scholarly journals Inhibition of apolipoprotein B net synthesis and secretion from cultured rat hepatocytes by the calcium-channel blocker diltiazem

1989 ◽  
Vol 263 (2) ◽  
pp. 411-415 ◽  
Author(s):  
T C Kwong ◽  
J D Sparks ◽  
D J Pryce ◽  
J F Cianci ◽  
C E Sparks
Keyword(s):  
Molecular Mass ◽  
Apolipoprotein B ◽  
Channel Blocker ◽  
Rat Hepatocytes ◽  
Acetate Incorporation ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Apo B ◽  
Cellular Lipids ◽  
Mass Form

1. The effect of the Ca2+-channel blocker diltiazem on hepatic apolipoprotein B (apo B) synthesis and secretion was studied in 12-18 h cultures of collagenase-dispersed rat hepatocytes. 2. The presence of diltiazem in the medium decreased apo B secretion by hepatocytes in a concentration-dependent manner. At 25 microM, diltiazem inhibited apo B secretion by approx. 36%, but there was no evidence of intracellular accumulation of apo B. 3. The inhibition of apo B secretion by hepatocytes was significantly correlated with cell-associated diltiazem (r = 0.72, P less than 0.01). 4. The rate of apo B secretion remained linear over 16 h even in the presence of 50 microM-diltiazem. 5. At diltiazem concentrations in the medium which were inhibitory for apo B secretion, [14C]acetate incorporation into cellular lipids and [35S]methionine incorporation into protein were enhanced. 6. Diltiazem inhibited the secretion of the apo B variants with a preferential inhibition of the higher-molecular-mass form of apo B (apo BH) over the lower-molecular-mass form (apo BL) at diltiazem concentrations in the medium greater than 25 microM. 7. Together, these results suggest that Ca2+ may play an important role in the synthesis and secretion of apo B-containing lipoproteins.

Vasorelaxing Activity of Stilbenoid and Phenanthrene Derivatives from Brasiliorchis porphyrostele: Involvement of Smooth Muscle CaV1.2 Channels

Planta Medica ◽  
2020 ◽  
Vol 86 (09) ◽  
pp. 631-642
Author(s):  
Watcharee Waratchareeyakul ◽  
Fabio Fusi ◽  
Miriam Durante ◽  
Amer Ahmed ◽  
Walter Knirsch ◽  
...  
Keyword(s):  
Channel Blocker ◽  
Antihypertensive Agents ◽  
Dependent Manner ◽  
Vascular Effects ◽  
Concentration Dependent Manner ◽  
Spasmolytic Activity ◽  
Depth Analysis ◽  
Cav1.2 Channels ◽  
Vasorelaxant Activity

AbstractFive compounds, 3,4′-dihydroxy-3′,5,5′-trimethoxydihydrostilbene, 1; 3,4′-ihydroxy-3′,5′-dimethoxydihydrostilbene, 2; 3,4′-dihydroxy-5,5′-dimethoxydihydrostilbene, 3; 9,10-dihydro-2,7-dihydroxy-4,6-dimethoxyphenanthrene, 4; and the previously unreported 1,2,6,7-tetrahydroxy-4-methoxyphenanthrene, 5 were isolated from the South American orchid, Brasiliorchis porphyrostele. An in-depth analysis of their vascular effects was performed on in vitro rat aorta rings and tail main artery myocytes. Compounds 1 – 4 were shown to possess vasorelaxant activity on rings pre-contracted by the α 1 receptor agonist phenylephrine, the CaV1.2 stimulator (S)-(−)-Bay K 8644, or depolarized with high K+ concentrations. However, compound 5 was active solely on rings stimulated by 25 mM but not 60 mM K+. The spasmolytic activity of compounds 1 and 4 was significantly affected by the presence of an intact endothelium. The KATP channel blocker glibenclamide and the KV channel blocker 4-aminopyridine significantly antagonized the vasorelaxant activity of compounds 4 and 1, respectively. In patch-clamp experiments, compounds 1 – 4 inhibited Ba2+ current through CaV1.2 channels in a concentration-dependent manner, whereas neither compound 4 nor compound 1 affected K+ currents through KATP and KV channels, respectively. The present in vitro, comprehensive study demonstrates that Brasiliorchis porphyrostele may represent a source of vasoactive agents potentially useful for the development of novel antihypertensive agents that has now to be validated in vivo in animal models of hypertension.

