Induction of decay accelerating factor and membrane cofactor protein by resveratrol attenuates complement deposition in human coronary artery endothelial cells

The involvement of complement activation in various forms of cardiovascular disease renders it an important factor for disease progression and therapeutic intervention. The protective effect of resveratrol against cardiovascular disease via moderate red wine consumption has been established but the exact mechanisms are still under investigation. The current study utilised human coronary artery endothelial cells (HCAECs) in order to assess the extent to which the protective effect of resveratrol, at concentrations present in red wine, can be attributed to the upregulation of complement regulatory proteins through heme-oxygenase (HO)-1 induction. Resveratrol at concentrations as low as 0.001 μΜ increased HO-1 expression as well as membrane cofactor protein (MCP, CD46) and decay-accelerating factor (DAF, CD55) expression with no-effect on CD59. Silencing of HO-1 expression by HO-1 siRNAs abrogated both DAF and MCP protein expression with no effect on CD59. Resveratrol-mediated induction of DAF and MCP reduced C3b deposition following incubation of HCAECs with 10% normal human serum or normal rat serum as a source of complement. Incubation of HCAECs, with either a DAF blocking antibody or following transfection with HO-1 siRNAs, in the presence of 10% normal rat serum increased C3b deposition, indicating that both DAF and HO-1 are required for C3b reduction. These observations support a novel mechanism for the protective effect of resveratrol against cardiovascular disease and confirm the important role of HO-1 in the regulation of the complement cascade.     

Protein biomarkers for cardiorenal syndrome

Introduction: The term cardiorenal syndrome (CRS) describes the progressive pathology and interactions that develop upon heart and kidney failure. The definition of CRS is not firmly established and has evolved gradually during the last decade. The main clinical challenges associated with CRS are the lack of tools for early disease diagnosis and the inability to predict the development of cardiorenal pathophysiology. Currently several biomarkers have been proposed for improving CRS patient management. However, validation studies are needed to implement these initial findings to the clinical setting.

Areas covered: In this review the database PubMed was used for a literature search on the definition and classification of CRS as well as biomarkers for CRS diagnosis and prognosis.

Expert opinion: A universally acceptable classification system for CRS is not available. Thus, acquiring mechanistic insights relative to the pathophysiology of the disease is challenging. Reported biomarkers include well-established markers for heart/renal dysfunction and inflammation. Some proteins expressed in both organs have also been associated with CRS, yet their link to disease pathophysiology and organ cross-talk is missing. Establishing the link between deregulated molecular pathways and CRS phenotypes is required to define biological relevance of existing findings and ultimately biology-driven markers and targets.     

Drug Resistance in Natural Isolates of Leishmania donovani s.l. Promastigotes Is Dependent of Pgp170 Expression

Resistance of pathogens to drugs is a growing concern regarding many diseases. Parasites like Leishmania, Plasmodium and Entamoeba histolytica; and neoplastic cells, present the multidrug-resistant phenotype rendering chemotherapy ineffective. The acquired resistance of Leishmania to antimony has generated intense research on the mechanisms involved but the question has not yet been resolved. To test the hypothesis that drug efflux in Leishmania, as measured by flow cytometry using the fluorescent dye Rhodamine-123, is largely dependent on the number of efflux pumps an isolate can express, the amount of Pgp 170 molecules was assessed in ten field isolates (5 “resistant” and 5 “susceptible”) using: Western Blotting, Confocal and Transmission Electron Microscopy, and proteomics. Their survival after exposure to three antileishmanial drugs, in vitro, was evaluated and clinical data were compared to the in vitro results. All isolates were resistant to Glucantime but susceptible to Miltefosine, whilst Amphotericin B was more effective on the “susceptible” isolates. The MDR gene, expressing the transmembrane efflux pump Pgp 170, appears to play a key role in the phenomenon of drug resistance. When “susceptible” versus “resistant” parasites were compared, it was shown that the higher the number of Pgp 170 molecules the higher the Rhodamine-123 efflux from the parasite body and, when exposed to the drug, the number of efflux pumps increased. However, the rate of this increase was not linear and it is possible that there is a maximum number of Pgp 170 molecules an isolate can express. Nevertheless, the phenomenon is a complex one and other factors and proteins are involved in which the HSP-70 group proteins, detected in the “resistant” isolates, may play a significant role.     

Adenosine A2A receptors are required for glutamate mGluR5‐ and dopamine D1 receptor‐evoked ERK1/2 phosphorylation in rat hippocampus: involvement of NMDA receptor

