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Biomedical subjects

A Brunetti

Publications and source records attributed to A Brunetti.

At least 91 records · Page 5Linked to original sources
PubMed1989

Insulin receptor monoclonal antibodies that mimic insulin action without activating tyrosine kinase.

HTC rat hepatoma cells were transfected with human insulin receptor cDNA to a level of 40,000 receptors/cell. In these cells, as well as in nontransfected cells, insulin stimulated the uptake of alpha-aminoisobutyric acid. Two monoclonal antibodies directed against the human insulin receptor alpha subunit, like insulin, stimulated amino acid uptake in transfected HTC cells, but not in nontransfected HTC cells. The antibodies, in contrast to insulin, failed to stimulate insulin receptor tyrosine kinase activity, both in intact transfected cells and in cell free extracts prepared from them. These data suggest, therefore, that activation of insulin receptor tyrosine kinase may not be an obligatory step in all of the transmembrane signaling mechanisms of the insulin receptor.

Aminoisobutyric Acids↗
PubMed1989

Evaluation of a newly discovered water suppression pulse sequence for high-field in vivo 1H surface coil NMR spectroscopy.

The use of a water-suppressing spin-echo pulse sequence reported recently (V. Sklenar and A. Bax, J. Magn. Reson. 74, 469 (1987); M. von Kienlin, M. DeCorps, J. P. Albrand, M. F. Foray, and P. Blondet, J. Magn. Reson. 76, 169 (1987)) was evaluated for in vivo brain proton surface coil NMR spectroscopy. The studies were performed on cat brain using surface coils at 4.7 T. The sequence produced brain spectra with adequate water suppression, and a broader excitation profile than sequences which form spin echoes using 1331 pulses (P. J. Hore, J. Magn. Reson. 54, 539 (1983); H. P. Hetherington, M. J. Avison, and R. G. Shulman, Proc. Natl. Acad. Sci. USA 82, 3115 (1985)). The phase artifacts were smaller than those produced in 1331 methods, but theoretical analysis showed they should not be completely absent. The effectiveness of lengthening the spin-echo delay in the new sequence for suppression of unwanted lipid resonances was demonstrated. The sequence was shown to be capable of detecting lactate formation and clearance in a global cerebral ischemia experiment.

Animals↗
PubMed1989

On the origin of paramagnetic inhomogeneity effects in blood.

Hydrogen, sodium, and fluorine (added F-) NMR spectra of venous and oxygenated blood were measured. The fluorine resonance was seen as a single peak in both samples, and all three resonances exhibited the same deoxy-oxy shift. Because F- exchanges slowly across the red cell membrane, and because sodium is 95% extracellular, these results suggest that the intra-extracellular field difference delta B is less than 0.1 ppm. A small value of delta B tends to rule out transmembrane exchange as an important contributor to relaxation in MRI of blood and hematomas. However, the broadening of the resonances with deoxygenation, by 0.3-0.4 ppm, indicates that both intra- and extracellular gradients are of comparable and sufficient magnitude to produce the T2-weighted hypointensity seen in clinical magnetic resonance images of hematomas at high fields.

Blood↗
PubMed1989

Phase-distortion-free in vivo surface coil proton spectroscopy.

A 1-11 water suppression pulse sequence for phase-distortion-free surface coil NMR spectroscopy was constructed and tested in vivo on cat brain. The sequence produces brain spectra free of phase distortions, but some degradation of water suppression quality compared to closely related sequences which produce phase distortions was observed.

Animals↗
PubMed1989

Assessment of postischemic cerebral energy metabolism in cat by 31P NMR: the cumulative effects of secondary hypoxia and ischemia.

