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The chemical composition of suberin in apoplastic barriers affects radial hydraulic conductivity differently in the roots of rice (Oryza sativa L. cv. IR64) and corn (Zea mays L. cv. Helix).

Apoplastic transport barriers in the roots of rice (Oryza sativa L. cv. IR64) and corn (Zea mays L. cv. Helix) were isolated enzymatically. Following chemical degradation (monomerization, derivatization), the amounts of aliphatic and aromatic suberin monomers were analysed quantitatively by gas chromatography and mass spectrometry. In corn, suberin was determined for isolated endodermal (ECW) and rhizo-hypodermal (RHCW) cell walls. In rice, the strong lignification of the central cylinder (CC), did not allow the isolation of endodermal cell walls. Similarly, exodermal walls could not be separated from the rhizodermal and sclerenchyma cell layers. Suberin analyses of ECW and RHCW of rice, thus, refer to either the entire CC or to the entire outer part of the root (OPR), the latter lacking the inner cortical cell layer. In both species, aromatic suberin was mainly composed of coumaric and ferulic acids. Aliphatic suberin monomers released from rice and corn belonged to five substance classes: primary fatty acids, primary alcohols, diacids, omega-hydroxy fatty acids, and 2-hydroxy fatty acids, with omega-hydroxy fatty acids being the most prominent substance class. Qualitative composition of aliphatic suberin of rice was different from that of corn; (i) it was much less diverse, and (ii) besides monomers with chain lengths of C(16), a second maximum of C(28) was evident. In corn, C(24) monomers represented the most prominent class of chain lengths. When suberin quantities were related to surface areas of the respective tissues of interest (hypodermis and/or exodermis and endodermis), exodermal cell walls of rice contained, on average, six-times more aliphatic suberin than those of corn. In endodermal cell walls, amounts were 34 times greater in rice than in corn. Significantly higher amounts of suberin detected in the apoplastic barriers of rice corresponded with a substantially lower root hydraulic conductivity (Lp(r)) compared with corn, when water flow was driven by hydrostatic pressure gradients across the apoplast. As the OPR of rice is highly porous and permeable to water, it is argued that this holds true only for the endodermis. The results imply that some caution is required when discussing the role of suberin in terms of an efficient transport barrier for water. The simple view that only the quantity of suberin present is important, may not hold. A more detailed consideration of both the chemical nature of suberins and of the microstructure of deposits is required, i.e. how suberins impregnate wall pores.

Cell Wall↗

Expression of the 12-oxophytodienoic acid 10,11-reductase gene in the compatible interaction between pea and fungal pathogen.

Suppressors produced by Mycosphaerella pinodes are glycopeptides to block pea defense responses induced by elicitors. A clone, S64, was isolated as cDNA for suppressor-inducible gene from pea epicotyls. The treatment of pea epicotyls with suppressor alone induced an increase of S64 mRNA within 1 h, and it reached a maximum level at 3 h after treatment. The induction was not affected by application of the elicitor, indicating that the suppressor has a dominant action to regulate S64 gene expression. S64 was also induced by inoculation with a virulent pathogen, M. pinodes, but not by inoculation with a non-pathogen, Ascochyta rabiei, nor by treatment with fungal elicitor. The deduced structure of S64 showed high homology to 12-oxophytodienoic acid reductase (OPR) in Arabidopsis thaliana. A recombinant protein derived from S64 had OPR activity, suggesting compatibility-specific activation of the octadecanoid pathway in plants. Treatment with jasmonic acid (JA) or methyl jasmonic acid, end products of the octadecanoid pathway, inhibited the elicitor-induced accumulation of PAL mRNA in pea. These results indicate that the suppressor-induced S64 gene expression leads to the production of JA or related compounds, which might contribute to the establishment of compatibility by inhibiting the phenylpropanoid biosynthetic pathway.

Acetates↗

Anti-arrhythmic effects of (-)-carnitine chloride and its acetyl analogue on canine late ventricular arrhythmia induced by ligation of the coronary artery as related to improvement of mitochondrial function.

