Indicators of acute and persistent renal damage in adult thrombotic microangiopathy

Background

Thrombotic microangiopathies (TMA) in adults such as thrombotic thrombocytopenic purpura (TTP) and hemolytic uremic syndrome (HUS) are life-threatening disorders if untreated. Clinical presentation is highly variable and prognostic factors for clinical course and outcome are not well established.

Methods

We performed a retrospective observational study of 62 patients with TMA, 22 males and 40 females aged 16 to 76 years, treated with plasma exchange at one center to identify clinical risk factors for the development of renal insufficiency.

Results

On admission, 39 of 62 patients (63%) had acute renal failure (ARF) with 32 patients (52%) requiring dialysis treatment. High systolic arterial pressure (SAP, p = 0.009) or mean arterial pressure (MAP, p = 0.027) on admission was associated with acute renal failure. Patients with SAP>140 mmHg on admission had a sevenfold increased risk of severe kidney disease (OR 7.464, CI 2.097–26.565). MAP>100 mmHg indicated a fourfold increased risk for acute renal failure (OR 4.261, CI 1.400–12.972). High SAP, diastolic arterial pressure (DAP), and MAP on admission were also independent risk factors for persistent renal insufficiency with the strongest correlation for high MAP. Moreover, a high C-reactive protein (CRP) level on admission correlated with renal failure in the course of the disease (p = 0.003). At discharge, renal function in 11 of 39 patients (28%) had fully recovered, 14 patients (23%) remained on dialysis, and 14 patients (23%) had non-dialysis-dependent chronic kidney disease. Seven patients (11%) died. We identified an older age as risk factor for death.

Conclusions

High blood pressure as well as high CRP serum levels on admission are associated with renal insufficiency in TMA. High blood pressure on admission is also a strong predictor of sustained renal insufficiency. Thus, adult TMA patients with high blood pressure may require special attention to prevent persistent renal failure.     

Determination of the glycosaminoglycan and collagen contents in tissue samples by high-resolution 1H NMR spectroscopy after DCl-induced hydrolysis

The determination of the collagen and glycosaminoglycan (GAG) contents of native and particularly bioengineered tissues is of considerable interest because the collagen-to-GAG ratio determines the water content of the tissue, which is crucial regarding its mechanical properties. (1)H NMR spectroscopy subsequent to the hydrolysis of the sample by aqueous 6 M DCl at 353 K is used to determine the GAG and collagen contents simultaneously. Under these strongly acidic conditions the biopolymers of the extracellular matrix, collagen, and GAG are fragmented into their individual monomers, that is, free amino acids from collagen and monosaccharides from the polymer repeat units of GAGs. The amino acid amount can be easily determined in the presence of an internal standard by (1)H NMR spectroscopy because amino acids proved to be stable under acidic conditions. The carbohydrates are subject to charring in the presence of concentrated DCl, but glucosamine and galactosamine were found to be sufficiently stable for quantification under the chosen conditions.     

Comparative toxicity of nanoparticulate CuO and ZnO to soil bacterial communities

The increasing industrial application of metal oxide Engineered Nano-Particles (ENPs) is likely to increase their environmental release to soils. While the potential of metal oxide ENPs as environmental toxicants has been shown, lack of suitable control treatments have compromised the power of many previous assessments. We evaluated the ecotoxicity of ENP (nano) forms of Zn and Cu oxides in two different soils by measuring their ability to inhibit bacterial growth. We could show a direct acute toxicity of nano-CuO acting on soil bacteria while the macroparticulate (bulk) form of CuO was not toxic. In comparison, CuSO(4) was more toxic than either oxide form. Unlike Cu, all forms of Zn were toxic to soil bacteria, and the bulk-ZnO was more toxic than the nano-ZnO. The ZnSO(4) addition was not consistently more toxic than the oxide forms. Consistently, we found a tight link between the dissolved concentration of metal in solution and the inhibition of bacterial growth. The inconsistent toxicological response between soils could be explained by different resulting concentrations of metals in soil solution. Our findings suggested that the principal mechanism of toxicity was dissolution of metal oxides and sulphates into a metal ion form known to be highly toxic to bacteria, and not a direct effect of nano-sized particles acting on bacteria. We propose that integrated efforts toward directly assessing bioavailable metal concentrations are more valuable than spending resources to reassess ecotoxicology of ENPs separately from general metal toxicity.     

