A total of 348 serum samples were collected from 301 hemodialysis patients with chronic renal failure of four healthcare settings in Western Bohemia. The sera were screened for the presence of ...hantavirus antibodies using ELISA kits (PROGEN Biotechnik GmbH) with Hantaan and Puumala antigens. Specific anti-Puumala antibodies were detected in five patients (1.7%). Although hantaviruses are known to cause primarily acute renal damage (interstitial nephritis) in Eurasia, chronic effects of hantavirus infection and the detection of specific antibodies in hemodialysis patients have also been reported. Nonetheless, the detection of seropositivity is not proof of an etiological link between hantaviruses and chronic renal failure. The hantavirus seropositivity rate in hemodialysis patients was not significantly higher than that in the general population. Our findings are consistent with the literature data and do not contradict the contribution of hantaviruses to the pathogenesis of chronic renal damage in the Czech Republic.
L-Carnitine is important in beta-oxidation of fatty acids. A lack of carnitine in hemodialysis patients is caused by insufficient carnitine synthesis and especially by its loss during dialysis. The ...aim of our study was to test the influence of carnitine supplementation on plasma lipids, red blood cell count, and metabolism of free radicals.
Twelve regularly dialyzed patients (average age 55.5 years, average dialysis treatment period 22.5 months) were given 15 mg/kg L-carnitine intravenously three times weekly (after each hemodialysis session) for 6 months. Laboratory markers of oxidative stress, lipid metabolism, and red blood cell count were measured before the supplementation and then controlled during two 3-month intervals. Nine patients were retested 3 months after the supplementation had ended.
All supplemented patients showed increased plasma free carnitine in comparison with the pretreatment values (113.3 +/- 11.2 vs. 62.3 +/- 16.7 micromol/l, p < 0.001). The proportion of decreased L-carnitine values after hemodialysis was reduced from 79 to 22%. Plasma total cholesterol (4.66 +/- 0.30 mmol/l after treatment vs. 5.65 +/- 1.53 mmol/l before treatment, p < 0.05) and LDL cholesterol (1.74 +/- 0.86 vs. 2.81 +/- 1.43 mmol/l, p < 0.05) decreased. The albumin concentration significantly increased from 34.8 +/- 7.3 to 46.0 +/-5.4 g/l (p < 0.05). Intraerythrocyte reduced glutathione increased from 1.65 +/- 0.25 to 2.23 +/- 0.16 mmol/l (p < 0.001), and the plasma antioxidant capacity increased from 1.65 +/- 0.09 to 2.06 +/- 0.17 mmol/l (p < 0.001). At the same time, plasma malondialdehyde decreased from 4.18 +/- 0.72 to 3.07 +/- 0.35 micromol/l (p < 0.001). The erythropoietin dose could be reduced from an average value of 5,500 to 3,500 U/week. No significant changes in the above-mentioned parameters were observed in a control group of dialyzed patients without L-carnitine supplementation.
Regular carnitine supplementation of hemodialysis patients can improve their lipid metabolism, protein nutrition, red blood cell count, and antioxidant status.
Replication stress (RS) fuels genomic instability and cancer development and may contribute to aging, raising the need to identify factors involved in cellular responses to such stress. Here, we ...present a strategy for identification of factors affecting the maintenance of common fragile sites (CFSs), which are genomic loci that are particularly sensitive to RS and suffer from increased breakage and rearrangements in tumors. A DNA probe designed to match the high flexibility island sequence typical for the commonly expressed CFS (FRA16D) was used as specific DNA affinity bait. Proteins significantly enriched at the FRA16D fragment under normal and replication stress conditions were identified using stable isotope labeling of amino acids in cell culture-based quantitative mass spectrometry. The identified proteins interacting with the FRA16D fragment included some known CFS stabilizers, thereby validating this screening approach. Among the hits from our screen so far not implicated in CFS maintenance, we chose Xeroderma pigmentosum protein group C (XPC) for further characterization. XPC is a key factor in the DNA repair pathway known as global genomic nucleotide excision repair (GG-NER), a mechanism whose several components were enriched at the FRA16D fragment in our screen. Functional experiments revealed defective checkpoint signaling and escape of DNA replication intermediates into mitosis and the next generation of XPC-depleted cells exposed to RS. Overall, our results provide insights into an unexpected biological role of XPC in response to replication stress and document the power of proteomics-based screening strategies to elucidate mechanisms of pathophysiological significance.
