Difference in coagulation markers in acute and one year post acute ischemic stroke

Background and purpose: Shifts of the balance between coagulation and fibrinolysis play a crucial role in pathogenesis of cerebral ischemia. However, its relevance to post-acute disease period requires further elucidation. This study aimed to characterize whether hypercoagulation markers differ for patients in acute and the same patients in the post acute phase of ischemic stroke. Materials and methods: Systemic generation of hemostasis markers was monitored and fraction composition was described during one year of disease development and treatment. Results: Increased concentration of the markers of hypercoagulation in blood plasma of patients in acute as well as one year past acute phase ischemic stroke were shown. Thus, not all markers of hypercoagulation become relevant to norm one year past ischemic stroke. It\u27s the first time characterization of the coagulation markers in acute and post acute period of the absolutely same group of patients was done. Our study demonstrates the difference in quantity and quality of the fractions of proteins with prothrombin origin and soluble fibrin monomer complexes which could be used as a potential target for the prevention of the stroke repetition. Conclusions: We assume that prothrombin plays the most important role in the development of pathology tested, hence it may provide future targets for therapeutic strategies.</p

Fragmenti kolagena male molekularne mase poboljšavaju masene i upalne parametre masnog tkiva pretilih štakora

Research background. Despite clearly recognized links between increased body mass and increased risk for various pathological conditions, therapeutic options to treat obesity are still very limited. The aim of the present study is to explore the effect of low-molecular-mass collagen fragments obtained from the scales of Antarctic wild marine fish on rats\u27 visceral and subcutaneous white adipose tissue in a high-calorie diet-induced obesity model. Experimental approach. The study was conducted on outbred rats, which were divided into 3 experimental groups: (i) control, consuming standard food (3.81 kcal/g), (ii) obese group, consuming a high-calorie diet (5.35 kcal/g), and (iii) obese group, consuming a high-calorie diet (5.35 kcal/g) with intragastric administration of low-molecular-mass collagen fragments (at a dose 1 g/kg of body mass during 6 weeks). Low-molecular-mass collagen fragments were obtained by a procedure that included collagen extraction from fish scales and enzymatic hydrolysis with pepsin. Apart from hematoxylin and eosin staining, fibrosis level was assessed by histochemical Van Gieson’s trichrome picrofuchsin staining, and mast cells were analysed by toluidine blue O staining. Results and conclusions. Group treated with low-molecular-mass fragments of collagen exhibited decreased rate of mass gain, relative mass, area occupied by collagen fibre of both visceral and subcutaneous adipose tissue, and cross-sectional area of both visceral and subcutaneous adipocytes. Treatment with low-molecular-mass fragments of collagen reduced the infiltration of immune cells, number of mast cells and their redistribution back to the septa. This was also accompanied by a decreased number of the crown-like structures formed by the immune cells, which are markers of chronic inflammation that accompanies obesity. Novelty and scientific contribution. This is the first study that reports the anti-obesity effect of low-molecular-mass fragments produced as a result of controlled hydrolysis of collagen from the scales of Antarctic wild marine fish in the in vivo model. Another novelty of this work is the observation that the tested collagen fragments not only reduce the body mass, but also improve the morphological and inflammatory parameters (decrease in the number of crown-like structures, immune cell infiltration, fibrosis and mast cells). Altogether, our work suggests that low-molecular-mass collagen fragments are a promising candidate for amelioration of some comorbidities linked to obesity.Pozadina istraživanja. Usprkos jasno utvrđenoj povezanosti povećane tjelesne mase s većim rizikom razvoja različitih patoloških oboljenja, mogućnosti liječenja pretilosti su još uvijek vrlo ograničene. Svrha je ovoga rada bila ispitati učinak fragmenata kolagena male molekularne mase, dobivenih iz ljuski morskih riba s područja Antarktika, na visceralno i potkožno masno tkivo štakora s dijabetesom uzrokovanim visokokaloričnom prehranom. Eksperimentalni pristup. Istraživanje je provedeno na nesrođenim sojevima štakora, podijeljenim u tri eksperimentalne skupine: (i) kontrolna skupina, koja je primala standardnu prehranu (3,81 kcal/g), (ii) pretila skupina koja je primala visokokaloričnu prehranu (5,35 kcal/g), i (iii) pretila skupina koja je primala visokokaloričnu prehranu (5,35 kcal/g) i intragastrično fragmente kolagena male molekularne mase (1 g po kg tjelesne mase tijekom 6 tjedana). Fragmenti kolagena male molekularne mase dobiveni su ekstrakcijom kolagena iz ljusaka ribe i njegovom enzimskom hidrolizom pomoću pepsina. Stupanj fibroze masnog tkiva je, osim bojenjem hematoksilin eozinom, utvrđen Van Giesonovim bojenjem, a mastociti su ispitani bojenjem toluidinskim modrilom. Rezultati i zaključci. Skupina štakora koja je primala fragmente kolagena male molekularne mase sporije je dobivala na masi i relativnoj masi, a kolagenska vlakna su zauzimala manju površinu njihovog visceralnog i potkožnog masnog tkiva te presjeka visceralnih i potkožnih adipocita. Intragastričnom primjenom fragmenata kolagena male molekularne mase smanjili su se infiltracija imunoloških stanica, broj mastocita i njihova redistribucija u septe. Također se smanjio broj krunastih struktura koje tvore makrofagi, a koje su markeri kronične upale povezane s pretilošću. Novina i znanstveni doprinos. Ovo je prvo istraživanje in vivo o utjecaju fragmenata kolagena dobivenih kontroliranom hidrolizom ljusaka morskih riba iz područja Antarktika na smanjenje tjelesne mase. Još jedna novina ovoga rada je opažanje da ispitani fragmenti kolagena nisu samo smanjili tjelesnu masu, već i poboljšali morfološke i upalne parametre (smanjili su se broj krunastih struktura, infiltracija imunostanica, fibroza i broj mastocita). Zaključno, rad je pokazao da fragmenti kolagena male molekularne mase mogu ublažiti neke komorbiditete povezane s pretilošću

