Total body irradiation is a crucial risk factor for developing secondary carcinomas after allogeneic hematopoietic stem cell transplantation in childhood.

Allogeneic hematopoietic stem cell transplantation (HSCT) has become a standard part of therapy for a variety of malignant and non-malignant disorders. With improved outcomes after HSCT, increasing attention has been drawn to late complications in long-term survivors. The development of secondary malignancies is recognized as one of the most serious complications. We have evaluated data from 426 patients (272 males, 154 females) who underwent allogeneic transplantation at a median age of 7.9 years from 1989 till 2017 and were alive more than one year after transplantation for the occurrence of secondary solid tumors. We have documented the occurrence of secondary solid tumors in 20 patients (4.7%). The median duration of the development of secondary solid cancer from HSCT was 11.7 (range, 5.4-21.5 years). 18 out of 20 patients (90%) had total body irradiation (TBI) 12-14.4 Gy as a part of a conditioning regimen. All but two had transplantation for malignant disease. All patients underwent surgery and/or chemo-radiotherapy. Eighteen are alive, and two died due to the progression of their secondary malignancy. The most frequent solid cancer was thyroid carcinoma (n=9). Cumulative incidence of secondary solid cancer in all groups was 15.2±3.9%, in a group using TBI based regimen 34.7±8.9%, in non-TBI (only chemo) group was 1.5±1.1%. Overall, the cumulative incidence is statistically significantly different between the TBI based and non-TBI (chemo only) group. The incidence and number of complications following allogeneic HSCT in childhood are increasing in time. The early diagnosis of secondary malignancies is one of the key tasks of long-life multidisciplinary post-transplant care.

Post-mortem analysis of Candida albicans breakthrough infection during echinocandin treatment in haematopoietic stem cell transplant recipient.

We present case of a girl deceased due to Candida albicans breakthrough invasive infection during the echinocandin treatment after undergoing allogeneic haematopoietic stem cell transplant for relaps of acute myeloid leukaemia. Candida albicans generally susceptible to all antifungal drugs wasn't considered for potential resistance and conventional blood culture positivity was too late to reveal the resistance to echinocandins. Due to severe organ toxicities (liver, kidneys) she received echinocandin as an antifungal prophylaxis, no change was made for the treatment of Candida albicans infection. Later, the molecular analysis proved the mutation S645P known as being responsible for the echinocandin resistance. The post mortem analysis of fungal burden in autopsy samples showed very high levels of Candida DNA in gut, liver, spleen and kidneys.

Animal model of posterior cingulate cortex hypometabolism implicated in amnestic MCI and AD.

The posterior cingulate cortex (PCC) is the brain region displaying the earliest sign of energy hypometabolism in patients with amnestic mild cognitive impairment (MCI) who develop Alzheimer's disease (AD). In particular, the activity of the mitochondrial respiratory enzyme cytochrome oxidase (C.O.) is selectively inhibited within the PCC in AD. The present study is the first experimental analysis designed to model in animals the localized cortical C.O. inhibition found as the earliest metabolic sign of early-stage AD in human neuroimaging studies. Rats were used to model local inhibition of C.O. by direct injection of the C.O. inhibitor sodium azide into the PCC. Learning and memory were examined in a spatial holeboard task and brains were analyzed using quantitative histochemical, morphological and biochemical techniques. Behavioral results showed that sodium azide-treated rats were impaired in their memory of the baited pattern in probe trials as compared to their training scores before treatment, without non-specific behavioral differences. Brain analyses showed that C.O. inhibition was specific to the PCC, and sodium azide increased lipid peroxidation, gliosis and neuron loss, and lead to a network functional disconnection between the PCC and interconnected hippocampal regions. It was concluded that impaired memory by local C.O. inhibition in the PCC may serve to model in animals a metabolic lesion similar to that found in patients with amnestic MCI and early-stage AD. This model may be useful as an in vivo testing platform to investigate neuroprotective strategies to prevent or reduce the amnestic effects produced by posterior cingulate energy hypometabolism.

Studies of electrorheological properties of blood.

