Leukocyte sequestration in pulmonary microvessels and lung injury following systemic complement activation in rabbits

Inflammatory reactions are associated with sequestration of leukocytes in the lung. Complement activation leads to accumulation of leukocytes in alveolar septa and alveoli, to lung edema and hemorrhage. Although in organs other than the lung leukocytes interact with the vascular endothelium only in postcapillary venules, alveolar capillaries are considered to be the site of leukocyte sequestration in the lung. However, pulmonary venules and arterioles have not been investigated systematically after complement activation so far, A closed thoracic window was implanted in anesthetized rabbits; leukocytes and red blood cells were stained, and the movement of these cells was measured in superficial pulmonary arterioles, venules and alveolar capillaries using fluorescence video microscopy before and 30 and 60 min after infusion of cobra venom factor (CVF). Erythrocyte velocity and macrohemodynamic conditions did not change after CVF infusion and were not different from the sham-treated controls. The number of sticking leukocytes increased significantly compared to baseline and control: by 150% in arterioles and in venules and by 740% in alveolar capillaries within 60 min after CVF infusion. The width of alveolar septa in vivo was significantly enlarged after CVF infusion, indicating interstitial pulmonary edema. At the end of the experiments, myeloperoxidase activity was higher in the CVF group, showing leukocyte sequestration in the whole organ. It is concluded that complement activation by CVF induces leukocyte sequestration in lung arterioles, venules and alveolar capillaries and leads to mild lung injury

The Female Cervicovaginal Mucosa Is a Unique Site for the Production of Autoantibodies Associated with Rheumatoid Arthritis

Purpose/Background: Women have a 3-fold higher incidence of rheumatoid arthritis (RA) and a lower likelihood of remission compared to men suggesting a gender disparity in the etiology of RA. In order to devise female specific prevention and treatment strategies, it is critical to understand the mechanism initiating the production of RA autoantibodies termed anti-citrullinated protein antibodies (ACPA). ACPA target proteins that are posttranslationally modified by a family of enzymes termed peptidylarginine deiminases (PADs), which convert arginine into citrulline. Research suggests that ACPA are generated at a mucosal site years before becoming systemic and causing clinical joint disease. Mucosal sites such as the lung, gut, and gingiva have been explored as sites of ACPA production, yet none of these account for the higher incidence of RA in women. We hypothesize that the cervicovaginal mucosa is a novel, sex-specific site for APCA production in women. Materials & Methods: To begin to test this hypothesis, healthy control (HC) women, women at risk for RA (AR), and those with clinical RA self-collected cervicovaginal fluid (CVF) at three time points during the menstrual cycle. CVF samples were examined for PAD activity, total citrulline concentration, and cyclic citrullinated peptides (CCP) as a marker for ACAP levels. Results: In naturally cycling HC women, CCP peak in early follicular phase (d5), dropped substantially by ovulation (d14), and remained low at the end of the luteal phase (d26). PAD enzymatic activity and total citrulline concentration also peak in CVF at d5 of the menstrual cycle, suggesting that changes in citrullinated proteins may drive local ACPA production. We next examined if CCP, PAD activity, and total citrulline concentration are increased in CVF from women at-risk (AR) for developing RA and women with RA. Although PAD activity and total citrulline concentration does not increase in these groups compared to health controls, CCP levels are significantly increased between the HC and RA CVF samples at d25. At issue is the identity of the citrullinated proteins in HC, AR and RA CVF, and if their abundance changes across the cycle and with disease progression. To address this, we performed mass spectrometry on CVF samples which identified a number of citrullinated proteins present in HC, AR, and RA women. Discussion/Conclusion: Our work suggests that citrullinated proteins and ACPA are produced in the cervicovaginal mucosa and may help explain why women have increased risk of developing RA

Technical Report: Using Static Analysis to Compute Benefit of Tolerating Consistency

Synchronization is the Achilles heel of concurrent programs. Synchronization requirement is often used to ensure that the execution of the concurrent program can be serialized. Without synchronization requirement, a program suffers from consistency violations. Recently, it was shown that if programs are designed to tolerate such consistency violation faults (\cvf{s}) then one can obtain substantial performance gain. Previous efforts to analyze the effect of \cvf-tolerance are limited to run-time analysis of the program to determine if tolerating \cvf{s} can improve the performance. Such run-time analysis is very expensive and provides limited insight. In this work, we consider the question, `Can static analysis of the program predict the benefit of \cvf-tolerance?' We find that the answer to this question is affirmative. Specifically, we use static analysis to evaluate the cost of a \cvf and demonstrate that it can be used to predict the benefit of \cvf-tolerance. We also find that when faced with a large state space, partial analysis of the state space (via sampling) also provides the required information to predict the benefit of \cvf-tolerance. Furthermore, we observe that the \cvf-cost distribution is exponential in nature, i.e., the probability that a \cvf has a cost of $c$ is $A.B^{-c}$, where $A$ and $B$ are constants, i.e., most \cvf{s} cause no/low perturbation whereas a small number of \cvf{s} cause a large perturbation. This opens up new aveneus to evaluate the benefit of \cvf-tolerance

Cervicovaginal fluid and semen block the microbicidal activity of hydrogen peroxide produced by vaginal lactobacilli

