Behavior of ordered sodium in enzymatically depleted cartilage tissue.

The onset of cartilage tissue disorders can be characterized by a loss of proteoglycans (PGs) and diagnosed by contrast-enhanced proton ((1)H) MRI techniques, as well as sodium MRI. The behavior of sodium located in anisotropic environments, is examined as a function of cartilage degeneration. PGs are proteolytically depleted from the cartilage samples, which gives rise to a decrease of the ordered sodium content. More surprisingly, however, the residual quadrupolar couplings are shown to increase with increasing depletion levels. Since the residual quadrupolar couplings are intimately related to local order and anisotropic motion, measuring their distribution in cartilage may provide insight into the structural changes that occur within the tissue upon degradation. In this study relatively mild orientational dependence of the couplings was found. Little or no free sodium was observed in the cartilage specimens under study.     

Hypophyseal-gonadal system during male aging

The concentration of sex and gonadotropic hormones in blood plasma of 280 reasonably healthy men aged 20-105 was determined using radioimmunoassay kits. Compared to men aged 20-39, the statistically significant decrease in testosterone level was registered in men aged 55-59, the increase in oestradiol in men aged 60-64, progesterone in men aged 55-59, and in LH and FSH in the group aged 65-69. The reactivity of central and peripheral links of the hypothalamic-gonadal system to direct and feed-back control influences alters with age.     

A dual role for interleukin-1 in LTP in mouse hippocampal slices

Interleukin-1 (IL-1) exerts numerous effects in the central nervous system and has been implicated in synaptic plasticity. The objective of this study was to investigate the role of endogenous as well as exogenous IL-1 on long-term potentiation (LTP). Hippocampal slices incubated at 34-36 degrees C show enhanced levels of IL-1alpha and IL-1beta compared to slices incubated at 21-24 degrees C. IL-1 inhibits LTP induced by theta-burst stimulation (TBS) at either temperature. IL-1 receptor antagonist (IL-1ra) had no effect on LTP at 21-24 degrees C, but displayed a concentration-dependent inhibition of LTP at 34-36 degrees C. Under control conditions, the magnitude of LTP was not temperature dependent. These data suggest that IL-1 is required for LTP under physiological conditions but at higher doses, as encountered in pathological conditions, IL-1 inhibits LTP.     

Heating of non-ferromagnetic total hip endoprostheses during magnetic resonance imaging in an animal model

To determine the heating effects of magnetic resonance imaging (MRI) on non-ferromagnetic total hip endoprostheses, hip endoprostheses were tested in vitro and in an ex vivo animal model. The MRI protocol on a 1.5 T scanner consisted of five sequences with a total duration of 23 min. The maximum heating of prosthetic material was 0.17 K in vitro and 0.90 K ex vivo; the maximum heating of bone surrounding tissue was 0.73 K ex vivo. Slight heating of the prostheses and the surrounding tissue was noted, which should not have any side effects in patients with titanium total hip endoprostheses.     

Characterizing folding, structure, molecular interactions and ligand gated activation of single sodium/proton antiporters

Using the example of sodium/proton antiporter from Escherichia coli NhaA, we review the capabilities of single-molecule atomic force microscopy and force spectroscopy to observe structural and functional insights of a membrane protein, which are not attainable by other traditional methods. While atomic force microscopy provides high-resolution topographs of single membrane proteins, their oligomeric state and assembly, single-molecule force spectroscopy experiments detect molecular interactions of the protein. The sensitivity of this method makes it possible to detect and locate interactions that stabilize secondary structures such as transmembrane alpha-helices, polypeptide loops and segments within them. Controlled refolding experiments using single-molecule force spectroscopy observed individual secondary structure segments folding into the functional protein. Various folding pathways of NhaA were detected, each one exhibiting a certain probability to be taken. Time-lapse refolding experiments enabled determining the folding kinetics and hierarchy of individual secondary structural elements. Recent examples detected and located the ligand binding of an antiporter. Similarly, inhibitor binding and location can be detected which in future may guide towards comparative studies of agonist and antagonist altering the functional state of a membrane protein. We review current and future potentials of these approaches to characterize the action of pharmacological molecules on the antiporter function.     

Mechanisms of neurohumoral regulation of heart function in aging.

Changes in hemodynamics and myocardial contractile capacity following the stimulation of extracardial nerves and administration of various doses of acetylcholine, norepinephrine, propranolol, phentolamin, atropine, serpasil and benzohexonium were studied in albino rats, rabbits and cats of various age. The shifts in cardiac acetylcholine and norepinephrine content and the activity of some enzymes of their catabolism were determined as well. Functional changes were compared with the structural shifts in intramural nervous system. In old animals there was a rise in threshold of voltage, which induced threshold reaction at stimulation of n. vagus and n. sympaticus and there was a change in heart sensitivity to acetycholine and norepinephrine, propranolol, atropine, serpasil and benzohexonium. A relationship was shown between age changes in nervous regulation of heart and transmitter metabolism. The intensity of acetylcholine and norepinephrine synthesis was decreased in old age, the transmitter metabolism in various parts of the heart was not uniformly altered, the sensitivity of epicardial chemoreceptors to nicotine, veratrum, acetylcholine was increased. The reflexes from cardiac mechanoreceptors were diminished.     

