Impact of Dextran-Sodium-Sulfate-Induced Enteritis on Murine Cytomegalovirus Reactivation

Abstract

(1) Background: Ulcerative colitis (UC) is an inflammatory bowel disease that causes inflammation of the intestines, which participates in human cytomegalovirus (HCMV) reactivation from its latent reservoir. CMV-associated colitis plays a pejorative role in the clinical course of UC. We took advantage of a model of chemically induced enteritis to study the viral reactivation of murine CMV (MCMV) in the context of gut inflammation. (2) Methods: Seven-week-old BALB/c mice were infected by 3 × 10³ plaque-forming units (PFU) of MCMV; 2.5% (w/v) DSS was administered in the drinking water from day (D) 30 to D37 post-infection to induce enteritis. (3) Results: MCMV DNA levels in the circulation decreased from D21 after infection until resolution of the acute infection. DSS administration resulted in weight loss, high disease activity index, elevated Nancy index shortening of the colon length and increase in fecal lipocalin. However, chemically induced enteritis had no impact on MCMV reactivation as determined by qPCR and immunohistochemistry of intestinal tissues. (4) Conclusions: Despite the persistence of MCMV in the digestive tissues after the acute phase of infection, the gut inflammation induced by DSS did not induce MCMV reactivation in intestinal tissues, thus failing to recapitulate inflammation-driven HCMV reactivation in human UC.

Keywords: DSS-induced enteritis; mouse model; murine cytomegalovirus; viral reactivation.

Figure 1

Flowchart of the study. Seven-week-old female BALB/c mice were divided into four groups (5 animals per group): group 1 = negative controls; group 2 = mice treated by dextran sodium sulfate (DSS) in the drinking water from D30 to D37; group 3 = mice infected by murine cytomegalovirus (MCMV) at D0; and group 4 = mice infected by MCMV at D0 then treated by DSS in the drinking water from D30 to D37.

Figure 2

Kinetics of blood MCMV DNA load measured by qPCR for infected mice. Group 3 (red curve, (A)) and group 4 (purple curve, (B)). D0 corresponds to the time of infection with MCMV and D37 represents the time of sacrifice. The arrow denotes the beginning of dextran sodium sulfate (DSS) treatment in group 4. The limit of PCR quantification is represented by the horizontal line. The statistical test that was used was a two-way ANOVA with Bonferroni correction for panels. Standard variations are too low to be seen in the figures.

Figure 3

Monitoring of weight loss, clinical score and gut inflammation during DSS-induced colitis of MCMV-infected mice. Monitoring of clinical, biological and histopathological parameters. Mice were infected at D0 or not by murine cytomegalovirus (MCMV) and treated or not by dextran sodium sulfate (DSS) from D30 to D37 post-infection (p.i.) before being euthanized. (A)—Longitudinal weight loss assessment from D30 to D37. (B)—Evolution of the mean Disease Activity Index (DAI) from D30 to D37 p.i. (C)—Length (in cm) of the colon at the time of sacrifice, expressed as a mean. (D)—Longitudinal assessment of faecal lipocalin levels (in pg/mL) post treatment by DSS, values are expressed as means. (E)—Determination of the Nancy histological score (from 0 to 4) of the small intestine specimen from individual mice at sacrifice (D37 p.i.). (F)—Determination of the Nancy histological score (from 0 to 4) of the colon specimen. Statistical tests that were used were a two-way ANOVA with Bonferroni correction for panels (A,B,D), and a one-way ANOVA for panel (C,E,F). ns: not statistically significant, * p < 0.05, ** p < 0.01, *** p < 0.001.

Figure 4

Exploration of MCMV reactivation in the gut of mice. (A)—Proportion of mice for which murine cytomegalovirus (MCMV) DNA was detected in small intestine specimens by PCR; positive mice are shown in black and negative ones in grey. (B)—For colon specimens. (C)—Immunohistochemistry at D37 (day of euthanasia) on gut specimens of a mouse of group 3 (upper part) or of group 4 (lower part). Staining corresponds to positive zones marked by horseradish peroxidase and revealed by diaminobenzidine substrate. The negative control was performed by omitting the primary antibody. The positive control was performed by using anti-alpha smooth muscle actin (anti-αSMA) as the primary antibody. The presence of a replicative virus was looked for by using an anti-immediate early (IE) 1 protein of MCMV as primary antibody. The scale bar on images corresponds to 50 µm. Photographs were taken at 20× objective in Zeiss Axioimager Apotome 3 optical microscope with a Zeiss quadriCCD Axiocam camera.