Low-density lipoprotein promotes microvascular thrombosis by enhancing von Willebrand factor self-association.

von Willebrand factor (VWF) mediates primary hemostasis and thrombosis in response to hydrodynamic forces. We previously showed that high shear promoted self-association of VWF into hyperadhesive strands, which can be attenuated by high-density lipoprotein (HDL) and apolipoprotein A-I. In this study, we show that low-density lipoprotein (LDL) binds VWF under shear and enhances self-association. Vortexing VWF in tubes resulted in its loss from the solution and deposition onto tube surfaces, which was prevented by HDL. At a stabilizing HDL concentration of 1.2 mg/mL, increasing concentrations of LDL progressively increased VWF loss, the effect correlating with the LDL-to-HDL ratio and not the absolute concentration of the lipoproteins. Similarly, HDL diminished deposition of VWF in a post-in-channel microfluidic device, whereas LDL increased both the rate and extent of strand deposition, with both purified VWF and plasma. Hypercholesterolemic human plasma also displayed accelerated VWF accumulation in the microfluidic device. The initial rate of accumulation correlated linearly with the LDL-to-HDL ratio. In Adamts13-/- and Adamts13-/-LDLR-/- mice, high LDL levels enhanced VWF and platelet adhesion to the myocardial microvasculature, reducing cardiac perfusion, impairing systolic function, and producing early signs of cardiomyopathy. In wild-type mice, high plasma LDL concentrations also increased the size and persistence of VWF-platelet thrombi in ionophore-treated mesenteric microvessels, exceeding the accumulation seen in similarly treated ADAMTS13-deficient mice that did not receive LDL infusion. We propose that targeting the interaction of VWF with itself and with LDL may improve the course of thrombotic microangiopathies, atherosclerosis, and other disorders with defective microvascular circulation.

Quantitative analysis of eco-economic benefits of reclaimed water for controlling urban dust.

With the acceleration of urbanization and industrialization, urban air pollution, especially dust pollution, has become a global problem. The traditional method to control dust problem is to spray roads with tap water. However, this method will inevitably lead to a huge waste of water resources. Using reclaimed water instead of tap water for dust control can not only achieve the same effect of reducing dust and haze, but also save water resources. In addition, the saved tap water can be used for production and life, thereby obtaining additional benefits. In order to quantify the eco-economic benefits of sprinkling water for dust control, a quantitative method was proposed based on the emergy theory of ecological economics. It was used to calculate the cost of different water resources, the cooling and humidifying benefit, the dust control and haze reduction benefit, and the other use benefit. Taking the 2017 data of Zhengzhou as an example, the results indicated that the cost of using reclaimed water to control dust was reduced by 54%. The total benefit of using reclaimed water was about 1.30-1.80 times that of using tap water, and the net benefit was 4.65-7.17 times. Therefore, the use of reclaimed water instead of tap water for road dust control has advantages of low cost and high eco-economic benefits. The method proposed in this paper can provide quantitative basis for the popularization of using reclaimed water to dust control in cities with serious air pollution and water resource shortage.

Quantitative assessment of ecological compensation for groundwater overexploitation based on emergy theory.

Ecological compensation is an important economic tool for the control and management of ecological and environmental problems. The accurate assessment of the amount of compensation is the key factor in the implementation of ecological compensation schemes. However, due to the complex and various ecological and environmental problems caused by groundwater overexploitation, there is no scientific quantitative method. Emergy theory is a new method to quantitatively study the relationship between environmental resources and social economy. Based on the literature review of ecological compensation for groundwater overexploitation, this paper puts forward a new evaluation method for using emergy loss as groundwater overexploitation ecological compensation. The emergy system diagram of environmental problems caused by overexploitation of groundwater is constructed. And the calculation methods of the emergy loss of eco-environmental problems caused by groundwater overexploitation, such as land subsidence (collapse), salt water intrusion, surface runoff reduction, vegetation deterioration, and groundwater pollution, were presented, respectively. Taking Zhengzhou as an example, the total amount of ecological compensation for groundwater overexploitation in 2014 was equivalent to 853 × 106 US dollars. The largest loss of land subsidence is 816 × 106 US dollars, which accounts for 95.64% of the total loss. It can be seen that land subsidence is the most serious in the eco-environmental problems caused by overexploitation of groundwater in Zhengzhou.

