Olgica Nedić

Molecular effects of advanced glycation end products on cell signalling pathways, ageing and pathophysiology.

Abstract Advanced glycation end-products (AGEs) are a heterogeneous group of compounds formed by the Maillard chemical process of non- enzymatic glycation of free amino groups of proteins, lipids and nucleic acids. This chemical modification of biomolecules is triggered by endogeneous hyperglycaemic or oxidative stress-related processes. Additionally, AGEs can derive from exogenous, mostly diet-related, sources. Considering that AGE accumulation in tissues correlates with ageing and is a hallmark in several age-related diseases it is not surprising that the role of AGEs in ageing and pathology has become increasingly evident. The receptor for AGEs (RAGE) is a single transmembrane protein being expressed in a wide variety of human cells. RAGE binds a broad repertoire of extracellular ligands and mediates responses to stress conditions by activating multiple signal transduction pathways being mostly responsible for acute and/or chronic inflammation. RAGE activation has been implicated in ageing as well as in a number of age-related diseases, including atherosclerosis, neurodegeneration, arthritis, stoke, diabetes and cancer. Here we present a synopsis of findings that relate to AGEs-reported implication in cell signalling pathways and ageing, as well as in pathology. Potential implications and opportunities for translational research and the development of new therapies are also discussed.

Serum insulin-like growth factor (IGF)-II is more closely associated with liver dysfunction than is IGF-I in patients with cirrhosis

The aim of this investigation was to determine the total concentrations of the insulin-like growth factors (IGF-I and IGF-II) in the blood serum of patients with liver cirrhosis and to evaluate their association with the condition. Cirrhosis was alcohol induced (n=27), of viral origin (n=17) or due to combined or other causes (n=21) and was moderate or severe in similar numbers of cases (Child A: n=21; Child B: n=21; Child C: n=23). While serum levels of both peptides were lower in patients than in age-matched healthy subjects (n=81), there was considerable overlap into the lower normal range for IGF-I. Moreover, no correlation between disease severity (Child score) and serum IGF-I was observed. Since a total of 78% of the results for IGF-II were outside the normal range (95% confidence interval) and serum concentrations were correlated with Child score (P=0.007), it is suggested that serum IGF-II concentrations may reflect compromised hepatic function more closely than IGF-I.

Analysis of protein carbonylation — pitfalls and promise in commonly used methods

Abstract Oxidation of proteins has received a lot of attention in the last decades due to the fact that they have been shown to accumulate and to be implicated in the progression and the pathophysiology of several diseases such as Alzheimer, coronary heart diseases, etc. This has also resulted in the fact that research scientists are becoming more eager to be able to measure accurately the level of oxidized protein in biological materials, and to determine the precise site of the oxidative attack on the protein, in order to get insights into the molecular mechanisms involved in the progression of diseases. Several methods for measuring protein carbonylation have been implemented in different laboratories around the world. However, to date no methods prevail as the most accurate, reliable, and robust. The present paper aims at giving an overview of the common methods used to determine protein carbonylation in biological material as well as to highlight the limitations and the potential. The ultimate goal is to give quick tips for a rapid decision making when a method has to be selected and taking into consideration the advantage and drawback of the methods.

