Higher Serum Concentrations of N-Terminal Pro-B-Type Natriuretic Peptide Associate with Prevalent Hypertension whereas Lower Associate with Incident Hypertension

BackgroundThe role of the natriuretic peptides (NPs) in hypertension is complex. Thus, a higher blood NP concentration is a robust marker of pressure-induced cardiac damage in patients with hypertension, whereas genetically elevated NP concentrations are associated with a reduced risk of hypertension and overweight individuals presumably at high risk of hypertension have lower NP concentrations.ObjectiveTo investigate the associations between serum N-terminal pro-B-type natriuretic peptide (NT-proBNP), used as a surrogate marker for active BNP, and prevalent as well as 5-year incident hypertension in a Danish general population sample.MethodsCross-sectional and prospective population-based study.ResultsAt baseline, among 5,307 participants (51.3% women, mean age 46.0±7.9 years) with a complete set of data, we recorded 1,979 cases with prevalent hypertension (PHT). Among 2,389 normotensive participants at baseline with a complete set of data, we recorded 324 cases with incident hypertension (IHT) on follow-up 5 years later. In models adjusted for age, sex, lifestyle, social, dietary, anthropometric, pulmonic, lipid, metabolic and renal risk factors, as well as heart rate and baseline blood pressure (only incident model), one standard deviation increase in baseline log-transformed NT-proBNP concentrations was on one side associated with a 21% higher risk of PHT (odds ratio [OR]: 1.21 [95% confidence interval (CI): 1.13-1.30], P<0.001), and on the other side with a 14% lower risk of IHT (OR: 0.86 [95%CI:0.76-0.98], P = 0.020).ConclusionsHigher serum concentrations of NT-proBNP associate with PHT whereas lower concentrations associate with IHT. This suggests that a lower amount of circulating BNP, resulting in diminished vasodilation and natriuresis, could be involved in the pathogenesis of hypertension in its early stages.     

Cell Sorting of Neural Stem and Progenitor Cells from the Adult Mouse Subventricular Zone and Live-imaging of their Cell Cycle Dynamics

Neural stem cells (NSCs) in the subventricular zone of the lateral ventricles (SVZ) sustain olfactory neurogenesis throughout life in the mammalian brain. They successively generate transit amplifying cells (TACs) and neuroblasts that differentiate into neurons once they integrate the olfactory bulbs. Emerging fluorescent activated cell sorting (FACS) techniques have allowed the isolation of NSCs as well as their progeny and have started to shed light on gene regulatory networks in adult neurogenic niches. We report here a cell sorting technique that allows to follow and distinguish the cell cycle dynamics of the above-mentioned cell populations from the adult SVZ with a LeX/EGFR/CD24 triple staining. Isolated cells are then plated as adherent cells to explore in details their cell cycle progression by time-lapse video microscopy. To this end, we use transgenic Fluorescence Ubiquitination Cell Cycle Indicator (FUCCI) mice in which cells are red-fluorescent during G₁ phase due to a G₁ specific red-Cdt1 reporter. This method has recently revealed that proliferating NSCs progressively lengthen their G₁ phase during aging, leading to neurogenesis impairment. This method is easily transposable to other systems and could be of great interest for the study of the cell cycle dynamics of brain cells in the context of brain pathologies.     

A mutational analysis reveals new functional interactions between domains of the Oxa1 protein in Saccharomyces cerevisiae

The Oxa1/YidC/Alb3 family plays a key role in the biogenesis of the respiratory and photosynthetic complexes in bacteria and organelles. In Saccharomyces cerevisiae, Oxa1 mediates the co‐translational insertion of mitochondrially encoded subunits of the three respiratory complexes III, IV and V within the inner membrane and also controls a late step in complex V assembly. No crystal structure of YidC or Oxa1 is available and little is known about the respective role of each transmembrane segment (TM) and hydrophilic loop of this polytopic protein on the biogenesis of the three complexes. Here, we have generated a collection of random point mutations located in the hydrophobic and hydrophilic domains of the protein and characterized their effects on the assembly of the three respiratory complexes. Our results show mutant‐dependent differential effects, particularly on complex V. In order to identify tertiary interactions within Oxa1, we have also isolated revertants carrying second‐site compensatory mutations able to restore respiration. This analysis reveals the existence of functional interactions between TM2 and TM5, TM4 and TM5 as well as between TM4 and loop 2, highlighting the key position of TM4 and TM5 in the Oxa1 protein.     

