Mode-coupling instability in a fluid two-dimensional complex plasma.

A theory of the mode-coupling instability (MCI) in a fluid two-dimensional complex plasma is developed. In analogy to the point-wake model of the wake-mediated interactions commonly used to describe MCI in two-dimensional crystals, the layer-wake model is employed for fluids. It is demonstrated that the wake-induced coupling of wave modes occurs in both crystalline and fluid complex plasmas, but the confinement-density threshold, which determines the MCI onset in crystals, virtually disappears in fluids. The theory shows excellent qualitative agreement with available experiments and provides certain predictions to be verified.

Icilin reduces voltage-gated calcium channel currents in naïve and injured DRG neurons in the rat spinal nerve ligation model.

Recently, the transient receptor potential (TRP) channels TRPM8 and TRPA1 have been identified as molecular sensors for cold, and it has been suggested that they play a crucial role in allodynia by modulating voltage-gated calcium channel currents (ICa(V)). The aim of this study was to analyze the modulation of ICa(V) by the TRPM8-agonist icilin in vitro and to investigate the analgesic effect of icilin in a neuropathic pain model in vivo. Whole cell patch-clamp recordings were performed on isolated naïve and injured rat dorsal root ganglia (DRG) neurons, and the analgesic efficacy of icilin applied topically to the paws or intrathecally was tested in rats after spinal nerve ligation (SNL). ICa(V) (depolarization from -80 to 0mV) in naïve DRG neurons was reduced dose dependently (0.002-200µM) by icilin (18-80%). Subtype isolation of calcium channels show a marked reduction of L-type channel currents compared to N-type channel currents. The effects of icilin on ICa(V) were not significantly different in non-injured and SNL-injured DRG neurons. In vivo, neither topical (10-200µM) nor intrathecal application of icilin (0.1nM to 1µM) affected tactile allodynia or thermal hyperalgesia after SNL, but it increases cold allodynia 6h after application. We conclude that the icilin-induced modulation of ICa(V) in DRG neurons is unlikely to mediate analgesic effects or contribute directly to the pathogenesis of cold allodynia in the rat SNL model, but it is a potential mechanism for the analgesic effects of icilin in other pain models.

Identification and evaluation of a combination of chlamydial antigens to support the diagnosis of severe and invasive Chlamydia trachomatis infections.

Chlamydia trachomatis is the most common sexually transmitted organism in industrialized countries. Nucleic acid amplification testing, using non-invasively collected specimens, is considered to be the method of choice for diagnosis of chlamydial infections of the urethra and the lower genital tract. Serological testing has the potential to circumvent the problem of specimen sampling in invasive C. trachomatis infections of the upper genital tract. However, only a few defined chlamydial antigens have been used in a standardized diagnostic assay format. In this study, we used serological two-dimensional proteomic analysis to broaden the spectrum of diagnostically relevant C. trachomatis proteins. The genes encoding an assortment of already known chlamydial antigens, as well as immunogenic proteins that have not been described before, were cloned, and the recombinant proteins were purified in order to compare their diagnostic usefulness in parallel with a newly developed line immunoassay. With 189 sera collected from patients with and without C. trachomatis infection, recombinant major outer membrane protein (MOMP), chlamydial protease-like activity factor (CPAF), outer membrane protein 2 (OMP2), translocated actin-recruiting protein, and polymorphic membrane protein D (PmpD) showed the highest level of diagnostic sensitivity and specificity. In patients suffering from ascending and invasive C. trachomatis infections, such as pelvic inflammatory disease and lymphogranuloma venereum, the sensitivity reached with these proteins ranged between 71% (PmpD) and 94% (OMP2), and the specificity ranged between 82% (PmpD) and 100% (MOMP and OMP2). Recombinant thio-specific antioxidant peroxidase, ribosomal protein S1 (RpsA) and hypothetical protein 17 showed lower sensitivity but comparably high specificity, ranging from 94% to 100%. The novel line immunoassay based on defined recombinant antigens has promise for improved serodiagnosis in severe and invasive C. trachomatis infections.

