The liquid Kangfuxin (KFX) has efficient antifungal activity and can be used in the treatment of vulvovaginal candidiasis in mice

Vulvovaginal candidiasis (VVC) is an infectious disease caused mainly by Candida albicans. Kangfuxin (KFX) is a traditional Chinese medicine preparation made from Periplaneta americana extracts, which promotes wound healing and enhances body immunity and also acts as an antifungal agent. Here, we evaluated the effect of KFX in the treatment of VVC in vitro and in vivo. The minimum inhibitory concentration (MIC₅₀) of KFX against C. albicans ranged from 7·65 to 20·57%. In addition, KFX was more efficient than fluconazole (FLC) in inhibiting the drug-resistant C. albicans, and the effect was more intense after 8 h. The KFX treatment also exhibited good activity in vivo. It restored the body weight and reduced the vulvovaginal symptoms in mice induced with VVC. It downregulated the expression of the hyphae-related gene, HWP1, thus inhibiting the growth and development of C. albicans hyphae. It also increased the number of neutrophils and promoted the secretion of interleukin-17A (IL-17A); however, the levels of interleukin-8 (IL-8) and interleukin-1β (IL-1β) decreased in mice with VVC. We deduce that KFX effectively treats vaginal candidiasis in two ways: by inhibiting the growth and development of mycelia to reduce colonization of C. albicans and by promoting the secretion and release of IL-17A and neutrophils in high numbers to fight C. albicans infection. This study provides a theoretical basis for the use of KFX for the clinical treatment of VVC.     

The effects of recombinant interleukin-1 and recombinant tumor necrosis factor on non-specific resistance to infection

Natural and synthetic immunomodulators that increase non-specific resistance to infection induce the production of interleukin-1 (IL-1) and tumor necrosis factor (TNF). Therefore, we investigated the effect of IL-1 and of TNF on the survival of lethally-infected mice. Mice were injected with 1 x 10(6) Klebsiella pneumoniae in the thigh muscle. When recombinant human IL-1 beta was given as a single i.p. injection 24 h before the infection, survival was increased. Using 80 ng IL-1 beta per mouse, survival compared to control animals was 80% versus 20% 48 h after the infection (p less than 0.001). No effect of IL-1 was observed when it was given 1/2 h before or 6 h after the infection. IL-1 alpha proved to be at least as potent as IL-1 beta. Numbers of bacteria cultured from the blood, thigh muscle, liver, spleen, and kidney were similar in IL-1-treated and control animals. Protection against death by IL-1 was also investigated in granulocytopenic mice with a Pseudomonas aeruginosa infection. Administration of the cyclooxygenase-inhibitor, ibuprofen, did not affect the beneficial effect of IL-1. In this model human recombinant TNF was at least tenfold less active than IL-1 beta. Pretreatment with IL-1 also had a significant effect on survival of mice that received a high dose of bacterial lipopolysaccharide.     

The effects of recombinant interleukin-1 and recombinant tumor necrosis factor on non-specific resistance to infection

Natural and synthetic immunomodulators that increase non-specific resistance to infection induce the production of interleukin-1 (IL-1) and tumor necrosis factor (TNF). Therefore, we investigated the effect of IL-1 and of TNF on the survival of lethally-infected mice. Mice were injected with 1 × 10⁶ Klebsiella pneumoniae in the thigh muscle. When recombinant human IL-1 was given as a single i.p. injection 24 h before the infection, survival was increased. Using 80 ng IL-l per mouse, survival compared to control animals was 80% versus 20% 48 h after the infection (p < 0.001). No effect of IL-1 was observed when it was given &frac; h before or 6 h after the infection. IL-l proved to be at least as potent as IL-1.Numbers of bacteria cultured from the blood, thigh muscle, liver, spleen, and kidney were similar in IL-1-treated and control animals. Protection against death by IL-1 was also investigated in granulocytopenic mice with aPseudomonas aeruginosa infection. Administration of the cyclooxygenase-inhibitor, ibuprofen, did not affect the beneficial effect of IL-1. In this model human recombinant TNF was at least tenfold less active than IL-1.Pretreatment with IL-1 also had a significant effect on survival of mice that received a high dose of bacterial lipopolysaccharide.     

