Intervention time decides the status of autophagy, NLRP3 activity and apoptosis in macrophages induced by ox-LDL

Background: It has been determined through extensive studies that Autophagy, the Nucleotide-binding oligomerization domain-like receptor containing pyrin domain 3 (NLRP3) inflammasome and apoptotic responses in macrophages jointly contribute to atherogenesis and its development in the presence of lipid abnormalities. Few studies have investigated in full-scale if the intervention time for lipids abnormality or NLRP3 activation have a significant effect on Autophagy, NLRP3 or the apoptotic status in macrophages.

Methods: Human THP-1 monocyte-derived macrophages were established by challenging THP-1 monocytes with 80 µg/ml oxidized low-density lipoprotein (ox-LDL) for specific durations. Foam cell formation was observed by Oil Red O (ORO) staining. Western blots were employed to determine protein expression. Transmission electron microscope (TEM) and immunofluorescence microscopy were applied to observe the autophagic status of cells. Cell Apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL).

Results: The cells were treated with ox-LDL for 12 h and 36 h, which were considered to represent early and advanced stages of atherogenesis for this study. The results showed that inhibition of ox-LDL phagocytosis by cytochalasin D in the early stage improved autophagic status, reduced NLRP3 activation and the apoptotic response significantly. In contrast, cytochalasin D had little effect on blocking the detrimental effect of ox-LDL at the advanced stage. Moreover, the changes in Autophagy, Apoptosis and NLRP3 expression after treatment with small interfering (si) RNA targeting NLRP3 in the early and advanced stages of atherogenesis were consistent with the above data.

Conclusions: Interventions against lipid disorders or inflammatory reactions in the early or advanced stages of atherogenesis may have different results depending on when they are applied during the process of atherosclerotic pathogenesis. These results may help improve therapeutic strategies for atherosclerosis prevention. Furthermore, a healthy lifestyle should still be recommended as the most important and inexpensive measure to prevent atherogenesis.

Apoptosis; oxidized low-density lipoprotein; Atherosclerosis; Autophagy; Macrophage; Nucleotide-binding oligomerization domain-like receptor containing pyrin domain 3.