A specific plasminogen activator inhibitor‐1 antagonist derived from inactivated urokinase

Fibrinolysis is a process responsible for the dissolution of formed thrombi to re‐establish blood flow after thrombus formation. Plasminogen activator inhibitor‐1 (PAI‐1) inhibits urokinase‐type and tissue‐type plasminogen activator (uPA and tPA) and is the major negative regulator of fibrinolysis. Inhibition of PAI‐1 activity prevents thrombosis and accelerates fibrinolysis. However, a specific antagonist of PAI‐1 is currently unavailable for therapeutic use. We screened a panel of uPA variants with mutations at and near the active site to maximize their binding to PAI‐1 and identified a potent PAI‐1 antagonist, PAItrap. PAItrap is the serine protease domain of urokinase containing active‐site mutation (S195A) and four additional mutations (G37bR–R217L–C122A–N145Q). PAItrap inhibits human recombinant PAI‐1 with high potency (K_d = 0.15 nM) and high specificity. In vitro using human plasma, PAItrap showed significant thrombolytic activity by inhibiting endogenous PAI‐1. In addition, PAItrap inhibits both human and murine PAI‐1, allowing the evaluation in murine models. In vivo, using a laser‐induced thrombosis mouse model in which thrombus formation and fibrinolysis are monitored by intravital microscopy, PAItrap reduced fibrin generation and inhibited platelet accumulation following vascular injury. Therefore, this work demonstrates the feasibility to generate PAI‐1 inhibitors using inactivated urokinase.     

The Cu/Zn superoxide dismutase +35A/C (rs2234694) variant correlates with altered levels of protein carbonyls and glutathione and associates with severity of COPD in a Tunisian population

Chronic obstructive pulmonary disease (COPD) is a major cause of mortality that has been associated with inflammation and oxidative stress. The purpose of the present case–control study was to determine the relationships between oxidative stress-related genetic variants and the risk and severity of COPD, as well as, the influence of these variants on inflammatory and oxidative stress parameters. Genotyping of superoxide dismutase 1 (SOD1) + 35 A/C (rs2234694), catalase [A-21T (rs7943316), C-262T (rs1001179)] and glutathione peroxidase 1 (reduced glutathione (GSH)-Px1) 198Pro/Leu (rs1050450) was carried out in 143 patients with COPD and 216 healthy controls using PCR-RFLP. Serum levels of IL-6 and TNF-α were determined by enzyme-linked immunosorbent assays (ELISA), while the levels of reduced GSH, total antioxidant status (TAS), H₂O₂, lipid peroxides (TBARS) and protein carbonyls (PCs) were determined using spectrophotometric methods. We also evaluated the activities of GSH-Px, catalase, and superoxide dismutase (SOD) in both plasma and erythrocytes. We did not observe significant differences in the genotype and allele frequencies of chosen variants between COPD patients and healthy controls. A significant correlation was retrieved between the SOD1?+?35A/C variant and disease severity (odds ratios (OR) = 0.15, p?=?0.04). In addition, patients having the +35AC genotype presented increased plasma levels of GSH and a reduced level of PCs (p?=?0.03, p?=?0.04, respectively). The present data highlighted the important role of antioxidant enzymes and their genetic variants in the oxidative stress-mediated pathogenesis and progression of COPD.     

SubCellBarCode: Proteome-wide Mapping of Protein Localization and Relocalization

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Involvement of the Adhesion GPCRs Latrophilins in the Regulation of Insulin Release

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Conformational solution studies of the SDS micelle-bound 11-28 fragment of two Alzheimer's beta-amyloid variants (E22K and A21G) using CD, NMR, and MD techniques

The beta-amyloid (Abeta) is the major peptide constituent of neuritic plaques in Alzheimer's disease (AD) and its aggregation is believed to play a central role in the pathogenesis of the disease. Naturally occurring mutations resulting in changes in the Abeta sequence (pos. 21-23) are associated with familial AD-like diseases with extensive cerebrovascular pathology. It was proved that the mutations alter the aggregation ability of Abeta and its neurotoxicity. Among five mutations at positions 21-23 there are two mutations with distinct clinical characteristics and potentially distinct pathogenic mechanism-the Italian (E22K) and the Flemish (A21G) mutations. In our studies we have examined the structures of the 11-28 fragment of the Italian and Flemish Abeta variants. The fragment was chosen because it has been shown to be the most important for amyloid fibril formation. The detailed structure of both variants Abeta(11-28) was determined using CD, 2D NMR, and molecular dynamics techniques under water-SDS micelle conditions. The NMR analysis revealed two distinct sets of proton resonances for the peptides. The studies of both peptides pointed out the existence of well-defined alpha-helical conformation in the Italian mutant, whereas the Flemish was found to be unstructured with the possibility of a bent structure in the central part of the peptide.     

Leishmania (Viannia) braziliensis: epidemiology of canine cutaneous leishmaniasis in the State of Paraná (Brazil)

The present study examines the role that dogs play in the maintenance of the Leishmania cycle in the State of Paraná, Southern Brazil. Dogs were examined in three regions where cutaneous leishmaniasis is endemic or epidemic (R1-Vale da Ribeira; R2-Central region of Paraná State and R3-Northern region). To determine serum prevalence rates ELISA was used. In regions endemic for Trypanosoma cruzi (R1 and R3), serum from dogs seroreactive towards Leishmania antigen was subjected to T. cruzi adsorption in order to eliminate cross-reaction with common antigen epitopes. Concomitantly, dogs with cutaneous lesions were biopsied to isolate and identify parasites using RAPD. Leishmania were classified by the phenetic method using the Jaccard coefficient of similarity, and grouped by Unweighted Pair-Group Method using an Arithmetic Average (UPGMA). A total of 410 dogs were studied. In R1 (Vale da Ribeira) 159 dogs were evaluated of which 10 had anti-Leishmania antibody. In R2 (Central Paraná), 39 animals were examined of which 8 were seropositive. In R3 (the North) 212 dogs were evaluated of which 39 animals were seropositive. Thirteen dogs had cutaneous lesions and the parasites were isolated from a dog with mucocutaneous lesion in R1, two animals with simple skin lesions in R2 and 10 dogs with multiple lesions in R3. The identification of the parasite by molecular methods showed it to be L. (Viannia) braziliensis. Based on this information, the role of domestic dogs in Leishmania infection of cutaneous leishmaniasis in Paraná is discussed.     

The significance of multiple mating in the social wasp Vespula maculifrons

The evolution of the complex societies displayed by social insects depended partly on high relatedness among interacting group members. Therefore, behaviors that depress group relatedness, such as multiple mating by reproductive females (polyandry), are unexpected in social insects. Nevertheless, the queens of several social insect species mate multiply, suggesting that polyandry provides some benefits that counteract the costs. However, few studies have obtained evidence for links between rates of polyandry and fitness in naturally occurring social insect populations. We investigated if polyandry was beneficial in the social wasp Vespula maculifrons. We used genetic markers to estimate queen mate number in V. maculifrons colonies and assessed colony fitness by counting the number of cells that colonies produced. Our results indicated that queen mate number was directly, strongly, and significantly correlated with the number of queen cells produced by colonies. Because V. maculifrons queens are necessarily reared in queen cells, our results demonstrate that high levels of polyandry are associated with colonies capable of producing many new queens. These data are consistent with the explanation that polyandry is adaptive in V. maculifrons because it provides a fitness advantage to queens. Our research may provide a rare example of an association between polyandry and fitness in a natural social insect population and help explain why queens in this taxon mate multiply.