[Contraindications to DOACs in atrial fibrillation]. / Contra-indicaties voor DOAC's bij atriumfibrilleren.

Sometimes there is doubt as to whether or not anticoagulants should be initiated, and if so which ones, in patients with atrial fibrillation and advanced age, increased frailty, or fall risk, kidney, or liver impairment, alcohol abuse, uncontrolled hypertension, or a history of major bleeding. These subgroups have increased risk of haemorrhage as well as thromboembolism. Treatment with anticoagulants is indicated in the vital elderly, preferably with direct oral anticoagulants as demonstrated by robust data. The available study results for the other subgroups may not be (fully) generalisable to clinical practice. In such patients, a comprehensive risk assessment is therefore advised; as is discussing the pros and cons of (not) using anticoagulants and of both type of anticoagulants. Only in exceptional cases is it justified not to use anticoagulants.

CSF neurofilament proteins in the differential diagnosis of dementia.

BACKGROUND: Neurofilament (NF) proteins are major cytoskeletal constituents of neurons. Increased CSF NF levels may reflect neuronal degeneration. OBJECTIVE: To investigate the diagnostic value of CSF NF analysis to discriminate in relatively young dementia patients between frontotemporal lobe degeneration (FTLD) and early onset Alzheimer's disease (EAD; onset < or = 65 years of age), and in elderly dementia patients between dementia with Lewy bodies (DLB) and late onset AD (LAD; onset > 65 years of age). METHODS: In CSF of 28 FTLD, 37 EAD, 18 DLB and 33 LAD patients, and 26 control subjects, we analysed NF light chain (NFL), phosphorylated NF heavy chain (pNFH), amyloid beta42 protein (Abeta42), total tau and tau phosphorylated at threonine 181 (p-tau181). RESULTS: CSF NFL levels were higher in FTLD patients compared with EAD patients (p<0.001), and diagnostic accuracy of p-tau181 and Abeta42 analysis improved with addition of NFL analysis (sensitivity 86%, specificity 100%). CSF pNFH levels were elevated in DLB, LAD and FTLD compared with controls (p<0.05) but no significant differences were found between the dementia groups. CONCLUSIONS: In the diagnostic workup of relatively young dementia patients, CSF NFL levels may play a role in the discrimination between FTLD and EAD, especially in combination with Abeta42 and p-tau181 analysis.

Reproducibility of cerebral blood volume measurements by near infrared spectroscopy in 16 healthy elderly subjects.

Near infrared spectroscopy (NIRS) is a non-invasive method to monitor cerebral haemodynamics. Used either alone or in combination with other non-invasive methods such as transcranial Doppler sonography, this technique is well suited for use in cerebrovascular research in ageing. Reproducibility of NIRS, however, has only been determined in neonates and adults. We applied controlled desaturation (the O(2)-method) to measure the cerebral blood volume (CBV) with NIRS in 16 healthy subjects aged 65 to 88. This method uses deoxygenated haemoglobin (the concentration of which is manipulated by desaturation) as an intravascular tracer for NIRS. We determined repeatability (between tests interval: 2 min), short-term reproducibility (intervals of 20 and 40 min) and long-term reproducibility (interval > 2 weeks). We found a coefficient of variation (CV) of 12.5% for repeatability and a CV of 11.7% for short-term reproducibility. The CV for long-term reproducibility was 15%. We conclude that NIRS can reproducibly measure CBV in subjects aged 65 and older, using the O(2)-method. In this group of healthy subjects, this method was well tolerated.

[Elderly heart failure patients and the role of beta-blocker therapy]. / De oudere patiënt met hartfalen en de rol van bètablokkers bij de behandeling.

In this article different aspects of chronic heart failure in old age are described. We mainly focus on the place of beta-blocker therapy in chronic heart failure. Beta-blockers are recommended for the treatment of stable chronic heart failure with left ventricular systolic dysfunction. There is additional information from recent studies that there is proven efficacy for beta-blocker therapy in patients with heart failure up to the age of 80 years. For patients with heart failure aged 80 and over the evidence to prescribe beta-blockers is limited. However, it is known that also in very elderly patients beta-blocker therapy is well tolerated. In patients with heart failure with preserved systolic ventricular function there is still no evidence that there is a beneficial effect of beta-blockers. It is still not clear if there are differences between beta-blocking agents. Of all beta-blockers, only bisoprolol, carvedilol, nebivolol and metoprolol CR are proven effective in stable chronic heart failure with impaired left ventricular systolic function and can be recommended in elderly patients on standard treatment with diuretics and ACE inhibition.

