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Abstract

This study investigated the molecular mechanism of the effect of sulfur dioxide (SO₂) on the expression of adenosine triphosphate (ATP)-sensitive potassium ion (K⁺; K_ATP) channel, big-conductance calcium ion (Ca²⁺)-activated K⁺ (BK_Ca) channel, and L-type (L-Ca²⁺) channel subunits in rat aortas with quantitative reverse transcription polymerase chain reaction (qRT-PCR) and Western blot. The results showed that the messenger RNA and protein levels of the K_ATP channel subunits Kir6.1, Kir6.2, and sulfonylurea receptor 2B (SUR2B) of rat aortas were significantly increased by SO₂ at 14 mg/m³, whereas the levels of SUR2A were not changed. SO₂ at all the treated concentrations markedly raised the expression of the BK_Ca channel subunits α and β1. SO₂ at 14 mg/m³ significantly decreased the expression of the L-Ca²⁺ channel Ca_v1.2 and Ca_v1.3. The histological examination of rat aorta tissues showed moderate injury of tunica media in the presence of SO₂ at 14 mg/m³. These suggest that SO₂ can activate the K_ATP and BK_Ca channels by upregulating the expression of Kir6.1, Kir6.2, SUR2B, BK_Ca α, and BK_Ca β1, while inhibit the L-Ca²⁺ channels by downregulating the expression of Ca_v1.2 and Ca_v1.3 in rat aortas. The molecular mechanism of SO₂-induced vasorelaxant effect might be linked to the changes in expression of these channel subunits, which plays an important role in the pathogenesis of SO₂-associated cardiovascular diseases.

Keywords

Sulfur dioxide molecular mechanism rat aortas cardiovascular diseases

Introduction

Sulfur dioxide (SO₂) is the main product by the combustion of sulfur compounds and is of significant environmental concern. Epidemiological investigations have revealed that SO₂ exposure is linked to cardiovascular diseases.^1,2 In recent years, our studies indicated that SO₂ is a systemic toxic agent because it could cause many kinds of toxic effects, including induction of chromosomal aberrations (CA), micronuclei, and sister chromatid exchanges, DNA damage, gene mutagenesis, lipid peroxidation, and changes of cytokine levels and genome expressions.^3
–5,6 SO₂ caused relaxation of rat thoracic aortic rings in a concentration-dependent manner (from 1 μM to 2000 μM) and had toxicological effects on rat cardiovascular system.⁷ However, SO₂ is also generated during the normal processing of sulfur-containing amino acids.⁸ Our previous study found that the concentrations of SO₂ in rat plasma and thoracic aortic tissues were 16.77 ± 8.24 μM and 127.76 ± 31.34 μM, respectively.⁶

The mechanism of SO₂-induced vasorelaxation might be related to adenosine triphosphate (ATP)-sensitive potassium ion (K⁺; K_ATP) channel, big-conductance calcium ion (Ca²⁺)-activated K⁺ (BK_Ca) channel, and L-type Ca²⁺ (L-Ca²⁺) channel.⁸ K_ATP channels were found in many tissues like cardiac myocytes, vascular smooth muscle cells (VSMCs), neurons, skeletal muscles, and pancreatic β-cells.⁹ These channels play a vital role in the regulation of vascular tone. BK_Ca channels constitute an important physiological link between electrical signaling and cellular Ca²⁺ signaling at the plasma membrane. They are believed to maintain the membrane potential in vessels and are activated by changes in membrane depolarization and the concentration of intracellular Ca²⁺.^10
–12 L-Ca²⁺ channels are membrane proteins that mediate Ca²⁺ influx into electrically excitable cells. These channels are sensitive to 1,4-dihydropyridines, some of them increasing (Bay K8644) and some blocking (nicardipine and nifedipine) Ca²⁺ current, and are called dihydropyridine receptors. They are distributed in many tissues, namely, heart, neurons, skeletal muscle, endocrine cells, VSMCs, and brain.¹³

Our previous study found that the mechanism of SO₂-induced vasorelaxation was linked to K_ATP, BK_Ca, and L-Ca²⁺ channels.⁸ However, this study only indirectly proved the correlation between SO₂-induced vasorelaxation and ion channels using blocking agents of these ion channels. The molecular mechanism by which SO₂ produces this effect through K⁺ and Ca²⁺ channels is still unclear. Therefore, the purpose of this study is to investigate the effect of SO₂ on the expression of these channel subunits in rat aortas.

