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Biomedical subjects

P Nowotny

Publications and source records attributed to P Nowotny.

At least 37 records · Page 2Linked to original sources
PubMed2001

Lipid-dependent control of hepatic glycogen stores in healthy humans.

AIMS/HYPOTHESIS: Non-esterified fatty acids and glycerol could stimulate gluconeogenesis and also contribute to regulating hepatic glycogen stores. We examined their effect on liver glycogen breakdown in humans. METHODS: After an overnight fast healthy subjects participated in three protocols with lipid/heparin (plasma non-esterified fatty acids: 2.2 +/- 0.1 mol/l; plasma glycerol: 0.5 +/- 0.03 mol/l; n = 7), glycerol (0.4 +/- 0.1 mol/l; 1.5 +/- 0.2 mol/l; n = 5) and saline infusion (control; 0.5 +/- 0.1 mol/l; 0.2 +/- 0.02 mol/l; n = 7). Net rates of glycogen breakdown were calculated from the decrease of liver glycogen within 9 h using 13C nuclear magnetic resonance spectroscopy. Endogenous glucose production was measured with infusion of D-[6,6-2H2]glucose. RESULTS: Endogenous glucose production decreased by about 25 % during lipid and saline infusion (p < 0.005) but not during glycerol infusion (p < 0.001 vs lipid, saline). An increase of plasma non-esterified fatty acids or glycerol reduced the net glycogen breakdown by about 84 % to 0.6 +/- 0.3 micromol x kg(-1) x min(-1) (p < 0.001 vs saline: 3.7 +/- 0.5 micromol x kg(-1) x min(-1)) and by about 46 % to 2.0 +/- 0.4 micromol x kg(-1) x min(-1) (p < 0.01 vs saline and lipid), respectively. Rates of gluconeogenesis increased to 11.5 +/- 0.8 micromol x kg(-1) x min(-1) (p < 0.01) and 12.8 +/- 1.0 micromol x kg(-1) min(-1) (p < 0.01 vs saline: 8.2 +/- 0.7 micromol x l(-1) x min(-1)), respectively. CONCLUSION/INTERPRETATION: An increase of non-esterifled fatty acid leads to a pronounced inhibition of net hepatic glycogen breakdown and increases gluconeogenesis whereas glucose production does not differ from the control condition. We suggest that this effect is not due to increased availability of glycerol alone but rather to lipid-dependent control of hepatic glycogen stores.

Adult↗
PubMed2001

SNP analysis to dissect human traits.

The analysis of complex human diseases has been spurred by the number of published genomic sequence variants - many identified in the course of sequencing the human genome. But, to be useful for genetic analysis, variants have to be mapped accurately, their frequencies in various populations determined, and automated high-throughput assay techniques developed. Recently proposed methods address these issues: the use of 'reduced representation shotgun' methods for more efficient detection of single nucleotide polymorphisms (SNPs), the employment of high-throughput genotyping techniques, the development of SNP maps that incorporate information about linkage disequilibrium, and the use of SNPs in identifying susceptibility genes for common illnesses.

Chromosome Mapping↗
PubMed2001

Mutational spectrum of the steroid 21-hydroxylase gene in Austria: identification of a novel missense mutation.

This study attempted an analysis of the mutational spectrum of 21-hydroxylase deficiency in 79 unrelated Austrian patients with classical and nonclassical forms of congenital adrenal hyperplasia and their respective 112 family members. Apparent large gene deletions/conversions were present in 31% of the 158 unrelated congenital adrenal hyperplasia alleles, whereas the most frequent point mutations were intron 2 splice (22.8%), I172N (15.8%), V281L (12%), and P30L (7.6%), in line with the frequencies reported for other countries. In 5 of the 12 congenital adrenal hyperplasia alleles carrying a P30L mutation the aberration is based on a single base substitution, whereas the remaining 7 represent part of a CYP21B conversion (1 allele) or CYP21B/21A hybrid gene (6 alleles), the latter characterized by a junction site before intron 2 as indicated by Southern blot, PCR, and sequence analyses. Previously described mutations were not present in 1.2% of unrelated congenital adrenal hyperplasia alleles, including one female patient presenting with severe genital virilization. Sequence analysis of the complete functional 21-hydroxylase gene revealed an as yet undescribed mutation in exon 10-Arg(426)His, which has not yet been described to represent a common pseudogene sequence. In vitro expression experiments showed the Arg(426)His mutant to exhibit only low enzyme activity toward the natural substrate 17-hydroxyprogesterone corresponding to the degree of disease manifestation in the patient in whom it was found.