Interaction between mGluR5 and NMDA receptors (NMDAR ) is vital for synaptic plasticity and cognition. We recently demonstrated that stimulation of mGluR5 enhances NMDAR responses in hippocampus by phosphorylating NR2B(Tyr1472) subunit, and this reaction was enabled by adenosine A_2A receptors (A_2AR) (J Neurochem, 135, 2015, 714). In this study, by using in vitro phosphorylation and western blot analysis in hippocampal slices of male Wistar rats, we show that mGluR5 stimulation or mGluR5/NMDAR s co‐stimulation synergistically activate ERK 1/2 signaling leading to c‐Fos expression. Interestingly, both reactions are under the permissive control of endogenous adenosine acting through A_2ARs. Moreover, mGluR5‐mediated ERK 1/2 phosphorylation depends on NMDAR , which however exhibits a metabotropic way of function, since no ion influx through its ion channel is required. Furthermore, our results demonstrate that mGluR5 and mGluR5/NMDAR ‐evoked ERK 1/2 activation correlates well with the mGluR5/NMDAR ‐evoked NR2B(Tyr1472) phosphorylation, since both phenomena coincide temporally, are Src dependent, and are both enabled by A_2ARs. This indicates a functional involvement of NR2B(Tyr1472) phosphorylation in the ERK 1/2 activation. Our biochemical results are supported by electrophysiological data showing that in CA 1 region of hippocampus, the theta burst stimulation (TBS)‐induced long‐term potentiation coincides temporally with an increase in ERK 1/2 activation and both phenomena are dependent on the tripartite A_2A, mGlu5, and NMDAR s. Furthermore, we show that the dopamine D1 receptors evoked ERK 1/2 activation as well as the NR2B(Tyr1472) phosphorylation are also regulated by endogenous adenosine and A_2ARs. In conclusion, our results highlight the A_2ARs as a crucial regulator not only for NMDAR responses, but also for regulating ERK 1/2 signaling and its downstream pathways, leading to gene expression, synaptic plasticity, and memory consolidation.

     

Influence of training on the response to exercise of adrenocorticotropin and growth hormone plasma concentrations in human swimmers

The aim of the present study was to evaluate the response of adrenocorticotropin ([ACTH]) and growth hormone ([GH]) concentrations to a typical aerobic swimming set during a training season. Nine top-level male endurance swimmers (age range 17–23 years) were tested during three training sessions occurring 6, 12 and 18 weeks after the beginning of the season. During each session, after a standard warm-up, the swimmers performed a training set of 15 × 200-m freestyle, with 20 s of rest between repetitions, at a predetermined individual speed. Blood samples were collected before warm-up and at the end of the training set. A few days before each session, the individual swimming velocity corresponding to the 4 mmol · l⁻¹ blood lactate concentration (v ₄) was assessed as a standard of aerobic performance. Aerobic training affected v ₄ levels, which were highest 18 weeks after the beginning of the season; at the same time, while [ACTH] response was attenuated, [GH] response was enhanced. These results could be considered as adaptations to the exercise intensity. In our training programme, these adaptations seemed to have occurred between the 12th and 18th weeks of the training season. Accepted: 21 April 1998     

Micropropagation and molecular characterization of Thymus sibthorpii Benth. (Lamiaceae), an aromatic-medicinal thyme with ornamental value and conservation concern

Thymus sibthorpii Benth. (Lamiaceae), with accession number 01,1796-22, is a biotype of native Greek thyme with ascending stems and potential use as a new medicinal-aromatic crop and ornamental plant. An efficient and reliable protocol for in vitro clonal propagation of T. sibthorpii from nodes and meristem tip explants was developed. Shoot proliferation succeeded on a new basal medium (BB) without plant growth regulators, as prior experiments with 6-benzyladenine generated hyperhydricity. Eight different basal media were compared; on two formulations using the new BB 5.9 and 5.6 shoots per explant were produced. Regenerated single shoots were rooted in the BB medium, supplemented with 5 μM of indole-3-butyric acid, and produced 3.1 roots along with 2.5 adventitious shoots. Three types of acclimatization were assessed: in vitro, using two different systems (no significant differences); ex vitro, using eight soil substrates under greenhouse and outdoor nursery conditions (in two of them, 100% of plantlets survived); and in field cultivations, established at eight geographically distant areas of Greece (100% survival rate at all locations). Molecular characterization of T. sibthorpii was evaluated with one nuclear ribosomal DNA and seven chloroplast DNA markers, followed by DNA sequence comparisons with a total of 30 different Thymus species, subspecies, and varieties. The trnH/psbA, trnL/trnF, and matK genes were the most efficient markers for molecular characterization of T. sibthorpii. The molecular markers rpoC1 and petB/petD did not match to any Thymus species and therefore, these DNA sequences provide new sequence information for entire Thymus taxa.

     

Measuring Symmetry,Asymmetry and Randomness in Neural Network Connectivity

Cognitive functions are stored in the connectome, the wiring diagram of the brain, which exhibits non-random features, so-called motifs. In this work, we focus on bidirectional, symmetric motifs, i.e. two neurons that project to each other via connections of equal strength, and unidirectional, non-symmetric motifs, i.e. within a pair of neurons only one neuron projects to the other. We hypothesise that such motifs have been shaped via activity dependent synaptic plasticity processes. As a consequence, learning moves the distribution of the synaptic connections away from randomness. Our aim is to provide a global, macroscopic, single parameter characterisation of the statistical occurrence of bidirectional and unidirectional motifs. To this end we define a symmetry measure that does not require any a priori thresholding of the weights or knowledge of their maximal value. We calculate its mean and variance for random uniform or Gaussian distributions, which allows us to introduce a confidence measure of how significantly symmetric or asymmetric a specific configuration is, i.e. how likely it is that the configuration is the result of chance. We demonstrate the discriminatory power of our symmetry measure by inspecting the eigenvalues of different types of connectivity matrices. We show that a Gaussian weight distribution biases the connectivity motifs to more symmetric configurations than a uniform distribution and that introducing a random synaptic pruning, mimicking developmental regulation in synaptogenesis, biases the connectivity motifs to more asymmetric configurations, regardless of the distribution. We expect that our work will benefit the computational modelling community, by providing a systematic way to characterise symmetry and asymmetry in network structures. Further, our symmetry measure will be of use to electrophysiologists that investigate symmetry of network connectivity.