The electrorheological (ER) properties of blood indicate changes in the blood rheological behaviour due to imposition of electric field. The present work identifies and quantifies ER properties of blood at different shear rates and at different local structure of the flow field. A concurrent measurement system, based on a Contraves Low Shear 30 rotational rheometer was used in this study. It includes a pair of cylindrically shaped platinum electrodes, embedded into the wall of a resin replica of the Couette type flow chamber of the rheometer, constructed for conductivity measurement, and associated software (Data acquisition system). The relationship between the whole blood and plasma conductivity (the main active component of blood impedance) was studied in parallel with the changes in the rheological behaviour under steady and transient flow conditions. The time variation of blood conductivity at different flow regimes and the dependences of the apparent whole blood and plasma viscosity were investigated in the presence and absence of an electric field of 2 kHz. The results show that blood conductivity is strongly dependent on the blood factors considered and that any application using blood conductivity measurement should take into account the effect of flow, shear rates and hematocrit. The results also show that valuable information on the mechanical properties of blood can be obtained, in particular concerning the structuring and kinetics of "rouleaux formation".

Transient hypobaric hypoxia improves spatial orientation in young rats.

To achieve a better understanding of learning and declarative memory under mild transient stress, we investigated the effect of brief hypobaric hypoxia on spatial orientation in rats. Young male Wistar rats aged 30 days were exposed for 60 min to hypobaric hypoxia, simulating an altitude of 7,000 m (23,000 ft) either shortly prior to attempting or after mastering an allothetic navigation task in the Morris water maze with a submerged platform. The post-hypoxic group performed significantly better in the navigation task than the control animals (the mean difference in escape latencies was 11 seconds; P=0.0033, two-way ANOVA with repeated measures, group x session). The experimental group also achieved a remarkably higher search efficiency (calculated as a percentage of successful trials per session), especially during the first four days following hypoxic stress (P=0.0018). During the subsequent training, the post-hypoxic group performed better than the control animals, whilst the efficiency levels of both groups progressively converged. Spatial memory retention and recall of well-trained rats were not affected by the transient hypobaric hypoxia. These results indicate that brief hypobaric hypoxia enhances rats' spatial orientation. Our findings are consistent with several studies, which also suggested that mild transient stress improves learning.

Functionalized Multi-Walled Carbon Nanotube Paper for Monitoring Chemical Vapors.

The multi-walled carbon nanotube paper is prepared by vacuum filtration of pure nanotubes and their functionalized forms prepared by KMnO4 and H2O2 oxidation or by grafting with Poly(methyl methacrylate) (PMMA) and polypyrrole to form sensory nanotubes layer for detection of volatile organic compounds in air. The selected compounds for experimentation (acetone, diethyl ether, isopentane, methanol, tetrahydrofuran) have different polarities and volume fractions of saturated vapors. The sensing is measured by electrical resistance of the paper, which increases when exposed to vapors. A reversible reaction is observed when the paper is removed from the vapors. The functionalized nanotubes differ in their sensitivity to selected organics solvents. For example, KMnO4 oxidized paper has differentiated response to all used vapors, so the measured data may indicate clearly the type of the vapor. On the other hand, the MWCNT/PMMA composite has nearly the same response to acetone, diethyl ether and tetrahydrofuran and different response to isopentane and methanol. The investigation can lead to construction of sensory unit which could be capable of detecting and identifying different vapors in the air.

The cooperative effect of L-selectin clusters and velocity-dependent bond formation that stabilizes leukocyte rolling.

The role of clustered of L-selectin receptors on the leukocyte surface is discussed in connection with the postulated velocity-dependent formation of selectin-ligand tether bonds to interpret the mechanism of leukocyte tethering to, and rolling along, the vascular endothelium. The distinct feature of this step-wise process is a weak dependence of leukocyte rolling velocity on the hydrodynamic forces of ambient flow due to the increased number of selectin bonds with increasing flow shear rate and also their clustering. The contact zones on the leukocyte surface are separated by distances with distribution which corresponds to the distribution of distances of the observed L-selectin clusters. It suggests that the localization of L-selectin receptors to clusters and the way of their approach to the ligand molecules creates such conditions for binding of L-selectin and ligand molecules that resulting number of bonds stabilizes rolling velocity.

Coagulation and hemorheology.

The application of principles of hemorheology on a process of blood coagulation was suggested by H.H. Hartert - inventor of the thromboelastograph which he introduced in 1947. These principles of applying of a mechanical force and detecting the response of coagulating blood enable to determine a functional evidence of the physiological process producing a hemostatic clot. The thromboelastography, as well as other rheological techniques used for this determination, are introduced evaluating the process in terms of the rate of coagulation, clot elasticity, deformability and permeability. Recent applications of the techniques to the diagnosis of disorders of hemostasis and the study of drug effects on coagulation are presented as well.