Materials and methods

Chemicals

Maxima SYBR Green quantitative polymerase chain reaction (qPCR) Master Mix kit, PrimeScript reverse transcription (RT) reagent kit, and TRIzol reagent were purchased from TaKaRa (Dalian, China). Goat polyclonal BK_Ca α, BK_Ca β1, SUR2A, SUR2B, Kir6.1, and Kir6.2 and rabbit polyclonal Ca_v1.2, Ca_v1.3, and β-actin antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, California, USA). Liquid SO₂ (purity: 99.99%) was purchased from the Beijing He-Pu-Bei-Fen Gas (Beijing, China).

Animals and treatment protocols

Male Wistar rats (220–250 g) were bought from Heibei Medical University (Shijiazhuang, China). All animal procedures complied with the Animal Care Committee of Shanxi University. The rats were divided randomly into four groups with six in each group. Three groups were exposed to 3.5 ± 0.35, 7.0 ± 1.08, and 14.1 ± 2.04 mg/m³ SO₂ in 1-m³ exposure chambers for 4 h/day for 30 consecutive days, respectively, while the control group was placed in another identical chamber, which was continually pumped with filtered air for the same period of time. The SO₂ was diluted with fresh air at the intake port of the chamber to yield the desired SO₂ concentrations. The SO₂ within the chambers was measured every 30 min by pararosaniline hydrochloride spectrophotometry.¹⁴ When not being treated, the rats had free access to food and water. Rats were killed 24 h after the last treatment by anesthetic overdose (intraperitoneal injection of 90 mg/kg of pentobarbital sodium). The aortas were removed immediately and cleared of any connective tissues, then cut into rings of around 1 mm in length. For the hematoxylin and eosin (HE) staining assay, some rings were fixed in 10% formaldehyde in phosphate-buffered saline (PBS). The other rings were frozen in liquid nitrogen for protein and messenger RNA (mRNA) assay.

RNA isolation and qRT-PCR

According to the manufacturer’s instructions, total RNA was isolated from rat aorta tissues using TRIzol reagent. To assure RNA quality 1% gel electrophoresis (28S/18S RNA) was used. The concentration of RNA was measured by spectrophotometric analysis at 260 nm. The First Strand cDNA Synthesis kit (TakaRa, Dalian, China) was used to synthesize the first-strand complementary DNA (cDNA). Then the cDNA product was stored at −80°C before use.

All PCR primers were designed by Primer designer software (Primer Premier 5.0) (Table 1). The quantitative reverse transcription PCR (qRT-PCR) was carried out using an iQ5 iCycler thermal cycler (Bio-Rad, Hercules, California, USA) and Maxima SYBR Green qPCR Master Mix kit. Each sample established a melting curve. The conditions of PCR were as follows: 95°C for 3 min followed by 40 cycles of 20 s at 94°C, 20 s at 55–60°C, and 20 s at 72°C. The fluorescence data were obtained at the 72°C step. The quantities of target genes were calculated from separate standard curves gained for each experiment. The relative quantification of the expression was measured by dividing the quantities of the target gene with the quantity acquired for β-actin as an internal reference gene.

Table 1.

Primers sequences used in qRT-PCR.

