Neutrophil gelatinase-associated lipocalin (NGAL) and matrix metalloproteinases as novel stress markers in children and young adults on chronic dialysis

Phenomena related to chronic kidney disease, such as atherosclerosis, aggravate with the introduction of dialysis. Matrix metalloproteinases (MMP) and factors modifying their activity, such as their tissue inhibitors (TIMP) or neutrophil gelatinase-associated lipocalin (NGAL), take part in the matrix turnover and the endothelial damage characteristic for atherogenesis. However, there are no data on the associations between these parameters and other known pro-atherogenic factors, or on the impact of various dialysis modalities on them. The aim of our study was to assess the serum concentrations of NGAL, MMP-7, MMP-9, and TIMP-1, as well as their correlations with human heat shock proteins (Hsp90α, anti-Hsp60), endothelial dysfunction (sE-selectin), and inflammation (hsCRP) in pediatric patients chronically dialyzed. Twenty-two children on automated peritoneal dialysis (APD), 17 patients on hemodialysis (HD) and 24 controls were examined. The serum concentrations of NGAL, MMP-7, MMP-9, TIMP-1, Hsp90α, anti-Hsp60, and sE-selectin were assessed by enzyme-linked immunosorbent assay (ELISA). The median values of NGAL, MMP-7, MMP-9, TIMP-1, and MMP-9/NGAL ratio were significantly elevated in all dialyzed children vs. controls and were higher in HD than in APD. The values of MMP-9/TIMP-1 and MMP-7/TIMP-1 ratios in the HD subjects were lower than those in the APD children. Hsp90α and anti-Hsp60 predicted the values of NGAL, MMPs, and TIMP-1. Additionally, sE-selectin was a predictor of NGAL levels, whereas NGAL predicted the MMP and TIMP-1 concentrations. The increased concentrations of examined parameters indicate the dysfunction of MMP/TIMP/NGAL system in the dialyzed children, more pronounced on hemodialysis. The discrepancies between dialysis modalities and correlations with heat shock proteins (HSPs) suggest that NGAL may be considered a novel stress protein, whereas MMP-7, MMP-9, and TIMP-1 may be regarded as indicators of stress response in the pediatric population on chronic dialysis.     

Search for anticonvulsant and analgesic active derivatives of dihydrofuran-2(3H)-one

A series of derivatives of dihydrofuran-2(3H)-one (γ-butyrolactone, GBL) was synthesized and tested for anticonvulsant, neurotoxic and analgesic activity. In the anticonvulsant screening 10 lactones were effective in the maximal electroshock test (MES) at the highest doses (300 and 100 mg/kg, 0.5 h, ip, mice). Statistical analysis showed correlation between the anticonvulsant activity and relative lipophilicity parameters determined by experimental and computational methods (R_M0, C log P and M log P). Preliminary antinociceptive evaluation of selected derivatives revealed strong analgesic activity. The majority of the tested compounds showed high efficacy in animal models of acute pain (hot plate and writhing tests) and strong local anesthetic activity (modified tail immersion test). The obtained ED₅₀ values were comparable with such analgesics as acetylsalicylic acid and morphine.     

Malondialdehyde plasma concentration correlates with declarative and working memory in patients with recurrent depressive disorder

Oxidative stress has been implicated in the cognitive decline, especially in memory impairment. The purpose of this study was to determine the concentration of malondialdehyde (MDA) in patients with recurrent depressive disorders (rDD) and to define relationship between plasma levels of MDA and the cognitive performance. The study comprised 46 patients meeting criteria for rDD. Cognitive function assessment was based on: The Trail Making Test , The Stroop Test, Verbal Fluency Test and Auditory-Verbal Learning Test. The severity of depression symptoms was assessed using the Hamilton Depression Rating Scale (HDRS). Statistically significant differences were found in the intensity of depression symptoms, measured by the HDRS on therapy onset versus the examination results after 8 weeks of treatment (P < 0.001). Considering the 8-week pharmacotherapy period, rDD patients presented better outcomes in cognitive function tests. There was no statistically significant correlation between plasma MDA levels, and the age, disease duration, number of previous depressive episodes and the results in HDRS applied on admission and on discharge. Elevated levels of MDA adversely affected the efficiency of visual-spatial and auditory-verbal working memory, short-term declarative memory and the delayed recall declarative memory. 1. Higher concentration of plasma MDA in rDD patients is associated with the severity of depressive symptoms, both at the beginning of antidepressants pharmacotherapy, and after 8 weeks of its duration. 2. Elevated levels of plasma MDA are related to the impairment of visual-spatial and auditory-verbal working memory and short-term and delayed declarative memory.     

