Influence of atherosclerosis on the molecular expression of the TRPC1/BK signal complex in the aortic smooth muscles of mice

Abstract

Atherosclerosis (AS) is one a disease that seriously endangers human health. Previous studies have demonstrated that transient receptor potential channel-1 (TRPC1)/large conductance Ca²⁺ activated K⁺ channel (BK) signal complex is widely distributed in arteries. Therefore, it was hypothesized that TRPC1-BK signal complex may be a new target for the treatment of AS-related diseases. Apolipoprotein E^-/- (ApoE^-/-) mice were used to establish an atherosclerotic animal model in the present study, and the association between AS and the TRPC1-BK signal complex was examined. The present study aimed to compare the differences in the expression levels of mRNAs and proteins of the TRPC1-BK signal complex expressed in the aortic vascular smooth muscle tissue, between mice with AS and control mice. There were 10 mice in each group. Reverse transcription PCR, western blotting and immunohistochemistry were used to detect the differences in the mRNA and protein expression levels of TRPC1, BKα (the α subunit of BK) and BKβ₁ (the β₁ subunit of BK). The mRNA expression level of TRPC1 in AS model mice was significantly higher compared with that in the control group (P<0.05). However, the mRNA expression levels of BKα and BKβ₁ were lower compared with those in the controls (both P<0.01). The mice in the ApoE^-/- group successfully developed AS. In this group, the protein expression level of TRPC1 was significantly higher than that in the control group (P<0.01), while the protein expression levels of BKα and BKβ₁ were lower compared with those in the control group (P<0.01 and P<0.05, respectively). Collectively, it was identified that the protein and mRNA expression levels of the TRPC1/BK signal complex in the aortic vascular smooth muscle tissue could be influenced by the development of AS in mice. Hence, the TRPC1/BK signal complex may be a potential therapeutic target for the prevention and treatment of AS-related complications in the future.

Keywords: atherosclerosis; gene knockout techniques; large-conductance calcium-activated potassium channels; myocytes; smooth muscle.

Figure 1

Optical microscopy H&E staining was used to detect AS formation in the aortic tissue of the two groups (n=10). (A) Aortic lumen of mice in the control group. (B) Aortic lumen of the experimental apolipoprotein E^-/- group of mice, with thickening of the vessel wall and cholesterol crystal deposits under the intima. Arrows refer to the atherosclerotic lesions, the general optical microscope (magnification, x100). AS, atherosclerosis.

Figure 2

Reverse transcription PCR was used to detect the different mRNA expression levels in the aortic smooth muscle of the two groups (n=10). (A) The RNA band of TRPC1 in the AS group was brighter than that in the control group, whereas RNA bands of BKα and BKβ₁ in the AS group were darker than those in the control group. ‘C’ indicates the control group, ‘M’ indicates the marker. (B) Histogram showing that there are statistical differences between the AS group and the control group RNA. ^*P<0.05, ^**P<0.01 vs. respective control group. AS, atherosclerosis; TRPC1, transient receptor potential channel-1; BK, large conductance Ca²⁺ activated K⁺ channel.

Figure 3

Immunohistochemistry for the detection of the protein expression levels in the aortic smooth muscle of the two groups. Expression levels of (A,B) TRPC1, (C,D) BKα and (E,F) BKβ₁ in the aortic vascular smooth muscle of mice in each group were detected via immunohistochemistry (n=10). Panels A, C, E show results for the control group, while panels B, D and F are for the AS group. The protein expression was observed under an optical microscope (magnification, x400). The brown-yellow shows the protein localization, while the blue area shows the nuclei. (G) The MOD of the brown-yellow area was detected via Metamorph imaging software. ^*P<0.05, ^**P<0.01 vs. respective control group. TRPC1, transient receptor potential channel-1; BKα, large conductance calcium-activated potassium channel α subunit; BKβ₁, large conductance calcium-activated potassium channel β1 subunit; AS, atherosclerosis; MOD, mean optical density.

Figure 4

Western blotting was used to detect the expression of each protein in the aortic vascular smooth muscle of the two groups (n=10). (A) Western blotting bands. (B) Semi-quantification of western blotting results. ^*P<0.05, ^**P<0.01 vs. respective control group. TRPC1, transient receptor potential channel-1; BKα, large conductance calcium-activated potassium channel α subunit; BKβ₁, large conductance calcium-activated potassium channel β1 subunit; AS, atherosclerosis