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In this event, vascular smooth muscle cells VSMCs express, secrete, and deposit bone matrix-related proteins in the vessel wall.
Therefore, vascular calcification has also been indicated as a process of the phenotypic switching of VSMCs into osteoblast-like cells [ 14 , 15 , 16 ].
Bone morphogenetic proteins BMPs have been well-demonstrated to be the major inducer during bone formation. The most direct evidence is the presence of BMPs in vessel lesions and atherosclerotic plaques.
Moreover, accumulating evidence has also indicated the inducer role of BMPs in regulating the expression of bone matrix-related proteins in the VSMCs of vessel walls [ 21 , 26 ].
These pathogenic phenomena have been indicated to resemble those of atherosclerosis development. Moreover, accumulating clinical data has also found that KD patients with large aneurysms may be prone to the formation of coronary artery calcification [ 27 , 28 , 29 ].
However, the mechanism of vascular calcification within KD progression is still unclear. Our prior studies have shown that serum IP and IL levels are usually elevated in acute KD and may play a role in the disease pathogenesis [ 12 , 13 ].
In this study, we investigated whether the serum IP and IL of KD patients could regulate the development of vascular calcification, as well as the underlying mechanism.
Our in vitro findings indicated that vascular calcification could be a complication of coronary arteritis in KD patients.
The cells treated with FC plasma demonstrated no effect. The expressions of bone matrix-related proteins, i. Therefore, we examined whether KD plasma could induce bone matrix-related proteins, i.
The results in d — e are representative of three independent experiments with similar results. BMPs are potential regulators of vascular calcification development.
Runx2 is a BMP downstream transcription factor. Pretreating cells with BMP6-blocking antibodies Fig. The results in a — e are representative of three independent experiments with similar results.
Our in vivo data showed that the average secretion levels of BMP6 in the plasma of the KD patients KD serum has higher levels of secreted BMP6.
The presented in vitro findings provided new insights into the pathogenic effect of KD progression on vascular arteritis initiating vascular calcification in HCASMCs.
In an aneurysm, the phenotype of ECs, myofibroblasts, and SMCs can be modulated by the inflammatory KD-secreted cytokines and vasoactive factors [ 1 , 2 , 3 , 4 , 5 , 32 , 33 ].
Previous studies have found that vasoactive factors, including vascular endothelial growth factors, nitric oxide, and platelet-derived growth factors, etc.
Accumulating clinical data has further indicated that KD patients with large aneurysms may have an increased incidence of coronary artery calcification formation [ 27 , 28 , 29 ].
Moreover, it has been suggested that coronary artery calcification may be one of the sequelae of severe vasculitis occurring in acute KD patients [ 27 , 28 , 29 ].
The clinical symptoms of vascular calcification in KD patients has already been demonstrated, however its mechanism is still unclear.
Although it still lacks an in vivo animal model or direct clinical evidence, the elucidation of the mechanism in this study could provide a new direction to explain the cause of KD vascular calcification.
VSMC calcification is one of the crucial components of cardiovascular disease [ 14 , 16 ]. Previous studies have demonstrated that both IP and IL can be secreted into the plasma and subsequently may play a role in the development of coronary arteritis in KD [ 7 , 8 , 9 , 10 , 11 , 12 , 13 ].
Inflammation is an important factor in the formation of atherosclerosis, as well as KD. It has also been indicated that the coronary artery calcification of KD appears to be initiated by intensive inflammatory stimulation [ 29 ].
Both IP and IL can be secreted from immune cells, i. Although still slightly controversial, recent studies have further indicated that IP and IL can be recognized as proinflammatory factors that may be involved in the development of coronary arterial disease, including aneurysms and atherosclerosis.
BMPs are context-dependent growth factors and thus possess different functional activities in different cell types. In embryos, BMPs can control vascular angiogenesis and patterning [ 41 , 42 ].
In the present results, although higher levels of BMP6 were demonstrated in KD serum, we also found three samples from febrile patients that had similar results.
It has been suggested that more serum samples from KD and febrile patients are needed to be counted and studied.
Moreover, both KD and febrile disease have been considered as a kind of inflammation. Thus, our study also suggested that BMP6 may play a role in the inflammation regulation of KD and even febrile disease , however this role needs to be further examined in the future.
Additional in vivo experimental mouse models of KD and human studies will be needed to confirm the in vivo relevance of these in vitro findings. Therefore, this limitation will be addressed in our ongoing study and be further elucidated in future.
Overall, this in vitro study could provide new insights into the pathogenesis of vascular calcification in VSMCs in KD progression. All others reagents and chemicals were obtained from Sigma St.
Louis, MO. ARS an anthraquinone dye is a calcium-deposition detective reagent. ARS stain has been widely used to analyze calcium-rich deposits in mineralizing tissues and cells for many decades [ 46 , 47 , 48 ].
After the treatment, the cells were fixed formaldehyde and then were stained with ARS reagent. These stained cells were further extracted by adding the acetic acid and then were neutralized by adding the ammonium hydroxide.
Cell lysates were collected by lysing cells with 0. The nitrocellulose paper was further hybridized and analyzed with the indicated antibodies and the detection system.
The clinical plasma samples from both the FC and KD patients were recruited and harvested. In all the assays, at least three individual experiments were carried out.
The data that support the findings of this study are available from the corresponding author upon reasonable request.
Agarwal S, Agrawal DK. Kawasaki disease: etiopathogenesis and novel treatment strategies. Expert Rev Clin Immunol. Pathogenesis of Kawasaki disease.
Clin Exp Immunol. Am Fam Physician. Kawasaki disease: insights into pathogenesis and approaches to treatment. Nat Rev Rheumatol. Sundel R. Kawasaki disease.
Rheum Dis Clin North Am. Kawasaki disease: an update on diagnosis and treatment. Pediatr Neonatol. Serum and urinary interleukin-6 IL-6 levels as predicting factors of Kawasaki disease activity.
Kim DS. Serum interleukin-6 in Kawasaki disease. Yonsei Med J. Arthritis Rheum. Association of lower eosinophil-related T helper 2 Th2 cytokines with coronary artery lesions in Kawasaki disease.
Pediatr Allergy Immunol. J Clin Lab Anal. Th and Treg-related cytokine and mRNA expression are associated with acute and resolving Kawasaki disease.
Circ Res. The regulation of valvular and vascular sclerosis by osteogenic morphogens. Leszczynska A, Murphy JM. Front Bioeng Biotechnol.
Cell Mol Life Sci. Bone morphogenetic protein expression in human atherosclerotic lesions. J Clin Invest. BMP signaling in vascular diseases.
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