human growth hormone (Protropin) or a placebo. The course of ALS was assessed serially using a quantitative (TQNE) neuromuscular and manual exam (MRC) and laboratory chemistries. Average insulin-related growth factor (IGF-I) values increased from 1.2 to 2.3 U/mL in the treated group. Surprisingly, serum insulin levels did not increase. Hyperglycemia was noted in only 2 patients of the 38 patients receiving hGH, and this resolved with cessation of treatment. Over the 12 months of treatment there were 11 deaths (6 controls, 5 treated). Survival analysis, performed approximately 12 months following cessation of treatment, did not reveal a difference between the treatment and placebo group. The TQNE scores declined inexorably in both the control and treated group. Retrospective analysis of the TQNE data indicated a poor prognosis for patients who lost arm strength early. A correlation between the TQNE and MRC scores was evident at early stages of motor unit loss, less so when muscle weakness was advanced.

Adult↗
PubMed1993

Rat pituitary GC cell insulin-like growth factor-I receptor regulation.

Exposure of rat GC pituitary cells to insulin-like growth factor-I (IGF-I) resulted in a time- and concentration-dependent reduction in cell surface IGF-I receptor binding. Preincubation of cells with 6.5 nM IGF-I for 24 h caused an approximately 50% reduction of the number of cell surface receptors. Scatchard analysis revealed that the number of IGF-I cell surface receptors decreased from 18,500 to 9,100 sites/cell, with no apparent alteration in the affinity of IGF-I receptor for its ligand. The level of IGF-I receptor mRNA, however, was not altered by preincubating cells for up to 24 h with 6.5 nM IGF-I. The sequestered IGF-I receptors reappeared on the cell membrane and return to their preincubation surface distribution within 3 h after removal of IGF-I. These results support the notion that down-regulation of somatotroph IGF-I receptors involves translocation of cell surface receptors to the intracellular pool, and this effect is reversed by removal of IGF-I ligand.

Animals↗
PubMed1993

Biochemical assessment of bone formation and resorption in acromegaly.

The effects of chronic GH and insulin-like growth factor-I (IGF-I) excess on bone metabolism were examined by measuring serum markers of bone formation and urine markers of bone resorption as well as vertebral bone densities in patients with active acromegaly. Fasting serum GH levels were elevated in all 27 patients (31 +/- 11 micrograms/L). Serum calcium levels were within the normal range, except in 3 of 27 (10%) patients with mild hypercalcemia. Urinary calcium excretion, however, was increased in 6 (22%) patients despite mainly normal serum PTH and 1,25-dihydroxyvitamin D levels, suggesting a direct renal GH and/or IGF-I-mediated calciuric effect. Urinary hydroxyproline/creatinine excretion was increased in all except 1 patient and correlated with plasma IGF-I levels (r = 0.49; P < 0.02; n = 22). A more specific indicator of bone collagen turnover, urinary type I collagen cross-linked N-telopeptide, was elevated in all except 1 patient and correlated with serum GH (r = 0.47; P < 0.02), IGF-I (r = 0.60; P < 0.005), and urinary hydroxyproline/creatinine excretion (r = 0.62; P < 0.001). Serum bone Gla protein (osteocalcin), a specific marker of osteoblastic activity, was also increased in 50% of the patients and correlated with urinary N-telopeptide (r = 0.47; P < 0.02), but not with serum GH or IGF-I concentrations. Trabecular bone density, as determined by quantitative computerized tomography of the lumbar spine, was increased in only 1 patient; 13 others had subnormal bone density. The results suggest that in long-standing acromegaly, osteoblastic and osteoclastic activities are increased. Vertebral trabecular bone mass is usually reduced. Urinary collagen cross-links may serve as a more specific marker of bone resorption in acromegaly.

Acromegaly↗
PubMed1993

Molecular screening of pituitary adenomas for gene mutations and rearrangements.

Although pituitary tumors arise as benign monoclonal neoplasms, genetic alterations have not readily been identified in these adenomas. We studied restriction fragment abnormalities involving the GH gene locus, and mutations in the p53 and H-, K- and N-ras genes in 22 human GH cell adenomas. Twenty two nonsecretory adenomas were also examined for p53 and ras gene mutations. Seven prolactinoma DNA samples were tested for deletions in the multiple endocrine neoplasia-1 (MEN-1) locus, as well as for rearrangements in the hst gene, a member of the fibroblast growth factor family. Pituitary adenoma tissue and lymphocytes were obtained from patients at the time of transsphenoidal surgery. In DNA from GH-cell adenomas, identical GH restriction patterns were detected in both pituitary and lymphocyte DNA in all patients and in one patient with a mixed GH-TSH cell adenoma. Using polymerase chain reaction (PCR)-single stranded conformation polymorphism analysis, no mutations were detected in exons 5, 6, 7, and 8 of the p53 gene in GH cell adenomas nor in 22 nonsecretory adenomas. Codons 12/13 and 61 of H-ras, K-ras, and N-ras genes were also intact in GH cell adenomas and in nonsecretory adenomas. Site-specific probes for chromosome 11q13 including PYGM, D11S146, and INT2 were used in 7 sporadic PRL-secreting adenomas to detect deletions of the MEN-1 locus on chromosome 11. One patient was identified with a loss of 11p, and the remaining 6 patients did not demonstrate loss of heterozygosity in the pituitary 11q13 locus, compared to lymphocyte DNA. None of these patients, demonstrated hst gene rearrangements which also maps to this locus. These results show that p53 and ras gene mutations are not common events in the pathogenesis of acromegaly and nonsecretory tumors. Although hst gene rearrangements and deletions of 11q13 are not associated with sporadic PRL-cell adenoma formation, a single patient was detected with a partial loss of chromosome 11, including the putative MEN-1 site.

