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1.
A Arcangeli M Riccarda Del Bene R Poli L Ricupero M Olivotto 《Journal of cellular physiology》1989,139(1):1-8
This study deals with the modulation of the plasma membrane potential (delta psi p) of murine erythroleukemia (MEL) cells by cell-substratum or cell-cell contact. delta psi p was determined by measuring the distribution of tetraphenylphosphonium (TPP+) across the plasma membrane; it appeared strongly, and inversely, influenced by the two types of cell contacts. Contact with the culture surface produced a delta psi p hyperpolarization directly proportional to average distance among the ideal centers of the cells on this surface (d) within the range 10-80 microns. A detailed mathematical analysis of the function delta psi p = f(d) is presented, as well as experiments involving the use of ionophores (valinomycin and A23187) and the conditioning of the culture surface. We concluded that the d-dependent hyperpolarization (dDH) was the result of a complex interplay between the activating properties of substratum on Ca2+-dependent K+ channels (KCa) and some substratum-adherent factors that are shed by MEL cells and antagonize KCa activation (substratum-attached cellular factors = SACF). By contrast, contact of the cells with each other, obtained by incubating MEL cells at d smaller than the average cell diameter (phi = 10 microns), produced a marked delta psi p depolarization. This intercellular contact-dependent depolarization (ICDD) was unaffected by valinomycin; it was abolished by substituting Na+ in the external medium with a nondiffusible cation (choline), which shows that ICDD was sustained by Na+ influxes, probably mediated by stretch-activated (s.a.) cation channels. 相似文献
2.
Soil respiration (R s) plays a key role in any consideration of ecosystem carbon (C) balance. Based on the well-known temperature response of respiration in plant tissue and microbes, R s is often assumed to increase in a warmer climate. Yet, we assume that substrate availability (labile C input) is the dominant influence on R s rather than temperature. We present an analysis of NPP components and concurrent R s in temperate deciduous forests across an elevational gradient in Switzerland corresponding to a 6 K difference in mean annual temperature and a considerable difference in the length of the growing season (174 vs. 262 days). The sum of the short-lived NPP fractions (“canopy leaf litter,” “understory litter,” and “fine root litter”) did not differ across this thermal gradient (+6 % from cold to warm sites, n.s.), irrespective of the fact that estimated annual forest wood production was more than twice as high at low compared to high elevations (largely explained by the length of the growing season). Cumulative annual R s did not differ significantly between elevations (836 ± 5 g C m?2 a?1 and 933 ± 40 g C m?2 a?1 at cold and warm sites, +12 %). Annual soil CO2 release thus largely reflected the input of labile C and not temperature, despite the fact that R s showed the well-known short-term temperature response within each site. However, at any given temperature, R s was lower at the warm sites (downregulation). These results caution against assuming strong positive effects of climatic warming on R s, but support a close substrate relatedness of R s. 相似文献
3.
Obestatin regulates cardiovascular function and promotes cardioprotection through the nitric oxide pathway 下载免费PDF全文
Saveria Femminò Carmine Rocca Tommaso Angelone Maria C. Cerra Maria Pia Gallo Iacopo Gesmundo Alessandro Fanciulli Maria Felice Brizzi Pasquale Pagliaro Riccarda Granata 《Journal of cellular and molecular medicine》2017,21(12):3670-3678
Patients with ischaemic heart disease or chronic heart failure show altered levels of obestatin, suggesting a role for this peptide in human heart function. We have previously demonstrated that GH secretagogues and the ghrelin gene‐derived peptides, including obestatin, exert cardiovascular effects by modulating cardiac inotropism and vascular tone, and reducing cell death and contractile dysfunction in hearts subjected to ischaemia/reperfusion (I/R), through the Akt/nitric oxide (NO) pathway. However, the mechanisms underlying the cardiac actions of obestatin remain largely unknown. Thus, we suggested that obestatin‐induced activation of PI3K/Akt/NO and PKG signalling is implicated in protection of the myocardium when challenged by adrenergic, endothelinergic or I/R stress. We show that obestatin exerts an inhibitory tone on the performance of rat papillary muscle in both basal conditions and under β‐adrenergic overstimulation, through endothelial‐dependent NO/cGMP/PKG signalling. This pathway was also involved in the vasodilator effect of the peptide, used both alone and under stress induced by endothelin‐1. Moreover, when infused during early reperfusion, obestatin reduced infarct size in isolated I/R rat hearts, through an NO/PKG pathway, comprising ROS/PKC signalling, and converging on mitochondrial ATP‐sensitive potassium [mitoK(ATP)] channels. Overall, our results suggest that obestatin regulates cardiovascular function in stress conditions and induces cardioprotection by mechanisms dependent on activation of an NO/soluble guanylate cyclase (sGC)/PKG pathway. In fact, obestatin counteracts exaggerated β‐adrenergic and endothelin‐1 activity, relevant factors in heart failure, suggesting multiple positive effects of the peptide, including the lowering of cardiac afterload, thus representing a potential candidate in pharmacological post‐conditioning. 相似文献
4.