The structure of the oligomerization domain of Lsr2 from Mycobacterium tuberculosis reveals a mechanism for chromosome organization and protection

Lsr2 is a small DNA-binding protein present in mycobacteria and related actinobacteria that regulates gene expression and influences the organization of bacterial chromatin. Lsr2 is a dimer that binds to AT-rich regions of chromosomal DNA and physically protects DNA from damage by reactive oxygen intermediates (ROI). A recent structure of the C-terminal DNA-binding domain of Lsr2 provides a rationale for its interaction with the minor groove of DNA, its preference for AT-rich tracts, and its similarity to other bacterial nucleoid-associated DNA-binding domains. In contrast, the details of Lsr2 dimerization (and oligomerization) via its N-terminal domain, and the mechanism of Lsr2-mediated chromosomal cross-linking and protection is unknown. We have solved the structure of the N-terminal domain of Lsr2 (N-Lsr2) at 1.73 ? resolution using crystallographic ab initio approaches. The structure shows an intimate dimer of two ?-?-a motifs with no close homologues in the structural databases. The organization of individual N-Lsr2 dimers in the crystal also reveals a mechanism for oligomerization. Proteolytic removal of three N-terminal residues from Lsr2 results in the formation of an anti-parallel β-sheet between neighboring molecules and the formation of linear chains of N-Lsr2. Oligomerization can be artificially induced using low concentrations of trypsin and the arrangement of N-Lsr2 into long chains is observed in both monoclinic and hexagonal crystallographic space groups. In solution, oligomerization of N-Lsr2 is also observed following treatment with trypsin. A change in chromosomal topology after the addition of trypsin to full-length Lsr2-DNA complexes and protection of DNA towards DNAse digestion can be observed using electron microscopy and electrophoresis. These results suggest a mechanism for oligomerization of Lsr2 via protease-activation leading to chromosome compaction and protection, and concomitant down-regulation of large numbers of genes. This mechanism is likely to be relevant under conditions of stress where cellular proteases are known to be upregulated.     

Spatial and temporal determinants of genetic structure in Gentianella bohemica

The biennial plant Gentianella bohemica is a subendemic of the Bohemian Massif, where it occurs in seminatural grasslands. It has become rare in recent decades as a result of profound changes in land use. Using amplified fragment length polymorphisms (AFLP) fingerprint data, we investigated the genetic structure within and among populations of G. bohemica in Bavaria, the Czech Republic, and the Austrian border region. The aim of our study was (1) to analyze the genetic structure among populations and to discuss these findings in the context of present and historical patterns of connectivity and isolation of populations, (2) to analyze genetic structure among consecutive generations (cohorts of two consecutive years), and (3) to investigate relationships between intrapopulational diversity and effective population size (N(e)) as well as plant traits. (1) The German populations were strongly isolated from each other (pairwise F(ST)= 0.29-0.60) and from all other populations (F(ST)= 0.24-0.49). We found a pattern of near panmixis among the latter (F(ST)= 0.15-0.35) with geographical distance explaining only 8% of the genetic variance. These results were congruent with a principal coordinate analysis (PCoA) and analysis using STRUCTURE to identify genetically coherent groups. These findings are in line with the strong physical barrier and historical constraints, resulting in separation of the German populations from the others. (2) We found pronounced genetic differences between consecutive cohorts of the German populations (pairwise F(ST)= 0.23 and 0.31), which can be explained by local population history (land use, disturbance). (3) Genetic diversity within populations (Shannon index, H(Sh)) was significantly correlated with N(e) (R(S)= 0.733) and reflected a loss of diversity due to several demographic bottlenecks. Overall, we found that the genetic structure in G. bohemica is strongly influenced by historical periods of high connectivity and isolation as well as by marked demographic fluctuations in declining populations.     

