Alteration of Phosphoinositide Degradation by Cytosolic and Membrane-Bound Phospholipases after Forebrain Ischemia - Reperfussion in Gerbil: Effects of Amyloid Beta Peptide

The reperfusion of previously ischemic brain is associated with exacerbation of cellular injury. Reperfusion occasionally potentates release of intracellular enzymes, influx of Ca²⁺, breakdown of membrane phospholipids, accumulation of amyloid precursor protein or amyloid -(like) proteins, and apolipoprotein E. In this study, the effect of reperfusion injury on the activity of cerebral cortex enzymes acting on phosphatidyl [³H] inositol (PI) and [^l4C-arachidonoyl] PI was investigated. Moreover the effect of amyloid 25–35 on PI degradation by phospholipase(s) of normoxic brain and subjected to ichemia-reperfussion injury was determined. Brain ischemia in gerbils (Meriones unguiculatus) was induced by ligation of both common carotid arteries for 5 min and then brains were perfused for 15 min, 2 h and 7 days. Statistically significant activation of enzyme(s) involved in phosphatidylinositol degradation in gerbils subjected to ischemia-reperfusion injury was observed. Nearly all gerbils showed a higher activity of cytosolic PI phos-pholipase C (PLC) at 15 min after ischemia. Concomitantly, the significant enhancement of the level of DAG and AA radioactivity at this short reperfusion time confirmed the active PI degradation by phospholipase(s) in cerebral cortex and hippocampus. After a prolonged reperfusion time of 7 days after ischemia, both cytosolic and membrane-bound forms of PI-PLC were activated. The question arises if alteration of membranes by the degradation of phospholipids occurring after an ischemic episode potentates the effect of A on membrane-bound enzymes. A neuro-toxic fragment of amyloid, A 25–35, incubated in the presence of endogenous Ca²⁺, increased significantly the PI-PLC activity of normoxic brain. In its non-aggregated form, A 25–35 activates PI-PLC but in the aggregated form the enzymatic activity decreased. Thus, A 25–35 exerts a similar effect on the membrane-bound PI-PLC from normoxic brain or subjected to ischemia reperfussion injury. We conclude that the degradation of phosphatidylinositol by cytosolic phosphoinositide-phospholipase C may contribute to the pathophysiology of delayed neuronal death following cerebral ischemia. Thus, a specific inhibitor of this enzyme(s) may offer therapeutic strategies to protect the brain from damage triggered by ischemia. Ischemia-reperfusion injury had no effect on A-evoked alterations of synaptic plasma membrane-bound PI-PLC.     

连续性血浆滤过吸附对严重感染合并MODS患者炎症因子水平的影响

目的:探讨连续性血浆滤过吸附对严重感染合并多脏器功能障碍综合征(MODS)患者炎症因子水平的影响。方法:研究对象选取我院2013年3月-2016年3月收治严重感染合并MODS患者共160例,以随机数字表法分为对照组(80例)和观察组(80例),患者均先给予高容量血液滤过(HVHF)治疗,观察组加用连续性血浆滤过吸附辅助治疗,比较两组患者治疗前后平均动脉压(MAP)、心率(HR)、血氧饱和度(SpO_2)、碳酸氢根(HCO_3~-)、肌酐(Scr)、总胆红素(TBi L)、急性生理与慢性健康II(APACHE Ⅱ)评分、肿瘤坏死因子α(TNF-α)、超敏C反应蛋白(hs-CRP)及白介素6(IL-6)水平。结果:两组治疗后MAP、Scr、TBi L、TNF-α、IL-6、hs-CRP及APACHE Ⅱ评分均有所降低,而HR、SpO_2、HCO_3~-水平均上升,且观察组以上各指标改善更明显,差异均有统计学意义(P0.05)。结论:连续性血浆滤过吸附辅助治疗严重感染合并MODS可有效稳定生命体征,改善生化指标,延缓病情进展,并有助于降低机体炎症反应水平。     