The sensitivity of cerebral energy metabolism to ischemic and hypoxic stresses following global cerebral ischemia was evaluated in a cat model using 31P nuclear magnetic resonance (NMR) spectroscopic methods. Complete global cerebral ischemia of 5 to 10 min in length was produced at 1 h intervals by reversible arterial occlusion, permitting continuous monitoring of NMR and EEG. Ischemia appeared to produce slightly more severe energy failure in animals that had previously experienced an ischemic injury. Preischemic hypoxia (5% O2 for 5 min) resulted in minor changes in the levels of phosphocreatine and intracellular inorganic phosphate, which were slightly amplified in animals that previously experienced ischemia.

Animals↗
PubMed1989

Human insulin receptor radioimmunoassay: applicability to insulin-resistant states.

A radioimmunoassay of the human insulin receptor was developed employing a potent rabbit polyclonal antibody to the human insulin receptor and a highly purified human placental insulin receptor preparation. The receptor, obtained by sequential affinity chromatography with insulin receptor monoclonal antibody-agarose and wheat germ agglutinin-agarose, was radiolabeled with 125I-Bolton-Hunter reagent at specific activities of 2,100-3,300 Ci/mmol. Over 75% of this ligand was immunoprecipitable with the polyclonal antireceptor antibody and remained immunoprecipitable for greater than 45 days. The assay was sensitive to unlabeled receptor concentrations as low as 0.2 ng/0.5 ml; unlabeled insulin did not cross-react and unlabeled insulin-like growth factor (IGF)-I receptor cross-reacted weakly. The radioimmunoassay was applicable to the measurement of insulin receptors in tissues and cells that were extracted by solubilization in 1% Triton X-100; no purification of the extracted receptor was necessary. Of the three major target tissues for insulin action studied, liver had the highest concentration of receptors (47.6 ng/mg protein); fat and muscle had lower levels. Other studies with the radioimmunoassay indicated that insulin receptors were decreased both in monocytes from obese hyperinsulinemic subjects and in fibroblasts from patients with leprechaunism.

Female↗
PubMed1989

Muscle cell differentiation is associated with increased insulin receptor biosynthesis and messenger RNA levels.

Muscle is a major tissue for insulin action. To study the effect of muscle differentiation on insulin receptors, we employed cultured mouse muscle BC3H-1 and C2 cells. In both cell lines differentiation from myoblasts to myocytes was associated with a 5-10-fold increase in specific 125I-insulin binding to intact cells. When 125I-insulin binding was carried out on solubilized myocytes and myoblasts, 125I-insulin binding to myoblasts was low. After differentiation the number of insulin receptors increased 5-10-fold. In contrast to insulin binding, insulin growth factor I receptor binding was elevated in myoblasts and was decreased by 50% in myocytes. Specific radioimmunoassay of the insulin receptor indicated that the increase in insulin binding to myocytes was due to an increase in insulin receptor content. Studies employing [35S]methionine indicated that this increase in insulin-binding sites reflected an increase in insulin receptor biosynthesis. To study insulin receptor gene expression, myoblast and myocyte mRNA was isolated and analyzed on Northern and slot blots. Differentiation from myoblasts to myocytes was accompanied by a 5-10-fold increase in insulin receptor mRNA. These studies demonstrate, therefore that differentiation in muscle cells is accompanied by increased insulin receptor biosynthesis and gene expression.

Animals↗
PubMed1989

Defects in insulin-receptor internalization and processing in monocytes of obese subjects and obese NIDDM patients.