Evaluation of WLBU2 peptide and 3-O-octyl-sn-glycerol lipid as active ingredients for a topical microbicide formulation targeting Chlamydia trachomatis.

Topical microbicides for prevention of sexually transmitted diseases (STDs) would be especially useful for women who are not able to persuade their partner(s) to take precautions. Many topical microbicides are in various stages of development, based on a variety of active ingredients. We investigated the in vitro activity of an engineered antimicrobial peptide (WLBU2) and a lipid (3-O-octyl-sn-glycerol [3-OG]) which could potentially be used as active ingredients in such a product. Using commercially available cytotoxicity reagents [Alamar Blue, 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), and lactate dehydrogenase (LDH)], we first determined the toxicity of WLBU2 and 3-OG to the host cells in our assay procedure and excluded toxic concentrations from further testing. To determine activity against Chlamydia trachomatis, we used an assay previously developed by our laboratory in which chlamydial elementary bodies (EBs) were exposed to microbicides prior to contact with epithelial cells: the minimum (microbi)cidal concentration (MCC) assay. To further simulate conditions of transmission, we carried out the same assay in the presence of a simulated vaginal fluid, a simulated seminal fluid, human serum albumin, and a range of pH values which might be found in the human vagina at the time of exposure. Last, we tested WLBU2 and 3-OG in combination to determine if adding them together resulted in synergistic activity. We found that WLBU2 and 3-OG both have excellent activity in vitro against C. trachomatis and significantly more activity when added together. The simulated fluids reduced activity, but the synergy seen is good evidence that they would be effective when combined in a microbicide formulation.

ProACT for stress urinary incontinence after radical prostatectomy.

INTRODUCTION: Stress urinary incontinence is a bothersome complication of radical prostatectomy. Surgical treatment consists of the artificial urinary sphincter (AUS), the male sling and bulk injections. This study presents the results of the first series of implantations of ProACT in the Netherlands. MATERIALS AND METHODS: A non-validated questionnaire was sent to 29 male patients implanted with ProACT to determine Stamey score, pad count and questions about quality of life and satisfaction. Complications, revisions and explantations were registered. RESULTS: Mean follow-up was 41 months. Based on Stamey score four patients are continent at the end and nine patients according to the pad count. The average pad count decreased significantly. Remarkable was the high rate of dislocations and revisions and patients' satisfaction. CONCLUSIONS: ProACT is a less invasive treatment compared to the AUS. However, the procedure is associated with a substantial revision and explantation rate. ProACT can be part of a so-called step-up approach before opting for a more invasive treatment.

Deterministic and probabilistic approaches for tracking virus particles in time-lapse fluorescence microscopy image sequences.

Modern developments in time-lapse fluorescence microscopy enable the observation of a variety of processes exhibited by viruses. The dynamic nature of these processes requires the tracking of viruses over time to explore spatial-temporal relationships. In this work, we developed deterministic and probabilistic approaches for multiple virus tracking in multi-channel fluorescence microscopy images. The deterministic approaches follow a traditional two-step paradigm comprising particle localization based on either the spot-enhancing filter or 2D Gaussian fitting, as well as motion correspondence based on a global nearest neighbor scheme. Our probabilistic approaches are based on particle filters. We describe approaches based on a mixture of particle filters and based on independent particle filters. For the latter, we have developed a penalization strategy that prevents the problem of filter coalescence (merging) in cases where objects lie in close proximity. A quantitative comparison based on synthetic image sequences is carried out to evaluate the performance of our approaches. In total, eight different tracking approaches have been evaluated. We have also applied these approaches to real microscopy images of HIV-1 particles and have compared the tracking results with ground truth obtained from manual tracking. It turns out that the probabilistic approaches based on independent particle filters are superior to the deterministic schemes as well as to the approaches based on a mixture of particle filters.

Chlamydia trachomatis laboratory strains versus recent clinical isolates: implications for routine microbicide testing.

