Mouse Strain-Dependent Differences in Susceptibility to Neisseria gonorrhoeae Infection and Induction of Innate Immune Responses

Acute gonorrhea in women is characterized by a mucopurulent exudate that contains polymorphonuclear leukocytes (PMNs) with intracellular gonococci. Asymptomatic infections are also common. Information on the innate response to Neisseria gonorrhoeae in women is limited to studies with cultured cells, isolated immune cells, and analyses of cervicovaginal fluids. 17β-Estradiol-treated BALB/c mice can be experimentally infected with N. gonorrhoeae, and a vaginal PMN influx occurs in 50 to 80% of mice. Here, we compared the colonization loads and proinflammatory responses of BALB/c, C57BL/6 and C3H/HeN mice to N. gonorrhoeae. BALB/c and C57BL/6 mice were colonized at similar levels following inoculation with 10⁶ CFU of N. gonorrhoeae. BALB/c, but not C57BL/6, mice exhibited a marked vaginal PMN influx. Tumor necrosis factor alpha, interleukin-6, macrophage inflammatory protein 2 (MIP-2), and keratinocyte-derived chemokine were elevated in vaginal secretions from infected BALB/c mice, but not in those from C57BL/6 mice. MIP-2 levels positively correlated with a vaginal PMN influx. In contrast to BALB/c and C57BL/6 mice, C3H/HeN mice were resistant to infection, and there was no evidence of an inflammatory response. We conclude that N. gonorrhoeae causes a productive infection in BALB/c mice that is characterized by the induction of proinflammatory cytokines and chemokines and the recruitment of PMNs. Infection of C57BL/6 mice, in contrast, is more similar to asymptomatic infection. C3H/HeN mice are inherently resistant to N. gonorrhoeae infection, and this resistance is not due to an overwhelming inflammatory response to infection. Host genetic factors can therefore impact susceptibility and the immune response to N. gonorrhoeae.Uncomplicated gonorrhea is most commonly an infection of the urethra in men and the cervix. The female urethra may also be infected, and rectal and pharyngeal infection can occur in either sex. The hallmark of symptomatic gonococcal infection is the presence of a purulent exudate containing numerous polymorphonuclear leukocytes (PMNs), many of which contain intracellular gonococci. Asymptomatic infections are also common, particularly in females. Epithelial cells that line the genital mucosal surface are the first line of defense against this human-specific pathogen, and Neisseria gonorrhoeae produces a robust proinflammatory cytokine and chemokine response when incubated with cultured human vaginal, endocervical, ectocervical (12, 33), urethral (17), endometrial (3), and fallopian tube (31) tissue culture cells. Similarly, studies using the complex fallopian tube organ culture model suggest that N. gonorrhoeae induces the proinflammatory cytokines interleukin-1α (IL-1α) and tumor necrosis factor alpha (TNF-α) (29). Signaling through cellular receptors on epithelial cells results in the activation and recruitment of phagocytic cells, including PMNs and macrophages. Primary macrophages and peripheral blood mononuclear cells also elicit a proinflammatory response when incubated with N. gonorrhoeae (30, 34, 39). These innate immune cells further contribute to the array of proinflammatory cytokines and antimicrobial factors.Due to the multiple cell types that contribute to the host innate response to infection, it is important that whole model systems be utilized to measure the impact of N. gonorrhoeae infection on the host immune response. Experimental urethral colonization in male volunteers with N. gonorrhoeae evokes a strong innate response that is characterized by the production of proinflammatory cytokines (37, 38). Similar studies with female subjects are not feasible due to the risk of complications of gonococcal infection in women. Therefore, features of the innate response to N. gonorrhoeae in the female genital tract are predicted solely from tissue culture systems and the analysis of clinical samples. It is unclear whether women elicit a cytokine response to gonococcal infection. The reason why some infections are asymptomatic is also not known. Hedges et al. (18, 19) were unable to detect local proinflammatory cytokines in cervicovaginal secretions from infected women and detected a low anti-gonococcal antibody response. Based on these observations, it was proposed that N. gonorrhoeae fails to induce host