IRES-dependent translation of egr2 is induced under inflammatory conditions

Adjusting translation is crucial for cells to rapidly adapt to changing conditions. While pro-proliferative signaling via the PI3K-mTOR-pathway is known to induce cap-dependent translation, stress conditions, such as nutrient deprivation or hypoxia often activate alternative modes of translation, e.g., via internal ribosome entry sites (IRESs). As the effects of inflammatory conditions on translation are only poorly characterized, we aimed at identifying translationally deregulated targets in inflammatory settings. For this purpose, we cocultured breast tumor cells with conditioned medium of activated monocyte-derived macrophages (CM). Polysome profiling and microarray analysis identified early growth response-2 (egr2) to be regulated at the level of translation. Using bicistronic reporter assays, we found that egr2 contains an IRES within its 5′ UTR, which facilitated enhanced translation upon CM treatment. We further provide evidence that the activity of egr2-IRES was induced by IL-1β and p38-MAPK signaling. In addition, we identified several potential IRES trans-acting factors (ITAFs) such as polypyrimidine tract binding protein (PTB) and hnRNP-A1 that directly bind to the egr2-5′UTR. In summary, our data provide evidence that egr2 expression is translationally regulated via an IRES element, which is responsive to an inflammatory environment.     

Caenorhabditis elegans as a chemical screening tool for the study o f neuromuscular disorders. Manual and semi-automated methods

We previously reported the use of the cheap and fast-growing nematode Caenorhabditis elegans to search for molecules, which reduce muscle degeneration in a model for Duchenne Muscular Dystrophy (DMD). We showed that Prednisone, a steroid that is generally prescribed as a palliative treatment to DMD patients, also reduced muscle degeneration in the C. elegans DMD model. We further showed that this strategy could lead to the discovery of new and unsuspected small molecules, which have been further validated in a mammalian model of DMD, i.e. the mdx mouse model. These proof-of-principles demonstrate that C. elegans can serve as a screening tool to search for drugs against neuromuscular disorders. Here, we report and discuss two methodologies used to screen chemical libraries for drugs against muscle disorders in C. elegans. We first describe a manual method used to find drugs against DMD. We further present a semi-automated method, which is currently in use for the search of drugs against the Schwartz-Jampel Syndrome (SJS). Both assays are simple to implement and can be readily transposed and/or adapted to screens against other muscle/neuromuscular diseases, which can be modeled in the worm. Finally we discuss, with respect to our experience and knowledge, the different parameters that have to be taken into account before choosing one or the other method.     

"There are too many, but never enough": qualitative case study investigating routine coding of clinical information in depression

Background

We sought to understand how clinical information relating to the management of depression is routinely coded in different clinical settings and the perspectives of and implications for different stakeholders with a view to understanding how these may be aligned.

Materials and Methods

Qualitative investigation exploring the views of a purposefully selected range of healthcare professionals, managers, and clinical coders spanning primary and secondary care.

Results

Our dataset comprised 28 semi-structured interviews, a focus group, documents relating to clinical coding standards and participant observation of clinical coding activities. We identified a range of approaches to coding clinical information including templates and order entry systems. The challenges inherent in clearly establishing a diagnosis, identifying appropriate clinical codes and possible implications of diagnoses for patients were particularly prominent in primary care. Although a range of managerial and research benefits were identified, there were no direct benefits from coded clinical data for patients or professionals. Secondary care staff emphasized the role of clinical coders in ensuring data quality, which was at odds with the policy drive to increase real-time clinical coding.

Conclusions

There was overall no evidence of clear-cut direct patient care benefits to inform immediate care decisions, even in primary care where data on patients with depression were more extensively coded. A number of important secondary uses were recognized by healthcare staff, but the coding of clinical data to serve these ends was often poorly aligned with clinical practice and patient-centered considerations. The current international drive to encourage clinical coding by healthcare professionals during the clinical encounter may need to be critically examined.