J Vet Sci.  2016 Sep;17(3):413-419. 10.4142/jvs.2016.17.3.413.

Establishment of hydrochloric acid/lipopolysaccharide-induced pelvic inflammatory disease model

• Oh Y
• Lee J
• Kim HC
• Hahn TW
• Yoon BI
• Han JH
• Kwon YS
• Park JJ
• Koo DB
• Rhee KJ
• Jung BD

• KMID: 2413143
• DOI: http://doi.org/10.4142/jvs.2016.17.3.413

Abstract

Pelvic inflammatory disease (PID), which is one of the most problematic complications experienced by women with sexually transmitted diseases, frequently causes secondary infections after reproductive abnormalities in veterinary animals. Although the uterus is self-protective, it becomes fragile during periods or pregnancy. To investigate PID, bacteria or lipopolysaccharide (LPS) extracted from gram negative bacteria has been used to induce the disease in several animal models. However, when LPS is applied to the peritoneum, it often causes systemic sepsis leading to death and the PID was not consistently demonstrated. Hydrochloric acid (HCl) has been used to induce inflammation in the lungs and stomach but not tested for reproductive organs. In this study, we developed a PID model in mice by HCl and LPS sequential intracervical (i.c.) administration. The proinflammatory cytokines, interleukin (IL)-1β, IL-6 and tumor necrosis factor-α, were detected in the mouse uterus by western blot analysis and cytokine enzyme-linked immunosorbent assay after HCl (25 mg/kg) administration i.c. followed by four LPS (50 mg/kg) treatments. Moreover, mice exhibited increased infiltration of neutrophils in the endometrium and epithelial layer. These results suggest that ic co-administration of HCl and LPS induces PID in mice. This new model may provide a consistent and reproducible PID model for future research.

Keyword

hydrochloric acid; lipopolysaccharide; mice; pelvic inflammatory disease; pelvic inflammatory disease model

MeSH Terms

Animals
Blotting, Western/veterinary
Cytokines/*metabolism
*Disease Models, Animal
Female
Humans
*Hydrochloric Acid
Inflammation/chemically induced/*veterinary
*Lipopolysaccharides
*Mice
Mice, Inbred C57BL
Pelvic Inflammatory Disease/*chemically induced
Uterus/immunology/pathology
Cytokines
Lipopolysaccharides
Hydrochloric Acid

Figure

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  Fig. 1 Mean value of clinical sign scoring. ***Significantly more serious clinical signs in lipopolysaccharide (LPS) intraperitoneal administered animals compared with LPS or hydrochloric acid (HCl)-LPS intracervical (i.c.) administered animals (p < 0.001).

• 

  Fig. 2 Changes in cytokine expression in uterine tissue after co-administration of HCl and LPS. (A) Mice were administered i.c. with HCl (25 mg/kg) followed by four applications of LPS (25 mg/kg or 50 mg/kg) every 2 h. Two hours after the final injection, the uterus was removed and protein expression was assessed by Western blot. (B) Mice were administered i.c. with HCl (25 mg/kg, 1 N) followed by four applications of LPS (50 mg/kg) every 2 h.

• 

  Fig. 3 Bar graphs for inflammatory cytokines measured by enzyme-linked immunosorbent assay in the uterine tissues of different experimental groups and a normal control. *Significantly higher inflammatory cytokine levels than the HCl25 and Neg CTL groups (p < 0.05). ***Significantly higher inflammatory cytokine levels in the uteruses of HCl-LPS combined with i.c. administered animals (p < 0.001).

• 

  Fig. 4 The representative histopathological findings of the uterus with saline only (A) and co-administration of HCl and LPS (B). (B) Mice were administered i.c. with HCl (25 mg/kg. 1 N) followed by four applications of LPS (50 mg/kg) every 2 h. Note the neutrophils in the lumen, in the endometrium and in the surface epithelial layer (arrows). No inflammation was observed in (A). L, lumen. H&E stain. 200×.

• 

  Fig. 5 Bar graphs for histopathological scoring in different experimental groups. *Significantly higher histopathological scores in LPS25ic and LPS50ic groups than in LPS25ip, LPS50ip, HCl25, and Neg CTL groups (p < 0.05). ***Significantly higher histopathological scores in HCl-LPS25 and HCl-LPS 50 groups than with other groups (p < 0.001).

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  Fig. 6 Bar graphs for the number of infiltrated neutrophils per unit area (0.25 mm2) in uterine tissue of different experimental groups. *Significantly increased number of neutrophils in LPS25 and LPS50 groups than in LPS25ip, LPS50ip, and Negative control groups (p < 0.05). ***Significantly increased number of neutrophils in HCl-LPS25 and HCl-LPS 50 groups compared with other groups (p < 0.001).

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