Biomedical subjects

C Riethmüller

Publications and source records attributed to C Riethmüller.

7 recordsLinked to original sources

PubMed2006

Bradykinin shifts endothelial fluid passage from para- to transcellular routes.

The signalling peptide bradykinin (BK) is implicated in inflammation and angiogenesis. It promotes fluid transport from blood vessels to interstitial space, and thus facilitates oedema formation. To clarify whether paracellular or transcellular pathways mediate this effect, we investigated the BK-stimulated fluid transport across endothelial monolayers in vitro by comparison of electrical and fluorescence methods. Electrical cell impedance sensing monitored a biphasic response of cell layers to BK with high time resolution: a short decrease (18%, 1 min) was followed by a more sustained increase in paracellular resistance (30%, 10 min). The two phases can be assigned to second messengers of the BK-signalling pathway: Ca(2+) for the decrease and cyclic adenosine monophosphate for the rise of resistance, respectively. Despite tightening of the intercellular clefts, BK increased the fluid permeability by 39%, indicating transcellular fluid transport. Additionally, BK stimulated both in- and outwardly directed membrane trafficking as assessed by vesicular fluid uptake (by 49%) and secretion of von Willebrandt factor (by 85%). In conclusion, the combination of electrical and fluorescence data suggests that BK induces a shift from para- to transcellular fluid transport across endothelium.

Body Fluid Compartments↗

PubMed2006

17beta-estradiol increases volume, apical surface and elasticity of human endothelium mediated by Na+/H+ exchange.

OBJECTIVE: 17beta-estradiol is known to delay the onset of atherosclerosis in women but cellular mechanisms are still unclear. Estrogens bind to specific receptors and initiate a signaling cascade that involves the activation of plasma membrane Na(+)/H(+) exchange. We hypothesized that estrogens interfere with ion transport across the plasma membrane and thus control endothelial structure and function. Therefore, we investigated the effects of the sex steroids 17beta-estradiol, progesterone, and testosterone on volume, apical surface and elasticity in human endothelium. METHODS: The atomic force microscope was used as an imaging tool and as an elasticity sensor. We applied the antiestrogen tamoxifen, the Na(+)/H(+) exchange blocker cariporide and the epithelial Na(+)channel blocker amiloride to elucidate the role of transmembrane ion transport in hormone-treated human umbilical vein endothelial cells (HUVEC). RESULTS: Incubation with 17beta-estradiol for 72 h led to a dose-dependent increase of endothelial cell volume (41%), apical cell surface (22%), and cell elasticity (53%) as compared to non-17beta-estradiol treated controls. Block of the 17beta-estradiol receptor by tamoxifen and of plasma membrane Na(+)/H(+) exchange by cariporide prevented the hormone-induced changes. Progesterone and testosterone were ineffective. CONCLUSIONS: 17beta-estradiol increases HUVEC water content and HUVEC elasticity mediated by activated estrogen receptors. The estrogen response depends on the activation of plasma membrane Na(+)/H(+) exchange. The increase in endothelial cell elasticity could be one of the vasoprotective mechanisms postulated for 17beta-estradiol.

Amiloride↗

PubMed2003

Endothelial cell swelling by aldosterone.

There is accumulating evidence that mineralocorticoids not only act on kidney but also on the cardiovascular system. We investigated the response of human umbilical venous endothelial cells (HUVECs) to aldosterone at a time scale of 20 minutes in absence and presence of the aldosterone antagonist spironolactone or other transport inhibitors. We applied atomic force microscopy (AFM), which measures cell volume and volume shifts between cytosol and cell nucleus. We observed an immediate cell volume increase (about 10%) approximately 1 min after addition of aldosterone (0.1 micromol/l), approaching a maximum (about 18%) 10 min after aldosterone treatment. Cell volume returned to normal 20 min after hormone exposure. Spironolactone (1 micromol/l) or amiloride (1 micromol/l) prevented the late aldosterone-induced volume changes but not the immediate change observed 1 min after hormone exposure. AFM revealed nuclear swelling 5 min after aldosterone addition, followed by nuclear shrinkage 15 min later. The Na(+)/H(+) exchange blocker cariporide (10 micromol/l) was ineffective. We conclude: (i). Aldosterone induces immediate (1 min) swelling independently of plasma membrane Na(+) channels and intracellular mineralocorticoid receptors followed by late mineralocorticoid receptor- and Na(+)-channel-dependent swelling. (ii). Intracellular macromolecule shifts cause the changes in cell volume. (iii). Both amiloride and spironolactone may be useful for medical applications to prevent aldosterone-induced vasculopathies.