Adrenal Hyperplasia, Congenital↗
PubMed2001

Production rates of dihydrotestosterone in healthy men and women and in men with male pattern baldness: determination by stable isotope/dilution and mass spectrometry.

Production rates of dihydrotestosterone (DHT) were determined in healthy men (n = 8), in healthy women during the follicular phase of their menstrual cycle (n = 7), and in young men with male pattern baldness (n = 8) using the stable isotope dilution technique and mass spectrometry. [2,3,4-(13)C]DHT was infused for 10 h at doses of 15 microg/h (men) and 2 microg/h (women), and blood samples were obtained at 20-min intervals during the last 4 h of the observation period. Production rates estimated between April and June were 2.9 +/- 1.1 microg/h (women) and 17.8 +/- 6.2 microg/h (men). In men production rates of DHT were similar (16.2 +/- 7.7 microg/h) when the investigation was repeated between October and December. Mean production rates of DHT in young men with male pattern baldness (60 +/- 50 microg/h) were higher than those in healthy men (P < 0.005). Although this group included two individuals with normal production rates of DHT, the production rate of DHT was markedly elevated (range, 32.0-161.0 microg/h) in the remaining patients. Stable isotope-labeled infusions of DHT are suitable for clinical use in a routine setting to obtain analytically correct estimates of DHT production in vivo. In the majority of men with male pattern baldness endogenous production of DHT is markedly increased, providing a rationale for therapeutic 5 alpha-reductase inhibition in this disorder.

Adult↗
PubMed2001

Free fatty acids inhibit the glucose-stimulated increase of intramuscular glucose-6-phosphate concentration in humans.

To test Randle's hypothesis we examined whether free fatty acids (FFAs) affect glucose-stimulated glucose transport/phosphorylation and allosteric mediators of muscle glucose metabolism under conditions of fasting peripheral insulinemia. Seven healthy men were studied during somatostatin-glucose-insulin clamp tests [plasma insulin, 50 pmol/L; plasma glucose, 5 mmol/L (0-180 min), 10 mmol/L (180-300 min)] in the presence of low (0.05 mmol/L) and increased (2.6 mmol/L) plasma FFA concentrations. (31)P and (1)H nuclear magnetic resonance spectroscopy was used to determine intracellular concentrations of glucose-6-phosphate (G6P), inorganic phosphate, phosphocreatine, ADP, pH, and intramyocellular lipids. Rates of glucose turnover were measured using D-[6,6-(2)H(2)]glucose. Plasma FFA elevation reduced rates of glucose uptake at the end of the euglycemic period (R(d 150-180 min): 8.6 +/- 0.5 vs. 12.6 +/- 1.6 micromol/kg.min, P < 0.05) and during hyperglycemia (R(d 270-300 min): 9.9 +/- 0.6 vs. 22.3 +/- 1.7 micromol/kg.min, P < 0.01). Similarly, intramuscular G6P was lower at the end of both euglycemic (G6P(167-180 min): -22 +/- 7 vs. +24 +/- 7 micromol/L, P < 0.05) and hyperglycemic periods (G6P(287-300 min): -7 +/- 9 vs. +28 +/- 7 micromol/L, P < 0.05). Changes in intracellular inorganic phosphate exhibited a similar pattern, whereas FFA did not affect phosphocreatine, ADP, pH, and intramyocellular lipid contents. In conclusion, the lack of an increase in muscular G6P along with reduction of whole body glucose clearance indicates that FFA might directly inhibit glucose transport/phosphorylation in skeletal muscle.