A topical microbicide that women can use to prevent sexually transmitted diseases (STDs) is essential, and many microbicide candidates are being tested for activity against human immunodeficiency virus and other STDs, including Chlamydia trachomatis. Screening assays for assessing the activity of microbicides against C. trachomatis are typically done with laboratory-adapted strains, but it is possible that recent clinical isolates may have different susceptibilities to microbicides, as has been seen with Neisseria gonorrhoeae and Lactobacillus spp. (B. J. Moncla and S. L. Hillier, Sex. Transm. Dis. 32:491-494, 2005). We utilized three types of microbicides to help define this aspect of our assay to test microbicides against C. trachomatis in vitro. To simulate conditions of transmission, we used an assay that we previously developed in which we exposed chlamydial elementary bodies to microbicides prior to contact with epithelial cells. We first determined the toxicity of microbicides to the cells used to culture Chlamydia trachomatis in the assay and, if necessary, modified the assay to eliminate toxicity at the concentrations tested. We compared the sensitivities of recent clinical isolates of Chlamydia trachomatis versus laboratory strains of the same serovar and found major differences in sensitivity to nonoxynol-9 (non-9), but only minor differences were seen with the other microbicides. We thus conclude that when assessing activity of potential topical microbicides versus the obligate intracellular bacteria C. trachomatis, the use of recent clinical isolates may not be necessary to draw a conclusion about a microbicide's effectiveness. However, it is important to keep in mind that differences (like those seen with non-9) are possible and that clinical isolates could be included in later stages of testing.

Eight versus 16-week re-evaluation period in rheumatoid arthritis patients treated with leflunomide or methotrexate accompanied by moderate dose prednisone.

In a step-up approach of DMARD treatment of RA a fast response and an early DMARD switch in the case of non-response is important. Therefore, we performed an open trial in which we compared an 8-week and a 16-week observation period during treatment of RA with MTX or LEF, both given in intensified starting doses and accompanied by moderate dose prednisone. MTX and LEF naïve patients with RA (mean time since diagnosis: 2.3 years) were randomised to receive either LEF in a 3-day-loading dose of 100 mg/day followed by 20 mg/day (n = 19) or MTX intramuscularly in a dose of 25 mg once weekly (n = 21). All patients received concomitant treatment with oral prednisone in an initial dose of 20 mg/day with weekly dose reductions of 5 mg/day. The disease activity was re-evaluated 8 and 16 weeks after the start of the treatment. Mean DAS28 before the start of treatment was 5.36 +/- 0.8 for the MTX-group and 5.46 +/- 0.8 for the LEF-group. After 8 weeks of treatment the DAS28 in the MTX-group was 2.59 +/- 1.0 and 3.16 +/- 0.8 in the LEF group (difference not significant). The mean DAS28 at re-evaluation 16 weeks after the starting of treatment (2.58 +/- 1.5 for the MTX-group and 3.25 +/- 1.16 for the LEF-group) was significantly different neither in between the both treatment groups nor in comparison to the week 8 evaluation. Efficiency of RA treatment with MTX or LEF in intensified doses and in combination with moderate dose prednisone can be sufficiently judged 8 weeks after its initiation.

Susceptibility of Chlamydia trachomatis to excipients commonly used in topical microbicide formulations.

Commonly used "inactive" pharmaceutical excipients were tested in a previously developed minimum cidal concentration assay to assess their ability to kill Chlamydia trachomatis topically. Sixteen excipients were evaluated in these studies under various conditions. A range of activities was found among the excipients that could be tested in our assay system.

Development of new prostate specific monoclonal antibodies.

