Advertisement

Simvastatin down-regulates osteogenic response in cultured human aortic valve interstitial cells

Published:October 30, 2019DOI:https://doi.org/10.1016/j.jtcvs.2019.10.081

      Abstract

      Background

      Aortic valve interstitial cells have been implicated in the pathogenesis of aortic stenosis. In response to proinflammatory stimuli, aortic valve interstitial cells undergo an osteogenic phenotypic change. The purpose of this study was to determine whether the anti-inflammatory effects of statins prevent osteogenic activity in cultured aortic valve interstitial cells.

      Methods

      Human aortic valve interstitial cells were isolated from hearts explanted for cardiac transplantation. To test whether simvastatin down-regulates TLR4-induced osteogenic response, aortic valve interstitial cells were treated with simvastatin with and without TLR4 agonist lipopolysaccharide (LPS), and osteogenic markers were measured. Simvastatin's influence on in vitro calcium deposition was assessed by alizarin red staining. Knockdown of postreceptor signaling proteins (MyD88 and TRIF) was performed to determine which of 2 TLR4-associated pathways mediates the osteogenic response. Expression levels of TLR4-induced nuclear factor kappa light chain enhancer of activated B cells (NF-κB) and TLR4 expression were assessed after treatment with simvastatin. Statistical testing was done by analysis of variance (P < .05).

      Results

      Simvastatin decreased LPS-induced ALP and Runx2 expression and inhibited in vitro calcium deposition in aortic valve interstitial cells. Knockdown of MyD88 and TRIF attenuated the osteogenic response. Simvastatin attenuated TLR4-dependent NF-κB signaling and down-regulated TLR4 levels.

      Conclusions

      Simvastatin prevented TLR4-induced osteogenic phenotypic changes in isolated aortic valve interstitial cells via down-regulation of TLR4 and inhibition of NF-κB signaling. These data offer mechanistic insight into a possible therapeutic role for simvastatin in the prevention of aortic stenosis.

      Graphical abstract

      Key Words

      Abbreviations and Acronyms:

      ALP (alkaline phosphatase), DAMP (damage-associated molecular pattern), DMSO (dimethyl sulfoxide), ELISA (enzyme-linked immunosorbent assay), FBS (fetal bovine serum), LPS (lipopolysaccharide), MyD88 (myeloid differentiation primary response 88), NF-kB (nuclear factor kappa light chain enhancer of activated B cells), PAMP (pathogen-associated molecular pattern), Runx2 (runt-related transcription factor 2), shRNA (short hairpin RNA), TLR4 (Toll-like receptor 4), TRIF (TIR-domain-containing adapter inducing interferon-β)
      To read this article in full you will need to make a payment

