Mechanism of Vaginal Epithelial Cell Pyroptosis Induced by the NLRP3 Inflammasome in Vulvovaginal Candidiasis.

PROBLEM: Vulvovaginal candidiasis (VVC) is a common mucosal fungal infection, and Candida albicans is the main causative agent. The NLRP3 inflammasome plays an important role in VVC, but the underlying mechanism is unknown. METHOD OF STUDY: Vaginal epithelial cells were divided into three groups: control, C. albicans strain SC5314 (wild-type, WT), and WT+ Matt Cooper Compound 950 (MCC950, a specific NLRP3 inhibitor). After human vaginal epithelial cells were pretreated with 1 µmol/L MCC950 for 2 h, C. albicans (MOI = 1) was cocultured with the human vaginal epithelial cells for 12 h. The cell supernatants were collected, LDH was detected, and the IL-1ß and IL-18 levels were determined by ELISA. The expression of the pyroptosis-related proteins NLRP3, Caspase-1 p20 and GSDMD was measured by Western blotting analysis. The protein expression of the pyroptosis-related N-terminus of GSDMD (GSDMD-N) was detected by immunofluorescence. RESULTS: In this study, we showed that the WT C. albicans strain induced pyroptosis in vaginal epithelial cells, as indicated by the LDH and proinflammatory cytokine levels and the upregulated levels of the pyroptosis-related proteins NLRP3, Caspase-1 p20, and GSDMD-N. MCC950 reversed the changes in the expression of these proteins and proinflammatory cytokines in vaginal epithelial cells. CONCLUSION: C. albicans activated the NLRP3 inflammasome to induce vaginal epithelial cell pyroptosis. MCC950 inhibited the NLRP3 inflammasome, reduced vaginal epithelial cell pyroptosis, and decreased the release of inflammatory cytokines.

Candida pathogens induce protective mitochondria-associated type I interferon signalling and a damage-driven response in vaginal epithelial cells.

Vaginal candidiasis is an extremely common disease predominantly caused by four phylogenetically diverse species: Candida albicans; Candida glabrata; Candida parapsilosis; and Candida tropicalis. Using a time course infection model of vaginal epithelial cells and dual RNA sequencing, we show that these species exhibit distinct pathogenicity patterns, which are defined by highly species-specific transcriptional profiles during infection of vaginal epithelial cells. In contrast, host cells exhibit a homogeneous response to all species at the early stages of infection, which is characterized by sublethal mitochondrial signalling inducing a protective type I interferon response. At the later stages, the transcriptional response of the host diverges in a species-dependent manner. This divergence is primarily driven by the extent of epithelial damage elicited by species-specific mechanisms, such as secretion of the toxin candidalysin by C. albicans. Our results uncover a dynamic, biphasic response of vaginal epithelial cells to Candida species, which is characterized by protective mitochondria-associated type I interferon signalling and a species-specific damage-driven response.

Differential Expression of Local Immune Response Genes in the Vagina: Implication for the Diagnosis of Vaginal Infections.

Transcription profiles of genes of local immune response were determined in the vagina of women with bacterial vaginosis, aerobic vaginitis, and vulvovaginal candidosis for detection of the most specific immune markers for these vaginal infections. Laboratory diagnosis of the vaginal infections was performed microscopically; the inflammatory reaction in the vagina (leukorrhea) was defined as the presence of >10 white blood cells per field of view. Transcription profiles of IL1b, IL10, IL18, TNFα, TLR4, GATA3, and CD68 were determined using reverse-transcription quantitative real-time PCR. The strongest predictors of aerobic vaginitis were increased levels of IL1b and IL10 mRNA. Bacterial vaginosis was strongly associated with reduced levels of IL18 and GATA3 mRNA. Increased levels of IL1b and TLR4 transcripts showed significant discriminatory power for vulvovaginal candidosis and leukorrhea. The results of this study suggest differential expression of local immune response genes in the vagina of women with different vaginal infections. Detection of specific immune markers in the vagina using reverse-transcriptase PCR could supplement PCR detection of abnormal vaginal microflora for the diagnosis of vaginal infections.

