Low density lipoproteins of atherosclerotic patients have an increased potential to raise intracellular calcium in vascular smooth muscle cells.
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Animals↗
Biomedical subjects
R Locher
Publications and source records attributed to R Locher.
Oxidized low-density lipoproteins in atherogenesis: possible mechanisms of action.
The increased atherogenic potential of oxidized low-density lipoprotein (ox-LDL) is well documented. In the present study, we investigated possible mechanisms of action of the difference to native LDL. In vitro oxidation of LDL was determined by measurement of thiobarbituric acid-reacting substances and absorption at 234 nm. Copper (5 mumol/L) induced significant (p less than 0.01) oxidation in vitro. Furthermore, LDL isolated from atherosclerotic patients was slightly but significantly (p less than 0.05) more oxidized than LDL from normal controls (2.81 +/- 0.08 vs. 3.21 +/- 0.16 nmol of TBARS/mg of LDL protein). Ox-LDL caused significantly (p less than 0.01) more pronounced contractions of rat aortic rings in vitro compared to nonoxidized LDL expressed as a percentage of maximal contractions induced by 40 mmol/L of KCl (29.0 +/- 5.4% vs. 61.1 +/- 7.2%). Lysolecithin, which is a principal component of ox-LDL formed during oxidation, induced a dose-dependent increase in intracellular free calcium in vascular smooth muscle cells cultured from rat aorta. Doses from 2-25 micrograms/ml were tested and caused a maximum increase of more than 500% (25 micrograms/ml). In conclusion, this study provides further evidence for a higher biological activity of ox-LDL. Lysolecithin might be one of the active components formed during oxidation of LDL.
Animals↗
Oxidation of low density lipoprotein enhances its potential to increase intracellular free calcium concentration in vascular smooth muscle cells.
There have been suggestions that oxidation of low density lipoproteins (LDL) might increase their atherogenic potential. Because changes in intracellular free calcium concentration [Ca2+]i have been linked to atherogenesis, we compared the influence of oxidized LDL (Ox-LDL) and native LDL (N-LDL) on [Ca2+]i in vascular smooth muscle cells cultured from rat aortas. For determination of [Ca2+]i, fura-2 fluorescence was used. LDL was isolated by ultracentrifugation from the sera of human donors (n = 17). In N-LDL, oxidation was prevented by addition of antioxidants, whereas Ox-LDL was obtained by auto-oxidation. The extent of oxidation was assessed by measurement of thiobarbituric acid-reactive substances. Addition of Ox-LDL (20 micrograms protein/ml) to the vascular smooth muscle cells induced a mean increase of 129 +/- 13% in [Ca2+]i compared with 81 +/- 7% with N-LDL (p less than 0.01). Dose-response curves from 1 to 20 micrograms/ml (six experiments) confirmed this difference within the entire dose range. These results indicate that a more pronounced increase in [Ca2+]i induced by Ox-LDL might be one of the cellular mechanisms responsible for the higher atherogenic potential of Ox-LDL compared with N-LDL, as [Ca2+]i is an important second-messenger system involved in many atherogenic processes such as hypertrophy, cell migration, and cell damage.
Animals↗
Potentiation of vascular smooth muscle cell activity by cyclosporin A.
Pretreatment of rat vascular smooth muscle cells with cyclosporin A caused concentration- and time-dependent enhancement of both angiotensin II- and platelet-derived growth factor-stimulated cellular functions, which may be related to a rise in vascular tone. In particular, cyclosporin A increased cytosolic free calcium, and augmented the agonist-induced formation of inositol polyphosphate. In addition, it markedly increased the sensitivity of the cells to platelet-derived growth factor-mediated stimulation of thymidine incorporation. The potentiation of vascular smooth muscle cell activity by cyclosporin A may be a novel mechanism by which it exerts an adverse vasoconstrictor effect.
Angiotensin II↗
Generation of intracellular signals by low density lipoprotein is independent of the classical LDL receptor.
Low density lipoprotein cholesterol (LDL) and apolipoprotein B-100 (1 to 15 micrograms/mL) had no significant influence on the inositol-1,4,5-trisphosphate (InsP3) formation in vascular smooth muscle cells and fibroblasts. Low density lipoprotein cholesterol (15 micrograms/mL) induced an elevation of intracellular Ca2+ from 85 to approximately 210 nmol/L in vascular smooth muscle cells from rat aorta in the absence or in the presence of 15 micrograms/mL monoclonal antibodies against the classical low density lipoprotein receptor or in the presence of apolipoprotein B-100. Moreover, in both human cultured fibroblasts from normocholesterolemic individuals and from patients with familial hypercholesterolemia homozygote class 1, LDL induced a dose-dependent rise of free intracellular calcium and a biphasic change of intracellular pH. Since homozygote class 1 fibroblasts are classical LDL receptor negative, and as antibodies against this receptor, as well as apolipoprotein B-100, did not attenuate the LDL-induced elevation of cytosolic calcium, we conclude that LDL might modify vascular activity via the observed intracellular changes without involving the classical low density lipoprotein receptor.