Adenoma↗
PubMed1993

Structure-function of the human insulin-like growth factor-I receptor: a discordance of somatotroph internalization and signaling.

Insulin-like growth factor-I (IGF-I), a GH-dependent growth factor, suppresses GH secretion by pituitary cells. To clarify the role of ligand-mediated receptor internalization for IGF-I signaling to GH, human IGF-I receptor (IGF-IR) cDNAs mutated in the beta-subunit were stably transfected into GC rat pituitary cells. Overexpression of wild-type IGF-IR markedly enhanced IGF-I suppression of GH 4-fold (P < 0.005) compared to that of untransfected cells. A mutant IGF-IR with a 943Tyr-->Ala substitution in the IGF-IR submembrane domain only partially suppressed GH (73% of IGF-IR wild type), while replacement of 957Tyr-->Ala or 940Gly-->Ala produced IGF-IRs that retained enhanced IGF-I signaling to GH. Substitution of 950Tyr-->Ala or 1003Lys-->Ala in the human IGF-IR beta-subunit failed to enhance IGF-I signaling to GH above that of untransfected cells. Intracellular phosphorylation of insulin-responsive substrate-I by these mutant IGF-IRs paralleled the observed IGF-I suppression of GH, with no phosphorylation of IRS-I by 950Tyr-->Ala. Ligand-mediated receptor internalization, however, was not reduced by substitution of either 943Tyr-->Ala or 950Tyr-->Ala. In contrast, substitution of 957Tyr-->Ala reduced the internalization of labeled IGF-I to 35% that of wild-type IGF-IR. Substitution of 1003Lys-->Ala abolished IGF-IR internalization, as expected. These results demonstrate that both 950Tyr and 943Tyr are important for IGF-I signaling to GH and that IGF-IR internalization is discordant for IGF-I signaling to the GH gene.

Animals↗
PubMed1993

Interaction between insulin and thyroid hormone in rat pituitary tumour cells: insulin attenuates tri-iodothyronine-induced growth hormone mRNA levels.

Insulin has previously been shown to inhibit basal and stimulated rat GH (rGH) secretion as well as basal GH transcription in rat pituitary cells. The effect of physiological doses of insulin on tri-iodothyronine (T3)-stimulated GH mRNA levels in rat pituitary tumour cells was therefore examined. Insulin (7 nmol/l) suppressed T3-stimulated GH mRNA levels in GC and GH3 rat pituitary tumour cells by 58%. This inhibitory effect of insulin on T3-stimulated GH mRNA levels was already present after 24 h of treatment, and persisted for at least 48 h after insulin treatment was withdrawn. The effect of insulin on GH mRNA was selective, as rat prolactin mRNA was stimulated by insulin and T3 in the same cells. Treatment of cells with cycloheximide (10 mumol/l) did not alter the attenuation of GH mRNA levels by insulin, indicating that the insulin effect is independent of new protein synthesis. When de-novo mRNA synthesis was blocked with actinomycin D (4 micrograms/ml) for up to 7 h, an additional decrease in the relative amount of GH mRNA levels was observed after 24, 48 and 72 h of insulin treatment, indicating that an effect of insulin on GH mRNA stability is likely. The results show that physiological doses of insulin selectively attenuate the stimulatory effect of T3 on GH mRNA levels. This suppressive effect of insulin occurs independently of protein synthesis and is presumably mediated both at a transcriptional and post-transcriptional level.

Animals↗
PubMed1993

Medical management of acromegaly--what and when?

GH and IGF-I levels are elevated in patients with acromegaly. At the time of diagnosis patients present with macroadenomas with a high surgical failure rate or microadenomas. Administration of octreotide (100 micrograms, three times daily) suppressed GH secretion during 8 h to < 5 micrograms/l in 47% and to < 2 micrograms/l in 26% of acromegalic patients after two weeks. IGF-I levels were normalized in up to 70% of patients. Increasing the dose of octreotide to 250 micrograms three times daily did not further improve results. Tumour shrinkage occurred in 37% after six months of treatment, while symptoms improved in 70%. Transient diarrhoea and nausea were noted in 88%, but after six months only 10% reported these symptoms. Gallbladder sludge and gallstones were noted in 19% of patients. IGF-I levels were normalized in 82% of patients with microadenoma and in 50% with macroadenoma. Well-defined pituitary adenomas are usually surgically removed. Invasive tumours are difficult to remove surgically but preoperative octreotide may shrink adenomas and improve response. Octreotide therapy, unlike surgery and irradiation, does not compromise pituitary function. This study suggests that octreotide therapy could be a viable primary management of small discreet adenomas. Where surgery and octreotide fail, other treatments of adenoma include bromocriptine and radiotherapy.

Acromegaly↗