The Interleukin (IL) 17R/IL-22R Signaling Axis Is Dispensable for Vulvovaginal Candidiasis Regardless of Estrogen Status.

Candida albicans, a ubiquitous commensal fungus that colonizes human mucosal tissues and skin, can become pathogenic, clinically manifesting most commonly as oropharyngeal candidiasis and vulvovaginal candidiasis (VVC). Studies in mice and humans convincingly show that T-helper 17 (Th17)/interleukin 17 (IL-17)-driven immunity is essential to control oral and dermal candidiasis. However, the role of the IL-17 pathway during VVC remains controversial, with conflicting reports from human data and mouse models. Like others, we observed induction of a strong IL-17-related gene signature in the vagina during estrogen-dependent murine VVC. As estrogen increases susceptibility to vaginal colonization and resulting immunopathology, we asked whether estrogen use in the standard VVC model masks a role for the Th17/IL-17 axis. We demonstrate that mice lacking IL-17RA, Act1, or interleukin 22 showed no evidence for altered VVC susceptibility or immunopathology, regardless of estrogen administration. Hence, these data support the emerging consensus that Th17/IL-17 axis signaling is dispensable for the immunopathogenesis of VVC.

Therapeutic effectiveness of type I interferon in vulvovaginal candidiasis.

Vulvovaginal candidiasis (VVC) affects approximately 75% of all women of during their reproductive years. Previously, we reported that recombinant human IFN α-2b (rhIFNα-2b) protects vaginal epithelial cells from candidal injury in vitro. In the current study, we examined the effects of rhIFNα-2b (1.25 mg/mL, 10% inhibition concentration) on fungal clearance, immunocompetent cytokine responses, non-B IgG production, and tissue repair in a rat model of VVC. Following rhIFNα-2b treatment, the negative pathogen conversion rate reached 50.0% (3/6). Although rhIFNα-2b exhibited a limited ability to decrease inflammation and injury progression (P > 0.05), the Flameng mitochondrial injury scores were significantly reduced (P < 0.001) compared with those of the Model rats. After rhIFNα-2b treatment, the levels of IFN-γ and epithelial-derived IgG (tested by RP215) in vaginal tissues were significantly increased with those in the Control and Model groups (both P < 0.001), while there were no significant differences in the levels of IL-4 and IL-17 (P > 0.05). This is the first study to address the efficacy of rhIFNα-2b in treating VVC in a rat model, providing a theoretical basis for development of this promising treatment for clinical use.

Impaired Th17 cell proliferation and decreased pro-inflammatory cytokine production in CXCR3/CXCR4 double-deficient mice of vulvovaginal candidiasis.

Vulvovaginal candidiasis (VVC) is a common observed infection, affecting approximately 75% of women of reproductive age. Drug resistance represents a troublesome stumbling block associated with VVC therapy. Thus the aim of the present study was to provide information regarding the selection of potential drug targets for VVC. CXCR3-, CXCR4-, or CXCR/CXCR4 double-deficient mouse models of VVC were subsequently established, with changes to the load of Candida Albicans evaluated accordingly. The biological behaviors of the vaginal epithelial cells were characterized in response to the CXCR3-, CXCR4-, or CXCR3/CXCR4 double-knockout in vivo. Our initial observations revealed that in mice with VVC, CXCR3-, CXCR4-, or CXCR3 - CXCR4 double-knockout resulted in a decreased load of C. Albicans as well as reduced levels and proportion of Th17 cells. Proinflammatory cytokine production was found to be inhibited by CXCR3-, CXCR4-, or CXCR3/CXCR4 double-knockout whereby the mRNA and protein expressions CXCR3, CXCR4, IL-17, IL-6, and TNF-α exhibited decreased levels. CXCR3-, CXCR4-, or CXCR3/CXCR4 double-knockout appeared to function as positive proliferation factors, while playing a negative role in the processes of apoptosis and the cell cycle of vaginal epithelial cells. Taken together, the key findings of the study suggested that CXCR3/CXCR4 double-knockout could act to hinder the progression of VVC, highlighting its promise as a novel therapeutic target in the treatment of VVC. CXCR3 and CXCR4 genes may regulate Th17/IL-17 immune inflammatory pathways to participate in antifungal immunity.

