Extracellular HIV-1 Nef increases migration of monocytes

Infiltration of human immunodeficiency virus type 1 (HIV-1)-infected and uninfected monocytes/macrophages in organs and tissues is a general phenomenon observed in progression of acquired immunodeficiency syndrome (AIDS). HIV-1 protein Nef is considered as a progression factor in AIDS, and is released from HIV-1-infected cells. Here, we show that extracellular Nef increases migration of monocytes. This effect is (i) concentration-dependent, (ii) reaches the order of magnitude of that induced by formyl-methyonyl-leucyl-proline (fMLP) or CC chemokine ligand 2 (CCL2)/monocyte chemotactic protein (MCP)-1, (iii) inhibited by anti-Nef monoclonal antibodies as well as by heating, and (iv) depends on a concentration gradient of Nef. Further, Nef does not elicit monocytic THP-1 cells to express chemokines such as CCL2, macrophage inhibitory protein-1alpha (CCL3) and macrophage inhibitory protein-1beta (CCL4). These data suggest that extracellular Nef may contribute to disease progression as well as HIV-1 spreading through affecting migration of monocytes.     

Loop 3 of Short Neurotoxin II is an Additional Interaction Site with Membrane-bound Nicotinic Acetylcholine Receptor as Detected by Solid-state NMR Spectroscopy

The contact area of neurotoxin II from Naja naja oxiana when interacting with the membrane-bound nicotinic acetylcholine receptor from Torpedo californica was determined by solid-state, magic-angle spinning NMR spectroscopy. For this purpose, the carbon signals for more than 90% of the residues of the bound neurotoxin were assigned. Differences between the solution and solid-state chemical shifts of the free and bound form of the toxin are confined to distinct surface regions. Loop II of the short toxin was identified as the main interaction site. In addition, loop III of neurotoxin II shows several strong responses defining an additional interaction site. A comparison with the structures of α-cobratoxin bound to the acetylcholine-binding protein from snail species Lymnaea stagnalis and Aplysia californica, and of α-bungarotoxin bound to an extracellular domain of an α-subunit of the receptor reveals different contact areas for long and short α-neurotoxins.     

Butyrylcholinesterase Predicts Cardiac Mortality in Young Patients with Acute Coronary Syndrome

Background

The incidence of acute coronary syndrome (ACS) in young people (≤65 years) is continuously rising. While prognostic factors in ACS are well-investigated less attention has been paid to their age-dependent prognostic value and their particular relevance in younger patients. The aim of our study was to assess the age-dependent prognostic impact of butyrylcholinesterase (BChE).

Methods

Retrospective cohort study including 624 patients with ACS. Patients were stratified by age into equal groups (n = 208) corresponding to “young patients” (45–64 years), "middle-aged patients” (65–84 years) and “old patients” (85–100 years). Cox regression hazard analysis was used to assess the influence of BChE on survival.

Results

After a mean follow-up time of 4.0 (interquartile range [IQR] 2.0–6.4) years, 154 patients (24.7%) died due to a cardiac cause. In the overall cohort, BChE was indirectly associated with cardiac mortality-free survival (adjusted hazard ratio (HR): 0.70 (95% confidence interval [CI] 0.53–0.93, p = 0.01). The primary-analysis of BChE by age strata showed the strongest effect in the age group 45–64 years with an adjusted HR per 1-SD of 0.28 (95% CI 0.12–0.64, p = 0.003), a weaker association with mortality in middle aged (65–84 years: adjusted HR per 1-SD 0.66 [95% CI: 0.41–1.06], p = 0.087), and no association in older patients (85–100 years: adjusted HR per 1-SD 0.89 [95% CI: 0.58–1.38], p = 0.613).

Conclusion

BChE is a strong predictor for cardiac mortality specifically in younger patients with ACS aged between 45 and 64 years. No significant association of BChE with cardiac-mortality was detected in other age classes.     

