Biomedical subjects

S E Stred

Publications and source records attributed to S E Stred.

9 recordsLinked to original sources

PubMed1996

Interstitial 6q deletion and Prader-Willi-like phenotype.

A third case of an interstitial deletion of the long arm of chromosome 6 with clinical features mimicking Prader-Willi syndrome (PWS) is presented. Although preliminary clinical evaluation in each case suggested PWS, further review revealed that the features in all three cases are not completely compatible with the characteristic findings in Prader-Willi syndrome. Furthermore, the deletions in the three cases do not show a consistent region of overlap. Consequently, no particular band or region in 6q can be defined as associated with obesity. However, our findings confirm the suggestion of Villa et al. in 1995, that individuals with a PWS phenotype who are cytogenetically and molecularly negative for a deletion of 15q11-q13 should be examined for a deletion of 6q.

Child↗

PubMed1992

Stimulation by growth hormone (GH) of GH receptor-associated tyrosine kinase activity.

We have shown previously that GH receptors (GHRs) become phosphorylated on tyrosyl residues when GH-responsive cells are exposed to GH. In this work we investigate the molecular mechanism by which GH binding stimulates tyrosyl phosphorylation of GHR. To test whether in the presence of GH, GHR and the tyrosine kinase responsible for GHR phosphorylation are tightly associated in a complex or form a transient enzyme-substrate complex, the rate of in vitro phosphorylation of GHR was determined as a function of receptor concentration. GH-GHR complexes were purified from 3T3-F442A fibroblasts by immunoadsorption to and elution from immobilized phosphotyrosyl-binding antibody. The rate of in vitro tyrosyl phosphorylation of GH-GHR complexes was not substantially reduced when the GHR preparation was diluted 1:10 or 1:100 before phosphorylation, consistent with a tight association between kinase and substrate (GHR). When cells were labeled metabolically with 35S, and GHRs containing phosphorylated tyrosyl residues were isolated by immunoadsorption to and elution from phosphotyrosyl-binding antibody, the ability to immunoprecipitate 35S-labeled GHR with GHR antibodies was only evident when the cells had been incubated with GH. This indicates that in vivo, GHRs are phosphorylated on tyrosyl residues only when GH is bound. Additionally, when anti-GHR antibodies were used to immunoprecipitate GHR from solubilized cells, only GHRs isolated from GH-treated cells were phosphorylated when subjected to an in vitro kinase assay. The increased tyrosyl phosphorylation of GHR detected after incubation of cells with GH is consistent either with GH increasing tyrosine kinase activity associated with the GHR or with GH inducing a conformational change in GHR that renders it susceptible to tyrosyl phosphorylation. To discern whether GH binding increased GHR-associated kinase activity, we tested the ability of anti-GHR antibody immunoprecipitates to phosphorylate the synthetic substrate poly(Glu4,Tyr). GH treatment of cells resulted in a 2-fold increase in the rate of phosphorylation of poly(Glu4,Tyr). The increase in kinase activity was dose dependent, with half-maximal stimulation between 15-20 ng/ml GH. These results provide strong evidence that GH actually increases tyrosine kinase activity associated with the GHR. This would be consistent with GH-dependent complex formation between a constitutively activated kinase and GHR and/or activation of a constitutively associated or intrinsic kinase.

Animals↗

PubMed1991

Antibodies to cytoplasmic sequences of cloned liver growth hormone (GH) receptors recognize GH receptors associated with tyrosine kinase activity.

GH stimulates tyrosyl phosphorylation of GH receptors in 3T3-F442A fibroblasts, and highly purified GH receptor preparations exhibit tyrosine kinase activity. Paradoxically, however, the GH receptor cloned from liver exhibits no sequence similarity to receptors with known signal transduction mechanisms, including those exhibiting ligand-activated tyrosine kinase activity. These observations raise the possibility that there are two kinds of receptors for GH: the first represented by the cloned liver GH receptor, and the second by a tyrosine kinase-containing GH receptor. To inquire into the possibility of two distinct GH receptors, we determined whether the cloned liver GH receptor shares structural similarities with the tyrosine kinase-associated GH receptor. When the cloned rabbit liver GH receptor is expressed in human kidney 293 cells, it migrates with a mol wt appropriate for the tyrosine kinase-associated GH receptor, despite the calculated mol wt of the cloned GH receptor being 60,000 smaller than that of the tyrosine kinase-associated GH receptor. The recognition of tyrosine kinase-associated GH receptor by antipeptide antibodies to three different epitopes on the cytoplasmic domain of the cloned liver GH receptor was also tested. Tyrosyl phosphorylated [125I]human GH-receptor complexes were prepared by immunoprecipitation with phosphotyrosyl-binding antibody; this subpopulation of GH-receptor complexes was recognized by all three antipeptide antibodies. The antibodies also recognized similarly isolated tyrosyl phosphorylated GH-receptor complexes, which had been further phosphorylated in solution on tyrosyl residues upon addition of [gamma 32P] ATP. Furthermore, highly purified GH receptors prepared by sequential immunoprecipitation using phosphotyrosyl-binding antibody and any one of the three antipeptide antibodies incorporated 32P into tyrosyl residues upon the addition of [gamma 32P] ATP. These results provide evidence that tyrosine kinase-associated GH receptors share sequence similarity in the cytoplasmic domain with the cloned liver GH receptor. The cloned GH receptor and the tyrosine kinase-associated GH receptor, therefore, are likely to be the same receptor or related receptor isoforms.