Inhibition of CXCR4 by MicroRNA-1192 Reduces the Activation of Th17 Cells and Expression of Inflammation Factors in a Mouse Model of Vulvovaginal Candidiasis.

BACKGROUND/AIMS: Vulvovaginal candidiasis (VVC) is a disease commonly occurring in sexually active women. The involvement of microRNAs in several kinds of infectious diseases has been highlighted in a number of researches. Therefore, we conducted the present study in order to investigate whether microRNA-1192 (miR-1192) would significantly target CXCR4 in Th17 cells as well as inflammatory factors in mouse models suffering from VVC. METHODS: Seventy-five mice were selected as test subjects for this study, of which twenty-five were used as the normal control, while the rest were treated with estradiol or oil-treated in order to establish VVC mouse models (each n = 25). Protein expressions of CXCR4, IL-6, IL-17, and IL-23 were all measured using both an immunohistochemistry and ELISA. The Th17 cell percentage in peripheral blood and the expression of RORγt in Th17 cells were detected using a flow cytometry. Mouse vaginal epithelial cells were isolated from normal mice, after which the mice were treated with estradiol to regulate their estrogen, followed by treatments involving the miR-1192 mimic, miR-1192 inhibitor, siRNA-CXCR4, and miR-1192 inhibitor + si-CXCR4. The cell cycle, apoptosis, and proliferation were all examined by using an additional flow cytometry as well as the employment of the MTT assay. The miR-1192, CXCR4, IL-6, IL-17, and IL-23 expressions in tissues and cells were both measured using both RT-qPCR and western blot assay techniques. RESULTS: The mice treated with either estradiol or oil had presented to us lowered levels in miR-1192 expression as well as higher levels in both Th17 cell percentage and expression of RORγt in Th17 cells, along with mRNA and protein expressions of CXCR4, IL-6, IL-17, and IL-23. In cell experiments, the mouse vaginal epithelial cells that had been treated with miR-1192 inhibitor had shown us a decreased cell proliferation rate and contrarily increased expressions of CXCR4, IL-6, IL-17, and IL-23 mRNA, protein, and cell apoptosis rate; these results were opposite to the ones found in the mice treated with miR-1192 mimic. CONCLUSION: Our results provided significant evidence that miR-1192 could directly development and progression of VVC by restraining the CXCR4 gene in the VVC mice.

Nystatin enhances the immune response against Candida albicans and protects the ultrastructure of the vaginal epithelium in a rat model of vulvovaginal candidiasis.

BACKGROUND: Vulvovaginal candidiasis (VVC) is a common infectious disease of the lower genital tract. Nystatin, a polyene fungicidal antibiotic, is used as a topical antifungal agent for VVC treatment. The aim of the current study was to investigate the possible immunomodulatory effects of nystatin on the vaginal mucosal immune response during Candida albicans infection and examine its role in protection of vaginal epithelial cell (VEC) ultrastructure. RESULTS: Following infection with C. albicans, IFN-γ and IL-17 levels in VECs were significantly elevated, while the presence of IgG was markedly decreased as compared to uninfected controls (P <  0.05). No significant differences in IL4 expression were observed. After treatment with nystatin, the level of IFN-γ, IL-17 and IgG was dramatically increased in comparison to the untreated group (P <  0.05). Transmission electron microscopy revealed that C. albicans invades the vaginal epithelium by both induced endocytosis and active penetration. Nystatin treatment protects the ultrastructure of the vaginal epithelium. Compared with the untreated C. albicans-infected group, Flameng scores which measure mitochondrial damage of VECs were markedly decreased (P <  0.001) and the number of adhesive and invasive C. albicans was significantly reduced (P <  0.01) after treatment with nystatin. CONCLUSIONS: Nystatin plays a protective role in the host defense against C. albicans by up-regulating the IFN-γ-related cellular response, the IL-17 signaling pathway and possibly through enhancing VEC-derived IgG-mediated immunity. Furthermore, nystatin notably improves the ultramorphology of the vaginal mucosa, partially through the protection of mitochondria ultrastructure in VECs and inhibition of adhesion and invasion by C. albicans. Together, these effects enhance the immune response of the vaginal mucosa against C. albicans and protect the ultrastructure of vaginal epithelium in VVC rats.