We investigated intracellular processing of the insulin-receptor complex in monocytes from 12 healthy control subjects, 11 obese nondiabetic subjects, and 13 obese patients with non-insulin-dependent diabetes mellitus (NIDDM) by measuring receptor internalization, recovery of cell-surface insulin binding after receptor internalization, and the release of intracellular intact insulin (insulin retroendocytosis). When monocytes from the three groups of subjects were exposed to 100 nM unlabeled insulin for 30 min at 37 degrees C, the subsequent cell-surface 125I-labeled insulin binding was reduced, but the total number of insulin receptors, measured by radioimmunoassay, was not changed. These findings indicate a redistribution of insulin receptors from the surface to the cell interior. Insulin-receptor internalization was significantly lower in monocytes of obese NIDDM patients (mean +/- SE 17.8 +/- 4.7%) than in obese subjects and healthy control subjects (33.5 +/- 4.5%, P less than .05, and 34.4 +/- 3.7%, P less than .02, respectively). Moreover, in downregulated cells, a complete recovery of the initial insulin binding was observed in control subjects but not in obese NIDDM patients or obese nondiabetic subjects. The release of internalized insulin was also reduced in obese NIDDM patients and obese subjects (t 1/2 = 49.0 +/- 2.4 min, P less than .02; 47.4 +/- 5.7 min, P less than .05; and 32.9 +/- 3.8 in NIDDM patients, obese subjects, and control subjects, respectively). In the radioactivity released from monocytes of obese subjects and obese NIDDM patients, the percentage of intact insulin was higher (P less than .05) than in control subjects, suggesting reduced intracellular insulin degradation in obese subjects and obese NIDDM patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Diabetes Mellitus↗
PubMed1989

Renal artery stenosis detection by combined Gates' technique and captopril test in hypertensive patients.

We studied 11 hypertensive patients by a radionuclide technique using Gates' method with [99mTc]DTPA to investigate the acute effects of captopril on glomerular filtration rate (GFR). Five patients had hypertension with unilateral renal artery stenosis (RAS) angiographically documented and six patients had essential hypertension (EH). Total and split GFR were determined under control conditions and after oral administration of captopril (50 mg). In the patients with RAS, captopril induced a significant decrease of GFR in the stenotic kidneys (from 42.4 +/- 4 to 29.6 +/- 3 ml/min, p less than 0.01), while no changes were observed in the nonstenotic kidneys (from 61.2 +/- 3 to 61.6 +/- 5 ml/min, NS). Total GFR was 103.6 +/- 5 ml/min under control conditions and decreased to 91.8 +/- 6 ml/min after captopril (p less than 0.05). No significant changes of GFR were detected after captopril administration in patients with EH. In a separate group of ten patients with EH, good correlation between 24-hr creatinine clearance and fractional uptake of [99mTc]DTPA was obtained. Good reproducibility of this radionuclide technique was also shown. This study demonstrates that the computed radionuclide GFR determination coupled with the captopril test allows one to unmask angiotensin II-dependent renal function and hemodynamic changes. This technique can be useful in clinical practice for identifying patients with renovascular hypertension.

Adult↗
PubMed1989

Reversal of brain metabolic abnormalities following treatment of AIDS dementia complex with 3'-azido-2',3'-dideoxythymidine (AZT, zidovudine): a PET-FDG study.

Brain glucose metabolism was evaluated in four patients with acquired immunodeficiency syndrome (AIDS) dementia complex using [18F]fluorodeoxyglucose (FDG) and positron emission tomography (PET) scans at the beginning of therapy with 3'-azido-2',3'-dideoxythymidine (AZT, zidovudine), and later in the course of therapy. In two patients, baseline, large focal cortical abnormalities of glucose utilization were reversed during the course of therapy. In the other two patients, the initial PET study did not reveal pronounced focal alterations, while the post-treatment scans showed markedly increased cortical glucose metabolism. The improved cortical glucose utilization was accompanied in all patients by immunologic and neurologic improvement. PET-FDG studies can detect cortical metabolic abnormalities associated with AIDS dementia complex, and may be used to monitor the metabolic improvement in response to AZT treatment.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Comparison of iron-59, indium-111, and gallium-69 transferrin as a macromolecular tracer of vascular permeability and the transferrin receptor.