Thigmomorphogenesis in Solanum lycopersicum: Morphological and biochemical responses in stem after mechanical stimulation

The activation of the phenylpropanoid pathway in plants by environmental stimuli is one of the most universal biochemical stress responses known. In tomato plant, rubbing applied to a young internode inhibit elongation of the rubbed internode and his neighboring one. These morphological changes were correlated with an increase in lignification enzyme activities, phenylalanine ammonia-lyase (PAL), cinnamyl alcohol dehydrogenase (CAD) and peroxidases (POD), 24 hours after rubbing of the forth internode. Furthermore, a decrease in indole-3-acetic acid (IAA) content was detected in the rubbed internode and the upper one. Taken together, our results suggest that decrease in rubbed internode length is a consequence of IAA oxidation, increases in enzyme activities (PAL, CAD and POD), and cell wall rigidification associated with induction of lignification process.Key words: Mechanical stimulation, PAL, CAD, POD, IAAIn their environment, plants are constantly submitted to several stimuli such as wind, rain and wounding. The growth response of plants to such stimuli was termed thigmomorphogenesis and was observed in a wide range of plants.^1–3 The most common thigmomorphogenetic response is a retardation of tissue elongation accompanied by an increase in thickness.⁴ The plant response to mechanical perturbation is mainly restricted to the young developing internode, since no influence can be detected when the internode has reached its final length.^5,6 These plant growth modifications, which characterize thigmomorphogenesis, are related to biochemical events associated with lignification process⁷ and ethylene production.^8,9In tomato plant the length of internodes 4 (N4) and 5 (N5) was measured 14 days after rubbing of the fourth internode. Results reported in Figure 1 show that rubbing led to a significant reduction of elongation of the stressed internode (N4) (decrease of N4 length from 4.3 cm in the control plant to 2.9 in the rubbed one). This effect was not limited to the rubbed area but affected also the elongation of the neighboring internodes (N5) that were shorter in rubbed plants than in control ones.Open in a separate windowFigure 1Internode lengths of control and rubbed plants measured 14 day after mechanical stress applied to the fourth internode. Standard errors are indicated by vertical bars.Results reported in Figure 2 show an increase in PAL activity in both internodes N4 and N5, 24 hours after mechanical stress application as compared with corresponding controls. CAD activity was also investigated in N4 and N5, 24 h after rubbing of the fourth internode. Results presented in Figure 3 show that mechanical stress application induces a strong increase of CAD activity in the rubbed internode N4 (5.3 nkatal μg-1 protein) with an approximately two-fold increase when compared to control tomato internodes (2.3 nkatal μg-1 protein). Further, CAD activity in N5 was also increased in the rubbed internode (5.538 nkatal μg-1 protein) as compared with the control one (3.256 nkatal μg-1 protein).Open in a separate windowFigure 2PAL activity of internode 4, and 5 in control and rubbed plants 24 h after rubbing of the fourth internode. Standard errors are indicated by vertical bars.Open in a separate windowFigure 3CAD activity of internode 4, and 5 in control and rubbed plants 24 h after rubbing of the fourth internode. Standard errors are indicated by vertical bars.Syringaldazine (S-POD) and gaïacol (G-POD) peroxidase activities were measured in tomato N4 and N5. Results reported in Figure 4 show an increase in soluble peroxidase activity with both substrates in the rubbed internode N4 as compared with control plant. Enhancement in peroxidase activities in N4 was more pronounced with gaïacol (80.7 U) as an electron donor than syringaldazine (33.8 U). Similar results were observed in internode 5 as compared with control one (Fig. 4).Open in a separate windowFigure 4(A) Syringaldazine-POD (Syr-POD) activity of internode 4 and 5 in control and rubbed plants 24 h after rubbing of the fourth internode. Standard errors are indicated by vertical bars. (B) Gaiacol-POD (G-POD) activity of internode 4 and 5 in control and rubbed plants 24 h after rubbing of the fourth internode. Standard errors are indicated by vertical bars.IAA was quantified in control and rubbed plant internodes 24 h after rubbing of the fourth internode. Results reported in figure 5 show that in control sample and as expected, the content of IAA was found to be higher in the younger internode (N5) as compared to the older one (N4). Rubbing led to a significant decrease in IAA levels in N4 (5.06 nmol g⁻¹ MF⁻¹) as compared with corresponding controls (7.27 nmol g⁻¹ MF⁻¹). Similar results were observed in internode 5, where IAA content was reduced from 16.52 nmol g⁻¹ MF⁻¹ in control internode to 12.35 nmol g⁻¹ MF⁻¹ in the rubbed internode (Fig. 5).Open in a separate windowFigure 5IAA Level of internode 4 and 5 in control and rubbed plants 24 h after rubbing of the fourth internode. Standard errors are indicated by vertical bars.The results reported here establish an evident correlation between growth limitation of the rubbed internode and their degree of lignification, the increase in lignification enzymes activities and auxin degradation after mechanical stress application.Auxin seems to be involved in thigmomorphogenesis.