IL-10 in antilipopolysaccharide immunity against systemic Klebsiella infections

AIM: This study was undertaken in order to determine whether anti-inflammatory cytokine interleukin-10 is responsible for a previously described protection against Klebsiella infection mediated by antilipopolysaccharide antibodies. METHODS: BALB/c mice were infected intraperitoneally with a lethal challenge of Klebsiella pneumoniae Caroli. One group was protected with monoclonal antibodies prior to infection and the second was not. We measured plasma levels of interleukin-10 at different time points by enzyme immunoassay and analyzed the relation between interleukin-10 and proinflammatory cytokines interleukin-6 and tumor necrosis factor-alpha in order to determine the association of these ratios with the outcome of infection. MAJOR FINDINGS AND CONCLUSIONS: We found different pattern of interleukin-10 production in protected mice compared with unprotected ones. The difference is greatest 24 hours postinfection. The ratios between IL-10 and proinflammatory cytokines confirmed the suppressed proinflammatory response in protected animals, especially 24 hours postinfection. Hence the mortality in unprotected mice begins immediately after we conclude that such cytokine relation and IL-10 production are, at least partially, responsible for the destiny of infected animals and the outcome of infection.     

IL-1 Coordinates the Neutrophil Response to C. albicans in the Oral Mucosa

Mucosal infections with Candida albicans belong to the most frequent forms of fungal diseases. Host protection is conferred by cellular immunity; however, the induction of antifungal immunity is not well understood. Using a mouse model of oropharyngeal candidiasis (OPC) we show that interleukin-1 receptor (IL-1R) signaling is critical for fungal control at the onset of infection through its impact on neutrophils at two levels. We demonstrate that both the recruitment of circulating neutrophils to the site of infection and the mobilization of newly generated neutrophils from the bone marrow depended on IL-1R. Consistently, IL-1R-deficient mice displayed impaired chemokine production at the site of infection and defective secretion of granulocyte colony-stimulating factor (G-CSF) in the circulation in response to C. albicans. Strikingly, endothelial cells were identified as the primary cellular source of G-CSF during OPC, which responded to IL-1α that was released from keratinocytes in the infected tissue. The IL-1-dependent crosstalk between two different cellular subsets of the nonhematopoietic compartment was confirmed in vitro using a novel murine tongue-derived keratinocyte cell line and an established endothelial cell line. These data establish a new link between IL-1 and granulopoiesis in the context of fungal infection. Together, we identified two complementary mechanisms coordinating the neutrophil response in the oral mucosa, which is critical for preventing fungal growth and dissemination, and thus protects the host from disease.     

Thein vivo andin vitro effects of interleukin-1 and tumor necrosis factor on murine cytomegalovirus infection

The effects of tumor necrosis factor (TNF) and interleukin-1 (IL-1) on infection with murine cytomegalovirus (MCMV) were investigatedin vitro andin vivo. The addition of each of these cytokines (at 1 ng/ml) to tissue culture monolayers 24 hr prior to MCMV challenge produced a reproducible decrease in vital titer (from 1 × 10⁸ pfu to approximately 4 × 10⁶ pfu for both cytokines). There was no further increase in this effect when a 10 or 100 ng/ml of each of these cytokines was employed. Despite thesein vitro effects, the pretreatment of suckling, weanling, or adult mice with 80 or 400 ng of TNF or IL-1 alone, or 80 ng of each cytokine together, had no effect on the survival of mice following MCMV. Similarly, neither of these cytokines adversely influenced the protective effects of hyperimmune anti-MCMV antiserum; that is, they did not attenuate the protection conferred by the antiserum nor affect the protective effects of subtherapeutic doses of the antiserum. We conclude that despite promising antiviral effects against MCMVin vitro, these agents do not result in a useful therapeutic effectin vivo. Moreover, despite the ability of IL-1 to induce ACTH and corticosterone in mice, IL-1 treatment did not increase the mortality to CMV.     