Adipose Tissue↗

PubMed1990

Demonstration of growth hormone (GH) receptor-associated tyrosine kinase activity in multiple GH-responsive cell types.

Highly purified GH-receptor preparations from 3T3-F442A fibroblasts, whose differentiation into adipocytes is promoted by GH, have been shown to contain a tyrosine kinase capable of phosphorylating GH receptors. In the current work, characteristics of the tyrosine kinase responsible for the in vitro phosphorylation of GH receptors from cultured 3T3-F442A fibroblasts were examined, and the presence of this GH receptor-associated tyrosine kinase activity was demonstrated in multiple cell types. GH-receptor complexes from GH-treated cells were partially purified by immunoprecipitation using anti-GH antibodies and then incubated as an immune complex with [gamma 32P] ATP. Incorporation of 32P into the GH receptor from 3T3-F442A fibroblasts was apparent within 1 min at 30 C after the addition of [gamma 32P]ATP (5-10 microM). A divalent cation was requisite for the phosphorylation; Mn2+ was significantly more effective than Mg2+ and Co2+; Ba2+, Ca2+, or Zn2+ had no effect. Excess unlabeled ATP, but not cytosine triphosphate, GTP, or uridine triphosphate, abolished 32P incorporation into the GH receptor and [gamma 32P]GTP could not replace [gamma 32P]ATP as a source of 32P. At 5.5 mM Mn2+, phosphorylation exhibited a biphasic dose response to ATP, with maximal phosphorylation occurring at a concentration of 10 microM ATP. At more physiological concentrations of ATP (1 mM), phosphorylation of the GH receptor was also stimulated by lower concentrations of Mn2+ (as low as 500 nM). Optimal reaction conditions determined for the phosphorylation reaction in 3T3-F442A fibroblasts were used to demonstrate incorporation of 32P from [gamma 32P]ATP into partially purified GH receptors from cultured human IM-9 lymphocytes, murine 3T3-F442A adipocytes, rat H-35 hepatoma cells, and freshly isolated rat adipocytes. The 32P was shown to be incorporated into tyrosyl residues in receptors from the two cell types tested (IM-9 lymphocytes and rat adipocytes). Cross-linked [125I] hGH-receptor complexes solubilized from the four cell types (IM-9 lymphocytes, 3T3-F442A adipocytes, H-35 hepatoma cells, and freshly isolated rat adipocytes) bound to and could be eluted from phosphotyrosyl antibody, suggesting that tyrosyl phosphorylation of GH receptors in all of these cells occurs in vivo. The presence of tyrosine kinase activity associated with GH receptors in multiple cell types from different species is consistent with tyrosine kinase activity playing a role in the actions of GH.

Adenosine Triphosphate↗

PubMed1989

Phosphorylation of highly purified growth hormone receptors by a growth hormone receptor-associated tyrosine kinase.

We have shown previously that growth hormone (GH) promotes the phosphorylation of its receptor on tyrosyl residues (Foster, C. M., Shafer, J. A., Rozsa, F. W., Wang, X., Lewis, S. D., Renken, D. A., Natale, J. E., Schwartz, J., and Carter-Su, C. (1988) Biochemistry 27, 326-334). In the present study, we investigated the possibility that a tyrosine kinase is specifically associated with the GH receptor. GH-receptor complexes were first partially purified from GH-treated 3T3-F442A fibroblasts, a GH-responsive cell, by immunoprecipitation using anti-GH antiserum. 35S-Labeled proteins of Mr = 105,000-125,000 were observed in the immunoprecipitate from GH-treated cells labeled metabolically with 35S-amino-acids. These proteins were not observed in immunoprecipitates from cells not exposed to GH or when non-immune serum replaced the anti-GH antiserum, consistent with the proteins being GH receptors. GH receptors appeared to be phosphorylated, as evidenced by the presence of 32P-labeled bands, comigrating with the 105-125 kDa 35S-labeled proteins, in the immunoprecipitate of GH-treated cells labeled metabolically with [32P]Pi. When partially purified GH receptor preparation was incubated with [gamma-32P]ATP (7-15 microM) for 10 min at 30 degrees C in the presence of MnCl2, a protein of Mr = 121,000 was phosphorylated exclusively on tyrosyl residues. As expected for the GH receptor, this protein was not observed in immunoprecipitates when cells had not been treated with GH nor when non-immune serum replaced the anti-GH antiserum. GH-receptor complexes were also purified to near homogeneity by sequential immunoprecipitation with phosphotyrosyl-binding antibody followed by anti-GH antiserum. When cells were labeled metabolically with 35S-amino acids, the 35S label migrated almost exclusively as an Mr = 105,000-125,000 protein. This protein also incorporated 32P into tyrosyl residues when incubated in solution with [gamma-32P]ATP. These results show that highly purified GH receptor preparations undergo tyrosyl phosphorylation, suggesting that either the GH receptor itself is a tyrosine kinase or is tightly associated with a tyrosine kinase.

