Transmission Fourier transform infrared microspectroscopy allows simultaneous assessment of cutin and cell-wall polysaccharides of Arabidopsis petals.

A procedure for the simultaneous analysis of cell-wall polysaccharides, amides and aliphatic polyesters by transmission Fourier transform infrared microspectroscopy (FTIR) has been established for Arabidopsis petals. The combination of FTIR imaging with spectra derivatization revealed that petals, in contrast to other organs, have a characteristic chemical zoning with high amount of aliphatic compounds and esters in the lamina and of polysaccharides in the stalk of the petal. The hinge region of petals was particular rich in amides as well as in vibrations potentially associated with hemicellulose. In addition, a number of other distribution patterns have been identified. Analyses of mutants in cutin deposition confirmed that vibrations of aliphatic compounds and esters present in the lamina were largely associated with the cuticular polyester. Calculation of spectrotypes, including the standard deviation of intensities, allowed detailed comparison of the spectral features of various mutants. The spectrotypes not only revealed differences in the amount of polyesters in cutin mutants, but also changes in other compound classes. For example, in addition to the expected strong deficiencies in polyester content, the long-chain acyl CoA synthase 2 mutant showed increased intensities of vibrations in a wavelength range that is typical for polysaccharides. Identical spectral features were observed in quasimodo2, a cell-wall mutant of Arabidopsis with a defect in pectin formation that exhibits increased cellulose synthase activity. FTIR thus proved to be a convenient method for the identification and characterization of mutants affected in the deposition of cutin in petals.

Calcium antagonists as first-line therapy in hypertension: results of the Swiss Isradipine Study. Swiss Hypertension Society

In this study of the efficacy and safety of isradipine as first-line therapy in hypertension, 1,647 patients enrolled; 1,472 completed the 4-week placebo run-in period and began treatment with isradipine at 2.5 mg twice daily for 4 weeks. During placebo, 11% (n = 175) of the 1,647 patients withdrew because of normalization of blood pressure, side effects, noncompliance, violation of the study protocol, side effects from concomitant therapy, or other reasons. During isradipine therapy (n = 1,376), blood pressure decreased from 168 +/- 18/102 +/- 8 mm Hg at the end of the placebo period to 155 +/- 17/94 +/- 9 mm Hg after 2 weeks (p less than 0.001) and 151 +/- 16/92 +/- 9 mm Hg after 4 weeks (p less than 0.001). During active treatment, 6.4% (n = 94) were withdrawn because of flushing, headache, edema, palpitations, gastrointestinal side effects, skin rashes, or other side effects, and two patients because of lack of efficacy. The side effect score in the remaining patients worsened for flushing, remained unchanged for edema, but significantly improved for palpitations, fatigue, dizziness, headache, and nervousness. After 4 weeks, 60% of patients had diastolic blood pressures of less than or equal to 90 mm Hg. Thus, isradipine is effective and safe as first-line therapy in patients with primary hypertension as seen in general practice.

Effect of age on intraoperative cerebrovascular autoregulation and near-infrared spectroscopy-derived cerebral oxygenation.

Background. Age is an important risk factor for perioperative cerebral complications such as stroke, postoperative cognitive dysfunction, and delirium. We explored the hypothesis that intraoperative cerebrovascular autoregulation is less efficient and brain tissue oxygenation lower in elderly patients, thus, increasing the vulnerability of elderly brains to systemic insults such as hypotension.Methods. We monitored intraoperative cerebral perfusion in 50 patients aged 18-40 and 77 patients >65 yr at two Swiss university hospitals. Mean arterial pressure (MAP) was measured continuously using a plethysmographic method. An index of cerebrovascular autoregulation (Mx) was calculated based on changes in transcranial Doppler flow velocity due to changes in MAP. Cerebral oxygenation was assessed by the tissue oxygenation index (TOI) using near-infrared spectroscopy. End-tidal CO(2), O(2), and sevoflurane concentrations and peripheral oxygen saturation were recorded continuously. Standardized anaesthesia was administered in all patients (thiopental, sevoflurane, fentanyl, atracurium).Results. Autoregulation was less efficient in patients aged >65 yr [by 0.10 (SE 0.04; P=0.020)] in a multivariable linear regression analysis. This difference was not attributable to differences in MAP, end-tidal CO2, or higher doses of sevoflurane. TOI was not significantly associated with age, sevoflurane dose, or Mx but increased with increasing flow velocity [by 0.09 (SE 0.04; P=0.028)] and increasing MAP [by 0.11 (SE 0.05; P=0.043)].Conclusions. Our results do not support the hypothesis that older patients' brains are more vulnerable to systemic insults. The difference of autoregulation between the two groups was small and most likely clinically insignificant.

