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Sulfur Dioxide Protects Against Collagen Accumulation in Pulmonary Artery in Association With Downregulation of the Transforming Growth Factor β 1/Smad Pathway in Pulmonary Hypertensive Rats

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ABSTRACT

Background: We aimed to explore the role of endogenous sulfur dioxide (SO2) in pulmonary vascular collagen remodeling induced by monocrotaline and its mechanisms.

Methods and results: A rat model of monocrotaline‐induced pulmonary vascular collagen remodeling was developed and administered with l‐aspartate‐β‐hydroxamate or SO2 donor. The morphology of small pulmonary arteries and collagen metabolism were examined. Cultured pulmonary arterial fibroblasts stimulated by transforming growth factor β1 (TGF‐β1) were used to explore the mechanism. The results showed that in monocrotaline‐treated rats, mean pulmonary artery pressure increased markedly, small pulmonary arterial remodeling developed, and collagen deposition in lung tissue and pulmonary arteries increased significantly in association with elevated SO2 content, aspartate aminotransferase (AAT) activity, and expression of AAT1 compared with control rats. Interestingly, l‐aspartate‐β‐hydroxamate, an inhibitor of SO2 generation, further aggravated pulmonary vascular collagen remodeling in monocrotaline‐treated rats, and inhibition of SO2 in pulmonary artery smooth muscle cells activated collagen accumulation in pulmonary arterial fibroblasts. SO2 donor, however, alleviated pulmonary vascular collagen remodeling with inhibited collagen synthesis, augmented collagen degradation, and decreased TGF‐β1 expression of pulmonary arteries. Mechanistically, overexpression of AAT1, a key enzyme of SO2 production, prevented the activation of the TGF‐β/type I TGF‐β receptor/Smad2/3 signaling pathway and abnormal collagen synthesis in pulmonary arterial fibroblasts. In contrast, knockdown of AAT1 exacerbated Smad2/3 phosphorylation and deposition of collagen types I and III in TGF‐β1–treated pulmonary arterial fibroblasts.

Conclusions: Endogenous SO2 plays a protective role in pulmonary artery collagen accumulation induced by monocrotaline via inhibition of the TGF‐β/type I TGF‐β receptor/Smad2/3 pathway.

No MeSH data available.


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Endogenous SO2 inhibited the Smad2/3 signaling pathway and collagen deposition. A, AAT1 protein expression, SO2 content, and AAT activity in PAFs overexpressing AAT1. B, The phosphorylation of Smad2/3 induced by TGF‐β1 in PAFs overexpressing AAT1. C and D, Western blotting (C) and immunofluorescence analysis (D) of collagen types I and III induced by TGF‐β1 in PAFs with AAT1 overexpression. Results are expressed as mean±SE for 3 separate experiments (n=3), each in triplicate. *P<0.05 compared with the vehicle group; #P<0.05 compared with the vehicle plus TGF‐β1 group. AAT1 indicates aspartate aminotransferase 1; PAF, pulmonary arterial fibroblast; SO2, sulfur dioxide; TGF‐β1, transforming growth factor β1.
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jah31819-fig-0005: Endogenous SO2 inhibited the Smad2/3 signaling pathway and collagen deposition. A, AAT1 protein expression, SO2 content, and AAT activity in PAFs overexpressing AAT1. B, The phosphorylation of Smad2/3 induced by TGF‐β1 in PAFs overexpressing AAT1. C and D, Western blotting (C) and immunofluorescence analysis (D) of collagen types I and III induced by TGF‐β1 in PAFs with AAT1 overexpression. Results are expressed as mean±SE for 3 separate experiments (n=3), each in triplicate. *P<0.05 compared with the vehicle group; #P<0.05 compared with the vehicle plus TGF‐β1 group. AAT1 indicates aspartate aminotransferase 1; PAF, pulmonary arterial fibroblast; SO2, sulfur dioxide; TGF‐β1, transforming growth factor β1.

Mentions: To further verify the inhibitory effect of endogenous SO2 on collagen remodeling, PAFs were infected with lentivirus, which contains the cDNA encoding AAT1. After 72 hours, AAT activity, AAT1 protein expression, and SO2 concentration were detected. Compared with the vehicle‐lentivirus group and the control group, AAT1 protein expression was markedly enhanced in the group with AAT1 overexpression (P<0.05) (Figure 5A). Likewise, AAT activity in PAFs and SO2 concentration in the supernatant of PAFs were significantly increased in the group with AAT1 overexpression (P<0.05 for both) (Figure 5A).


