In vitro-in vivo correlation of voriconazole resistance due to G448S mutation (cyp51A gene) in Aspergillus fumigatus.

Invasive pulmonary aspergillosis continues to be associated with a high mortality despite timely and appropriate therapy. Although host immunity plays a major role in poor clinical response, antifungal drug resistance cannot be ignored. Our studies were aimed 1) to study the mechanism of drug resistance in voriconazole strains of Aspergillus fumigatus, 2) to establish a causal relationship between cyp51A mutation and voriconazole resistance (VRC-R), and 3) to determine whether VRC-R due to cyp51A mutation correlated with in vivo resistance. A point mutation (G448S) involving cyp51A gene in VRC-R isolate was associated with resistance to VRC but not to posaconazole (POS); POS had superior activity to VRC in reducing lung fungal burden and mortality in mice infected with a VRC-R mutant of A. fumigatus. Our study demonstrated that azole resistance is based on specific site of cyp51A mutation and that in vitro VRC-R correlates with in vivo resistance.

In-vitro activity of nikkomycin Z alone and in combination with polyenes, triazoles or echinocandins against Aspergillus fumigatus.

The in-vitro activity of nikkomycin Z was investigated in combination with polyenes, triazoles or echinocandins against 20 clinical isolates of Aspergillus fumigatus with the fractional inhibitory concentration index (FICI) method. The drug interactions were classified as synergic (FICI < or = 0.5), no interaction (FICI > 0.5, but FICI < or = 4) or antagonistic (FICI > 4). The fungicidal activity of nikkomycin Z alone and in combination with a representative echinocandin (caspofungin) or triazole (voriconazole) was also examined with time-kill experiments and fungal cell viability assays. Two-drug combinations of nikkomycin Z with amphotericin B (FICI 3.59 +/- 0.57), amphotericin B lipid complex (FICI 3.95 +/- 0.74), liposomal amphotericin B (FICI 3.62 +/- 0.98), itraconazole (FICI 2.0 +/- 0.0), voriconazole (FICI 1.07 +/- 0.37), posaconazole (FICI 2.20 +/- 0.44) or ravuconazole (FICI 1.76 +/- 0.44) showed no interactions, but the pairwise combination of nikkomycin Z with caspofungin (FICI 0.22 +/- 0.19) or micafungin (FICI 0.35 +/- 0.27) showed synergic activity against A. fumigatus. Time-kill studies and fungal cell viability assays showed that neither nikkomycin Z nor caspofungin alone possessed fungicidal activity against A. fumigatus, whereas a combination of these two drugs at concentrations > or = 2 mg/L (> or = 0.031 x the concentration of drug that produced no visible growth) killed germinated conidia within 24 h in a concentration-dependent manner. These data suggest that two-drug combinations of nikkomycin Z with echinocandins, but not with polyenes and triazoles, have a synergic effect against A. fumigatus.

Efficacy of voriconazole plus amphotericin B or micafungin in a guinea-pig model of invasive pulmonary aspergillosis.

The efficacy of voriconazole in combination with amphotericin B or micafungin was studied in a transiently neutropenic guinea-pig model of invasive pulmonary aspergillosis. Guinea-pigs treated with the antifungal drugs, alone or in two-drug combinations, had an improved survival rate and reduced fungal burden in the lungs compared to untreated control animals. The efficacy of monotherapy and combination therapy was similar; activity was neither enhanced nor reduced with the two-drug combinations. Further studies of efficacy, dosing and optimal regimens for antifungal combinations are warranted.

The effect of antibiotic therapy on intrauterine infection-induced preterm parturition in rabbits.