Gene	Accession no.	Sequences
β-Actin	NM_031144
	Forward primer	5′-CCT CTA TGC CAA CAC AGT GCT GTC T-3′
	Reverse primer	5′-GCT CAG GAG GAG CAA TGA TCT TGA-3′
SUR2A	D83598
	Forward primer	5′-GGA GTG CGA TAC TGG TCC AAA CCT-3′
	Reverse primer	5′-CCC GAT GCA GAG AAC GAG ACA CT-3′
SUR2B	AF087838
	Forward primer	5′-CAT AGC TCA TCG GGT TCA CAC CAT T-3′
	Reverse primer	5′-GCA TCG AGA CAC AGG TGC TGT TGT-3′
Kir6.1	NM_017099
	Forward primer	5′-ACG ACG CCA GAA GGA GAG-3′
	Reverse primer	5′-GCC ACT AGG AAG ATG TTA TTG C-3′
Kir6.2	NM_031358
	Forward primer	5′-CAA CGT CGC CCA CAA GAA CAT C-3′
	Reverse primer	5′-CCA GCT GCA CAG GAA GGA CAT G-3′
BK_Ca α	NM_031828
	Forward primer	5′-TTA CAG CAC TCC GCA GAC-3′
	Reverse primer	5′-CAC CAT AAC AAC CAC CAT CC-3′
BK_Ca β1	NM_019273
	Forward primer	5′-ATC CTC CTC TTC TCC TTC TTC TG-3′
	Reverse primer	5′-GCC GTT CCT GGT GAC TCC-3′
Ca_v1.1	NM_053873
	Forward primer	5′-ACA TCG CCC TGC TCG TCC TCT-3′
	Reverse primer	5′-CAC TCG CTG CCG TTG ATG G-3′
Ca_v1.2	NM_012517
	Forward primer	5′-CAT CAT CAT CAT TGC CTT CTT C-3′
	Reverse primer	5′-ACT GGT GCT GGT TCT TGG-3′
Ca_v1.3	NM_017298
	Forward primer	5′-CTT CCT CTT CAT CAT CAT CTT C-3′
	Reverse primer	5′-TCA TAC ATC ACC GCA TTC C-3′
Ca_v1.4	NM_053701
	Forward primer	5′-AAC TGG GCC TAG TGA TGA TGA TGG-3′
	Reverse primer	5′-AGG GAC TTC CTG TTC CTC ATT CTG-3′

qRT-PCR: quantitative reverse transcription polymerase chain reaction; SUR: sulfonylurea receptor; BK_Ca: calcium ion-activated potassium channel.

Western blot analysis

Proteins from aortas were extracted in ice-cold lysis buffer (25 μM phenylmethylsulfonyl fluoride, 5 μg/ml of pepstatin, 1% Nonidet P 40, 50 μg/ml of leupeptin, 1 mM ethylenediaminetetraacetic acid, 0.5 mM sodium vanadate, 2.5 μg/ml of aprotinin, 125 mM sodium fluoride, and 25 μg/ml of trypsin inhibitor). Bradford protein assay kit (Beyotime, Shanghai, China) was used to measure the protein concentration (milligrams protein per milliliter). The aorta lysates were centrifuged at 13,000 r/min for 15 min, and the supernatants were collected for protein separation by sodium dodecyl sulfate–polyacrylamide gel electrophoresis using precast 12% separating/4% stacking gels (Bio-Rad). The separated proteins were transferred to nitrocellulose membranes (Millipore, Billerica, Massachusetts, USA). Membranes were blocked in nonfat dry milk in PBS buffer and then were incubated in either goat polyclonal antibodies specific for rat Ca_v1.2, Ca_v1.3, and β-actin or rabbit polyclonal antibodies for BK_Ca α, BK_Ca β1, SUR2A, SUR2B, Kir6.1, and Kir6.2 at 4°C overnight. Fluorescently labeled anti-rabbit or anti-goat secondary antibody (IRDye 800CW rabbit anti-goat or goat anti-rabbit IgG (H + L), LI-COR) at a concentration of 1:5000 was added to membranes and detected with LI-COR Odyssey infrared fluorescent system (Lincoln, Nebraska, USA).