Adult↗
PubMed2001

Glucose metabolism in pregnancy at high altitude.

OBJECTIVE: To assess insulin sensitivity and beta-cell function associated with lower maternal fasting plasma glucose levels at high altitude compared with sea level. RESEARCH DESIGN AND METHODS: We studied 215 pregnant women at 8-42 weeks of gestation in Peru. The women were recruited from Cerro de Pasco, which is situated 4,370 m (14,340 feet) above sea level, and Lima, which is at sea level. We also examined 53 nonpregnant control subjects (22 in Cerro de Pasco and 31 in Lima). Fasting plasma glucose, insulin, C-peptide, and proinsulin concentrations were measured in samples obtained from the antecubital vein between 8:00 a.m. and 10:00 a.m. after an overnight period of fasting for 10-14 h Insulin resistance and beta-cell function were calculated using homeostasis model assessment. RESULTS: Fasting C-peptide levels and beta-cell function were similar, fasting concentrations of insulin and proinsulin were lower, and insulin sensitivity was higher at high altitude compared with sea level. CONCLUSIONS: Maternal fasting plasma glucose that is lower at high altitude than at sea level in the presence of similar insulin secretion is associated with higher peripheral insulin sensitivity. This may partly explain the lower birth weights at high altitudes.

Altitude↗
PubMed2000

Cloning and expression of secretagogin, a novel neuroendocrine- and pancreatic islet of Langerhans-specific Ca2+-binding protein.

We have cloned a novel pancreatic beta cell and neuroendocrine cell-specific calcium-binding protein termed secretagogin. The cDNA obtained by immunoscreening a human pancreatic cDNA library using the recently described murine monoclonal antibody D24 contains an open reading frame of 828 base pairs. This codes for a cytoplasmic protein with six putative EF finger hand calcium-binding motifs. The gene could be localized to chromosome 6 by alignment with GenBank genomic sequence data. Northern blot analysis demonstrated abundant expression of this protein in the pancreas and to a lesser extent in the thyroid, adrenal medulla, and cortex. In addition it was expressed in scant quantity in the gastrointestinal tract (stomach, small intestine, and colon). Thyroid tissue expression of secretagogin was restricted to C-cells. Using a sandwich capture enzyme-linked immunosorbent assay with a detection limit of 6.5 pg/ml, considerable amounts of constitutively secreted protein could be measured in tissue culture supernatants of stably transfected RIN-5F and dog insulinoma (INS-H1) cell clones; however, in stably transfected Jurkat cells, the protein was only secreted upon CD3 stimulation. Functional analysis of transfected cell lines expressing secretagogin revealed an influence on calcium flux and cell proliferation. In RIN-5F cells, the antiproliferative effect is possibly due to secretagogin-triggered down-regulation of substance P transcription.

Amino Acid Sequence↗
PubMed2000

Tau polymorphisms are not associated with Alzheimer's disease.

Alzheimer's disease (AD) is one of a number of neurodegenerative conditions including frontotemporal dementia and progressive supranuclear palsy that are associated with abnormal tau protein aggregates in neurons. Mutations in the tau gene cause familial forms of frontotemporal dementia and alleles of the tau gene have been associated with risk for progressive supranuclear palsy. However, studies evaluating whether polymorphic variation in tau is associated with AD have produced conflicting results. We investigated the role of the tau exon 2 polymorphism in a large sample of AD cases and controls and found no evidence that polymorphic variation in tau is associated AD.

Aged↗
PubMed2000

Posttranslational modification and plasma membrane localization of the Drosophila melanogaster presenilin.