Estrogen Receptor-Alpha (ESR1) Governs the Lower Female Reproductive Tract Vulnerability to Candida albicans.

Estradiol-based therapies predispose women to vaginal infections. Moreover, it has long been known that neutrophils are absent from the vaginal lumen during the ovulatory phase (high estradiol). However, the mechanisms that regulate neutrophil influx to the vagina remain unknown. We investigated the neutrophil transepithelial migration (TEM) into the vaginal lumen. We revealed that estradiol reduces the CD44 and CD47 epithelial expression in the vaginal ectocervix and fornix, which retain neutrophils at the apical epithelium through the estradiol receptor-alpha. In contrast, luteal progesterone increases epithelial expression of CD44 and CD47 to promote neutrophil migration into the vaginal lumen and Candida albicans destruction. Distinctive to vaginal mucosa, neutrophil infiltration is contingent to sex hormones to prevent sperm from neutrophil attack; although it may compromise immunity during ovulation. Thus, sex hormones orchestrate tolerance and immunity in the vaginal lumen by regulating neutrophil TEM.

The Case for an Expanded Concept of Trained Immunity.

Trained immunity was originally proposed as a program of innate immunity memory by innate immunity cells of hematopoietic origin such as the monocytes/macrophages and the NK cells. Here I discuss some old and new data justifying this program and some specific, still unanswered, questions it raises regarding the model fungus Candida albicans and the chronic, inflammatory vulvovaginal disease it causes. Building upon this well-established program, the recent reports that epithelial cells of mammals can also acquire memory from previous stimulations, and the apparent intrinsic ability of many living cells from bacteria to mammals to learn from experience, I suggest an expansion of the concept of trained immunity to include all cells of different lineages with the potential of memorizing previous microbial encounters. This expansion would better fit the complexity of innate immunity and the role it plays in infectious and inflammatory diseases.

Candidalysin Drives Epithelial Signaling, Neutrophil Recruitment, and Immunopathology at the Vaginal Mucosa.

Unlike other forms of candidiasis, vulvovaginal candidiasis, caused primarily by the fungal pathogen Candida albicans, is a disease of immunocompetent and otherwise healthy women. Despite its prevalence, the fungal factors responsible for initiating symptomatic infection remain poorly understood. One of the hallmarks of vaginal candidiasis is the robust recruitment of neutrophils to the site of infection, which seemingly do not clear the fungus, but rather exacerbate disease symptomatology. Candidalysin, a newly discovered peptide toxin secreted by C. albicans hyphae during invasion, drives epithelial damage, immune activation, and phagocyte attraction. Therefore, we hypothesized that Candidalysin is crucial for vulvovaginal candidiasis immunopathology. Anti-Candida immune responses are anatomical-site specific, as effective gastrointestinal, oral, and vaginal immunities are uniquely compartmentalized. Thus, we aimed to identify the immunopathologic role of Candidalysin and downstream signaling events at the vaginal mucosa. Microarray analysis of C. albicans-infected human vaginal epithelium in vitro revealed signaling pathways involved in epithelial damage responses, barrier repair, and leukocyte activation. Moreover, treatment of A431 vaginal epithelial cells with Candidalysin induced dose-dependent proinflammatory cytokine responses (including interleukin 1α [IL-1α], IL-1ß, and IL-8), damage, and activation of c-Fos and mitogen-activated protein kinase (MAPK) signaling, consistent with fungal challenge. Mice intravaginally challenged with C. albicans strains deficient in Candidalysin exhibited no differences in colonization compared to isogenic controls. However, significant decreases in neutrophil recruitment, damage, and proinflammatory cytokine expression were observed with these strains. Our findings demonstrate that Candidalysin is a key hypha-associated virulence determinant responsible for the immunopathogenesis of C. albicans vaginitis.