HE staining

For HE staining, the paraffin was used to embed the aortic rings fixed in 10% formaldehyde in PBS. After routine processing, the paraffin sections of every aortic ring were cut into 5 μm thickness, stained with HE, and observed under light microscopy (Olympus, Japan) with 400× magnification.¹⁵ According to the degree of injury, the findings were graded as 0 to +3, which corresponds to no change, slight, moderate, and severe changes, respectively. The endothelial cell loss, middle elastic plate damage, and inflammatory reaction in tissues were assessed.¹⁶

Statistical analysis

All values were expressed as mean ± standard deviation. The data were analyzed using one-way analysis of variance followed by a least significant difference post hoc test to evaluate whether the means were significantly different between the SO₂ groups and the control group. Statistical significance was set at p < 0.05.

Results

The mRNA and protein expression of K_ATP channel subunits in the rat aortas

Kir6.1 expression in the rat aortas

The mRNA levels of Kir6.1 were assessed in the aortas of rats exposed to SO₂ (Figure 1(a)). The Kir6.1 mRNA levels in the 14 mg/m³ SO₂ group were significantly higher than those in the control group. Moreover, the levels of Kir6.1 protein were significantly increased in the aortas treated with the highest concentration of SO₂ (14 mg/m³) relative to the control group (Figure 1(b)).

Figure 1.

Effects of SO₂ inhalation on the mRNA and protein expression of Kir6.1 (a and b), Kir6.2 (c and d), and SUR2B (e and f) in the rat aortas. The mean expression in each treated group is shown as a fold increase compared to the mean expression in the control, which has been ascribed an arbitrary value of 1. Each column and bar represent the mean ± standard deviation of six independent experiments.*p < 0.05; **p < 0.01; ***p < 0.001: compared with control group. SO₂: sulfur dioxide; mRNA: messenger RNA; Kir: inward-rectifying potassium channel; SUR2B: sulfonylurea receptor 2B.

Kir6.2 expression in the rat aortas

Kir6.2 mRNA levels were significantly increased in the aortas of rats treated with 14 mg/m³ SO₂ compared with the control group (Figure 1(c)). Similarly, Kir6.2 protein levels were statistically elevated after 14 mg/m³ SO₂ exposure (Figure 1(d)).

SUR2A expression in the rat aortas

The mRNA and protein expression of SUR2A in the aortas of rats were not significantly different in the aortas exposed to SO₂ as compared to the control group (data not shown).

SUR2B expression in the rat aortas

As shown in Figure 1(e) and (f), SUR2B mRNA and protein levels were statistically elevated by 14 mg/m³ SO₂.

The mRNA and protein expression of BK_Ca channel subunits in the rat aortas

BK_Ca α expression in the rat aortas

SO₂ at 3.5, 7, and 14 mg/m³ significantly increased the BK_Ca α mRNA and protein levels in the aortas (Figure 2(a) and (b)). As shown in Figure 2(c) and (d), BK_Ca β1 mRNA and protein levels in the aortas of rats were significantly elevated after 3.5, 7, and 14 mg/m³ SO₂ exposure.

Figure 2.

Effects of SO₂ inhalation on the mRNA and protein expression of BK_Ca α (a and b) and BK_Ca β1 (c and d) in the rat aortas. The mean expression in each treated group is shown as a fold increase compared to the mean expression in the control, which has been ascribed an arbitrary value of 1. Each column and bar represent the mean ± standard deviation of six independent experiments.*p < 0.05; **p < 0.01; ***p < 0.001: compared with control group. SO₂: sulfur dioxide; mRNA: messenger RNA; BK_Ca: calcium ion-activated potassium channel.

BK_Ca β1 expression in the rat aortas

The mRNA and protein expression of BK_Ca β1 in the aortas of rats from control and different SO₂ inhalation groups are shown in Figure 2(c) and (d). Statistically significant enhancement of BK_Ca β1 mRNA and protein levels occurred at all the treated concentrations.

The mRNA and protein expression of L-Ca²⁺ channel subunits in the rat aortas

Ca_v1.2 expression in the rat aortas

The mRNA and protein levels of Ca_v1.2 were assessed in the aortas of rats exposed to SO₂ (Figure 3(a) and (b)). The Ca_v1.2 mRNA and protein levels in the presence of 14 mg/m³ SO₂ were significantly lower than the control group.