The functional evolution of proteins advances through gene duplication followed by functional drift, whereas molecular evolution occurs through random mutational events. Over time, protein active-site structures or functional epitopes remain highly conserved, which enables relationships to be inferred between distant orthologs or paralogs. In this study, we present the first functional clustering and evolutionary analysis of the RCSB Protein Data Bank (RCSB PDB) based on similarities between active-site structures. All of the ligand-bound proteins within the RCSB PDB were scored using our Comparison of Protein Active-site Structures (CPASS) software and database ( http://cpass.unl.edu/ ). Principal component analysis was then used to identify 4431 representative structures to construct a phylogenetic tree based on the CPASS comparative scores ( http://itol.embl.de/shared/jcatazaro ). The resulting phylogenetic tree identified a sequential, step-wise evolution of protein active-sites and provides novel insights into the emergence of protein function or changes in substrate specificity based on subtle changes in geometry and amino acid composition. 相似文献
5.
Families of distantly related proteins typically have very low sequence identity, which hinders evolutionary analysis and functional annotation. Slowly evolving features of proteins, such as an active site, are therefore valuable for annotating putative and distantly related proteins. To date, a complete evolutionary analysis of the functional relationship of an entire enzyme family based on active‐site structural similarities has not yet been undertaken. Pyridoxal‐5′‐phosphate (PLP) dependent enzymes are primordial enzymes that diversified in the last universal ancestor. Using the comparison of protein active site structures (CPASS) software and database, we show that the active site structures of PLP‐dependent enzymes can be used to infer evolutionary relationships based on functional similarity. The enzymes successfully clustered together based on substrate specificity, function, and three‐dimensional‐fold. This study demonstrates the value of using active site structures for functional evolutionary analysis and the effectiveness of CPASS. Proteins 2014; 82:2597–2608. © 2014 Wiley Periodicals, Inc. 相似文献
6.
Baragli A Ghè C Arnoletti E Granata R Ghigo E Muccioli G 《Biochimica et biophysica acta》2011,1811(6):386-396
The acylated peptide ghrelin (AG) and its endogenous non-acylated isoform (UAG) protect cardiomyocytes, pancreatic β-cells, and preadipocytes from apoptosis, and induce preadipocytes differentiation into adipocytes. These events are mediated by AG and UAG binding to a still unidentified receptor, which determines the activation of phosphoinositide 3-kinase (PI3K), protein kinase B (AKT), and mitogen-activated protein kinase (MAPK) ERK1/2. AG and UAG also possess antilipolytic activity in vitro, but the underlying mechanism remains unknown. Thus, the objective of the current study was to characterize the molecular events involved in AG/UAG receptor signaling cascade. We treated rat primary visceral adipocytes with isoproterenol (ISO) and forskolin (FSK) to stimulate lipolysis, simultaneously incubating them with or without AG or UAG. Both peptides blocked ISO- and FSK-induced lipolysis. By direct measurement of cAMP intracellular content, we demonstrated that AG/UAG effect was associated to a reduction of ISO-induced cAMP accumulation. Moreover, the cAMP analog 8Br-cAMP abolished AG/UAG effect. As AG and UAG were ineffective against lipolysis induced by db-cAMP, another poorly hydrolyzable cAMP analog, phosphodiesterase (PDE) involvement was hypothesized. Indeed, cilostamide, a specific PDE3B inhibitor, blocked AG/UAG effect on ISO-induced lipolysis. Furthermore, the PI3K inhibitor wortmannin and AKT inhibitor 1,3-dihydro-1-(1-((4-(6-phenyl-1H-imidazo(4,5-g)quinoxalin-7-yl)phenyl)methyl)-4piperidinyl)-2H-benzimidazol-2-one trifluoroacetate also blocked AG/UAG action, suggesting a role in PDE3B activation. In particular, PI3K isoenzyme gamma (PI3Kγ) selective inhibition through the compound AS605240 prevented AG/UAG effect on ISO-stimulated lipolysis, hampering AKT phosphorylation on Ser(473). Taken together, these data demonstrate for the first time that AG/UAG attenuation of ISO-induced lipolysis involves PI3Kγ/AKT and PDE3B. 相似文献
7.