      References

        • Benjamin E.J.
        • Virani S.S.
        • Callaway C.W.
        • Chamberlain A.M.
        • Chang A.R.
        • Cheng S.
        • et al.
        Heart Disease and Stroke Statistics—2018 update: a report from the American Heart Association.
        Circulation. 2018; 137: e67-e492
        • Leopold J.A.
        Cellular mechanisms of aortic valve calcification.
        Circ Cardiovasc Interv. 2012; 5: 605-614
        • Venardos N.
        • Nadlonek N.A.
        • Zhan Q.
        • Weyant M.J.
        • Reece T.B.
        • Meng X.
        • et al.
        Aortic valve calcification is mediated by a differential response of aortic valve interstitial cells to inflammation.
        J Surg Res. 2014; 190: 1-8
        • Pawade T.A.
        • Newby D.E.
        • Dweck M.R.
        Calcification in aortic stenosis: the skeleton key.
        J Am Coll Cardiol. 2015; 66: 561-577
        • Garcia-Rodriguez C.
        • Parra-Izquierdo I.
        • Castaños-Mollor I.
        • López J.
        • San Román J.A.
        • Sánchez Crespo M.
        Toll-like receptors, inflammation, and calcific aortic valve disease.
        Front Physiol. 2018; 9: 201
        • Yao Q.
        • Song R.
        • Ao L.
        • Cleveland Jr., J.C.
        • Fullerton D.A.
        • Meng X.
        Neurotrophin 3 upregulates proliferation and collagen production in human aortic valve interstitial cells: a potential role in aortic valve sclerosis.
        Am J Physiol Cell Physiol. 2017; 312: C697-C706
        • Zeng Q.
        • Song R.
        • Fullerton D.A.
        • Ao L.
        • Zhai Y.
        • Li S.
        • et al.
        Interleukin-37 suppresses the osteogenic responses of human aortic valve interstitial cells in vitro and alleviates valve lesions in mice.
        Proc Natl Acad Sci U S A. 2017; 114: 1631-1636
        • Song R.
        • Fullerton D.A.
        • Ao L.
        • Zheng D.
        • Zhao K.S.
        • Meng X.
        BMP-2 and TGF-β1 mediate biglycan-induced pro-osteogenic reprogramming in aortic valve interstitial cells.
        J Mol Med (Berl). 2015; 93: 403-412
        • Yang X.
        • Fullerton D.A.
        • Su X.
        • Ao L.
        • Cleveland Jr., J.C.
        • Meng X.
        Pro-osteogenic phenotype of human aortic valve interstitial cells is associated with higher levels of Toll-like receptors 2 and 4 and enhanced expression of bone morphogenetic protein 2.
        J Am Coll Cardiol. 2009; 53: 491-500
        • Takeda K.
        • Akira S.
        Toll-like receptors.
        Curr Protoc Immunol. 2015; 109 (14.12.1-10)
        • Yamamoto M.
        • Sato S.
        • Hemmi H.
        • Hoshino K.
        • Kaisho T.
        • Sanjo H.
        • et al.
        Role of adaptor TRIF in the MyD88-independent Toll-like receptor signaling pathway.
        Science. 2003; 301: 640-643
        • Guven-Maiorov E.
        • Keskin O.
        • Gursoy A.
        • VanWaes C.
        • Chen Z.
        • Tsai C.J.
        • et al.
        The architecture of the TIR domain signalosome in the Toll-like receptor-4 signaling pathway.
        Sci Rep. 2015; 5: 13128
        • Kagan J.C.
        • Su T.
        • Horng T.
        • Chow A.
        • Akira S.
        • Medzhitov R.
        TRAM couples endocytosis of Toll-like receptor 4 to the induction of interferon-beta.
        Nat Immunol. 2008; 9: 361-368
        • Hodgkinson C.P.
        • Ye S.
        Statins inhibit Toll-like receptor 4-mediated lipopolysaccharide signaling and cytokine expression.
        Pharmacogenet Genomics. 2008; 18: 803-813
        • Venardos N.
        • Deng X.S.
        • Yao Q.
        • Weyant M.J.
        • Reece T.B.
        • Meng X.
        • et al.
        Simvastatin reduces the TLR4-induced inflammatory response in human aortic valve interstitial cells.
        J Surg Res. 2018; 230: 101-109
        • Meng X.
        • Ao L.
        • Song Y.
        • Babu A.
        • Yang X.
        • Wang M.
        • et al.
        Expression of functional Toll-like receptors 2 and 4 in human aortic valve interstitial cells: potential roles in aortic valve inflammation and stenosis.
        Am J Physiol Cell Physiol. 2008; 294: C29-C35
        • Cowan C.M.
        • Zhang X.
        • James A.W.
        • Kim T.M.
        • Sun N.
        • Wu B.
        • et al.
        NELL-1 increases pre-osteoblast mineralization using both phosphate transporter Pit1 and Pit2.
        Biochem Biophys Res Commun. 2012; 422: 351-357
        • Li F.
        • Song R.
        • Ao L.
        • Reece T.B.
        • Cleveland Jr., J.C.
        • Dong N.
        • et al.
        ADAMTS5 deficiency in calcified aortic valves is associated with elevated pro-osteogenic activity in valvular interstitial cells.
        Arterioscler Thromb Vasc Biol. 2017; 37: 1339-1351
        • Shen Y.
        • Wu H.
        • Wang C.
        • Shao H.
        • Huang H.
        • Jing H.
        • et al.
        Simvastatin attenuates cardiopulmonary bypass-induced myocardial inflammatory injury in rats by activating peroxisome proliferator-activated receptor γ.
        Eur J Pharmacol. 2010; 649: 255-262
        • Milajerdi A.
        • Larijani B.
        • Esmaillzadeh A.
        Statins influence biomarkers of low-grade inflammation in apparently healthy people or patients with chronic diseases: a systematic review and meta-analysis of randomized clinical trials.
        Cytokine. 2019; 123: 154752
        • Dobrzycka M.
        • Spychalski P.
        • Lachinski A.J.
        • Kobiela P.
        • Jedrusik P.
        • Kobiela J.
        Statins and colorectal cancer: a systematic review.
        Exp Clin Endocrinol Diabetes. August 27, 2018; ([Epub ahead of print])
        • Kohtz P.D.
        • Halpern A.L.
        • Eldeiry M.A.
        • Hazel K.
        • Kalatardi S.
        • Ao L.
        • et al.
        Toll-like receptor-4 is a mediator of proliferation in esophageal adenocarcinoma.
        Ann Thorac Surg. 2019; 107: 233-241
        • Chan K.L.
        • Teo K.
        • Dumesnil J.G.
        • Ni A.
        • Tam J.
        Effect of lipid lowering with rosuvastatin on progression of aortic stenosis: results of the aortic stenosis progression observation: measuring effects of rosuvastatin (ASTRONOMER) trial.
        Circulation. 2010; 121: 306-314
        • Rossebø A.B.
        • Pedersen T.R.
        • Boman K.
        • Brudi P.
        • Chambers J.B.
        • Egstrup K.
        • et al.
        Intensive lipid lowering with simvastatin and ezetimibe in aortic stenosis.
        N Engl J Med. 2008; 359: 1343-1356