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IJS, KILJ, NUK, PNG, UL, UM
Waste bovine hooves and horns were enzymatically hydrolysed into soluble products intended for foliar fertilizer. With the powdered keratin at 50℃ and pH 8 between 34 to nearly 60% of nitrogen was ...solubilized in 5 h, depending on the enzyme concentration. The reaction could further be improved by steam pretreatment of the keratin, resulting in 98% solubilisation of the nitrogen. The products of hydrolysis consisted of a mixture of soluble proteins, peptides, and free amino acids. Among the latter, 18 common amino acids were detected. Several of them were previously recognized to have a positive effect on plants. Nonpolar neutral, basic, and sulphur amino acids were present in relatively large amounts, while proline and tryptophan were not found. Comparison with other protein hydrolysates aimed for fertilizer suggests that keratin degradation products, obtained by enzymatic hydrolysis, have potential to be used for foliar fertilization, alone or in a combination with another complementary hydrolysate of a different source, such as skin or plant proteins.
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CEKLJ, EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
To determine antibiotic resistance and incidence of multidrug resistance among Nontyphoidal salmonellae serovars isolated from humans.
Consecutive Salmonella isolates from patients, recovered in 48 ...microbiology laboratories in May 2012, were analyzed in the respective reference laboratories at the National Institute of Public Health. Strains were re-identified and differentiated into serovars. Their minimum inhibitory concentrations (MICs) to 11 antibiotics were determined by the microdilution method.
Of 25 serovars identified among 637 strains of Salmonella enterica, the most frequent were Enteritidis (87.0 %), Typhimurium (4.9 %), and monophasic Typhimurium 4,5,12:i:- (2.0 %) and Mbandaka (0.6 %); other serovars were rare. Altogether 558 strains (87.6 %) were susceptible to all antibiotics tested and the remaining 79 strains were resistant to one or more antibiotics. The prevalence rates of resistance to individual antibiotics among 637 study strains were as follows: ampicillin 8.5%, tetracycline 5.7%, sulfamethoxazole 5.2%, cipro-floxacin 3.8%, and chloramphenicol 2.5%. Resistance to gentamicin, trimethoprim, and third and fourth generation cephalosporins was rare ( 0.5%) and none of the study strains showed resistance to meropenem. Three producers of extended spectrum beta-lactamase were multidrug resistant and two of them recovered from twins exhibited a different pattern of resistance. Resistant strains were most often assigned to the following serovars: Enteritidis (49.4%), Typhimurium (26.6%), and monophasic Typhimurium (15.2%). While only 7% (39 of 554 strains) of Enteritidis strains were resistant, the serovars Typhimurium and its monophasic variant 4,5,12:i:- showed high rates of resistance, i.e. 66.7 and 92.3%, respectively. Furthermore, resistance was revealed in all strains of the serovars Virchow (n = 3), Kentucky (n = 1), and Newport (n = 1), in two of three strains of the serovar Infantis, and in one of two strains of the serovar Stanley. All five blood isolates were assigned to the serovar Enteritidis and one of them showed resistance to ciprofloxacin. Of 79 resistant strains, 26.6% showed resistance to ampicillin only and 24.1% to ciprofloxacin only, with multidrug resistance, i.e. resistance to three or more antibiotics, confirmed in 43.0% of strains.
Despite a relatively low prevalence of resistance to the antibiotics tested among 637 study strains, the following alarming findings were made: Detection of Salmonella enterica strains resistant to ciprofloxacin as the drug of choice or to higher generation cephalosporins and multidrug resistance revealed in two thirds of the strains of the serovar Typhimurium and in all but one strains of its monophasic variant 4,5,12:i:-.