Figure 3.

Effects of SO₂ inhalation on the mRNA and protein expression of Ca_v1.2 (a and b) and Ca_v1.3 (c and d) in the rat aortas. The mean expression in each treated group is shown as a fold increase compared to the mean expression in the control, which has been ascribed an arbitrary value of 1. Each column and bar represents the mean ± standard deviation of six independent experiments.***p < 0.001: compared with control group. SO₂: sulfur dioxide; mRNA: messenger RNA.

Ca_v1.3 expression in the rat aortas

The effect of SO₂ on the mRNA and protein levels of Ca_v1.3 in the aortas detected by qRT-PCR and Western blot analysis are shown in Figure 3(c) and (d). The results indicated that SO₂ treatment at the highest concentration significantly decreased the Ca_v1.3 mRNA levels in the aortas of rats. By protein analysis, the levels of Ca_v1.3 were significantly decreased in the aortas only treated with the highest concentration of SO₂ (14 mg/m³) relative to the control group.

Histopathological observation analysis

The aortas of rats in the 3.5 and 7 mg/m³ SO₂ groups (Figure 4(b) and (c)) showed normal histological features of the tunica layers as did the control group (Figure 4(a)).

Figure 4.

HE staining in the aortas of rats exposed to SO₂. (A) The control group, (B) 3.5 mg/m³ SO₂ group, (C) 7 mg/m³ SO₂ group, and (D) 14 mg/m³ SO₂ group. In the control group, TI, TM, and TA appear normal. In the 14 mg/m³ SO₂ group, arrows indicate the sites of abnormal histological changes in aortas. At ×400 magnification. HE: hematoxylin and eosin; SO₂: sulfur dioxide; TI: tunica intima; TM: tunica media; TA: tunica adventitia.

However, SO₂ at 14 mg/m³ caused abnormal histological changes in the aortas compared with the control (Figure 4(d)). Comparisons of the histopathological changes in the control group and 14 mg/m³ SO₂ group rat aortas are depicted in Table 2. In the 14 mg/m³ SO₂ group, the middle elastic plates were significantly damaged and had loose, broken, or disappearing phenomenon.

Table 2.

Microscopic findings in the SO₂-exposed rat aortas.

Histopathological changes	Control group	14 mg/m³ SO₂ group
Middle elastic plate damage	0.4 ± 0.3	1.5 ± 0.6^a
Endothelial cell loss	0.3 ± 0.3	0.5 ± 0.4
Inflammatory reaction	None	None

SO₂: sulfur dioxide.

^ap < 0.05: compared with control group.

Discussion

Our previous study indicated that SO₂ could cause a decrease of rat blood pressure.³ The vasorelaxant effect of SO₂ at low concentrations was endothelium-dependent, which might be partly related to BK_Ca channel and nitric oxide/cyclic guanosine monophosphate pathway.¹⁷ The mechanism of SO₂-induced vasorelaxation at high concentrations was shown to be endothelium independent, which might be related to K_ATP channel and L-Ca²⁺ channel.⁸ The purpose of this study was to investigate the effect of gaseous SO₂ on the gene and protein expressions of K_ATP, BK_Ca, and L-Ca²⁺ channels in the rat aortas.

