uPA-silica-Particles (SP-uPA): a novel analytical system to investigate uPA-uPAR interaction and to test synthetic uPAR antagonists as potential cancer therapeutics

The urokinase-type plasminogen activation system, including the serine protease uPA (urokinase-type plasminogen activator) and its cell surface receptor (uPAR, CD87), are important key molecules in tumor invasion and metastasis. Besides its proteolytic function, binding of uPA to uPAR on tumor cells exerts various cell responses such as migration, adhesion, proliferation, and differentiation. Hence, the uPA/uPAR system is a potential target for tumor therapy. We have designed a new generation of uPA-derived synthetic cyclic peptides suited to interfere with the binding of uPA to uPAR and present a new technology involving micro silica particles coated with uPA (SP-uPA) and reacting with recombinant soluble uPAR (suPAR), to rapidly assess the antagonistic potential of uPA-peptides by flow cytofluorometry (FACS). For this, we used silica particles of 10 microm in diameter to which HMW-uPA is coupled using the EDC/NHS method. Soluble, recombinant suPAR was added and the interaction of SP-uPA with suPAR verified by reaction with monoclonal antibody HD13.1 directed to uPAR, followed by a cyan dye (cy5)-labeled antibody directed against mouse IgG. Thereby it was possible to test naturally occurring ligands of uPAR (HMW-uPA, ATF) as well as highly effective, synthetic cyclic uPA-derived peptides (cyclo21,29[D-Cys21Cys29]-UPA21-30, cyclo21,29[D-Cys21Nle28Cys29]-uPA21-30, cyclo21,29[D-Cys(21)2-Nal24Cys29]-uPA21-30, and cyclo21,29[D-Cys21Orn23Thi24Thi25Cys29]-uPA21-30. The results obtained with the noncellular SP-uPA/uPAR system are highly comparable to those obtained with a cellular system involving FITC-uPA and the promyeloid cell line U937 as the source of uPAR.     

Differential genotoxic effects of antitumor trans-[PtCl(2)(E-iminoether)(2)] and cisplatin in Escherichia coli

In the present work, we measured survival and the platinum on the genome after treatment of repair-proficient or repair-deficient Escherichia coli strains with trans-[PtCl₂(E-iminoether)₂] and compared these results with the effects of “classical” cisplatin. We found that toxicity of antitumor trans-[PtCl₂(E-iminoether)₂] in repair-deficient trains was much less than that of cisplatin. This markedly reduced toxicity was not a consequence of the reduced uptake or low levels of DNA binding in the bacteria cells but rather appeared to reflect DNA binding mode of this trans-platinum drug different from that of cisplatin.     

Ultrasonographic assessment of bone maturity in newborns

BACKGROUND: Traditionally, the bone maturity at birth has been estimated from the radiological presence and size of the ossified distal femoral epiphysis. This study was conducted in a search for a sonographic tool for the evaluation of neonatal bone maturity. METHODS: We examined sonographically 256 neonates within 24 h of birth. Gestational ages ranged from 36 to 42 weeks (mean: 39.4; median: 40). Birth weights ranged from 1,945 to 5,000 g (mean: 3,175; median: 3,180). The distal femoral epiphysis was imaged on the coronal plane sonogram of the distal femur with the knee at 90 degrees flexion and the distal femoral epiphysis maximal height was recorded. The acetabulum was imaged using Graf's method in the coronal plane image and the acetabular diameter recorded. RESULTS: It was found that plotting the distal femoral epiphysis against neonatal birth weight and gestational age provided a simple method for assessing the bone maturity. According to our study, a neonate can be regarded as bone maturity percentile X when plotting distal femoral epiphysis height or acetabulum diameter against birth weight and gestational age or when averaging the four readings. CONCLUSIONS: We suggest performing sonography of the distal femoral epiphysis as a bedside tool for the assessment of skeletal maturity in newborns.     

Adiponectin concentrations as a criterion of metabolic control in persons with type 2 diabetes mellitus?

Adiponectin (ADP) is an adipocytokin with many antiatherogenic properties; its decreased level is associated with numerous atherogenic diseases and syndromes (e.g. diabetes mellitus (DM), dyslipidemia, endothelial dysfunction, hypertension, and obesity). Decreased ADP values in blood may be an independent risk factor of atherosclerotic (ATS) complications. AIM OF THE STUDY: 1) Do persons with type 2 diabetes have lower ADP values than individuals without DM but with a high risk of ATS complications? 2) Do ADP values differ between persons with well controlled and persons with uncontrolled type 2 diabetes? We examined 109 patients of the Metabolic Center of Hospital Sternberk. Out of them, 58 had type 2 diabetes, others were individuals with variously expressed risk factors of early atherosclerosis (obesity, hypertension, age, family history, smoking, dyslipidemia, etc.). In all persons under this study the following parameters were determined in peripheral venous blood: adiponectin, resistin, leptin, ObRe, cholesterol, HDL-cholesterol, triacylglycerols, glucose, HbA1c, creatinine, urea, ALT, AST, CRP, homocysteine, thrombocyte aggregation after CPG induction. The whole group was divided according to the presence of type 2DM into two subgroups; persons with diabetes were divided into the well controlled and uncontrolled subgroups. All data obtained were processed statistically using the software SPSS for Windows and Medcalc. The adiponectin/BMI index correlated negatively with HbA1c value (correlation coefficient -0.37, p = 0.00053), triacylglycerols (-0.4, p = 0.000001), P-glucose (-0.3, p = 0.0017), uricemia (-0.35, p = 0.0007) and positively with HDL-cholesterol value (0.6, p=0.00001). Women had higher adiponectin values than men. Persons with hypertension and with diabetes mellitus, individuals with atherogenic lipotype or persons with inflammation signs had lower values than individuals without these diseases and syndromes. Persons with wellcontrolled diabetes mellitus had higher values than persons with uncontrolled diabetes (medians of the adiponectin/BMI index 9.7 vs. 6.7, p < 0.01). Persons with type 2 diabetes mellitus have lower ADP values than persons with a high ATS risk without diabetes mellitus. Persons with wellcontrolled diabetes mellitus (DM) and with satisfactory compensation have significantly higher ADP levels (independently of other metabolic parameters of DM control). ADP may be a new marker of metabolic control in persons with a high risk of atherosclerotic complications.     

