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The dihydropyridine calcium channel blocker benidipine prevents lysophosphatidylcholine-induced endothelial dysfunction in rat aorta.

Takayama M, Yao K, Wada M - J. Biomed. Sci. (2009)

Bottom Line: We examined the effects of benidipine on the impairment of endothelium-dependent relaxation induced by LPC.Additionally, the effects of benidipine on intracellular calcium concentration ([Ca2+]i) and membrane fluidity altered by LPC in primary cultured rat aortic endothelial cells were examined. [Ca2+]i was measured using the fluorescent calcium indicator fura-2.The present findings may provide new insights into the endothelial protective effects of benidipine.

View Article: PubMed Central - HTML - PubMed

Affiliation: Pharmaceutical Research Center, Kyowa Hakko Kirin Co, Ltd, Sunto-gun, Shizuoka-ken, Japan. makoto.takayama@kyowa-kirin.co.jp

ABSTRACT

Background: Lysophosphatidylcholine (LPC), an atherogenic component of oxidized low-density lipoprotein, has been shown to induce the attenuation of endothelium-dependent vascular relaxation. Although benidipine, a dihydropyridine-calcium channel blocker, is known to have endothelial protective effects, the effects of benidipine on LPC-induced endothelial dysfunction remain unknown. We examined the effects of benidipine on the impairment of endothelium-dependent relaxation induced by LPC.

Methods: Benidipine was administered orally to rats and aortas were then isolated. Aortic rings were treated with LPC and endothelial functions were then evaluated. Additionally, the effects of benidipine on intracellular calcium concentration ([Ca2+]i) and membrane fluidity altered by LPC in primary cultured rat aortic endothelial cells were examined. [Ca2+]i was measured using the fluorescent calcium indicator fura-2. Membrane fluidity was monitored by measuring fluorescence recovery after photobleaching.

Results: Treatment with LPC impaired endothelial function. Benidipine prevents the impairment of relaxation induced by LPC. Acetylcholine elicited an increase in [Ca2+]i in fura-2 loaded endothelial cells. The increase in [Ca2+]i was suppressed after exposure to LPC. Plasma membrane fluidity increased following incubation with LPC. Benidipine inhibited the LPC-induced increase in membrane fluidity and impairment of increase in [Ca2+]i.

Conclusion: These results suggest that benidipine inhibited LPC-induced endothelial dysfunction by maintaining increase in [Ca2+]i. Benidipine possesses membrane stabilization properties in LPC-treated endothelial cells. It is speculated that the preservation of membrane fluidity by benidipine may play a role in the retainment of calcium mobilization. The present findings may provide new insights into the endothelial protective effects of benidipine.

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Effects of benidipine with or without LPC on ACh-induced increases in [Ca2+]i. Endothelial cells were treated as described for Figure 3. Benidipine (1 or 10 nmol/L) or DMSO was simultaneously added with LPC or ethanol. Changes in ACh-induced increases in calcium are expressed as percentage fluorescence value of ACh-induced increases in calcium prior to LPC or ethanol treatment. Each value represents the mean ± S.E. of 7 cells. *P < 0.05, **P < 0.01, ***P < 0.001 compared between indicated groups.
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Figure 4: Effects of benidipine with or without LPC on ACh-induced increases in [Ca2+]i. Endothelial cells were treated as described for Figure 3. Benidipine (1 or 10 nmol/L) or DMSO was simultaneously added with LPC or ethanol. Changes in ACh-induced increases in calcium are expressed as percentage fluorescence value of ACh-induced increases in calcium prior to LPC or ethanol treatment. Each value represents the mean ± S.E. of 7 cells. *P < 0.05, **P < 0.01, ***P < 0.001 compared between indicated groups.

Mentions: In fura-2 loaded endothelial cells, ACh induced an increase in [Ca2+]i (Figure 3A). The increase in [Ca2+]i was inhibited by 3 μmol/L LPC (Figure 3B and 4), and the simultaneous addition of 10 nmol/L benidipine prevented LPC-induced suppression of increases in [Ca2+]i (Figure 3C and 4). In the absence of LPC, benidipine had no effect on ACh-induced increases in [Ca2+]i at 1 and 10 nmol/L (Figure 4). The PKC activators PMA and PDB did not alter ACh-induced increases in [Ca2+]i (Figure 5). The PKC inhibitors Ro-31-8220 and calphostin C did not affect the LPC-induced suppression of increases in [Ca2+]i (Figure 6). The antioxidants ascorbic acid, PDTC and BHT also did not affect the LPC-induced suppression of increases in [Ca2+]i (Figure 7).