A comparative study of neurotoxic potential of synthesized polysaccharidecoated and native ferritinbased magnetic nanoparticles

Aim To analyze the neurotoxic potential of synthesized magnetite nanoparticles coated by dextran, hydroxyethyl starch, oxidized hydroxyethyl starch, and chitosan, and magnetic nanoparticles combined with ferritin as a native protein. Methods The size of nanoparticles was analyzed using photon correlation spectroscopy, their effects on the conductance of planar lipid membrane by planar lipid bilayer technique, membrane potential and acidification of synaptic vesicles by spectrofluorimetry, and glutamate uptake and ambient level of glutamate in isolated rat brain nerve terminals (synaptosomes) by radiolabeled assay. Results Uncoated synthesized magnetite nanoparticles and nanoparticles coated by different polysaccharides had no significant effect on synaptic vesicle acidification, the initial velocity of L-[14C]glutamate uptake, ambient level of L-[14C]glutamate and the potential of the plasma membrane of synaptosomes, and conductance of planar lipid membrane. Native ferritin-based magnetic nanoparticles had no effect on the membrane potential but significantly reduced L-[14C]glutamate transport in synaptosomes and acidification of synaptic vesicles. Conclusions Our study indicates that synthesized magnetite nanoparticles in contrast to ferritin have no effects on the functional state and glutamate transport of nerve terminals, and so ferritin cannot be used as a prototype, analogue, or model of polysaccharide-coated magnetic nanoparticle in toxicity risk assessment and manipulation of nerve terminals by external magnetic fields. Still, the ability of ferritin to change the functional state of nerve terminals in combination with its magnetic properties suggests its biotechnological potential

Influence of the complex drug Cocarnit on the sciatic nerve in the development of diabetic polyneuropathy in rats

Ulcers and slow wound healing are common in diabetic polyneuropathy (DP), as well as shooting or burning pain, sensitivity to touch or lack of sensitivity, low oxygenation of nerve tissue, conductivity disorders and various vascular disorders. The mechanisms of DP development are complex and have not been completely studied. To take into account the role of B group vitamins, we investigated histological structure of nerve tissue, the level of different growth factors and the qualitative composition of active proteolytic enzymes in rats with DP and after the use of the metabolic drug Cocarnit for 9 days. This drug composition include nicotinamide, cocarboxylase, cyanocobalamin, adenosine triphosphate disodium trihydrate. We used an histological study of sciatic nerve; enzyme-linked immunosorbent assay and enzyme electrophoresis methods. In rats with DP, fragmentation of nerve tissue and their necrosis was established. Moreover, degraded forms of plasmin that has a fully functional serine proteinase domain are evident, and, therefore, it exhibits proteolytic properties. DP led to a decrease of neuron growth factor (NGF), vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). After treatment, the histological structure of nerve tissue was significantly improved, and the expression of growth factors NGF and bFGF was increased. Our study demonstrated that administration of Corcarnit brought about the complete restoration of the activation potential of plasmin and the almost disappearance of all degraded forms which were evident in the group with DP

Tlr2, Tjp1 genes expression during wound healing dynamics – with melanin treatment