Exclusion of venous thromboembolism: evaluation of D-Dimer PLUS for the quantitative determination of D-dimer.

The objective of this study was to evaluate if D-Dimer PLUS (Dade Behring, USA), a rapid fully automated assay, could be used as an initial screening test in the diagnosis of venous thromboembolism (VTE). Samples from 274 consecutive symptomatic patients with suspected pulmonary embolism (n=229; 79% outpatients, 21% inpatients), deep venous thrombosis (n=37; 84% outpatients, 16% inpatients) or suspected for both complications (n=8) were tested with this D-dimer assay with a Sysmex CA-1500 Coagulation Analyzer. Clinical probability for pulmonary embolism (PE) or deep venous thrombosis (DVT) was staged according to a pretest risk score proposed by Wells. Final diagnosis of PE and/or DVT was established by spiral-computed tomography of the pulmonary arteries or compression ultrasonography, respectively. PE was diagnosed in 13.5% of the patients, whereas DVT was confirmed in 17.7% of the patients. The optimal cut-off value for exclusion of venous thromboembolism was 130 mug/l, and sensitivity, specificity and negative predictive value (NPV) were 95.0% (95% CI: 92.4-97.6), 30.4% (95% CI: 25.0-35.8) and 97.2% (95% CI: 95.2-99.2), respectively. In fact, two patient with PE were missed using D-Dimer PLUS; both cases were outpatients. In conclusion, this assay appears to be safe when implemented in an algorithm based on clinical assessment, D-dimer concentration, and radiological diagnostic techniques to stratify the risk for PE or DVT. However, higher sensitivities and negative predictive values were claimed in the scarce published reports for the D-Dimer PLUS assay than found in this study.

Furosemide withdrawal improves postprandial hypotension in elderly patients with heart failure and preserved left ventricular systolic function.

OBJECTIVE: To assess the effects of furosemide withdrawal on postprandial blood pressure (BP) in elderly patients with heart failure and preserved left ventricular systolic function. METHODS: Noninvasive measurement of blood pressure (BP) and heart rate, computation of stroke volume and cardiac output (after a 1247-kJ (297-kcal) meal, and Doppler echocardiography before and 3 months after placebo-controlled withdrawal of furosemide therapy. RESULTS: Of 20 patients with heart failure (mean+/-SEM age, 75+/-1 years; left ventricular ejection fraction, 61%+/-3%), 13 were successfully able to discontinue furosemide therapy. At baseline, 11 (55%) of the 20 patients (had maximum postprandial systolic BP declines of 20 mm Hg or more. In the withdrawal group, the maximum systolic BP decline lessened from -25+/-4 to -11+/-2 mm Hg (P<.001) and the maximum diastolic BP from -18+/-3 to -9+/-1 mm Hg (P= .01), compared with no changes in the continuation group. In the withdrawal group, maximum postprandial declines in stroke volume and cardiac output decreased from -9+/-1 to -4+/-2 mL (P =.01) and from -0.6+/-0.2 to -0.2+/-0.1 L/min) (P = .04), respectively. The baseline maximum postprandial systolic BP decrease was correlated with the ratio of early to late flow (n = 20; Spearman rank correlation coefficient, 0.58; P = .007). For patients in the withdrawal group, the changes in postprandial systolic BP response were independently related to changes in peak velocity of early flow (n = 13; r2= 0.61; P = .003). CONCLUSIONS: Postprandial hypotension is common in elderly patients with heart failure and preserved left ventricular systolic function. The withdrawal of furosemide therapy ameliorates postprandial BP homeostasis in these patients, possibly by improving left ventricular diastolic filling.

Postprandial hypotension in elderly patients with unexplained syncope.