Von Willebrand Factor Adhesive Activity and ADAMTS13 Protease Activity in HIV-1-Infected Men.

Background: Endothelial activation caused by HIV-1 infection leads to release of von Willebrand factor (VWF), which enters the circulation or attaches to vessel walls and self-assembles into strings and fibers, enabling platelet adhesion; this adhesive activity is regulated by the VWF-cleaving protease ADAMTS13. Our objective was to assess VWF adhesive activity and ADAMTS13 protease activity in HIV-1 infection. Methods: We measured levels of VWF antigen, VWF activation factor (a measure of adhesive activity), ADAMTS13 antigen, ADAMTS13 activity, and apolipoprotein A1 (which interferes with VWF self-association) in serum samples from HIV-1-infected men whose infections were acute (n=10), chronic untreated (n=10), or chronic treated (n=10), compared to uninfected controls (n=10). Means across groups were compared using analysis of variance with contrasts, and Pearson correlations were calculated. Results: Plasma viral load was positively correlated with VWF adhesive activity, which was elevated in acute relative to chronic treated HIV-1 infection. ADAMTS13 antigen and activity were both positively correlated with plasma viral load, and ADAMTS13 activity was significantly higher in men with acute HIV infection than in uninfected controls, and in both acute and chronic untreated HIV infection relative to chronic treated infection. Conclusion: These findings suggest that even in the setting of increased ADAMTS13 protease activity, VWF in HIV-1 infection is hyperadhesive, which may favor development of microvascular and arterial thromboses and thereby contribute to increased cardiovascular risk in HIV-1-infected individuals.

Analysis of groundwater variation in the Jinci Spring area, Shanxi Province (China), under the influence of human activity.

Jinci Spring is one of the most famous karst spring in Northern China and is the main drinking and irrigation water source of Taiyuan city, Shanxi Province. It has special significance in terms of humanities and water resources. Because of the continuous over-exploitation of groundwater, the water level of groundwater has been decreased significantly. The flow of Jinci Spring has decreased year by year since the 1970s, and in 1994 it cut off completely. The flow cutoff has brought huge losses to the local economy and negatively impacted the ecological environment. From the hydrogeological conditions of Jinci Spring area to study, the dynamic change in karst groundwater level will provide a basis for the protection and rational exploitation of karst water in the spring area. The previous research on Jinci Spring area was limited to the spring flow and water level changes, lack of analysis to the overall spring area and the decay process of spring flow. This paper takes the overall spring area as research object. According to groundwater level data over 1961-2012, the period is divided into two phases of decline and recovery. Human activities in the region and corresponding groundwater evolution at various stages were systematically analyzed, and the reservoir capacity was calculated for each period of decline. The results show that the average reservoir capacity decreased from 920 (1954-1960) to 116 (1961-1977) to 31 (1978-1994) million m3. The important effect of "Water Resumption" project on groundwater in the area during the remediation period was also analyzed. In the light of observed increases in groundwater level during this stage, the project has a notably positive effect.

High-density lipoprotein modulates thrombosis by preventing von Willebrand factor self-association and subsequent platelet adhesion.

The ability of von Willebrand factor (VWF) to initiate platelet adhesion depends on the number of monomers in individual VWF multimers and on the self-association of individual VWF multimers into larger structures. VWF self-association is accelerated by shear stress. We observed that VWF self-association occurs during adsorption of VWF onto surfaces, assembly of secreted VWF into hyperadhesive VWF strings on the endothelial surface, and incorporation of fluid-phase VWF into VWF fibers. VWF adsorption under static conditions increased with increased VWF purity and was prevented by a component of plasma. We identified that component as high-density lipoprotein (HDL) and its major apolipoprotein ApoA-I. HDL and ApoA-I also prevented VWF on the endothelium from self-associating into longer strands and inhibited the attachment of fluid-phase VWF onto vessel wall strands. Platelet adhesion to VWF fibers was reduced in proportion to the reduction in self-associated VWF. In a mouse model of thrombotic microangiopathy, HDL also largely prevented the thrombocytopenia induced by injection of high doses of human VWF. Finally, a potential role for ApoA-I in microvascular occlusion associated with thrombotic thrombocytopenic purpura and sepsis was revealed by the inverse relationship between the concentration of ApoA-I and that of hyperadhesive VWF. These results suggest that interference with VWF self-association would be a new approach to treating thrombotic disorders.