: Serum amino grams and daily losses of glutamine (Gin) and other amino acids (AAs) into diafiltrate were measured during the first 5 days of continuous venovenous hemodiafiltration (CVVHDF) in 6 ICU ...patients with acute renal failure (ARF). Four patients had ARF as a part of multiple organ failure (MOF) of septic origin, and 2 patients had isolated ARF because of primary renal disease. During the study, all the patients received defined total parenteral nutrition (TPN). The mean daily AA losses into dialysate were relatively low (0.61 ± 0.1 gN) and reached 4.5% of the daily AA substitution. Gln represented 32.7 ± 5.9% of the total AA losses (0.19 ± 0.04 gN). Serum levels of Gin (p = 0.002) and of most other AAs were significantly lower in the patients than in the control subjects (AA analysis in 16 healthy volunteers). Phenylalanine (Phe) was the only AA that was increased significantly (p < 0.01) in the patients. The mean patient serum concentrations of Phe and tyrosine were significantly higher (p < 0.03) than the correspondent concentrations in dialysate, but the lysine concentration was higher in dialysate (p < 0.03). The serum and dialysate concentrations of other AAs did not differ. Gin in serum decreased significantly (p < 0.03) on the second day of CVVHDF but returned to the baseline levels subsequently. Serum concentrations of Phe increased on the second day of CVVHDF (p < 0.05). Serum concentrations of other AAs remained stable during the whole study. We conclude that Gin losses into dialysate during CVVHDF are relatively low, but CVVHDF itself may induce changes in Gin metabolism and distribution that are reflected by a decrease of serum Gin levels at the institution of this treatment. Therefore, the need for Gin supplementation in ICU patients is even greater in the first days of CVVHDF.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Cdc7 kinase regulates DNA replication. However, its role in DNA repair and recombination is poorly understood. Here we describe a pathway that stabilizes the human Cdc7-ASK (activator of S-phase ...kinase; also called Dbf4), its regulation, and its function in cellular responses to compromised DNA replication. Stalled DNA replication evoked stabilization of the Cdc7-ASK (Dbf4) complex in a manner dependent on ATR-Chk1-mediated checkpoint signaling and its interplay with the anaphase-promoting complex/cyclosome super(cdh1) (APC/C super(cdh1)) ubiquitin ligase. Mechanistically, Chk1 kinase inactivates APC/C super(cdh1) through degradation of Cdh1 upon replication block, thereby stabilizing APC/C super(cdh1) substrates, including Cdc7-ASK (Dbf4). Furthermore, motif C of ASK (Dbf4) interacts with the N-terminal region of RAD18 ubiquitin ligase, and this interaction is required for chromatin binding of RAD18. Impaired interaction of ASK (Dbf4) with RAD 18 disables foci formation by RAD 18 and hinders chromatin loading of translesion DNA polymerase eta . These findings define a novel mechanism that orchestrates replication checkpoint signaling and ubiquitin-proteasome machinery with the DNA damage bypass pathway to guard against replication collapse under conditions of replication stress.
Neuraminidase from
Arthrobacter nicotianae was purified by chromatography on DEAE-cellulose, gel filtration on Sephadex G-150, and chromatography on CM-cellulose. The molecular mass of the highly ...purified neuraminidase was found to be approximately 69 kDa by SDS-PAGE. The pH and temperature optima were 5.0 and 45°C, respectively. The enzyme was stable at temperatures up to 50°C, Ca
2+, Ba
2+, and Mg
2+ activated the enzyme, but Pb
2+, Fe
3+, and the chelating agent EDTA were inhibitory. The purified neuraminidase had a
K
m value of 3.17 m
M when glucomacropeptide was the substrate.
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IJS, IMTLJ, KILJ, KISLJ, NUK, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Treatment of anaemia with human recombinant erythropoietin (r-Hu EPO) substantially improves the quality of life of patients receiving regular dialysis treatment. The positive effect on the ...nutritional state of patients during r-Hu EPO treatment is not accepted unequivocally so far. The authors followed-up 9 patients (6 men and 3 women) who had regular dialysis treatment, and suffered from severe anaemia. The patients were treated for 12 months with r-Hu EPO (preparation Eprex Cilagd Co.) with an initial dose of 30 mu/kg 3x per week by s.c. route with a target haemoglobin (Hb) value of 90-100 g/l. Before treatment during the 6th and 12th month of treatment the basic nutritional parameters of the patients were followed up. In the course of the investigation the Hb level (x +/- SD) increased from 61 +/- 12 g/l to 98 +/- 8 g/l, p < 0.01. On comparison of the nutritional parameters before treatment and after 12 months of treatment with r-Hu EPO the authors recorded rising trends in the patients' body weight (kg) from 67.4 +/- 12.2 to 68.4 +/- 11.2 (n.s.), an increase of the skinfold thickness above the triceps brachii muscle (mm) from 8.6 +/- 5.8 to 9.8 +/- 4.3 (n.s.), a decline of skin reactivity to diphtheric antigen (mm) from 2.1 +/- 2.8 to 1.8 +/- 0.8 (n.s.), a rising trend of reactivity to Candida antigen (mm) from 2.9 +/- 2.2 to 3.9 +/- 2.4 (n.s.), to diphtheric antigen from 3.0 +/- 3.3 to 5.1 +/- 3.8 (n.s.), a rise of the transferrin level (g/l) from 2.5 +/- 0.3 to 2.8 +/- 1.3 (n.s.), a rise of serum urea (mmol/l) from 25.7 +/- 3.6 to 32.5 +/- 9.5 (n.s.), a rise of Mullen's nutritional index from 21.8 +/- 15.5 to 22.3 +/- 24.7 (n.s.).
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