Mutations in two genes, presenilin 1 (PS1) and presenilin 2, are linked to early onset cases of familial Alzheimer's disease. The presenilins are thought to contribute to the pathogenesis of Alzheimer's disease by directly or indirectly affecting the proteolytic processing of the amyloid precursor protein. They have also been implicated in the proteolytic processing of Notch. In PS1-deficient mammalian cells, the proteolytic release of the Notch intracellular domain is reduced. Likewise, loss-of-function mutations in Drosophila presenilin (Psn) prevent the production of the intracellular Notch signaling fragment and lead to phenotypes resembling Notch mutants. Here we characterize the Drosophila Psn protein and demonstrate that it undergoes a proteolytic cleavage. We describe Psn expression at different developmental stages of the fly and show Psn localization near both apical and basal plasma membranes. Furthermore, we demonstrate that portions of the Psn protein span the plasma membrane in S2 cells.

Alternative Splicing↗
PubMed2000

production rates of testosterone in patients with Cushing's syndrome.

Testosterone production rates were determined in 16 patients with Cushing's syndrome (4 men and 12 women) using the stable-isotope dilution technique and mass spectrometry. 1alpha,2alpha-D-Testosterone was infused for 10 hours at a dose of 20 microg/h (men) and 0.4 microg/h (women) and blood samples were obtained at 20-minute intervals during the last 4 hours of the observation period. Estimated production rates in men with Cushing's syndrome were 27, 73, 150, and 180 microg/h (mean, 106 +/- 70 microg/h; healthy men [n = 12], 210 +/- 70 microg/h). In the 12 women with Cushing's syndrome, testosterone production rates were 0.3 to 22.3 microg/h (healthy women [n = 5], 4.3 +/- 1.9 microg/h). There was no difference in testosterone production rates in female patients with central (n = 8) versus adrenal (n = 4) Cushing's syndrome. In summary, testosterone production rates are subnormal or low-normal in male patients with endogenous hypercortisolism, but not in female patients with the same disorder. We conclude that testosterone production in men, but not in women, is predominantly of gonadal origin and hence susceptible to a glucocorticoid-induced suppression of gonadotropin secretion.

Adult↗
PubMed2000

Relative hypoleptinemia in patients with type 1 and type 2 diabetes mellitus.

OBJECTIVE: To determine the relation between plasma leptin concentrations and metabolic control in human diabetes mellitus. DESIGN AND SUBJECTS: Cross sectional study consisting of 156 patients with diabetes mellitus type 1 (n=42), type 2 (n=114), and non-diabetic subjects (n=74). RESULTS: Plasma leptin concentrations were lower (P<0.05) in type 1 (8.3+/-1.7 ng/ml) and type 2 diabetic (14.9+/-1.8 ng/ml) than in non-diabetic humans (18.3+/-1.9 ng/ml). Only female type 1 and type 2 diabetic subjects also had decreased leptin/BMI ratios (P<0.05 vs non-diabetic females). The log rank test identified age-adjusted correlation of plasma leptin concentration with sex (P<0.0004) and body mass index (P<0.0218), but not with glycosylated haemoglobin A1c (P>0.5) in all groups. Plasma leptin was correlated with age (P<0.0058) and serum triglycerides (P<0.0199) in type 1 diabetic patients, and with serum cholesterol (P<0.0059) and LDL (P<0.0013) in type 2 diabetic patients. CONCLUSIONS: Defective leptin production and/or secretion might be present independently of metabolic control in female patients with type 1 or type 2 diabetes mellitus.

Adult↗
PubMed2000

Spuriously high serum concentrations of TSH due to the use of an automatic pipetting device.

In a clinically hyperthyroid patient a TSH-value of 0.75 mU/L was measured. Upon retesting TSH was <0.1 mU/L. The patient's sample initially had been preceded, in the pipetting device, by a grossly hypothyroid patient's sample. Analogous problems were subsequently documented in 10 cases when samples of other hyperthyroid patients were preceded by samples with modestly to markedly elevated TSH. The spurious elevation of TSH (0.12 to 1.18 vs. <0.1 mU/L) appears to be due to a carryover effect from the respective previous sample within the used automatic pipetting system. This carryover was much larger than previously suggested, and occurred unpredictably.