Using the two-stage coronary ligation method, first described by Harris, (1950), anti-arrhythmic effects (AAE) of (-)-carnitine chloride (LCC) and acetyl (-)-carnitine chloride (ALCC) were studied in conscious unrestrained dogs in comparison with those of disopyramide (D). Two-stage ligation of the coronary artery resulted in a significant decrease in the myocardial free carnitine content. Intravenous administration of LCC (300 mg kg-1) and D (5 mg kg-1) suppressed the ventricular arrhythmia induced by coronary ligation after 24 hours. ALCC (300 mg kg-1) was found to be less potent. An improvement of the mitochondrial function (respiratory control index (RCI) and oxidative phosphorylation rate (OPR) ) was noted with LCC and ALCC and there was a linear correlation between AAE expressed as reduction of arrhythmic ratio and improvement in the OPR, whereas there was no improvement in mitochondrial function after D. Plasma carnitine concentration was increased after administration of LCC, attaining the value of around 8 mM at 10 min after 300 mg kg-1. At 60 min, the plasma carnitine concentration was still about half as high as at 10 min. After ALCC, both acetyl carnitine and free carnitine were found in the plasma. The concentration of the former was decreased after attaining a peak value of around 0.2 mM at 10 min, while the plasma concentration of free carnitine was gradually increased. The anti-arrhythmic effects of LCC and ALCC were ascribed to the improvement of mitochondrial oxidative phosphorylation, while effects other than the improvement of the mitochondrial activity were suggested as mechanisms of anti-arrhythmic effects of D.

Acetylcarnitine↗

Differentiation of Aspergillus parasiticus from Aspergillus sojae by random amplification of polymorphic DNA.

Aspergillus parasiticus and Aspergillus sojae are two morphologically similar species belonging to the Aspergillus section flavi. A new method to distinguish the two species, the aflatoxin producer A. parasiticus and the koji mold A. sojae, was developed. Single primers with arbitrary sequences were used to generate random amplified polymorphic DNA (RAPD) markers from strains of these two species. Three decamers, OPA-04, OPB-10, and OPR-01, allowed adequate discrimination between strains of A. parasiticus and A. sojae in RAPD analyses. A. sojae was further separated into group I and group II with the three primers. On the other hand, A. parasiticus was divided into group A and group B when amplified with OPA-04 and OPR-10 primers. The previously misidentified stain CCRC 32423 and the misclassified strain CCRC 30227 were identified as Aspergillus flavus and A. sojae, respectively, on the basis of RAPD patterns and morphological characteristics. We suggest that the RAPD technique is a rapid and reliable tool to distinguish A. parasiticus from A. sojae.

Aspergillus↗

Multiplicity of oligopeptide transport systems in Escherichia coli.

The ability of Escherichia coli K-12 4212 to utilize a variety of oligopeptides as sources of required amino acids was examined. Triornithine-resistant mutants of this strain were oligopeptide permease deficient (Opp-) as judged by their inability to utilize (Lys)3 and (Lys)4 as sources of lysine and their resistance to the toxic tripeptide (Val)3. These same mutants were able to grow when Met-Met-Met, Met-Gly-Met, Met-Gly-Gly, Gly-Met-Gly, Gly-Gly-Met, Gly-Met-Met, Met-Met-Gly, or Leu-Leu-Leu were supplied in place of the requisite amino acid. The system mediating the uptake of these peptides, herein designated Opr I, was not able to transport N-alpha-acetylated peptides, nor the tetrapeptides Met-Gly-Met-Met, Met-Met-Gly-Met, or Met-Met-Met-Gly. Competition experiments indicated that trimethionine and trileucine enter E. coli K-12 via either Opp or Opr I. Analogous results were found using the methionine, leucine-requiring auxotroph E. coli B163. It appears that more than one oligopeptide transport system exists in E. coli and that the system mediating peptide uptake is complex.

Biological Transport, Active↗

The physiological role of relaxin in the pregnant rat. I. The influence of relaxin on parturition.