Interaction of Concanavalin A and a Divalent Derivative with Lymphocytes and Reconstituted Lymphocyte Membrane Glycoproteins

Both concanavalin A (con A) and its divalent derivative, succinyl-concanavalin A (S-con A) are mitogenic for porcine lymph node lymphocytes. We have compared the binding of these two lectins to intact porcine lymphocytes and phospholipid vesicles containing reconstituted lymphocyte membrane glycoproteins. Both con A and S-con A showed high- and low-affinity binding to intact cells, as indicated by LIGAND analysis of Scatchard plots of binding data. Despite the apparently identical saccharide specificities of the two lectins, high-affinity binding sites for S-con A were only one-third as numerous as high-affinity sites for the parent lectin. Large numbers of low-affinity binding sites existed for con A, while many fewer were present for S-con A. It is suggested that these sites result from hydrophobic association. Con A bound to lymphocytes in a positively cooperative fashion, while S-con A showed non-cooperative behavior. Lectin binding to large unilamellar phospholipid vesicles containing reconstituted lymphocyte membrane glycoproteins was measured using a rapid filtration assay, and was linear with the glycoprotein content of the vesicles. Almost all of the outward-facing glycoprotein was functional in terms of lectin binding. Reconstituted glycoproteins showed only a single class of high-affinity binding sites for both con A and S-con A, with association constants similar to those measured for intact cells. Con A, but not S-con A, showed positively cooperative binding to reconstituted vesicles. Cooperativity was observed in both gel phase and liquid crystalline phase lipid, and was thus not dependent on long-range lateral rearrangement of glycoprotein receptors. Results suggested that con A induces a microre-distribution of receptors on the lymphocyte membrane surface, leading to the exposure of glycoproteins that were previously inaccessible to the lectin. S-Con A does not cause glycoprotein redistribution, and a large fraction of the receptors remain cryptic.     

Kinin-mediated inflammation in neurodegenerative disorders

The mediatory role of kinins in both acute and chronic inflammation within nervous tissues has been widely described. Bradykinin, the major representative of these bioactive peptides, is one of a few mediators of inflammation that directly stimulates afferent nerves due to the broad expression of specific kinin receptors in cell types in these tissues. Moreover, kinins may be delivered to a site of injury not only after their production at the endothelium surface but also following their local production through the enzymatic degradation of kininogens at the surface of nerve cells. A strong correlation between inflammatory processes and neurodegeneration has been established. The activation of nerve cells, particularly microglia, in response to injury, trauma or infection initiates a number of reactions in the neuronal neighborhood that can lead to cell death after the prolonged action of inflammatory substances. In recent years, there has been a growing interest in the effects of kinins on neuronal destruction. In these studies, the overexpression of proteins involved in kinin generation or of kinin receptors has been observed in several neurologic disorders including neurodegenerative diseases such Alzheimer's disease and multiple sclerosis as well as disorders associated with a deficiency in cell communication such as epilepsy. This review is focused on recent findings that provide reliable evidence of the mediatory role of kinins in the inflammatory responses associated with different neurological disorders. A deeper understanding of the role of kinins in neurodegenerative diseases is likely to promote the future development of new therapeutic strategies for the control of these disorders. An example of this could be the prospective use of kinin receptor antagonists.     

Biophysical characterization of the interaction of bovine seminal plasma protein PDC-109 with phospholipid vesicles

