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Toni Darville

Publications and source records attributed to Toni Darville.

17 recordsLinked to original sources
PubMed2006

Recruitment of BAD by the Chlamydia trachomatis vacuole correlates with host-cell survival.

Chlamydiae replicate intracellularly in a vacuole called an inclusion. Chlamydial-infected host cells are protected from mitochondrion-dependent apoptosis, partly due to degradation of BH3-only proteins. The host-cell adapter protein 14-3-3beta can interact with host-cell apoptotic signaling pathways in a phosphorylation-dependent manner. In Chlamydia trachomatis-infected cells, 14-3-3beta co-localizes to the inclusion via direct interaction with a C. trachomatis-encoded inclusion membrane protein. We therefore explored the possibility that the phosphatidylinositol-3 kinase (PI3K) pathway may contribute to resistance of infected cells to apoptosis. We found that inhibition of PI3K renders C. trachomatis-infected cells sensitive to staurosporine-induced apoptosis, which is accompanied by mitochondrial cytochrome c release. 14-3-3beta does not associate with the Chlamydia pneumoniae inclusion, and inhibition of PI3K does not affect protection against apoptosis of C. pneumoniae-infected cells. In C. trachomatis-infected cells, the PI3K pathway activates AKT/protein kinase B, which leads to maintenance of the pro-apoptotic protein BAD in a phosphorylated state. Phosphorylated BAD is sequestered via 14-3-3beta to the inclusion, but it is released when PI3K is inhibited. Depletion of AKT through short-interfering RNA reverses the resistance to apoptosis of C. trachomatis-infected cells. BAD phosphorylation is not maintained and it is not recruited to the inclusion of Chlamydia muridarum, which protects poorly against apoptosis. Thus, sequestration of BAD away from mitochondria provides C. trachomatis with a mechanism to protect the host cell from apoptosis via the interaction of a C. trachomatis-encoded inclusion protein with a host-cell phosphoserine-binding protein.

Apoptosis↗
PubMed2006

Stimulation of the cytosolic receptor for peptidoglycan, Nod1, by infection with Chlamydia trachomatis or Chlamydia muridarum.

Infection of epithelial cells by the intracellular pathogen, Chlamydia trachomatis, leads to activation of NF-kappaB and secretion of pro-inflammatory cytokines. We find that overexpression of a dominant-negative Nod1 or depletion of Nod1 by RNA interference inhibits partially the activation of NF-kappaB during chlamydial infection in vitro, suggesting that Nod1 can detect the presence of Chlamydia. In parallel, there is a larger increase in the expression of pro-inflammatory genes following Chlamydia infection when primary fibroblasts are isolated from wild-type mice than from Nod1-deficient mice. The Chlamydia genome encodes all the putative enzymes required for proteoglycan synthesis, but proteoglycan from Chlamydia has never been detected biochemically. Since Nod1 is a ubiquitous cytosolic receptor for peptidoglycan from Gram-negative bacteria, our results suggest that C. trachomatis and C. muridarum do in fact produce at least the rudimentary proteoglycan motif recognized by Nod1. Nonetheless, Nod1 deficiency has no effect on the efficiency of infection, the intensity of cytokine secretion, or pathology in vaginally infected mice, compared with wild-type controls. Similarly, Rip2, a downstream mediator of Nod1, Toll-like receptor (TLR)-2, and TLR4, increases only slightly the intensity of chlamydial infection in vivo and has a very mild effect on the immune response and pathology. Thus, Chlamydia may not produce sufficient peptidoglycan to stimulate Nod1-dependent pathways efficiently in infected animals, or other receptors of the innate immune system may compensate for the absence of Nod1 during Chlamydia infection in vivo.

Adaptor Proteins, Signal Transducing↗
PubMed2005

Chlamydia trachomatis induces expression of IFN-gamma-inducible protein 10 and IFN-beta independent of TLR2 and TLR4, but largely dependent on MyD88.