Modelling production and biomasses of zoobenthos in lakes

This work presents a dynamic model to predict zoobenthos in lakes. The model has been developed within the framework of a more comprehensive lake ecosystem model, LakeWeb, which also accounts for the following functional groups of organisms, phytoplankton, bacterioplankton, two types of zooplankton (herbivorous and predatory), macrophytes, prey fish and predatory fish. This work also presents a new data-base for zoobenthos in lakes. Many of the lakes included in this study are situated in the former Soviet Union. They were investigated during the Soviet period and those results have been largely unknown in the West. Using this data-base, this work also presents new empirical models for zoobenthos. The new dynamic model gives seasonal variations (the calculation time, dt, is 1 week using Euler's method and enough iterations to get stable solutions). The basic aim of the dynamic model is that it should capture general functional and structural patterns in lakes. We have demonstrated by several model tests along limnological gradients (total phosphorus concentrations, pH, lake colour, latitude and lake size) that the dynamic model gives predictions that agree well with the values given by the empirical regressions, and also expected and requested divergences from these regressions when they do not provide sufficient resolution. It would have been very difficult indeed to carry out such tests regarding ecosystem responses using traditional methods with extensive field studies in a few lakes. We have given algorithms for (1) production of zoobenthos from eating macrophytes, benthic algae and sediments, (2) elimination (related to the turnover time of zooplankton), and (3) zoobenthos consumption by prey fish, and the factors influencing these processes/rates. The model is driven by data easily accessed from standard monitoring programs or maps a prerequisite for practical utility in contexts of lake management.     

Melting of cross-linked DNA v. cross-linking effect caused by local stabilization of the double helix

DNA interstrand cross-links are usually formed due to bidentate covalent or coordination binding of a cross-linking agent to nucleotides of different strands. However interstrand linkages can be also caused by any type of chemical modification that gives rise to a strong local stabilization of the double helix. These stabilized sites conserve their helical structure and prevent local and total strand separation at temperatures above the melting of ordinary AT and GC base pairs. This local stabilization makes DNA melting fully reversible and independent of strand concentration like ordinary covalent interstrand cross-links. The stabilization can be caused by all the types of chemical modifications (interstrand cross-links, intrastrand cross-links or monofunctional adducts) if they give rise to a strong enough local stabilization of the double helix. Our calculation demonstrates that an increase in stability by 25 to 30 kcal in the free energy of a single base pair of the double helix is sufficient for this "cross-linking effect" (i.e. conserving the helicity of this base pair and preventing strand separation after melting of ordinary base pairs). For the situation where there is more then one stabilized site in a DNA duplex (e.g., 1 stabilized site per 1000 bp), a lower stabilization per site is sufficient for the "cross-linking effect" (18 - 20 kcal). A substantial increase in DNA stability was found in various experimental studies for some metal-based anti-tumor compounds. These compounds may give rise to the effect described above. If ligand induced stabilization is distributed among several neighboring base pairs, a much lower minimum increase per stabilized base pair is sufficient to produce the cross-linking effect (1 bp- 24.4 kcal; 5 bp- 5.3 kcal; 10 bp- 2.9 kcal, 25 bp- 1.4 kcal; 50 bp- 1.0 kcal). The relatively weak non-covalent binding of histones or protamines that cover long regions of DNA (20- 40 bp) can also cause this effect if the salt concentration of the solution is sufficiently low to cause strong local stabilization of the double helix. Stretches of GC pairs more than 25 bp in length inserted into poly(AT) DNA also exhibit properties of stabilizing interstrand cross-links.     

Analysis of multidimensional neural activity via CNN-UM

In this paper we show that the Cellular Nonlinear Network Universal Machine (CNN-UM) is an excellent tool for analyzing time series of multidimensional binary signals. The developed algorithm is dedicated to process electrophysiological multi-neuron recordings: our aim is to find specific multidimensional activity patterns, which may reflect higher order functional cell-assemblies. The analysis consists of two parts: first, the occurrences of different patterns are counted, then the statistical significance of each occurrence frequency is calculated separately.