PubMed Health⌕ Search

Biomedical subjects

G Wirthensohn

Publications and source records attributed to G Wirthensohn.

At least 19 recordsLinked to original sources
PubMed1992

Carbohydrate and lipid metabolism of the renal tubule in diabetes mellitus.

Diabetic nephropathy not only involves vascular and glomerular changes but also affects tubular metabolism, structure and function. Under acute insulin withdrawal the tubular size increases with glomerular hyperfiltration. Insulin like growth factor 1 (IGF1) has been found to be a candidate mediator involved under these conditions. Tubular carbohydrate metabolism is characterized by gluconeogenesis in the proximal tubule, glycolytic enzymes in the distal segments and high aldose reductase activity in the structures of the renal papilla. In the diabetic state, gluconeogenesis is stimulated by changes of the acid base status. Mitochondrial glucose oxidation is decreased by inhibition of pyruvate dehydrogenase activity through preferential oxidation of fatty acids and ketone bodies. The increase in glycogen in distal tubule cells and sorbitol accumulated in papillary structures can be explained by the high extracellular glucose supply under diabetic conditions. Fatty acids taken up in excess of tubular energy needs accumulate in the nephron as triacylglycerols, mainly in the proximal convoluted tubule. Fatty acid oxidation is inhibited by ketone bodies in proximal and outer medullary tubules, leading to preferential oxidation of the latter under ketotic conditions. Ammonia formed during tubular metabolism of glutamine increases in metabolic acidosis but is suppressed by ketone bodies, leading to a nitrogen sparing effect of ketone bodies. All acute metabolic derangements are abolished, and normal metabolism reestablished by adequate insulin treatment in vivo.

Animals↗
PubMed1989

Regulation of organic osmolyte concentrations in tubules from rat renal inner medulla.

Glycerophosphorylcholine, inositol, and sorbitol were measured in rat kidney homogenates and tubules from inner medulla and papilla by enzymatic spectrophotometric techniques. Organic osmolytes exhibited their highest concentrations in the papillary tip. In contrast to glycerophosphorylcholine and sorbitol, inositol was of similar high concentrations in inner and outer medulla. Freshly prepared inner medullary tubules maintained tissue osmolyte concentrations under control, antidiuretic, and furosemide diuretic conditions. When tubules were incubated in vitro over 90 min, tubular organic osmolyte concentrations decreased as a function of extracellular NaCl, but not urea concentrations. Organic osmolyte disappearance from cells was quantitatively recovered from the medium. In contrast, medium lactate dehydrogenase activity did not rise in parallel and tubular ATP remained constant. Glucose up to a concentration of 200 mM increased tubule and medium sorbitol. The results obtained indicate that glycerophosphorylcholine, sorbitol, and inositol rapidly adapt their intracellular concentrations to extracellular NaCl osmolality by a change in tubular plasma membrane permeability. In addition sorbitol levels are regulated by the extracellular glucose concentration.

Animals↗
PubMed1987

Role and regulation of glycerophosphorylcholine in rat renal papilla.

Glycerophosphorylcholine (GPC)--an organic solute which is considered to be involved in cellular osmoregulation in the renal medulla--was determined by means of an enzymatic assay in various zones of the rat kidney and in papillary tubule suspensions. In antidiuresis, GPC content in cortex, outer medulla and papillary tip was 0.64, 14.6, and 108.9 mmol/kg fresh weight, respectively. Significant concentrations of GPC could not be detected in the urine or in the peripheral plasma. The sharp increase in GPC concentration from cortex to papillary tip was partially abolished by the induction of diuresis by either waterloading or furosemide. These manoeuvres, however, did not change cortical GPC content. Papillary tubule suspensions prepared from hydropenic rats contained only slightly less GPC per g protein than whole papillae from antidiuretic animals. Incubation of tubules over 120 min did not lead to a significant loss of GPC which is in accordance with the low activity of GPC degrading enzymes in papillary tissue. The results confirm the intracellular localization of GPC and provide further evidence that this substance plays a substantial role in the osmoregulation of renal papillary cells.

Animals↗
PubMed1987

Kallikrein (kininogenase) in the mouse nephron: effect of dietary potassium.

Kininogenase activity of kallikrein was measured in microdissected mouse nephron segments using kininogen from dog plasma and a radioimmunoassay for bradykinin. When single nephron segments were examined, results showed a large scatter. This was found to be due to heterogeneity of distal convoluted tubules (DCT) from different nephrons, since replicate measurements in pools of DCT structures did not show this degree of variation. Nearly 20% of activity was accessible to extracellular substrate when freshly dissected segments were incubated in isoosmotic media. Freezing and thawing which markedly releases activity of intracellular enzymes, did not significantly elevate kininogenase activity. On the other hand deoxycholate and trypsin treatment increased tubular kininogenase activity in an additive fashion. A detailed analysis of microdissected tubule fragments revealed that kallikrein is concentrated in late distal convoluted tubule before entering a branching point (connecting tubule). In contrast initial portions of distal convoluted tubules and cortical collecting tubules contained only little kallikrein activity. Potassium rich diet increased basal and total activity 5-fold, when compared to a potassium poor diet.