Effect of caries preventive products on the growth of bacterial biofilm on titanium surface

Fluorides may affect the oxide layer on titanium surface. Caries preventive mouthwashes or gels contain fluorides and are applied at low pH. The aim of the present work was to study whether various concentrations of fluoride at acidic pH cause changes in the surface structure on the polished region of Ti implants, and alter the adherence and colonization of bacteria. Commercially pure Ti grade 4 discs with a polished surface were treated with a mouthwash containing 0.025% fluoride, a gel containing 1.25% fluoride or a 1% aqueous solution of NaF (pH 4.5). The change of surface roughness of the samples and the colonization of Porphyromonas gingivalis strains were studied by scanning electron microscopy after 5 days of anaerobic incubation. The quantity of the bacterial protein was determined by protein assay analysis. Agents with high fluoride concentration at acidic pH increased the roughness of the Ti surface. A slight increase in the amount of bacteria was found on the surfaces treated with 1% NaF and gel in comparison with the control surface. This study suggested that a high fluoride concentration at acidic pH may hinder the development of a healthy transgingival epithelial junction on Ti implants, due to bacterial colonization.     

Delayed fluorescence study on P*QA→ P+QA − charge separation energetics linked to protons and salt in reaction centers from Rhodobacter sphaeroides

The energetics of the first stable charge separated state, P⁺Q_A– relative to that of P–Q_A was examined in isolated RC from Rhodobacter sphaeroides by delayed fluorescence. The temperature dependence of the delayed fluorescence indicates that the charge separation is a highly enthalpy-driven process (H = – 818 ± 20 meV at pH 8) and the free energy gap between P–Q_A and P⁺Q_A– drops with increasing pH (40 ± 4 meV between pH 6 and 10). The pH-dependence of the free energy change of the P⁺Q_A– state runs parallel to the (integrated) net proton uptake due to the PQ_A/P⁺Q_A– redox change in a wide pH range and under different ionic conditions. Elevation of the ionic strength increases the delayed fluorescence intensity and decreases the (dark and light) pK_a values as well as the light-induced pK_a changes of the protonatable groups of the protein. The observed dependence of the energetics of P⁺Q_A– on the concentration and composition of mobile ions is discussed in terms of binding and screening of protonatable groups and surface charges as dominant modes of electrostatic interaction between RC and salt.     

Facies and diagenetic evaluation of the Permian–Triassic boundary interval and basal Triassic carbonates: shallow and deep ramp sections,Hungary

The Permian–Triassic boundary and basal Triassic shallow-marine successions were studied and correlated in sections of two structural units in Hungary (Transdanubian Range and Bükk units). Core sections in the Transdanubian Range unit recovered inner ramp deposits whereas outcrops in the Bükk unit expose deposits of the deeper ramp area of the western Tethys. The inner ramp section (studied ca. 10 m in thickness) is characterized by a succession of dolomites overlain by bioclastic limestones, peloidal grainstones (which recorded the biotic decline) and oolites with finely crystalline limestone interlayers. The deeper ramp section (studied ca. 15 m in thickness) is characterized by a succession of bioclastic limestones and marlstones, mudstone beds (recording the first biotic decline), the ‘boundary shales’ (recording the second biotic decline and the stable carbon isotope marker), mudstones with wackestone laminae, and stromatolite boundstones. Accordingly, oolite formation and microbial micrite precipitation represent carbonate sedimentary responses of end-Permian mass extinction on the carbonate shelf. In both successions, mudstones predominate the upsection, suggesting a relative sea-level rise. The succession of the deep ramp area exhibits a continuous sediment accumulation and the diagenesis here was influenced by marine and marine-derived pore water. The δ¹³C curve shows a continuous change towards more negative values, starting in bioclastic limestones, followed by a sharp symmetric negative peak at the second biotic decline that is a chemostratigraphic marker of the boundary event. Facies and microfacies trend of the inner ramp carbonates in the Transdanubian Range unit exhibits close similarities to that found in many South Alpine sections. Relict peloidal deposits, formed cemented submarine hardground substrate, indicate the extinction level. Sedimentary and diagenetic features of the overlying oolite bedset revealed slightly different depositional environments in the two studied Transdanubian Range unit sections. Petrography of the oolites highlighted shallow burial diagenetic alterations which includes marine cementation, marine-burial replacement and dolomitization. A lack of the specific negative peak in the δ¹³C values is most likely due to the multiple redeposition events of the sedimentary grains. This led to the conclusion that the deeper ramp deposits (e.g., in Bükk unit) have greater potential for recognizing trends in processes, affecting the marine environments and related to the end-Permian mass extinction, at the western Tethys.     

