Korean J Physiol Pharmacol.  2018 Jul;22(4):437-445. 10.4196/kjpp.2018.22.4.437.

α-Isocubebene modulates vascular tone by inhibiting myosin light chain phosphorylation in murine thoracic aorta

• Ye BH
• Kim EJ
• Baek SE
• Choi YW
• Park SY
• Kim CD

• KMID: 2414268
• DOI: http://doi.org/10.4196/kjpp.2018.22.4.437

Abstract

α-Iso-cubebene (ICB) is a dibenzocyclooctadiene lignin contained in Schisandra chinensis (SC), a well-known medicinal herb that ameliorates cardiovascular symptoms, but the mechanism responsible for this activity has not been determined. To determine the role played by ICB on the regulation of vascular tone, we investigated the inhibitory effects of ICB on vascular contractile responses by adrenergic α-receptor agonists. In addition, we investigated the role on myosin light chain (MLC) phosphorylation and cytosolic calcium concentration in vascular smooth muscle cells (VSMC). In aortic rings isolated from C57BL/6J mice, ICB significantly attenuated the contraction induced by phenylephrine (PE) and norepinephrine (NE), whereas ICB had no effects on KCl (60 mM)-induced contraction. In vasculatures precontracted with PE, ICB caused marked relaxation of aortic rings with or without endothelium, suggesting a direct effect on VSMC. In cultured rat VSMC, PE or NE increased MLC phosphorylation and increased cytosolic calcium levels. Both of these effects were significantly suppressed by ICB. In conclusion, our results showed that ICB regulated vascular tone by inhibiting MLC phosphorylation and calcium flux into VSMC, and suggest that ICB has anti-hypertensive properties and therapeutic potential for cardiovascular disorders related to vascular hypertension.

Keyword

α-Iso-cubebene; Myosin light chain; Vascular tone; Vascular smooth muscle cells

MeSH Terms

Animals
Aorta, Thoracic*
Calcium
Cytosol
Endothelium
Hypertension
Lignin
Mice
Muscle, Smooth, Vascular
Myosin Light Chains*
Myosins*
Norepinephrine
Phenylephrine
Phosphorylation*
Plants, Medicinal
Rats
Relaxation
Schisandra
Calcium
Lignin
Myosin Light Chains
Myosins
Norepinephrine
Phenylephrine

Figure

• 

  Fig. 1 Effects of α-isocubebene (ICB) on phenylephrine (PE)- and norepinephrine (NE)-induced contractions in mouse thoracic aorta.Under pretreatment with vehicle or ICB (1–10 µg/ml), PE (10−7−3×10−6 M) (A) or NE (10−8−3×10−7 M) (B) was added in a step-wise manner to elicit vasocontraction. Tracings are representative of 4–5 independent experiments. Bottom: percentage vasocontraction represents contraction relative to that elicited by 60 mM KCl. Results were presented as the means±SEM of 4–5 independent experiments. *p<0.05; **p<0.01 vs. corresponding value in Vehicle.

• 

  Fig. 2 Effects of α-isocubebene (ICB) on acetylcholine (Ach)- or sodium nitroprusside (SNP)-induced vasodilation in mouse thoracic aorta pre-contracted with phenylephrine (PE, 1 µM).Under pretreatment with vehicle or ICB (1–10 µg/ml), Ach (1 µM) or SNP (0.1 µM) was added to elicit vasodilation. Tracings are representative of 5 independent experiments (A). Percentage vasodilation was presented as the means±SEM of 5 independent experiments (B).

• 

  Fig. 3 Effects of α-isocubebene (ICB) on vascular tension in mouse thoracic aorta pre-contracted with phenylephrine (PE, 1 µM).In mice thoracic aorta with (ED+) or without (ED−) endothelium, ICB (3–30 µg/ml) was added cumulatively to elicit vasodilation. Tracings are representative of 6 independent experiments (A). Percentage vasodilation represents relative value to vasodilation induced by SNP (0.1 µM). Data was presented as the means±SEM of 6 independent experiments (B).

• 

  Fig. 4 Inhibitory effect of α-isocubebene (ICB) on MLC phosphorylation in phenylephrine (PE) (A)- or norepinephrine (NE) (B)-treated VSMC.After pretreating cells with vehicle or ICB (10 µg/ml), PE (1 µM) or NE (1 µM) was added to elicit MLC phosphorylation. Blots are representative of 5–6 independent experiments. Densitometric results of Western blot are shown in the bottom panel. The p-MLC to t-MLC ratios were presented as the means±SEM of 5–6 independent experiments. **p<0.01 vs. corresponding value in control. ##p<0.01 vs. corresponding value in vehicle.

• 

  Fig. 5 Effects of α-isocubebene (ICB) on cytosolic Ca2+ levels in vascular smooth muscle cells.After pretreating cells with vehicle or ICB (10 µg/ml), phenylephrine (PE, 1 µM) (A) or norepinephrine (NE, 0.1 µM) (B) was added to induce calcium influx. Tracings are representative of 4–5 independent experiments (A). Bottom panel: Relative fluorescence ratio was presented as the means±SEM of 4–5 independent experiments. **p<0.01 vs. vehicle.

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