Adenosine Triphosphate↗

PubMed1989

Increased growth hormone pulse frequency in acromegaly.

To investigate whether GH secretion in acromegaly is subject to regulatory control by the hypothalamic GH-releasing hormone (GHRH) we studied GH secretion in 22 patients with acromegaly. Parameters of pulsatile GH secretion were assessed using frequent blood sampling (every 20 or 10 min for 24 h). Acute GH responses to GHRH-44 (0.1, 0.33, and 1.0 micrograms/kg BW, iv) were measured, and GH secretion during therapy with the long-acting somatostatin analog SMS 201-995 (Sandoz) was assessed. The results were compared to those in normal volunteers. Spontaneous GH pulse frequency was greater in patients with acromegaly than in 6 control subjects (8.6 +/- 0.6 vs. 4.3 +/- 1.1 pulses/24 h), as estimated by the 20-min sampling frequency. The 10-min sampling frequency revealed 12.9 +/- 0.7 pulses/24 h in acromegalics. Spontaneous GH pulse amplitude and acute GH rises in response to GHRH did not differ between control and acromegalic subjects. A similar degree of nocturnal augmentation of GH secretion was observed in both groups, and it persisted during SMS 201-995 therapy in patients with acromegaly. These observations suggest that GH secretion in acromegaly remains under stimulatory control by GHRH, which may be released at an abnormally high rate.

Acromegaly↗

PubMed1987

Effect of growth hormone on growth and glucose tolerance of normal rats.

To determine the effect of growth hormone (GH) administration on growth rate, ultimate size, and glucose tolerance of intact rats, we administered ovine GH to male Sprague-Dawley weanling rats for six weeks. Growth hormone, in a dose sufficient to double the growth rate and increase sixfold the somatomedin C (SM-C) levels of hypophysectomized rats, failed to increase the rate of growth or size of visceral organs of intact rats. Their SM-C levels increased only 10%. Following glucose administration, the mean blood glucose levels were higher at every time point measured in those treated with GH compared with the intact control animals. We conclude that, although there was no difference in somatic or visceral growth, the dose of exogenous GH administered increased SM-C levels and decreased glucose tolerance in the intact rats. While a larger GH dose might increase the final body size of non-GH-deficient animals, impaired glucose tolerance could be a significant side effect.

Animals↗

PubMed1986

Coarctation of the midaortic arch presenting with monoparesis.

A 6 1/2-month-old infant was seen with right-arm monoparesis, right upper extremity hypertension, and findings of a coarctation located in the midaortic arch between the innominate and left common carotid arteries. Associated left cerebral atrophy (Dyke-Davidoff syndrome) was documented by computed tomography. Excision of the coarctation segment with primary repair of the arch and reanastomosis of the left carotid to the innominate artery was accomplished.

Aorta, Thoracic↗

PubMed1984

Regulation of glutamine synthetase and glutaminase activities in cultured skeletal muscle cells.

Glutamine is synthesized in skeletal muscle, released to the circulation, and transported to other tissues, where it may provide important substrate for gluconeogenesis, ammoniagenesis, and energy-yielding pathways. With the ultimate goal of delineating the factors that control glutamine production and release by skeletal muscle, we have studied the regulation of two key enzymes, glutamine synthetase and glutaminase, in the L6 line of rat skeletal muscle cells grown in monolayer culture. The cultured myotubes were found to have glutamine synthetase and phosphate-dependent glutaminase activities. Glutamine synthetase activity was increased following incubation (1) in glutamine-free medium (threefold); (2) in medium containing high glutamic acid concentrations (fourfold); and (3) in medium supplemented with dexamethasone (threefold). In each case the increase in glutamine synthetase activity required several hours to reach a maximum and was prevented by cycloheximide, suggesting that the change occurred through increased enzyme biosynthesis. No substances tested were found to affect glutaminase activity. We conclude that glutamine synthetase in cultured skeletal muscle is responsive to substrate, product, and hormonal regulation.

Animals↗