BACKGROUND: Syncope in older patients may be caused by a variety of disorders, including hypotension, but frequently remains unexplained. Postprandial hypotension is a common disorder of blood pressure regulation in the elderly. OBJECTIVE: To determine the pathogenic mechanisms and potential role of postprandial hypotension in elderly patients with otherwise unexplained syncope. METHODS: We studied 16 elderly patients with unexplained syncope and nine elderly controls. Blood pressure, heart rate, forearm vascular resistance, plasma norepinephrine level, and cardiac and splanchnic blood volumes were measured before and after a 1680-kJ meal. RESULTS: Eight elderly patients with syncope had postprandial hypotension, with a decline in supine mean arterial blood pressure of 17 +/- 2 mm Hg after a meal (P < .001). The blood pressure remained unchanged after the meal in the other patients with syncope and the controls. In patients with postprandial hypotension, systemic vascular resistance fell after the meal, while it remained unchanged in the other groups. Heart rate and plasma norepinephrine level increased to a similar extent in all three groups. Forearm vascular resistance increased only in the control subjects. Splanchnic blood volume increased by 26% (P < .01) in patients with syncope who had postprandial hypotension and by 22% (P < .01) in control subjects. Splanchnic blood volume remained unchanged in the patients with syncope without postprandial hypotension. CONCLUSIONS: Postprandial hypotension may be an important causative factor in elderly patients with unexplained syncope. The evaluation of syncope in elderly patients should therefore include blood pressure measurements surrounding a meal. Elderly patients with syncope who have postprandial hypotension fail to maintain systemic vascular resistance, probably because of splanchnic blood pooling without a compensatory increase in peripheral vascular resistance.

Lower body positive pressure by anti-G garment inflation: a suitable method to increase pulmonary capillary wedge pressure in healthy elderly subjects.

In a study on non-invasive assessment of pulmonary capillary wedge pressure (PCWP), we sought a method to increase PCWP non-invasively. We hypothesized that inflation of an anti-G garment was suitable to increase PCWP non-invasively in healthy elderly subjects. In 20 subjects, aged 70 +/- 4 years (mean +/- SD), before, immediately after, and 4 min after anti-G garment inflation to 52 mmHg, PCWP and mean pulmonary artery pressure (MPAP) were measured with a Swan-Ganz catheter, and mean arterial blood pressure (MAP) with Finapres, in supine and semi-recumbent position. Supine, PCWP (mmHg, mean +/- SD) increased from 9.9 +/- 2.1 to 15.5 +/- 3.9** immediately after inflation and 13.4 +/- 3.7** at 4 min; semi-recumbent from 8.9 +/- 2.0 to 17.5 +/- 3.3** and 14.7 +/- 2.9** (*P<0.05, **P< 0.001 versus before inflation). MPAP (mmHg) increased after inflation: supine 16.9 +/- 2.3 to 22.3 +/- 4.6** and 20.6 +/- 3.9** and semi-recumbent 15.7 +/- 2.8 to 24.3 +/- 5.1** and 22.5 +/- 3.5**, suggesting that increased preload was the primary effect of anti-G garment inflation. Supine MAP (mmHg) increased from 96.0 +/- 11.3 to 101.4 +/- 13.4** and 100.5 +/- 12.7* and semi-recumbent from 102.0 +/- 8.9 to 108.3 +/- 11.4** and 106.0 +/- 11.3*, suggesting an effect of increased afterload as well. The latter was supported by an increase in total peripheral resistance (d s cm(-5)) from 1346 +/- 299 to 1441 +/- 384 after 4 min (P = 0.057) and from 1461 +/- 341 to 1532 +/- 406 (P = 0.054), supine and semi-recumbent respectively, while cardiac output remained unchanged. Complications did not occur. We conclude that in healthy elderly subjects, anti-G garment inflation is a safe, non-invasive, method to induce a significant increase in PCWP. Our findings justify its application in future studies in which non-invasive temporary increase in PCWP is required.

Normal mammary cells in long term culture. I. development of hormone-dependent functional monolayer cultures and assay of alpha-lactalbumin production.

