Pseudorabies virus causes splenic injury via inducing oxidative stress and apoptosis related factors in mice

Abstract

Pseudorabies virus (PRV) is an immunosuppressive virus that causes significant damage to the pig industry. This study aimed to investigate the effects of PRV on oxidative stress and apoptotic related in the spleen of mice to provide basis knowledge for further research on the pathogenesis of PRV in mice model. 36 mice were randomly two groups, the control group which only received 200 μL PBS and infection group which was subcutaneously infected with 200 μL of 1 × 10³ TCID₅₀/100 μL PRV, respectively. Spleen tissues in each group were collected for further experiments at 48, 72, and 96 h post-infection (hpi). Pathological observation was performed by hematoxylin and eosin Y staining. Biochemical and Flow cytometry methods were used to determine the reactive oxygen species profile and apoptosis of the spleen post-infection and apoptosis detection. In addition, q-PCR and Western blot were adopted to measure the apoptotic conditions of the spleen infected with PRV. The results indicated that the reactive oxygen species (ROS) level in the PRV infection group was remarkedly increased (p < 0.01) at a time-dependent pattern. Furthermore, the Malondialdehyde levels in the spleen of mice in the infection group increased (p < 0.01) in a time-dependent mode. However, the activity of Catalase, Superoxide dismutase, and glutathione peroxidase and the content of Glutathione in the infection group were decreased with the control group (p < 0.01) at a time-dependent manner. In addition, the ratio of splenocyte apoptosis in the infection group significantly increased (p < 0.01) in a time-dependent manner. In conclusion, PRV infection causes apoptosis of the spleen via oxidative stress in mice.

Figure 1

The pathological observation on Spleen (400 × , Bar = 50 μm). (A) Control; (B) 48 hpi; (C) 72 hpi; (D) 96 hpi congestion (yellow arrow) and infiltration of neutrophils (green arrow).

Figure 2

The inducement of PRV on production of ROS in the spleen. (A) The results of ROS detected by flow cytometry; (B) ROS statistical analysis histogram. **p < 0.01 versus mock infection, the same as below.

Figure 3

The effect of PRV infection mitochondrial oxidative stress-related factors. (A) MDA; (B) CAT; (C) SOD; (D) GSH; (E) GSH-Px. *p < 0.05 versus mock infection, the same as below.

Figure 4

Effect of PRV on the percentage of splenocyte apoptosis. (A) The results of apoptosis detected by flow cytometry; (B) Apoptosis statistical analysis histogram.

Figure 5

The relative mRNA expression of apoptosis-related factors caused by PRV.

Figure 6

The expression levels of apoptosis-related factors induced by PRV in protein. (A) The relative protein amount of Caspase-3, Caspase-9, Bax, and Bcl-2 to β-actin. (B) Protein expression related to apoptosis detected by western blot.