# The Role of Macrophage Dynamics in Atherosclerosis Analyzed Using a Petri Net-Based Model
## 3 Results and Discussion
### Simulation-Based Knockout Analysis
: _The effect of inhibiting the cytokine M-CSF._  
Colony-stimulating factors (CSFs) are a group of cytokines that are important for forming blood cells, functioning leukocytes, and maintaining immune health. This family consists of M-CSF, G-CSF, and GM-CSF, where M-CSF and G-CSF have a specific influence on the survival, growth, differentiation, and functioning of monocytes/macrophages and neutrophils, respectively. Each member of this family plays a unique role in the development and activity of various immune cells [63]. M-CSF is constantly produced by different cell types, such as endothelial cells, osteoblasts, and macrophages, and plays a crucial role in the regulation of the functions of mononuclear phagocytes, including their survival, proliferation, and differentiation. It contributes to inflammation, which in turn leads to macrophage destruction and the progression of atherosclerosis. Furthermore, M-CSF triggers tissue factor production, which can result in plaque thrombosis and instability, which can lead to acute coronary syndrome [44]. Elevated levels of M-CSF have been observed in the bloodstream not only in cardiovascular diseases (CVDs) but also in various other diseases, such as tumors [45]. To investigate the impact of inhibiting M-CSF, we removed transition \(t_{21}\) (local production of M-CSF in the vessel wall) from the model under study. As a result, it was possible to observe that the atherosclerotic plaque formation process was stopped in the analyzed model (see Figure 4). However, the altered cholesterol homeostasis balance still causes an accumulation of apoB lipoproteins, leading to lipid accumulation. This process triggers cellular responses within the artery wall and, subsequently, increases local inflammation. However, it is important to note that this accumulation does not directly cause plaque development in the model. If the transition \(t_{20}\) (defective cholesterol homeostasis) is also blocked, then these processes stop. Furthermore, the secretion of pro-inflammatory cytokines such as IL-17 and IL-23 is inhibited, affecting the G-CSF cytokine secretion process, which IL-17 usually stimulates. Similarly, whether \(t_{21}\) is inhibited alone or in conjunction with \(t_{20}\), the level of interleukin-10 (IL-10) produced by M2 macrophages, a cytokine with wide-ranging anti-inflammatory properties [2], remains unchanged (\(p_{34}\)). Simultaneously, the M1 phenotype of the pro-inflammatory macrophage is inhibited (\(t_{0}\) (monocyte differentiation from Lyc6C\({}^{high}\)) and \(p_{0}\) (M1 macrophages)). The results presented in this scenario align with the findings of a previous study [46], which demonstrated that the absence of M-CSF leads to a decrease in macrophage proliferation, an increase in macrophage apoptosis within lesions, and a significant reduction in the development of atherosclerosis. **Scenario 3**. : _The impact of inhibition of IL-23 cytokine._ \begin{table}
\begin{tabular}{l l} \hline
**Disabled Transitions** & **Average Chance of Firing for \(t_{41}\)** \\ \hline \hline Nothing is knocked out in the model & 4.03\% (reference value) \\ \hline \(t_{32}\), \(t_{33}\) & 3.97\% (\(-\)0.05\%) \\ \hline \(t_{12}\), \(t_{13}\) & 3.36\% (\(-\)0.67\%) \\ \hline \(t_{11}\), \(t_{16}\), \(t_{17}\), \(t_{18}\), \(t_{19}\) & 2.69\% (\(-\)1.34\%) \\ \hline \(t_{9}\) & 1.29\% (\(-\)2.74\%) \\ \hline \(t_{0}\), \(t_{1}\), \(t_{2}\), \(t_{3}\), \(t_{4}\), \(t_{5}\), \(t_{6}\), \(t_{7}\), \(t_{10}\), \(t_{20}\), \(t_{21}\), \(t_{22}\), \(t_{23}\), \(t_{40}\) & 0.0\% (\(-\)4.03\%) \\ \hline \end{tabular}
\end{table} 
Table 5: The effect of knocking out selected transitions on the atherosclerosis process (\(t_{41}\)) is shown. The change in firing probability is indicated in parentheses, represented as a percentage point difference compared to the reference firing value for \(t_{41}\), which is provided in the first row (where no knockouts occurred).