Acylated ghrelin (AG) is a 28 amino acid gastric peptide a natural ligand for the growth hormone secretagogue (GHS) receptor type 1a (GHS-R1a), endowed with GH-secreting and orexigenic properties. Besides, ghrelin exerts several peripheral metabolic actions, including modulation of glucose homeostasis and stimulation of adipogenesis. Notably, AG administration causes hyperglycemia in rodents as in humans. Ghrelin pleiotropy is supported by a widespread expression of the ghrelin gene, of GHS-R1a and other unknown ghrelin binding sites. The existence of alternative receptors for AG, of several natural ligands for GHS-R1a and of acylation-independent ghrelin non-neuroendocrine activities, suggests that there might be a complex ‘ghrelin system’ not yet completely explored. Moreover, the patho-physiological implications of unacylated ghrelin (UAG), and obestatin (Ob), the other two ghrelin gene-derived peptides, need to be clarified. Within the next few years, we may better understand the ‘ghrelin system’, where we might envisage clinical applications. 相似文献
8.
This paper describes a new amperometric biosensor for glucose monitoring. The biosensor is based on the activity of glucose dehydrogenase (GDH) and diaphorase (DI) co-immobilized with NAD(+) into a carbon nanotube paste (CNTP) electrode modified with an osmium functionalized polymer. This mediator was demonstrated to shuttle the electron transfer between the immobilized diaphorase and the CNTP electrode, thus, showing a good electrocatalytic activity towards NADH oxidation at potentials around +0.2V versus Ag|AgCl, where interfering reactions are less prone to occur. The biosensor exhibits a detection limit of 10 micromol L(-1), linearity up to 8 x 10(-4) mol L(-1), a sensitivity of 13.4 microA cm(-2)mmol(-1)L(-1), a good reproducibility (R.S.D. 2.1%, n=6) and a stability of about 1 week when stored dry at 4 degrees C. Finally, the proposed biosensor was applied for the determination of glucose in different samples of sweet wine and validated with a commercial spectrophotometric enzymatic kit. 相似文献
9.
Tobias T. H?gi Sabrina Klemensberger Riccarda Bereiter Sandor Nietzsche Raluca Cosgarea Simon Flury Adrian Lussi Anton Sculean Sigrun Eick 《PloS one》2015,10(6)
Background and Aim
There is a lack of suitable in vitro models to evaluate various treatment modalities intending to remove subgingival bacterial biofilm. Consequently, the aims of this in vitro-study were: a) to establish a pocket model enabling mechanical removal of biofilm and b) to evaluate repeated non-surgical periodontal treatment with respect to biofilm removal and reformation, surface alterations, tooth hard-substance-loss, and attachment of periodontal ligament (PDL) fibroblasts.Material and Methods
Standardized human dentin specimens were colonized by multi-species biofilms for 3.5 days and subsequently placed into artificially created pockets. Non-surgical periodontal treatment was performed as follows: a) hand-instrumentation with curettes (CUR), b) ultrasonication (US), c) subgingival air-polishing using erythritol (EAP) and d) subgingival air-polishing using erythritol combined with chlorhexidine digluconate (EAP-CHX). The reduction and recolonization of bacterial counts, surface roughness (Ra and Rz), the caused tooth substance-loss (thickness) as well as the attachment of PDL fibroblasts were evaluated and statistically analyzed by means of ANOVA with Post-Hoc LSD.Results
After 5 treatments, bacterial reduction in biofilms was highest when applying EAP-CHX (4 log10). The lowest reduction was found after CUR (2 log10). Additionally, substance-loss was the highest when using CUR (128±40 µm) in comparison with US (14±12 µm), EAP (6±7 µm) and EAP-CHX (11±10) µm). Surface was roughened when using CUR and US. Surfaces exposed to US and to EAP attracted the highest numbers of PDL fibroblasts.Conclusion
The established biofilm model simulating a periodontal pocket combined with interchangeable placements of test specimens with multi-species biofilms enables the evaluation of different non-surgical treatment modalities on biofilm removal and surface alterations. Compared to hand instrumentation the application of ultrasonication and of air-polishing with erythritol prevents from substance-loss and results in a smooth surface with nearly no residual biofilm that promotes the reattachment of PDL fibroblasts. 相似文献10.
Stephan Seiler Helena Schmidt Anita Lechner Thomas Benke Guenter Sanin Gerhard Ransmayr Riccarda Lehner Peter Dal-Bianco Peter Santer Patricia Linortner Christian Eggers Bernhard Haider Margarete Uranues Josef Marksteiner Friedrich Leblhuber Peter Kapeller Christian Bancher Reinhold Schmidt PRODEM Study Group 《PloS one》2012,7(12)