New strategies to measure intracellular sodium concentrations

Fluorescent ion indicators are widely used to measure ion concentrations in living cells. However, despite considerable efforts in synthesizing new compounds, no ratiometric sodium indicator is available that can be excited at visible wavelengths. Ratiometric indicators have an advantage in that measured fluorescence intensities can be corrected for fluctuations of the indicator concentration and the illumination intensity, which is not possible when non-ratiometric indicators are used. One way to circumvent this problem is to measure fluorescence lifetimes, which are independent of these factors. Another way to overcome the disadvantages of a non-ratiometric indicator dye is to embed it, together with a reference dye, into nanoparticles. By relating the indicator fluorescence to the fluorescence of the reference dye, inhomogeneities in the nanosensor concentration or the illumination intensity can be cancelled out reliably. In this study we compare the benefits and drawbacks of these approaches. © 2010 Copyright SPIE - The International Society for Optical Engineering

A Robust and Universal Metaproteomics Workflow for Research Studies and Routine Diagnostics Within 24 h Using Phenol Extraction, FASP Digest, and the MetaProteomeAnalyzer

The investigation of microbial proteins by mass spectrometry (metaproteomics) is a key technology for simultaneously assessing the taxonomic composition and the functionality of microbial communities in medical, environmental, and biotechnological applications. We present an improved metaproteomics workflow using an updated sample preparation and a new version of the MetaProteomeAnalyzer software for data analysis. High resolution by multidimensional separation (GeLC, MudPIT) was sacrificed to aim at fast analysis of a broad range of different samples in less than 24 h. The improved workflow generated at least two times as many protein identifications than our previous workflow, and a drastic increase of taxonomic and functional annotations. Improvements of all aspects of the workflow, particularly the speed, are first steps toward potential routine clinical diagnostics (i.e., fecal samples) and analysis of technical and environmental samples. The MetaProteomeAnalyzer is provided to the scientific community as a central remote server solution at www.mpa.ovgu.de.Peer Reviewe

Effects of telmisartan and ramipril on adiponectin and blood pressure in patients with type 2 diabetes

<b>Background:</b> Adiponectin is secreted by adipose tissue and may play a role in cardiovascular disease. We examined adiponectin levels in patients with type 2 diabetes who participated in the Telmisartan vs. Ramipril in Renal Endothelial Dysfunction (TRENDY) study. <b>Methods</b> A total of 87 patients were assessed at baseline and following 9 weeks treatment with the angiotensin-receptor blocker telmisartan (final dose, 80 mg; n = 45) or the angiotensin-converting enzyme inhibitor ramipril (final dose, 10 mg; n = 42). Adiponectin levels were measured in plasma by radioimmunoassay. <b>Results:</b> Adiponectin levels were inversely correlated with systolic (SBP; r = -0.240, P < 0.05) and diastolic (DBP; r = -0.227, P < 0.05) blood pressure at baseline and following treatment with telmisartan or ramipril (SBP: r = -0.228, P < 0.05; DBP: r = -0.286, P < 0.05). Changes in adiponectin levels were related to changes in SBP (r = -0.357, P < 0.01) and DBP (r = -0.286, P < 0.01). There was a significant increase in adiponectin levels in the telmisartan (0.68 (95% confidence interval (CI), 0.27 to 1.10) <sup>µ</sup>g/ml, P < 0.01) but not in the ramipril group (0.17 (95% CI, -0.56 to 0.90) <sup>µ</sup>g/ml, P = 0.67). Blood pressure reduction in the telmisartan group (DeltaSBP: -13.5 (95% CI, -17.0 to -10.0) mm Hg; ΔDBP: -7.6 (95% CI, -9.8 to -5.3) mm Hg, each P < 0.001) was significantly (P less than or equal to 0.01 for SBP and P < 0.01 for DBP) greater than in the ramipril group (ΔSBP: -6.1 (95% CI, -6.2 to -2.0) mm Hg; ΔDBP: -2.7 (95% CI, -5.0 to -0.5) mm Hg; P < 0.01 and P < 0.05, respectively). <b>Conclusion:</b> Adiponectin is correlated with blood pressure in patients with type 2 diabetes. Whether increased adiponectin contributes to the blood pressure–lowering effect of telmisartan needs further study

Nocardia macrotermitis sp. nov. and Nocardia aurantia sp. nov., isolated from the gut of the fungus-growing termite Macrotermes natalensis