K_ATP channels are widely distributed in a number of tissues and play a vital role in the regulation of vascular tone. They connect membrane excitability with cellular metabolism.⁹ K_ATP channels are tetrameric ion-channel complexes and consist of two subunits: a SUR subunit of the ATP-binding cassette protein superfamily and a subunit of the inward-rectifying K⁺ (Kir6.x) channel family.^18,19 The SUR subunits afford different drug sensitivities to the channel complex, whereas the Kir6.x subunits are the pore-forming structures through which K⁺ transverse the membrane. In this study, the results showed that SO₂ at 14 mg/m³ significantly increased the expression levels of K_ATP channel subunits SUR2B, Kir6.1, and Kir6.2 of the rat aortas. The findings suggest that the SO₂-induced vasorelaxant effect might be linked to the increased expression of K_ATP channel subunits that caused the opening of K⁺ channels. These results fit in with a previous study that suggested that SO₂ was an endothelium-derived hyperpolarizing factor that resulted in relaxation of vascular smooth muscle, probably through opening K_ATP channels.²⁰ The membrane potential hyperpolarization in VSMC caused by opening of K⁺ channels is an important mechanism in vasodilation.^10,21 In addition, co-expression of SUR2B with Kir6.1 or Kir6.2 in the K_ATP channels in VSMC has been found to produce channels with the properties of native K_ATP channels.^22,23

BK_Ca channels play an important role in the dynamic control of smooth muscle tone in arteries and are characterized by their synergistic activation by the electrical depolarization of membrane and cytoplasmic Ca²⁺ to result in rapid K⁺ efflux under physiological conditions. The rapid K⁺ efflux can be tested electrically as a large outward current, and this accordingly yield a rapid hyperpolarization of the membrane.^10
–12 BK_Ca channels are formed from a pore-forming α subunit and a regulatory β subunit.^24
–26 The α subunit includes six transmembrane-spanning domains (S1–S6), including a voltage sensor (S4), which form the pore.¹⁰ The β subunit contains four β-subunit isoforms (β1–4), which may be linked with the α subunits in a 1:1 ratio.²⁷ The important function of the β subunits is to raise the Ca²⁺ sensitivity of the channel.^11,28 β1 subunit is the primary isoform in VSMC.^26,29 The results showed that the mRNA and protein levels of the BK_Ca channel subunits α and β1 of the rat aortas were significantly raised by SO₂ at all the treated concentrations. Similarly, some previous studies suggested that BK_Ca channel subunits α and β1 play an important role in the vasorelaxant effect caused by carbon monoxide and NO.³⁰

It has been reported that SO₂ derivatives can increase the outward K⁺ current in VSMC.³¹ Moreover, some studies showed that SO₂ derivatives effectively increased K⁺ currents in rat hippocampal CA1 neurons and the isolated adult rat ventricular myocyte.³² These results indicate that the increase of K⁺ currents is correlated with the increased expression of K⁺ channels subunits caused by SO₂. The functions of each channel should be further studied.

Ca²⁺ influx plays a crucial physiological role in regulating the contraction of VSMC. The opening of L-Ca²⁺ channels permits Ca²⁺ influx and triggers intracellular Ca-induced Ca²⁺ release, leading to cell contraction.³³ L-Ca²⁺ channels have quite high conductance and slow time- or voltage-dependent inactivation. There is a big variety of L-Ca²⁺ channels, and the same cell can express several types of L-Ca²⁺ channel.¹³ The channels consist of five polypeptide subunits (α1, β, α2/δ, and γ). Of the five subunits, the α1 subunit is the most important polypeptide of the Ca²⁺ channel-forming proteins. α1 subunit forms the channel pore for ion flow and is responsible for voltage-dependent Ca²⁺ channel opening and channel selectivity for Ca²⁺. This subunit is divided into Ca_v1.1, Ca_v1.2, Ca_v1.3, and Ca_v1.4.¹³ Our results presented that the mRNA of Ca_v1.1 and Ca_v1.4 was not expressed in the rat aortas. This study also showed that SO₂ at 14 mg/m³ significantly decreased the expression levels of Ca_v1.2 and Ca_v1.3, which indicated the possible involvement of L-Ca²⁺ channels by downregulating the expression levels of Ca_v1.2 and Ca_v1.3 in the vasorelaxant effect of SO₂. Some studies also showed that strong mRNA expression for Ca_v1.1 and Ca_v1.4 was detected in rat cerebellum, skeletal muscle, brain cortex and striatum, and retina, respectively, while Ca_v1.2 and Ca_v1.3 were strongly expressed in rat artery.^34
–36 Our results support their findings.