Artifacts↗
PubMed2000

Metabolic response to anisoosmolarity of rat skeletal muscle in vitro.

Isolated rat hepatocytes exhibit an insulin-like anabolic response to hypoosmotic incubation and a glucagon-like catabolic response to hyperosmotic incubation. Recently, a distinct glycogenic response to hypoosmotic treatment was likewise reported for cultured rat myotubes. The present study examines the effects of anisoosmolar exposure on glucose metabolism in freshly isolated rat soleus muscle strips. Under the same experimental conditions as used for cultured myotubes, hypoosmolarity reduced net glycogen synthesis to 52%, while hyperosmolarity stimulated glycogen storage to 231% of isoosmolar control (nmol glucose incorporated into glycogen g(-1) x h(-1): hypoosmolar, 34+/-3; isoosmolar, 65+/-8; hyperosmolar, 150+/-11; p<0.01 each vs. isoosmolar). The responses of native skeletal muscle to anisoosmolarity are therefore in opposition to what has been described for hepatocytes and cultured myotubes. Further experiments on rat skeletal muscle revealed that the observed lack of a glycogenic response to hypoosmolarity persisted independent of medium composition, specifically with regard to prevailing glucose and K+ concentrations. In conclusion, hypoosmotic exposure inhibits glycogen synthesis in isolated rat soleus muscle, which clearly argues against the hypothesis that osmotic changes and cell swelling may be physiologically relevant stimulators of muscle glycogen synthesis.

Animals↗
PubMed2000

Hormonal and metabolic counterregulation during and after high-dose insulin-induced hypoglycemia in diabetes mellitus type 2.

Non-obese type 2 diabetic subjects in good metabolic control (n=6, HbA1c 7.0 +/- 0.3%, mean diabetes duration: 5.7 +/- 1 years) and matched non-diabetic subjects (control; n = 6) were studied during hyperinsulinemic (approximately 3 nmol/l)-hypoglycemic (approximately 3.1 mmol/l) clamp tests (0-120 min) and the subsequent recovery period (120-240 min). Plasma glucagon rose gradually but not significantly, whereas norepinephrine and epinephrine similarly increased approximately 2 and approximately 25-fold in both groups. Islet amyloid polypeptide (IAPP) decreased to approximately 41% and approximately 24% of basal values during hypoglycemia and rapidly rose approximately 4.7-fold during the recovery period, while plasma C-peptide remained suppressed in both groups. Within 140 min, plasma free fatty acids similarly decreased to approximately 70 micromol/l (p < 0.05), but then rose to values being approximately 50% higher in diabetic than in control subjects (240 min: 907 +/- 93 vs. 602 +/- 90 micromol/l; p < 0.05). Glucose infusion rates were comparable during hypoglycemia, but approximately 40% lower during recovery in diabetic patients (1.88 +/- 0.27 vs. 3.44 +/- 0.27 mg x kg(-1) x min(-1), p < 0.001). These results demonstrate that (i) hypoglycemia induced by high-dose insulin largely abolishes the counterregulatory response of glucagon, but not of catecholamines in nondiabetic and well-controlled type 2 diabetic subjects, (ii) the rapid posthypoglycemic increase of plasma IAPP occurs independently of plasma insulin, and (iii) the superior rise in plasma free fatty acids may account at least in part for the posthypoglycemic insulin resistance of type 2 diabetic patients.

Adult↗
PubMed2000

Effects of free fatty acid elevation on postabsorptive endogenous glucose production and gluconeogenesis in humans.

