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Biomedical subjects

G Hennemann

Publications and source records attributed to G Hennemann.

At least 55 records · Page 3Linked to original sources
PubMed1993

Evidence for carrier-mediated uptake of triiodothyronine in cultured anterior pituitary cells of euthyroid rats.

T3 uptake and TSH secretion were investigated in anterior pituitary cells isolated from adult fed Wistar rats and cultured for 3 days in medium containing 10% fetal calf serum. TSH release during culture increased linearly with the number of cells in the range of 80,000-800,000 cells/well. Uptake and incubation experiments were performed at 37 C in medium containing 0.5% BSA. Incubation with TRH (0.1 microM) for 2 h stimulated TSH release 2.6-fold, and this effect was partly (approximately 45%) suppressed by preexposure for 2 h to T3 (0.01-1 microM) or T4 (1 microM). Similar concentrations of T3 and T4 reduced the cellular uptake of [125I]T3 (50 pM) during 1 h of incubation by 55%. After 15 min of incubation, [125I]T3 uptake (percent dose) amounted to 1.26 +/- 0.05% (mean +/- SE; n = 9)/500,000 cells. The major part (75%) of the [125I]T3 was found in the extranuclear fraction. Simultaneous incubation with unlabeled T3 (1 or 10 microM) reduced [125I]T3 uptake by 43% (n = 3; P < 0.001) and 52% (n = 6; P < 0.001), respectively. Reduction of the temperature to 20 C diminished the T3-suppressible fraction of [125I]T3 uptake approximately 3-fold. After preincubation (30 min) and incubation (15 min) with monodansylcadaverine (100 microM), the uptake of [125I]T3 was reduced by 32% (n = 3; P < 0.01). When the Na+ gradient was reduced by preincubation and incubation with ouabain (0.5 mM) or monensin (10 or 100 microM), T3 uptake was inhibited by 25% (n = 5; P < 0.01), 37% (n = 6; P < 0.001), and 61% (n = 3; P < 0.001), respectively. It is concluded that 1) T3 is taken up by the pituitary by a carrier-mediated mechanism, and 2) this uptake is at least partly dependent on the Na+ gradient.

Animals↗
PubMed1993

A furan fatty acid and indoxyl sulfate are the putative inhibitors of thyroxine hepatocyte transport in uremia.

We studied the effects of 3-carboxy-4-methyl-5-propyl-2-furan-propanoic acid (CMPF), indoxyl sulfate, and hippuric acid on iodide production by rat hepatocytes in primary cultures. We questioned whether these substances could explain the alteration of serum thyroid hormone parameters observed in renal failure. Iodide production from [125I]T4 by rat hepatocytes was significantly inhibited in the presence of serum from uremic patients. Serum concentrations of CMPF, indoxyl sulfate, and hippuric acid were markedly elevated in uremic patients. The minimum concentration that inhibited iodide production, when expressed as a molar ratio of the inhibitor to BSA, was 0.13 for CMPF, 0.53 for indoxyl sulfate, and 1.33 for hippuric acid. This molar ratio was lower than the corresponding mean molar ratio in uremic sera for CMPF (0.38) and indoxyl sulfate (0.63), while it was higher than that found for hippuric acid (0.85). The inhibition was reproduced when the inhibitors were added to normal human serum. The decreased iodide production was not due to the inhibition of deiodinase activity. The deiodination of rT3 by rat liver microsomes was unaffected by these inhibitors. Charcoal adsorption of uremic serum normalized the iodide production by hepatocytes. This normalization coincided with almost complete removal of CMPF and indoxyl sulfate, with a concomitant reduction of the free T4 fraction. Dialysis of uremic serum only partially restored iodide production. Even though indoxyl sulfate and hippuric acid were no longer detectable, a high concentration of CMPF remained in the serum. The serum free T4 fraction remained elevated in uremic patients after dialysis. Our studies indicate that CMPF and indoxyl sulfate in concentrations normally present in the serum of uremic patients inhibit cellular transport and subsequent deiodination of T4. These substances may account for the low total T3 level in uremic patients.

Adsorption↗
PubMed1993

Inhibition of thyroxine transport into cultured rat hepatocytes by serum of nonuremic critically ill patients: effects of bilirubin and nonesterified fatty acids.