In the elastic arteries, the tunica media is composed of lamellae and elastic material with intervening layers comprising VSMC, collagenous fibers, and ground substance. The histopathological observation of rat aortas showed that the middle elastic plates were significantly damaged in the presence of SO₂ at 14 mg/m³. These results suggest that SO₂ might cause potential damage on the aortas. However, the results of this study can’t explain whether there are any relationships between the histopathological change and the expression alterations of these channel subunits in rat aortas.

Closure of K⁺ channels depolarizes the plasma membrane leading to the opening of more Ca²⁺ channels, increased intracellular Ca²⁺ levels, and vasoconstriction. Conversely, opening of K⁺ channels caused by pharmacological agents or endogenous stimuli results in an efflux of K⁺ from VSMC, hyperpolarization of the plasma membrane, closure of Ca²⁺ channels, reduced intracellular Ca²⁺ levels, and eventually vasodilation.¹⁰ The opening of BK_Ca channel causes K⁺ efflux and membrane hyperpolarization. This change of membrane potential closes Ca_V1.2, which in turn decreases [Ca²⁺]_i and enables vasorelaxation.³⁷ Abnormalities in the function and expression of K⁺ and/or Ca²⁺ channels in VSMC were observed in human hypertension and in several experimental models of hypertension.^38,39 According to these findings, our results showed that the SO₂-induced vasorelaxant effect might be linked to the increased expression of K_ATP and BK_Ca channel subunits, which led to the opening of K⁺ channels, and the decreased expression of L-Ca²⁺ channel subunits, which led to the closure of Ca²⁺ channels. After the VSMC hyperpolarization, the voltage-gated Ca²⁺ channels are closed, resulting in a decrease of intracellular Ca²⁺ concentrations and subsequent vasodilation.

In conclusion, the results suggest that the molecular mechanism of SO₂-mediated vasorelaxant effect might be linked to the changes in expression of BK_Ca, K_ATP, and L-Ca²⁺ channel subunits. The changes in expression of these channel subunits and histopathological damage induced by SO₂ may play a crucial role in the pathogenesis of SO₂-induced cardiovascular diseases. Further study is needed to investigate the interaction of these channels in the vasorelaxant effect induced by SO₂.

Footnotes

Conflict of interest

The authors declare that there are no conflicts of interest.

Funding

The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: This research was supported by National Natural Science Foundation of China (grant number 21107064), Natural Science Foundation of Shanxi Province (grant numbers 2012021033-4 and 2012011036-4), and Scientific and Technological Innovation Programs of Higher Education Institutions in Shanxi (STIP, grant number 20120010501).

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The molecular mechanism of the effect of sulfur dioxide inhalation on the potassium and calcium ion channels in rat aortas

Abstract

Keywords

Introduction

Materials and methods

Chemicals

Animals and treatment protocols

RNA isolation and qRT-PCR

Western blot analysis

HE staining

Statistical analysis

Results

The mRNA and protein expression of KATP channel subunits in the rat aortas

Kir6.1 expression in the rat aortas

Kir6.2 expression in the rat aortas

SUR2A expression in the rat aortas

SUR2B expression in the rat aortas

The mRNA and protein expression of BKCa channel subunits in the rat aortas

BKCa α expression in the rat aortas

BKCa β1 expression in the rat aortas

The mRNA and protein expression of L-Ca2+ channel subunits in the rat aortas

Cav1.2 expression in the rat aortas

Cav1.3 expression in the rat aortas

Histopathological observation analysis

Discussion

Footnotes

Conflict of interest

Funding

References

The mRNA and protein expression of K_ATP channel subunits in the rat aortas

The mRNA and protein expression of BK_Ca channel subunits in the rat aortas

BK_Ca α expression in the rat aortas

BK_Ca β1 expression in the rat aortas

The mRNA and protein expression of L-Ca²⁺ channel subunits in the rat aortas

Ca_v1.2 expression in the rat aortas

Ca_v1.3 expression in the rat aortas