Filaria-induced IL-10 suppresses murine cerebral malaria

Filarial nematodes achieve long survival in their hosts due to their capacity to modulate immune responses. Therefore, immunomodulation by filarial nematodes may alter responses to concomitant infections such as malaria. Cerebral malaria (CM), a severe complication of Plasmodium falciparum infections, is triggered as a consequence of the immune response developed against malaria parasites. The question arises whether prior infection with helminth parasites is beneficial against CM. In the present work a murine model for subsequent has been used to assess this hypothesis. C57BL/6 mice were infected with the rodent filarial parasite Litomosoides sigmodontis and the murine model parasite for CM, Plasmodium berghei ANKA. Previously filaria-infected C57BL/6 mice showed significantly reduced CM rates. CD8⁺ T cell recruitment to the brain, a hallmark for CM development, was reduced in protected mice. Furthermore, in contrast to P. berghei single-infected animals, filaria-infected mice had significantly higher levels of circulating IL-10. The requirement for IL-10 in CM protection was demonstrated by the lack of protection in IL-10 KO mice. This suggests that the anti-inflammatory IL-10 elicited by filarial nematodes is able to suppress the overwhelming inflammatory reaction otherwise triggered against malaria parasites in C57BL/6 mice, preventing full progress to CM.     

IL-33/ST2 contributes to severe symptoms in Plasmodium chabaudi-infected BALB/c mice

It has been reported that IL-33 contributes to potentiation of Th2 inflammatory diseases and protection against helminth infection. Increased plasma IL-33 levels have been observed in patients with severe falciparum malaria, however, the role of IL-33 in malaria remains unclear. Here we report that IL-33 enhances inflammatory responses in malaria infection. ST2-deficiency altered severity of inflammation in the liver and serum levels of pro-inflammatory cytokines such as TNF-α and IL-6, and IL-13 that is a Th2 cytokine during Plasmodium chabaudi infection. IL-13-deficient mice have similar phenotype with ST2-deficient mice during P. chabaudi infection. Furthermore, ST2- and IL-13-deficiency reduced mortality from P. chabaudi infection. These results indicate that IL-33/ST2 can induce production of proinflammatory cytokines, such as TNF-α and IL-6, through production of IL-13 in P. chabaudi-infected BALB/c mice, suggesting that IL-33/ST2 play a critical role in inflammatory responses to malaria infection. Thus, these findings may define a novel therapeutic target for patients with severe malaria.     

The pathological activity of metabolic products of candida albicans on newborn mice

Summary Concentrated filtrates ofCandida albicans have an inhibiting effect on the growth of newborn mice. Occasionally, this inhibition of growth is very distinct and the clinical picture corresponds to human progeria.Increased dosages of subcutaneous injections of filtrates did not increase the frequency of appearance of distinctly retarded growth in mice, but the mice showed a great area of baldness.In the number of mice, which did not show progeria, there were degenerative changes in liver and kidneys morphologically suggesting glycogenosis. The same type of changes were also observed when contents of disruptedC. albicans cells were injected into newborn mice. In the later case there was positive histochemical staining for the deposits of glycogen.The induction of pathological changes through fungus filtrates is a newly described biological phenomenon, which may play a greater role in pathology.     

Filarial infection induces protection against P. berghei liver stages in mice

Chronic helminth infections such as filariasis in human hosts can be life long, since parasites are equipped with a repertoire of immune evasion strategies. In many areas where helminths are prevalent, other infections such as malaria are co-endemic. It is still an ongoing debate, how one parasite alters immune responses against another. To dissect the relationships between two different parasites residing in the same host, we established a murine model of co-infection with the filarial nematode Litomosoides sigmodontis and the malaria parasite Plasmodium berghei (ANKA strain). We found that filarial infection of BALB/c mice leads to protection against a subsequent P. berghei sporozoite infection in one-third of co-infected mice, which did not develop blood-stage malaria. This finding did not correlate with adult worm loads, however it did correlate with the presence of microfilariae in blood. Interestingly, protection was abrogated in IL-10-deficient mice. Thus, murine filariasis, in particular when it is a patent infection, is able to modify the immunological balance to induce protection against an otherwise deadly Plasmodium infection and is therefore able to influence the course of malaria in favour of the host.     