BACKGROUND: Despite the need for new prostate-specific diagnostic and therapeutic targets, very few unique prostate (cancer) specific antigens have been characterized. Monoclonal antibody (mAb) technology is a powerful tool to identify specific antigenic markers, which could be potential targets for cancer diagnostics or therapy. METHODS: Splenocytes from mice immunized with prostate cancer (PCa) homogenates of different origin were fused using standard techniques. Employing a differential high-throughput screening method followed by immediate screening in immunohistochemistry (IHC) a large number of hybridomas were screened for prostate (cancer) specificity. RESULTS: From 25 successful fusions approximately 300 clones were identified excreting PCa-reactive antibodies. Subsequent immunohistochemical fine-specificity analysis reduced this number to 26. Eventually, after extensive fine-specificity analysis, the number of mAbs appearing to define prostate-specific antigenic structures that might serve as new diagnostic or therapeutic targets was reduced to three. CONCLUSIONS: Using mAb technology combined with a high throughput screening method we have developed three mAbs (1.8, 2.26, and 3.10) directed against prostate associated antigens that might identify potential new therapeutic targets.

In vitro microbicidal activities of cecropin peptides D2A21 and D4E1 and gel formulations containing 0.1 to 2% D2A21 against Chlamydia trachomatis.

Topically applied microbicides that eradicate pathogens at the time of initial exposure represent a powerful strategy for the prevention of sexually transmitted infections. To aid in the further development of an effective topical microbicide, we assessed the minimum cidal concentration (MCC) of two cecropin peptides, D2A21 and D4E1, and gel formulations containing 0.1 to 2% D2A21 against Chlamydia trachomatis in vitro. The MCC of peptide D2A21 was 5 microM (18.32 microg/ml), and that of peptide D4E1 was 7.5 microM (21.69 microg/ml). The MCC of gel formulations containing 2% D2A21 was 0.2 mM (0.7 mg/ml), and that of gel formulations containing 0.5% D2A21 was 0.2 mM (0.7 mg/ml). There was no significant variation in the results when two different C. trachomatis strains were tested, and the addition of 10% human blood did not significantly alter the MCCs. pH values above and below 7 reduced the activity of the D2A21 peptide alone, but the MCC of the 2% D2A21 gel formulation was only slightly altered at the various pHs tested. Ultrastructural studies indicated that C. trachomatis membranes were disrupted after D2A21 exposure, resulting in leakage of the cytoplasmic contents. These in vitro results suggest that these cecropin peptides may be an effective topical microbicide against C. trachomatis and support the need for further evaluation.

Effect of trapped ions on shielding of a charged spherical object in a plasma.

Collisions have traditionally been neglected in calculating the shielding around a small spherical collector in a plasma, and the plasma flow to the collector. We show analytically that, in dusty plasmas under typical discharge conditions, ion charge-exchange collisions lead to the buildup of negative-energy trapped ions which dominate the shielding cloud in the nonlinear region near a dust grain and substantially increase the ion current to the grain, even when the mean-free path is much greater than the Debye length.

CD8+ T cells recognize an inclusion membrane-associated protein from the vacuolar pathogen Chlamydia trachomatis.

During infection with Chlamydia trachomatis, CD8(+) T cells are primed, even though the bacteria remain confined to a host cell vacuole throughout their developmental cycle. Because CD8(+) T cells recognize antigens processed from cytosolic proteins, the Chlamydia antigens recognized by these CD8(+) T cells very likely have access to the host cell cytoplasm during infection. The identity of these C. trachomatis proteins has remained elusive, even though their localization suggests they may play important roles in the biology of the organism. Here we use a retroviral expression system to identify Cap1, a 31-kDa protein from C. trachomatis recognized by protective CD8(+) T cells. Cap1 contains no strong homology to any known protein. Immunofluorescence microscopy by using Cap1-specific antibody demonstrates that this protein is localized to the vacuolar membrane. Cap1 is virtually identical among the human C. trachomatis serovars, suggesting that a vaccine incorporating Cap1 might enable the vaccine to protect against all C. trachomatis serovars. The identification of proteins such as Cap1 that associate with the inclusion membrane will be required to fully understand the interaction of C. trachomatis with its host cell.

New mothers' knowledge and attitudes about perinatal human immunodeficiency virus infection.