Animals↗
PubMed1987

Quantification of kininogen in human renal medulla.

Renal kininogen was detected in human medullary tissue as well as human medullary tubule suspensions. After treatment with pig pancreatic kallikrein or human renal cortical homogenate liberated kinin was measured by bradykinin radioimmunoassay. In the absence of inhibitors kinins were degraded by kininases located in the same part of the kidney. Several known inhibitors of kininase I and II did not inhibit this activity. Endogenous medullary kininase was inhibited by preincubation of homogenates at 56 degrees C for one hour or by addition of 0.25 mmol/l HgCl2. Under these conditions endogenous medullary kinin release amounted to 9-26 nmol/g protein. The action of renal cortical kininogenase on kinin formation from papillary kininogen was completely inhibited by addition of 1 mumol/l aprotinin. Kininogen examined in renal tubule suspensions revealed an increase in amount per g protein compared to homogenates, confirming the tubular localization of renal kininogen.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1985

Stimulation of phospholipid turnover by angiotensin II and phenylephrine in proximal convoluted tubules microdissected from mouse nephron.

Angiotensin II and phenylephrine significantly increased phospholipid turnover in cortical tubule suspensions. To further localize this effect we studied the 32P incorporation into phosphatidylcholine (PC) and phosphatidylinositol (PI) in single dissected proximal convoluted tubules of mouse nephron. Both hormones significantly stimulated 32P labeling of PC and PI indicating that the proximal tubule is a target site of angiotensin II and phenylephrine action.

Angiotensin II↗
PubMed1984

Phospholipid metabolism in rat kidney cortical tubules. I. Effect of renal substrates.

Renal phospholipid metabolism was studied in rat cortical tubule suspensions by combining net measurements with precursor incorporation studies in order to be able to calculate the turnover of the different phospholipid species. Net amounts of tubular phosphatidylcholine (PC), phosphatidylinositol (PI) and phosphatidylethanolamine (PE) were 96.7 +/- 3.9, 29.2 +/- 2.9 and 74.1 +/- 6.0, mumol per g protein, respectively. Incubation of tubules in the absence or presence of renal substrates did not change net phospholipid contents. The predominant fatty acids were palmitate (47%) in PC and stearate in PI (56%) and PE (41%). Highest [14C]palmitate and [14C]glycerol incorporation rates were found in PC. In contrast, 32P-labeling was highest in phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate exceeding PI-labeling by a factor of 2.4 and 5.2, respectively. In contrast, [3H]inositol incorporation into phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate was only 20% compared to that into PI. Addition of renal substrates like lactate, glutamine, glycerol and fatty acids increased precursor incorporation. The stimulating effects of gluconeogenic precursors and fatty acids were more than additive. Comparison of rates of [14C]glycerol, [32P]- and [3H]inositol incorporation suggests that de novo phospholipid biosynthesis is represented by these measurements, while the fatty acid fraction in addition turns over by exchange processes. According to our conclusions PE, PC, PI and phosphoinositides turn over with a half-life of 139, 16, 15 and 0.7 h, respectively, under optimal in vitro conditions.

Animals↗
PubMed1984

Phospholipid metabolism in rat kidney cortical tubules. II. Effects of hormones on 32P incorporation.

To evaluate the effect of hormones on renal phospholipid metabolism and turnover, we studied the changes in 32P-labeling of phospholipids in rat cortical tubule suspension. Angiotensin II, phenylephrine and parathyroid hormone (PTH) stimulate 32P incorporation into PC by 25, 29 and 26% and into PI by 189, 328 and 33% above control rates, respectively, whereas phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate labeling was not affected. However, when phospholipids were prelabeled with [32P]Pi, addition of angiotensin II led to a significant decrease in phosphatidylinositol 4,5-bisphosphate labeling in the first 2 min with no effect on the other phospholipid fractions. The phenylephrine effect on phospholipid labeling was blocked by prazosin but not by yohimbine, indicating an alpha 1-mediated action. In contrast, the effect of angiotensin II was not inhibited by either antagonist. The stimulating effect of substrates on 32P incorporation reported in the preceding paper was additive to that of hormones. Our results confirm previous studies on renal gluconeogenesis that catecholamines act by an alpha 1-type receptor on proximal tubules, and indicate that phenylephrine and angiotensin II act by different receptor sites exerting the same metabolic effect. The additivity of hormone effects with that of renal substrates indicates that the former are not secondary to release of precursors for phospholipid biosynthesis. The rapid decrease in phosphatidylinositol 4,5-bisphosphate labeling after angiotensin II suggests that the polyphosphoinositide is degraded after hormone binding to the receptor and that PI labeling is a secondary event.