Infection of Female BWF1 Lupus Mice with Malaria Parasite Attenuates B Cell Autoreactivity by Modulating the CXCL12/CXCR4 Axis and Its Downstream Signals PI3K/AKT,NFκB and ERK

Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease characterized by abnormal autoreactivity in B cells. Lymphocytes and their soluble mediators contribute to the disease pathogenesis. We recently demonstrated that infecting lupus mice with malaria confers protection against lupus nephritis by attenuating oxidative stress in both liver and kidney tissues. In the current study, we further investigated B cell autoreactivity in female BWF1 lupus mice after infection with either live or gamma-irradiated malaria, using ELISA, flow cytometry and Western blot analysis. The lupus mice exhibited a significant elevation in plasma levels of IL-4, IL-6, IL-7, IL-12, IL-17, IFN-α, IFN-γ, TGF-β, BAFF and APRIL and a marked elevation of IgG2a, IgG3 and ant-dsDNA autoantibodies compared with normal healthy mice. Infecting lupus mice with live but not gamma-irradiated malaria parasite partially and significantly restored the levels of the soluble mediators that contribute to the progression of lupus. Furthermore, the B cells of lupus mice exhibited an increased proliferative capacity; aberrant overexpression of the chemokine receptor CXCR4; and a marked elevation in responsiveness to their cognate ligand (CXCL12) via aberrant activation of the PI3K/AKT, NFκB and ERK signaling pathways. Interestingly, infecting lupus mice with live but not gamma-irradiated malaria parasite restored a normal proliferative capacity, surface expression of CXCR4 and B cell response to CXCL-12. Taken together, our data present interesting findings that clarify, for the first time, the molecular mechanisms of how infection of lupus mice with malaria parasite controls B cell autoreactivity and thus confers protection against lupus severity.     

RNase-sensitive and RNase-insensitive protective components isolated fromListeria monocytogenes

Crude ribosomes were isolated fromListeria monocytogenes serotype 4b and separated into two fractions by molecular sieve chromatography. Chemical analysis indicated that fraction I contained cell envelope components while fraction II contained the ribosomes. Both fractions protected mice againstListeria, but only in combination with the adjuvant dimethyldioctadecylammonium bromide (DDA). RNase-treatment; but not proteinase K-treatment destroyed the protective properties of fraction II, and RNA purified from fraction II also induced protection. Protection induced by fraction I was not affected by either RNase- or proteinase K-treatment. Both subcutaneous and intraperitoneal, but not intravenous administration of fraction I, fraction 11, or purified RNA induced significant protection against intraperitoneal infection, the intraperitoneal route of administration being the most effective. All preparations induced high levels of protection 3 to 7 days after administration, but protection was already decreased after 14 days. Protection induced with RNA appeared to be biphasic, because it also protected mice 1 day, but not 2 days after administration. Protection induced with both fraction I and RNA was at least in part nonspecific, because both preparations also protected mice againstL. monocytogenes serotype 3,Streptococcus pneumoniae andPseudomonas aeruginosa. Results are discussed in relation to previous work with analogous preparations fromP. aeruginosa.     

IL-27 Receptor Signalling Restricts the Formation of Pathogenic,Terminally Differentiated Th1 Cells during Malaria Infection by Repressing IL-12 Dependent Signals

