# The Role of Macrophage Dynamics in Atherosclerosis Analyzed Using a Petri Net-Based Model
## 3 Results and Discussion
### Simulation-Based Knockout Analysis
II-23 is recognized for its significant role in chronic inflammation and its critical involvement in the production of IL-17 family cytokines by Th17 cells, thus establishing the IL-23/IL-17 axis. Additionally, it is known for its pro-atherogenic effects. This is further supported by several clinical research studies that have documented a notable increase in IL-23 levels in individuals with atherosclerosis [31,36]. Figure 4: A graphical depiction is presented to illustrate the impact of knockout on the development of atherosclerosis, specifically focusing on transition \(t_{21}\) regarding the formation of atherosclerotic plaques (\(t_{41}\)). In the graphical representation, the inactive transitions identified through simulation knockout are denoted by red circles. Active transitions are represented by rectangles filled with either green or yellow, indicating whether the activity of the transition has decreased (partially filled) or remained unchanged (fully filled) compared to the reference set. To model the effects of inhibiting the IL-23 cytokine, we have excluded transition \(t_{19}\) (IL-23 secretion) from the net. In a physiological context, the proposed effect might be achievable via two distinct but interconnected biological mechanisms. The first involves an increase in HDL-C levels [64], while the second strategy focuses on the direct inhibition of the IL-23 cytokine [65]. Consequently, in the model, we observed that one of the mechanisms leading to HSPCs' mobilization, specifically the G-CSF-induced one, has been inhibited. Another mechanism involved in the mobilization of HSPCs in the model is the CXCL12/CXCR4 pathway, which also plays a significant role in atherosclerosis. The interaction between CXCL12 and CXCR4 has been shown to activate a signaling pathway that inhibits the efflux of ABCA1-dependent macrophage cholesterol, thus intensifying atherosclerosis [66]. Therefore, if transition \(t_{9}\) (cleavage of CXCR4 on HSPCs via increasing proteolytic activity caused by neutrophils) is further removed, the mobilization of HSPCs becomes blocked, ultimately leading to inhibition of atherosclerotic plaque formation in the model (see Figure 5). **Scenario 4**. : _Inhibition of M1 macrophages' polarization while simultaneously promoting M2 macrophages' polarization._  
The immune system activates a defensive inflammatory response when it detects danger signals. This response includes several processes, with macrophages playing a crucial role. Macrophages, derived from monocytes in the bone marrow, adjust their characteristics and function based on the surrounding conditions, a process called polarization. There are two main types of macrophages: pro-inflammatory M1 and anti-inflammatory M2 [67]. M1 macrophages, stimulated by IFN-\(\gamma\) and LPS, release pro-inflammatory substances and have a significant impact on the advancement of atherosclerosis and chronic inflammation. This leads to the formation of foam cells and the development of plaque [51]. To simulate the impact of the inhibition of macrophage differentiation in the M1 type, we removed the transition \(t_{40}\) (increased local inflammation) from the model. As a result, the development of atherosclerotic plaque in the model was observed to stop (similarly to that in Figure 4). Furthermore, it was noticed that the firing probability of the transition \(t_{27}\) (monocyte differentiation from Lyc6C\({}^{low}\)), as compared to the reference firing value, increased by 4%, indicating a significant influence on macrophage differentiation toward the M2 phenotype. The results obtained are consistent with the findings described in [48], where it was demonstrated that by decreasing the polarization of M1 macrophages, it was possible to inhibit the formation of foam cells and the production of pro-inflammatory cytokines. Consequently, this led to a decrease in the development of atherosclerotic plaque.