Wound healing is the complex and dynamic process of replacing devitalized and missing cellular structures and tissue layers. We have previously shown that melanin, herein, produced by the Antarctic black yeast fungi Pseudonadsoniella brunea (Nadsoniella nigra sp. X-1), has expressed a cyto-protective effect, promoted rapid wound healing of various ethiology and can be offered as a new dermatropic drug. The current study was conducted on a rat model of purulent necrotic wound. In each model, one group was a control, while in the others, wound healing occurred without drug application or with administration of 0,5% carbopol or with both 0,5% carbopol and 0,1% melanin. The pro-oxidant-antioxidant balance in skin gomogenate in dynamics on 3, 6, 9, 14 and day of full epithelization was estimated using the spectrophotometric biochemical method. Moreover, so as to understand the role played by the Tlr2 and Tjp1 in the process of wound healing and scar formation, Tlr2, Tjp1gene expression and genetic mRNA was determined with quantitative RT-PCR. The application of our pharmacological composition stimulated the decrease of Tlr2 and Tjp1 gene expression against the background of suppression of free radical processes (reduction of superoxide anion radical content) with epithelization and without scarring. The results of this study have shown the positive effects of melanin on wound healing. The obtained results indicate the advisability of applying melanin for the treatment of inflammatory processes

Manipulation of isolated brain nerve terminals by an external magnetic field using D-mannose-coated γ-Fe2O3 nano-sized particles and assessment of their effects on glutamate transport

The manipulation of brain nerve terminals by an external magnetic field promises breakthroughs in nano-neurotechnology. D-Mannose-coated superparamagnetic nanoparticles were synthesized by coprecipitation of Fe(II) and Fe(III) salts followed by oxidation with sodium hypochlorite and addition of D-mannose. Effects of D-mannose-coated superparamagnetic maghemite (γ-Fe2O3) nanoparticles on key characteristics of the glutamatergic neurotransmission were analysed. Using radiolabeled L-[14C]glutamate, it was shown that D-mannose-coated γ-Fe2O3 nanoparticles did not affect high-affinity Na+-dependent uptake, tonic release and the extracellular level of L-[14C]glutamate in isolated rat brain nerve terminals (synaptosomes). Also, the membrane potential of synaptosomes and acidification of synaptic vesicles was not changed as a result of the application of D-mannose-coated γ-Fe2O3 nanoparticles. This was demonstrated with the potential-sensitive fluorescent dye rhodamine 6G and the pH-sensitive dye acridine orange. The study also focused on the analysis of the potential use of these nanoparticles for manipulation of nerve terminals by an external magnetic field. It was shown that more than 84.3 ± 5.0% of L-[14C]glutamate-loaded synaptosomes (1 mg of protein/mL) incubated for 5 min with D-mannose-coated γ-Fe2O3 nanoparticles (250 µg/mL) moved to an area, in which the magnet (250 mT, gradient 5.5 Т/m) was applied compared to 33.5 ± 3.0% of the control and 48.6 ± 3.0% of samples that were treated with uncoated nanoparticles. Therefore, isolated brain nerve terminals can be easily manipulated by an external magnetic field using D-mannose-coated γ-Fe2O3 nanoparticles, while the key characteristics of glutamatergic neurotransmission are not affected. In other words, functionally active synaptosomes labeled with D-mannose-coated γ-Fe2O3 nanoparticles were obtained

Proteolytic parameter changes in the plasma of patients with bladder cancer – depending on tumor stage

Bladder cancer (BC) is a worldwide common disease with a high mortality rate. Recognizing the dynamic changes in plasma that proteases and their inhibitors undergo might be valuable in understanding the carcinogenesis of invasive bladder cancer and in identifying BC patients with poor prognosis. This study aims to determine the activity of the proteolytic enzyme system and their inhibitors in patients with BC. In this paper, the total proteolytic activity, the activity of matrix metalloproteases (MMPs) and serine proteases was analyzed by the method of caseinolytic activity. For detection of activity of some inhibitors of proteolysis, we used the unified method for determining the activity of alpha-1-antitrypsin (α1A) and alpha-2-Macroglobulin (α2M) in human plasma. The level of medium-mass molecules (MMM) was assessed spectrophotometrically by applying the Nikolaichik method

