Formation of polyelectrolyte multilayers and polyelectrolyte complexes in dual polymer treatment of papermaking pulp

The goal of this study was to find a new approach to modify chemically the properties of paper by improving fiber quality. This Master’s thesis includes the multiple polymer treatment in general and themeasurement methods with which the formation of multilayers and complexes can be noticed. The treatment by an oppositely charged dual polymer system is a good approach to increase paper strength. In this work, starch, a cationic polymer, and carboxymethyl cellulose (CMC), an anionic polymer, were used step-by-step to improve paper strength. The adsorption of cationic starch and CMC on cellulose fibers were analyzed via polyelectrolyte titration. The results showed that paper strength was enhanced slightly with a layer-by-layer assembly of the polymers. However, if the washing stage, which was required for layer-by-layer assembly, was eliminated, the starch/CMC complex was deposited on fibers more efficiently, and the paper strength was improved more significantly.

Seed germination of a rare neotropical canopy tree dormancy and the effects of abiotic factors

The purpose of this study was to examine if germination is a critical phase on Enterolobium glaziovii regeneration. Hence, the germinative response of E. glaziovii seeds was investigated in relation to some of the main environmental factors (temperature, light and water stress) to which its seeds are subjected in the forest, as well as its dormancy and the longevity of its burial seeds. According to our results, its seeds may be regarded as photoblastic neutral. They do not need alternating temperatures to germinate and can germinate under a broad range of water stress. However, only about 10% of E. glaziovii seeds remain viable after one year. In other words, the annual fruiting, instead seed longevity, seems to maintain the long-term seed availability of this species. Consequently, the seed longevity could be a critical phase of E. glaziovii germination.

Molecular diagnostics of rare inherited SYNDROMES with a view on diagnostic test development

CHARGE syndrome, Sotos syndrome and 3p deletion syndrome are examples of rare inherited syndromes that have been recognized for decades but for which the molecular diagnostics only have been made possible by recent advances in genomic research. Despite these advances, development of diagnostic tests for rare syndromes has been hindered by diagnostic laboratories having limited funds for test development, and their prioritization of tests for which a (relatively) high demand can be expected. In this study, the molecular diagnostic tests for CHARGE syndrome and Sotos syndrome were developed, resulting in their successful translation into routine diagnostic testing in the laboratory of Medical Genetics (UTUlab). In the CHARGE syndrome group, mutation was identified in 40.5% of the patients and in the Sotos syndrome group, in 34%, reflecting the use of the tests in routine diagnostics in differential diagnostics. In CHARGE syndrome, the low prevalence of structural aberrations was also confirmed. In 3p deletion syndrome, it was shown that small terminal deletions are not causative for the syndrome, and that testing with arraybased analysis provides a reliable estimate of the deletion size but benign copy number variants complicate result interpretation. During the development of the tests, it was discovered that finding an optimal molecular diagnostic strategy for a given syndrome is always a compromise between the sensitivity, specificity and feasibility of applying a new method. In addition, the clinical utility of the test should be considered prior to test development: sometimes a test performing well in a laboratory has limited utility for the patient, whereas a test performing poorly in the laboratory may have a great impact on the patient and their family. At present, the development of next generation sequencing methods is changing the concept of molecular diagnostics of rare diseases from single tests towards whole-genome analysis.

Probabilistic contact preferences in protein-ligand and protein-protein complexes