Adaptation, Physiological↗

PubMed2000

Nitric oxide-induced autoinhibition of neuronal nitric oxide synthase in the presence of the autoxidation-resistant pteridine 5-methyltetrahydrobiopterin.

Nitric oxide synthase (NOS) catalysis results in formation of NO or superoxide (O(2)(-.)) depending on the presence or absence of the cofactor tetrahydrobiopterin (BH4). In the absence of O(2)(-.) scavengers, net NO formation cannot be detected even at saturating BH4 concentrations, which is thought to be due to O(2)(-.) production by BH4 autoxidation. Because the N-5-methylated analogue of BH4 (5-Me-BH4) sustains NOS catalysis and is autoxidation-resistant, net NO formation by the neuronal isoform of NOS (nNOS) can be observed at saturating 5-Me-BH4 concentrations. Here we compare the effects of 5-Me-BH4 on L-citrulline formation, NADPH oxidation, H(2)O(2) production and soluble guanylate cyclase (sGC) stimulation. All activities were stimulated biphasically (EC(50) approx. 0.2 microM and more than 1 mM), with an intermediate inhibitory phase at the same pterin concentration as that required for net NO generation and sGC stimulation (4 microM). Concomitantly with inhibition, the NADP(+)/L-citrulline stoichiometry decreased from 2.0 to 1.6. Inhibition occurred only at high enzyme concentrations (IC(50) approx. 10 nM nNOS) and was antagonized by oxyhaemoglobin and by BH4. We ascribe the first stimulatory phase to high-affinity binding of 5-Me-BH4. The inhibitory phase is due to low-affinity binding, resulting in fully coupled catalysis, complete inhibition of O(2)(-.) production and net NO formation. At high enzyme concentrations and thus high NO levels, this causes autoinhibition. NO scavenging by 5-Me-BH4 at concentrations above 1 mM, resulting in the antagonization of inhibition of NOS, explains the second stimulatory phase. In agreement with these assignments 5-Me-BH4 was found to stimulate formation of a haem-NO complex during NOS catalysis. The observation of inhibition with 5-Me-BH4 but not with BH4 implies that, unless O(2)(-.) scavengers are present, a physiological role for NO-induced autoinhibition is unlikely.

Animals↗

PubMed1999

Activation of neuronal nitric-oxide synthase by the 5-methyl analog of tetrahydrobiopterin. Functional evidence against reductive oxygen activation by the pterin cofactor.

Tetrahydrobiopterin ((6R)-5,6,7,8-tetrahydro-L-biopterin (H4biopterin)) is an essential cofactor of nitric-oxide synthases (NOSs), but its role in enzyme function is not known. Binding of the pterin affects the electronic structure of the prosthetic heme group in the oxygenase domain and results in a pronounced stabilization of the active homodimeric structure of the protein. However, these allosteric effects are also produced by the potent pterin antagonist of NOS, 4-amino-H4biopterin, suggesting that the natural cofactor has an additional, as yet unknown catalytic function. Here we show that the 5-methyl analog of H4biopterin, which does not react with O2, is a functionally active pterin cofactor of neuronal NOS. Activation of the H4biopterin-free enzyme occurred in a biphasic manner with half-maximally effective concentrations of approximately 0.2 microM and 10 mM 5-methyl-H4biopterin. Thus, the affinity of the 5-methyl compound was 3 orders of magnitude lower than that of the natural cofactor, allowing the direct demonstration of the functional anticooperativity of the two pterin binding sites of dimeric NOS. In contrast to H4biopterin, which inactivates nitric oxide (NO) through nonenzymatic superoxide formation, up to 1 mM of the 5-methyl derivative did not consume O2 and had no effect on NO steady-state concentrations measured electrochemically with a Clark-type NO electrode. Therefore, reconstitution with 5-methyl-H4biopterin allowed, for the first time, the detection of enzymatic NO formation in the absence of superoxide or NO scavengers. These results unequivocally identify free NO as a NOS product and indicate that reductive O2 activation by the pterin cofactor is not essential to NO biosynthesis.