scholarly journals Determination of the molecular mass of apolipoprotein B-100 A chemical approach

1986 ◽  
Vol 239 (3) ◽  
pp. 777-780 ◽  
Author(s):  
C Y Yang ◽  
F S Lee ◽  
L Chan ◽  
D A Sparrow ◽  
J T Sparrow ◽  
...  
Keyword(s):  
Molecular Mass ◽  
Apolipoprotein B ◽  
Cell Surface Receptor ◽  
Specific Cell ◽  
Apo B ◽  
Chemical Approach ◽  
Surface Receptor ◽  
Cnbr Cleavage ◽  
Specific Cell Surface

Apolipoprotein B-100 (apo B-100) is the protein ligand in low-density lipoproteins that binds to a specific cell-surface receptor. Its molecular mass has been a subject of controversy. We have determined the molecular mass of the protein by a chemical approach. After complete CNBr cleavage, the C-terminal fragment of apo B-100 was purified by reverse-phase h.p.l.c. Amino acid N- and C-terminal analyses confirm that this peptide represents the C-terminal peptide as deduced from the DNA sequence of a human apo B-100 cDNA clone. A chemically synthesized peptide was used to determine the recovery of the peptide (74.72%). On the basis of these data, the molecular mass of apo B-100 was determined to be 496.82 +/- 24.84 kDa.

scholarly journals Citrus bergamiaRisso Elevates Intracellular Ca2+in Human Vascular Endothelial Cells due to Release of Ca2+from Primary Intracellular Stores

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Purum Kang ◽  
Seung Ho Han ◽  
Hea Kyung Moon ◽  
Jeong-Min Lee ◽  
Hyo-Keun Kim ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Channel Blocker ◽  
Vascular Endothelial Cells ◽  
Umbilical Vein ◽  
Dependent Manner ◽  
Vascular Endothelial ◽  
Concentration Dependent Manner ◽  
Carbonyl Cyanide ◽  
Intracellular Ca ◽  
Umbilical Vein Endothelial Cells

The purpose of the present study is to examine the effects of essential oil ofCitrus bergamiaRisso (bergamot, BEO) on intracellular Ca2+in human umbilical vein endothelial cells. Fura-2 fluorescence was used to examine changes in intracellular Ca2+concentration[Ca2+]i. In the presence of extracellular Ca2+, BEO increased[Ca2+]i, which was partially inhibited by a nonselective Ca2+channel blocker La3+. In Ca2+-free extracellular solutions, BEO increased[Ca2+]iin a concentration-dependent manner, suggesting that BEO mobilizes intracellular Ca2+. BEO-induced[Ca2+]iincrease was partially inhibited by a Ca2+-induced Ca2+release inhibitor dantrolene, a phospholipase C inhibitor U73122, and an inositol 1,4,5-triphosphate (IP3)-gated Ca2+channel blocker, 2-aminoethoxydiphenyl borane (2-APB). BEO also increased[Ca2+]iin the presence of carbonyl cyanide m-chlorophenylhydrazone, an inhibitor of mitochondrial Ca2+uptake. In addition, store-operated Ca2+entry (SOC) was potentiated by BEO. These results suggest that BEO mobilizes Ca2+from primary intracellular stores via Ca2+-induced and IP3-mediated Ca2+release and affect promotion of Ca2+influx, likely via an SOC mechanism.

