A population level computational model of the basal ganglia that generates parkinsonian local field potential activity

Recordings from the basal ganglia’s subthalamic nucleus are acquired via microelectrodes immediately prior to the application of Deep Brain Stimulation (DBS) treatment for Parkinson’s Disease (PD) to assist in the selection of the final point for the implantation of the DBS electrode. The acquired recordings reveal a persistent characteristic beta band peak in the power spectral density function of the Local Field Potential (LFP) signals. This peak is considered to lie at the core of the causality–effect relationships of the parkinsonian pathophysiology. Based on LFPs acquired from human subjects during DBS for PD, we constructed a computational model of the basal ganglia on the population level that generates LFPs to identify the critical pathophysiological alterations that lead to the expression of the beta band peak. To this end, we used experimental data reporting that the strengths of the synaptic connections are modified under dopamine depletion. The hypothesis that the altered dopaminergic modulation may affect both the amplitude and the time course of the postsynaptic potentials is validated by the model. The results suggest a pivotal role of both of these parameters to the pathophysiology of PD.     

The endocytic recycling protein EHD2 interacts with myoferlin to regulate myoblast fusion

Skeletal muscle is a multinucleated syncytium that develops and is maintained by the fusion of myoblasts to the syncytium. Myoblast fusion involves the regulated coalescence of two apposed membranes. Myoferlin is a membrane-anchored, multiple C2 domain-containing protein that is highly expressed in fusing myoblasts and required for efficient myoblast fusion to myotubes. We found that myoferlin binds directly to the eps15 homology domain protein, EHD2. Members of the EHD family have been previously implicated in endocytosis as well as endocytic recycling, a process where membrane proteins internalized by endocytosis are returned to the plasma membrane. EHD2 binds directly to the second C2 domain of myoferlin, and EHD2 is reduced in myoferlin null myoblasts. In contrast to normal myoblasts, myoferlin null myoblasts accumulate labeled transferrin and have delayed recycling. Introduction of dominant negative EHD2 into myoblasts leads to the sequestration of myoferlin and inhibition of myoblast fusion. The interaction of myoferlin with EHD2 identifies molecular overlap between the endocytic recycling pathway and the machinery that regulates myoblast membrane fusion.     

A novel Leishmania infantum nuclear phosphoprotein Lepp12 which stimulates IL1-beta synthesis in THP-1 transfectants

Background

Tuberculosis remains a serious world-wide health threat which requires the characterisation of novel drug targets for the development of future antimycobacterials. One of the key obstacles in the definition of new targets is the large variety of metabolic alterations that occur between cells in the active growth and chronic/dormant phases of tuberculosis. The ideal biochemical target should be active in both growth phases. Methionine adenosyltransferase, which catalyses the formation of S-adenosylmethionine from methionine and ATP, is involved in polyamine biosynthesis during active growth and is also required for the methylation and cyclopropylation of mycolipids necessary for survival in the chronic phase.

Results

The gene encoding methionine adenosyltransferase has been cloned from Mycobacterium tuberculosis and the model organism M. smegmatis. Both enzymes retained all amino acids known to be involved in catalysing the reaction. While the M. smegmatis enzyme could be functionally expressed, the M. tuberculosis homologue was insoluble and inactive under a large variety of expression conditions. For the M. smegmatis enzyme, the Vmax for S-adenosylmethionine formation was 1.30 μmol/min/mg protein and the Km for methionine and ATP was 288 μM and 76 μM respectively. In addition, the enzyme was competitively inhibited by 8-azaguanine and azathioprine with a Ki of 4.7 mM and 3.7 mM respectively. Azathioprine inhibited the in vitro growth of M. smegmatis with a minimal inhibitory concentration (MIC) of 500 μM, while the MIC for 8-azaguanine was >1.0 mM.

Conclusion

The methionine adenosyltransferase from both organisms had a primary structure very similar those previously characterised in other prokaryotic and eukaryotic organisms. The kinetic properties of the M. smegmatis enzyme were also similar to known prokaryotic methionine adenosyltransferases. Inhibition of the enzyme by 8-azaguanine and azathioprine provides a starting point for the synthesis of higher affinity purine-based inhibitors.     

Active remodeling in idiopathic interstitial pneumonias: evaluation of collagen types XII and XIV.

Fibril-associated collagens with interrupted triple helices (FACITs) XII and XIV act as fibril organizers and assist in the maintenance of uniform fibril size. We investigated the spatial expression patterns of collagens XII and XIV in cryptogenic organizing pneumonia (COP)/organizing pneumonia (OP) and in idiopathic pulmonary fibrosis (IPF)/usual interstitial pneumonia (UIP) and compared them to normal human lung. Study subjects included 10 patients with COP/OP, 10 patients with IPF/UIP, and 8 control subjects. Immunostaining for collagens XII and XIV was carried out in paraffin-embedded human lung tissue sections. Picrosirius red histochemical staining for collagen I expression and electron microcopy to evaluate fibril diameter were also performed. In normal lung, collagens XII and XIV were expressed in perivascular and subpleural connective tissue. In COP/OP, both collagens showed intense staining in perivascular connective tissue, thickened alveolar septae, and subpleural areas. In IPF/UIP, XII and XIV were expressed in perivascular connective tissue, in areas of established fibrosis, and in areas of subpleural thickening. Only collagen XII was expressed in granulation tissue plugs in COP/OP and in fibroblastic foci in IPF/UIP. Collagen type I was overexpressed in fibrotic areas. Electron micrographs revealed obvious fibril diameter alteration and fusion in the same areas. FACITs XII and XIV are expressed in normal and fibrotic lung. Unlike collagen XIV, collagen XII was expressed in granulation tissue plugs in COP/OP and in fibroblast foci in IPF/UIP. This may suggest a possible distinct role for both collagens in the modulation of the extracellular matrix during the onset of fibrotic process.     