OBJECTIVE: The purpose of this study was to determine whether early antibiotic administration to pregnant rabbits with intrauterine infection could prevent preterm delivery and perinatal mortality. STUDY DESIGN: Under hysteroscopic guidance, pregnant rabbits at 70% gestation (21 days) were allocated to three groups: (1) control group, transcervical inoculation of 0.2 ml phosphate-buffered saline (n = 16); (2) infection group, transcervical inoculation of 0.2 ml of 10(5) colony-forming units (CFU) of Escherichia coli (n = 21); (3) infection and antibiotics group, transcervical inoculations of 0.2 ml of 10(5) CFU of E. coli and ampicillin-sulbactam 150 mg/kg every 8 h intramuscularly (n = 32). To examine the consequences of treatment delay, animals in the latter group were subdivided to receive antibiotics at different time intervals of 0, 6, 11 and 18 h after bacterial inoculation. The intervals from bacterial inoculation to delivery and litter survival were documented. Systemic (rectal) temperatures were recorded at 4 h intervals through the first 36 h and every 12 h until delivery. A p value of < 0.05 was considered significant. RESULTS: All rabbits inoculated with E. coli without antibiotic treatment delivered prematurely. The median inoculation-to-delivery interval was significantly shorter in the infected group than in the control group (median 32 h, range 14.9-76.5 h vs. median 219 h, range 173-246 h, respectively; p < 0.0001). Antibiotic administration within 12 h of inoculation, but not after 18 h, increased duration of pregnancy (by reducing the rate of preterm delivery) and neonatal survival (0% vs. 71%; p < 0.0001). The mean temperatures at delivery of animals whose treatments began at 6 and 11 h post-inoculation were significantly lower than those untreated with antibiotics or those treated at 18 h post-inoculation (p < 0.0001 for each comparison). CONCLUSIONS: Antibiotic administration can prolong pregnancy and reduce perinatal mortality if administered early (within 12 h of microbial inoculation) in a rabbit model of ascending intrauterine infection.

17-beta estradiol can reduce secondary ischemic damage and mortality of subarachnoid hemorrhage.

Subarachnoid hemorrhage (SAH) is a unique disorder commonly occurring when an aneurysm ruptures, leading to bleeding and clot formation, with a higher incidence in females. To evaluate the influence of 17-beta estradiol (E2) in the outcome of subarachnoid hemorrhage, SAH was induced by endovascular puncture of the intracranial segment of internal carotid artery in 15 intact females (INT), 19 ovariectomized females (OVX), and 13 ovariectomized female rats with E2 replacement (OVX + E2). Cerebral blood flow was recorded before and after SAH. All animals were decapitated immediately after death or 24 hours after SAH for clot area analysis. Brains were sliced and stained with 2,3,5-triphenyltetrazolium chloride (TTC) for secondary ischemic lesion analysis. The cortical cerebral blood flow (CBF), which was measured by a laser-Doppler flowmeter, decreased to 29.6%+/-17.7%, 22.8%+/-8.3%, and 43.5%+/-22.9% on the ipsilateral side (P = 0.01), and decreased to 63.4%+/-14.1%, 57.4%+/-11.0%, and 66.6%+/-17.9% on the contralateral side (P = 0.26) in INT, OVX, and OVX + E2, respectively. The subcortical CBF, which were measured by the H2 clearance method, were 7.77+/-12.03, 7.80+/-8.65, and 20.58+/-8.96 mL 100 g(-1) min(-1) on the ipsilateral side (P < 0.01), and 21.53+/-2.94, 25.13+/-3.01, and 25.30+/-3.23 mL 100 g(-1) min(-1) on the contralateral side in INT, OVX, and OVX + E2, respectively. The mortality was 53.3%, 68.4%, and 15.4% in INT, OVX, and OVX + E2, respectively (P = 0.01), whereas no significant difference in clot area was noted among the groups. The secondary ischemic lesion volume was 9.3%+/-8.4%, 24.3%+/-16.3%. and 7.0%+/-6.4% in INT, OVX, and OVX + E2, respectively (P < 0.01). This study demonstrated that E2 can reduce the mortality and secondary ischemic damage in a SAH model without affecting the clot volume.

Candida-specific systemic cell-mediated immune reactivities in human immunodeficiency virus-positive persons with mucosal candidiasis.

Oropharyngeal candidiasis (OPC), as opposed to vulvovaginal candidiasis (VVC), is a common opportunistic infection in human immunodeficiency virus (HIV)-positive persons that correlates with reduced CD4 T cell counts. Although cell-mediated immunity (CMI) by CD4 Th1-type cells is considered to be the predominant host defense against mucosal candidiasis, the immune factors associated with susceptibility to OPC in HIV-positive persons are not well understood. This study investigated Candida-specific systemic CMI in HIV-positive persons with OPC and/or VVC. Reductions in delayed skin test reactivity to Candida antigen were observed in HIV-positive persons with CD4 cell counts <200 cells/microL, irrespective of the presence of mucosal infection. Likewise, despite the correlate of OPC with reduced CD4 cell counts in HIV-positive persons, differences in Candida-specific peripheral blood mononuclear cell proliferation and Th1/Th2 cytokine production between HIV-positive and HIV-negative persons were not consistent in a manner to suggest that deficiencies in Candida-specific systemic CMI account solely for the susceptibility to OPC.

