Int J Biol Sci 2020; 16(15):3062-3074. doi:10.7150/ijbs.49332 This issue

Research Paper

Monocytes enhance the inflammatory response to TLR2 stimulation in aortic valve interstitial cells through paracrine up-regulation of TLR2 level

Peijian Zhang1,2, Erlinda The1, Balachandar Nedumaran1, Lihua Ao1, Michael J. Jarrett1, Dingli Xu2, David A. Fullerton1, Xianzhong Meng1✉

1. Department of Surgery, University of Colorado Denver, Aurora, CO 80045.
2. Department of Cardiology, Nanfang Hospital, Southern Medical University, Guangzhou, China.

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Zhang P, The E, Nedumaran B, Ao L, Jarrett MJ, Xu D, Fullerton DA, Meng X. Monocytes enhance the inflammatory response to TLR2 stimulation in aortic valve interstitial cells through paracrine up-regulation of TLR2 level. Int J Biol Sci 2020; 16(15):3062-3074. doi:10.7150/ijbs.49332. Available from

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Graphic abstract

Background and Objectives: Chronic valvular inflammation associated with monocyte infiltration promotes calcific aortic valve disease (CAVD) progression. Further, innate immunity in aortic valve interstitial cells (AVICs), mediated by Toll-like receptors (TLRs), up-regulates cellular inflammatory, fibrogenic and osteogenic activities. Currently, the pro-inflammatory communication between monocytes and AVICs and the underlying mechanism are unclear. We hypothesized that monocytes up-regulate AVIC inflammatory activity. This study sought to characterize the interaction between monocytes and AVICs and to elucidate the mechanism underlying cell-to-cell communication.

Methods and Results: AVICs, monocytes and co-cultures were exposed to a low concentration of TLR2 activator Pam3CSK4 (0.03 µg/ml). The TLR2 activator at this dose induced a marked increase in AVIC production of ICAM-1 and VCAM-1 only when co-cultured with monocytes. Adding conditioned medium from Pam3CSK4-treated monocytes (Pam3 CM, containing 0.1 µg/ml of Pam3CSK4) to AVIC culture (30% vol/vol; diluting Pam3CSK4 to 0.03 µg/ml) greatly increased the expression of adhesion molecules while adding conditioned medium from untreated monocytes (control CM) had no effect. Inhibition or knockdown of TLR2 in AVICs markedly reduced ICAM-1 and VCAM-1 expression induced by Pam3 CM. Further, Pam3 CM increased TLR2 levels in AVICs. Multiplex-ELISA analysis of Pam3 CM identified greater levels of TNF-α. Neutralization of TNF-α abolished the effect of Pam3 CM on AVIC TLR2 levels, resulting in marked attenuation of its potency in the induction of adhesion molecule expression.

Conclusions: This study demonstrates that activated monocytes use paracrine signaling to sensitize AVICs for inflammatory responses to a low level of TLR2 activator. The mechanism of sensitization involves up-regulation of AVIC TLR2 levels by TNF-α from monocytes. Infiltrated monocytes in aortic valve tissue may exacerbate valvular inflammation by rendering AVICs hypersensitive to TLR2 activators.

Keywords: Aortic valves, inflammation, adhesion molecules, monocytes, Toll-like receptor