Expression of matrix metalloproteinases subsequent to urogenital Chlamydia muridarum infection of mice.

The central hypothesis of this study was that matrix metalloproteinases (MMPs) would be enhanced following murine chlamydial infection and that their expression would vary in mouse strains that differ in their susceptibility to chronic chlamydia-induced disease. To address this hypothesis, female C3H/HeN and C57BL/6 mice were infected intravaginally with Chlamydia muridarum. Uterine and oviduct tissues were assessed for transcription of MMP genes and their tissue inhibitors. An increased activity of MMP genes relative to preinfection tissues was observed in the C3H/HeN mice when compared to C57BL/6 mice. Using gelatin zymography, we detected constitutive MMP-2 activity in both strains of mice but an increase in MMP-9. Casein zymography indicated the presence of two elastase-like activities consistent with MMP-12 and possibly MMP-7. Western blotting and antigen capture enzyme-linked immunoassay also confirmed an increase in MMP-9 but constitutive MMP-2 expression subsequent to the infection in both strains of mice. In C57BL/6 mice, MMP-9 was present in monomer and dimer form throughout the 56-day monitoring period. C3H/HeN mice produced dimeric MMP-9, but increases in the monomer form were also observed through day 14. Post-translational modification of MMP-9 between the two strains also differed. Immunohistochemistry revealed neutrophils as a prominent source for MMP-9 in both strains of mice. We conclude that differences in the relative expression and activity of MMPs, particularly MMP-9, occur in mice differing in their susceptibility to the development of chronic chlamydial disease. These differences may account for disparate outcomes with regard to chronic sequelae of the disease.

Inducible nitric oxide synthase regulates production of isoprostanes in vivo during chlamydial genital infection in mice.

Urinary nitrite and F(2)-isoprostanes, an index of oxidant stress, were elevated during chlamydial genital infection of mice. Enhancement of urinary nitrite and F(2)-isoprostanes was observed in phagocyte oxidase-deficient mice. Inhibition of inducible nitric oxide synthase reduced isoprostane excretion. We conclude that nitrogen radicals induce F(2)-isoprostane production and excretion during murine chlamydial genital infection.

Role for inducible nitric oxide synthase in protection from chronic Chlamydia trachomatis urogenital disease in mice and its regulation by oxygen free radicals.

It has been previously reported that although inducible nitric oxide synthase (iNOS) gene knockout (NOS2(-/-)) mice resolve Chlamydia trachomatis genital infection, the production of reactive nitrogen species (RNS) via iNOS protects a significant proportion of mice from hydrosalpinx formation and infertility. We now report that higher in vivo RNS production correlates with mouse strain-related innate resistance to hydrosalpinx formation. We also show that mice with a deletion of a key component of phagocyte NADPH oxidase (p47(phox-/-)) resolve infection, produce greater amounts of RNS in vivo, and sustain lower rates of hydrosalpinx formation than both wild-type (WT) NOS2(+/+) and NOS2(-/-) controls. When we induced an in vivo chemical block in iNOS activity in p47(phox-/-) mice using N(G)-monomethyl-L-arginine (L-NMMA), a large proportion of these mice eventually succumbed to opportunistic infections, but not before they resolved their chlamydial infections. Interestingly, when compared to WT and untreated p47(phox-/-) controls, L-NMMA-treated p47(phox-/-) mice resolved their infections more rapidly. However, L-NMMA-treated p47(phox-/-) mice lost resistance to chronic chlamydial disease, as evidenced by an increased rate of hydrosalpinx formation that was comparable to that for NOS2(-/-) mice. We conclude that phagocyte oxidase-derived reactive oxygen species (ROS) regulate RNS during chlamydial urogenital infection in the mouse. We further conclude that while neither phagocyte oxidase-derived ROS nor iNOS-derived RNS are essential for resolution of infection, RNS protect from chronic chlamydial disease in this model.

Chlamydia trachomatis persistence in the female mouse genital tract: inducible nitric oxide synthase and infection outcome.

It was previously reported that female mice resolve a primary Chlamydia trachomatis urogenital infection independent of inducible nitric oxide synthase (iNOS). We now report that although iNOS-deficient (NOS2(-/-)) mice resolve culture-apparent infection in a fashion similar to that of normal control (NOS2(+/+)) mice, they sustain significantly increased rates of disease, as assessed by hydrosalpinx formation. PCR amplification of ompA followed by Southern blot detection of amplicands revealed the presence of chlamydial DNA in the lower genital tracts of both NOS2(-/-) and NOS2(+/+) mice at > or =120 days postinfection and in upper genital tract tissues at >120 days postinfection. However, only NOS2(-/-) mice shed low numbers of viable chlamydiae from the lower genital tract after immunosuppressive treatment at 120 days postinfection. When cultured primary murine lung fibroblasts were activated in the presence of gamma interferon (IFN-gamma), inhibition of chlamydial growth occurred in both NOS2(+/+) and NOS2(-/-) cells, but the inhibition was reversible after removal of the cytokine in the NOS2(-/-) primary cell culture only. The iNOS-independent inhibition was microbistatic but was independent of 2,3-indoleamine dioxygenase activity. We conclude that chlamydial DNA and antigens persist in mice subsequent to culture-apparent resolution. In addition, IFN-gamma induces in vivo inhibition of chlamydial growth through microbistatic mechanisms in the absence of iNOS activity, but in the presence of iNOS activity, IFN-gamma is microbicidal and effects eradication.