Aspergillus: rising frequency of clinical isolation and continued susceptibility to antifungal agents, 1994-1999.

We investigated the frequency of clinical isolation and the in vitro susceptibility to antifungal agents of Aspergillus species obtained from patients at the Detroit Medical Center from January 1994 to December 1999. During this period, 593 clinical isolates of Aspergillus species [406 A. fumigatus, 68%; 82 A. niger, 14%; 42 A. flavus, 7%; 63 Aspergillus spp., 11%] were recovered from hospitalized patients. From January 1996 to December 1999, approximately 2.5-4.5 fold yearly increase of the number of aspergillus isolates occurred compared to that of 1994. Conidial suspensions from clinical isolates were prepared and their in vitro susceptibility to amphotericin B and three azoles were determined. All four agents examined were extremely active. The minimum inhibitory concentrations (MIC(90)) (microg/mL) of amphotericin B, itraconazole, voriconazole and posaconazole for A. fumigatus (n = 406) were 0.5, 1.0, 0.5 and 0.25. Similar values were noted for non-A. fumigatus isolates. A year-to-year comparison of the MIC(90) of the four agents for A. fumigatus and non-A. fumigatus isolates over the 6-year study period showed no significant differences. Our study showed a steady increase in the frequency of clinical isolation of Aspergillus species; and the organism has remained susceptible to amphotericin B/triazoles without any change in susceptibility levels during the 6-year study period.

A comparative study of the in vitro susceptibilities of clinical and laboratory-selected resistant isolates of Aspergillus spp. to amphotericin B, itraconazole, voriconazole and posaconazole (SCH 56592).

We investigated the in vitro susceptibilities of clinical and laboratory-selected Aspergillus spp. to posaconazole, and compared the results with those obtained for amphotericin B, itraconazole and voriconazole. Conidial suspensions from clinical isolates (284 Aspergillus fumigatus, 66 Aspergillus niger, 31 Aspergillus flavus and 43 Aspergillus spp.) and laboratory-selected resistant A. fumigatus isolates (15 resistant to amphotericin B, 25 to itraconazole and 12 to voriconazole) were prepared and their susceptibilities to various antifungal agents determined using a previously described broth macrodilution technique. The geometric mean MICs (mg/L) of posaconazole for A. fumigatus (0.17 +/- 0.11) and non-A. fumigatus aspergilli (0.16 +/- 0.28) were significantly lower (P 0.05) than those for amphotericin B, itraconazole and voriconazole. Amphotericin B-resistant A. fumigatus isolates were as susceptible to posaconazole as the parental strain. Itraconazole- and voriconazole-resistant isolates showed low-level (two- to three-fold increase in MICs) cross-resistance to posaconazole. The minimum fungicidal concentrations (mg/L) of posaconazole for A. fumigatus (n = 58) and non-A. fumigatus aspergilli (n = 40) were 4. 45 +/- 2.70 (range 0.25-8) and 4.14 +/- 3.03 (range 0.5-8), respectively. Time-kill studies showed that the fungicidal activity of posaconazole against A. fumigatus is time- and concentration-dependent (for example, posaconazole 4 mg/L killed >99% of A. fumigatus conidia within 24 h). These results suggest that overall, posaconazole has better activity and a smaller range of MICs for Aspergillus spp., including those with reduced susceptibility to amphotericin B, itraconazole and voriconazole.

Efficacy of voriconazole against invasive pulmonary aspergillosis in a guinea-pig model.

We compared the efficacies of amphotericin B and voriconazole against invasive pulmonary aspergillosis in a guinea-pig model. A susceptible isolate of Aspergillus fumigatus was used to produce the infection. Voriconazole-treated animals had significantly better survival and decreased fungal burden in the lungs as compared with controls. Although no statistical difference was seen between the efficacies of voriconazole and amphotericin B, a trend favouring voriconazole was noted. Thus, voriconazole, with its cidal activity, may be an attractive alternative to potentially toxic amphotericin B in the treatment of invasive pulmonary aspergillosis.