Animals↗

PubMed1997

Efficacy of antihistamine pretreatment in the prevention of adverse reactions to Hymenoptera immunotherapy: a prospective, randomized, placebo-controlled trial.

BACKGROUND: Some clinical studies suggest that a combination of an H1- and H2-antagonist may be effective in the prophylaxis of allergic reactions. OBJECTIVE: The efficacy of pretreatment with an H1/H2-antagonist combination, H1-antagonist alone, or placebo in the prophylaxis of local and systemic adverse reactions to specific immunotherapy with Hymenoptera venom was compared. METHODS: In a prospective, randomized, double-blind, placebo-controlled study, 121 patients with Hymenoptera venom allergy were treated with rush immunotherapy and pretreatment with one of the following: 120 mg of terfenadine plus 300 mg of ranitidine, 120 mg of terfenadine alone, or placebo. The incidence of unwanted systemic adverse and local reactions was recorded for up to 50 weeks. RESULTS: In seven patients (6%), six in the placebo group and one in the terfenadine group, systemic side effects required cessation of therapy (p = 0.005). Subjective symptoms occurred in four patients (10%) in the terfenadine plus ranitidine group and in three patients (7%) in the terfenadine group. Regarding local reactions, significantly fewer patients treated with a combination of terfenadine and ranitidine and with terfenadine alone as compared with placebo had severe local symptoms of erythema (29%, 29%, and 49%), edema (24%, 18%, and 41%), and pruritus (13%, 11%, and 31%) at week 1 (p < 0.05). This therapeutic benefit was limited to the first 4 weeks of treatment. Treatment with a combination of terfenadine and ranitidine was not superior to treatment with terfenadine alone. CONCLUSIONS: Pretreatment with H1-antihistamines with or without H2-antihistamines significantly reduced local and systemic adverse reactions to immunotherapy with Hymenoptera venom and may therefore be helpful in the management of immunotherapy.

Administration, Oral↗

PubMedDate not supplied

Aldosterone remodels human endothelium.

AIM: In response to aldosterone endothelial cells swell and stiffen. Although amiloride-sensitive sodium and water uptake is known to be involved, the underlying mechanisms are yet unclear. We tested the hypothesis whether the intracellular accumulation of water or organic matter is responsible for the structural and functional alterations. METHODS: Atomic force microscopy was used as an imaging tool and a mechanical nanosensor. Cell water, organic cell matter and cell pressure was measured at single cell level in human umbilical vein endothelial cells (HUVEC). Furthermore, we tested by means of a miniature perfusion chamber in vitro the physical robustness to blood flow of the aldosterone-treated endothelium. RESULTS: In response to a three-day treatment with 1 nM aldosterone HUVEC swell. To our surprise, cell water decreased from 82+/-6% to 71+/-5% while intracellular organic matter increased from 18+/-1.8% to 29+/-3.0%. These changes were paralleled by a rise in cell pressure of 114%, measured in living HUVEC in vitro. Blood flow across the endothelium was found significantly altered after aldosterone treatment. Imaging the endothelial monolayer after blood perfusion disclosed large gaps between cells treated with aldosterone. The mineralocorticoid receptor blockers, spironolactone and eplerenone could prevent the aldosterone actions. CONCLUSION: Mild aldosteronism causes intracellular accumulation of organic matter at the cost of cell water. This makes endothelium stiff and vulnerable to shear stress. The measurements could explain clinical observations that high blood pressure combined with high plasma aldosterone concentration may damage the endothelium of blood vessels.

Aldosterone↗