Table 2.

List of molecules interfering with valvular interstitial cells in progression of AS based on Rutkovskiy et al. [31]. AKT—protein kinase B; BMP—bone morphogenetic protein; BSP—bone sialoprotein; GPIIb/IIIa—glycoprotein IIb/IIIa; HIF—hypoxia inducible factor; HOTAIR—HOX transcript antisense RNA; IFN—interferon; LPS—lipopolysaccharide; MMP—matrix metalloproteinase; NFκB—Nuclear factor kappa-light-chain-enhancer of activated B cells; OSP—osteopontin, PALMD—palmdelphin; P2Y2—purinoceptor 2; shRNA—short hairpin RNA; SORT—sortilin; STAT—signal transducer and activator of transcription; TGFβ—transforming growth factor β; TNF—tumor necrosis factor; TRAIL—TNF-related apoptosis-inducing ligand; VCAM—vascular cell adhesion molecule, V-LPP—VCAM-1 targeted lipopolyplexes.

Author, Year	Factor	Key Result, End Point
Osman, 2006 [32]	TGF β family cytokines and statins	Cytokines from the TGFβ family promote the differentiation of osteoblasts, whereas atorvastatin inhibits this process.
Osman, 2006 [33]	Adenosine triphosphate and statins	The activation of osteoblast differentiation is facilitated by adenosine triphosphate, but this effect is counteracted by atorvastatin.
Yang, 2009 [34]	LPS and peptidoglycan	Osteoblast differentiation is prompted by lipopolysaccharides (LPS) and peptidoglycan through the activation of toll-like receptors 2 and 4.
Yang, 2009 [35]	BMP 2	BMP2 triggers the early phases of osteoblast differentiation through both canonical and non-canonical pathways.
Yu, 2011 [36]	TNF α and BMP2	Tumor necrosis factor α exclusively triggers osteoblast differentiation in calcified VICs via BMP2 and NFkB signaling.
Carthy, 2012 [37]	Versican	VICs secrete versican in the wound assay; inhibiting its receptor CD44 leads to a reduction in stress fiber (αSMA) formation during VIC migration and inhibits collagen gel contraction.
Song, 2012 [38]	Biglycan	VICs derived from calcified valves exhibit elevated levels of biglycan expression. Biglycan, in turn, promotes osteoblast differentiation through the toll-like receptor 2 and ERK signaling pathways. The expression of biglycan and the calcification process are further stimulated by oxidized low-density lipopolysaccharides.
Zeng, 2012 [39]	LPS, toll-like receptor 4, and Notch	LPS activates an inflammatory phenotype through toll-like receptor 4 (TLR4). In calcified VICs, Notch1 enhances the responsiveness of toll-like receptor 4 to LPS through NFκB signaling.
Poggio, 2013 [40]	BMP 4	Bone morphogenetic protein 4 exclusively initiates osteoblast differentiation in non-calcified VICs, leading to higher levels of differentiation compared to osteogenic medium alone.
Zeng, 2013 [41]	LPS, Notch1	LPS stimulates the cleavage and nuclear translocation of the Notch1 intracellular domain, which subsequently triggers osteoblast differentiation via the activation of ERK and NFκB signaling pathways.
Nadlonek, 2013 [42]	Interleukin-1β	Interleukin-1β induces an inflammatory phenotype in VIC via NFκB.
Zhang, 2014 [43]	MicroRNA 30b	BMP2 initiates osteoblastic differentiation in VICs and suppresses the expression of microRNA 30b. MicroRNA 30b, in turn, inhibits osteoblastic differentiation and apoptosis.
Galeone, 2013 [44]	TNF-related apoptosis-inducing ligand (TRAIL)	Calcified VICs exhibit the presence of TRAIL receptors. The addition of TRAIL to the osteogenic medium enhances the formation of calcified nodules and promotes apoptosis.
El Husseini, 2014 [45]	AKT kinase and P2Y2 receptor	NFκB pathway is involved in inhibiting the expression of IL-6, which is a necessary factor for mineralization. Both AKT kinase and P2Y2 receptor activate this pathway, thereby suppressing IL-6 expression. Cells derived from P2Y2−/− mice are prone to osteoblast differentiation.
Zhang, 2014 [46]	Transcription factor Twist	The osteogenic medium leads to the upregulation of Twist. This process leads to a decrease in the expression of other calcification-related genes. Conversely, the use of Twist siRNA induces osteoblast differentiation.
Carrion, 2014 [47]	Long noncoding RNA HOTAIR	Stretching downregulates HOTAIR through the Wnt signaling pathway. When siRNA is used to target HOTAIR, it leads to the upregulation of BMP2 and alkaline phosphatase expression.
Zeng, 2014 [48]	Oxidized low-density lipoproteins, LPS, and Notch1	Oxidized low-density lipoproteins enhance LPS-induced osteoblastic differentiation through the activation of NFκB and cleavage of Notch1.
Witt, 2014 [49]	Polyunsaturated fatty acids	Several polyunsaturated fatty acids can temporarily inhibit myofibroblast activation through the suppression of Rho kinase and ROCK kinase.
Song, 2014 [50]	Biglycan	Biglycan acts as a ligand for toll-like receptors 2 and 4, contributing to the activation of inflammation in VICs. This effect is mediated through NFκB and ERK pathways
Bouchareb, 2019 [5]	Autotaxin and lysophosphatidic acid	The release of autotaxin by VICs was induced by adenosine diphosphate derived from platelets. Autotaxin, in turn, bound to GPIIb/IIIa receptors on platelets, resulting in the generation of lysophosphatidic acid, which possesses pro-osteogenic properties.
Parra-Izquierdo, 2019 [51]	HIF-1α	HIF-1α activation via STAT1 in valve cells results in the proangiogenic, proinflammatory, and pro-osteogenic effects of IFN-γ
Wang, 2022 [52]	PALMD (Palmdelphin)	PALMD, a protein involved in myoblast differentiation, enhancing VIC osteogenic differentiation and inflammation through the activation of NF-κB.
Voicu, 2022 [53]	V-LPP/shRunx2 lipopolyplexes	VCAM-1 targeted lipopolyplexes, which downregulate the Runx2 gene and decrease the expression of osteogenic molecules OSP, BSP, and BMP-2 in VICs
Liu, 2022 [54]	MMP9	MMP9 expression was distinctly increased in AS, and its inhibition attenuated the calcification of valve interstitial cells by suppressing mitochondrial damage and oxidative stress.
Iqbal, 2023 [55]	Sortilin (SORT1)	Sortilin enhances fibrosis and calcification in aortic valve disease via the transformation of valvular interstitial cells into pathological phenotypes