Hypochlorous acid generated by neutrophils inactivates ADAMTS13: an oxidative mechanism for regulating ADAMTS13 proteolytic activity during inflammation.

ADAMTS13 is a plasma metalloproteinase that cleaves large multimeric forms of von Willebrand factor (VWF) to smaller, less adhesive forms. ADAMTS13 activity is reduced in systemic inflammatory syndromes, but the cause is unknown. Here, we examined whether neutrophil-derived oxidants can regulate ADAMTS13 activity. We exposed ADAMTS13 to hypochlorous acid (HOCl), produced by a myeloperoxidase-H2O2-Cl(-) system, and determined its residual proteolytic activity using both a VWF A2 peptide substrate and multimeric plasma VWF. Treatment with 25 nm myeloperoxidase plus 50 µm H2O2 reduced ADAMTS13 activity by >85%. Using mass spectrometry, we demonstrated that Met(249), Met(331), and Met(496) in important functional domains of ADAMTS13 were oxidized to methionine sulfoxide in an HOCl concentration-dependent manner. The loss of enzyme activity correlated with the extent of oxidation of these residues. These Met residues were also oxidized in ADAMTS13 exposed to activated human neutrophils, accompanied by reduced enzyme activity. ADAMTS13 treated with either neutrophil elastase or plasmin was inhibited to a lesser extent, especially in the presence of plasma. These observations suggest that oxidation could be an important mechanism for ADAMTS13 inactivation during inflammation and contribute to the prothrombotic tendency associated with inflammation.

Structural and functional analysis of the pro-domain of human cathelicidin, LL-37.

Cathelicidins form a family of small host defense peptides distinct from another class of cationic antimicrobial peptides, the defensins. They are expressed as large precursor molecules with a highly conserved pro-domain known as the cathelin-like domain (CLD). CLDs have high degrees of sequence homology to cathelin, a protein isolated from pig leukocytes and belonging to the cystatin family of cysteine protease inhibitors. In this report, we describe for the first time the X-ray crystal structure of the human CLD (hCLD) of the sole human cathelicidin, LL-37. The structure of the hCLD, determined at 1.93 Å resolution, shows the cystatin-like fold and is highly similar to the structure of the CLD of the pig cathelicidin, protegrin-3. We assayed the in vitro antibacterial activities of the hCLD, LL-37, and the precursor form, pro-cathelicidin (also known as hCAP18), and we found that the unprocessed protein inhibited the growth of Gram-negative bacteria with efficiencies comparable to that of the mature peptide, LL-37. In addition, the antibacterial activity of LL-37 was not inhibited by the hCLD intermolecularly, because exogenously added hCLD had no effect on the bactericidal activity of the mature peptide. The hCLD itself lacked antimicrobial function and did not inhibit the cysteine protease, cathepsin L. Our results contrast with previous reports of hCLD activity. A comparative structural analysis between the hCLD and the cysteine protease inhibitor stefin A showed why the hCLD is unable to function as an inhibitor of cysteine proteases. In this respect, the cystatin scaffold represents an ancestral structural platform from which proteins evolved divergently, with some losing inhibitory functions.

Simultaneous exposure of sites in von Willebrand factor for glycoprotein Ib binding and ADAMTS13 cleavage: studies with ristocetin.

OBJECTIVE: Platelet-bound von Willebrand factor (VWF) was recently demonstrated to be a better substrate for ADAMTS13, suggesting that 1 conformational change exposes both the glycoprotein Ibα binding site in the A1 domain and the ADAMTS13 cleavage site in the A2 domain. Because ristocetin induces VWF to bind glycoprotein Ibα in the absence of shear stress, we evaluated whether it could also enhance ADAMTS13 proteolysis of VWF. METHODS AND RESULTS: We used several VWF sources: plasma, purified plasma VWF, recombinant VWF fragments encompassing A1A2A3, A1A2, and 2 A2 domains, 1 containing a ristocetin-binding site (Asp1459-His1472) and the other lacking it. Ristocetin accelerated ADAMTS13 cleavage of multimeric VWF and of each of the recombinant VWF fragments except for the A2 domain lacking the ristocetin-binding site. We also examined the effect of ristocetin on the conformation of the A2 domain by assessing its effect on the susceptibility of Met1606 at the ADAMTS13 cleavage site to be oxidized by hypochlorous acid. Ristocetin markedly enhanced oxidation of Met1606 and Met1521 of the A2 domain. CONCLUSIONS: These data indicate that exposure of the sites for glycoprotein Ibα and ADAMTS13 are coupled, explaining why platelet-bound VWF is a better ADAMTS13 substrate and why enhanced proteolysis is often observed in type 2B von Willebrand disease.