Pharmacological evidence for N-methyl-D-aspartate receptors on nigrostriatal dopaminergic nerve terminals

1991 ◽  
Vol 69 (10) ◽  
pp. 1416-1421 ◽  
Author(s):  
Kenneth M. Johnson ◽  
Yow-Juin Jeng
Keyword(s):  
Nmda Receptors ◽  
Phosphonic Acid ◽  
Channel Blocker ◽  
Nmda Antagonist ◽  
Nerve Terminals ◽  
Dependent Manner ◽  
Additional Mechanism ◽  
Concentration Dependent Manner ◽  
Nmda Channel ◽  
Da Release

The efflux of tritium from rat striatal synaptosomes labelled with [3H]dopamine was utilized as an index of dopamine (DA) release for the purpose of characterizing the receptors underlying the effects of L-glutamate. N-Methyl-D-aspartate (NMDA), α-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA), and kainate each induced DA release in the absence of Mg2+, though NMDA was much more efficacious and only the NMDA response was inhibited by Mg2+. The response to L-glutamate was potentiated in a concentration-dependent manner by glycine. Further, it was completely inhibited by the competitive NMDA antagonist 3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid and by the NMDA channel blocker phencyclidine. Finally, the response to L-glutamate was unaffected by either tetrodotoxin or the kainite–AMPA antagonist 6-cyano-7-nitroquinoxaline-2,3-dione. These data demonstrate the presence of NMDA receptors on dopaminergic nerve terminals that mediate the ability of L-glutamate to release DA and suggest an additional mechanism by which information from the nigrostriatal and corticostriatal pathways may be integrated.Key words: N-methyl-D-aspartate, receptors, dopamine, nigrostriatal, synaptosomes.

Inhibition of adenosine-induced coronary vasodilation by block of large-conductance Ca(2+)-activated K+ channels

1994 ◽  
Vol 267 (4) ◽  
pp. H1455-H1460 ◽  
Author(s):  
F. Cabell ◽  
D. S. Weiss ◽  
J. M. Price
Keyword(s):  
Coronary Arteries ◽  
Channel Blocker ◽  
Dependent Manner ◽  
Maximal Inhibition ◽  
Concentration Dependent Manner ◽  
Channel Modulation ◽  
Cyclic Monophosphate ◽  
Kca Channels ◽  
Thromboxane A2 Analogue

The aim of the present study was to investigate the contribution of large-conductance calcium-activated potassium (large-conductance KCa) channels to adenosine (Ado)- and nitroprusside-mediated relaxation in small coronary arteries. Canine subepicardial arteries (170 +/- 23 microns at 120 mmHg) were studied as in vitro pressurized vessels. Pressure-diameter experiments showed myogenic tone over a physiological range of pressures. Tone was increased with the thromboxane A2 analogue 9,11-dideoxy-11 alpha,9 alpha-epoxy-methanoprostaglandin F2 alpha (U-46619). Tetraethylammonium (TEA+; 1 mM) significantly inhibited Ado-induced [and by implication, adenosine 3',5'-cyclic monophosphate (cAMP)-induced] relaxations at Ado concentrations ranging from 0.1 to 10 microM with maximal inhibition (61 +/- 8%) at 1 microM Ado. The large-conductance KCa-channel blocker iberiotoxin (IbTX; 0.01-0.1 microM) inhibited Ado-mediated relaxation in a concentration-dependent manner. Inhibition by IbTX increased with increasing vessel pressure (i.e., 45 +/- 12% at 40 mmHg and 83 +/- 20% at 120 mmHg). TEA+ had a minimal effect (8 +/- 3%) on relaxation induced by nitroprusside. Similar results were found with acetylcholine and bradykinin. These results suggest that (in dog coronary arteries with diameter < 200 microns) large-conductance KCa-channel modulation may play a major role in cAMP-mediated relaxation but is not significant in guanosine 3',5'-cyclic monophosphate-mediated relaxation.