Inhibition of mitochondrial fusion by α‐synuclein is rescued by PINK1, Parkin and DJ‐1

Aggregation of α‐synuclein (αS) is involved in the pathogenesis of Parkinson's disease (PD) and a variety of related neurodegenerative disorders. The physiological function of αS is largely unknown. We demonstrate with in vitro vesicle fusion experiments that αS has an inhibitory function on membrane fusion. Upon increased expression in cultured cells and in Caenorhabditis elegans, αS binds to mitochondria and leads to mitochondrial fragmentation. In C. elegans age‐dependent fragmentation of mitochondria is enhanced and shifted to an earlier time point upon expression of exogenous αS. In contrast, siRNA‐mediated downregulation of αS results in elongated mitochondria in cell culture. αS can act independently of mitochondrial fusion and fission proteins in shifting the dynamic morphologic equilibrium of mitochondria towards reduced fusion. Upon cellular fusion, αS prevents fusion of differently labelled mitochondrial populations. Thus, αS inhibits fusion due to its unique membrane interaction. Finally, mitochondrial fragmentation induced by expression of αS is rescued by coexpression of PINK1, parkin or DJ‐1 but not the PD‐associated mutations PINK1 G309D and parkin Δ1–79 or by DJ‐1 C106A.     

L1 is sequentially processed by two differently activated metalloproteases and presenilin/gamma-secretase and regulates neural cell adhesion, cell migration, and neurite outgrowth

The immunoglobulin superfamily recognition molecule L1 plays important functional roles in the developing and adult nervous system. Metalloprotease-mediated cleavage of this adhesion molecule has been shown to stimulate cellular migration and neurite outgrowth. We demonstrate here that L1 cleavage is mediated by two distinct members of the disintegrin and metalloprotease family, ADAM10 and ADAM17. This cleavage is differently regulated and leads to the generation of a membrane bound C-terminal fragment, which is further processed through gamma-secretase activity. Pharmacological approaches with two hydroxamate-based inhibitors with different preferences in blocking ADAM10 and ADAM17, as well as loss of function and gain of function studies in murine embryonic fibroblasts, showed that constitutive shedding of L1 is mediated by ADAM10 while phorbol ester stimulation or cholesterol depletion led to ADAM17-mediated L1 cleavage. In contrast, N-methyl-d-aspartate treatment of primary neurons stimulated ADAM10-mediated L1 shedding. Both proteases were able to affect L1-mediated adhesion and haptotactic migration of neuronal cells. In particular, both proteases were involved in L1-dependent neurite outgrowth of cerebellar neurons. Thus, our data identify ADAM10 and ADAM17 as differentially regulated L1 membrane sheddases, both critically affecting the physiological functions of this adhesion protein.     

5-Fluoro-4-thiouridine phosphoramidite: new synthon for introducing photoaffinity label into oligodeoxynucleotides

The synthesis of phosphoramidite of 5-fluoro-4-thio-2'-O-methyluridine is described. An appropriate set of protecting groups was optimized including the 4-thio function introduced via 4-triazolyl as the 4-(2-cyanoethyl)thio derivative, and the t-butyldimethyl silyl for 2' and 3' hydroxyl protection, enabling efficient synthesis of the phosphoramidite. These protecting groups prevented unwanted side reactions during oligonucleotide synthesis. The utility of the proposed synthetic route was proven by the preparation of several oligonucleotides via automated synthesis. Photochemical experiments confirmed the utility of the synthon.     

Activation of Wnt/β-Catenin Pathway in Monocytes Derived from Chronic Kidney Disease Patients

Patients with chronic kidney disease (CKD) have significantly increased morbidity and mortality resulting from infections and cardiovascular diseases. Since monocytes play an essential role in host immunity, this study was directed to explore the gene expression profile in order to identify differences in activated pathways in monocytes relevant to the pathophysiology of atherosclerosis and increased susceptibility to infections. Monocytes from CKD patients (stages 4 and 5, estimated GFR <20 ml/min/1.73 m²) and healthy donors were collected from peripheral blood. Microarray gene expression profile was performed and data were interpreted by GeneSpring software and by PANTHER tool. Western blot was done to validate the pathway members. The results demonstrated that 600 and 272 genes were differentially up- and down regulated respectively in the patient group. Pathways involved in the inflammatory response were highly expressed and the Wnt/β-catenin signaling pathway was the most significant pathway expressed in the patient group. Since this pathway has been attributed to a variety of inflammatory manifestations, the current findings may contribute to dysfunctional monocytes in CKD patients. Strategies to interfere with this pathway may improve host immunity and prevent cardiovascular complications in CKD patients.