Treatment of inflammatory arthritis via targeting of tristetraprolin, a master regulator of pro-inflammatory gene expression.

OBJECTIVES: Tristetraprolin (TTP), a negative regulator of many pro-inflammatory genes, is strongly expressed in rheumatoid synovial cells. The mitogen-activated protein kinase (MAPK) p38 pathway mediates the inactivation of TTP via phosphorylation of two serine residues. We wished to test the hypothesis that these phosphorylations contribute to the development of inflammatory arthritis, and that, conversely, joint inflammation may be inhibited by promoting the dephosphorylation and activation of TTP. METHODS: The expression of TTP and its relationship with MAPK p38 activity were examined in non-inflamed and rheumatoid arthritis (RA) synovial tissue. Experimental arthritis was induced in a genetically modified mouse strain, in which endogenous TTP cannot be phosphorylated and inactivated. In vitro and in vivo experiments were performed to test anti-inflammatory effects of compounds that activate the protein phosphatase 2A (PP2A) and promote dephosphorylation of TTP. RESULTS: TTP expression was significantly higher in RA than non-inflamed synovium, detected in macrophages, vascular endothelial cells and some fibroblasts and co-localised with MAPK p38 activation. Substitution of TTP phosphorylation sites conferred dramatic protection against inflammatory arthritis in mice. Two distinct PP2A agonists also reduced inflammation and prevented bone erosion. In vitro anti-inflammatory effects of PP2A agonism were mediated by TTP activation. CONCLUSIONS: The phosphorylation state of TTP is a critical determinant of inflammatory responses, and a tractable target for novel anti-inflammatory treatments.

Effect of ultrasound-facilitated fixation on oral mucosa density and morphology.

We investigated the effects of ultrasound-facilitated fixation on oral mucosal morphology. Bovine dorsal tongue and porcine buccal (cheek) mucosa were sonicated for 0, 5, 10 or 15 min in a modified methacarn fixative, then incubated at 25° C for 4 h. Initial mass, volume and density of each specimen were measured before and after treatment and fixation. Morphometric analysis of the scanning electron micrographs was used to quantify changes in mucosal structure and microtexture. Statistical methods were used to describe the relation between sonication time, tissue density and relative change in tissue density. Our results indicate a linear correlation between sonication time and density of the dorsal tongue specimens. The treatment caused contraction of the tongue tissue and expansion of the buccal mucosa. Differences between initial and final tongue densities and the relative change in tissue density of the tongue vs. buccal mucosa were statistically significant. Differences observed between initial and final buccal mucosa densities were not statistically significant. Changes in buccal mucosa density correlated inversely with sonication time by contrast to the tongue density, which was correlated directly with this factor. Our study illustrates that while preservation of mucosal morphology and biopolymers can be achieved by ultrasound-facilitated fixation, its effects on tissue density are both time-dependent and specific to certain regions of the mouth.

Homochiral xanthine quintet networks self-assembled on Au(111) surfaces.

Xanthine molecule is an intermediate in nucleic acid degradation from the deamination of guanine and is also a compound present in the ancient solar system that is found in high concentrations in extraterrestrial meteorites. The self-assembly of xanthine molecules on inorganic surfaces is therefore of interest for the study of biochemical processes, and it may also be relevant to the fundamental understanding of prebiotic biosynthesis. Using a combination of high-resolution scanning tunneling microscopy (STM) and density functional theory (DFT) calculations, two new homochiral xanthine structures have been found on Au(111) under ultrahigh vacuum conditions. Xanthine molecules are found to be self-assembled into two extended homochiral networks tiled by two types of di-pentamer units and stabilized by intermolecular double hydrogen bonding. Our findings indicate that the deamination of guanine into xanthine leads to a very different base pairing potential and the chemical properties of the base which may be of relevance to the function of the cell and potential development of human diseases. Moreover, the adsorption of xanthine molecules on inorganic surfaces leading to homochiral assemblies may be of interest for the fundamental understanding of the emerged chirality at early stages of life.

A new form of myeloma "kidney": shortened hemofilter survival and implications for membrane filtration plasmapheresis.