Lipase activity in Thermus thermophilus HB8: Purification and characterization of the extracellular enzyme

In this study, the lipolytic activity of Thermus thermophilus HB8 was examined. The addition of various oils increased the production of extracellular lipolytic activity, while a combination of olive oil and glucose increased both extracellular and intracellular lipolytic activity. The oxygen transfer rate had a significant influence on both biomass and production of extra- or intra-cellular lipolytic activity. The formation of white halos due to the hydrolysis of oleic acid ester (Tween 80) in agar plates containing Nile Blue and the formation of Ca²⁺-oleate indicated the secretion of lipase. When the cell-free supernatant of cells grown in basal reach medium or the corresponding intracellular extract were electrophoresed under denatured and renatured conditions, using ??-naphthyl acetate and Fast Blue RR, major bands at 56 kDa or 62 and 32 kDa were observed, respectively. The 56 kDa extracellular enzyme was partial purified and characterized. Its peak of activity occurred at 80°C and pH 7.0, while the T_1/2 was 1 h at 100°C. The K _m of the partial purified enzyme was 1 mM and the V _max was 0.044 U/mL/min when using p-nitrophenyl laurate as substrate. The presence of Ca²⁺ and Hg²⁺ stimulated lipase activity, whereas Zn²⁺, Co²⁺, or EDTA inhibited lipase activity. The highest activity was observed in the presence of coconut oil and p-nitrophenyl laurate (pNPL). Purified lipase was the most stable in the presence of various organic solvents, such as pentanol, chloroform and n-dodecane. Because of the superior thermostability and stability in the presence of organic solvents of T. thermophilus extracellular lipase, this lipase holds great promise for use in industrial applications.     

SALL4 controls cell fate in response to DNA base composition

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Towards analyzing the potential of exosomes to deliver microRNA therapeutics

Exosome selectivity mechanisms underlying exosome–target cell interactions and the specific traits affecting their capability to communicate still remain unclear. Moreover, the capacity of exosomes to efficiently deliver their molecular cargos intracellularly needs precise investigation towards establishing functional exosome‐based delivery platforms exploitable in the clinical practice. The current study focuses on: (a) exosome production from normal MRC‐5 and Vero cells growing in culture, (b) physicochemical characterization by dynamic light scattering (DLS) and cryo‐transmission electron microscopy; (c) cellular uptake studies of rhodamine‐labeled exosomes in normal and cancer cells, providing to exosomes either “autologous” or “heterologous” cellular delivery environments; and (d) loading exogenous Alexa Fluor 488‐labeled siRNA into exosomes for the assessment of their delivering capacity by immunofluorescence in a panel of recipient cells. The data obtained thus far indicate that MRC‐5 and Vero exosomes, indeed exhibit an interesting delivering profile, as promising “bio‐shuttles,” being pharmacologically exploitable in the context of theranostic applications.     

Monocyte attachment to laminin in diabetes mellitus: The role of ATP

Monocyte-extracellular matrix interactions have been implicated in atherosclerosis pathophysiology. Laminin, the main basement membrane protein contains cell binding domains that can be cryptic, presented only after protein modification. In the present study we evaluated monocyte attachment to laminin-1 in the presence of ATP. Monocytes were derived from either healthy volunteers or patients with diabetes mellitus type II. For the estimation of monocyte attachment to laminin the myeloperoxidase assay was used. Monocytes derived from diabetic patients, showed an increased ability to attach to laminin (p = 0.0055). The presence of ATP increased the attachment of control monocytes to laminin (p = 0.0022). On the contrary, the presence of ATP did not affect the attachment of monocytes derived from diabetic patients to laminin. Our results indicate a modified interaction between monocytes and laminin-1 in diabetes mellitusKey words: monocytes, ATP, laminin-1, diabetes mellitus, attachment     

Butterfly phenology in Mediterranean mountains using space‐for‐time substitution

Inferring species' responses to climate change in the absence of long‐term time series data is a challenge, but can be achieved by substituting space for time. For example, thermal elevational gradients represent suitable proxies to study phenological responses to warming. We used butterfly data from two Mediterranean mountain areas to test whether mean dates of appearance of communities and individual species show a delay with increasing altitude, and an accompanying shortening in the duration of flight periods. We found a 14‐day delay in the mean date of appearance per kilometer increase in altitude for butterfly communities overall, and an average 23‐day shift for 26 selected species, alongside average summer temperature lapse rates of 3°C per km. At higher elevations, there was a shortening of the flight period for the community of 3 days/km, with an 8.8‐day average decline per km for individual species. Rates of phenological delay differed significantly between the two mountain ranges, although this did not seem to result from the respective temperature lapse rates. These results suggest that climate warming could lead to advanced and lengthened flight periods for Mediterranean mountain butterfly communities. However, although multivoltine species showed the expected response of delayed and shortened flight periods at higher elevations, univoltine species showed more pronounced delays in terms of species appearance. Hence, while projections of overall community responses to climate change may benefit from space‐for‐time substitutions, understanding species‐specific responses to local features of habitat and climate may be needed to accurately predict the effects of climate change on phenology.