¹⁰ It was proposed that MIS (Mechanically-induced stress) has opposite effects on auxin levels in the two species studied to date, Phaseolus vulgaris¹⁰ and Bryonia dioica.^11,12 Auxin level as measured by bioassay, increased in Phaseolus vulgaris following rubbing of the stem.¹⁰ It was proposed that a build up of auxin may result from the reduced polar transport of IAA at the rubbed internode, causing a build up of IAA in the stem tissue. Exogenous IAA did not reverse the MIS inhibition of growth in Phaseolus vulgaris and high levels of IAA retarded growth in non-stressed plants.¹⁰ Thus, retardation of extension growth in Phaseolus vulgaris may have been caused by high levels of endogenous auxin and the increase in stem diameter by increased ethylene production.⁴ However, ethylene increases radial growth only if auxin is present.¹³Boyer¹¹ reported a decrease in auxinlike activity in Bryonia dioica following MIS and this was confirmed in the same species by Hofinger et al.¹² who reported a decrease in IAA using gas chromatography-mass spectrometry. Auxin catabolism was accompanied with changes in both soluble and ionically bound cell wall basic peroxidases¹⁴ and the appearance of an additional peroxidase. This can suggest that in Bryonia, auxin catabolism is hastened by mechanical stimulated peroxidase. In addition, Boyer et al.¹⁵ reported that lithium pre-treatment prevents both thigmomorphogenesis and appearance of specific cathodic isoperoxidase in Bryonia plants subjected to MIS. This is give further credence to the possibility that the peroxidase-auxin system is involved in Bryonia thigmomorphogenesis. In addition, ethylene increases peroxidase activity which reduces the auxin content in the tissue to a level low enough not to support normal growth. We have evidence that decrease of auxin level contribute to mechanism leading to tomato internode inhibition subjected to mechanical stress.Growth inhibition has been suggested to be the result of tissues lignification.⁶ As the initial enzyme in the monolignol biosynthesis pathway, PAL has a direct influence on lignin accumulation.¹⁶ The characteristics of lignin differ among cell wall tissues and plant organs.¹⁷ It comprises polyphenolic polymers derived from the oxidative polymerization of different monolignols, including p-coumaryl, coniferyl and sinapyl alcohols via a side pathway of phenylalanine metabolism leading to lignin synthesis.¹⁸ The increase in lignin content in the rubbed tomato internode could be a response mechanism to mechanical damage caused by rubbing.³ It is known that plants create a natural barrier that includes lignin and suberin synthesis, components directly linked to support systems.^19,20The increase in lignin content of rubbed tomato internode³ is paralleled by a rise in CAD activity and whilst such direct proportionality between CAD activity and lignin accumulation does not always agree with the results in the literature, it clearly is responding in ways similar to those of the other enzymes in the pathway.²¹Mechanical stress-induced membrane depolarization would generate different species of free radicals and peroxides, which in turn initiate lipid peroxidation.²² The degradation of cell membranes is suggested to bring about rapid changes in ionic flux, especially release of K⁺ which would result in an enhanced endogenous Ca/K ratio and in leakage of solutes, among them electron donors such as ascorbic acid and phenolic substances. The increased intracellular relative calcium level activated secretion of basic peroxidases²³ into the free space where, in association with the electron donors and may be with the circulating IAA, they eliminate the peroxides, and facilitated binding of basic peroxidases to membrane structures allowing a role as 1-aminocyclopropane-1-carboxylic acid (ACC)-oxidases. The resulting IAA and ACC oxidase-mediated changes in ethylene production²⁴ would further induce (this time through the protein synthesis machinery) an increase in activity of phenylalanine ammonia-lyase and peroxidases. The resulting lignification and cell wall rigidification determines the growth response of tomato internode to the mechanical stress.     