We investigated bilirubin and oleic acid as causes of low plasma T3 in nonuremic critically ill patients with gross changes in serum thyroid hormone levels (T4, < or = 60; T3, < or = 1.1; rT3, > or = 0.45 nmol/L) and elevated bilirubin concentrations (> or = 33 mumol/L). Iodide production from [125I]T4 was inhibited by 42% when rat hepatocytes in primary cultures were incubated with 10% serum from these patients. The mean serum concentration of albumin was reduced by 41%, while the concentrations of bilirubin and nonesterified fatty acids (NEFA) were increased by 2022% and 115%, respectively, in the patients. The molar ratios of bilirubin/albumin and NEFA/albumin in the patients were 0.42 and 3.18, respectively. Addition of oleic acid (50-400 mumol/L) and bilirubin (3-130 mumol/L) to 10% normal human serum (albumin, 70 mumol/L; NEFA, 54 mumol/L; bilirubin, 1.1 mumol/L) progressively inhibited the production of iodide by rat hepatocytes. The decreased iodide production was presumed to be caused by inhibition of T4 transport into hepatocytes. The deiodination of rT3 by rat liver microsomes was unaltered by free bilirubin and free oleic acid concentrations up to 0.1 mumol/L. These free concentrations are at least 1 order of magnitude higher than that attained in nonthyroidal illness. The inhibition of iodide production by the sera of critically ill patients (n = 12) was significantly correlated with the molar ratios of bilirubin/albumin (r = 0.72; P < 0.01) and NEFA/albumin (r = 0.58; P < 0.05). Extensive dialysis or treatment of the sera with charcoal did not completely remove the inhibitory activity on iodide production. Serum concentrations of indoxyl sulfate, 3-carboxy-4-methyl-5-propyl-2-furan propanoic acid, and hippuric acid in the critically ill patients (other known T4 transport inhibitors into hepatocytes) were similar to those in the normal subjects. This study together with the well known effects of carbohydrate on T3 neogenesis suggest that elevated bilirubin and NEFA and the low albumin level in non-uremic critical illness may be at least partly responsible for the T4 transport inhibition in T3-producing tissues (e.g. the liver) and, thus, the low plasma T3 levels in these critically ill patients. The question of whether inhibitors of T4 transport into the hepatocytes are also present in other patients with nonthyroidal illness who show only mild changes in thyroid hormone levels and have low concentrations of bilirubin and NEFA remains to be determined.

Adsorption↗
PubMed1993

Transport and metabolism of iodothyronines in cultured human hepatocytes.

Thyroid hormone uptake into cultured human hepatocytes was studied using measurement of cell-associated radioactivity of radioiodinated thyroid hormones after 10-min incubation in culture medium with 0.5% BSA. Furthermore, 20-h incubations were performed to study transport and further intracellular metabolism. The results indicate the presence of saturable active uptake systems for T4, T3, and rT3, as addition of the unlabeled hormone (1, 5, and 2 mumol/L, respectively) to the medium resulted in a decrease in cell-associated radioactivity of 20-30%. Inhibition was also achieved after 30-min preincubation with fructose (10 mmol/L), which induces a decrease in intracellular ATP or ouabain (0.5 mmol/L), indicating energy dependence and the necessity for a sodium gradient for at least part of the transport process, respectively. After 20-h incubation, iodide production was inhibited in the presence of ouabain (0.5 mmol/L), propylthiouracil (100 mumol/L), or a monoclonal antibody (81-1A1-10; ascites dilution, 1:200) directed against thyroid hormone transport systems in rat hepatocytes. These data indicate that there is a high degree of similarity between the properties of the uptake process and subsequent conversion of thyroid hormones in human and rat hepatocytes, although the rates of uptake and conversion are lower in human hepatocytes. Furthermore, regulation of thyroid hormone uptake at the level of the plasma membrane may also be operative in human hepatocytes.

Adenosine Triphosphate↗
PubMed1993

Decreased peripheral 3,5,3'-triiodothyronine (T3) production from thyroxine (T4): a syndrome of impaired thyroid hormone activation due to transport inhibition of T4- into T3-producing tissues.