Effects of free fatty acids (FFAs) on endogenous glucose production (EGP) and gluconeogenesis (GNG) were examined in healthy subjects (n = 6) during stepwise increased Intralipid/heparin infusion (plasma FFAs 0.8+/-0.1, 1.8+/-0.2, and 2.8+/-0.3 mmol/l) and during glycerol infusion (plasma FFAs approximately 0.5 mmol/l). Rates of EGP were determined with D-[6,6-2H2]glucose and contributions of GNG from 2H enrichments in carbons 2 and 5 of blood glucose after 2H2O ingestion. Plasma glucose concentrations decreased by approximately 10% (P < 0.01), whereas plasma insulin increased by approximately 47% (P = 0.02) after 9 h of lipid infusion. EGP declined from 9.3+/-0.5 (lipid) and 9.0+/-0.8 pmol x kg(-1) x min(-1) (glycerol) to 8.4+/-0.5 and 8.2+/-0.7 micromol x kg(-1) x min(-1), respectively (P < 0.01). Contribution of GNG similarly rose (P < 0.01) from 46+/-4 and 52+/-3% to 65+/-8 and 78+/-7%. To exclude interaction of FFAs with insulin secretion, the study was repeated at fasting plasma insulin (approximately 35 pmol/l) and glucagon (approximately 90 ng/ml) concentrations using somatostatin-insulin-glucagon clamps. Plasma glucose increased by approximately 50% (P < 0.005) during lipid but decreased by approximately 12% during glycerol infusion (P < 0.005). EGP remained unchanged over the 9-h period (9.9+/-1.2 vs. 9.0+/-1.1 micromol x kg(-1) x min(-1)). GNG accounted for 62+/-5 (lipid) and 60+/-6% (glycerol) of EGP at time 0 and rose to 74+/-3% during lipid infusion only (P < 0.05 vs. glycerol: 64+/-4%). In conclusion, high plasma FFA concentrations increase the percent contribution of GNG to EGP and may contribute to increased rates of GNG in patients with type 2 diabetes.

Absorption↗
PubMed2000

Prevention of in vitro lipolysis by tetrahydrolipstatin.

BACKGROUND: Metabolic effects of free fatty acids (FFAs) frequently are tested using combined infusion of triglycerides and heparin, which stimulates lipolysis in vivo. Ongoing in vitro lipolysis, however, probably produces falsely high plasma FFA concentrations under these conditions. Therefore, this study aims to assess the efficacy of tetrahydrolipstatin (THL) in inhibiting plasma lipolytic activity and to improve plasma FFA determination. METHODS: Plasma concentrations of FFAs and glycerol were measured in five healthy subjects in the presence and absence of THL. Blood was drawn at baseline, during infusion of a triglyceride emulsion (1.5 mL/min), and during infusion of triglycerides plus heparin (0.2 IU. kg(-1). min(-1)). In addition, the effects of storage temperature of the samples were analyzed. RESULTS: In samples frozen immediately after collection, plasma FFAs were 28% lower in the presence of THL than in its absence (P = 0.008). When THL-free plasma was incubated for 3 h on ice or at room temperature, plasma FFAs were 22% (P = 0.02) and 91% (P = 0.0004) higher, respectively, than in samples frozen immediately. The addition of THL blunted temperature-dependent in vitro lipolysis by 88% (P<0.01) and 89% (P <0.001) after incubation on ice and at room temperature, respectively. Changes in plasma glycerol concentrations exhibited similar behavior. CONCLUSIONS: THL, which is safe and easy to handle, is a potent inhibitor of in vitro lipolysis and could, therefore, be added to blood samples drawn during triglyceride/heparin infusions to allow more accurate determination of plasma FFA concentrations.

Adult↗
PubMed1999

Construction of a detailed physical and transcript map of the FTDP-17 candidate region on chromosome 17q21.

Frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17) is an autosomal dominant condition clinically characterized by behavioral, cognitive, and motor disturbances. Until now, at least 13 different FTDP-17 families that show linkage to chromosome 17q21 have been described. To characterize the FTDP-17 candidate region, flanked by the markers D17S1789 and D17S1804, we constructed a physical map in P1 and PAC clones. A detailed transcript map was generated by positioning known genes and EST clusters to the physical map. In total, we investigated 150 STSs mapped to this region. In addition, novel transcripts were isolated by exon-trapping. We were able to localize 19 known genes and a number of ESTs to this chromosomal region. Furthermore, seven novel genes were identified for which we isolated the full-length sequence.

Base Sequence↗