The IL-27R, WSX-1, is required to limit IFN-γ production by effector CD4⁺ T cells in a number of different inflammatory conditions but the molecular basis of WSX-1-mediated regulation of Th1 responses in vivo during infection has not been investigated in detail. In this study we demonstrate that WSX-1 signalling suppresses the development of pathogenic, terminally differentiated (KLRG-1⁺) Th1 cells during malaria infection and establishes a restrictive threshold to constrain the emergent Th1 response. Importantly, we show that WSX-1 regulates cell-intrinsic responsiveness to IL-12 and IL-2, but the fate of the effector CD4⁺ T cell pool during malaria infection is controlled primarily through IL-12 dependent signals. Finally, we show that WSX-1 regulates Th1 cell terminal differentiation during malaria infection through IL-10 and Foxp3 independent mechanisms; the kinetics and magnitude of the Th1 response, and the degree of Th1 cell terminal differentiation, were comparable in WT, IL-10R1^−/− and IL-10^−/− mice and the numbers and phenotype of Foxp3⁺ cells were largely unaltered in WSX-1^−/− mice during infection. As expected, depletion of Foxp3⁺ cells did not enhance Th1 cell polarisation or terminal differentiation during malaria infection. Our results significantly expand our understanding of how IL-27 regulates Th1 responses in vivo during inflammatory conditions and establishes WSX-1 as a critical and non-redundant regulator of the emergent Th1 effector response during malaria infection.     

Cervical mucins carry α(1,2)fucosylated glycans that partly protect from experimental vaginal candidiasis

Cervical mucins are glycosylated proteins that form a protective cervical mucus. To understand the role of mucin glycans in Candida albicans infection, oligosaccharides from mouse cervical mucins were analyzed by liquid chromatography-mass spectrometry. Cervical mucins carry multiple α(1-2)fucosylated glycans, but α(1,2)fucosyltransferase Fut2-null mice are devoid of these epitopes. Epithelial cells in vaginal lavages from Fut2-null mice lacked Ulex europaeus agglutinin-1 (UEA-I) staining for α(1-2)fucosylated glycans. Hysterectomy to remove cervical mucus eliminated UEA-I and acid mucin staining in vaginal epithelial cells from wild type mice indicating the cervix as the source of UEA-I positive epithelial cells. To assess binding of α(1-2) fucosylated glycans on C. albicans infection, an in vitro adhesion assay was performed with vaginal epithelial cells from wild type and Fut2-null mice. Vaginal epithelial cells from Fut2-null mice were found to bind increased numbers of C. albicans compared to vaginal epithelial cells obtained from wild type mice. Hysterectomy lessened the difference between Fut2-null and wild type mice in binding of C. ablicans in vitro and susceptibility to experimental C. albicans vaginitis in vivo. We generated a recombinant fucosylated MUC1 glycanpolymer to test whether the relative protection of wild type mice compared to Fut2-null mice could be mimicked with exogenous mucin. While a small portion of the recombinant MUC1 epitopes displayed α(1-2)fucosylated glycans, the predominant epitopes were sialylated due to endogenous sialyltransferases in the cultured cells. Intravaginal instillation of recombinant MUC1 glycanpolymer partially reduced experimental yeast vaginitis suggesting that a large glycanpolymer, with different glycan epitopes, may affect fungal burden.     

Prophylactic Effect of Enterococcus faecalis FK-23 Preparation on Experimental Candidiasis in Mice

The prophylactic effects of heat-killed cells of Enterococcus faecalis FK-23 (FK-23 preparation) on experimental candidiasis were investigated in normal and leukopenic mice. In cyclophosphamide-induced leukopenic mice, oral or intraperitoneal administration of the FK-23 preparation at a daily dose of 1.25 or 5 mg/mouse for 3 consecutive days prior to Candida albicans infection significantly prolonged survival periods of the infected mice, and decreased viable counts of C. albicans recovered from their kidneys. In normal mice, the FK-23 preparation administered at dosages ranging from 0.63 to 10 mg/mouse/day for 3 consecutive days was ineffective, while in leukopenic mice, the FK-23 administered orally caused a facilitated recovery in the number of white blood cells including neutrophils. Furthermore, intraperitoneal administration of the FK-23 preparation into mice augmented the anti-Candida activity of immunocompromised peritoneal exudate cells obtained from the animals. These results suggested the potential usefulness of the FK-23 preparation as a prophylactic agent for the management of patients with opportunistic fungal infections.     