Modeller för intermolekulär växelvärkan utnyttjas brett inom biologin. Analys av kontakter mellan proteiner och läkemedelsforskning representerar typiska tillämpningsområden för dylika modeller. En modell som beskriver sådana molekylära växelverkningar kan utformas med hjälp av biofysisk teori, vilket tenderar att resultera i ytterst tung beräkningsbörda även för enkla tillämpningar. Ett alternativt sätt att formulera modeller är att utnyttja stora databaser som innehåller strukturmätningar gjorda med hjälp av till exempel röntgendiffraktion. Då man använder sig av empiriska mätdata direkt, möjliggör en statistisk modell att osäkerheten och inexaktheten i datat tas till hänsyn på ett adekvat sätt, samtidigt som beräkningsbördan håller sig på en rimligare nivå jämfört med kvantmekaniska metoder som i princip borde ge de optimala resultaten. I avhandlingen utvecklades en 3D modell för numerisk undersökning av intermolekulär växelverkan baserad på Bayesiansk statistik. Modellens syfte är att åstadkomma prognoser för det hurdana eller vilka molekylstrukturer prefereras i en given kontext, d.v.s. är mer sannolika inom ramen för interaktion. Modellen testades i essentiella molekyläromgivningar - en liten molekyl vid sin bindningsplats hos ett protein och en gränsyta mellan proteinerna i ett komplex. De erhållna numeriska resultaten motsvarar väl experimentella resultat som tidigare rapporterats i litteraturen, exempelvis kvalitativa bindningsaffiniteter och kemisk kännedom av vissa aminosyrors rumsliga förmågor att utgöra bindningar. I avhandlingen gjordes ytterligare preliminära tester av den statistiska ansatsen för modellering av den centrala molekylära strukturella anpassningsbarheten. I praktiken är den utvecklade modellen ämnad som ett led i en mer omfattande analysmetod, så som en s.k. farmakofor modell. Molekyylivuorovaikutusten mallintamista hyödynnetään laajasti biologisten kysymysten tarkastelussa. Tyypillisiä esimerkkejä sovelluskohteista ovat proteiinien väliset kontaktit ja lääkesuunnittelu. Vuorovaikutuksia kuvaavan mallin lähtökohta voi olla molekyyleihin liittyvä teoria, jolloin soveltamiseen liittyvä laskenta saattaa olla erityisen raskasta, tai suuri havaintojoukko joka on saatu aikaan esimerkiksi mittaamalla rakenteita röntgendiffraktio menetelmällä. Tilastollinen malli mahdollistaa havaintoaineistossa olevan epätarkkuuden ja epävarmuuden huomioimisen, samalla pitäen laskennallisen kuorman pienempänä verrattuna periaatteessa parhaan tuloksen antavaan kvanttimekaaniseen mallinnukseen. Väitöstyössä kehitettiin bayesiläiseen tilastotieteeseen perustuva 3D malli molekyylien välisten vuorovaikutusten laskennalliseen tarkasteluun. Mallin tehtävä on tuottaa ennusteita sen suhteen, minkä tai millaisten molekyylirakenteiden väliset kompleksit ovat etusijalla, toisin sanoen todennäköisempiä, vuorovaikutustilanteessa. Työssä kehitetyn menetelmän toimivuutta testattiin käyttötarkoituksen suhteen olennaisissa molekyyliympäristöissä - pieni molekyyli sitoutumiskohdassaan proteiinissa sekä rajapinta kahden proteiinin välilllä proteiinikompleksissa. Saadut laskennalliset tulokset vastasivat hyvin vertailuun käytettyjä kirjallisuudesta saatuja kokeellisia tuloksia, kuten laadullisia sitoutumisaffiniteetteja, sekä kemiallista tietoa esimerkiksi tiettyjen aminohappojen avaruudellisesta sidoksenmuodostuksesta. Väitöstyössä myös alustavasti testattiin tilastollista lähestymistapaa tärkeän molekyylien rakenteellisen mukautuvuuden mallintamiseen. Käytännössä malli on tarkoitettu osaksi jotakin laajempaa analyysimenetelmää, kuten farmakoforimallia.

Structural studies and in vivo characterisation of Oct4:PORE and Oct4/Sox2:UTF1 complexes

Monikykyiset kantasolut pystyvät erilaistumaan alkion kehityksen aikana miksi tahansa kehittyvän eläimen kudokseksi. Alkion kehitystä on tutkittu vuosikymmeniä ja ensimmäisen kerran kantasoluja viljeltiin 1980-luvulla. Noin vuosikymmen myöhemmin kloonattiin ensimmäinen nisäkäs, Dolly-lammas, mikä oli läpimurto perimän kloonauksessa. Vajaa kymmenen vuotta sitten japanilaiset tutkijat pystyivät ensimmäisen kerran tuottamaan indusoituja monikykyisiä kantasoluja. Nämä kantasolut tuotettiin ilmentämällä neljää geeniä, jotka uudelleenohjelmoivat erilaistuneet solut takaisin kantasoluiksi. Geenien ilmentymistä säätelevä ja DNA:han sitoutuvan proteiinin Oct4:n on todettu olevan tärkeä tekijä uudelleenohjelmissa. Oct4 kuuluu transkriptiotekijäperheeseen, joka säätelee geenien ilmentymistä kantasoluissa. Tässä työssä kiteytin Oct4-proteiinin sen kohde-DNA:n kanssa sekä tutkin sen sitoutumista ja atomitason rakennetta röntgensädekristallografian avulla. Tutkimuksessamme paljastui useita Oct4:n atomitason rakenteellisia ominaisuuksia, jotka selittävät sen säätelymekanismeja kantasoluissa.

Mimosa splendida Barneby (Mimosoideae, Leguminosae) rediscovered in Central Brazil: preliminary studies for conservation of a rare species

The poorly known Mimosa splendida Barneby has been rediscovered in the Chapada dos Veadeiros, Brazil, 105 years after its single record in 1895. Data on its habitat description, demography, reproductive parameters, and geographic range are presented. Preliminary results suggest that the rarity of M. splendida in herbarium collections is associated with a narrow geographic range, and a specialized habitat preference on rocky slopes.

Juvenile dynamics of the endemic and rare Enterolobium glaziovii Benth. (Mimosaceae) around reproductive trees in the Atlantic forest, Brazil

Studies on the regeneration and seedling mortality of rare tree species are important, but scarce. The aim of this study was to investigate the annual variation in recruitment, growth and mortality of juveniles of Enterolobium glaziovii Benth., a rare tree species from the Brazilian Atlantic Rain Forest. All seedlings and juveniles around four reproductive trees were labeled and their fate was followed from 1996 to 1999. There were no annual differences in juveniles' recruitment below and beyond the parental crown, but juveniles' survival and growth were lower below than beyond of the parental tree crowns. Small individuals (< 15 cm tall) showed the greatest mortality and the lowest growth, followed by medium (from 15 to 50 cm tall) and large ones (> 50 cm tall). Large juveniles were more widely dispersed from the conspecific parental tree than were medium and small ones. This suggests that distance dependent mortality of juveniles mediated by the parental tree is an important cause of spacing shifts associated with the growth of small individuals of E. glaziovii into large ones. Widely dispersed juveniles may escape the high mortality associated with pathogens, herbivores or seed predators concentrated around adult conspecifics. The negative influence of the parental tree on its juveniles may explain the sparse distribution of its adults in the forest.