IFN-gamma-inducible protein 10 (IP-10) is a chemokine important in the attraction of T cells, which are essential for resolution of chlamydial genital tract infection. During infections with Gram-negative bacteria, the IP-10 response mediated through type I IFNs usually occurs as a result of TLR4 stimulation by bacterial LPS. However, we found that levels of IP-10 in genital tract secretions of Chlamydia trachomatis-infected female wild-type mice were similar to those of infected TLR2- and TLR4-deficient mice but significantly greater than those of infected MyD88-deficient mice. We investigated the mechanism of IP-10 and IFN-beta induction during chlamydial infection using mouse macrophages and fibroblasts infected ex vivo. The induction of IP-10 and IFN-beta was unchanged in Chlamydia-infected TLR2- and TLR4-deficient cells compared with wild-type cells. However, infection of MyD88-deficient cells resulted in significantly decreased responses. These results suggest a role for MyD88-dependent pathways in induction of IP-10 and IFN-beta during chlamydial infection. Furthermore, treatment of infected macrophages with an endosomal maturation inhibitor significantly reduced chlamydial-induced IFN-beta. Because endosomal maturation is required for MyD88-dependent intracellular pathogen recognition receptors to function, our data suggest a role for the intracellular pathogen recognition receptor(s) in induction of IFN-beta and IP-10 during chlamydial infection. Furthermore, the intracellular pathways that lead to chlamydial-induced IFN-beta function through TANK-binding kinase mediated phosphorylation and nuclear translocation of IFN regulatory factor-3.

Adaptor Proteins, Signal Transducing↗
PubMed2005

Can chlamydia be stopped?

Chlamydia is a rampant sexually transmitted disease, the world's leading cause of preventable blindness and a possible contributor to heart disease. Recent discoveries are suggesting new ways to curtail its spread

Anti-Bacterial Agents↗
PubMed2005

Chlamydia trachomatis infections in neonates and young children.

In 1911, Lindner and colleagues identified intracytoplasmic inclusions in infants with a nongonococcal form of ophthalmia neonatorum called inclusion conjunctivitis of the newborn (ICN). Mothers of affected infants were found to have inclusions in their cervical epithelial cells, fathers of such infants had inclusions in their urethral cells, and the epidemiology of sexually transmitted chlamydial infections was revealed. Fifty years later, chlamydial isolation procedures were developed, and studies again demonstrated Chlamydia trachomatis as an etiology of ICN and the female birth canal as the reservoir. In the late 1970s, a report by Beem and Saxon described respiratory tract colonization and a distinct pneumonia syndrome in infected infants. Genital chlamydial infection is recognized as the world's most common sexually transmitted disease, with estimates of greater than 4 million new infections occurring annually in the United States. Although most C. trachomatis infections in men and women are asymptomatic, infection can lead to severe reproductive complications in women. The high prevalence in women of child-bearing age results in exposure of an estimated 100,000 neonates in the United States annually. Many of these infants develop conjunctivitis, pneumonia, or both in the first few months of life. Clinical features, diagnosis, treatment, and approaches to prevention of conjunctivitis and pneumonia in the newborn and young infant are reviewed here. Appropriate testing for chlamydial infection in a pediatric victim of sexual assault and the implications of identifying C. trachomatis in suspected cases of childhood sexual abuse also are reviewed.

Adult↗
PubMed2005

Relationship between intracranial granulomas and cerebrospinal fluid levels of gamma interferon and interleukin-10 in patients with tuberculous meningitis.

Cerebrospinal fluid gamma interferon (IFN-gamma) and interleukin-10 levels in 39 patients with tuberculous meningitis were serially measured. Cytokine levels did not predict intracranial granuloma (IG) development, but IFN-gamma levels in the top quartile after 1 month of therapy were highly associated (odds ratio = 18) with detection of an IG by computed tomography scanning.

Adolescent↗
PubMed2004

Protection against Chlamydia trachomatis infection in vitro and modulation of inflammatory response in vivo by membrane-bound glycosaminoglycans.

Glycosaminoglycans (GAG) efficiently inhibit adherence of several strains of Chlamydia trachomatis to cell lines in vitro, but none of the GAG have been able to inhibit infections in vivo. One possible cause for failure of GAG inhibition in vivo is the inability to deliver a sustained concentration of GAG at the mucosal surface. We tested the possibility of enhancing cell protection by increasing the cell-surface concentration of GAG using membrane-anchored GAG (MAG), composed of phosphatidylethanolamine (PE)-linked GAG. These lipid conjugates were originally designed as extracellular phospholipase A2 (PLA2) inhibitors and exhibit a dual effect: the lipid moiety incorporates into the cell membrane, interfering with the action of PLA2 on cell membranes, and the anchored GAG protects the cell membrane from exogenous inflammatory mediators. We tested the ability of MAG to block chlamydia infection in vitro and in vivo. The MAG blocked infection of epithelial cells in vitro when added to the cells at the same time or before infection, but not if added after the bacteria had already invaded the host cells. One of the MAG led to the production of aberrant Chlamydia vacuoles, suggesting it may inhibit intracellular PLA2 associated with development of the vacuole. Although the MAG did not inhibit vaginal infection of mice, they decreased significantly the level of secretion of the inflammatory cytokines TNF-alpha and IFN-gamma but had no effect on secretion of the neutrophil chemokine, macrophage inflammatory protein-2 (MIP-2). Acute and chronic inflammatory cell infiltrates were not altered by MAG treatment. These findings suggest that lipid conjugation of GAG could be used as a novel approach for increasing cell-surface concentrations of GAG. The inconclusive in vivo results might be due to the physical properties of the tested MAG or an insufficient application protocol, and their improvement might provide the desired inhibitory effects.