Transient Loss of Protection Afforded by a Live Attenuated Non-typhoidal Salmonella Vaccine in Mice Co-infected with Malaria

In immunocompetent individuals, non-typhoidal Salmonella serovars (NTS) are associated with gastroenteritis, however, there is currently an epidemic of NTS bloodstream infections in sub-Saharan Africa. Plasmodium falciparum malaria is an important risk factor for invasive NTS bloodstream in African children. Here we investigated whether a live, attenuated Salmonella vaccine could be protective in mice, in the setting of concurrent malaria. Surprisingly, mice acutely infected with the nonlethal malaria parasite Plasmodium yoelii 17XNL exhibited a profound loss of protective immunity to NTS, but vaccine-mediated protection was restored after resolution of malaria. Absence of protective immunity during acute malaria correlated with maintenance of antibodies to NTS, but a marked reduction in effector capability of Salmonella-specific CD4 and CD8 T cells. Further, increased expression of the inhibitory molecule PD1 was identified on memory CD4 T cells induced by vaccination. Blockade of IL-10 restored protection against S. Typhimurium, without restoring CD4 T cell effector function. Simultaneous blockade of CTLA-4, LAG3, and PDL1 restored IFN-γ production by vaccine-induced memory CD4 T cells but was not sufficient to restore protection. Together, these data demonstrate that malaria parasite infection induces a temporary loss of an established adaptive immune response via multiple mechanisms, and suggest that in the setting of acute malaria, protection against NTS mediated by live vaccines may be interrupted.     

Allicin enhances host pro-inflammatory immune responses and protects against acute murine malaria infection

ABSTRACT: BACKGROUND: During malaria infection, multiple pro-inflammatory mediators including IFN-gamma, TNF and nitric oxide (NO) play a crucial role in the protection against the parasites. Modulation of host immunity is an important strategy to improve the outcome of malaria infection. Allicin is the major biologically active component of garlic and shows anti-microbial activity. Allicin is also active against protozoan parasites including Plasmodium, which is thought to be mediated by inhibiting cysteine proteases. In this study, the immunomodulatory activities of allicin were assessed during acute malaria infection using a rodent malaria model Plasmodium yoelii 17XL. METHODS: To determine whether allicin modulates host immune responses against malaria infection, mice were treated with allicin after infection with P. yoelii 17XL. Mortality was checked daily and parasitaemia was determined every other day. Pro-inflammatory mediators and IL-4 were quantified by ELISA, while NO level was determined by the Griess method. The populations of dendritic cells (DCs), macrophages, CD4+ T and regulatory T cells (Treg) were assessed by FACS. RESULTS: Allicin reduced parasitaemia and prolonged survival of the host in a dose-dependent manner. This effect is at least partially due to improved host immune responses. Results showed that allicin treatment enhanced the production of pro-inflammatory mediators such as IFN-gamma, TNF, IL-12 and NO. The absolute numbers of CD4+ T cells, DCs and macrophages were significantly higher in allicin-treated mice. In addition, allicin promoted the maturation of CD11c+ DCs, whereas it did not cause major changes in IL-4 and the level of anti-inflammatory cytokine IL-10. CONCLUSIONS: Allicin could partially protect host against P. yoelii 17XL through enhancement of the host innate and adaptive immune responses.     

Efficacy of Rhodotorula glutinis and Spirulina platensis carotenoids in immunopotentiation of mice infected with Candida albicans SC5314 and Pseudomonas aeruginosa 35

Enhancement of the immune response leading to protection against bacterial and fungal infections was shown using different schedules of immunization with microbial pigments and a polysaccharide. The group of mice given carotenoids of Rhodotorula glutinis (preparation I) and polysaccharide of Spitulina platensis (IV) survived for 2 weeks after Pseudomonas aeruginosa infection. The groups of mice given carotenoids (I), polysaccharide (IV), I+IV and with the crude phycocyanin of S. platensis (III)+IV survived for 2 weeks after Candida albicans infection. All other groups recorded a maximum level of mortality reaching 2 mice per group either after immunization or post-infection. Adding the carotenoids, phycocyanin and polysaccharides to food as additives might therefore enhance the human immune response against microbial infections.