The intricate ballet of inflammation and autophagy: Insights from Mycoplasma gallisepticum-infected HD11 cells

Abstract

Objectives

Mycoplasma gallisepticum (M. gallisepticum) infection often leads to inflammatory damage and immunosuppression. Macrophages play a crucial role as the primary immune defense in chickens, with their inflammatory response and autophagy levels critical. This study aimed to explore the relationship between NLRP3 inflammasome and autophagy in HD11 cells within 12 h after M. gallisepticum infection.

Methods

To investigate this, the HD11 cell model of M. gallisepticum infection was established using the CCK8 (Cell Counting Kit-8) method in this study. The study observed changes in M. gallisepticum-induced inflammation, oxidative stress, and autophagy levels through various methods including transmission electron microscopy (TEM), RT-qPCR, ELISA (enzyme linked immunosorbent assay), 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA), JC-1 and Western blot.

Results

TEM revealed that the nuclear membrane was damaged, the number of damaged mitochondria increased, and autophagosomes were detected at 4 and 8 h after M. gallisepticum infection. ELISA and RT-qPCR results indicated that M. gallisepticum induced oxidative stress and inflammation damage. Fluorescence analysis demonstrated an increase in intracellular reactive oxygen species (ROS) content and a continuous decrease in mitochondrial membrane potential (MMP) post-M. gallisepticum infection. Additionally, the NF-κB/NLRP3 signaling pathway remained consistently activated during M. gallisepticum infection. After M. gallisepticum infection, autophagy levels decreased significantly at 1 and 12 h, but increased significantly at 4 and 8 h.

Conclusions

M. gallisepticum infection triggers the activation of the NLRP3 inflammasome in HD11 cells, leading to inflammatory damage. Additionally, it causes fluctuations in autophagy levels, characterized by a wavy pattern of decrease, increase, and subsequent decrease.