Mammary cells from normal tissue of virginal, pregnant, and lactating rats have been adapted to long term monolayer culture in plastic culture dishes with retention of hormone-responsive functional activity. The addition of PRL, insulin, and corticosterone resulted in an increase in the proportion of epithelial cells. the development of intracellular lipid droplets, and the ordered aggregations of these cells. In the presence of these hormones, the milk protein, alpha-lactalbumin (a-LA), was secreted into the growth medium at rates of 20-100 ng/mg cellular protein . 24 h. A double antibody RIA for a-LA capable of measuring 0.1 ng a-LA/100 microliter growth medium was developed in our laboratory for these studies. Both intracellular and extracellular a-LA fell below detectability within 2-3 weeks after hormone withdrawal. Intracellular a-LA accounted for less than 3% of the total a-LA accumulated in each culture in 24 h. the production rate of cells continuously given hormones increased 4- to 7-fold over a period of several months in culture, and their output was greater than 100-fold above that of cells not given hormones. These cells were obtained by overnight digestion and dispersion of tissue using selected batches of collagenase in the presence of 5% fetal calf serum. Plating densities of at least 3 X 10(4) cells/cm2 in Minimum Essential Medium supplemented with 14% fetal calf serum were required for optimal functional activity. Despite several months without added hormones, these cultures can retain their hormone responsiveness, since subsequent hormone addition resulted in detectable a-LA production beginning within 7-14 days. Our studies demonstrate for the first time that normal mammary cells can be maintained in a functional hormone-responsive state for extended periods in primary cell culture. These long term cell cultures provide a system with which the effects of these and other hormones on milk production and cell differentiation can be assessed under conditions which minimize the influence of the prior in vivo hormonal millieu. (Endocrinology 108: 573, 1981)

A complex noncoordinate regulation of alpha-lactalbumin and 25 K beta-casein by corticosterone, prolactin, and insulin in long term cultures of normal rat mammary cells.

The concentrations of PRL, corticosterone, and insulin required by long term cultures of normal rat mammary cells to produce alpha-lactalbumin (alpha LA) and the 25,000 mol wt beta-casein were evaluated with a variety of hormone ratios and concentrations. For these studies a double antibody RIA for beta-casein capable of measuring 0.5 ng beta-casein/100 microliter growth media was developed and used along with our previously reported RIA for alpha LA. PRL was active at physiological levels (0.05-0.15 micrograms/ml) and quantitatively stimulated beta-casein more than alpha LA, whereas physiological levels of corticosterone (0.05-0.15 micrograms/ml) quantitatively stimulated alpha LA more than beta-casein. The concentration of corticosterone greatly altered the magnitude of the cells' response to insulin and PRL for alpha LA output by cells from either virginal or midpregnant rats. Insulin also enhanced production of these milk proteins, but very little effect was measured in the physiological range. alpha LA was increased more by insulin than by PRL, and beta-casein was enhanced more by insulin than by corticosterone. Cells from midpregnant rats required less insulin to stimulate beta-casein production than to stimulate alpha LA. Cells from virginal rats required a supraphysiological insulin level to stimulate both beta-casein and alpha LA under these conditions. These cells generally require 5-6 weeks to achieve a steady-state rate of milk protein output. The complexities of our observations help explain some of the conflicting reports in the literature concerning which hormone is of prime importance for quantitatively increasing the synthesis of a particular milk protein, particularly since high hormone levels are often employed and time in culture varies considerably among reports. We conclude that lower levels of all these hormones can and should be used in vitro. Our messenger RNA (mRNA) studies using cloned complementary DNA probes for two rat casein mRNAs show that cells grown for 2 months with hormones contain significant amounts of both alpha- and beta-casein mRNAs. Simultaneous quantification of beta-casein mRNA levels and rates of beta-casein protein production in these long term cell cultures indicated that a substantial portion of their beta-casein protein production is regulated by the amount of its mRNA. This could be controlled by mRNA synthesis and/or mRNA degradation.(ABSTRACT TRUNCATED AT 400 WORDS)

The influence of glycyrrhetinic acid on plasma cortisol and cortisone in healthy young volunteers.

Based on studies in laboratory animals and on measurements of the urinary metabolites (allo)tetrahydrocortisol and tetrahydrocortisone in human volunteers it has been claimed that liquorice-induced mineralocorticoid excess is caused by a unique defect in the conversion of cortisol to cortisone. To further evaluate this hypothesis we have investigated the influence of glycyrrhetinic acid (GA), the mineralocorticoid-active constituent of liquorice, on plasma cortisol and cortisone in 10 healthy young normotensive volunteers. Pure GA (500 mg/day), administered orally from days 3-10 of the study, exerted pronounced mineralocorticoid activity. Ingestion of GA resulted in an elevated urinary excretion of free cortisol and virtually unchanged plasma cortisol levels in the presence of markedly decreased levels of both plasma cortisone and urinary free cortisone. These results provide direct clinical support for the hypothesis that GA induces an inhibition of the activity of 11 beta-dehydrogenase, resulting in a blockade in the conversion of cortisol to cortisone.