The dihydropyridine calcium channel blocker benidipine prevents lysophosphatidylcholine-induced endothelial dysfunction in rat aorta.

Takayama M, Yao K, Wada M - J. Biomed. Sci. (2009)

Effects of benidipine with or without LPC on ACh-induced increases in [Ca2+]i. Endothelial cells were treated as described for Figure 3. Benidipine (1 or 10 nmol/L) or DMSO was simultaneously added with LPC or ethanol. Changes in ACh-induced increases in calcium are expressed as percentage fluorescence value of ACh-induced increases in calcium prior to LPC or ethanol treatment. Each value represents the mean ± S.E. of 7 cells. *P < 0.05, **P < 0.01, ***P < 0.001 compared between indicated groups.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2710319&req=5

Figure 4: Effects of benidipine with or without LPC on ACh-induced increases in [Ca2+]i. Endothelial cells were treated as described for Figure 3. Benidipine (1 or 10 nmol/L) or DMSO was simultaneously added with LPC or ethanol. Changes in ACh-induced increases in calcium are expressed as percentage fluorescence value of ACh-induced increases in calcium prior to LPC or ethanol treatment. Each value represents the mean ± S.E. of 7 cells. *P < 0.05, **P < 0.01, ***P < 0.001 compared between indicated groups.
Mentions: In fura-2 loaded endothelial cells, ACh induced an increase in [Ca2+]i (Figure 3A). The increase in [Ca2+]i was inhibited by 3 μmol/L LPC (Figure 3B and 4), and the simultaneous addition of 10 nmol/L benidipine prevented LPC-induced suppression of increases in [Ca2+]i (Figure 3C and 4). In the absence of LPC, benidipine had no effect on ACh-induced increases in [Ca2+]i at 1 and 10 nmol/L (Figure 4). The PKC activators PMA and PDB did not alter ACh-induced increases in [Ca2+]i (Figure 5). The PKC inhibitors Ro-31-8220 and calphostin C did not affect the LPC-induced suppression of increases in [Ca2+]i (Figure 6). The antioxidants ascorbic acid, PDTC and BHT also did not affect the LPC-induced suppression of increases in [Ca2+]i (Figure 7).

Bottom Line: We examined the effects of benidipine on the impairment of endothelium-dependent relaxation induced by LPC.Additionally, the effects of benidipine on intracellular calcium concentration ([Ca2+]i) and membrane fluidity altered by LPC in primary cultured rat aortic endothelial cells were examined. [Ca2+]i was measured using the fluorescent calcium indicator fura-2.The present findings may provide new insights into the endothelial protective effects of benidipine.

View Article: PubMed Central - HTML - PubMed

Affiliation: Pharmaceutical Research Center, Kyowa Hakko Kirin Co, Ltd, Sunto-gun, Shizuoka-ken, Japan. makoto.takayama@kyowa-kirin.co.jp

ABSTRACT

Background: Lysophosphatidylcholine (LPC), an atherogenic component of oxidized low-density lipoprotein, has been shown to induce the attenuation of endothelium-dependent vascular relaxation. Although benidipine, a dihydropyridine-calcium channel blocker, is known to have endothelial protective effects, the effects of benidipine on LPC-induced endothelial dysfunction remain unknown. We examined the effects of benidipine on the impairment of endothelium-dependent relaxation induced by LPC.

Methods: Benidipine was administered orally to rats and aortas were then isolated. Aortic rings were treated with LPC and endothelial functions were then evaluated. Additionally, the effects of benidipine on intracellular calcium concentration ([Ca2+]i) and membrane fluidity altered by LPC in primary cultured rat aortic endothelial cells were examined. [Ca2+]i was measured using the fluorescent calcium indicator fura-2. Membrane fluidity was monitored by measuring fluorescence recovery after photobleaching.

Results: Treatment with LPC impaired endothelial function. Benidipine prevents the impairment of relaxation induced by LPC. Acetylcholine elicited an increase in [Ca2+]i in fura-2 loaded endothelial cells. The increase in [Ca2+]i was suppressed after exposure to LPC. Plasma membrane fluidity increased following incubation with LPC. Benidipine inhibited the LPC-induced increase in membrane fluidity and impairment of increase in [Ca2+]i.

Conclusion: These results suggest that benidipine inhibited LPC-induced endothelial dysfunction by maintaining increase in [Ca2+]i. Benidipine possesses membrane stabilization properties in LPC-treated endothelial cells. It is speculated that the preservation of membrane fluidity by benidipine may play a role in the retainment of calcium mobilization. The present findings may provide new insights into the endothelial protective effects of benidipine.

Show MeSH
Related in: MedlinePlus