BACKGROUND: Despite rigorous focus on extracorporeal CRRT parameters such as access, blood flow, hemoconcentration, and anticoagulation, some patients have unexpected repetitive hemofilter clotting. We instead explored patient or disease-related factors that could be responsible, and present a case of plasma cell dyscrasia in which paraproteins caused hollow fiber failure. METHODS: A patient with IgG kappa chain multiple myeloma complicated by sepsis and acute renal failure was started on CVVH with a regional citrate anticoagulation protocol that typically yields filter life of >50 h. Polysulfone hemofilters repetitively clotted every 2-4 h, even after excluding circuit-related problems. Failed filters were examined by light, electron (EM), and immunofluorescence (IF) microscopy. RESULTS: Imaging of the hemofilters revealed several ultrastructural features typical for myeloma-associated alterations in native human tissue. Red cell rouleaux formation occurred within the hollow fibers. There was extensive protein layering on the luminal surface of the fibers with some extension into their walls: these deposits were IF+ for IgG kappa, and had a fibrillary substructure on EM. CONCLUSION: Extracorporeal hollow fiber phenomena recapitulate many intra-corporeal paraprotein effects such as those described in the kidney with plasma cell dyscrasias. Rapid protein layering suggests fouling of the membrane, decreased solute clearance before total device failure, and raises the theoretical concern that this might also occur during filtration plasmapheresis: we thus suggest serial serum free light chain levels to confirm their removal when using that technique. These findings emphasize the importance of disease rather than circuit-related factors that are under-appreciated causes of premature hemofilter failure.

Intra-abdominal hypertension can be monitored with femoral vein catheters during CRRT and may cause access recirculation.

OBJECTIVES: Intra-abdominal hypertension (IAH) is an increasingly recognized disorder, and its diagnosis depends on accurate pressure monitoring. Bladder-based protocols are favored, but are not always clinically feasible. Abdominal venous (i.e. vena cava) pressure measurements are an alternative but are logistically challenging. We hypothesized that for patients suffering acute kidney injury, transducers built into renal replacement therapy (RRT) machines offer a simple opportunity to monitor pressures using catheters inserted via femoral veins. DESIGN: We performed in vitro testing of the accuracy of pressure transducers incorporated into continuous RRT devices, using highly calibrated instrumentation in the IAH-relevant range of 0 to +50 mmHg. We developed a protocol for using this modality in vivo, by stopping all pumps so as to allow equilibration of pressures: clinical application in a patient with femoral vein catheters and IAH was then described. RESULTS: In vitro analyses showed accuracy of the extracorporeal pressure transducers with an r² of 0.998, p < 0.001. In the patient case, the pressure transduced at the RRT device was identical to those obtained from bladder catheters. IAH also led to access recirculation and ineffective therapy. CONCLUSIONS: Pressure sensors incorporated into continuous RRT machines can be accurate in the IAH physiologic range, and thus may be used to easily measure intra-abdominal pressure via appropriate-length femoral vein-inserted access catheters. If not relieved, IAH can be an under-appreciated cause of access recirculation and ineffective clearance for any RRT modality (continuous or intermittent) using femoral catheters.

Homopairings of the artificial nucleobase 1H-benzoimidazole-4,7-dione.

All planar homopairings of the artificial nucleobase 1H-benzoimidazole-4,7-dione are reported for the first time in this study. Using the idea of binding sites discussed in our previous work and an ab initio density functional theory method we predict 13 homopairs. The stabilization energies of the homopairs vary from -0.13 to -0.69 eV. The collected data on all the planar homopairs reported here may be useful when constructing assemblies of this artificial base on various solid substrates.

Lanthanum carbonate vs. sevelamer hydrochloride for the reduction of serum phosphorus in hemodialysis patients: a crossover study.

AIMS: The aim of this crossover study was to compare the reduction of serum phosphorus (SP) with fixed doses of the non-calcium-containing phosphate binders lanthanum carbonate (LC) and sevelamer hydrochloride (SH) in hemodialysis patients. METHODS: Following washout (2 - 3 weeks), 182 patients with SP >or= 6.0 mg/dl and calcium >or= 8.4 mg/dl were randomized (1:1) to receive LC (2,250 to 3,000 mg/day) or SH (4,800 to 6,400 mg/day) for 4 weeks. Patients underwent a second washout (2 weeks) and switched to the alternative binder for 4 weeks. RESULTS: At the end of treatment, LC had reduced SP by 1.7 +/- 0.1 mg/dl, compared with 1.4 +/- 0.1 mg/dl for SH; the difference was not statistically significant in the primary analysis (LOCF, p = 0.133). However, the reduction with LC was significantly greater than with SH in a prespecified key secondary analysis of patients who completed 4 weeks of treatment with each binder (0.5 mg/dl difference, p = 0.007). The reduction of SP was also greater with LC than SH after 1 week of treatment (p = 0.024). CONCLUSIONS: Although the primary analysis found no difference between LC and SH in the reduction of SP, a significant difference in favor of LC was observed in patients who completed treatment. The results of this study show interesting trends with respect to onset and duration of action that warrant further investigation in longer-term studies.