Thyroid Hormone Synthesis and Storage in the Thyroid Gland of Human Neonates

The aim of this study was to evaluate the morphological and biochemical maturation of the thyroid gland in human neonates. The mean iodine concentration in the thyroid gland of very premature infants (less than 32 weeks gestational age, 0-3 days survival, n = 12) was significantly lower than in the older group (34-41 weeks gestational age, 0-30 days survival; n = 15; p < 0.05). For the whole group of neonates there was a statistically significant linear correlation between duration of life, i.e. gestational age and survival, and iodine concentration (r = 0.64, p < 0.01). Although there was wide dispersion of the results the same tendency was seen for thyroglobulin (Tg) concentration in the thyroid gland (r = 0.52, n = 21; p < 0.05). Comparative histological examination of the fetal thyroids gave results in accordance with the biochemical data as intrafollicular colloid appeared to be more abundant in more mature thyroids. The iodine content in Tg was found to be 0.63 +/- 0.22% in very preterm neonates and was slightly but not significantly lower than that found in the thyroids of the older group (0.82 +/- 0.14%; p = 0.055). The content of T4 and T3 per Tg molecule in the neonates was related to the iodine content. The differences in mean values of T4/Tg and T3/Tg molar ratios between the two groups were not significant: T4: 2.8 +/- 1.8 mol/ mol, T3: 0.29 +/- 0.12 mol/mol in very preterm neonates; and T4: 3.5 +/- 0.7 mol/mol, T3: 0.34 +/- 0.09 mol/mol in the older group. These results offer useful information for further analysis of the development of thyroid function in the human neonate.

Alterations of IGF-binding proteins in patients with alcoholic liver cirrhosis

The protein synthetic activity of the liver is diminished in cirrhosis. The aim of this study was to investigate possible changes in the serum IGF-IGFBP system among patients with alcoholic liver cirrhosis (ALC). The results obtained demonstrated that serum IGF-I and IGF-II concentrations were significantly lower in patients with ALC than in healthy persons (P=0.0008 for IGF-I and 0.0002 for IGF-II). The IGFBP profile was markedly altered and the 34 kDa IGFBP from patients had higher affinity towards 125I-IGF-II compared to the 34 kDa IGFBP of control individuals. Moreover, the 40-45 kDa IGFBP (in isolated complex with 125I-IGF-II) exhibited diminished interaction with concanavalin A, wheat germ, and breadfruit lectins. Modification of the glyco-component of the 40-45 kDa IGFBP seems to be an early event in ALC since change in reactivity towards lectins was noticed in patients with ALC classified as Child score A, whose serum IGF-I and IGF-II levels were within reference limits (the existence of carbohydrate microheterogeneity of this IGFBP was also assessed by lectin-affinity electrophoresis). It is possible that these biochemical alterations may affect the functional activity of the IGFs by changing the dynamics and distribution of these growth factors in the organism.

Peripheral circulating insulin-like growth factor-I and -II in cattle

Interrelationships between circulating concentrations of the insulin-like growth factors (IGF-I and IGF-II) were investigated in 235 blood samples taken from 145 healthy beef or dairy calves, bulls and cows of different breeds and ages. Autoradiography of Western ligand blots indicated different IGF binding protein (IGFBP) profiles between sera from different categories of cattle. Each IGF radioimmunoassay was validated by determining the effects of IGFBPs, ligand and contraligand, as well as serial dilution and comparison with results obtained after molecular sieve chromatography in acid. In female cattle mean values for IGF-I varied from 5.1 nmol/l in postparturient Holstein cows to 18.5-20.5 nmol/l in growing beef heifers, while mean IGF-II concentrations ranged from 30.0 nmol/l in the cows to 14.7-15.7 nmol/l in the beef heifer calves. In male cattle mean serum IGF-I ranged widely from 8.2 nmol/l in 1-day-old Holstein calves to 67.4 nmol/l in 16-month-old Simmental-type bulls. Mean IGF-II concentrations decreased from 22.9 nmol/l in 1-day-old Holstein bull calves to 11.9 nmol/l in 12-month-old beef bulls. Thus, total molar IGF concentrations were fairly stable in female cattle (24.7-35.1 nmol/l) but extended from 27.3 nmol/l to 81.8 nmol/l in the male cattle. The tendency for a reciprocal relationship between serum concentrations of these growth factors was most obvious in the periparturient cows.

Organic cesium carrier(s) in lichen

Most of the radiocesium was extracted from the lichen Cetraria islandica in aqueous medium (91.5%) without significant destruction of the lichen itself. Fractional precipitation with ethanol demonstrated that essential biomacromolecules were contaminated slightly with cesium. Using thin-layer and column chromatography at least three cesium-containing compounds were found in the residual solution. Two of them absorbed light at 270 nm. Further analysis of the main peak demonstrated instability resulting in newly formed compounds with smaller molecular masses. From this it was concluded that a significant amount of radiocesium in lichen is associated with organic molecules which tend to decompose upon isolation.