[Vulvovaginal candidiasis: An old disease with new challenges]. / Candidiasis vulvovaginal: una antigua enfermedad con nuevos desafíos.

Vulvovaginal candidiasis is an old disease that, even in a modern world, continues to have a high incidence. Despite the therapeutic advances, treatments are not always effective, and our understanding of the pathogenesis of this fungal infection is still incomplete. A discussion is presented in this article on the most significant developments related to the fungal virulence factors, the role of the immunological mechanisms involved in the vaginal protection, and the genetic alterations that confer susceptibility to the recurrent form of this mycosis. Current treatments, the use of new agents with antifungal activity, as well as the development of strategies, such as vaccination, are approached in the context of the complex scenario that governs the interactions between Candida and its host.

[Study on the production of IgG derived from vaginal epithelial cells and the effect of anti-Candida albicans].

Objective: To investigate the function of IgG secreted by vaginal epithelial cells in natural resistance to vulvovaginal candidiasis. Methods: (1)Immunohistochemical method was used to determine the expression of IgG secreted by normal vaginal epithelial cells VK2/E6E7.(2)Samples were divided into three groups by different proportions of VK2/E6E7 cells to Candida albicans ,including Candida albicans: VK2/E6E7 cells were 1∶10, 1∶1[yeast+ cells(1∶10)group and yeast+ cells(1∶1)group]and VK2/E6E7 cells as blank control group. The growth status of 3 groups were observed under inverted microscope after 24 hours. ELISA method was used to detect the production of IgG in 3 groups after 0, 3, 6, 12, 24, 48 hours. Results: (1)Immunohistochemical method showed normal vaginal epithelial cells were polygonal with pale blue nucleus and cytoplasm were distributed by brown granules, which indicated that IgG were strongly positive. While negative control group just had light blue nuclei.(2)Inverted microscope observation represented that control group had a clear outline, strong refraction and large nuclei with cobblestone-like appearance. After yeast+cells(1∶10)group co-cultured for 24 hours, Candida albicans begin to sprout and transformed to hyphae. VK2/E6E7 cells and Candida albicans were close to each other with vacuoles and small black granules in the cytoplasm. The morphology of cells were complete. Yeast+ cells(1∶1)group showed obvious invasion effect of Candida albicans to VK2/E6E7 cells with vigorous growth of hyphae, the decreased number and incomplete morphology of cells. Moreover, the connection of cells were loose. ELISA assay showed that there were statistically significant difference of IgG secretions between the 3 groups after 0, 3, 6, 12, 24, 48 hours(P<0.05). After stimulation of Candida albicans, secretion of IgG was significantly lower than that in the control group. The statistical difference of IgG secretions between yeast+ cells(1∶10)group and yeast+ cells(1∶1)group after 3, 6, 12, 24 hours were dramatic(P<0.05). The peak of IgG production showed when the ratio of Candida albicans and VK2/E6E7 was 1∶1 after 24 hours co-cultured(1.61±0.05)µg/ml. Conclusions: Candida albicans has a significant invasion role on epithelial cells. With increasing concentrations of Candida albicans, the invasion effect will be enhenced. While, after the vaginal epithelial cells co-cultured with Candida albicans, the secretion of IgG decreased significantly.