The Cdc48-Ufd1-Npl4 complex is central in ubiquitin-proteasome triggered catabolite degradation of fructose-1,6-bisphosphatase

The switch from gluconeogenesis to glycolysis in yeast has been shown to require ubiquitin-proteasome dependent elimination of the key enzyme fructose-1,6-bisphosphatase (FBPase). Prior to proteasomal degradation, polyubiquitination of the enzyme occurs via the ubiquitin-conjugating enzymes Ubc1, Ubc4, Ubc5 and Ubc8 in conjunction with a novel multi-subunit ubiquitin ligase, the Gid complex. As an additional machinery required for the catabolite degradation process, we identified the trimeric Cdc48^Ufd1-Npl4 complex and the ubiquitin receptors Dsk2 and Rad23. We show that this machinery acts between polyubiquitination of FBPase and its degradation by the proteasome.     

Both caffeoyl Coenzyme A 3-<Emphasis Type="Italic">O</Emphasis>-methyltransferase 1 and caffeic acid <Emphasis Type="Italic">O</Emphasis>-methyltransferase 1 are involved in redundant functions for lignin,flavonoids and sinapoyl malate biosynthesis in Arabidopsis

Two methylation steps are necessary for the biosynthesis of monolignols, the lignin precursors. Caffeic acid O-methyltransferase (COMT) O-methylates at the C5 position of the phenolic ring. COMT is responsible for the biosynthesis of sinapyl alcohol, the precursor of syringyl lignin units. The O-methylation at the C3 position of the phenolic ring involves the Caffeoyl CoA 3-O-methyltransferase (CCoAOMT). The CCoAOMT 1 gene (At4g34050) is believed to encode the enzyme responsible for the first O-methylation in Arabidopsis thaliana. A CCoAOMT1 promoter-GUS fusion and immunolocalization experiments revealed that this gene is strongly and exclusively expressed in the vascular tissues of stems and roots. An Arabidopsis T-DNA null mutant named ccomt 1 was identified and characterised. The mutant stems are slightly smaller than wild-type stems in short-day growth conditions and has collapsed xylem elements. The lignin content of the stem is low and the S/G ratio is high mainly due to fewer G units. These results suggest that this O-methyltransferase is involved in G-unit biosynthesis but does not act alone to perform this step in monolignol biosynthesis. To determine which O-methyltransferase assists CCoAOMT 1, a comt 1 ccomt1 double mutant was generated and studied. The development of comt 1 ccomt1 is arrested at the plantlet stage in our growth conditions. Lignins of these plantlets are mainly composed of p-hydroxyphenyl units. Moreover, the double mutant does not synthesize sinapoyl malate, a soluble phenolic. These results suggest that CCoAOMT 1 and COMT 1 act together to methylate the C3 position of the phenolic ring of monolignols in Arabidopsis. In addition, they are both involved in the formation of sinapoyl malate and isorhamnetin.     