Angiotensin II↗
PubMed1984

Bioluminescence procedures for the measurement of NAD(P) dependent enzyme catalytic activities in submicrogram quantities of rabbit and human nephron structures.

Reduced flavin mononucleotide dependent luciferase (EC 1.14.14.3) from Photobacterium fischeri has been used to measure NAD(P) dependent enzymes in submicrogram quantities of tissue homogenates and isolated structures of rabbit and human kidney. The procedure for measuring NAD(P)H was optimized, with internal standardization, to give a linear constant signal between 1 and 100 pmol. This method was applied to the measurement of glucose-6-phosphate dehydrogenase (EC 1.1.1.49) and 3-hydroxybutyrate dehydrogenase (EC 1.1.1.30) along the various structures of the rabbit nephron, microdissected from fresh tissue slices. Blank and recovery measurements were performed on each structure, and enzyme catalytic activities were calculated on the basis of tubular length and protein. Glucose-6-phosphate dehydrogenase was found to be present in all nephron structures with highest catalytic activities in glomeruli, thin limbs of Henle's loop and medullary collecting tubules. Lowest catalytic activities were detected in the pars recta of proximal tubules, the distal convoluted and the connecting tubule. The mitochondrial 3-hydroxybutyrate dehydrogenase exhibited a different distribution pattern: Highest catalytic activities were found in the cortical ascending limbs of Henle's loop, the proximal and distal convoluted tubule. An unusual internephron heterogeneity for this enzyme was found in the distal convoluted tubule. Catalytic activities in thin limbs of Henle's loop, the medullary ascending limb, and the cortical and medullary collecting tubule were not significantly different from blank activities. The results obtained in isolated nephron segments agreed with those calculated from cortex and medullary homogenates. In a preliminary experiment on human kidney it could be demonstrated that the procedures can be applied to fresh human biopsy samples. The bioluminescence method offers several advantages (simplicity, rapidity) in comparison with the classical techniques used for ultramicro analysis of tissue enzymes (enzymatic cycling, radiochemical tests). The results confirm the biochemical heterogeneity of nephron structures and allow conclusions about species dependent functional differences of the pentose-phosphate cycle and renal ketone body metabolism along the nephron.

Animals↗
PubMed1983

Phosphatidylcholine biosynthesis in rabbit kidney tubule suspensions. Effect of metabolic substrates on precursor incorporation.

To gain insight in the mechanisms that regulate phosphatidylcholine (PC) biosynthesis we studied the incorporation of [14C]choline and 14C-labelled fatty acids into PC of tubule suspensions of rabbit kidney cortex and outer medulla, in vitro. Moreover, the influence of renal substrates on these incorporation rates were examined. When incubated with saturating concentration of 1 mM [14C]choline tubules incorporated 4.40 +/- 0.36 mumol choline per g protein per h. Addition of fatty acids such as palmitate and oleate led to a significant increase in [14C]choline incorporation into PC. Further addition of renal substrates such as glucose, propionate, proline, glutamine and glutamate further enhanced this stimulated incorporation significantly. Maximal rates of [14C]choline incorporation were found to be 9.94 mumol per g protein per h in presence of choline, oleate and glucose. In tubules of the outer medulla only glucose showed a stimulatory effect. On the other hand, 14C-labelled fatty acids were incorporated into tubular PC to a similar extent as was found for [14C]choline incorporation (4.54 +/- 0.62 and 5.52 +/- 0.77 mumol/g protein per h for oleate and palmitate, respectively). Addition of choline or renal substrates increased fatty acid incorporation in an additive manner. Maximal rates of 14C-labelled fatty acid incorporation were found to be 17.7 mumol per g protein per h in presence of oleate, choline and propionate. In contrast to isotope studies, net PC content decreased rather than increased under all experimental conditions, indicating that results may represent membrane lipid turnover. The observed rates are sufficient to replace total renal membrane PC in 1 day.

Animals↗
PubMed1983

Renal ketone body metabolism. Distribution of 3-oxoacid CoA-transferase and 3-hydroxybutyrate dehydrogenase along the mouse nephron.

