Vaginal epithelial cells have the capacity to respond to an encounter with C. albicans by production of pro-inflammatory cytokines that can recruit host defenses to the infected tissue . The influx of inflammatory cells results in local tissue damage that contributes to the morbidity of vaginal yeast infections. A review of clinical trials with lactobacilli to treat VVC currently reveals a lack of efficacy in curing C. albicans infections . Wira, et al., suggest that specific strains, like L. rhamnosus GR-1 and L. reuteri RC-14, may show some promise for treatment, but larger clinical studies are needed to know for sure.
Even if they do not cure VVC, the probiotic lactobacilli, L. rhamnosus GR-1 and L. reuteri RC-14, reduce the symptoms of vaginal yeast infections in conjunction with fluconazole treatment . This suggests that there could be a host response effect by these strains beyond their role in colonization resistance. The present study was conducted to determine if there is a mechanism by which pro-inflammatory VK2 cell responses to C. albicans are affected by these Lactobacillus spp. strains. Since estrogen plays a critical role in the pathogenesis of VVC, the role of estrogen in the VK2 cell response to C. albicans was also evaluated.
Vaginal epithelial cells may detect C. albicans surface molecules with Toll-like receptors TLR2 and TLR4 , and possibly TLR6 [9, 18], which signal through the NF-κB signal transduction pathway to initiate transcription of mRNA for pro-inflammatory cytokines. In the present study, probiotic bacteria suppressed expression of genes associated with the NF-κB signal transduction pathway and expression of TNFα and IL-8 mRNA in C. albicans-infected VK2 cells. Expression of TLR2 and TLR6 were induced in VK2 cells by contact with C. albicans, which has also been observed in normal human gingival epithelial cells infected with Candida famata. These receptors appear to be important for epithelial cell recognition of C. albicans. 17β-estradiol decreased TLR2 and TLR6 expression by VK2 cells, suggesting that the hormone might leave cells less able to respond to the presence of C. albicans through production of immune recruitment cytokines. The presence of the probiotic lactobacilli also decreased TLR2 and TLR6 mRNA.
Since Toll-like receptors activate proinflammatory cytokine production via the NF-κB signal transduction pathway [8, 9], it seemed pertinent to observe how expression of genes involved in this pathway and the cytokines they induce were affected by C. albicans, estrogen, and the probiotic lactobacilli. The increased VK2 cell expression of NF-κB-associated genes and cytokines ELK1, Iκκβ, IL-6, and TNFα by C. albicans challenge is consistent with induction of the TLR2/6-mediated transduction of signals through the NF-κB pathway to produce inflammatory signals, e.g., IL6 and TNFα.
Vaginal epithelial cells appear to have the capacity to produce a number of different cytokines that can recruit innate and adaptive immune responses. For example, antigens from C. albicans induce vaginal epithelial cell line A431 production of inflammatory cytokines: IL-1α, IL-1β, IL-6, IL-8, IL-10, GM-CSF, IFNγ, and TNFα . In our present study, the VK2 (E6/E7) cell line produced measurable amounts of IL-1α, IL-1β, IL-6, IL-8, and TNFα, but not IFNγ and IL-10 when challenged with C. albicans. IL-13 was also detected by immunoassay. Therefore, this cell line is capable of producing cytokines that can recruit innate immunity but not IFNγ and IL-10, which are mostly produced by lymphocytes. Experiments conducted with the same VK2 cell line and Lactobacillus spp. strains as the present study reported stimulation of IL-1α expression by C. albicans challenge, but not after subsequent challenge with the lactobacilli . The latter study reported an increase in IL-8 expression by VK2 cells when the lactobacilli were present, which is what our present study shows in the absence of 17β-estradiol. VK2 cells responded differently to 17β-estradiol when C. albicans was not present, as they had increased expression of IL-1β and reduced expression of IL-8, suggesting a complicated regulatory process during the simultaneous influence of C. albicans and estrogen. Experimental examples show the complexity of C. albicans surface molecules and estrogen effects on cytokine production. It is known that bacterial surface molecules induce IL-1β, IL-6, and IL-8 in vaginal secretions . C. albicans surface molecules induce Il-8 and TNFα mRNA by vaginal epithelial cells, which store translated cytokine proteins for secretion in a temporally dissociated manner . This may explain why our experiments did not show increased amounts of IL-1β and IL-8 in the supernatants from VK2 cells with mRNA induction. Estrogen differentially regulates expression of 3000 genes in the vaginal epithelium, including the repression of IL-8 . In polarized uterine epithelial cells, estradiol represses poly (I:C)-induced expression of IL-8 and reverses IL-1β induction of TNFα and IL-8 expression . Similar experiments have not been described for vaginal epithelial cells until the present study.
The probiotic lactobacilli suppressed expression of NF-κB pathway-associated genes Iκκα, and ELK1, which were induced by C. albicans infection of the VK2 cells. They also inhibited the additive effect of 17β-estradiol on C. albicans-induced TNFα. These results support the role of C. albicans acting through Toll-like receptors and the NF-κB signal transduction pathway in VK2 cells to activate expression of inflammatory cytokines, which is suppressed by the presence of probiotic lactobacilli. Although the probiotic lactobacilli appear to suppress proinflammatory responses, their presence is associated with increased VK2 cell expression of IL-1α and IL-1β, which can recruit cellular mediators of immunity. The pro-inflammatory cytokines IL-1α and IL-1β were expressed in greater amounts as a result of the probiotic bacteria and their expression was not abrogated by the NF-κB pathway inhibitor, curcumin. An alternative signal transduction response pathway to NF-κB, such as the MAPK pathway via the AP-1 transcription factor, could elicit an immunological cell recruitment response to C. albicans. It has been shown that IL-1β is involved in the stimulation of β-defensin expression by esophageal epithelial cells in Candida esophagitis via both the NF-κB and MAPK/AP-1 signal transduction pathways . The conclusion based on these results is that probiotic lactobacilli suppress the TLR2/6-initiated NF-κB-mediated induction of some pro-inflammatory cytokines, while stimulating another signal transduction pathway that induces IL-1α and IL-1β expression. The importance of IL-1β in stimulating an anti-Candida spp. response is exemplified by the apparent implementation of multiple signal transduction pathways for induction of its expression.