Tracer amounts of [59Fe++]citrate, [111In+++]chloride, and [68Ga+++]chloride were complexed with autologous plasma transferrin. Each of these complexes were co-administered with [125I]albumin by i.v. injection and their biodistribution was studied in Wistar rats. The plasma clearance of 59Fe and [125I]albumin was monoexponential with half-times of 49-70 and 277 min, respectively. The plasma clearance of 68Ga and 111In was biexponential with second component half-times of 157 and 232 min, respectively. Indium-111 tissue distribution was similar to that of [125I]albumin in heart, lung, muscle, brain and Walker-256 allograft. Iron-59 distribution spaces were generally the highest of the metal complexes in all tissues except muscle, where the 68Ga space was highest. The effects of transferrin-specific receptor-mediated endocytosis can be avoided in many organs and Walker-256 allografts by using the indium-transferrin complex, and the radiolabeled complex may be a convenient macromolecular tracer to estimate vascular permeability and vessel pore size in tumor and systemic tissue. In contrast, the iron-transferrin complex may be useful for measuring and imaging transferrin-specific receptors in brain and tumor tissue.

Animals↗
PubMed1988

Insulin degradation into monocytes from normal subjects: a high performance liquid chromatographic analysis.

In this work the fate of A14-125 I-insulin inside human cells has been investigated by the complementary use of gel permeation and reversed-phase high performance liquid chromatography to obtain a better resolution of the cell processed radioactive material resulting from the internalization of labeled insulin. Mononuclear leukocytes from 12 normals were incubated with pure A14-125 I insulin at 37 C and internalized radioactivity was characterized after 2, 15 and 60 min. Nearly 14% of intracellular radioactivity was associated to materials with a molecular weight of approximately 300,000. The remaining 86% had a molecular weight lower than 20,000. High molecular weight material showed an elution profile very similar to that obtained from purified human placental insulin receptor and was partially precipitable with antireceptor antibody. The reversed phase high performance liquid chromatography analysis of the low molecular weight material showed two main peaks corresponding to 125 I and A14-125 I-insulin and three intermediate peaks, a, b, c, accounting for about 8% of the recovered radioactivity. By increasing the incubation time of A14-125 I-insulin with monocytes a decrease of insulin peak (2 min: 38 +/- 18%; 15 min: 25 +/- 11%; 60 min: 6 +/- 4%) and a corresponding increase of iodide peak was observed. Immunoprecipitability with anti-insulin antibody was 0% for iodide and a peaks, 60% for peak b, 78% for peak c and 90% for A14-insulin peak. Our results show that intracellular insulin degradation procedes rapidly and in a time-dependent manner and that this process produces insulin derivatives which partially retain the immunological properties of intact A14-125 I insulin.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1988

Gallium-transferrin as a macromolecular tracer of vascular permeability.

Labeling of plasma transferrin with gallium was investigated to determine whether the gallium-transferrin complex could be effectively used as a macromolecular tracer in studies of capillary permeability using Positron Emission Tomography (PET). Three gallium-plasma preparations were tested and 2 h biodistribution studies were performed in rats. The three preparations gave similar blood clearance and tissue distribution data, but the methods used for evaluating gallium-transferrin binding were found to be suboptimal. Gallium clearance from blood was biexpoential with both components faster than that of 125I-albumin. Gallium distribution spaces in all tissues including intracerebral Walker-256 tumors were larger than those of albumin. These results indicate a relative instability of the gallium-transferrin complex in vivo, which appears to preclude its use as an acceptable radiolabeled protein for vascular permeability studies using PET.

Animals↗
PubMed1988

Metabolic imaging of human extremity musculoskeletal tumors by PET.