Posttranslational modifications of the insulin-like growth factor-binding protein 3 in patients with type 2 diabetes mellitus assessed by affinity chromatography.

Structural and ligand-binding properties of the insulin-like growth factor-binding protein (IGFBP)-3 in patients with poorly controlled diabetes mellitus type 2 were investigated using boronic acid- and lectin-affinity chromatography. IGFBP-3 species separated by chromatography were analyzed by immunoblotting and surface-enhanced laser desorption/ionization-time of flight mass spectrometry (SELDI-TOF MS). Increased IGFBP-3 binding to boronic acid in patients was shown to be accompanied by the increased ligand-binding. Increased binding of IGFBP-3 forms to lectins from Sambucus nigra (SNA) and Canavalia ensiformis (ConA) in patients, on the other hand, was either not accompanied by altered ligand-binding (in the case of ConA) or it was reduced (in the case of SNA). Strong and opposite effects of glycation and additional sialylation on ligand binding qualify them as factors that may be involved in the regulation of the amount of free, physiologically active IGFs, and modulation of processes that accompany development and progression of diabetes. SELDI-TOF MS analysis revealed a fragment of 13.9 kDa as representative for the non-glycosylated form of IGFBP-3, whereas a fragment of 28.0 kDa profiled as typical for the glycosylated/glycated IGFBP-3 species. The same fragmentation pattern found in healthy persons and in patients indicates that the same degradation process predominantly occurs in both groups of individuals.

Review time in peer review: quantitative analysis and modelling of editorial workflows

In this paper, we undertake a data-driven theoretical investigation of editorial workflows. We analyse a dataset containing information about 58 papers submitted to the Biochemistry and Biotechnology section of the Journal of the Serbian Chemical Society. We separate the peer review process into stages that each paper has to go through and introduce the notion of completion rate - the probability that an invitation sent to a potential reviewer will result in a finished review. Using empirical transition probabilities and probability distributions of the duration of each stage we create a directed weighted network, the analysis of which allows us to obtain the theoretical probability distributions of review time for different classes of reviewers. These theoretical distributions underlie our numerical simulations of different editorial strategies. Through these simulations, we test the impact of some modifications of the editorial policy on the efficiency of the whole review process. We discover that the distribution of review time is similar for all classes of reviewers, and that the completion rate of reviewers known personally by the editor is very high, which means that they are much more likely to answer the invitation and finish the review than other reviewers. Thus, the completion rate is the key factor that determines the efficiency of each editorial policy. Our results may be of great importance for editors and act as a guide in determining the optimal number of reviewers.

The N-glycan profile of placental membrane glycoproteins alters during gestation and aging

Alterations in the glycosylation of few membrane proteins from human placenta during gestation have been documented, but data on N-glycome of placental membrane proteins are still missing. The primary goal of this study was to obtain N-glycan profiles of human placental membrane proteins using a reliable, simple and high-throughput method. The second goal was to examine whether the N-glycan profile alters during gestation. Placental membrane proteins were isolated from women of different ages after first and third trimesters of pregnancy. The N-glycan fingerprint of membrane proteins was obtained using DNA sequencer-assisted fluorophore-assisted carbohydrate electrophoresis (DSA-FACE). Lectin blotting was used to confirm DSA-FACE results. Observed gestation-related alterations were: greater abundance of core-fucosylated and multiantennary N-glycans, but lower amounts of bisected biantennary N-glycans together with a decrease in α2,3-sialylation. Age-related alterations were: more core Fuc and more α2,3-Sia in first trimester placentas from older women than in those from younger women; also less core Fuc and less α2,6-Sia in third trimester placentas from older women compared to those from younger women. This study represents the first N-glycan profiling of placental cell membrane proteins. These data represent a basis for future research on the N-glycome of placental proteins in different (patho)physiological conditions.