Length of gestation, duration of labor and delivery, and fetal survival were determined in control intact pregnant rats (group C). Pregnant rats were bilaterally ovariectomized on day 9 and given progesterone (P) implants and, in addition, one of the following injection regimens; estrogen (E; group OPE), E and porcine relaxin (R; group OPER), or porcine R (group OPR). Hormone treatments were given in doses designed to produce serum levels of these hormones similar to those observed in intact pregnant rats. The P implants were removed during the evening of day 21 to mimic the decline in serum P levels that normally occurs as a result of luteolysis. Animals in groups OPE and OPR exhibited significantly prolonged gestation, prolonged duration of labor and delivery, and reduced fetal survival compared with controls. Group OPER animals exhibited slightly but not significantly shorter length of gestation, similar duration of labor and delivery, and similar rate of fetal survival compared to control values. Group OPER animals showed normal maternal behavior and were able to suckle their young.

Animals↗

The physiological role of relaxin in the pregnant rat. II. The influence of relaxin on uterine contractile activity.

In control intact rats (group C), the frequency of intrauterine pressure cycles (IUPC) declines steadily during pregnancy from 80-130 cycles/3 h on day 10 (D10) to 20-30 cycles/3 h on D20. The decline in frequency is due to increasingly prolonged periods of myometrial quiescence, which increase from 30-90 min/3 h on D10 to 120-150 min/3 h by D20. During the 24 h preceding labor, the frequency of IUPC remains at less than 15 cycles/h until 3 h prepartum, when there is an abrupt increase to 30 cycles/h. Ovariectomized pregnant animals treated with progesterone (P) and estrogen (E; group OPE) exhibit significantly greater frequency of IUPC and significantly lower incidence of myometrial quiescence than intact pregnant rats from D12 and for the remainder of pregnancy. Ovariectomized pregnant rats that received P, E, and porcine relaxin (R: group OPER) or P and porcine R only (group OPR) exhibited declining frequency of IUPC similar to that observed in group C animals. Group OPER rats exhibited prolonged periods of myometrial quiescence of similar duration to those observed in control intact rats. In group OPR animals, however, the periods of myometrial quiescence were considerably diminished during late pregnancy. Group OPER animals exhibited a pattern of myometrial activity during labor and postpartum similar to that of control animals. The results suggest an important role for R in the control of myometrial activity during the second half of pregnancy and parturition in the rat.

Animals↗

Hormonal control of the cervix in pregnant gilts. II. Relaxin promotes changes in the physical properties of the cervix in ovariectomized hormone-treated pregnant gilts.

The effects of relaxin on the physical properties of both the uterine and vaginal portions of the cervix were studied between days 80 and 110 of pregnancy in ovariectomized gilts given progesterone to maintain pregnancy. In controls gilts the extensibility, lumen diameter, and wet weight of the uterine portion of the cervix were less than those of the vaginal portion of the cervix on day 80, but increased thereafter, and by day 110 did not differ (extensibility and wet weight) or approached (lumen diameter) those values of the vaginal portion of the cervix. When gilts were ovariectomized on either days 80 or 100 and given progesterone only (groups OP-100 and OP-110) extensibility, lumen diameter, and wet weight failed to increase in both the uterine and vaginal portions of the cervix. After replacement therapy with progesterone plus relaxin (groups OPR-100 and OPR-110), extensibility, lumen diameter, and wet weight increased to values as least as large as those in control groups in both portions of the cervix. In all groups the uterine portion of the cervix changed more dramatically than the vaginal portion. After hormone replacement therapy, blood levels of relaxin were moderately higher, and blood levels of progesterone were somewhat lower than those in control gilts. There was little or no difference in estrogen levels between ovariectomized and control gilts. We conclude that relaxin plays an important role in promoting increased growth and extensibility in both the uterine and vaginal portions of the cervix after day 80 of gestation in the gilt.

Animals↗

Effect of relaxin on parturition and frequency of live births in pigs.

Gilts on Day 105 of gestation were sham ovariectomized (Group C, N = 5); ovariectomized and given i.m. injections of 100 mg progesterone twice daily from Day 105 to 112 (Group OP, N = 5); or ovariectomized and given progesterone and i.m. injections of 1 mg highly purified porcine relaxin 4 times/day from Day 105 until the end of parturition (Group OPR, N = 5). Concentrations of progesterone in peripheral plasma of gilts in Group OP were similar to those in Group C. Relaxin was undetectable in peripheral plasma of gilts in Group OP which also showed prolonged parturition (P less than 0.001) and impaired frequency of live births (P less than 0.001), although onset of lactation was not affected. In Group OPR the duration of parturition and frequency of live births were similar to those observed for gilts in Group C. These results indicate that the ovarian hormone relaxin is necessary for normal duration of parturition and frequency of live births and that the onset of lactation is not prevented by an absence of relaxin.