Effects of reproductive hormones on experimental vaginal candidiasis.

Vulvovaginal candidiasis (VVC) is an opportunistic mucosal infection caused by Candida albicans that affects large numbers of otherwise healthy women of childbearing age. Acute episodes of VVC often occur during pregnancy and during the luteal phase of the menstrual cycle, when levels of progesterone and estrogen are elevated. Although estrogen-dependent experimental rodent models of C. albicans vaginal infection are used for many applications, the role of reproductive hormones and/or their limits in the acquisition of vaginal candidiasis remain unclear. This study examined the effects of estrogen and progesterone on several aspects of an experimental infection together with relative cell-mediated immune responses. Results showed that while decreasing estrogen concentrations eventually influenced infection-induced vaginal titers of C. albicans and rates of infection in inoculated animals, the experimental infection could not be achieved in mice treated with various concentrations of progesterone alone. Furthermore, progesterone had no effect on (i) the induction and persistence of the infection in the presence of estrogen, (ii) delayed-type hypersensitivity in primary-infected mice, or (iii) the partial protection from a secondary vaginal infection under pseudoestrus conditions. Other results with estrogen showed that a persistent infection could be established with a wide range of C. albicans inocula under supraphysiologic and near-physiologic (at estrus) concentrations of estrogen and that vaginal fungus titers or rates of infection were similar if pseudoestrus was initiated several days before or after inoculation. However, the pseudoestrus state had to be maintained for the infection to persist. Finally, estrogen was found to reduce the ability of vaginal epithelial cells to inhibit the growth of C. albicans. These results suggest that estrogen, but not progesterone, is an important factor in hormone-associated susceptibility to C. albicans vaginitis.

In vitro susceptibilities of Aspergillus species to voriconazole, itraconazole, and amphotericin B.

We studied the in vitro activity of voriconazole (VCZ) itraconazole (ITZ) and amphotericin B (AMB) against 216 clinical isolates of Aspergillus spp. (142 Aspergillus fumigatus and 74 nonfumigatus Aspergillus spp. isolates) using a broth macrodilution method. The MICs (microgram/mL) (mean, range) for A. fumigatus were: VCZ 0.88, 0.25-4; ITZ 0.54, 0.25-4; AMB 2.16, 0.5-8. MIC90s were: VCZ 2, ITZ 1, AMB 4. MICs for nonfumigatus Aspergillus spp. were: VCZ 1.57, 0.25-4; ITZ 1.74, 0.25-4; AMB 2.88, 0.5-8. MIC90s for this group were: VCZ 4, ITZ 4, AMB 4. We also studied the susceptibility to VCZ of 18 AMB-resistant (mean, MIC 6.0 micrograms/mL) and 28 ITZ-resistant (mean, MIC 13.28 micrograms/mL) A. fumigatus isolates selected in the laboratory. The mean MICs of VCZ were 0.59 microgram/mL for AMB-resistant and 1.32 micrograms/mL for ITZ-resistant isolates. Our study showed that VCZ and ITZ had comparable in vitro activity against the isolates studied, except against A. fumigatus, where the MIC of ITZ was lower. The azoles had better in vitro activity than AMB against A. fumigatus and non-fumigatus spp. The non-fumigatus Aspergillus spp. were less susceptible to all three antifungals evaluated. When tested against ITZ- or AMB-resistant A. fumigatus strains, VCZ retained good activity, showing only a modest rise in the MIC against ITZ-resistant strains.

Comparative study of susceptibilities of germinated and ungerminated conidia of Aspergillus fumigatus to various antifungal agents.

Conidia are used as inocula for the in vitro susceptibility testing of Aspergillus fumigatus. Since the MIC is defined on the basis of visible mycelial growth, conidia should germinate and produce sporelings (germinated conidia) for monitoring of the growth inhibition and fungicidal activity of a drug. If a compound is capable of inhibiting germination of conidia while affecting or not affecting the growth of the organism, the MIC obtained will be the concentration of the drug required for the inhibition of conidial germination but not necessarily that required for inhibition of the growth of the organism. We investigated the susceptibility of germinated and ungerminated conidia to amphotericin B, itraconazole, voriconazole, and SCH56592. The MICs of various antifungal agents for germinated conidia were almost identical to those obtained for ungerminated conidia. In addition, both the germinated and ungerminated conidia were killed with almost equal efficiency by all of the compounds tested when exposed to the drugs for 24 h. These results suggest that either germinated or ungerminated conidia could be used as inocula for in vitro susceptibility studies of A. fumigatus with identical results.