Ketone bodies alter dinitrophenol-induced glucose uptake through AMPK inhibition and oxidative stress generation in adult cardiomyocytes

In aerobic conditions, the heart preferentially oxidizes fatty acids. However, during metabolic stress, glucose becomes the major energy source, and enhanced glucose uptake has a protective effect on heart function and cardiomyocyte survival. Thus abnormal regulation of glucose uptake may contribute to the development of cardiac disease in diabetics. Ketone bodies are often elevated in poorly controlled diabetics and are associated with increased cellular oxidative stress. Thus we sought to determine the effect of the ketone body beta-hydroxybutyrate (OHB) on cardiac glucose uptake during metabolic stress. We used 2,4-dinitrophenol (DNP), an uncoupler of the mitochondrial oxidative chain, to mimic hypoxia in cardiomyocytes. Our data demonstrated that chronic exposure to OHB provoked a concentration-dependent decrease of DNP action, resulting in 56% inhibition of DNP-mediated glucose uptake at 5 mM OHB. This was paralleled by a diminution of DNP-mediated AMP-activated protein kinase (AMPK) and p38 MAPK phosphorylation. Chronic exposure to OHB also increased reactive oxygen species (ROS) production by 1.9-fold compared with control cells. To further understand the role of ROS in OHB action, cardiomyocytes were incubated with H(2)O(2). Our results demonstrated that this treatment diminished DNP-induced glucose uptake without altering activation of the AMPK/p38 MAPK signaling pathway. Incubation with the antioxidant N-acetylcysteine partially restored DNP-mediated glucose but not AMPK/p38 MAPK activation. In conclusion, these results suggest that ketone bodies, through inhibition of the AMPK/p38 MAPK signaling pathway and ROS overproduction, regulate DNP action and thus cardiac glucose uptake. Altered glucose uptake in hyperketonemic states during metabolic stress may contribute to diabetic cardiomyopathy.     

Abnormal muscle and hematopoietic gene expression may be important for clinical morbidity in primary hyperparathyroidism

In primary hyperparathyroidism (PHPT), excess PTH secretion by adenomatous or hyperplastic parathyroid glands leads to elevated serum [Ca(2+)]. Patients present complex symptoms of muscular fatigue, various neuropsychiatric, neuromuscular, and cardiovascular manifestations, and, in advanced disease, kidney stones and metabolic bone disease. Our objective was to characterize changes in muscle and hematopoietic gene expression in patients with reversible mild PHPT after parathyroidectomy and possibly link molecular pathology to symptoms. Global mRNA profiling using Affymetrix gene chips was carried out in biopsies obtained before and 1 yr after parathyroidectomy in seven patients discovered by routine blood [Ca(2+)] screening. The tissue distribution of PTH receptor (PTHR1 and PTHR2) mRNAs were quantitated using real-time RT-PCR in unrelated persons to define PTH target tissues. Of about 10,000 expressed genes, 175 muscle, 169 hematological, and 99 bone-associated mRNAs were affected. Notably, the major part of muscle-related mRNAs was increased whereas hematological mRNAs were predominantly decreased during disease. Functional and molecular network analysis demonstrated major alterations of several tissue characteristic groups of mRNAs as well as those belonging to common cell signaling and major metabolic pathways. PTHR1 and PTHR2 mRNAs were more abundantly expressed in muscle and brain than in hematopoietic cells. We suggest that sustained stimulation of PTH receptors present in brain, muscle, and hematopoietic cells have to be considered as one independent, important cause of molecular disease in PHPT leading to profound alterations in gene expression that may help explain symptoms like muscle fatigue, cardiovascular pathology, and precipitation of psychiatric illness.