Sulfur Dioxide Protects Against Collagen Accumulation in Pulmonary Artery in Association With Downregulation of the Transforming Growth Factor β 1/Smad Pathway in Pulmonary Hypertensive Rats
Endogenous SO2 inhibited the Smad2/3 signaling pathway and collagen deposition. A, AAT1 protein expression, SO2 content, and AAT activity in PAFs overexpressing AAT1. B, The phosphorylation of Smad2/3 induced by TGF‐β1 in PAFs overexpressing AAT1. C and D, Western blotting (C) and immunofluorescence analysis (D) of collagen types I and III induced by TGF‐β1 in PAFs with AAT1 overexpression. Results are expressed as mean±SE for 3 separate experiments (n=3), each in triplicate. *P<0.05 compared with the vehicle group; #P<0.05 compared with the vehicle plus TGF‐β1 group. AAT1 indicates aspartate aminotransferase 1; PAF, pulmonary arterial fibroblast; SO2, sulfur dioxide; TGF‐β1, transforming growth factor β1.
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jah31819-fig-0005: Endogenous SO2 inhibited the Smad2/3 signaling pathway and collagen deposition. A, AAT1 protein expression, SO2 content, and AAT activity in PAFs overexpressing AAT1. B, The phosphorylation of Smad2/3 induced by TGF‐β1 in PAFs overexpressing AAT1. C and D, Western blotting (C) and immunofluorescence analysis (D) of collagen types I and III induced by TGF‐β1 in PAFs with AAT1 overexpression. Results are expressed as mean±SE for 3 separate experiments (n=3), each in triplicate. *P<0.05 compared with the vehicle group; #P<0.05 compared with the vehicle plus TGF‐β1 group. AAT1 indicates aspartate aminotransferase 1; PAF, pulmonary arterial fibroblast; SO2, sulfur dioxide; TGF‐β1, transforming growth factor β1.
Mentions: To further verify the inhibitory effect of endogenous SO2 on collagen remodeling, PAFs were infected with lentivirus, which contains the cDNA encoding AAT1. After 72 hours, AAT activity, AAT1 protein expression, and SO2 concentration were detected. Compared with the vehicle‐lentivirus group and the control group, AAT1 protein expression was markedly enhanced in the group with AAT1 overexpression (P<0.05) (Figure 5A). Likewise, AAT activity in PAFs and SO2 concentration in the supernatant of PAFs were significantly increased in the group with AAT1 overexpression (P<0.05 for both) (Figure 5A).

View Article: PubMed Central - PubMed

ABSTRACT

Background: We aimed to explore the role of endogenous sulfur dioxide (SO2) in pulmonary vascular collagen remodeling induced by monocrotaline and its mechanisms.

Methods and results: A rat model of monocrotaline&#8208;induced pulmonary vascular collagen remodeling was developed and administered with l&#8208;aspartate&#8208;&beta;&#8208;hydroxamate or SO2 donor. The morphology of small pulmonary arteries and collagen metabolism were examined. Cultured pulmonary arterial fibroblasts stimulated by transforming growth factor &beta;1 (TGF&#8208;&beta;1) were used to explore the mechanism. The results showed that in monocrotaline&#8208;treated rats, mean pulmonary artery pressure increased markedly, small pulmonary arterial remodeling developed, and collagen deposition in lung tissue and pulmonary arteries increased significantly in association with elevated SO2 content, aspartate aminotransferase (AAT) activity, and expression of AAT1 compared with control rats. Interestingly, l&#8208;aspartate&#8208;&beta;&#8208;hydroxamate, an inhibitor of SO2 generation, further aggravated pulmonary vascular collagen remodeling in monocrotaline&#8208;treated rats, and inhibition of SO2 in pulmonary artery smooth muscle cells activated collagen accumulation in pulmonary arterial fibroblasts. SO2 donor, however, alleviated pulmonary vascular collagen remodeling with inhibited collagen synthesis, augmented collagen degradation, and decreased TGF&#8208;&beta;1 expression of pulmonary arteries. Mechanistically, overexpression of AAT1, a key enzyme of SO2 production, prevented the activation of the TGF&#8208;&beta;/type I TGF&#8208;&beta; receptor/Smad2/3 signaling pathway and abnormal collagen synthesis in pulmonary arterial fibroblasts. In contrast, knockdown of AAT1 exacerbated Smad2/3 phosphorylation and deposition of collagen types I and III in TGF&#8208;&beta;1&ndash;treated pulmonary arterial fibroblasts.

Conclusions: Endogenous SO2 plays a protective role in pulmonary artery collagen accumulation induced by monocrotaline via inhibition of the TGF&#8208;&beta;/type I TGF&#8208;&beta; receptor/Smad2/3 pathway.

No MeSH data available.


Related in: MedlinePlus