Self-assembly of artificial nucleobase 1H-benzimidazole-4,7-dione at the liquid/solid interface.

Self-assembly at the liquid/solid interface of an electrochemically active DNA nucleobase analogue, 1H-benzoimidazole-4,7-dione (Q), has been studied by means of scanning tunneling microscopy (STM). High-resolution STM images revealed the formation of well-ordered two-dimensional (2D) supramolecular nanostructures when the Q molecules are adsorbed onto the graphite surface from a 1-octanol solution. Detailed analysis shows that the observed 2D nanostructures are mainly dominated by hydrogen-bonded Q molecules. Since Q can be considered as a molecule mimicking the nucleobase guanine (G), which is known to form Watson-Crick base pairs with its complementary nucleobase cytosine (C), we have examined the binding ability of Q with C realized by available hydrogen-bonding sites on both Q and C molecules. Upon deposition of a mixture of Q and C molecules onto a graphite surface, one might expect that hydrogen-bonded QC dimers were observed in a new 2D self-assembled structure governed by inter- and intramolecular hydrogen-bonding interactions between Q and C molecules. However, our STM experiments showed that no well-ordered structures are formed and instead phase separation occurs where large-scale homodomains are formed consisting of the individual QQ and CC dimers. To gain further insight into the possible molecular arrangements of the Q and C nucleobases in the mixture phase, the high-resolution STM images are compared with the results from ab initio density functional theory (DFT) calculations.

Bilateral renal-cell carcinoma associated with an acquired VHL mutation and long-term trichloroethylene exposure.

BACKGROUND/AIMS: The genetic basis for clear-cell renal carcinomas has been established in familial and many sporadic forms. Whether the latter can be induced by environmental carcinogens remains controversial, with concern over solvents such as trichloroethylene (TCE). To study this putative relationship, we analyzed the VHL gene from a patient with long-term TCE exposure. METHODS: PCR amplification and sequencing of VHL exons 1 - 3 were performed on peripheral blood and tumor tissue. RESULTS: The tumor alone had a previously undescribed mutation in exon 1 of the VHL gene: deletion of a cytidine residue at position 291 relative to the first ATG start codon of the wild-type sequence. This deletion causes a frameshift and predicts an altered protein sequence from position 98 onwards. CONCLUSION: The affected amino acids are in the functionally important beta-domain of the VHL protein that is implicated in substrate binding for ubiquitylation, and we hypothesize the mutation lowers that affinity. There is loss of suppressor function when substrates such as hypoxia-inducible factor have impaired degradation: they accumulate and ultimately cause uncontrolled cell turnover. This association of a proposed occupational cause and occurrence of renal-cell carcinoma emphasizes the availability and use of VHL sequencing for both studying the pathophysiology of malignant transformation and potentially playing a clinical role in genetic counseling or risk assessment.

Ultrafiltration for decompensated heart failure: renal implications.

The negative prognostic impact of worsening renal function in patients with decompensated heart failure has been widely recognised. As diuretics are thought to contribute to deterioration of kidney function in this setting, attempts have been made to either spare or suppress the diuretic-related pathophysiological mechanisms involved in this phenomenon. In this regard, extracorporeal ultrafiltration represents a novel therapy for patients with heart failure, lacking the adverse impacts of diuretics on kidney function (eg, activation of tubuloglomerular feedback). Consequently, besides its other positive clinical outcomes, there has been much hope for ultrafiltration therapy to play a protective role against negative effects of diuretics in patients with decompensated heart failure. However, the potential biological advantage has not been confirmed by clinical studies; currently available data from recent clinical trials have so far failed to demonstrate such expected positive results possibly due to counterbalance of the potential negative effects and other not well-known mechanisms. This paper briefly reviews the relevant pathophysiological mechanisms as well as existing evidence in this area and emphasises on the need for further studies specifically designed to explore the impact of ultrafiltration on kidney function in patients with decompensated heart failure.

Adenine monolayers on the Au(111) surface: structure identification by scanning tunneling microscopy experiment and ab initio calculations.