The effect of rouleaux formation on blood coagulation.

The coagulation of whole blood resulting a blood clot is initiated in vitro for studying the effect of rouleaux formation on the kinetics of coagulation and clot formation. The clot formation is monitored in the rotational viscosimeter and in the erythroaggregometer using a light backscattered technique. The intensity of flow as well as hematocrit were changed in a way to determine a tendency in the effect of rouleaux size on the rate of coagulation. The results indicate that the clot formation is promoted when size of rouleaux is small at higher shear rates and low hematocrit in comparison with the coagulation at low shear rates and high hematocrit when the size of rouleaux is increased.

Elasticity and fracture strain of whole blood clots.

The measurement of clot elasticity and fracture strain is carried out using a rotational rheometer with a controlled stress system. The elasticity is quantified by a shear elastic modulus and fracture strain by a critical strain (deformation) when the clot begins to break up. The results indicate a decrease of elasticity and increase of fracture strain of the clot with increasing hematocrit. Moreover, the elastic modulus of the clot is not constant but increasing with deformation.

[Leukocyte adhesion on a fibrinogen-coated surface under static conditions: experimentation and creation of a model]. / Adhésion des leucocytes sur une surface recouverte de fibrinogène en conditions statiques: expíence et modélisation]

The adhesion of polymorphonuclear leukocytes (PMNs) on the vascular endothelium is a complex process that occurs during different biological and pathological events and involves numerous molecules. The adhesion cascade is induced after PMN stimulation by various molecular or cellular signals. Fibrinogen is one of the substrates for CD11b/CD18 B2-integrins expressed at the PMN surface; fibrinogen-neutrophil binding is induced by inflammatory reactions. In order to understand this process, we have carried out studies on the basis of preliminary experiments on red blood cells and synthetic particles. The modelization of quiescent PMNs adhesion on a fibrinogen substrate was investigated with a sedimentation cell chamber. Two different physiological conditions were tested: the activated state of PMN by a synthetic pro-inflammatory activator (FMLP). The activated state of PMNs was both quantified by flow cytometry and controlled by fluorescence microscopy. The results suggest that quiescent neutrophils deposit in accordance with the ballistic deposition model. This random adsorption model differs from random sequential adsorption (RSA) in that the cells arriving at the surface are able to roll along cells previously adsorbed introducing the notion of gravitational attraction of cells. The preliminary results obtained with stimulated PMN do not allow to choose between one of this two deposition models. Nevertheless, the qualitative and quantitative effects of FMLP on neutrophils were demonstrated by modifications of adhesion molecules expression.

Summary of the Czech medical resources in the Internet.

The article describes the most important features of the computer network Internet. Various information services are presented such as E-mail, Listserv, Gopher, FTP and World Wide Web. The Internet contribution to medical education, research contacts, and information exchange is particularly emphasized. Medical information sources from the Czech and Slovak Republics are summarized in appendices.

Effects of chronic NMDA-NR2b inhibition in the median eminence of the reproductive senescent female rat.

Gonadotrophin-releasing hormone (GnRH) neurones of the hypothalamic-pituitary-gonadal (HPG) axis drive reproductive function and undergo age-related decreases in activation during the transition to reproductive senescence. Decreased GnRH secretion from the median eminence (ME) partially arises from attenuated glutamatergic signalling via the NMDA receptor (NMDAR) and may be a result of changing NMDAR stoichiometry to favour NR2b over NR2a subunit expression with ageing. We have previously shown that the systemic inhibition of NR2b-containing receptors with ifenprodil, an NR2b-specific antagonist, stimulates parameters of luteinising hormone (used as a proxy for GnRH) release in both young and middle-aged females. In the present study, we chronically administered ifenprodil, an NR2b-specific antagonist, at the site of GnRH terminals in the ME or at GnRH perikarya in the preoptic area, in reproductively senescent middle-aged female rats, aiming to determine whether NR2b antagonism could restore aspects of reproductive functionality. Effects on oestrous cyclicity, serum hormones, and protein expression of GnRH, NR2b and phosphorylated NR2b (Tyr-1472) in the ME were measured. Chronic ifenprodil treatment in the ME (but not the preoptic area) altered oestrous cyclicity by increasing the percentage of days spent in pro-oestrus. This was accompanied by increased GnRH fluorescence intensity in the external ME zone and a greater proportion of GnRH terminals that co-labelled with pNR2b with treatment. We also observed changes in the relationships between protein immunofluorescence, serum hormone levels and other aspects of reproductive physiology in acyclic females, as revealed by bionetwork analysis. Together, these data support the hypothesis that NMDAR-NR2b expression and phosphorylation state play a role in reproductive senescence and highlight the ME as a major player in reproductive ageing.

