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Lipopolysaccharide Induces Interleukin-8 Production By Human Cervical Smooth Muscle Cells

Departments of Obstetrics and gynecology and Molecular Biochemistry, Hokkaido University School of Medicine, Sapporo, Hokkaido, Japan; Center for Reserch on Reproduction and Women's Health, University of Pennsylvania Medical Center, Philadelphia, Pennsylvania

Hidemichi Watari, MD, PhD

Departments of Obstetrics and gynecology and Molecular Biochemistry, Hokkaido University School of Medicine, Sapporo, Hokkaido, Japan; Center for Reserch on Reproduction and Women's Health, University of Pennsylvania Medical Center, Philadelphia, Pennsylvania; hmwatari{at}med.hokudai.ac.jp

Toshio Fujimoto, MD, PhD

Hideto Yamada, MD, PhD

Jun Nishihira, MD, PhD

Jerome f. Strauss, III, MD, PhD

Seiichiro Fujimoto, MD, PhD

Departments of Obstetrics and gynecology and Molecular Biochemistry, Hokkaido University School of Medicine, Sapporo, Hokkaido, Japan; Center for Reserch on Reproduction and Women's Health, University of Pennsylvania Medical Center, Philadelphia, Pennsylvania

Objective: We examined the effect of lipopolysaccharide (LPS), a component of the outer wall of gram-negative bacteria, on expression of the neutrophil chemoattractant interleukin-8 (IL-8) and the effects of IL-8 treatment on release of matrix metabolizing enzymes in human cervical smooth muscle cells (CSMCs).

Methods: Human CSMCs were exposed to Escherichia coli LPS, and the expression of IL-8 mRNA was analyzed by Northern blotting. The Il-8 promoter activity was examined by dual luciferase assay, and the IL-8 concentration was assessed by enzyme-linked immunosorbent assay. We also treated the CSMCs with human IL-8 and examined the expression of matrix-degrading enzymes.

Results: E coli LPS (100 ng/mL) increased the expression of IL-8 mRNA 12.8-fold after 3 hours. This up-regulation was maintained for up to 24 hours. Lipopolysaccharide treatment produced a fivefold increase in IL-8 promoter activity in CSMCs transfected with an Il-8 promoter-reporter construct. IL-8 concentrations in conditioned medium of CSMC cultures treated with E coli LPS increased approximately 18-fold compared with the control culture. Northern blot analysis and zymography revealed that exogenous human IL-8 had no significant effect on the expression of matrix metalloproteinase (MMP)-1, -3, and tissue inhibitor of metalloproteinase (TIMP)-1 mRNAs, and on the secretion MMP-2 and -9 in CSMCs.

Conclusion: We conclude that CSMCs respond to LPS with increased expression of IL-8 mRNA and secreted IL-8, and that expression of matrix metabolizing enzymes in CSMCs is not directly affected by IL-8. IL-8 produced by CSMCs in reponse to gram-negative infection may promote neutrophil invasion, and release of neutrophil matrix-degrading enzymes may participate in the matrix remodeling associated with parturition.

Key Words: Bacterial vaginosis • cervical ripening • human cervical smooth muscle cells • interleukin-8 • lipopolysaccharide

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