Animals↗

[Olfactory dysfunction: its diagnosis and surgical treatment].

This paper discusses the results of diagnosis and surgical therapy of olfactory dysfunction. It describes a method of qualitative and quantitative olfactometry which involves olfactory investigation by means of an olfactory unit and an olfactometer based on the smelling principle. The olfactory unit and olfactometer permit qualitative and quantitative olfactory measurements. Objective olfactometry was performed in 34 patients using the galvanic skin reflex, olfactory-pupillary reflex (OPR), pneumography, and ECG. The method, developed by the present author for recording pupils responses by means of an optic pupillometer, allows differentiation of olfactory and trigeminal sensitivity during OPR examination. Objective olfactometry demonstrated that olfactory investigation with the aid of a custom built olfactometer gives reliable data. The procedure of endonasal operation of concha nasalis media in the olfactory fissure area in the case of hypertropic rhinitis accompanied by olfactory dysfunction is described.

Humans↗

[Synthesis of oligosaccharides related to HNK-1 antigen. 2. Synthesis of 3'''-O-(3-O-sulfo-beta-D-glucuronopyranosyl)-lacto-N-neotetrao se beta-propylglycoside].

A derivative of allyl 3"-O-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-beta-lactoside with a free OH group at C-4GlcNAc was glycosylated with trichloroacetimidate of selectively protected GlcA(beta 1-->3)Gal alpha disaccharide in dichloromethane in the presence of trimethylsilyl triflate resulting in a pentasaccharide product with an 82% yield. This product was converted to monohydroxy derivative with a free OH group at C-3GlcA via the formation and the subsequent opening of the 6,3-lactone ring in the glucuronic acid residue. The 3"'-O-sulfation of the monohydroxy derivative, the removal of the protective groups, and the reduction of the allyl aglycon yielded the pentasaccharide propyl glycoside NaSO3-3GlcA(beta 1-->3)Gal(beta 1-->4)GlcNAc(beta 1-->3)Gal(beta 1-->4)Glc beta-Opr comprising the oligosaccharide chain of the SGGL-1 glycolipid, which is recognized by HNK-1 antibodies. NaSO3-3GlcA(beta 1--> 3)Gal beta OAll, GlcA(beta 1-->3)Gal(beta 1-->4)GlcNAc(beta 1-->3)Gal(beta 1-->4)Glc beta-OPr and GlcA(beta 1-->3)Gal beta OAll were synthesized in a similar way.

CD57 Antigens↗

Establishment of a CRISPR-Cas9 Library for Indica Rice and Identification of OsOPR5 (LOC_Os06g11210) as a Regulator of Root Architecture.

Functional characterization of a large number of rice genes remains a major challenge despite the availability of genome sequences and large-scale transcriptomic datasets. CRISPR-Cas9 library is a powerful approach for high-throughput targeted mutagenesis; however, its application in indica rice cultivars remains limited due to low transformation and regeneration efficiencies. In this study, we developed a CRISPR-Cas9 library targeting 12,000 rice genes and evaluated its utility for functional genomics in the indica cultivar MTU-1010. Sanger sequencing and NGS analysis of the plasmid library revealed high sgRNA coverage and more than 80% accuracy. Transformation of the developed library into the indica cultivar MTU-1010 resulted in a high target editing efficiency, with 90% of analyzed transgenic plants carrying mutations at the intended target site. Functional analysis of one homozygous mutant identified a previously uncharacterized role for OsOPR5 (LOC_Os06g11210), a member of the 12-oxophytodienoate reductase family in root architecture. The opr5 mutants exhibited significant reductions in lateral root number, seminal and crown root number, and root length, demonstrating that OsOPR5 positively regulates root system architecture in rice. Notably, endogenous jasmonic acid (JA) and JA-isoleucine levels were not significantly altered in the mutant, suggesting potential functional specialization or redundancy among rice OPR family members for JA accumulation. The root system architecture is a key determinant of water and nutrient acquisition; our results suggest that OsOPR5 may play an important role in adaptation under adverse environmental conditions. Collectively, this study establishes an efficient genome-editing platform for indica rice and identifies OsOPR5 as a novel regulator of root development.