OBJECTIVES: To assess new mothers' attitudes toward perinatal human immunodeficiency virus (HIV) testing, their knowledge about perinatal HIV, and their trust of government and scientists. METHODS: In a cross-sectional survey of 1362 postpartum women at four United States locations in 1997, a standardized interview was administered to new mothers 24-48 hours postpartum to determine their HIV test acceptance, attitudes, and knowledge. RESULTS: Seventy-five percent of women who were offered HIV tests reported being tested. Although 95% of women were aware of perinatal HIV transmission, only 60% knew that HIV can be transmitted through breast-feeding, and only 51% knew of medication to prevent perinatal transmission. Eighty-four percent of women thought that all pregnant women should be tested for HIV, and 60% thought that prenatal HIV testing should be legally mandated. Twenty percent of women indicated mistrust of government and scientists regarding origins of HIV and potential cures for AIDS. Knowledge about perinatal transmission was unrelated to receipt of prenatal HIV tests. When other factors were controlled for, mistrust was not significantly associated with getting tested. CONCLUSION: Incomplete knowledge of prevention of perinatal HIV transmission and mistrust were prevalent among new mothers. Knowledge deficits or mistrust did not appear to reduce reported prenatal test rates, but our data suggest that future public health efforts need to educate women about methods of preventing perinatal HIV transmission and at enhancing their trust in the public health system.

Lack of evidence of mitochondrial dysfunction in the offspring of HIV-infected women. Retrospective review of perinatal exposure to antiretroviral drugs in the Perinatal AIDS Collaborative Transmission Study.

A recent report suggesting mitochondrial dysfunction among eight HIV-exposed but uninfected children exposed perinatally to nucleoside reverse transcriptase inhibitors (NRTIs) prompted a review within the Perinatal AIDS Collaborative Transmission Study (PACTS). A standardized retrospective review was conducted of 118 deaths at < 5 years. Deaths were classified as unrelated to mitochondrial dysfunction (Class 1), unlikely related (Class 2), possibly related (Class 3), or likely related or proven (Class 4). Among 35 deaths recorded in HIV-uninfected or indeterminate children, none were classified in either Class 2, 3, or 4. We also reviewed signs or symptoms consistent with possible mitochondrial dysfunction among 1,954 living uninfected children. Only one child was in Class 3 and two siblings were in Class 2; none had perinatal antiretroviral drug exposure. We found no evidence indicating that uninfected infants exposed to perinatal NRTIs died of mitochondrial disorders or that living exposed children had symptoms of mitochondrial dysfunction.

Fragile X syndrome and an isodicentric X chromosome in a woman with multiple anomalies, developmental delay, and normal pubertal development.

We report on an individual with developmental delays, short stature, skeletal abnormalities, normal pubertal development, expansion of the fragile X triplet repeat, as well as an isodicentric X chromosome. S is a 19-year-old woman who presented for evaluation of developmental delay. Pregnancy was complicated by a threatened miscarriage. She was a healthy child with intellectual impairment noted in infancy. Although she had global delays, speech was noted to be disproportionately delayed with few words until age 3.5 years. Facial appearance was consistent with fragile X syndrome. Age of onset of menses was 11 years with normal breast development. A maternal male second cousin had been identified with fragile X syndrome based on DNA studies. The mother of this child (S's maternal first cousin) and the grandfather (S's maternal uncle) were both intellectually normal but were identified as carrying triplet expansions in the premutation range. S's mother had some school difficulties but was not identified as having global delays. Molecular analysis of S's fragile X alleles noted an expansion of more than 400 CGG repeats in one allele. Routine cytogenetic studies of peripheral blood noted the presence of an isodicentric X in 81of 86 cells scored. Five of 86 cells were noted to be 45,X. Cytogenetic fra(X) studies from peripheral blood showed that the structurally normal chromosome had the fragile site in approximately 16% of the cells. Analysis of maternal fragile X alleles identified an allele with an expansion to approximately 110 repeats. FMRP studies detected the expression of the protein in 24% of cells studied. To our knowledge, this is the first patient reported with an isodicentric X and fragile X syndrome. Whereas her clinical phenotype is suggestive of fragile X syndrome, her skeletal abnormalities may represent the presence of the isodicentric X. Treatment of S with 20 mg/day of Prozac improved her behavior. In the climate of cost con trol, this individual reinforces the recommendation of obtaining chromosomes on individuals with developmental delay even with a family history of fragile X syndrome.