Experimental evaluation of mechanical and electrical properties of RBC suspensions in Dextran and PEG under flow II. Role of RBC deformability and morphology.

Mechanical and electrical properties of the normal RBCs suspensions and of hardened after treatment with glutaraldehyde (0.01-2.5%) RBCs in isotonic physiological solution and Dextran 70,000 (Dextran 70) and Polyethylene glycol 35,000 (PEG) and adjusted to hematocrit of 40%, were evaluated. Apparent viscosity and conductivity were measured under steady and transient flow regimes at low shear rates and at different local structure of the flow at 37 °C. A time course of conductivity was recorded in parallel with the rheological properties of the RBC suspensions and conductivity and apparent viscosity dependences on shear rates were studied and compared at different concentrations of Dextran 70, PEG and glutaraldehyde. Low shear viscosity decreased after RBCs treatment with glutaraldehyde and at 0.5-2.5% it is constant. Echinocytes are observed at low Dextran 70 and PEG concentrations while spherocytes are found mainly in smears treated with higher concentrations. The results show that the apparent viscosity and conductivity of RBCs suspensions in Dextran 70 and PEG are strongly influenced by flow, shear rates, concentration, cell deformability and morphology and the method is sensitive to study the mechanical and electrical properties of RBC suspension and to provide experimental description of RBCs and other cell-to-cell interactions.

Experimental evaluation of mechanical and electrical properties of RBC suspensions under flow. Role of RBC aggregating agent.

Mechanical and electrical properties of red blood cells (RBC) suspensions in dextran 70 (Dx70), dextran 150 (Dx150), dextran 500 (Dx500) and polyethileneglycol (PEG) 35,000 with different concentrations were evaluated through apparent viscosity and conductivity measurements under steady and unsteady flow conditions. RBCs suspensions of the washed RBS in PBS (control) and Dx70, Dx150, Dx500 and PEG in PBS with different concentrations, adjusted to the same hematocrit of 40% were used for the experiments. Conductivity time and shear rate dependences in parallel with the rheological properties of the samples were studied under transient flow regimes at different local structure of the uniform Couette flow. Their relationships on dextrans and PEG concentrations were evaluated too. Low shear viscosity increased and conductivity decreased of RBC suspensions, compared to non-aggregating suspensions, depending on dextrans and PEG concentrations. A time course of blood conductivity recorded under different flow conditions provides experimental description of RBC aggregation-disaggregation processes and other cell-cell interactions. The results show that the blood conductivity is strongly dependent on the considered blood factors and is influenced by flow, shear rates and concentration of dextran and PEG solutions.

Effect of fibrin polymerzation on flow properties of coagulating blood.

The evolution of stress in coagulating blood is described by aMaxwell-like constitutive model. The evolution is essentiallyaffected by conversion of fibrinogen to fibrin monomer, fibrinpolymerization and its crosslinking. The modifying effects of theprocess result from the active constituents of blood plasma,blood cell concentration and intensity of flow. Interrelationof stress evolution and the kinetics of polymerization species these effects in terms of the order of the kinetics ofpolymerization and degree of fibrin polymerization and values of the constitutive coefficients. The results are presented for a normal blood, averaging the effect of blood constituents and blood platelets on coagulation.

The strength of integrin binding between neutrophils and endothelial cells.

The firm adhesion of activated polymorphonuclear neutrophils to endothelial cells in blood vessels is achieved through binding of the integrin intercellular adhesion molecule. To contribute to the better understanding of this adhesion step, our investigation is aimed at the relationship between integrin expression and the strength of neutrophil binding to endothelial cells. Flow cytometry and 3D scanning microscopy are used to study integrin expression and distribution, respectively. It is found that CD11b/CD18 integrin expression is localized in clusters distributed irregularly over the neutrophil surface. After cell activation, the cluster distribution polarizes, increasing the local CD11b/CD18 density concurrently with nearly doubled integrin expression. The neutrophil adhesion efficiency is measured in a flow chamber coated successively by various substrates, including endothelial cells in an activated state. Analysis of the flow dependence of the number of attached cells reveals the prevailing number of neutrophils with stronger binding to the endothelium when both cells are in the activated state in comparison with non-activated cells.