Somatostatin analog octreotide (SMS 201-995) prevents the decrease in blood pressure after oral glucose loading in the elderly.

In elderly subjects blood pressure (BP) may fall after a meal. The mechanism of this phenomenon is unknown, but it has been suggested that it may be mediated by insulin and/or vasoactive gut hormones. We studied in normo- and hypertensive elderly subjects the effects of the synthetic long-acting somatostatin analog octreotide (SMS 201-995) on the BP reduction that follows oral glucose administration in subjects who are recumbent and on their postglucose plasma vasoactive intestinal polypeptide (VIP) and insulin concentrations. After placebo treatment, mean arterial pressure fell by 15 +/- 1 mm Hg (P less than 0.001) in the 10 hypertensive subjects and by 7 +/- 2 mm Hg (P less than 0.01) in the 10 normotensive subjects. In contrast, when 50 micrograms octreotide were given sc, BP did not change significantly in either group. Oral glucose did not induce a rise in plasma VIP after either octreotide or placebo administration. The postglucose rises in plasma glucose concentrations were similar after octreotide and placebo treatments in both groups. After placebo administration the postglucose plasma insulin levels increased from 79 to 519 pmol/L in the hypertensive subjects and from 63 to 464 pmol/L in the normotensive subjects, whereas after octreotide treatment plasma insulin increased little in either group. These data indicate that treatment with octreotide holds promise for patients with symptomatic postprandial hypotension, and that VIP does not seem to play a role in this phenomenon.

Nitrendipine versus hydrochlorothiazide in hypertensive patients over 70 years of age.

The effects of nitrendipine and hydrochlorothiazide were studied in hypertensive elderly patients. Blood pressure was reduced (p less than 0.01) by both nitrendipine (13/10 +/- 4/3 mm Hg [n = 15], mean +/- SE) and hydrochlorothiazide (25/11 +/- 4/2 mm Hg [n = 16]). After hydrochlorothiazide, plasma glucose, uric acid, and renin activity increased and plasma potassium levels decreased. Edema and flushing were the main adverse reactions during nitrendipine. The response of blood pressure and heart rate to head-up tilt were not significantly different under both treatments. However, the effects of both drugs on diastolic blood pressure and norepinephrine responses to head-up tilt differed significantly. We conclude that, in the elderly, hydrochlorothiazide lowers systolic blood pressure more effectively than nitrendipine. However, nitrendipine does not have any of the potentially harmful metabolic side effects that were found during hydrochlorothiazide therapy. The clinical significance of a lower vasoreactivity during nitrendipine, as was found with the head-up tilt test, has to be established.

Neoglycoproteins as carriers for antiviral drugs: synthesis and analysis of protein-drug conjugates.

In order to investigate whether neoglycoproteins can potentially act as carriers for targeting of antiviral drugs to certain cell types in the body, various neoglycoproteins were synthesized using thiophosgene-activated p-aminophenyl sugar derivatives. These neoglycoproteins were conjugated with the 5'-monophosphate form of the antiviral drug AZT. For a proper characterization of these preparations, both protein and drug content have to be determined. Comparison of the Lowry and the Bio-Rad protein assays revealed that for both the neoglycoprotein carriers themselves and the AZTMP conjugates, the Lowry assay yielded the most reliable and reproducible results. It was demonstrated that both the reagent used for drug conjugation (ECDI) as well as the introduction of phenyl-sugar groups in the protein interfered with the analysis of bound nucleotide as based on spectral differences between protein and protein-drug conjugate. Therefore, we developed a rapid HPLC system for determination of the drug-protein coupling ratio through acid hydrolysis of the covalently bound nucleotide. With the ECDI-mediated conjugation of 5'-monophosphate drug derivatives to neoglycoproteins, products with molar ratios of drug to protein ranging from 1.2 to 5.6 were obtained. The drug-neoglycoprotein conjugates appeared to be fairly stable during storage, in lyophilized form, at -20 degrees C. The anti-HIV-1 activity of the neoglycoprotein-drug conjugates, as determined in vitro in MT-4 cells, was shown to be dependent on glycosylation of the albumin and also on the kind of sugar present in the neoglycoprotein. The anti-HIV-1 activity of the AZTMP-mannose-albumin conjugate exceeded that of the parent drug by more than 4 times.

Blood pressure reduction after oral glucose loading and its relation to age, blood pressure and insulin.