Research on emergy evaluation method of ecological water use efficiency based on comprehensive benefits.

Scientifically evaluating ecological water use efficiency (EWUE) is an effective means to regulate the level of ecological water use in a country or a region. It is also a basic work to achieve high-efficiency use of ecological water under the current situation of water shortage. However, there were few researches on EWUE, and existing studies only focus on eco-environmental benefits generated by ecological water, without considering its impact on economy and society. An emergy evaluation method for EWUE based on comprehensive benefits was proposed in this paper innovatively. Considering the impact of ecological water use on society, economy, and eco-environment, the concept of EWUE could be defined. Then, comprehensive benefits of ecological water use (CBEW) were quantified by emergy method, and EWUE was evaluated by the comprehensive benefits of unit ecological water use. Taking Zhengzhou City as an example for calculation, from 2011 to 2020, CBEW increased from 5.20 × 1019 sej to 6.72 × 1020 sej, showing an overall upward trend, and EWUE rose from 2.71 × 1011 sej/m3 (1.27ï¿¥/m3) to 1.32 × 1012 sej/m3 (8.10ï¿¥/m3) with fluctuation. It showed that Zhengzhou City has paid enough attention to the allocation of ecological water and EWUE at a high level. The method proposed in this paper provides a new idea to evaluate EWUE scientifically, and the results can provide guidance to allocate ecological water resources to achieve sustainable development.

Oxidative modification of von Willebrand factor by neutrophil oxidants inhibits its cleavage by ADAMTS13.

Elevated plasma von Willebrand factor (VWF) and low ADAMTS13 activity have been reported in several inflammatory states, including sepsis and acute respiratory distress syndrome. One hallmark of inflammation is neutrophil activation and production of reactive oxygen species, including superoxide radical, hydrogen peroxide, and hypochlorous acid (HOCl). HOCl is produced from hydrogen peroxide and chloride ions through the action of myeloperoxidase. HOCl can oxidize methionine to methionine sulfoxide and tyrosine to chlorotyrosine. This is of interest because the ADAMTS13 cleavage site in VWF, the Tyr(1605)-Met(1606) peptide bond, contains both oxidation-prone residues. We hypothesized that HOCl would oxidize either or both of these residues and possibly inhibit ADAMTS13-mediated cleavage. We therefore treated ADAMTS13 substrates with HOCl and examined their oxidative modification by mass spectrometry. Met(1606) was oxidized to the sulfoxide in a concentration-dependent manner, with complete oxidation at 75muM HOCl, whereas only a miniscule percentage of Tyr(1605) was converted to chlorotyrosine. The oxidized substrates were cleaved much more slowly by ADAMTS13 than the nonoxidized substrates. A similar result was obtained with multimeric VWF. Taken together, these findings indicate that reactive oxygen species released by activated neutrophils have a prothrombotic effect, mediated in part by inhibition of VWF cleavage by ADAMTS13.

Assessment of eco-economic effects of urban water system connectivity project.

As one of the large ecological infrastructures, the urban water system connectivity (UWSC) project is an important part of urban ecosystem construction. It is helpful for the scientific planning and construction of the project to systematically evaluate the effects. However, due to the complex and various effects of UWSC project, there is no complete effect system and quantitative method. Against this backdrop, the composition and mechanism of positive and negative effects of ecological economics of UWSC project were deeply analyzed to improve the composition system of eco-economic effects in this study. At the same time, the emergy theory was used to put forward the quantification method of eco-economic effect system. Taking the UWSC project in Xuchang as an example, its ecological, social, and economic effects were evaluated. The result showed that the average eco-economic effect of the project is 49.97 million dollars/year. Economic effect and ecological effect are significant, accounting for 82.49% and 15.89% of total effect, respectively. This study can provide reference for comprehensive and unified assessment of eco-economic effects of UWSC project.