Animals↗
PubMed2004

Cell death, BAX activation, and HMGB1 release during infection with Chlamydia.

Infection by a number of Chlamydia species leads to resistance of the host cell to apoptosis, followed by induction of host-cell death. In a population of infected cells that displays protection against staurosporine-induced apoptosis among the adherent cells, we find that cells that had been recovered from the supernatant share characteristics of both apoptosis and necrosis, as assayed by the propidium iodide (PI)-annexin V double-labeling technique. Cell death was observed in both an epithelial cell line and primary fibroblasts, although the primary cells had a higher propensity to die through apoptosis than the immortalized cell line. Staurosporine-mediated activation of the pro-apoptotic BCL-2 family member, BAX, was inhibited in the epithelial cell line infected for 32 h with the lymphogranuloma venereum (LGV/L2) but not the murine pneumonitis (MoPn) strain of C. trachomatis, but inhibition of staurosporine-mediated BAX activation disappeared after 48 h of infection with the LGV/L2 strain. Conversely, infection with MoPn (C. muridarum) but not LGV/L2 led to BAX activation after 72 h, as previously reported for shorter (48 h) infection with the guinea pig inclusion conjunctivitis (GPIC) serovar of C. psittaci (C. caviae). These results suggest that the ability to inhibit staurosporine-mediated BAX activation or to activate BAX due to the infection itself may vary as a function of the chlamydial strain. Interestingly, both the epithelial cells and the fibroblasts also released high mobility group box 1 protein (HMGB1) during infection, although much less HMGB1 was released from fibroblasts, consistent with the higher level of apoptosis observed in the primary cells. HMGB1 is released preferentially by necrotic or permeabilized viable cells, but not apoptotic cells. In the extracellular space, HMGB1 promotes inflammation through interaction with specific cell-surface receptors. Higher levels of HMGB1 were also measured in the genital-tract secretions of mice infected vaginally with C. trachomatis, compared to uninfected controls. These results suggest that cells infected with Chlamydia release intracellular factors that may contribute to the inflammatory response observed in vivo.

Animals↗
PubMed2004

Nonadherence with pediatric human immunodeficiency virus therapy as medical neglect.

OBJECTIVE: To examine the results of an interventionist approach applied to human immunodeficiency virus (HIV)-infected children for whom caregiver nonadherence was suspected as the cause of treatment failure. METHODS: The medical records of a cohort of 16 perinatally HIV-infected children whose care was managed at the Arkansas Children's Hospital Pediatric HIV Clinic for an uninterrupted period of >or=3 years were reviewed through July 2003. Data collected included date of birth, dates of and explanations for clinic visits and hospitalizations, dates of laboratory evaluations, CD4(+) T cell percentages, plasma HIV-1 RNA levels, antiretroviral medications, viral resistance tests (eg, phenotype and genotype), and physician-initiated interventions to enhance adherence to the medication regimen. A stepwise interventionist approach was undertaken when patients continued to demonstrate high viral loads, despite documented viral sensitivity to the medication regimen and caregivers' insistence that medications were being administered regularly. Step 1 was prescribing a home health nurse referral, step 2 was administering directly observed therapy (DOT) while the patient was hospitalized for 4 days, and step 3 was submitting a physician-initiated medical neglect report to the Arkansas Department of Human Services. RESULTS: The results for 6 patients for whom this stepwise approach was initiated are reported. Home health nurse referrals failed to result in sustained improvements in adherence in all 6 cases. Viral load assays performed before and after DOT provided an objective measure of the effect of adherence, with 12 hospitalizations resulting in a mean +/- SD decrease in HIV RNA levels of 1.09 +/- 0.5 log(10) copies per mL, with a range of 0.6 to 2.1 log(10) copies per mL. Four families responded to DOT hospitalization, and sustained decreases in the respective patients' viral loads were noted. In 2 cases, medical neglect reports were submitted when DOT did not result in improved adherence. These patients were eventually placed in foster care, with subsequent improvements in their viral loads and CD4(+) T cell percentages. CONCLUSIONS: Nonadherence with antiretroviral therapy can be established on the basis of persistently elevated HIV RNA levels that decrease with DOT. Nonadherence poses a danger to the child that is grave and potentially irreversible. Caregivers should be offered all available resources to help them adhere to a sound treatment plan. In cases of demonstrated inability to provide needed care, it is necessary to consider seeking child protection, even for apparently healthy children.