From an interplay between scanning tunneling microscopy (STM) and ab initio density functional theory (DFT) we have identified and characterized two different self-assembled adenine (A) structures formed on the Au(111) surface. The STM observations reveal that both structures have a hexagonal geometry in which each molecule forms double hydrogen bonds with three nearest neighbors. One of the A structures, with four molecules in the primitive cell, has p2gg space group symmetry, while the other one, with two molecules in the cell, has p2 symmetry. The first structure is observed more frequently and is found to be the dominating structure after annealing. Experimental as well as theoretical findings indicate that the interaction of A molecules with the gold surface is rather weak and smooth across the surface. This enabled us to unequivocally characterize the observed structures, systematically predict all structural possibilities, based on all known A-A dimers, and provisionally optimize positions of the A molecules in the cell prior to full-scale DFT calculations. The theoretical method is a considerable improvement compared to the approach suggested previously by Kelly and Kantorovich [Surf. Sci. 589, 139 (2005)]. We propose that the less ordered p2gg symmetry structure is observed more frequently due to kinetic effects during island formation upon deposition at room temperature.

Understanding the disorder of the DNA base cytosine on the Au(111) surface.

Using ultrahigh vacuum scanning tunneling microscopy (STM) and ab initio density functional theory, we have investigated in detail structures formed by cytosine on the Au(111) surface in clean ultrahigh vacuum conditions. In spite of the fact that the ground state of this DNA base on the surface is shown to be an ordered arrangement of cytosine one-dimensional branches (filaments), this structure has never been observed in our STM experiments. Instead, disordered structures are observed, which can be explained by only a few elementary structural motifs: filaments, five- and sixfold rings, which randomly interconnect with each other forming bent chains, T junctions, and nanocages. The latter may have trapped smaller structures inside. The formation of such an unusual assembly is explained by simple kinetic arguments as a liquid-glass transition.

An investigation into the interactions between self-assembled adenine molecules and a Au(111) surface.

Two molecular phases of the DNA base adenine (A) on a Au(111) surface are observed by using STM under ultrahigh-vacuum conditions. One of these phases is reported for the first time. A systematic approach that considers all possible gas-phase two-dimensional arrangements of A molecules connected by double hydrogen bonds with each other and subsequent ab initio DFT calculations are used to characterize and identify the two phases. The influence of the gold surface on the structure of A assemblies is also discussed. DFT is found to predict a smooth corrugation potential of the gold surface that will enable A molecules to move freely across the surface at room temperature. This conclusion remains unchanged if van der Waals interaction between A and gold is also approximately taken into account. DFT calculations of the A pairs on the Au(111) surface show its negligible effect on the hydrogen bonding between the molecules. These results justify the gas-phase analysis of possible assemblies on flat metal surfaces. Nevertheless, the fact that it is not the most stable gas-phase monolayer that is actually observed on the gold surface indicates that the surface still plays a subtle role, which needs to be properly addressed.

New ab initio potential energy surface and the vibration-rotation-tunneling levels of (H2O)2 and (D2O)2.

We report a new full-dimensional potential energy surface (PES) for the water dimer, based on fitting energies at roughly 30,000 configurations obtained with the coupled-cluster single and double, and perturbative treatment of triple excitations method using an augmented, correlation consistent, polarized triple zeta basis set. A global dipole moment surface based on Moller-Plesset perturbation theory results at these configurations is also reported. The PES is used in rigorous quantum calculations of intermolecular vibrational frequencies, tunneling splittings, and rotational constants for (H2O)2 and (D2O)2, using the rigid monomer approximation. Agreement with experiment is excellent and is at the highest level reported to date. The validity of this approximation is examined by comparing tunneling barriers within that model with those from fully relaxed calculations.

Two-dimensional supramolecular nanopatterns formed by the coadsorption of guanine and uracil at the liquid/solid interface.

A novel supramolecular nanostructure formed by the coadsorption of the complementary nucleobases guanine (G) and uracil (U) at the liquid (1-octanol solvent)/solid (graphite) interface is revealed by scanning tunneling microscopy (STM). The GU supramolecular structure is distinctly different from the structures observed by STM when the individual nucleobases (NB) are adsorbed on graphite in the control experiments. Using a systematic methodology and ab initio density functional theory (DFT), an atomistic structural model is proposed for the supramolecular coadsorbed GU structure, which consists of a periodic repetition of cyclic units based on the strongest GU base pairing.

Elementary structural motifs in a random network of cytosine adsorbed on a gold(111) surface.

Nonsymmetrical organic molecules adsorbed on solid surfaces may assemble into random networks, thereby providing model systems for organic glasses that can be directly observed by scanning tunneling microscopy (STM). We investigated the structure of a disordered cytosine network on a gold(111) surface created by thermal quenching, to temperatures below 150 K, of the two-dimensional fluid present on the surface at room temperature. Comparison of STM images to density functional theory calculations allowed us to identify three elementary structural motifs (zigzag filaments and five- and six-membered rings) that underlie the whole supramolecular random network. The identification of elementary structural motifs may provide a new framework for understanding medium-range order in amorphous and glassy systems.