PDC-109 is the major protein of bovine seminal plasma. It binds to the bovine sperm surface at ejaculation and modulates sperm capacitation. PDC-109 displays phosphorylcholine- and heparin-binding activities which are thought to account for its sperm surface coating and glycosaminoglycan-induced sperm capacitating activities, respectively. We have characterized the interaction of isolated PDC-109 with membranes of phospholipid vesicles using a biophysical approach. Our results show that PDC-109 interacts not only with the solvent-exposed phosphorylcholine head group but also with the hydrophobic core of liposomes. Binding of PDC-109 to membranes is a very rapid, biphasic process with half times of less than one second. Maximal binding of PDC-109 to small unilamellar vesicles was achieved with a stoichiometric ratio of 10–11 phosphatidylcholine molecules/PDC-109 molecule. Incorporation of phosphatidylethanolamine or phosphatidylserine into phosphatidylcholine vesicles reduced the binding of PDC-109, suggesting that both the density of phosphorylcholine groups and the surface charge determine the interaction of the seminal plasma protein with the surface of the membrane. Electron spin resonance measurements showed that binding of PDC-109 to phosphatidylcholine vesicles caused a rigidification of the membrane. The relevance of the data for describing the role of PDC-109 in the modulation of sperm capacitation is discussed. Received: 16 June 1997 / Accepted: 10 September 1997     

A Combination of Proteomic Approaches Identifies A Panel of Circulating Extracellular Vesicle Proteins Related to the Risk of Suffering Cardiovascular Disease in Obese Patients

Plasma‐derived extracellular vesicles (EVs) have been extensively described as putative biomarkers in different diseases. Interestingly, increased levels of EVs subpopulations are well known to associate with obesity. The goal of this study is to identify EVs‐derived biomarkers in plasma from obese patients in order to predict the development of pathological events associated with obesity. Samples are obtained from 22 obese patients and their lean‐matched controls are divided into two cohorts: one for a 2D fluorescence difference gel electrophoresis (2D‐DIGE)‐based study, and the other one for a label free LC–MS/MS‐based approach. EVs are isolated following a serial ultracentrifugation protocol. Twenty‐two and 23 differentially regulated features are detected from 2D‐DIGE and label free LC–MS/MS, respectively; most of them involve in the coagulation and complement cascades. Remarkably, there is an upregulation of complement C4, complement C3, and fibrinogen in obese patients following both approaches, the latter two also validated by 2D‐western‐blotting in an independent cohort. These results correlate with a proinflammatory and prothrombotic state of those individuals. On the other hand, a downregulation of adiponectin leading to an increased risk of suffering cardiovascular diseases has been shown. The results suggest the relevance of plasma‐derived‐EVs proteins as a source of potential biomarkers for the development of atherothrombotic events in obesity.     

A SXRF method for determining the relative concentration of trace elements in plasma protein affected by cisplatin

This article presents an analytical method for the determination of the relative concentrations of trace elements in plasma protein by gel chromatography combined with SXRF (synchrotron radiation X-ray fluorescence). The fraction of plasma protein of male Kunming mice (body weight 24.2±0.3 g), treated with a cisplatin ip injection at a dose of 10 mg/kg, was obtained by the separation of a Sephadex G-50 column (buffered with ammonium acetate, pH 5.7). The SXRF experiments were performed at the Beijing Electron Positron Collider synchrotron radiation facility. The elements (Pt, S, Ca, Fe, Ni, Cu, Zn, Se, Br, and Sr) in the fraction of the plasma proteins (>22 kDa) were assayed using highly sensitive SXRF. The relative concentrations of elements were calculated by a normalization of Compton scattering intensity around 22 keV, after the normalization for the collection time of the X-ray spectrum and the counting of the ion chamber, and subtracting the contribution of the polycarbonate film for the supporting sample. The determination could prove that the element Pt in plasma was bound with macromolecular protein. Cu and S were present in the fraction of the protein in mice treated with cisplatin increase, and their ratios of treated/control were 1.66±0.06 and 1.78±0.33, respectively; Zn decreased to a ratio of 0.78±0.09. Our results are in agreement with others that cisplatin exposure leads to a marked loss of kidney copper and a moderate rise in kidney zinc. However, this article primarily describes one of the analytical methods used; it does not emphasize the results of the effect of cisplatin on trace elements in plasma protein.     

Synergistic effect of conjugated linolenic acid isomers against induced oxidative stress,inflammation and erythrocyte membrane disintegrity in rat model

Background

α-Eleostearic acid and punicic acid, two typical conjugated linolenic acid (CLnA) isomers present in bitter gourd and snake gourd oil respectively, exhibit contrasting cis-trans configuration which made them biologically important.