Neutrophil Gelatinase-Associated Lipocalin Concentration in Vaginal Fluid: Relation to Bacterial Vaginosis and Vulvovaginal Candidiasis.

OBJECTIVE: Neutrophil gelatinase-associated lipocalin (NGAL) is a component of innate immunity that prevents iron uptake by microorganisms. We evaluated whether NGAL was present in vaginal fluid and whether concentrations were altered in women with bacterial vaginosis (BV) or vulvovaginal candidiasis (VVC). METHODS: Vaginal secretions from 52 women with VVC, 43 with BV, and 77 healthy controls were assayed by enzyme-linked immunosorbent assay for NGAL and for concentrations of L-lactic acid. RESULTS: The median concentration of NGAL in vaginal fluid was significantly higher in control women (561 pg/mL) than in women with BV (402 pg/mL; P = .0116) and lower in women with VVC (741 pg/mL; P = .0017). Median lactic acid levels were similar in controls (0.11 mmol/L) and women with VVC (0.13 mmol/L) and were lower in women with BV (0.02 mmol/L; P < .0001). The NGAL and lactic acid concentrations were highly correlated (P < .0001). CONCLUSION: A decrease in Lactobacilli and/or lactic acid plus the absence of leukocytes results in lower vaginal NGAL levels that might facilitate the growth of bacteria associated with BV.

Serum interleukin-6 levels in murine models of Candida albicans infection.

Two Balb/C mouse models of Candida infection were used to detect serum interleukin-6 (IL-6) responses. The first model used systemic infection by Candida albicans ATCC 10231 strain infected through the lateral tail vein of mice without any specific pretreatment. The median Candida burdens of the kidneys were 1.5 × 106 CFU/ml 24 h postinoculation (p.i.) and 1.2 × 107 CFU/ml 72 h p.i., while median serum IL-6 levels were 479.3 pg/ml and 934.5 pg/ml, respectively. The Candida burden showed significant correlation with serum IL-6 24 h p.i. (R2 = 0.6358; P = 0.0082) but not 72 h p.i.The second model was a mouse vaginitis model applying intravaginal inoculation of mice pretreated with subcutaneous estradiol-valerate (10 mg/ml) 3 days before infection. Candida cell count in vaginal lavage fluid was 2.8 × 106 CFU/ml 24 h p.i. and 1.4 × 108 CFU/ml 72 h p.i. Serum IL-6 response was detected in 4 of 15 mice 24 h p.i. and 9 of 15 mice 72 h p.i. Even the responders had low IL-6 serum levels (mean values 29.9 pg/ml and 60.1 pg/ml, respectively) not correlating with Candida cell count in vaginal lavage fluid.In conclusion, serum IL-6 had strong relationship with systemic C. albicans infection while the local C. albicans infection of the vagina led to partial, prolonged and limited serum IL-6 response.

Vaginal epithelial cell-derived S100 alarmins induced by Candida albicans via pattern recognition receptor interactions are sufficient but not necessary for the acute neutrophil response during experimental vaginal candidiasis.