Evaluation of water quality and its driving forces in the Shaying River Basin with the grey relational analysis based on combination weighting.

The water quality of the Shaying River Basin and even the entire Huai River Basin has been widely concerned. Based on the water quality data acquired in flood and non-flood seasons from 2012 to 2016, the Shaying River Basin was selected as the research object. First, the principal component analysis method was used to identify the main pollution indices. Then, grey relational analysis combined with an analytic hierarchy process and entropy weight method was used to evaluate the water quality of the upper, middle, and lower reaches of the Shaying River Basin, while the single factor evaluation method was used for comparative analysis. Finally, the driving forces of water quality were analyzed and discussed from natural and human aspects. The results show that the main pollutants in the Shaying River Basin are total nitrogen, total phosphorus, and ammonium nitrogen. While the basin is seriously polluted by nitrogen and phosphorus, the spatial and temporal distribution of the pollution varies, although the overall trend toward improving water quality conditions is significant. The midstream region had the poorest water quality, which fluctuated between Classes III and V. The downstream region had generally good water quality, which could be ranked as Class III most of the time. And the upstream region had the best water quality with well-developed ecological conditions; all the water samples were ranked as Class I or II. The water quality improves significantly during the flood season when compared with that in the non-flood season. Seasonal climate variation, non-point source pollution emissions, the release of water from sluices and dams, and water resource management activities are the main reasons for the variations in water quality across the Shaying River Basin.

Cloning and Sequence Analyzing the Gene of Trichosanthes Trypsin Inhibitor.

A DNA probe for Trichosanthes trypsin inhibitor was amplified from the cDNA library of Trichosanthes kirilowii by polymerase chain reaction. Using this probe to screen the cDNA library, a full-length cDNA sequence of the inhibitor was obtained. The open reading frame encodes a Pre-Pro-inhibitor with Pre-, Pro, and mature peptides of 24, 14, and 27 residues, respectively. The deduced amino acid sequence of the mature peptide is consistent with that determined by the primary structure analysis of the protein.

cDNA Sequences of Two Anti-mammals Neurotoxins from Scorpion Buthus martensii Karsch.

The total mRNA was prepared and purified from the venom gland of scorpion Buthus martensii Karsch (BmK). The cDNA library of this venom gland was then constructed using reverse transcription. Two cDNAs encoding the anti-mammalian neurotoxins, named as BmK M1 and BmK M9, were selectively amplified by PCR and sequenced. Both have open reading frames of 252 bp encoding 84 residues of the toxin precursors, including a 19 amino acid signal peptide, the mature toxin with 64 residues, and an additional Arg tail at the carboxyl-terminus which was removed during post-protein processing. The deduced amino acid sequence of BmK M1 was completely identical with the known sequence of the natural toxin BmK M4, whereas the other BmK M9 shared a high homology with the natural toxin BmK M4, differing only in two residues. The cDNA of BmK M1 showed 87.2% homology with that of the African scorpion Lqh alphaIT.

Structural basis of type 2A von Willebrand disease investigated by molecular dynamics simulations and experiments.

The hemostatic function of von Willebrand factor is downregulated by the metalloprotease ADAMTS13, which cleaves at a unique site normally buried in the A2 domain. Exposure of the proteolytic site is induced in the wild-type by shear stress as von Willebrand factor circulates in blood. Mutations in the A2 domain, which increase its susceptibility to cleavage, cause type 2A von Willebrand disease. In this study, molecular dynamics simulations suggest that the A2 domain unfolds under tensile force progressively through a series of steps. The simulation results also indicated that three type 2A mutations in the C-terminal half of the A2 domain, L1657I, I1628T and E1638K, destabilize the native state fold of the protein. Furthermore, all three type 2A mutations lowered in silico the tensile force necessary to undock the C-terminal helix α6 from the rest of the A2 domain, the first event in the unfolding pathway. The mutations F1520A, I1651A and A1661G were also predicted by simulations to destabilize the A2 domain and facilitate exposure of the cleavage site. Recombinant A2 domain proteins were expressed and cleavage assays were performed with the wild-type and single-point mutants. All three type 2A and two of the three predicted mutations exhibited increased rate of cleavage by ADAMTS13. These results confirm that destabilization of the helix α6 in the A2 domain facilitates exposure of the cleavage site and increases the rate of cleavage by ADAMTS13.