The taxonomic positions of two novel aerobic, Gram-stain-positive Actinobacteria, designated RB20$^{T}$ and RB56$^{T}$, were determined using a polyphasic approach. Both were isolated from the fungus-farming termite Macrotermes natalensis. Results of 16S rRNA gene sequence analysis revealed that both strains are members of the genus Nocardia with the closest phylogenetic neighbours Nocardia miyunensis JCM12860$^{T}$ (98.9 %) and Nocardia nova DSM44481$^{T}$ (98.5 %) for RB20$^{T}$ and Nocardia takedensis DSM 44801$^{T}$ (98.3 %), Nocardia pseudobrasiliensis DSM 44290$^{T}$ (98.3 %) and Nocardia rayongensis JCM 19832$^{T}$ (98.2 %) for RB56$^{T}$. Digital DNA–DNA hybridization (DDH) between RB20$^{T}$ and N. miyunensis JCM12860$^{T}$ and N. nova DSM 44481$^{T}$ resulted in similarity values of 33.9 and 22.0 %, respectively. DDH between RB56$^{T}$ and N. takedensis DSM44801$^{T}$ and N. pseudobrasiliensis DSM44290$^{T}$ showed similarity values of 20.7 and 22.3 %, respectively. In addition, wet-lab DDH between RB56$^{T}$ and N. rayongensis JCM19832$^{T}$ resulted in 10.2 % (14.5 %) similarity. Both strains showed morphological and chemotaxonomic features typical for the genus Nocardia , such as the presence of meso-diaminopimelic acid (A$_{2}$pm) within the cell wall, arabinose and galactose as major sugar components within whole cell-wall hydrolysates, the presence of mycolic acids and major phospholipids (diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol), and the predominant menaquinone MK-8 (H4, ω-cyclo). The main fatty acids for both strains were hexadecanoic acid (C$_{16 : 0}$), 10-methyloctadecanoic acid (10-methyl C$_{18 : 0}$) and cis-9-octadecenoic acid (C$_{18 : 1}$ ω9c). We propose two novel species within the genus Nocardia : Nocardia macrotermitis sp. nov. with the type strain RB20$^{T}$ (=VKM Ac-2841$^{T}$=NRRL B65541$^{T}$) and Nocardia aurantia sp. nov. with the type strain RB56$^{T}$ (=VKM Ac-2842$^{T}$=NRRL B65542$^{T}$)

Metaproteome analysis reveals that syntrophy, competition, and phage-host interaction shape microbial communities in biogas plants

Background: In biogas plants, complex microbial communities produce methane and carbon dioxide by anaerobic digestion of biomass. For the characterization of the microbial functional networks, samples of 11 reactors were analyzed using a high-resolution metaproteomics pipeline. Results: Examined methanogenesis archaeal communities were either mixotrophic or strictly hydrogenotrophic in syntrophy with bacterial acetate oxidizers. Mapping of identified metaproteins with process steps described by the Anaerobic Digestion Model 1 confirmed its main assumptions and also proposed some extensions such as syntrophic acetate oxidation or fermentation of alcohols. Results indicate that the microbial communities were shaped by syntrophy as well as competition and phage-host interactions causing cell lysis. For the families Bacillaceae, Enterobacteriaceae, and Clostridiaceae, the number of phages exceeded up to 20-fold the number of host cells. Conclusion: Phage-induced cell lysis might slow down the conversion of substrates to biogas, though, it could support the growth of auxotrophic microbes by cycling of nutrients. © 2019 The Author(s)

Hsp27 regulates podocyte cytoskeletal changes in an in vitro model of podocyte process retraction

Nephrotic syndrome (NS) is characterized by structural changes in the actin‐rich foot processes of glomerular podocytes. We previously identified high concentrations of the small heat shock protein hsp27 within podocytes as well as increased glomerular accumulation and phosphorylation of hsp27 in puromycin aminonucleoside (PAN) ‐induced experimental NS. Here we analyzed murine podocytes stably transfected with hsp27 sense, antisense, and vector control constructs using a newly developed in vitro PAN model system. Cell morphology and the microfilament structure of untreated sense and antisense transfectants were altered compared with controls. Vector cell survival, polymerized actin content, cell area, and hsp27 content increased after 1.25 μg/ml PAN treatment and decreased after 5.0 μg/ml treatment. In contrast, sense cells were unaffected by 1.25 μg/ml PAN treatment whereas antisense cells showed decreases or no changes in all parameters. Treatment of sense cells with 5.0 μ g/ml PAN resulted in increased cell survival and cell area whereas antisense cells underwent significant decreases in all parameters. Hsp27 provided dramatic protection against PAN‐induced microfilament disruption in sense > vector > antisense cells. We conclude that hsp27 is able to regulate both the morphological and actin cytoskeletal response of podocytes in an in vitro model of podocyte injury.—Smoyer, W. E., Ransom, R. F. Hsp27 regulates podocyte cytoskeletal changes in an in vitro model of podocyte process retraction. FASEB J. 16, 315–326 (2002)Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/154256/1/fsb2fj010681com.pd

A Cognitive Model of an Epistemic Community: Mapping the Dynamics of Shallow Lake Ecosystems

We used fuzzy cognitive mapping (FCM) to develop a generic shallow lake ecosystem model by augmenting the individual cognitive maps drawn by 8 scientists working in the area of shallow lake ecology. We calculated graph theoretical indices of the individual cognitive maps and the collective cognitive map produced by augmentation. The graph theoretical indices revealed internal cycles showing non-linear dynamics in the shallow lake ecosystem. The ecological processes were organized democratically without a top-down hierarchical structure. The steady state condition of the generic model was a characteristic turbid shallow lake ecosystem since there were no dynamic environmental changes that could cause shifts between a turbid and a clearwater state, and the generic model indicated that only a dynamic disturbance regime could maintain the clearwater state. The model developed herein captured the empirical behavior of shallow lakes, and contained the basic model of the Alternative Stable States Theory. In addition, our model expanded the basic model by quantifying the relative effects of connections and by extending it. In our expanded model we ran 4 simulations: harvesting submerged plants, nutrient reduction, fish removal without nutrient reduction, and biomanipulation. Only biomanipulation, which included fish removal and nutrient reduction, had the potential to shift the turbid state into clearwater state. The structure and relationships in the generic model as well as the outcomes of the management simulations were supported by actual field studies in shallow lake ecosystems. Thus, fuzzy cognitive mapping methodology enabled us to understand the complex structure of shallow lake ecosystems as a whole and obtain a valid generic model based on tacit knowledge of experts in the field.Comment: 24 pages, 5 Figure