T4, rT3, and T3 kinetic studies have been performed in a T4-substituted female who showed an increased serum T4/T3 ratio and substantially increased rT3 serum levels in the presence of normal serum thyroid hormone binding properties. The kinetic studies were performed to discriminate between T4 transport inhibition into plasma T3-producing tissues and inhibition of extrathyroidal T4 to T3 conversion. The principal findings were that both T4 and rT3 transport were inhibited into the rapid equilibrating pool (REP), which mainly consists of the liver. The plasma T3 production rate was decreased. Despite an elevated serum free T4 level, serum TSH was elevated, pointing to T4 transport inhibition at the level of the thyrotroph as well. Transport of T4 and rT3 was normal to the slowly equilibrating pool, whereas no transport inhibition of T3 was found to either pool. Because T4 into T3 conversion efficiency in the REP (the main source of plasma T3 production) was normal, it was concluded that the lowered T3 production in the subject was caused by transport inhibition of T4 into the liver. Although the occurrence of the syndrome is rare, its significance is of general importance, in that it shows that transport of thyroid hormone may vary at the tissue level. Furthermore, as T3 is the principal biologically active thyroid hormone, regulation of transport of T4 into the REP may play a (patho)physiological role in the ultimate determination of thyroid hormone activity in the tissues.

Aged↗
PubMed1993

Free thyroxine assessed with three assays in sera of patients with nonthyroidal illness and of subjects with abnormal concentrations of thyroxine-binding proteins.

Three methods for estimating free thyroxine (FT4) in serum were studied: equilibrium dialysis, the SPAC-ET FT4 radioimmunoassay kit, and the Amerlite MAB FT4 luminometric assay. Serum samples from 10 subjects with above-normal thyroxine-binding globulin (TBG), 6 with low TBG, 30 with familial dysalbuminemic hyperthyroxinemia (FDH), 13 with nonesterified fatty acids (NEFA) concentrations in serum > 1.0 mmol/L, and 178 patients with various degrees of nonthyroidal illness (NTI) were measured and compared with samples from 42 euthyroid blood donors. The Amerlite MAB FT4 assay compared well with equilibrium dialysis, whereas the SPAC-ET assay averaged 40% lower. All three assays were not influenced by changes in TBG and showed no or only little changes in the presence of NEFA. Mean FT4 values in the FDH samples were somewhat higher than in controls when measured with the SPAC-ET assay, about equal with equilibrium dialysis, and somewhat below the mean control value with the Amerlite MAB FT4 assay, although individual results were within the control reference range. In NTI patients, no FT4 values were below the control reference range by the Amerlite MAB FT4 assay, 4 of 178 were below this range by equilibrium dialysis, and 1 of 178 was below this range by the SPAC-ET assay. In all assays a large proportion of the NTI samples showed FT4 values above the control reference range, a result that will interfere with the efficacy of these assays for assessing thyroid function in NTI patients.

Dialysis↗
PubMed1992

Transport of 3,5,3'-triiodothyronine into the perfused rat liver and subsequent metabolism are inhibited by fasting.

The effects of 48-h fasting on transport of T3 and subsequent metabolism in the isolated perfused rat liver were investigated. Tracer T3 disappearance curves from the recirculating medium consisted of a fast component (FC) and a slow component (SC). Using a two-compartment model, both transport [expressed as the fractional transport rate constant from medium to liver (k21)] and disposal of T3 were calculated. After fasting, k21, total metabolism, and metabolism corrected for differences in mass transfer were diminished, pointing to both decreased transport and metabolism, presumably caused by depletion of liver ATP. Concerning transport, it was shown that only transport into the intracellular liver compartment and not transport to the extracellular liver compartment was decreased after fasting. As for metabolism, T3 glucuronidation was diminished; T3 sulfation and subsequent deiodination were not affected. All mentioned decreased parameters normalized after the addition of a combination of insulin, cortisol, and/or glucose to the medium, possibly by (partially) restoration of cellular energy stores.

Adenosine Triphosphate↗
PubMed1991

Euro-Collins solution versus UW-solution for long-term liver preservation in the isolated rat-liver perfusion model.