Oryza↗

Electrochemically induced sol-gel deposition of zirconia thin films.

A novel electrochemical method for deposition of ZrO(2) thin films is described. The films, 50-600 nm thick, were obtained by applying moderate positive or negative potentials (+2.5 V to -1.5 V versus SHE) on conducting surfaces immersed in a 2-propanol solution of zirconium tetra-n-propoxide [Zr(OPr)(4)] in the presence of minute quantities of water (water/monomer molar ratios in the range of 10(-5) to 10(-1)), which was the limiting reagent. Oxidative electrochemical formation of solvated H(+) and reductive formation of OH(-) catalyze the hydrolysis and condensation of the metal alkoxide precursor. The magnitude of the applied potential and its duration provide a convenient way of controlling the film thickness. The films consist of an amorphous phase, as revealed by XRD measurements. The effects of different parameters, such as the applied potential and its duration, the amount of added water and the current-time characteristics, were studied. A mechanism for the electrodeposition of the zirconia films which is in accordance with our findings is proposed.

Journal Article↗

Minimal heparin/heparan sulfate sequences for binding to fibroblast growth factor-1.

The glycosaminoglycans heparin and heparan sulfate (HS) bind to fibroblast growth factor FGF1 and promote its dimerization, a proposed prerequisite for binding to a cellular receptor and triggering mitogenic signals. The problem of minimal structural requirements for heparin/HS sequences to bind FGF1 was approached by surface plasmon resonance (SPR), NMR spectroscopy, and MALDI mass spectrometry studies using the three synthetic tetrasaccharides GlcNSO(3)6OR-IdoA2SO(3)-GlcNSO(3)6OR'-IdoA2SO(3)OPr (AA, R = R' = SO(3); BA, R = H, R' = SO(3); BB, R = R' = H; Pr, propyl). AA and BA significantly interact with the protein, whereas BB is practically inactive. The NMR spectra show that, whereas the interaction of AA primarily involves the GlcNSO(3)6SO(3)IdoA2SO(3) disaccharide moiety at its nonreducing end, residues at both the nonreducing (NR) and reducing side (R) appear to be involved in the weaker complex of BA. Furthermore, MALDI experiments show that, in addition to 1:1 protein:tetrasaccharide complexes, AA and BA are able to form 2:1 complexes, indicating that heparin/HS-induced dimerization of FGF1 requires only one 6-OSO(3) group per tetrasaccharide.

Binding, Competitive↗

Ozone chemonucleolysis in non-contained lumbar disc herniations: a pilot study with 12 months follow-up.

STUDY DESIGN: Prospective case series with six and twelve months follow up. OBJECTIVE: To observe clinical and morphological results of the intradiscal ozone chemionucleolysis in patients affected by non-contained lumbar disc herniations. METHODS: 30 patients were included in the study on the base of precise inclusion and exclusion criteria. The patients were followed on 6 and 12 months period by Visual Analogic Scale (VAS), Roland Morris Disability Questionnaire (RMDQ) and Overall Patient Rating Scale (OPRS). Disc herniation volume morphology was evaluated at 5 months by control MRI scanning. RESULTS: Twenty-seven patients (90%) showed a statistically significant improvement in pain (P < 0.001, Wilcoxon test) and function (P < 0.001, Wilcoxon test), on VAS and RMDQ evaluation, respectively. The mean satisfaction with the treatment on OPSR was 79.3%, with 24 patients referring satisfaction equal or greater than 80%. There were no major complications related to the procedure. CONCLUSIONS: The results of this study indicate the ozone chemonucleolysis as a possibly effective modality of treatment in patients affected by signs and symptoms of non-contained lumbar disc herniations that have overpassed conservative measures and have not yet fulfilled the indications for open surgical treatment.