Animals↗
Intracellular free calcium concentration and thromboxane A2 formation of vascular smooth muscle cells are influenced by fish oil and n-3 eicosapentaenoic acid.
UNLABELLED: The effect of fish oil and n-3 eicosapentaenoic acid (EPA) on intracellular free calcium concentration ([Ca2+]i) and thromboxane B2 (TXB2) formation in resting and stimulated cultured rat vascular smooth muscle cells (VSMC) was examined. In resting control cells [Ca2+]i was 147 +/- 15 nmol l-1 (mean +/- SEM, n = 4). After pretreatment of the cells with fish oil or EPA for 24 days the resting [Ca2+]i was decreased to 126 +/- 10 nmol l-1 and 84 +/- 8 nmol-1, respectively. After stimulation of untreated control cells with either 100 nmol l-1 angiotensin II (AII), 40 micrograms ml-1 low-density lipoprotein (LDL), or 100 ng ml-1 of recombinant platelet-derived growth factor (PDGFAB), [Ca2+]i was (in nmol l-1) 306 +/- 31, 217 +/- 25 and 213 +/- 16. Treatment of cells with fish oil or EPA reduced the stimulatory effect of the agonists, and the following [Ca2+]i values (in nmol l-1) were found: 199 +/- 21, 131 +/- 10, 148 +/- 13; and 175 +/- 11, 98 +/- 12, and 103 +/- 6, respectively. PDGFAB induced a four fold increase in TXB2-generation (270 +/- 28 pg mg-1 cell protein compared with 61 +/- 8.2 pg mg-1 in unstimulated control cells) within 6 min. In cells pretreated with fish oil or EPA, TXB2-formation was reduced by 54% and 44%, respectively. IN CONCLUSION: in rat VSMC stimulated by a variety of vasoactive agonist, fish oil and EPA can markedly attenuate intracellular mechanisms related to changes of cytosolic calcium concentration and eicosanoid production.(ABSTRACT TRUNCATED AT 250 WORDS)
Angiotensin II↗
Oxidation of low-density lipoprotein increases vasoconstriction in vitro.
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Animals↗
The maximal stimulatory effect of low-density lipoprotein on the cytosolic free calcium ion depends on the presence of its lipid and protein component.
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Animals↗
The platelet-derived growth factor isomers, PDGF-AA, PDGF-AB and PDGF-BB, induce contraction of vascular smooth muscle cells by different intracellular mechanisms.
The effect of human recombinant platelet-derived growth factor (PDGF) isoforms, (r)PDGF-AA, PDGF-AB and PDGF-BB, on contractility of rat aortic rings as well as on intracellular free Ca2+ ([Ca2+]i), intracellular pHi (pHi) and thromboxane A2 (TXA2) formation in cultured vascular smooth muscle cells (VSMC) was examined. PDGF-BB behaved similar to PDGF-AB and both have features characteristic of conventional vasoconstrictor-agonists that directly increase [Ca2+]i, activate the Na+/H+ exchanger, stimulate the TXA2 formation, and induced contraction in VSMC whereas PDGF-AA induced contraction without increasing of [Ca2+]i, pHi, and TXA2 formation.
Animals↗
Different effects of platelet-derived growth factor isoforms on rat vascular smooth muscle cells.
The response of rat aortic smooth muscle cells to all three isoforms of platelet-derived growth factor (PDGF) was studied. 5,000 binding sites/cell were estimated for rPDGF-AA (Kd 0.22 nM), 45,000 for rPDGF-AB and (Kd 0.4 nM), and 31,000 for rPDGF-BB (Kd 0.29 nM). rPDGF-AB and -BB stimulated effectively [3H]thymidine incorporation, inositol 1,4,5-trisphosphate release, diacylglycerol productions, [Ca2+]i increase, and pHi changes at concentration in the range from 3 to 10 ng/ml. The extent of DNA synthesis stimulated by rPDGF-AA varied considerably, and in all cases higher concentrations than 10 ng/ml were required. rPDGF-AA did not stimulate inositol-1,4,5-trisphosphate release, [Ca2+]i increase or pHi changes but induced the production of diacylglycerol, although with a different kinetic compared with that observed with rPDGF-AB or -BB. Apparently rPDGF-AA acts via a different mechanism, generating diacylglycerol without the release of inositol-1,4,5-trisphosphate.