Recently it was shown that blood pressure (BP) in the elderly may decrease after a meal. The pathophysiologic mechanism of this phenomenon is unknown. It has been suggested that a failure of insulin-mediated sympathetic nervous system activation plays a role. To evaluate the role of endogenous insulin, the effects of oral glucose and oral fructose loading on BP, heart rate and norepinephrine levels were studied in 10 young normotensive volunteers (YN), 10 young hypertensive patients (YH), 10 elderly normotensive volunteers (EN) and 10 elderly hypertensive patients (EH). Fructose, 75 g/300 ml of water, elicited--in contrast to the same amount of glucose--only a small increase in insulin and glucose levels. After glucose loading, mean arterial BP decreased by 17 mm Hg in the EH group (p less than 0.001), 6 mm Hg in the EN group (p less than 0.01) and 7 mm Hg in the YH group (p less than 0.001), and did not change in the YN group. After oral fructose loading, BP did not change in any group. In all groups except the YN group, the increases in norepinephrine level and heart rate after both tests were not significantly different. These findings suggest that the BP reduction after glucose loading is related to glucose-mediated factors. A failure of insulin-mediated sympathetic nervous system activation does not appear to play a major role.

Furosemide withdrawal in elderly heart failure patients with preserved left ventricular systolic function.

To explore the possibilities of furosemide withdrawal in elderly heart failure (HF) patients with intact left ventricular (LV) systolic function and assess its effects on functional status and orthostatic blood pressure homeostasis, we performed a placebo-controlled pilot trial of furosemide withdrawal with 3 months of follow-up in 32 HF patients (aged 75.1 +/- 0.7 years [mean +/- SEM]) with a LV ejection fraction of 60 +/- 2% and without overt congestion. Investigations included repeated clinical assessment, spirometry, standardized 6-minute walking test, and chest x-rays. Measurements of blood pressure response on active standing and Doppler echocardiography were performed before and 3 months after furosemide withdrawal. Recurrent congestive HF occurred in 2 of 21 patients (10%) who discontinued furosemide use, and in 1 of 11 patients (9%) who continued furosemide (p = NS). Three patients restarted furosemide for ankle edema and 1 for blood pressure levels >180/100 mm Hg. After 3 months, there were no differences regarding HF symptom scores, blood pressure, heart rate, spirometric results, 6-minute walking distance, or quality of life scores between patients who discontinued use and patients who continued the therapy. In patients successfully withdrawn, Doppler E/A ratio increased from 0.68 +/- 0.05 to 0.79 +/- 0.06 after withdrawal (p <0.01), and maximum blood pressure decrease on active standing changed from -8 +/- 5 mm Hg to +5 +/- 3 mm Hg systolic (p <0.05). Thus, in this pilot investigation of furosemide withdrawal in elderly HF patients without overt congestion and with a normal LV systolic function, withdrawal was successful in almost all patients and was associated with improvement of LV diastolic filling and blood pressure homeostasis on active standing.

Hepatic and intrahepatic targeting of an anti-inflammatory agent with human serum albumin and neoglycoproteins as carrier molecules.

The anti-inflammatory agent naproxen (Nap) was covalently coupled to human serum albumin (HSA) and to the neoglycoproteins, galactose and mannose terminated HSA, to deliver this drug selectively to different cell types of the liver. Disposition of Nap20-HSA was studied in rats and compared to that of equivalent doses of mixtures of uncoupled drug and protein. The liver to kidney ratios of the drug (L/K-Nap) and the protein (L/K-prot.) were increased, indicating an improved delivery of both protein and drug to the target site. After injection of 10 micrograms Nap20-HSA the L/K-prot. was increased 15.0 +/- 0.21-fold as measured 1 hr after injection. Even after injection of 5 mg of the conjugate, the L/K-prot. was enhanced 5.6 +/- 0.34-fold and the L/K-Nap 4.6 +/- 0.23-fold as measured 1 hr after injection. Immunohistochemical staining of liver slices revealed that the endothelial cells were the main sites for hepatic uptake. Further pharmacokinetic studies of Nap20-HSA in isolated perfused rat livers showed a saturable uptake process (Vmax = 2.46 micrograms/min/10.0 g liver and Km = 4.27 x 10(-6) M). The uptake in the liver could be inhibited by various polyanionic probes, indicating the major involvement of a scavenger receptor system in the internalization mechanism of Nap20-HSA. This endothelial uptake via the scavenger receptor system is likely to be related to the increased negative charge of the Nap-albumin conjugate as was revealed by anion exchange chromatography. Studies in the intact organ and in purified liver lysosomal lysates indicate that after internalization of Nap20-HSA the conjugate is proteolytically degraded leading to the formation of the lysine conjugate of Nap. This amino acid conjugate of Nap was shown in a previous study by us to be equipotent to Nap itself with regard to prostaglandin-E2 synthesis inhibition. A pronounced altered intrahepatic distribution was observed when Nap was coupled to lactosaminated and mannosylated HSA (Lact-HSA and Man-HSA, respectively). Coupling of Nap to Lact27-HSA and Man10-HSA resulted in a major shift in intrahepatic distribution from endothelial cells to the hepatocytes and Kupffer cells, respectively. We conclude that conjugation of Nap to HSA itself results in a selective delivery to endothelial cells and that the local proteolysis of the conjugate produces an active catabolite. Selective delivery to other cell types of the liver can be achieved by attaching naproxen to neoglycoproteins with an appropriate type and number of sugar groups.