inflammatory responses or is actively immunosuppressive. This finding is in marked contrast with the robust induction of proinflammatory cytokines observed from in vitro cell lines that constitute the female genital tract. The absence of various other cell types in tissue culture cell models could influence the cytokine response to infection; alternatively, the timing of sample collection from infected subjects may also influence the data. Therefore, a systematic analysis of cytokine induction over the course of infection in a female animal model is needed.The 17β-estradiol-treated mouse model is the only small-animal model available for studying the immune response to N. gonorrhoeae genital tract infection. While the mechanism by which estradiol promotes long-term colonization in female mice is not known, it is likely that promotion of an estrus-like state is beneficial for the gonococcus based on the fact that untreated mice can be transiently colonized with N. gonorrhoeae provided they are inoculated in the proestrus stage of the reproductive cycle (7, 46). The 17β-estradiol-treated mouse model has been a useful system for studying many aspects of gonococcal infection, including gonococcal evasion of PMN killing (43, 49) and antimicrobial peptides (23, 48), antigenic variation in vivo (41), and interactions between N. gonorrhoeae and commensal flora (32). This model is based on the use of BALB/c mice. Approximately 50 to 80% of infected BALB/c mice that are treated with a slow-release estradiol pellet exhibit a significant vaginal PMN response following inoculation with N. gonorrhoeae strain FA1090 based on examination of stained vaginal smears (21, 22, 43), and PMNs and macrophages are also found in vaginal and cervical tissue samples from infected mice (44). Gonococci are localized within vaginal and cervical tissue, and similar to that which occurs in humans, an insignificant and transient humoral response to N. gonorrhoeae occurs which was not protective against reinfection with the same strain (44). A recent modification of the model utilizes water-soluble estradiol to reduce the length of time that mice are exposed to nonphysiological concentrations of estradiol. A vaginal PMN influx also occurs during infection of mice treated with water-soluble estradiol, and as with pelleted mice, infection persists despite the presence of PMNs (44).One advantage of using inbred mouse strains for studies of infectious diseases is that environmental and genetic components can be controlled. Interestingly, the susceptibility to infectious agents can often vary with the genetic background of the mouse. One example in the area of sexually transmitted infections is that genetically controlled differences in the development of infertility in inbred mouse strains following inoculation with chlamydia have been reported, with pregnancy rates following infection of C3H/HeN mice being significantly lower than those of C57BL/6 mice (10). Darville and colleagues (8) found similar results, and their data suggested that an earlier and more severe acute inflammatory response in the C57BL/6 strain may lead to earlier eradication of the infection, thus protecting the upper tract from disease. Numerous examples of vulnerability have been found for other infectious agents, including Leishmania major, Listeria monocytogenes, Salmonella enterica serovar Typhimurium, Plasmodium chabaudi, Legionella pneumophila, and Mycobacterium tuberculosis (reviewed by Kramnik and Boyartchuk [27]).In this study, we sought to characterize in greater detail the cytokine and inflammatory response to genital tract infection with N. gonorrhoeae in 17β-estradiol-treated BALB/c mice and to determine if susceptibility to colonization and the host inflammatory response to infection vary between inbred mouse strains. Our data demonstrate that BALB/c, C57BL/6, and C3H/HeN mouse strains differ widely in their response to infection. While both BALB/c and C57BL/6 strains support colonization with the gonococcus, only the BALB/c strain appears to mount an inflammatory response. In contrast, the C3H/HeN strain appears to be resistant to colonization with the gonococcus. These data demonstrate significant divergence among inbred mouse strains in terms of susceptibility and inflammatory response to gonococcal infection, and they suggest that future studies can be designed to correlate genetic markers with the host response.