Effects of Mg2+ on Ca2+ waves and Ca2+ transients of rat ventricular myocytes

1996 ◽  
Vol 270 (3) ◽  
pp. H907-H914 ◽  
Author(s):  
H. Terada ◽  
H. Hayashi ◽  
N. Noda ◽  
H. Satoh ◽  
H. Katoh ◽  
...  
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Channel Blocker ◽  
Ventricular Myocytes ◽  
Inhibitory Effect ◽  
Antiarrhythmic Action ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Triggered Activity ◽  
Rat Ventricular Myocytes ◽  
High Concentrations

It has been shown that the occurrence of the transient inward current, which is responsible for triggered activity, was often associated with propagating regions of increased intracellular Ca2+ concentration ([Ca2+]i), i.e., the “Ca2+ wave.” To investigate the mechanism of antiarrhythmic action of Mg2+, we have studied effects of high concentrations of Mg2+ on Ca2+ waves in isolated rat ventricular myocytes. [Ca2+]i was estimated using the Ca(2+)-indicating probe indo 1. Ca2+ waves in myocytes, stimulated at 0.2 Hz, were induced by perfusion of isoproterenol (10(-7) M). High Mg2+ concentration suppressed Ca2+ waves in a concentration-dependent manner (36% at 4 mM, 70% at 8 mM, and 82% at 12 mM). The Ca2+ channel blocker verapamil also suppressed Ca2+ waves in a similar way. In contrast with marked depression of Ca2+ transients by verapamil, Ca2+ transients were not affected by high Mg2+ concentration (8 mM). High Mg2+ concentration also reduced frequencies of Ca2+ waves in the absence of electrical stimulation, whereas verapamil failed to reduce frequencies of Ca2+ waves. Reduction in frequency of Ca2+ waves by high Mg2+ concentration was associated with slowing of propagation velocity of Ca2+ waves. To examine whether suppressive effects of high Mg2+ concentration on Ca2+ waves were related to an increase in intracellular Mg2+ concentration ([Mg2+]i), the effect of high-Mg2+ solution on [Mg2+]i was examined in myocytes loaded with mag-fura 2. An increase in extracellular Mg2+ concentration from 1 to 12 mM increased [Mg2+]i from 1.06 +/- 0.16 to 1.87 +/- 0.22 mM (P < 0.01) in 30 min. To examine the effect of high Mg2+ concentration on amount of releasable Ca2+ in the sarcoplasmic reticulum, the effect of high Mg2+ concentration on the Ca2+ transient induced by a rapid application of caffeine was examined. High-Mg2+ solution increased the peak of the caffeine-induced Ca2+ transient. These results suggest that the inhibitory effect of Mg2+ on Ca2+ waves was not due to inhibition of the sarcolemmal Ca2+ channel but could be due to a decreased propensity for the sarcoplasmic reticulum to divest itself of excess Ca2+.

Hepatocytes are a rich source of novel aspirin-triggered 15-epi-lipoxin A4

1999 ◽  
Vol 277 (5) ◽  
pp. C870-C877 ◽  
Author(s):  
Esther Titos ◽  
Nan Chiang ◽  
Charles N. Serhan ◽  
Mario Romano ◽  
Joan Gaya ◽  
...  
Keyword(s):  
Rat Hepatocytes ◽  
Arachidonic Acid Metabolism ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Potent Inducer ◽  
Biologically Relevant ◽  
In Vivo Experiments ◽  
Cox 2 ◽  
Cytochrome P 450