Targeting of antiviral drugs to T4-lymphocytes. Anti-HIV activity of neoglycoprotein-AZTMP conjugates in vitro.

The delivery of the anti-HIV agent 3'-azido-3'-deoxythymidine (AZT), in its 5'-monophosphate form, (in) to human T-lymphocyte MT-4 cells in vitro through covalent coupling to neoglycoproteins was investigated. In vivo application of this drug targeting concept may lead to increased efficacy and/or diminished side effects caused by AZT during the treatment of AIDS and ARC patients. The rationale for the design of the neoglycoprotein carriers is based on the existence of sugar recognizing lectins on T-lymphocytes. Using a phenyl-linkage between sugar and Human Serum Albumin (HSA), various mannose-, fucose-, galactose-and glucose-containing neoglycoproteins were synthesized. The intrinsic anti-HIV activity of these neoglycoproteins was tested in vitro in HIV-1 infected MT-4 cells. Only the derivative having 40 moles mannose per mole protein (Man40HSA) shows pronounced anti-HIV-1 activity itself. This effect may be caused by interference of the Man40HSA with the gp120-CD4 mediated virus/MT-4 cell interaction. After conjugation with AZTMP, the mannose- as well as the fucose- and galactose-containing conjugates exhibited a pronounced activity. Conjugates of glucose-HSA and HSA displayed much less activity in spite of the fact that drug loading was considerably higher, compared with the galactose, mannose and fucose derivatives. In the series of mannose-neoglycoproteins, the Man22HSA-AZTMP conjugate was shown to be more than 30 times as active against HIV-1 compared to HSA-AZTMP. Selectivity indices of Man7 and Man22HSA-AZTMP were exceeding the AZT and AZTMP indices, indicating that these conjugates possess a more selective action. Stability experiments indicate that the potent action of the galactose-, mannose- and fucose-HSA-AZTMP conjugates is not due to a complete extracellular hydrolysis of the covalent drug-protein bond. Since Man22HSA has no intrinsic activity in the concentration range used, the antiviral effect is unlikely to be explained by synergism of the neoglycoprotein by a component of the cell membrane and subsequent internalization and release of the drug from the conjugate may play a role.

The influence of oral glucose loading on baroreflex function in the elderly.

Blood pressure in the elderly may decrease after a meal or after oral glucose loading. It has been suggested that eating may affect blood pressure homeostasis through an insulin-induced blunting of baroreflex sensitivity. We investigated the effects of oral glucose loading on baroreflex sensitivity in young normotensives and in elderly normo-and hypertensive subjects. Blood pressure was measured by a new noninvasive device, Finapres, which measures blood pressure continuously in the finger. Baroreflex sensitivity was estimated with the phenylephrine and nitroglycerin method. In both elderly groups mean arterial pressure fell significantly after the glucose load (11 +/- 1 mm Hg, P less than .001 in the hypertensives, and 8 +/- 2 mm Hg, P less than .01 in the normotensive subjects), whereas no change in blood pressure was found in the young group. Baroreflex sensitivity was lower in both elderly groups than in young normotensives. Glucose loading had no influence on baroreflex sensitivity in the three groups. Therefore, we conclude that other factors are involved in the phenomenon of postprandial hypotension in the elderly.