Staphylococcal biofilm formation on the surface of three different calcium phosphate bone grafts: a qualitative and quantitative in vivo analysis.

Differences in physico-chemical characteristics of bone grafts to fill bone defects have been demonstrated to influence in vitro bacterial biofilm formation. Aim of the study was to investigate in vivo staphylococcal biofilm formation on different calcium phosphate bone substitutes. A foreign-body guinea-pig infection model was used. Teflon cages prefilled with β-tricalcium phosphate, calcium-deficient hydroxyapatite, or dicalcium phosphate (DCP) scaffold were implanted subcutaneously. Scaffolds were infected with 2 × 10(3) colony-forming unit of Staphylococcus aureus (two strains) or S. epidermidis and explanted after 3, 24 or 72 h of biofilm formation. Quantitative and qualitative biofilm analysis was performed by sonication followed by viable counts, and microcalorimetry, respectively. Independently of the material, S. aureus formed increasing amounts of biofilm on the surface of all scaffolds over time as determined by both methods. For S. epidermidis, the biofilm amount decreased over time, and no biofilm was detected by microcalorimetry on the DCP scaffolds after 72 h of infection. However, when using a higher S. epidermidis inoculum, increasing amounts of biofilm were formed on all scaffolds as determined by microcalorimetry. No significant variation in staphylococcal in vivo biofilm formation was observed between the different materials tested. This study highlights the importance of in vivo studies, in addition to in vitro studies, when investigating biofilm formation of bone grafts.

ATP release due to Thy-1-integrin binding induces P2X7-mediated calcium entry required for focal adhesion formation.

Thy-1, an abundant mammalian glycoprotein, interacts with αvβ3 integrin and syndecan-4 in astrocytes and thus triggers signaling events that involve RhoA and its effector p160ROCK, thereby increasing astrocyte adhesion to the extracellular matrix. The signaling cascade includes calcium-dependent activation of protein kinase Cα upstream of Rho; however, what causes the intracellular calcium transients required to promote adhesion remains unclear. Purinergic P2X7 receptors are important for astrocyte function and form large non-selective cation pores upon binding to their ligand, ATP. Thus, we evaluated whether the intracellular calcium required for Thy-1-induced cell adhesion stems from influx mediated by ATP-activated P2X7 receptors. Results show that adhesion induced by the fusion protein Thy-1-Fc was preceded by both ATP release and sustained intracellular calcium elevation. Elimination of extracellular ATP with Apyrase, chelation of extracellular calcium with EGTA, or inhibition of P2X7 with oxidized ATP, all individually blocked intracellular calcium increase and Thy-1-stimulated adhesion. Moreover, Thy-1 mutated in the integrin-binding site did not trigger ATP release, and silencing of P2X7 with specific siRNA blocked Thy-1-induced adhesion. This study is the first to demonstrate a functional link between αvβ3 integrin and P2X7 receptors, and to reveal an important, hitherto unanticipated, role for P2X7 in calcium-dependent signaling required for Thy-1-stimulated astrocyte adhesion.

The heat shock response in moss plants is regulated by specific calcium-permeable channels in the plasma membrane.

Land plants are prone to strong thermal variations and must therefore sense early moderate temperature increments to induce appropriate cellular defenses, such as molecular chaperones, in anticipation of upcoming noxious temperatures. To investigate how plants perceive mild changes in ambient temperature, we monitored in recombinant lines of the moss Physcomitrella patens the activation of a heat-inducible promoter, the integrity of a thermolabile enzyme, and the fluctuations of cytoplasmic calcium. Mild temperature increments, or isothermal treatments with membrane fluidizers or Hsp90 inhibitors, induced a heat shock response (HSR) that critically depended on a preceding Ca(2+) transient through the plasma membrane. Electrophysiological experiments revealed the presence of a Ca(2+)-permeable channel in the plasma membrane that is transiently activated by mild temperature increments or chemical perturbations of membrane fluidity. The amplitude of the Ca(2+) influx during the first minutes of a temperature stress modulated the intensity of the HSR, and Ca(2+) channel blockers prevented HSR and the onset of thermotolerance. Our data suggest that early sensing of mild temperature increments occurs at the plasma membrane of plant cells independently from cytosolic protein unfolding. The heat signal is translated into an effective HSR by way of a specific membrane-regulated Ca(2+) influx, leading to thermotolerance.

Individual responses to converting enzyme inhibitors and calcium antagonists.