Methods

Rats were divided into six groups. Group 1 was control and group 2 was treated control. Rats in the groups 3 and 4 were treated with mixture of α-eleostearic acid and punicic acid (1:1) (0.5% and 1.0% respectively) while rats in the groups 5 and 6 were treated with 0.5% of α-eleostearic acid and 0.5% of punicic acid respectively along with sodium arsenite by oral gavage once per day.

Results

Results showed that increase in nitric oxide synthase (NOS) activity, inflammatory markers expression, platelet aggregation, lipid peroxidation, protein oxidation, DNA damage and altered expression of liver X receptor-α (LXR-α) after arsenite treatment were restored with the supplementation of oils containing CLnA isomers. Altered activities of different antioxidant enzymes such as superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase and ferric reducing ability of plasma (FRAP) also restored after oil supplementation. Altered morphology and fluidity of erythrocyte membrane studied by atomic force and scanning electron microscopy, after stress induction were significantly improved due to amelioration in cholesterol/phospholipid ratio and fatty acid profile of membrane. Oils treatment also improved morphology of liver and fatty acid composition of hepatic lipid.

Conclusions

Overall two isomers showed synergistic antioxidant and anti-inflammatory effect against induced perturbations and membrane disintegrity.

General significance

Synergistic antioxidant and anti-inflammatory role of these CLnA isomers were established by this study.     

Laser-induced Breakdown Spectroscopy: A New Approach for Nanoparticle's Mapping and Quantification in Organ Tissue

Emission spectroscopy of laser-induced plasma was applied to elemental analysis of biological samples. Laser-induced breakdown spectroscopy (LIBS) performed on thin sections of rodent tissues: kidneys and tumor, allows the detection of inorganic elements such as (i) Na, Ca, Cu, Mg, P, and Fe, naturally present in the body and (ii) Si and Gd, detected after the injection of gadolinium-based nanoparticles. The animals were euthanized 1 to 24 hr after intravenous injection of particles. A two-dimensional scan of the sample, performed using a motorized micrometric 3D-stage, allowed the infrared laser beam exploring the surface with a lateral resolution less than 100 μm. Quantitative chemical images of Gd element inside the organ were obtained with sub-mM sensitivity. LIBS offers a simple and robust method to study the distribution of inorganic materials without any specific labeling. Moreover, the compatibility of the setup with standard optical microscopy emphasizes its potential to provide multiple images of the same biological tissue with different types of response: elemental, molecular, or cellular.     

Effects of Salinity Stress on the Levels of Covalently and Noncovalently Conjugated Polyamines in Plasma Membrane and Tonoplast Isolated from Barley Seedlings

When the 7-d old barley ( Hordeum vulgare L. ) seedlings were treated with different concentrations of NaC1 for 3 d, the levels of the noncovalemly conjugated polyamines (PAs) in the plasma membrane and tonoplast vesicles and the covalently conjugated PAs in the membrane proteins were promoted by NaC1 of low concentrations and suppressed by NaC1 of high concentrations. Among the noncovalenfly conjugated PAs in the vesicles, spennidine (Spd) level was the most abundant, while putrescine (Put) content was predominant among the covalenfly conjugated PAs, accounted for 40%– 70%, 35% – 60%, respectively. In addition, the TLC (thin-layer chromatography) profiles of the benzoylated PAs presented an unknown polyamine with Rf = 0.92 (X0.92), which conjugated covalently and noncovalently in root tonoplast and its content changed as well as Spd with NaC1 treatment. The total PA contents in the roots were higher than that in the leaves, and the types and contents of covalenfly and noncovalently conjugated PAs in the tonoplast were higher than those in the plasma membrane. The results showed that the above two PAs associated with the membrane might be essential in salt adaption of cells and the maintenance of membrane function.