Novel aspirin (ASA)-triggered 15-epi-lipoxins (ATL) comprise new potent bioactive eicosanoids that may contribute to the therapeutic effect of this drug. ATL biosynthesis is initiated by ASA acetylation of cyclooxygenase (COX)-2 and was originally identified during the interaction of leukocytes with either endothelial or epithelial cells. Here, we examined ATL biosynthesis in rat hepatocytes either alone or in coincubation with nonparenchymal liver cells (NPC) and in liver homogenates from ASA-treated rats. Rat hepatocytes and CC-1 cells, a rat hepatocyte cell line, displayed COX-1 but not COX-2 mRNA expression and predominantly produced thromboxane A2(TXA2) and 15-hydroxyeicosatetraenoic acid (15-HETE). In these cells, ASA shifted the arachidonic acid metabolism from TXA2 to 15-HETE in a concentration-dependent manner. In contrast, neither indomethacin, ibuprofen, valeryl salicylate, nor nimesulide was able to trigger 15-HETE biosynthesis. SKF-525A, a cytochrome P-450 inhibitor, significantly reduced the effect of ASA on 15-HETE biosynthesis. Furthermore, phenobarbital, a potent inducer of cytochrome P-450 activity, further increased ASA-induced 15-HETE production. ASA treatment of hepatocyte-NPC coincubations resulted in the generation of significant amounts of ATL. In addition, in vivo experiments demonstrated augmented hepatic levels of 15-epi-lipoxin A4 in ASA-treated rats. Taken together and considering that ASA is hydrolyzed on its first pass through the portal circulation, these data indicate that, during ASA's consumption, liver tissue generates biologically relevant amounts of ATL by COX-2-independent mechanisms.

scholarly journals Synchronized synthesis and intracellular transport of serum albumin and apolipoprotein B in cultured rat hepatocytes as studied by double immunofluorescence.

1986 ◽  
Vol 34 (9) ◽  
pp. 1223-1230 ◽  
Author(s):  
G A Keller ◽  
C Glass ◽  
D Louvard ◽  
D Steinberg ◽  
S J Singer
Keyword(s):  
Serum Albumin ◽  
Apolipoprotein B ◽  
Intracellular Transport ◽  
Primary Cultures ◽  
Rat Hepatocytes ◽  
Cell System ◽  
Secretory Proteins ◽  
Transport Pathway ◽  
Apo B ◽  
Normal Rat

Synthesis and intracellular transport of two secretory proteins, serum albumin (SA) and apolipoprotein B (apo B) have been synchronized in primary cultures of normal rat hepatocytes to make possible immunocytochemical study of the transport pathway. Under appropriate conditions of cycloheximide treatment, synthesis of new protein was inhibited and, by double immunofluorescent labeling, the cells were found to be largely depleted of the SA and apo B previously synthesized. Re-initiation of protein synthesis led to sequential appearance of SA and apo B, first in the endoplasmic reticulum, then in the Golgi complex, and finally at the cell surface. These results indicate that it should be feasible to use this cell system for high-resolution investigation of the sequence of structures involved in intracellular transport of SA and apo B by corresponding immunolabeling experiments as observed by electron microscopy.

scholarly journals Glucosamine Hydrochloride Specifically Inhibits COX-2 by Preventing COX-2 N-Glycosylation and by Increasing COX-2 Protein Turnover in a Proteasome-dependent Manner

2007 ◽  
Vol 282 (38) ◽  
pp. 27622-27632 ◽  
Author(s):  
Byeong-Churl Jang ◽  
Su-Haeng Sung ◽  
Jong-Gu Park ◽  
Jong-Wook Park ◽  
Jae Hoon Bae ◽  
...  
Keyword(s):  
Molecular Mass ◽  
Protein Turnover ◽  
A549 Cells ◽  
Lung Epithelial Cells ◽  
Prostaglandin E ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Glucosamine Hydrochloride ◽  
Cox 2 ◽  
Factor Α