This study was designed to assess whether the acute blood pressure response of an individual hypertensive patient to a calcium antagonist or an angiotensin converting enzyme (ACE) inhibitor is a good predictor of the long-term efficacy of these drug classes in this particular patient. The concept that good responses to ACE inhibitors and calcium antagonists may be mutually exclusive was also tested. Sixteen patients were included in a randomized crossover trial of enalapril, 20 mg daily, and diltiazem, 120 mg daily, for 6 weeks each. Blood pressure was measured by ambulatory blood pressure recording. During the washout phase, the acute effect of nifedipine, 10 mg p.o., and enalaprilat, 5 mg i.v., was evaluated. Nifedipine and enalaprilat reduced blood pressure equally well. The long-term blood pressure reduction induced by enalapril and diltiazem was similar. The acute blood pressure response to a given drug was not a good predictor of the result obtained with long-term therapy. No age dependency of the antihypertensive effect of either drug class was apparent. There was no evidence that a good response to one drug excluded a similarly good response to the other.

Patterns of calcium-binding proteins support parallel and hierarchical organization of human auditory areas.

The human primary auditory cortex (AI) is surrounded by several other auditory areas, which can be identified by cyto-, myelo- and chemoarchitectonic criteria. We report here on the pattern of calcium-binding protein immunoreactivity within these areas. The supratemporal regions of four normal human brains (eight hemispheres) were processed histologically, and serial sections were stained for parvalbumin, calretinin or calbindin. Each calcium-binding protein yielded a specific pattern of labelling, which differed between auditory areas. In AI, defined as area TC [see C. von Economo and L. Horn (1930) Z. Ges. Neurol. Psychiatr.,130, 678-757], parvalbumin labelling was dark in layer IV; several parvalbumin-positive multipolar neurons were distributed in layers III and IV. Calbindin yielded dark labelling in layers I-III and V; it revealed numerous multipolar and pyramidal neurons in layers II and III. Calretinin labelling was lighter than that of parvalbumin or calbindin in AI; calretinin-positive bipolar and bitufted neurons were present in supragranular layers. In non-primary auditory areas, the intensity of labelling tended to become progressively lighter while moving away from AI, with qualitative differences between the cytoarchitectonically defined areas. In analogy to non-human primates, our results suggest differences in intrinsic organization between auditory areas that are compatible with parallel and hierarchical processing of auditory information.

Differential vulnerability of neurochemically identified subpopulations of retinal neurons in a monkey model of glaucoma.

The vulnerability of subpopulations of retinal neurons delineated by their content of cytoskeletal or calcium-binding proteins was evaluated in the retinas of cynomolgus monkeys in which glaucoma was produced with an argon laser. We quantitatively compared the number of neurons containing either neurofilament (NF) protein, parvalbumin, calbindin or calretinin immunoreactivity in central and peripheral portions of the nasal and temporal quadrants of the retina from glaucomatous and fellow non-glaucomatous eyes. There was no significant difference between the proportion of amacrine, horizontal and bipolar cells labeled with antibodies to the calcium-binding proteins comparing the two eyes. NF triplet immunoreactivity was present in a subpopulation of retinal ganglion cells, many of which, but not all, likely correspond to large ganglion cells that subserve the magnocellular visual pathway. Loss of NF protein-containing retinal ganglion cells was widespread throughout the central (59-77% loss) and peripheral (96-97%) nasal and temporal quadrants and was associated with the loss of NF-immunoreactive optic nerve fibers in the glaucomatous eyes. Comparison of counts of NF-immunoreactive neurons with total cell loss evaluated by Nissl staining indicated that NF protein-immunoreactive cells represent a large proportion of the cells that degenerate in the glaucomatous eyes, particularly in the peripheral regions of the retina. Such data may be useful in determining the cellular basis for sensitivity to this pathologic process and may also be helpful in the design of diagnostic tests that may be sensitive to the loss of the subset of NF-immunoreactive ganglion cells.

Serum calcium levels are associated with novel cardiometabolic risk factors in the population-based CoLaus study.

BACKGROUND: Associations of serum calcium levels with the metabolic syndrome and other novel cardio-metabolic risk factors not classically included in the metabolic syndrome, such as those involved in oxidative stress, are largely unexplored. We analyzed the association of albumin-corrected serum calcium levels with conventional and non-conventional cardio-metabolic risk factors in a general adult population. METHODOLOGY/PRINCIPAL FINDINGS: The CoLaus study is a population-based study including Caucasians from Lausanne, Switzerland. The metabolic syndrome was defined using the Adult Treatment Panel III criteria. Non-conventional cardio-metabolic risk factors considered included: fat mass, leptin, LDL particle size, apolipoprotein B, fasting insulin, adiponectin, ultrasensitive CRP, serum uric acid, homocysteine, and gamma-glutamyltransferase. We used adjusted standardized multivariable regression to compare the association of each cardio-metabolic risk factor with albumin-corrected serum calcium. We assessed associations of albumin-corrected serum calcium with the cumulative number of non-conventional cardio-metabolic risk factors. We analyzed 4,231 subjects aged 35 to 75 years. Corrected serum calcium increased with both the number of the metabolic syndrome components and the number of non-conventional cardio-metabolic risk factors, independently of the metabolic syndrome and BMI. Among conventional and non-conventional cardio-metabolic risk factors, the strongest positive associations were found for factors related to oxidative stress (uric acid, homocysteine and gamma-glutamyltransferase). Adiponectin had the strongest negative association with corrected serum calcium. CONCLUSIONS/SIGNIFICANCE: Serum calcium was associated with the metabolic syndrome and with non-conventional cardio-metabolic risk factors independently of the metabolic syndrome. Associations with uric acid, homocysteine and gamma-glutamyltransferase were the strongest. These novel findings suggest that serum calcium levels may be associated with cardiovascular risk via oxidative stress.