In-vitro and in-vivo susceptibility of Aspergillus fumigatus to a novel conjugated styryl ketone.

We investigated the in-vitro and in-vivo susceptibility of Aspergillus fumigatus to the novel conjugated styryl ketone NC1175 and the results were compared with those obtained for amphotericin B and itraconazole. All 20 clinical isolates of A. fumigatus examined were susceptible to NC1175 (MIC = 5.54 +/- 2.48 mg/L; range 2.92-11.68 mg/L), and the minimum lethal concentration (MLC) was only twice the MIC, suggesting that NC1175 is fungicidal. The mean MIC values of amphotericin B (1.22 +/- 0.58 mg/L; range 0.5-4 mg/L) and itraconazole (0.37 +/- 0.11 mg/L; range 0.125-0.5 mg/L) were approximately nine- and 22-fold, respectively, lower than that of NC1175. Both amphotericin B-resistant (n = 18) and itraconazole-resistant (n = 28) isolates of A. fumigatus were as susceptible to NC1175 as amphotericin B-, and itraconazole-susceptible isolates. Kill curve experiments revealed that NC1175 at 23.35 mg/L (approximately four times the MIC) killed > or = 99% of conidia within 24 h of exposure to the drug. The in-vivo susceptibility of A. fumigatus to NC1175 was investigated using a murine pulmonary aspergillosis model. Treatment of infected mice with amphotericin B or NC1175 did not result in significant improvement of the mean survival (amphotericin B, 7.05 +/- 0.07 days; NC1175, 6.65 +/- 1.25 days) of the animals compared with that of the placebo group (7.21 +/- 1.20 days). However, semiquantitative organ culture revealed that clearance of A. fumigatus occurred in 16.6%, 50% and 66.6% of the mice treated with placebo, NC1175 and amphotericin B, respectively (P value for the control and the treated groups <0.01). These results suggest that NC1175 has in-vivo and in-vitro activity against A. fumigatus and can be used as a prototypic molecule for further development as an antifungal agent.

Organism-dependent fungicidal activities of azoles.

We investigated the antifungal activities of itraconazole and voriconazole on Aspergillus species by time kill studies, and the results were compared with those obtained for Candida species. Exposure of Aspergillus fumigatus conidia to varying concentrations (1.25 to 10 microg/ml) of itraconazole and voriconazole resulted in cellular death; the cytocidal effect was time and concentration dependent. In contrast, no killing of Candida albicans occurred in the presence of itraconazole and voriconazole at concentrations as high as 10 microg/ml, although candidal growth was inhibited compared to the drug-free control. Amphotericin B (1.25 to 10 microg/ml), on the other hand, killed both A. fumigatus and C. albicans. Similar results were obtained for non-A. fumigatus aspergilli and non-C. albicans Candida species. These observations indicate that both itraconazole and voriconazole are cytocidal agents for Aspergillus species but not for Candida species, suggesting that azoles possess organism-dependent fungicidal activities.

Vaginal-associated immunity in women with recurrent vulvovaginal candidiasis: evidence for vaginal Th1-type responses following intravaginal challenge with Candida antigen.

Studies from women with recurrent vulvovaginal candidiasis (RVVC) and from an animal model of experimental vaginitis suggest that deficiencies in immune function should be examined at the local rather than systemic level. Evidence of vaginal cell-mediated immunity (CMI) was evaluated for the first time in cervicovaginal lavage (CVL) fluid from RVVC patients. Results showed that although constitutive Th1- and Th2-type cytokine expression was detectable in CVL fluid from normal women, and differences in cytokines were observed in RVVC patients, limitations in experimental design of such de novo analyses urged caution in interpretation. Alternatively, attempts were made to establish conditions in control subjects whereby vaginal immunity could be detected after intravaginal challenge with Candida antigen. Preliminary results showed that Th1-type cytokines (interleukin-2 and -12, interferon-gamma) and histamine were increased 16-18 h after intravaginal introduction of Candida skin test antigen. Intravaginal antigenic challenge represents a novel approach for studying Candida-specific vaginal CMI.