Antiretroviral Therapy, Highly Active↗
PubMed2003

Toll-like receptor-2, but not Toll-like receptor-4, is essential for development of oviduct pathology in chlamydial genital tract infection.

The roles of Toll-like receptor (TLR) 2 and TLR4 in the host inflammatory response to infection caused by Chlamydia trachomatis have not been elucidated. We examined production of TNF-alpha and IL-6 in wild-type TLR2 knockout (KO), and TLR4 KO murine peritoneal macrophages infected with the mouse pneumonitis strain of C. trachomatis. Furthermore, we compared the outcomes of genital tract infection in control, TLR2 KO, and TLR4 KO mice. Macrophages lacking TLR2 produced significantly less TNF-alpha and IL6 in response to active infection. In contrast, macrophages from TLR4 KO mice consistently produced higher TNF-alpha and IL-6 responses than those from normal mice on in vitro infection. Infected TLR2-deficient fibroblasts had less mRNA for IL-1, IL-6, and macrophage-inflammatory protein-2, but TLR4-deficient cells had increased mRNA levels for these cytokines compared with controls, suggesting that ligation of TLR4 by whole chlamydiae may down-modulate signaling by other TLRs. In TLR2 KO mice, although the course of genital tract infection was not different from that of controls, significantly lower levels of TNF-alpha and macrophage-inflammatory protein-2 were detected in genital tract secretions during the first week of infection, and there was a significant reduction in oviduct and mesosalpinx pathology at late time points. TLR4 KO mice responded to in vivo infection similarly to wild-type controls and developed similar pathology. TLR2 is an important mediator in the innate immune response to C. trachomatis infection and appears to play a role in both early production of inflammatory mediators and development of chronic inflammatory pathology.

Animals↗
PubMed2003

Inhibition of chlamydial infectious activity due to P2X7R-dependent phospholipase D activation.

Chlamydia trachomatis survives within host cells by inhibiting fusion between Chlamydia vacuoles and lysosomes. We show here that treatment of infected macrophages with ATP leads to killing of chlamydiae through ligation of the purinergic receptor, P2X(7)R. Chlamydial killing required phospholipase D (PLD) activation, as PLD inhibition led to rescue of chlamydiae in ATP-treated macrophages. However, there was no PLD activation nor chlamydial killing in ATP-treated P2X(7)R-deficient macrophages. P2X(7)R ligation exerts its effects by promoting fusion between Chlamydia vacuoles and lysosomes. P2X(7)R stimulation also resulted in macrophage death, but fusion with lysosomes preceded macrophage death and PLD inhibition did not prevent macrophage death. These results suggest that P2X(7)R ligation leads to PLD activation, which is directly responsible for inhibition of infection.

Adenosine Triphosphate↗
PubMed2003

Infective endocarditis in Arkansan children from 1990 through 2002.

BACKGROUND: Recent medical and surgical advances have increased the potential risk of endocarditis. Epidemiology, pathogens, morbidity and/or mortality may have evolved in the past decade in pediatric patients diagnosed with endocarditis. METHODS: Retrospective reviews of medical records from Arkansas Children's Hospital were done to identify patients with infective endocarditis from January 1990 through December 2002. Basic demographic and clinical data were gathered and reviewed. RESULTS: Of 76 patients identified, 57 were included based on diagnostic criteria. Congenital heart disease with (56%) and without (25%) correction were the most common risk factors. Echocardiograms revealed vegetations in 67%. Echocardiograms remain relatively deficient in patients with complex cyanotic heart disease (50% positive) vs. those with normal anatomy (83% positive). Streptococci (30%) and Staphylococcus aureus (21%) were the most common organisms identified overall; however, we observed a predominance of enterococci in infants <2 years of age (41%). Our study revealed a reduced rate of complications, especially of the central nervous system (7% vs. 25 and 33% in prior studies); however, our rate of S. aureus was lower (21% vs. 32 and 39%), and this pathogen continues to be associated with a higher incidence of complications. CONCLUSIONS: Infective endocarditis continues to present a difficult challenge in pediatrics; its associated pathogens and outcomes have changed little in the past decade.

Age Distribution↗
PubMed2003

Characterization of chlamydial genital infection resulting from sexual transmission from male to female guinea pigs and determination of infectious dose.