PTH and 1.25 vitamin D response to a low-calcium diet is associated with bone mineral density in renal stone formers.

BACKGROUND: Renal calcium stones and hypercalciuria are associated with a reduced bone mineral density (BMD). Therefore, the effect of changes in calcium homeostasis is of interest for both stones and bones. We hypothesized that the response of calciuria, parathyroid hormone (PTH) and 1.25 vitamin D to changes in dietary calcium might be related to BMD. METHODS: A single-centre prospective interventional study of 94 hyper- and non-hypercalciuric calcium stone formers consecutively retrieved from our stone clinic. The patients were investigated on a free-choice diet, a low-calcium diet, while fasting and after an oral calcium load. Patient groups were defined according to lumbar BMD (z-score) obtained by dual X-ray absorptiometry (group 1: z-score <-0.5, n = 30; group 2: z-score -0.5-0.5, n = 36; group 3: z-score >0.5, n = 28). The effect of the dietary interventions on calciuria, 1.25 vitamin D and PTH in relation to BMD was measured. RESULTS: An inverse relationship between BMD and calciuria was observed on all four calcium intakes (P = 0.009). On a free-choice diet, 1.25 vitamin D and PTH levels were identical in the three patient groups. However, the relative responses of 1.25 vitamin D and PTH to the low-calcium diet were opposite in the three groups with the highest increase of 1.25 vitamin D in group 1 and the lowest in group 3, whereas PTH increase was most pronounced in group 3 and least in group 1. CONCLUSION: Calcium stone formers with a low lumbar BMD exhibit a blunted response of PTH release and an apparently overshooting production of 1.25 vitamin D following a low-calcium diet.

Calcium-sensing receptors modulate renin release in vivo and in vitro in the rat.

OBJECTIVES: Calcium-sensing receptors (CaSRs) have been localized in the juxtaglomerular apparatus where they may contribute to the regulation of renin release. In the present study, we investigated the in-vitro and in-vivo effects of the calcimimetic R-568 on renin release. METHODS: In vitro, the effect of calcimimetics on renin release was assessed by incubating freshly isolated rat juxtaglomerular cells with or without R-568 (1 and 10 mumol/l) in serum-free medium in the presence or absence of forskolin or CaCl2. In vivo, we measured the impact of R-568 (20 ng/min intravenously) on the acute changes in plasma renin activity (PRA) induced by either a 90 min infusion of the angiotensin-converting enzyme inhibitor captopril, or the beta-receptor agonist isoproterenol, or of a vehicle in or after a furosemide challenge in conscious Wistar rats. RESULTS: In vitro, R-568 dose-dependently blunted renin release, but also reduced the increase in renin due to forskolin (P < 0.01). Both isoproterenol and enalapril increased in vivo PRA to 3.1 +/- 0.3 and 3.7 +/- 0.5 ng Ang I/ml per h, respectively (P < 0.01), compared with vehicle (1.5 +/- 0.2 ng Ang I/ml per h). R-568 significantly reduced PRA to 2.1 +/- 0.1 ng/ml per h in isoproterenol-treated rats and to 1.6 +/- 0.2 ng/ml per h in enalapril-treated rats (P < 0.05). In low-salt treated animals, acute infusion of furosemide increased PRA from 8.7 +/- 3.2 to 18.6 +/- 2.3, whereas R-568 partially blunted this rise to 11.2 +/- 1.5 (P = 0.02). In vivo, R-568 significantly lowered serum calcium and PTH1-84, but the drug-induced changes in PRA were independent of the changes in calcium and parathyroid hormone. CONCLUSION: After the recent discovery of CaSRs in juxtaglomerular cells of mice, our results confirm the presence of such receptors in rats and demonstrate that these receptors modulate renin release both in vitro and in vivo. This suggests that CaSRs play a role as a regulatory pathway of renin release.