Two enzymes of ketone body metabolism, 3-oxoacid CoA-transferase and 3-hydroxybutyrate dehydrogenase were measured with ultramicromethods to map their activities along single, dissected segments of mouse nephron. 3-Oxoacid CoA-transferase activity was assayed with a radiochemical procedure by separating [14C]succinyl-CoA, formed in the presence of acetoacetyl-CoA from [1,4-14C]succinate. This procedure, when compared to the spectrophotometric method, resulted in similar activities in mouse organ homogenates. 3-Hydroxybutyrate dehydrogenase activity was determined with sufficient sensitivity by using NADH-dependent luciferase. The dehydrogenase exhibited only 5% of the activity of the 3-oxoacid CoA-transferase when measured in mouse kidney cortex homogenates. Both enzymes were found to be present in all nephron structures studies. The specific activity related to tubular protein, which was determined in parallel in all segments dissected, exhibited a typical distribution pattern along the nephron. Both enzymes roughly paralleled each other along the structures of the distal nephron. They exhibited high activities in the thick ascending limb of Henle's loop and the distal convoluted tubule, but decreased to nearly 20% in the segments of the collecting tubule. In the proximal convoluted and straight tubule 3-oxoacid CoA-transferase activity was almost equal. In contrast, 3-hydroxybutyrate dehydrogenase increased by a factor of 5 from the convoluted to the straight portion. This heterogeneity along the proximal tubule remained when pure D-hydroxybutyrate was used as substrate, but was not found when rat or rabbit nephron segments were analysed. Lowest activities of enzymes were recovered from glomeruli and thin descending limbs of Henle's loop. The results obtained allow for the conclusion that 3-hydroxybutyrate and acetoacetate can be metabolized in all structures of the mouse nephron with different capacities in various segments. With the exception of the relatively low activity of 3-hydroxybutyrate dehydrogenase in the proximal convoluted tubule, the distribution pattern mirrors the distribution of mitochondria along the rat nephron. The results point to the possible role of ketone bodies as energy fuels for nephron segments performing active transport processes.

Animals↗
PubMed1982

Choline kinase activity along the rabbit nephron.

Choline kinase catalyzes the phosphorylation of choline to phosphorylcholine which is thus made available for phosphatidylcholine biosynthesis. Choline kinase activity was determined in defined microdissected structures of rabbit nephron with a radiochemical microprocedure. Enzyme activity was present in all segments tested. When referred to tubular length, the highest activities were found in proximal convoluted tubules. Due to the lower protein content of distal structures these segments exhibited higher enzyme activities when referred to micrograms of protein. From this distribution pattern it was concluded that all nephron segments are able to use extracellular choline for phosphatidylcholine biosynthesis.

Animals↗
PubMed1981

Renal glycerol metabolism and the distribution of glycerol kinase in rabbit nephron.

Glycerol and dihydroxyacetone are metabolized by rabbit kidney-cortex tubules, isolated by collagenase treatment. Half-maximal concentrations of both substrates were determined with regard to uptake rates and product formations. Maximal uptake rates were 643 and 329 mumol/h per g of protein for dihydroxyacetone and glycerol respectively. Glucose and lactate were found as major metabolic products. Glycerol kinase, the enzyme catalysing the first step in renal glycerol and dihydroxyacetone metabolism, was measured radiochemically as described by Newsholme, Robinson & Taylor [(1967) Biochim, Biophys. Acta 132, 338-346] and adapted for studies of the localization of this enzyme along the different structures of rabbit nephron. The results show that glycerol kinase is located exclusively in the proximal segments, i.e. the proximal convoluted tubules and the pars recta, but is negligible in the other structures studied. The activities were close to the maximal dihydroxyacetone uptake rates measured in tubule suspensions.

Animals↗
PubMed1981

Distribution of hexokinase and phosphoenolpyruvate carboxykinase along the rabbit nephron.

Radiochemical microprocedures were developed for the determination of hexokinase and phosphoenolpyruvate carboxykinase (PEPCK) activity in single microdissected segments of the mature rabbit nephron dissected from fresh tissue after collagenase treatment. All results were related to tubular length and tubular protein content. Hexokinase activity was found to be lowest in the proximal convoluted tubule and to increase along the following nephron segments, with highest activity in the connecting tubule. The gluconeogenic enzyme PEPCK, on the other hand, was exclusively found in the proximal tubule. Early and late portions of the convoluted segment exhibited the same specific activity, but only 50% was found in the pars recta. All other renal structures exhibited only insignificant activity of PEPCK. The results show that renal glucose metabolism and gluconeogenesis are clearly separated. As previously shown for the cytosolic rat enzyme, rabbit mitochondrial PEPCK is also exclusively a proximal tubular enzyme, thus confirming the dominant role of this segment in mammalian renal gluconeogenesis. The high activity of hexokinase in the segments of the distal tubule points to the role of glucose as metabolic fuel, glycogen precursor, and other glucose-6-phosphate-using pathways in these structures.

Animals↗