The measurement of glucose utilization rate (GUR) by positron emission tomography (PET) using 18F-2-fluoro-2-deoxy-D-glucose (FDG) is a valuable method to assess the grade of malignancy of brain tumors. We have designed a feasibility trial to determine whether PET could be used to image and predict the grade of malignancy of human extremity musculoskeletal tumors. Five patients with extremity tumors (four soft-tissue tumors and one osteogenic tumor) were studied. Peak and mean apparent GURs were determined in the tumor region. All tumors were subsequently resected and graded in a standard fashion using the NCI grading system. Peak apparent GURs ranged from 3.3 mg/100 g/min to 15.2 mg/100 g/min, with the highest values found in the high grade tumors. Although the number of patients studied was small, a good correspondence was shown between GURs and histopathologic grading. Our results indicate that PET can be used to image and evaluate the metabolic activity of human musculoskeletal tumors.

Adolescent↗
PubMed1987

Response of human-immunodeficiency-virus-associated neurological disease to 3'-azido-3'-deoxythymidine.

Four patients with human-immuno-deficiency-virus-associated neurological disease were treated with 3'-azido-3'-deoxythymidine (AZT). Three (two with chronic dementia, and one with chronic dementia and peripheral neuropathy) improved as assessed by clinical examination, psychometric tests, nerve conduction studies, and/or positron emission tomography; there was no improvement in the fourth patient who presented with paraplegia. These results support the hypothesis that certain AIDS-virus-associated neurological abnormalities are reversible by antiretroviral chemotherapy.

Acquired Immunodeficiency Syndrome↗
PubMed1987

Direct effects of biguanides on glucose utilization in vitro.

The effect of the biguanides metformin and phenformin on glucose utilization in isolated cells was studied with IM-9 human lymphocytes. Both agents stimulated glucose consumption from the incubation media. Detectable effects of metformin were seen at 33 mumol/L and detectable effects of phenformin were seen at 1.7 mumol/L. Both agents, at similar concentrations, also stimulated [3H] 2-deoxy-D-glucose uptake. Studies with phenformin indicated that biguanides increase the Vmax of uptake without changing the Km. In contrast to the biguanides, IM-9 cells insulin did not influence either glucose consumption or [3H] 2-deoxy-D-glucose uptake. These data provide evidence, therefore, that biguanides may directly influence the cellular utilization of glucose.

Blood Glucose↗
PubMed1987

Intracellular insulin processing is altered in monocytes from patients with type II diabetes mellitus.

We studied total cell-associated A14-[125I]insulin radioactivity (including surface-bound and internalized radioactivity), insulin internalization, and its intracellular degradation at 37 C in monocytes from nonobese type II untreated diabetic patients (n = 9) and normal subjects (n = 7). Total cell-associated radioactivity was decreased in diabetic patients [2.65 +/- 1.21% (+/- SD) vs. 4.47 +/- 1.04% of total radioactivity; P less than 0.01]. Insulin internalization was also reduced in diabetic patients (34.0 +/- 6.8% vs. 59.0 +/- 11.3% of cell-associated radioactivity; P less than 0.01). Using high performance liquid chromatography six intracellular forms of radioactivity derived from A14-[125I] insulin were identified; 10-20% of intracellular radioactivity had approximately 300,000 mol wt and was identified as radioactivity bound to the insulin receptor, and the remaining intracellular radioactivity included intact A14-[125I]insulin, [125I]iodide, or [125I]tyrosine, and three intermediate compounds. A progressive reduction of intact insulin and a corresponding increase in iodine were found when the incubation time was prolonged. Intracellular insulin degradation was reduced in monocytes from diabetic patients; intracellular intact insulin was 65.6 +/- 18.1% vs. 37.4 +/- 18.0% of intracellular radioactivity (P less than 0.01) after 2 min and 23.6 +/- 22.3% vs. 3.9 +/- 2.3% (P less than 0.01) after 60 min in diabetic patients vs. normal subjects, respectively. In conclusion, 1) human monocytes internalize and degrade insulin in the intracellular compartment in a stepwise time-dependent manner; and 2) in monocytes from type II diabetic patients total cell-associated radioactivity, insulin internalization, and insulin degradation are significantly reduced. These defects may be related to the cellular insulin resistance present in these patients.

Aged↗