Efficacy of D0870 treatment of experimental Candida vaginitis.

In this study, oral administration of the triazole D0870 was compared to oral administration of fluconazole in the treatment of experimental vaginal candidiasis. With an estrogen-dependent murine model of Candida albicans vaginal infection, the effects of D0870 on several isolates, including fluconazole-susceptible and -resistant isolates, were tested. D0870, at doses of 0.5 and 2.5 mg/kg of body weight given once over the course of a 10-day infection, was effective in eradicating vaginitis caused by fluconazole-susceptible laboratory and clinical isolates, respectively. In contrast, a stricter treatment regimen (every 24 to 48 h) with 10 and 25 mg of fluconazole per kg was required to achieve similar reductions in vaginal fungal titers induced by the same isolates. Whereas fluconazole was consistently ineffective in infections induced by fluconazole-resistant isolates, as predicted by in vitro susceptibility tests, D0870 was effective, although a daily regimen of 25 mg/kg was required. Additional studies showed that despite the in vitro activity of D0870 against two clinical Candida glabrata isolates, neither D0870 nor fluconazole was effective at daily doses as high as 100 and 125 mg/kg, respectively. Taken together, although D0870 failed to show efficacy against experimental C. glabrata vaginitis, D0870 was superior to fluconazole in the treatment of experimental C. albicans vaginitis caused by isolates that were either susceptible or resistant to fluconazole.

Treatment with the interleukin-I receptor antagonist and soluble tumor necrosis factor receptor Fc fusion protein does not prevent endotoxin-induced preterm parturition in mice.

OBJECTIVE: To determine whether the administration of anticytokine agents, the interleukin-1 receptor antagonist (IL-1ra) and a soluble tumor necrosis factor receptor Fc fusion protein (sTNFR-Fc), prevents endotoxin-induced preterm delivery in mice. METHODS: C3H/HeN pregnant mice at 15 days of gestation (70% gestation) were randomized to receive phosphate-buffered saline (PBS) or lipopolysaccharide (LPS) (50 micrograms/mouse) intraperitoneally (i.p.). Randomly selected PBS- or LPS-treated mice were additionally treated intravenously (i.v.), i.p., or subcutaneously (s.c.) every 3 hours with IL-1ra (1-50 mg) or every 12 hours with sTNFR-Fc (200-400 micrograms) beginning 1 hour before LPS injection. Animals were observed for vaginal bleeding and preterm delivery. RESULTS: Mice treated i.p. with 50 micrograms LPS (n = 13) had a shorter injection-to-delivery interval than mice treated similarly with PBS (n = 19) (median 13.5 hours, range 10-105 versus median 86.8 hours, range 53-120, respectively; P < .001). Saline-treated mice given 10 mg IL-1ra every 3 hours i.p. (n = 3) or 200 micrograms sTNFR-Fc every 12 hours i.v. (n = 4) had similar injection-to-delivery intervals as PBS-treated control mice (median 70 hours, range 70-76 versus median 58 hours, range 50-120, respectively). Similarly, LPS-treated mice given PBS every 3 hours (n = 20) had injection-to-delivery intervals comparable to LPS-treated mice (n = 13) (median 15.5 hours, range 9.8-92 versus median 13.5 hours, range 10-105, respectively). Lipopolysaccharide-treated mice given i.p. injections of 1 (n = 4), 10 (n = 31), or 50 (n = 15) mg of IL-1ra every 3 hours did not have longer injection-to-delivery intervals compared with LPS-treated mice (n = 13) (medians 11.6, 15, 14.5 and 13.5 hours; ranges 10.8-12, 8-95, 11-92, and 10-105, respectively). Lipopolysaccharide-treated mice given i.v. injections of 200 (n = 4) or 400 (n = 9) micrograms sTNFR-Fc every 12 hours did not have longer injection-to-delivery intervals compared with LPS-treated mice (n = 8) (medians 23.3, 22.5, and 21.9 hours; ranges 14.8-33, 15-95.5, and 15.5-44, respectively). The median injection-to-delivery interval of LPS-treated mice given both IL-1ra (10 mg) every 3 hours i.p. and sTNFR-Fc (200 micrograms) every 12 hours i.v. (n = 5) was not different from that of LPS-treated mice (median 26 hours, range 24.5-72 versus median 13.5 hours, range 10-105, respectively; P > .05). CONCLUSION: The anticytokine agents IL-1ra and sTNFR-Fc did not prevent preterm delivery or prolong pregnancy in endotoxin-induced preterm labor in mice.

