Ivermectin reduces in vivo coronavirus infection in a mouse experimental model

Abstract

The objective of this study was to test the effectiveness of ivermectin for the treatment of mouse hepatitis virus (MHV), a type 2 family RNA coronavirus similar to SARS-CoV-2. Female BALB/cJ mice were infected with 6,000 PFU of MHV-A59 (group infected, n = 20) or infected and then immediately treated with a single dose of 500 µg/kg ivermectin (group infected + IVM, n = 20) or were not infected and treated with PBS (control group, n = 16). Five days after infection/treatment, the mice were euthanized and the tissues were sampled to assess their general health status and infection levels. Overall, the results demonstrated that viral infection induced typical MHV-caused disease, with the livers showing severe hepatocellular necrosis surrounded by a severe lymphoplasmacytic inflammatory infiltration associated with a high hepatic viral load (52,158 AU), while mice treated with ivermectin showed a better health status with a lower viral load (23,192 AU; p < 0.05), with only a few having histopathological liver damage (p < 0.05). No significant differences were found between the group infected + IVM and control group mice (P = NS). Furthermore, serum transaminase levels (aspartate aminotransferase and alanine aminotransferase) were significantly lower in the treated mice than in the infected animals. In conclusion, ivermectin diminished the MHV viral load and disease in the mice, being a useful model for further understanding this therapy against coronavirus diseases.

Figure 1

Representative liver and spleen from each group: (A) infected; (B) infected + IVM; (C) control. Upper panel: abdominal cavity at necropsy; middle panel: dissected liver and spleen; lower panel: HE histological sections of livers. White arrows indicate white spotted patterns in the liver from infected mice and severe hepatocellular necrosis and lymphoplasmacytic inflammatory infiltration in the histological images (A). IVM ivermectin.

Figure 2

Body weight at the beginning and the end of the experiment (a) and organ weight and liver appearance at necropsy five days postinfection (b,c,d, respectively) in MHV-infected (infected group, n = 20), infected and treated with ivermectin (infected + IVM group, n = 20), and noninfected untreated mice (control group, n = 16) (mean ± SD). Different letters indicate significant differences (p < 0.05) between pre- and postinfection timepoints for a and significant differences (p < 0.05) between the indicated groups for (b–d).

Figure 3

Histopathological scores (grades 0 to 3, where 0 is no damage and 3 is the most damaged) of livers from the MHV-infected (infected group, n = 20), infected and treated with ivermectin (infected + IVM group, n = 20), and noninfected untreated mice (control group, n = 16). The distribution of animals for each score showed significant differences between the groups (p < 0.05).

Figure 4

Hepatic viral load measured by qPCR in liver samples from MHV-infected (infected group, n = 20), infected and treated with ivermectin (infected + IVM group, n = 20), and noninfected untreated mice (control group, n = 16) (mean ± SD). Different letters indicate significant differences (p < 0.05) among the groups.

Figure 5

Protein levels measured in the blood of mice in the MHV-infected (infected group, n = 20), infected and treated with ivermectin (infected + IVM group, n = 20), and noninfected untreated mice (control group, n = 16) before and after infection with MHV A-59. (a) Total proteins; (b) albumin; (c) globulin, (d) albumin/globulin ratio and (e) total bilirubin (mean ± SD). Different letters indicate significant differences (p < 0.05) between the pre- and post-infection timepoints; asterisks (*) refer to significant differences (p < 0.05) between indicated groups.

Figure 6

Hepatic enzyme levels in the blood of MHV-infected (infected group, n = 20), infected and treated with ivermectin (infected + IVM group, n = 20), and noninfected untreated mice (control group, n = 16) before and after virus infection. (a) Alanine aminotransferase; (b) aspartate aminotransferase and (c) gamma glutamyl transpeptidase (mean ± SD). Different letters indicate significant differences (p < 0.05) between pre- and postinfection timepoints; asterisks (*) refer to significant differences (p < 0.05) between the indicated groups.

Figure 7

Blood urea nitrogen (BUN), creatinine (CRE), BUN/CRE ratio and glucose (GLU) levels were measured before and after infection in MHV-infected (infected group, n = 20), infected and treated with ivermectin (infected + IVM group, n = 20), and noninfected untreated mice (control group, n = 16). The results are shown in (a–d) (mean ± SD). Different letters indicate significant differences (p < 0.05) between pre- and postinfection timepoints; asterisks (*) refer to significant differences (p < 0.05) between the indicated groups.

Figure 8

Evaluation of hematological parameters in mice. (a) White blood cell (WBC) count, (c,d) neutrophil, lymphocyte and monocyte counts and percentages were determined in blood samples before and after infection for each experimental group (infected, infected + IVM, and control) (mean ± SD). (b) Lymphocyte staining for the expression of surface markers for B and T cells at the end point of the experiment for all groups (infected, infected + IVM, and control) (mean ± SD). Different letters indicate significant differences (p < 0.05) between pre- and postinfection timepoints; asterisks (*) refer to significant differences (p < 0.05) between the indicated groups.

Figure 9

Detection of cytokines in the plasma of all experimental groups (infected, infected + IVM, and control). Murine plasma was obtained 5 days postinfection, and cytokine concentrations were determined by multiplex bead array (mean ± SD). Different letters indicate statistically significant differences (p < 0.05).