Central auditory processing disorder (CAPD) in children with specific language impairment (SLI). Central auditory tests

The aim of this project is to use central auditory tests for diagnosis of central auditory processing disorder (CAPD) in children with specific language impairment (SLI), in order to confirm relationship between speech-language impairment and central auditory processing. We attempted to establish special dichotic binaural tests in Czech language modified for younger children. Tests are based on behavioral audiometry using dichotic listening (different auditory stimuli that presented to each ear simultaneously). The experimental tasks consisted of three auditory measures (test 1-3)-dichotic listening of two-syllable words presented like binaural interaction tests. Children with SLI are unable to create simple sentences from two words that are heard separately but simultaneously. Results in our group of 90 pre-school children (6-7 years old) confirmed integration deficit and problems with quality of short-term memory. Average rate of success of children with specific language impairment was 56% in test 1, 64% in test 2 and 63% in test 3. Results of control group: 92% in test 1, 93% in test 2 and 92% in test 3 (p<0.001). Our results indicate the relationship between disorders of speech-language perception and central auditory processing disorders.     

Evidence of the milder phenotypic spectrum of c.1582G>A PIGT variant: Delineation based on seven novel Polish patients

PIGT is one of over 29 glycosylphosphatidylinositol biosynthesis defect genes. Mutations cause genetically determined disorders characterized mainly by epilepsy with fever-sensitivity, central hypotonia, psychomotor delay and congenital malformations. The disease is known as multiple congenital anomalies-hypotonia-seizures syndrome 3 (MCAHS3) or glycosylphosphatidylinositol biosynthesis defect-7. Twenty-eight cases have been reported until today. We present seven novel Polish patients, all harboring 1582G>A variant in a homozygous or compound heterozygous state which seems to cause a milder phenotype of the disease.     

Phosphates form spectroscopically dark state assemblies in common aqueous solutions

Phosphates and polyphosphates play ubiquitous roles in biology as integral structural components of cell membranes and bone, or as vehicles of energy storage via adenosine triphosphate and phosphocreatine. The solution phase space of phosphate species appears more complex than previously known. We present nuclear magnetic resonance (NMR) and cryogenic transmission electron microscopy (cryo-TEM) experiments that suggest phosphate species including orthophosphates, pyrophosphates, and adenosine phosphates associate into dynamic assemblies in dilute solutions that are spectroscopically “dark.” Cryo-TEM provides visual evidence of the formation of spherical assemblies tens of nanometers in size, while NMR indicates that a majority population of phosphates remain as unassociated ions in exchange with spectroscopically invisible assemblies. The formation of these assemblies is reversibly and entropically driven by the partial dehydration of phosphate groups, as verified by diffusion-ordered spectroscopy (DOSY), indicating a thermodynamic state of assembly held together by multivalent interactions between the phosphates. Molecular dynamics simulations further corroborate that orthophosphates readily cluster in aqueous solutions. This study presents the surprising discovery that phosphate-containing molecules, ubiquitously present in the biological milieu, can readily form dynamic assemblies under a wide range of commonly used solution conditions, highlighting a hitherto unreported property of phosphate’s native state in biological solutions.

Phosphate-containing species are in constant flux throughout the phosphorus cycle and accumulate within the cells of all living organisms. Cellular energy is primarily harvested through the dynamical formation and breakage of phosphoanhydride chemical bonds of adenosine phosphates (1, 2). Free phosphates and their subsequent assembly are also involved in bone formation and growth (3–5); however, the underlying assembly mechanisms of phosphate species and other ions that lead to bone formation processes are not well understood. An understanding of the equilibrium between free phosphates and higher-order phosphate assemblies in the form of polyphosphates and phosphate clusters would provide further insight into the mechanisms involving biological energy storage and/or the engineering of biological structures.³¹P nuclear magnetic resonance (NMR) offers useful information about the composition, dynamics, and structural properties of lipid membrane interfaces (6–8), phosphorylated biomolecules (9–11), polyphosphates (12, 13), and precursors of bone formation (14). We performed ³¹P NMR to investigate the native state of phosphate species as a function of temperature with the initial intent to subsequently study the formation processes of calcium phosphate clusters. In this process, we encountered peculiar ³¹P NMR line broadening with increasing temperature of aqueous solution of pure phosphates. Such characteristics cannot be explained by the usual temperature-dependent T₂ relaxation due to increasing molecular tumbling of small molecules with increasing temperature. ³¹P NMR line broadening as a function of pH, phosphate concentration, and counter-cation species has been described in the literature (15–17); however, line broadening with increasing temperature has not been reported before.Underscoring these unexpected observations, we present experimental results showing that phosphate-containing species, including orthophosphate, pyrophosphate, and adenosine diphosphate assemble into hitherto unreported spectroscopically “dark” species, whose fractional population increases with increasing temperature. This observation is shown to be consistent with the dehydration entropy-driven formation of dynamic phosphate assemblies. ³¹P NMR chemical exchange saturation transfer (CEST) reveals that phosphates assemble into species with broad spectroscopic signatures, whosepopulation is in exchange with NMR-detectable phosphate species. A subpopulation of these assemblies is also observed in cryogenic transmission electron microscopy (cryo-TEM) images to exhibit droplet-like spherical assemblies up to 50 nm in diameter. The discovery that common phosphate-containing molecules can readily assemble into higher-order species in water under physiological conditions in the absence of biologically activated processes should be relevant to a variety of biological and biochemical processes that use phosphate-containing species as building blocks, energy sources, or reactants in an aqueous environment.