Disturbances of extracellular protein metabolism in ceruleininduced pancreatitis

Chronic pancreatitis (CP) is still a serious clinical problem due to the significant difficulties in its diagnosis, especially in the initial stages of development. Among the mechanisms that mediate the pathogenesis of CP and lead to pancreatitis-related disorders is unregulated activation of proteolytic enzymes, namely, matrix metalloproteinases (MMPs). The aim of our study was to determine the disturbances of protein metabolism under the conditions of CP alone or in combination with diabetes type 1 (CP+DT1). Herein, CP was induced in the nonlinear male rats by intraperitoneal injection of cerulein (5 µg·kg−1 of body weight; five times during fives day). DT1 was modeled in the rats with CP by a single intraperitoneal injection of streptozotocin (65 mg·kg−1 of the body weight). The levels of MMP-2 and -9 were determined by enzyme-linked immune sorbent assay, and the level of low and middle molecular weight (LMMW) substance was measured spectrophotometrically, while the peptide fractions were analyzed by size exclusion chromatography. The present study revealed a significant increase of MMP-2 and MMP-9 levels in the serum, liver and pancreas of the rats with CP and CP+DT1. Elevated levels of MMPS may act as a factor for the initiation of subsequent cascade of events resulting in the development of pancreatitis-associated complications. Pathogenesis of chronic pancreatitis alone or in combination with diabetes type 1 has been accompanied by the formation and accumulation of LMMW substance, changes in peptide composition and level of individual peptides in the tissues of the rats. Such alterations are among key triggers of amplification of metabolic disorders under chronic pancreatitis

Low-Molecular-Mass Fragments of Collagen Improve Parameters Related to Mass and Inflammation of the Adipose Tissue in the Obese Rat

Research background. Despite clearly recognized links between increased body mass and increased risk for various pathological conditions, therapeutic options to treat obesity are still very limited. The aim of the present study is to explore the effect of low-molecular-mass collagen fragments obtained from the scales of Antarctic wild marine fish on rats' visceral and subcutaneous white adipose tissue in a high-calorie diet-induced obesity model. Experimental approach. The study was conducted on outbred rats, which were divided into 3 experimental groups: (i) control, consuming standard food (3.81 kcal/g), (ii) obese group, consuming a high-calorie diet (5.35 kcal/g), and (iii) obese group, consuming a high-calorie diet (5.35 kcal/g) with intragastric administration of low-molecular-mass collagen fragments (at a dose 1 g/kg of body mass during 6 weeks). Low-molecular-mass collagen fragments were obtained by a procedure that included collagen extraction from fish scales and enzymatic hydrolysis with pepsin. Apart from hematoxylin and eosin staining, fibrosis level was assessed by histochemical Van Gieson’s trichrome picrofuchsin staining, and mast cells were analysed by toluidine blue O staining. Results and conclusions. Group treated with low-molecular-mass fragments of collagen exhibited decreased rate of mass gain, relative mass, area occupied by collagen fibre of both visceral and subcutaneous adipose tissue, and cross-sectional area of both visceral and subcutaneous adipocytes. Treatment with low-molecular-mass fragments of collagen reduced the infiltration of immune cells, number of mast cells and their redistribution back to the septa. This was also accompanied by a decreased number of the crown-like structures formed by the immune cells, which are markers of chronic inflammation that accompanies obesity. Novelty and scientific contribution. This is the first study that reports the anti-obesity effect of low-molecular-mass fragments produced as a result of controlled hydrolysis of collagen from the scales of Antarctic wild marine fish in the in vivo model. Another novelty of this work is the observation that the tested collagen fragments not only reduce the body mass, but also improve the morphological and inflammatory parameters (decrease in the number of crown-like structures, immune cell infiltration, fibrosis and mast cells). Altogether, our work suggests that low-molecular-mass collagen fragments are a promising candidate for amelioration of some comorbidities linked to obesity

Hypoglycemic activity of Phaseolus vulgaris (L.) aqueous extract in type 1 diabetic rats

The aim of the present study was to evaluate the hypoglycemic activity of the aqueous extract from the fruit walls of Phaseolus vulgaris pods and to examine the potential mechanism underlying the improvement of the glycemic level. In the course of the study, diabetes mellitus was induced in rats with a single intraperitoneal injection of streptozotocin (45 mg·kg−1 b.w.). Diabetic and control rats were then orally administered with a single-dose or repeated-dose (28 day) of P. vulgaris extract (200 mg·kg−1). Results show that the extract was found to possess significant hypoglycemic activity, and the study of glucose utilization by isolated rat hemidiaphragm suggests that the aqueous extract may enhance the peripheral utilization of glucose. The subsequent experiments have revealed that the P. vulgaris extract could increase glucose transporter 4 (GLUT-4) content in skeletal muscle cells of control and diabetic rats. Our data also indicate that the P. vulgaris extract did not affect the content of the insulin receptor, but significantly reduced the total tyrosine kinase activity in skeletal muscle cells of both experimental groups of rats. The present results clearly indicated that P. vulgaris extract may be beneficial for reducing hyperglycemia through its potency in regulation of glucose utilization via GLUT-4, but the current mechanism remains to be unidentified