To compare UW-solution (UW) and Euro-Collins (EC) for long-term liver preservation we investigated the morphology and metabolic capacity of rat liver after 18 and 42-hours cold-storage in either UW or EC. After harvesting the rat liver was transferred to a perfusion chamber where it was perfused for 10 min with UW or EC at 4 degrees C. Thereafter livers were stored at 4 degrees C in UW or EC for 18 hours (both groups n = 6) or for 42 hours (both groups n = 8). After 18-hr or 42-hr cold-storage a 2-hr warm perfusion (37 degrees C) was started with Krebs-Ringer solution with carbogen to which 125Iodine-triiodothyronine (T3) was added. Control livers (n = 8) were immediately perfused with Krebs-Ringer without cold-storage. The following parameters were assessed: ASAT-levels in the perfusate, T3-metabolites in the bile and the perfusate, the perfusion pressure, the volume of bile secreted and light-microscopical morphology at the end of the warm perfusion period. After cold storage in UW-solution the ASAT-levels in the perfusate were lower than after storage in EC as well as the perfusion pressures. These livers demonstrated a better T3-metabolism and secreted more bile than EC-stored livers. Histological examination showed more tissue damage in the EC-stored livers than in the UW stored livers. We conclude that cold-storage of rat liver in UW-solution resulted in a better morphology and metabolic capacity as compared with EC-solution.

Adenosine↗
PubMed1991

Thyroxine and 3,3',5-triiodothyronine are glucuronidated in rat liver by different uridine diphosphate-glucuronyltransferases.

Male Wistar rats were treated with 50 mg 3,3',4,4'-tetrachlorobiphenyl (TCB)/kg BW or vehicle. After 4 days, the livers were isolated and perfused for 90 min with 2 nM [125I]T3 or 10 nM [125I]T4 in Krebs-Ringer medium containing 1% albumin. Deiodination and conjugation products and remaining substrates were determined in bile and medium samples by Sephadex LH-20 chromatography and HPLC. TCB treatment did not affect hepatic uptake and metabolism of T3. However, biliary excretion of T4 glucuronide was strongly increased by TCB, resulting in an augmented T4 disappearance from the medium, although initial hepatic uptake of T4 was not altered. Measurement of the microsomal UDP-glucuronyltransferase (UDPGT) activities confirmed that T4 UDPGT was induced by TCB, whereas T3 glucuronidation was unaffected. T3 UDPGT activity showed a discontinuous variation, which completely matched the genetic heterogeneity in androsterone glucuronidation in Wistar rats. These results indicate that different isozymes catalyze the glucuronidation of T3 and T4.

Animals↗
PubMed1990

Development and use of a mathematical two-pool model of distribution and metabolism of 3,3',5-triiodothyronine in a recirculating rat liver perfusion system: albumin does not play a role in cellular transport.

To describe the T3 kinetics in a recirculating rat liver perfusion system, we have developed a mathematical two-pool model consisting of medium and liver. It appeared that all parameters of the model could be fully resolved by using the time-dependent disappearance of radioactive T3 (2 nM) from the medium only. The model calculates the T3 medium pool, the T3 liver pool, and the amount of hormone metabolized at different times after the start of the perfusion. To check the validity of the model, metabolism was also estimated from the appearance of labeled metabolites (glucuronides, sulfates, and I-) in the medium and the cumulative excretion of T3 and metabolites into the bile. The medium pool was also estimated by the product of medium volume and remaining T3 concentration, and the liver pool as the amount of T3 at time zero minus medium pool minus T3 metabolized). These results were in excellent agreement with the predicted values from the model. Taking the metabolites appearing in medium and bile together, about 38% of the total amount of T3 metabolized during 60 min was converted into T3 glucuronide, 12% into T3 sulfate, and 48% into I-, respectively, while about 3% was excreted in the bile unaltered. The results show that not all T3 transported to the liver is being metabolized, but part is bound outside the cellular compartment. This latter pool of T3 is dependent on the albumin concentration in the medium. The amount of T3 metabolized is solely determined by the free T3 concentration and is independent of total T3 or albumin concentration in the medium.

Animals↗
PubMed1989

Congenital hypothyroidism and partial thyroid hormone unresponsiveness of the pituitary in a patient with congenital thyroxine binding albumin elevation.

We describe a girl who presented at the age of 6 weeks with cardiogenic shock due to congenital hypothyroidism (serum thyroxine (T4) less than 12 nmol/l). Thyroxine replacement therapy was instituted. In spite of high total serum T4 levels, thyroid stimulating hormone (TSH) serum values remained elevated. The raised serum T4 levels were the result of congenital elevation of thyroid binding albumin (TBA). Toxic doses of both T4 and triiodothyronine (T3) normalized the elevated TSH levels indicating that the pituitary is responsive to thyroid hormone, albeit at a higher threshold. In patients with congenital TBA elevation and an altered T4 pituitary response requiring thyroid replacement therapy, the measurement of serum free T4 levels is the parameter of choice to monitor treatment.