A major problem in the study of chlamydial genital infections in animal models has been the use of varied doses of chlamydiae for infection in different laboratories. It is clearly desirable to use a dose which approximates that of natural sexual infection, but that dose to date has not been determined because of the inability of researchers to quantify chlamydiae in semen. Fortunately, sexual transmission of chlamydiae has been described for the guinea pig model of infection with the chlamydial agent of guinea pig inclusion conjunctivitis (GPIC). In this study, we undertook to determine the approximate infection dose in actual sexual transmission by comparing the kinetics of infection in female guinea pigs acquired via sexual contact to those of genital infections induced artificially with known quantities of chlamydiae. Groups of guinea pigs were infected intravaginally with 10(4), 10(3), 10(2), and 10(1) inclusion-forming units (IFU) of GPIC, and the kinetics of the infection were determined. Infection with 10(2) IFU produced infections with lower peak levels than those in animals receiving 10(4) or 10(3) IFU. Seventy percent of animals receiving 10(2) IFU became infected, while 100 and 79% of animals receiving 10(4) and 10(3) IFU, respectively, became infected. Animals receiving 10(2) IFU also had a longer incubation period. Of 19 animals that mated with infected males, 63.2% became infected, with an infection course which was not significantly different than that of the 10(2)-IFU-infected group. The data suggest that female guinea pigs received approximately 10(2) IFU by sexual transmission. Of interest was the observation that the guinea pigs infected by sexual transmission shed organisms for a significantly shorter time period than that of any group that was artificially infected. This result suggests that there may be factors associated with semen which passively transfer antimicrobial activity to the female or enhance the innate host response in the female. Immunization of females with an inactivated vaccine was also found to elicit a protective immune response against sexual challenge, demonstrating that the model can be used in the evaluation of possible vaccine candidates and/or methodologies. There is currently no other animal model available for any sexually transmitted disease in which the disease or the ability to prevent the disease may be studied in animals infected by the natural means.

Animals↗
PubMed2002

Role of proapoptotic BAX in propagation of Chlamydia muridarum (the mouse pneumonitis strain of Chlamydia trachomatis) and the host inflammatory response.

The BCL-2 family member BAX plays a critical role in regulating apoptosis. Surprisingly, bax-deficient mice display limited phenotypic abnormalities. Here we investigate the effect of BAX on infection by the sexually transmitted pathogen, Chlamydia muridarum (the mouse pneumonitis strain of Chlamydia trachomatis). Bax(-/-) cells are relatively resistant to Chlamydia-induced apoptosis, and fewer bacteria are recovered after two infection cycles from Bax(-/-) cells than from wild-type cells. These results suggest that BAX-dependent apoptosis may be used to initiate a new round of infection, most likely by releasing Chlamydia-containing apoptotic bodies from infected cells that could be internalized by neighboring uninfected cells. Nonetheless, infected Bax(-/-) cells die through necrosis, which is normally associated with inflammation, more often than infected wild-type cells. These studies were confirmed in mice infected intravaginally with C. muridarum; since the infection disappears more quickly from Bax(-/-) mice than from wild-type mice, secretion of proinflammatory cytokines is increased in Bax(-/-) mice, and large granulomas are present in the genital tract of Bax(-/-) mice. Taken together, these data suggest that chlamydia-induced apoptosis via BAX contributes to bacterial propagation and decreases inflammation. Bax deficiency results in lower infection and an increased inflammatory cytokine response associated with more severe pathology.

Animals↗
PubMed2002

Inhibition of apoptosis by gamma interferon in cells and mice infected with Chlamydia muridarum (the mouse pneumonitis strain of Chlamydia trachomatis).

The effect of gamma interferon (IFN-gamma) on apoptosis due to infection by Chlamydia muridarum (the mouse pneumonitis strain of Chlamydia trachomatis) was studied in epithelial cells in culture and in the genital tracts of mice. IFN-gamma concentrations that induce the formation of aberrant, persistent chlamydiae inhibit apoptosis due to C. muridarum infection. In cells treated with an IFN-gamma concentration that leads to the development of a heterogenous population of normal and aberrant Chlamydia vacuoles, apoptosis was inhibited preferentially in cells that contained the aberrant vacuoles. The inhibitory effect of IFN-gamma appears to be due in part to expression of host cell indoleamine 2,3-dioxygenase activity, since inhibition of apoptosis could be partially reversed through coincubation with exogenous tryptophan. Apoptotic cells were observed in the genital tracts of wild-type mice infected with C. muridarum, and a significantly larger number of apoptotic cells was detected in infected IFN-gamma-deficient mice. These results suggest that IFN-gamma may contribute to pathogenesis of persistent Chlamydia infections in vivo by preventing apoptosis of infected cells.

Animals↗