Animals↗
Low-density lipoprotein elevates intracellular calcium and pH in vascular smooth muscle cells and fibroblasts without mediation of LDL receptor.
Low-density lipoprotein (7 micrograms/ml) induced in the absence or in the presence of 7, 35, 70 micrograms/ml monoclonal antibodies against the specific Low-density lipoprotein receptor an elevation of intracellular Ca2+ from 105 to approximately 210 nM in vascular smooth muscle cells from rat aorta. Moreover, in both human cultured fibroblasts from normocholesterolemic individuals and from patients with familial hypercholesterolemia homozygote class 1, Low-density lipoprotein (7 micrograms/ml) induced a rise of free intracellular calcium and a biphasic change of intracellular pH. Low-density lipoprotein (1,7,15,30 micrograms/ml) had no significant influence on the phosphatidylinositol-turnover in vascular smooth muscle cells and fibroblasts. Since homozygote class 1 fibroblasts lack specific Low-density lipoprotein receptors, and as antibodies against this receptor did not attenuate the Low-density lipoprotein-induced elevation of cytosolic calcium and pH, we conclude that these intracellular changes are independent from the classical Low-density lipoprotein receptor.
Calcium↗
Attenuation of the low-density-lipoprotein-activated phosphoinositide signalling system by calcium blockers in human lymphocytes.
1. The capacity of low-density lipoprotein-cholesterol (40 micrograms/ml) to stimulate both the phosphoinositide cycle and the dislocation of the phospholipid- and calcium-dependent protein kinase C activity from the cytosolic to the membranous compartment in human circulating lymphocytes has been studied. 2. Pretreatment of lymphocytes with pertussis toxin inhibited significantly the low-density-lipoprotein-induced formation of inositol trisphosphate. 3. The calcium-channel blockers verapamil, nifedipine and diltiazem, each in a concentration range from 1 mumol/l to 1 nmol/l, attenuated the low-density lipoprotein-induced formation of inositol trisphosphate in a dose-dependent manner. Similar results were observed when so-called non-specific calcium antagonists, such as flunarizine and molsidomine, were used. 4. The results suggest that low-density lipoprotein activates the phosphoinositide cycle in circulating lymphocytes through mechanisms sensitive to calcium blockers and pertussis toxin.
Calcium Channel Blockers↗
Novel cellular activities for low density lipoprotein in vascular smooth muscle cells.
Hyperlipidemia and hypertension play important roles in the pathogenesis of atherosclerosis. To investigate the underlying intracellular mechanisms, we studied the effect of various concentrations of low density lipoprotein from normolipidemic subjects on concentrations of free intracellular calcium, intracellular pH, DNA synthesis, and vascular tone in vascular smooth muscle cells and rings from rat aortas. Low density lipoprotein in the range of 1-15 micrograms/ml induced a dose-dependent increase of concentration of free intracellular calcium and a biphasic change of the intracellular pH. Similar concentrations of low density lipoprotein led to an enhanced DNA synthesis. Furthermore, cumulative addition of 1-15 micrograms/ml low density lipoprotein produced a dose-dependent increase in contractile tension of thoracic aortic rings from rats. The maximal low density lipoprotein-induced contractile response was approximately 70% of that induced by 40 mM KCl. These findings indicate that low concentrations of low density lipoprotein occurring, for example, in the extravascular fluid might contribute to the pathogenesis of cardiovascular diseases by enhancing cell proliferation and vasoconstriction by changing intracellular calcium and intracellular pH.
Animals↗
In vitro activity of oral cephalosporin BAY v 3522 compared with other oral cephalosporins.
Minimal inhibitory concentrations (MICs) of the oral cephalosporin BAY v 3522, and of cephprozyl, cefaclor, cefixime, cefuroxime, cefetamet, cefpodoxime and cefotaxime were determined against Gram-positive and Gram-negative clinical isolates with the NCCLS agar dilution procedures. BAY was the most active drug against Gram-positive organisms. MICs ranged from 0.01 mg/l against group A streptococci to 16 mg/l against S. faecium. Although mean MICs of BAY against methicillin-resistant S. aureus and S. epidermidis were between 0.9-1.8 mg/l, respectively, such strains showed typical heteroresistance in population studies. In addition, the biochemical correlate of methicillin-resistance, the PBP-2', showed similar low affinity to BAY as methicillin. beta-lactamase-producing H. influenzae and B. catarrhalis were inhibited by 2-8 and 0.25-2 mg/l, respectively, whereas non-producers were inhibited by 0.25-2 and 0.12-1 mg/l of the drug. The activity of BAY against enterobacteriaceae was rather low. Ampicillin-susceptible E. coli strains were inhibited by 2-8 and resistant strains by 8-32 mg/l. The mean MIC against cephalothin-susceptible K. pneumoniae strains was 2.8, and that against resistant strains 27.4 mg/l. MICs against beta-lactamase-producing enterobacteriaceae determined in broth dilution were 4-8 times higher than those determined in agar dilution. Bactericidal activity was measured in killing-curve experiments at 4 times the MIC. BAY killed equally well as standard control drugs.