BACKGROUND: H(2)O(2 )produced by vaginal lactobacilli is believed to protect against infection, and H(2)O(2)-producing lactobacilli inactivate pathogens in vitro in protein-free salt solution. However, cervicovaginal fluid (CVF) and semen have significant H(2)O(2)-blocking activity. METHODS: We measured the H(2)O(2 )concentration of CVF and the H(2)O(2)-blocking activity of CVF and semen using fluorescence and in vitro bacterial-exposure experiments. RESULTS: The mean H(2)O(2 )measured in fully aerobic CVF was 23 ± 5 μM; however, 50 μM H(2)O(2 )in salt solution showed no in vitro inactivation of HSV-2, Neisseria gonorrhoeae, Hemophilus ducreyii, or any of six BV-associated bacteria. CVF reduced 1 mM added H(2)O(2 )to an undetectable level, while semen reduced 10 mM added H(2)O(2 )to undetectable. Moreover, the addition of just 1% CVF supernatant abolished in vitro pathogen-inactivation by H(2)O(2)-producing lactobacilli. CONCLUSIONS: Given the H(2)O(2)-blocking activity of CVF and semen, it is implausible that H(2)O(2)-production by vaginal lactobacilli is a significant mechanism of protection in vivo

Cervicovaginal fluid acetate: a metabolite marker of preterm birth in symptomatic pregnant women

Changes in vaginal microbiota that is associated with preterm birth (PTB) leave specific metabolite fingerprints that can be detected in the cervicovaginal fluid (CVF) using metabolomics techniques. In this study, we characterize and validate the CVF metabolite profile of pregnant women presenting with symptoms of threatened preterm labor (PTL) by both 1H-nuclear magnetic resonance spectroscopy (NMR) and enzyme-based spectrophotometry. We also determine their predictive capacity for PTB, singly, and in combination, with current clinical screening tools – cervicovaginal fetal fibronectin (FFN) and ultrasound cervical length (CL). CVF was obtained by high-vaginal swabs from 82 pregnant women with intact fetal membranes presenting between 24 and 36 weeks gestation with symptoms of threatened, but not established, PTL. Dissolved CVF samples were scanned with a 400 MHz NMR spectrometer. Acetate and other metabolites were identified in the NMR spectrum, integrated for peak area, and normalized to the total spectrum integral. To confirm and validate our observations, acetate concentrations (AceConc) were also determined from a randomly-selected subset of the same samples (n = 57), by spectrophotometric absorption of NADH using an acetic acid assay kit. CVF FFN level, transvaginal ultrasound CL, and vaginal pH were also ascertained. Acetate normalized integral and AceConc were significantly higher in the women who delivered preterm compared to their term counterparts (P = 0.002 and P = 0.006, respectively). The 1H-NMR-derived acetate integrals were strongly correlated with the AceConc estimated by spectrophotometry (r = 0.69; P 0.53 g/l), and of delivery within 2 weeks of the index assessment (acetate integral: AUC = 0.77, 95% CI = 0.58–0.96; AceConc: AUC = 0.68, 95% CI = 0.5–0.9). The predictive accuracy of CVF acetate was similar to CL and FFN. The combination of CVF acetate, FFN, and ultrasound CL in a binary logistic regression model improved the prediction of PTB compared to the three markers individually, but CVF acetate offered no predictive improvement over ultrasound CL combined with CVF FFN. Elevated CVF acetate in women with symptoms of PTL appears predictive of preterm delivery, as well as delivery within 2 weeks of presentation. An assay of acetate in CVF may prove of clinical utility for predicting PTB

Comprehensive proteomic analysis of human cervical-vaginal fluid using colposcopy samples

<p>Abstract</p> <p>Background</p> <p>Cervical-vaginal fluid (CVF) plays an important role in the prevention of gynecological infections, although little is known about the contribution of CVF proteins to the immunity of the lower female genital tract. In order to analyze the protein composition of human CVF, we used CVF samples that are routinely collected during colposcopy, but are usually discarded. Since these samples are available in large quantities we aimed to analyze their usefulness for proteomics experiments. The samples were analyzed using different prefractionation techniques (ultrafiltration and C₄(RP)-LC protein separation) followed by C₁₈(RP)-LC peptide separation and identification by MALDI-TOF-TOF mass spectrometry. To determine the reproducibility of this proteomics platform we analyzed three technical replicates. Using spectral counting, protein abundances were estimated in a semiquantitative way. We also compared the results obtained in this study with those from previous studies derived from patients with different physiological conditions in order to determine an overlapping protein set.</p> <p>Results</p> <p>In total, we were able to identify 339 proteins in human CVF of which 151 proteins were not identified in any other proteomics study on human CVF so far. Those included antimicrobial peptides, such as human beta-defensin 2 and cathelicidin, which were known to be present in CVF, and endometrial proteins such as glycodelin and ribonucleoprotein A. Comparison of our results with previously published data led to the identification of a common protein set of 136 proteins. This overlapping protein set shows increased fractions of immunological and extracellular proteins, confirming the extracellular immunological role of CVF.</p> <p>Conclusion</p> <p>We demonstrated here that CVF colposcopy samples can be used in proteomics experiments and hence are applicable for biomarker discovery experiments. The delineation of an overlapping set of proteins that is identified in most proteomics studies on CVF may help in the description of a reference proteome when performing proteomics studies on human CVF.</p

CVF Open House invitation

Open House at Milestone Tower