FMRP expression as a potential prognostic indicator in fragile X syndrome.

Absence or deficit of FMR1 protein (FMRP) resulting from methylation of full mutation genes is the fundamental defect in fragile X syndrome. We used FMRP immunocytochemistry and detailed phenotypic assessment to investigate the relationship between degree of FMRP expression and the broad clinical spectrum of impairment in 80 individuals affected with fragile X syndrome. FMRP expression correlated with IQ in mosaic males (P=0.043), males with a partially methylated full mutation (P=0.0005), and females with a full mutation (P=0.046). In the females, FMRP expression also correlated with the number of fragile X physical features (P=0.0003). Even modest deficits in FMRP result in some manifestations of fragile X syndrome. In this initial study of 53 males, FMRP expression testing had a very high positive predictive value (100%, confidence interval of 29-100%) for a nonretarded IQ among males with expression of FMRP in > or = 50% of lymphocytes (3 males), suggesting that FMRP expression may have potential as a prognostic indicator in males with fragile X syndrome.

Electrodermal responses to sensory stimuli in individuals with fragile X syndrome: a preliminary report.

The fragile X mutation and fragile X syndrome are associated with hyperarousal, hyperactivity, aggression, and anxiety. These may be related to strong reactions to auditory, tactile, visual, and olfactory stimuli [Hagerman, 1996b; Hagerman and Cronister, 1996]. However, almost no data exist describing hyperarousal and sensory sensitivity in individuals with the fragile X mutation. This study establishes a reliable laboratory paradigm for examining reactions to sensory stimuli. We found the pattern of electrodermal responses (EDRs) to stimulation in one sensory modality predicted the pattern of EDRs in four other sensory systems. In addition, the EDR pattern of individuals with the fragile X mutation was related to their FMR-protein expression. Finally, EDRs in individuals with fragile X syndrome were significantly different from those of normal controls, demonstrating greater magnitude, more responses per stimulation, responses on a greater proportion of trials, and lower rates of habituation. The findings support the theory that individuals with fragile X syndrome have a physiologically based enhancement of reactions to sensations. Because electrodermal activity indexes sympathetic nervous system activity, the data suggest that the over-arousal to sensation may involve the sympathetic system.

Gamma interferon production by cytotoxic T lymphocytes is required for resolution of Chlamydia trachomatis infection.

In this study, we used mice in which the gene for gamma interferon (IFN-gamma) has been disrupted (IFN-gamma-/- mice) to study the role of this cytokine in the resolution of Chlamydia trachomatis infection. We show that IFN-gamma-/- mice are impaired in the ability to clear infection with C. trachomatis compared to IFN-gamma+/+ control mice. Activated CD8(+) cytotoxic T lymphocytes (CTL) secrete IFN-gamma in response to intracellular infection, and we have shown previously that a Chlamydia-specific CTL line can reduce C. trachomatis infection when adoptively transferred into infected mice. In the present study, we found that when these IFN-gamma+/+ CTL lines are transferred into Chlamydia-infected IFN-gamma-/- mice, the transferred CTL cannot overcome the immune defect seen in the IFN-gamma-/- mice. We also show that Chlamydia-specific CTL can be cultured from IFN-gamma-deficient mice infected with C. trachomatis; however, the adoptive transfer of IFN-gamma-/- CTL into infected IFN-gamma+/+ mice does not reduce the level of infection. These results suggest that IFN-gamma production by CTL is not sufficient to overcome the defect that IFN-gamma-/- mice have in the resolution of Chlamydia infection, yet IFN-gamma production by CTL is required for the protective effect seen upon adoptive transfer of CTL into IFN-gamma+/+ mice.