Vulvovaginal candidiasis (VVC), caused by Candida albicans, affects women worldwide. Animal and clinical studies suggest that the immunopathogenic inflammatory condition of VVC is initiated by S100 alarmins in response to C. albicans, which stimulate polymorphonuclear neutrophil (PMN) migration to the vagina. The purpose of this study was to extend previous in vitro data and determine the requirement for the alarmin S100A8 in the PMN response and to evaluate pattern recognition receptors (PRRs) that initiate the response. For the former, PMN migration was evaluated in vitro or in vivo in the presence or absence of S100 alarmins initiated by several approaches. For the latter, vaginal epithelial cells were evaluated for PRR expression and C. albicans-induced S100A8 and S100A9 mRNAs, followed by evaluation of the PMN response in inoculated PRR-deficient mice. Results revealed that, consistent with previously reported in vitro data, eukaryote-derived S100A8, but not prokaryote-derived recombinant S100A8, induced significant PMN chemotaxis in vivo. Conversely, a lack of biologically active S100A8 alarmin, achieved by antibody neutralization or by using S100A9(-/-) mice, had no effect on the PMN response in vivo. In PRR analyses, whereas Toll-like receptor 4 (TLR4)- and SIGNR1-deficient vaginal epithelial cells showed a dramatic reduction in C. albicans-induced S100A8/S100A9 mRNAs in vitro, inoculated mice deficient in these PRRs showed PMN migration similar to that in wild-type controls. These results suggest that S100A8 alarmin is sufficient, but not necessary, to induce PMN migration during VVC and that the vaginal PMN response to C. albicans involves PRRs in addition to SIGNR1 and TLR4, or other induction pathways.

Female sex hormones regulate the Th17 immune response to sperm and Candida albicans.

STUDY QUESTION: What role do female sex hormones play in the antisperm immune response? SUMMARY ANSWER: We found that sperm induce a Th17 immune response and that estradiol down-regulates the antisperm Th17 response by dendritic cells. WHAT IS KNOWN ALREADY: Estradiol down-regulates the immune response to several pathogens and impairs the triggering of dendritic cell maturation by microbial products. STUDY DESIGN, SIZE, DURATION: Ex vivo and in vivo murine models of vaginal infection with sperm and Candida albicans were used to study the induction of Th17 and its hormonal regulation. PARTICIPANTS/MATERIALS, SETTING, METHODS: We analyzed the induction of Th17 cytokines and T cells in splenocytes obtained from BALB/c mice challenged with sperm and C. albicans. For the in vivo vaginal infection models, we used ovariectomized mice treated with vehicle, estradiol or progesterone, and we assessed the effect of these hormones on the immune response in the lymph nodes. MAIN RESULTS AND THE ROLE OF CHANCE: Th17 cytokines and T cells were induced by sperm antigens in both ex vivo and in vivo experiments. Estrus levels of estradiol down-regulated the Th17 response to sperm and C. albicans in vivo. LIMITATIONS, REASONS FOR CAUTION: This study was conducted using murine models; whether or not the results are applicable to humans is not known. WIDER IMPLICATIONS OF THE FINDINGS: Our results describe an adaptive mechanism that reconciles immunity and reproduction and further explains why unregulated Th17 could be linked to infertility and recurrent infections. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by research grants from the Instituto de Salud Carlos III (ISCIII) (PI10/00897) and Fundación Mutua Madrileña to M.R. M.R. holds a Miguel Servet contract from the ISCIII (CP08/00228). M.A.M.-F. was supported by (ISCIII) INTRASALUD PI09/02029. We have no conflicts of interest to declare. TRIAL REGISTRATION NUMBER: Not required.

Immune deviation in recurrent vulvovaginal candidiasis: correlation with iron deficiency anemia.