Adult↗

The protective effect of desferal on rat myocardial mitochondria is not prolonged after withdrawal of desferal.

Reperfusion of ischaemic myocardium is necessary to sustain tissue viability (without it the tissue becomes necrotic), but reperfusion, on the other hand, can damage cells which have survived ischaemia. There is now considerable evidence that oxygen radicals, especially hydroxyl radicals produced via the Haber-Weiss and Fenton reactions, are responsible for reperfusion damage. Various investigators have reported that desferal, an iron chelator, has a beneficial effect on the myocardium during ischaemia and reperfusion. The aim of this study was two-fold: i) whether superoxide anions in the absence of LMWI can impair mitochondrial function, and ii) whether the protective effect of desferal on the mitochondrial function persists after withdrawal of desferal. Experiments were done on isolated rat hearts subjected to normothermic ischaemic cardiac arrest (NICA), with or without desferal, followed by 15-min reperfusion with desferal, followed by 15-min perfusion without desferal, or a hypoxanthine/xanthine oxidase medium that generates superoxide anions (with or without desferrioxamine (desferal) in the perfusate). Mitochondrial function (QO2 (state 3), ADP/O and OPR) as well as LMWI were measured. Our results indicated that i) superoxide anions and/or hydrogen peroxide can, independently of LMWI, damage the mitochondria, and ii) withdrawal of desferal after the respiratory burst resulted in the same or more severe mitochondrial damage than without any desferal.

Adenosine Diphosphate↗

The active analog approach applied to the pharmacophore identification of benzodiazepine receptor ligands.

Applied to seven potent benzodiazepine-receptor ligands belonging to chemically different classes, the active analog approach allowed the stepwise identification of the pharmacophoric pattern associated with the recognition by the benzodiazepine receptor. A unique pharmacophore model was derived which involves six critical zones: (a) a pi-electron rich aromatic (PAR) zone; (b) two electron-rich zones delta 1 and delta 2 placed at 5.0 and 4.5 A respectively from the reference centroid in the PAR zone; (c) a freely rotating aromatic ring (FRA) region; (d) an out-of-plane region (OPR), strongly associated with agonist properties; and (e) an additional hydrophobic region (AHR). The model accommodates all presently known ligands of the benzodiazepine receptor, identifies sensitivity to steric hindrance close to the delta 1 zone, accounts for R and S differential affinities and distinguishes requirements for agonist versus non-agonist activity profiles.

Carbolines↗

Characterization, application and potential uses of biotin-tagged inhibitors for lymphocyte serine proteases (granzymes).

Cytotoxic lymphocytes and natural killer cells are able to kill their target cells in minutes. The death of the target cell occurs after the release of cytoplasmic granules from the effector cell. These granules contain the pore-forming protein perforin and serine proteases (granzymes). To date 10 genes encoding lymphocyte granzymes have been discovered; of these only four have been purified and characterized for their substrate specificity. Several are predicted to have a common chymase, like specificity which is found in the granule extracts. Others may need to be enriched as active enzymes before they can be evaluated for substrate hydrolysis. Due to the limitations of detection by substrate hydrolysis, a more sensitive method for the detection of dilute granules was needed. We report the differing reactivities of seven biotin (Bi)-tagged isocoumarin (IC) inhibitors for Asp-ase, chymase, tryptase and Met-ase granzymes. The inhibitors contained different substituents at their no. 3 position: methoxy (OMe), ethoxy (OEt), propoxy (OPr) or 2-phenylethoxy (OEtPh) groups. The OMe group conferred general reactivity, whereas the OEtPh group conferred selective reactivity with chymase granzymes. The inhibitors that contained the longest aminocaproyl (Aca) spacers between the biotin-tag and the isocoumarin ring mediated the most stable granzyme inactivation. These inhibitors were the most effective at blocking lysis of red blood cells by the granule extracts. The inhibitors were used in protein blotting experiments where the biotin was detected with an avidin-enzyme complex. Over 10 granzymes were labelled by the inhibitor Bi-Aca-Aca-IC-OMe. The inhibitors detected granzymes when they were not readily detected by substrate hydrolysis.

Amino Acid Sequence↗