COX-2 and its products, including prostaglandin E2, are involved in many inflammatory processes. Glucosamine (GS) is an amino monosaccharide and has been widely used for alternative regimen of (osteo) arthritis. However, the mechanism of action of GS on COX-2 expression remains unclear. Here we describe a new action mechanism of glucosamine hydrochloride (GS-HCl) to tackle endogenous and agonistdriven COX-2 at protein level. GS-HCl (but not GS sulfate, N-acetyl GS, or galactosamine HCl) resulted in a shift in the molecular mass of COX-2 from 72–74 to 66–70 kDa and concomitant inhibition of prostaglandin E2 production in a concentration-dependent manner in interleukin (IL)-1β-treated A549 human lung epithelial cells. Remarkably, GS-HCl-mediated decrease in COX-2 molecular mass was associated with inhibition of COX-2 N-glycosylation during translation, as assessed by the effect of tunicamycin, the protein N-glycosylation inhibitor, or of cycloheximide, the translation inhibitor, on COX-2 modification. Specifically, the effect of low concentration of GS-HCl (1 mm) or of tunicamycin (0.1 μg/ml) to produce the aglycosylated COX-2 was rescued by the proteasomal inhibitor MG132 but not by the lysosomal or caspase inhibitors. However, the proteasomal inhibitors did not show an effect at 5 mm GS-HCl, which produced the aglycosylated or completely deglycosylated form of COX-2. Notably, GS-HCl (5 mm) also facilitated degradation of the higher molecular species of COX-2 in IL-1β-treated A549 cells that was retarded by MG132. GS-HCl (5 mm) was also able to decrease the molecular mass of endogenous and IL-1β- or tumor necrosis factor-α-driven COX-2 in different human cell lines, including Hep2 (bronchial) and H292 (laryngeal). However, GS-HCl did not affect COX-1 protein expression. These results demonstrate for the first time that GS-HCl inhibits COX-2 activity by preventing COX-2 co-translational N-glycosylation and by facilitating COX-2 protein turnover during translation in a proteasome-dependent manner.

scholarly journals Insulin-mediated inhibition of apolipoprotein B secretion requires an intracellular trafficking event and phosphatidylinositol 3-kinase activation: studies with brefeldin A and wortmannin in primary cultures of rat hepatocytes

1996 ◽  
Vol 313 (2) ◽  
pp. 567-574 ◽  
Author(s):  
Janet D. SPARKS ◽  
Thuy L. PHUNG ◽  
Mary BOLOGNINO ◽  
Charles E. SPARKS
Keyword(s):  
Apolipoprotein B ◽  
Enzyme Inhibitor ◽  
Primary Cultures ◽  
Brefeldin A ◽  
Rat Hepatocytes ◽  
Phosphatidylinositol 3 Kinase ◽  
Apo B ◽  
Insulin Effect ◽  
Insulin Inhibition ◽  
Phosphatidylinositol 3

Insulin inhibition of the secretion of apolipoprotein B (apo B) was studied in primary cultures of rat hepatocytes by using brefeldin A (BFA), an inhibitor of protein transport from the endoplasmic reticulum (ER) to the Golgi apparatus, and by using the phosphatidylinositol 3-kinase (PI 3-K) inhibitor wortmannin. Incubation of hepatocytes with BFA (10 μg/ml) for 1 h inhibited the subsequent secretion of apo B, albumin and transferrin for up to 3 h. BFA treatment resulted in the time-dependent accumulation in cells of [14C]leucine-labelled proteins and apo B. Under conditions where insulin decreased total apo B (cell plus secreted), BFA blocked the insulin-dependent effect. These results suggest that export of apo B from the ER is a prerequisite for the observed insulin effect. Treatment of hepatocytes with wortmannin for 20 min abolished insulin inhibition of apo B secretion, suggesting that the insulin effect on the apo B pathway involves activation of PI 3-K. Enzyme inhibitor studies indicate that chymostatin and (+)-(2S,3S)-3-[(S)-methyl-1-(3-m e t h y l b u t y l c a r b a m o y l) - b u t y l c a r b a m o y l] - 2-oxiranecarboxylate (E-64-c) partially block insulin effects on apo B compared with leupeptin, which had no discernible effect. The cell-permeable derivative of E-64-c, EST, and N-Ac-Leu-Leu-norleucinal (ALLN) were most effective in blocking insulin effects on apo B. These results suggest that insulin action on apo B in primary rat hepatocytes involves (1) vesicular movement of apo B from the ER; (2) activation of PI 3-K and (3) a cellular protease that is either a cysteine- or calcium-activated neutral protease.

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