BACKGROUND: Iron Deficiency Anemia (IDA) has been controversially linked to IL-4 production in previous studies. A predominant Th1 response leads to resistance against recurrent vulvovaginal candidiasis (RVVC), whereas a Th2 response exacerbates the disease. OBJECTIVE: To investigate the possible effect of iron deficiency on the host's susceptibility to RVVC as a result of the Th1/Th2 cytokine polarization. METHODS: We conducted a case-control study of 92 women in 4 groups based on strict inclusion and exclusion criteria: RVVC+IDA+ group consisted of 23 women with RVVC and IDA; RVVC+ IDA- group consisted of 23 women with RVVC without IDA; RVVC-IDA+ group consisted of 23 women without RVVC and with IDA and RVVC- IDA- group consisted of 23 healthy women. The iron parameters and key cytokines (IFN-γ, IL-10, IL-12, IL-4) were measured in blood samples. RESULTS: Comparison of IL-4 production between RVVC+ IDA+ (12.2 ± 1.3 pg/ml) and RVVC+ IDA- (2.4 ± 4.0 pg/ml) groups (p=0.044), between RVVC- IDA+ (14.6 ± 1.7 pg/ml) and RVVC- IDA- (1.28 ± 3.6 pg/ml) groups (p=0.006), between RVVC- IDA+ (14.6 ± 1.7 pg/ml) and RVVC+ IDA-) 2.4 ± 4.0 pg/ml) groups (p=0.009) and also between RVVC+ IDA+ and RVVC- IDA- (1.28 ± 3.6 pg/ml) groups (p=0.03) showed significant differences. We found a significant positive correlation between IL-4 and total iron binding capacity (TIBC, p=0.046) and between serum IL-10 and Hb levels (p=0.041) in the RVVC+ IDA- group. There was also a significant negative correlation between serum IL-4 and levels of serum iron (SI, p=0.041) in the RVVC- IDA- group. CONCLUSION: It seems that IDA determines the balance between and the intensity of Th1 and Th2 arms of the immune response and leads to a deviation toward Th2 response which could contribute to recurrence of candidiasis.

The synthetic melanocortin (CKPV)2 exerts anti-fungal and anti-inflammatory effects against Candida albicans vaginitis via inducing macrophage M2 polarization.

In this study, we examined anti-fungal and anti-inflammatory effects of the synthetic melanocortin peptide (Ac-Cys-Lys-Pro-Val-NH2)2 or (CKPV)2 against Candida albicans vaginitis. Our in vitro results showed that (CKPV)2 dose-dependently inhibited Candida albicans colonies formation. In a rat Candida albicans vaginitis model, (CKPV)2 significantly inhibited vaginal Candida albicans survival and macrophages sub-epithelial mucosa infiltration. For mechanisms study, we observed that (CKPV)2 inhibited macrophages phagocytosis of Candida albicans. Meanwhile, (CKPV)2 administration inhibited macrophage pro-inflammatory cytokines (TNF-α, IL-1ß and IL-6) release, while increasing the arginase activity and anti-inflammatory cytokine IL-10 production, suggesting macrophages M1 to M2 polarization. Cyclic AMP (cAMP) production was also induced by (CKPV)2 administration in macrophages. These above effects on macrophages by (CKPV)2 were almost reversed by melanocortin receptor-1(MC1R) siRNA knockdown, indicating the requirement of MC1R in the process. Altogether, our results suggest that (CKPV)2 exerted anti-fungal and anti-inflammatory activities against Candida albicans vaginitis probably through inducing macrophages M1 to M2 polarization and MC1R activation.

Mouse strain-dependent differences in estrogen sensitivity during vaginal candidiasis.

The animal models available for studying the immune response to genital tract infection require induction of a pseudo estrous state, usually achieved by administration of 17-ß-estradiol. In our experimental model of vaginal candidiasis, under pseudo estrus, different strains of mice were used. We observed major differences in the clearance of Candida albicans infection among the different strains, ascribable to differing susceptibility to estradiol treatment. In the early phase of infection CD1, BALB/c, C57BL/6 albino and C57BL/6 mice were colonized to similar levels, while in the late phase of infection, BALB/c mice, which are considered genetically resistant to C. albicans infection, exhibited greater susceptibility to vaginal candidiasis than CD1 and C57BL/6 albino strains of mice. This was because estradiol induced "per se" enlarged and fluid-filled uteri, more pronounced in infected mice and consistently more evident in BALB/c and C57BL/6 mice than in CD1 mice. Unlike CD1, BALB/c and C57BL/6 mice showed a heavy fungal colonization of the uterus, even though C57BL/6 mice apparently cleared C. albicans from the vagina. The presence of C. albicans in the vagina and uterus was accompanied by a heavy bacterial load. Collectively these observations prompted us to carry out a careful analysis of estradiol effects in a mouse model of vaginal infection.