Administration, Oral↗
Induction of cell proliferation and vasoconstriction by low density lipoprotein.
Hyperlipidemia and hypertension are risk factors for cardiovascular diseases. To obtain insight into intracellular mechanisms underlying these phenomenon, the influence of low density lipoprotein (LDL) on the intracellular free calcium concentration, the intracellular pH, the calcium influx as well as proliferation rate of cultured vascular smooth muscle cells was studied. 1-15 micrograms/ml of LDL increased dose-dependently the concentration of intracellular free calcium and led to biphasic pH-shifts. 3-7 micrograms/ml of LDL augmented thymidine incorporation into cell DNA by 70% and enhanced calcium influx by 80%. LDL (1-15 micrograms/ml) clearly increased contractile response of aortic rings. The findings indicate that low concentrations of LDL may contribute to the pathogenesis of cardiovascular diseases by enhancing cell proliferation and vasoconstriction via changing intracellular calcium and intracellular pH.
Animals↗
Vasoconstriction: a novel activity for low density lipoprotein.
Low density lipoprotein plays an important role in the pathogenesis of atherosclerosis. Cumulative addition of 1-30 micrograms/ml of LDL from normolipidemic subjects produced a dose-dependent increase in contractile tension of thoracic aortic rings from rats. The maximal LDL-induced contractile response was approximately 30% of that induced by 1 microM norepinephrine. Similar concentrations of LDL induced a dose-dependent transient increase of the concentration of intracellular free calcium, and a biphasic change of the intracellular pH in cultured rat vascular smooth muscle cells. We conclude that low density lipoprotein occurring for example in the extravascular fluid can mediate vasoconstriction by changes in cytosolic calcium and intracellular pH.
Animals↗
Fish oil affects phosphoinositide turnover and thromboxane A metabolism in cultured vascular muscle cells.
Fish oil has been reported as having beneficial effects on cardiovascular diseases. Elevated serum lipoproteins, prostaglandins and intracellular free calcium concentrations [( Ca2+]i) of the vasculature and thus the phosphoinositide (PI) turnover may be involved in the pathogenesis of these disorders. Therefore, the effect of fish oil on the potency of both low-density lipoprotein (LDL) and angiotensin II (AII) to stimulate the PI turnover in cultured rat vascular smooth muscle cells (VSMC) has been studied. Furthermore, a possible link between PI turnover activity and thromboxane A2 (TXA2) metabolism in these cells has been investigated. In VSMC cultured for up to 7 weeks with either fish oil or n-3 eicosapentaenoic acid (EPA) a decrease to 5-48% of the LDL-induced inositol trisphosphate (IP3) formation (= 100%) was found. A similar range of decreased IP3 synthesis was observed, when AII was used instead of LDL. Both LDL- and AII-stimulated TXA2 synthesis was suppressed concomitantly within the range 34-60%. Blockade of VSMC TXA2 biosynthesis with either indomethacin or TXA2 synthetase blocker (SQ-80338) inhibited LDL-induced formation of IP3 in a dose-dependent manner. Similar results were obtained, when TXA2 receptor coupling antagonists (SQ-27427 or BM-13177) were used. However, blockers of TXA2 synthesis and of TXA2 receptor binding failed to affect AII-induced formation of IP3.
Angiotensin II↗
Effect of low-density lipoprotein on intracellular calcium, intracellular pH and DNA synthesis in cultured vascular smooth muscle cells.
Low-density lipoprotein has proliferative effects on vascular smooth muscle cells and thus may play a role in the pathogenesis of atherosclerosis and hypertension. In order to investigate the underlying mechanisms, we studied the influence of low-density lipoprotein on intracellular free calcium, intracellular pH and the rate of DNA synthesis in cultured vascular smooth muscle cells of the rat aorta. The low-density lipoprotein (1-30 micrograms/ml) caused a dose-dependent increase in intracellular calcium and affected the intracellular pH accordingly. In addition, a marked low-density lipoprotein-induced increase in 3H-thymidine incorporation into DNA was observed. It is concluded that low-density lipoprotein, apart from its classical role as a cholesterol-transporting vehicle, enhances cellular processes leading to cell proliferation.
Animals↗