Resident phenotypically modulated vascular smooth muscle cells in healthy human arteries

Vascular interstitial cells (VICs) are non‐contractile cells with filopodia previously described in healthy blood vessels of rodents and their function remains unknown. The objective of this study was to identify VICs in human arteries and to ascertain their role. VICs were identified in the wall of human gastro‐omental arteries using transmission electron microscopy. Isolated VICs showed ability to form new and elongate existing filopodia and actively change body shape. Most importantly sprouting VICs were also observed in cell dispersal. RT‐PCR performed on separately collected contractile vascular smooth muscle cells (VSMCs) and VICs showed that both cell types expressed the gene for smooth muscle myosin heavy chain (SM‐MHC). Immunofluorescent labelling showed that both VSMCs and VICs had similar fluorescence for SM‐MHC and αSM‐actin, VICs, however, had significantly lower fluorescence for smoothelin, myosin light chain kinase, h‐calponin and SM22α. It was also found that VICs do not have cytoskeleton as rigid as in contractile VSMCs. VICs express number of VSMC‐specific proteins and display features of phenotypically modulated VSMCs with increased migratory abilities. VICs, therefore represent resident phenotypically modulated VSMCs that are present in human arteries under normal physiological conditions.     

Granulocyte-dependent Autoantibody-induced Skin Blistering

Autoimmune phenomena occur in healthy individuals, but when self-tolerance fails, the autoimmune response may result in specific pathology. According to Witebsky''s postulates, one of the criteria in diagnosing a disease as autoimmune is the reproduction of the disease in experimental animals by the passive transfer of autoantibodies. For epidermolysis bullosa acquisita (EBA), a prototypic organ-specific autoimmune disease of skin and mucous membranes, several experimental models were recently established. In the animal model described in our present work, purified IgG antibodies against a stretch of 200 amino acids (aa 757-967) of collagen VII are injected repeatedly into mice reproducing the blistering phenotype as well as the histo- and immunopathological features characteristic to human EBA ¹. Full-blown widespread disease is usually seen 5-6 days after the first injection and the extent of the disease correlates with the dose of the administered collagen VII-specific IgG. The tissue damage (blister formation) in the experimental EBA is depending on the recruitment and activation of granulocytes by tissue-bound autoantibodies ^2,-4. Therefore, this model allows for the dissection of the granulocyte-dependent inflammatory pathway involved in the autoimmune tissue damage, as the model reproduces only the T cell-independent phase of the efferent autoimmune response. Furthermore, its value is underlined by a number of studies demonstrating the blister-inducing potential of autoantibodies in vivo and investigating the mechanism of the blister formation in EBA ^1,3,-6. Finally, this model will greatly facilitate the development of new anti-inflammatory therapies in autoantibody-induced diseases. Overall, the passive transfer animal model of EBA is an accessible and instructive disease model and will help researchers to analyze not only EBA pathogenesis but to answer fundamental biologically and clinically essential autoimmunity questions.     

Compound heterozygosity for loss-of-function lysyl-tRNA synthetase mutations in a patient with peripheral neuropathy

Charcot-Marie-Tooth (CMT) disease comprises a genetically and clinically heterogeneous group of peripheral nerve disorders characterized by impaired distal motor and sensory function. Mutations in three genes encoding aminoacyl-tRNA synthetases (ARSs) have been implicated in CMT disease primarily associated with an axonal pathology. ARSs are ubiquitously expressed, essential enzymes responsible for charging tRNA molecules with their cognate amino acids. To further explore the role of ARSs in CMT disease, we performed a large-scale mutation screen of the 37 human ARS genes in a cohort of 355 patients with a phenotype consistent with CMT. Here we describe three variants (p.Leu133His, p.Tyr173SerfsX7, and p.Ile302Met) in the lysyl-tRNA synthetase (KARS) gene in two patients from this cohort. Functional analyses revealed that two of these mutations (p.Leu133His and p.Tyr173SerfsX7) severely affect enzyme activity. Interestingly, both functional variants were found in a single patient with CMT disease and additional neurological and non-neurological sequelae. Based on these data, KARS becomes the fourth ARS gene associated with CMT disease, indicating that this family of enzymes is specifically critical for axon function.