A murine model of Candida glabrata vaginitis.

Vaginal Candida glabrata infections have increased significantly in recent years and are particularly common in women with uncontrolled diabetes mellitus. Efforts to understand the pathogenesis and treatment of this infection have been hindered by the lack of experimental animal models. Before onset of hyperglycemia, nonobese diabetic (NOD) mice inoculated intravaginally with clinical C. glabrata isolates were shown to support high vaginal titers of C. glabrata for > 14 days with evidence for superficial invasion of vaginal epithelial tissue. In contrast, congenic diabetic-resistant mice and mice susceptible to Candida albicans infections were significantly less susceptible to vaginal infection by C. glabrata, suggesting a potential link between the susceptibility of NOD mice to diabetes and their susceptibility to vaginal C. glabrata infections. This animal model of C. glabrata vaginitis provides a means to study the genetics and pathogenesis of C. glabrata infections and to evaluate the efficacy of antimycotic agents against C. glabrata.

Effects of systemic cell-mediated immunity on vaginal candidiasis in mice resistant and susceptible to Candida albicans infections.

Studies to date with CBA/J mice suggest a limited role for systemic cell-mediated immunity (CMI) against vaginal Candida albicans infections. The results of the present study show that preinduced Candida-specific systemic CMI was equally nonprotective against C. albicans vaginal infections in mice with high (BALB/cJ), low (DBA/2), or intermediate (CBA/J) resistance to C. albicans infections. Similarly, the locally acquired partial protection against a second C. albicans vaginal infection was equally observed with BALB/cJ, DBA/2, and CBA/J mice. These results indicate that observations made previously with CBA/J mice were not murine strain specific and provide additional support for the hypothesis that systemic CMI does not represent a dominant host defense mechanism at the vaginal mucosa.

Interleukin-1 receptor antagonist (IL-1ra) production by human amnion, chorion, and decidua.

PROBLEM: This study was conducted to determine whether (1) conditioned media from unstimulated primary cultures of human amnion, chorion, or decidua contain detectable concentrations of IL-1ra in vitro, and (2) bacterial endotoxin (LPS), tumor necrosis factor-alpha (TNF-alpha), or IL-1-beta (IL-1 beta) stimulate amnion, chorion, or decidua to produce increased amounts of IL-1ra. METHOD: Placentae were obtained from women at term with intact membranes before the onset of labor. Amnion, chorion, and decidual cells were isolated by standard procedures and grown to confluence. Cells were then cultured in quadruplicate for 16 h in tissue culture medium supplemented with 10% fetal calf serum or, additionally, with various concentrations of Escherichia coli LPS, TNF-alpha, or IL-1 beta. Culture supernatants were collected, and concentrations of IL-1ra were quantitated by a sensitive and specific enzyme-linked immunosorbent assay for IL-1ra. RESULTS: Results showed that primary cultures of amnion and chorion from 4 of 9 and decidua from 10 of 12 placentae had detectable rates of production of IL-1ra (ranges: 0.08-6.5, 0.42-12.1, and 1.55-96.5 pg IL-1ra/microgram protein/16 h, respectively). In addition, LPS (10-1,000 ng/ml) and IL-1 beta (0.1-10 ng/ml), but not TNF-alpha (0.01-100 ng/ml), stimulated decidual cells to release/secrete increased amounts of IL-1ra compared with media alone (range: 2.5-400 pg IL-1ra/microgram protein/16 h, P < 0.0001). In contrast, neither LPS, TNF-alpha, or IL-1 beta could stimulate amnion or chorion to release/secrete IL-1ra. CONCLUSIONS: These results indicate (1) that amnion, chorion, and predominantly decidua, can release or secrete IL-1ra in vitro, and (2) that LPS and IL-1 beta can stimulate decidual cells to produce increased amounts of IL-1ra.