Congenital Hypothyroidism↗
PubMed1988

Three-compartmental analysis of effects of D-propranolol on thyroid hormone kinetics.

Tracer thyroxine (T4), 3.3',5-triiodothyronine (T3), and 3,3',5'-triiodothyronine (rT3) kinetic studies were performed in normal T4 substituted subjects before and during oral D-propranolol treatment to determine whether changes in thyroid hormone metabolism in a propranolol-induced low-T3 syndrome result from inhibition of 5'-deiodination or inhibition of transport of iodothyronines into tissues. Data were analyzed according to a three-compartmental model of distribution and metabolism. T4 plasma appearance rate decreased by 16% (P less than 0.01), reflecting a decreased intestinal absorption of orally administered T4 during propranolol. Serum T4 and free T4 levels increased significantly by 14%, whereas T4 metabolic clearance rate (MCR) was lowered by 26% (P less than 0.001). No changes were observed in size of the three T4 compartments or in fractional and mass transfer rates of T4 from plasma to the rapidly (REP) and slowly (SEP) equilibrating pools. Serum T3, free T3, T3 plasma pool, T3 mass transfer rate to REP and SEP, and the T3 pool masses were all significantly decreased during propranolol to a similar extent as the T3 plasma production rate (PR). T3 MCR decreased by 14% (P less than 0.05). Serum total and free rT3 increased, whereas the rT3 MCR was substantially lowered during propranolol (P less than 0.001). The rT3 plasma pool, rT3 REP and SEP, and the mass transfer rates to REP and SEP increased, whereas no alterations were observed in rT3 PR and fractional transfer rates of rT3 to REP and SEP.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1988

Inhibition of uptake of thyroid hormone into rat hepatocytes by preincubation with N-bromoacetyl-3,3',5-triiodothyronine.

To investigate whether affinity coupling of N-bromoacetyl-T3 (BrAcT3) to the T3 membrane carrier results in an inhibition of transport of T3 into the cell, rat hepatocytes in monolayer were incubated for 2 h at 21 C with 1.3 mumol/liter BrAcT3 in medium without protein. After extensive washing, cells were incubated during 20 h at 37 C with [125I]T3 in medium with 0.5% BSA, and products in supernatants were analyzed by LH-20 column chromatography. In addition the apparent affinity constant (Km) and maximal uptake velocity (Vmax) of the high affinity uptake process were estimated using 1 min incubations of hepatocytes with various concentrations of T3. In control experiments (i.e. without BrAcT3 affinity coupling) about 57% of the added T3 was cleared from the medium and further metabolized, 85% of the cleared T3 reappeared in the medium as I-, 15% as conjugates. Addition of propylthiouracil during the 20 h incubation with T3 strongly inhibited deiodination, without a change in T3 clearance. Because T3 is sulfated before deiodination, a concomitant rise in conjugates was observed. Addition of ouabain to control cells during the 20 h incubation with T3 strongly inhibited uptake, with a parallel decrease in I- and conjugate formation. After affinity coupling of BrAcT3, T3 clearance was inhibited (by 30% P less than 0.001). Since I- production was more depressed (by 73%) than T3 clearance, with some rise in conjugate formation (P less than 0.001), inhibition of deiodinase by BrAcT3 also took place. The effects of BrAcT3 and ouabain on uptake of T3 appeared to be additive as were the effects of propylthiouracil and BrAcT3 on deiodination. After affinity coupling of BrAcT3, the Km of T3 uptake did not change significantly; however Vmax was 54% lower (P less than 0.025) indicating a noncompetitive inhibition of the transport system. Preincubation of the cells with N-acetyl-T3 does not alter the characteristics of uptake of T3 by rat hepatocytes as compared to controls, indicating that no binding of this compound occurs. It is concluded that preincubation of hepatocytes with BrAcT3 diminished I- formation from T3; 50% of this inhibition is due to decreased membrane transport and 50% by reduction of deiodination. Inhibition of membrane transport by BrAcT3 is substantiated by a 54% lower Vmax without a significant change in Km as compared to control. The effect of transport of thyroid hormone on metabolism stresses the importance of the membrane carrier in the translocation process.

Affinity Labels↗