Occurrence of the rare genus Microcrocis P. Richter (Chroococcales, Cyanobacteria) in a coastal lagoon from southern Brazil

The paper records the first occurrence of the genus Microcrocis P. Richter (Chroococcales, Cyanobacteria), represented by M. pulchella (Buell) Geitler, in Brazil. The species was found in two zones, one with freshwater and the other with brackish water, of a coastal lagoon of Rio Grande do Sul State (31°15’-31°30’ S and 50°54’-51°09’ W). Comparison between M. pulchella and its most closely related species is presented. Up to now this species had occurrence records limited to freshwater systems in temperate regions. Its presence in a subtropical coastal lagoon from southernmost Brazil, either in fresh or in brackish water, broadened the knowledge of the distribution area of M. pulchella.

Rare adverse events associated with oral poliovirus vaccine in Brazil

Oral poliovirus vaccine (OPV) developed by A. Sabin has been effectively used to control poliomyelitis in Brazil, and the last case with the isolation of a wild poliovirus strain occurred in March 1989. Although the vaccine controlled the circulation of wild strains and poliomyelitis cases associated with these strains were not detected during the last eight years, rare cases classified as vaccine-associated paralytic poliomyelitis (VAPP) have been detected. Molecular characterization studies of poliovirus strains isolated from VAPP cases and from healthy contacts have confirmed that the isolates are derived from the Sabin vaccine strains and also detected genomic modifications known or suspected to increase neurovirulence such as mutations and recombination. The molecular characterization of polioviruses isolated during the last eight years from paralysis cases classified as Guillain-Barré (GBS) syndrome and transverse myelitits (TM), and from facial paralysis (FP) cases also confirmed the vaccine origin of the strains and demonstrated mutations known to increase neurovirulence. Analysis of the epidemiologic data of these GBS, TM and FP cases demonstrated that in most of them the last OPV dose was given months or years before the onset of the disease and the isolation of the polioviruses. The temporal association between the isolation of these strains and the GBS, TM and FP suggested that the Sabin vaccine-derived poliovirus strains could also rarely trigger the diseases.

Precipitated immune complexes of IgM induce the generation of reactive oxygen species by rabbit polymorphonuclear leucocytes

We report that immune complexes of IgM (ICIgM) antibodies and ovalbumin in the form of a precipitate from the equivalence zone induce the generation of reactive oxygen species by rabbit blood polymorphonuclear leucocytes (PMN), as measured by the chemiluminescence (CL) production in the presence of luminol. The kinetics of CL generation induced by ICIgM is quite different from that induced by precipitated immune complexes of IgG (ICIgG): the maximum rate of CL production for ICIgM occurs around 14 min, whereas for ICIgG it occurs about 5 min after incubation with the cells. Also the triggering of the process requires a higher concentration of ICIgM than of ICIgG. Evidence is presented that these effects are not mediated by interaction of the antigen (ovalbumin) with the cell, since immune precipitates of ovalbumin and the F(ab')2 fragment had no effect. Our observations that precipitated ICIgM can also be an effective stimulus for CL generation and thus for O2- production reveal a new functional capability of PMN. These results may have implications for the understanding of the participation of ICIgM (as well as of ICIgG) in inflammatory reactions mediated by PMN in immune complex diseases, and in the mechanisms of defense against microbes and other non-self agents.

The role of complement in the modulation by fluid-phase IgG of the production of reactive oxygen species by polymorphonuclear leukocytes stimulated with IgG immune complexes

The production of reactive oxygen species (ROS) by polymorphonuclear leukocytes (PMN) can be induced by immune complexes and is an important component of phagocytosis in the killing of microorganisms, but can also be involved in inflammatory reactions when immune complexes are deposited in tissues. We have observed that fluid-phase IgG can inhibit the generation of ROS by rabbit PMN stimulated with precipitated immune complexes of IgG (ICIgG) in a dose-dependent manner, acting as a modulatory factor in the range of physiological IgG concentrations. This inhibitory effect is compatible with the known affinity (Kd) of monomeric IgG for the receptors involved (FcRII and FcRIII). The presence of complement components in the immune complexes results in a higher stimulation of ROS production. In this case, however, there is no inhibition by fluid-phase IgG. The effect of complement is strongly dependent on the presence of divalent cations (Ca2+ or Mg2+) in the medium, whereas the stimulation of ICIgG (without complement) does not depend on these cations. We have obtained some evidence indicating that iC3b should be the component involved in the effect of complement through interaction with the CR3 receptor. The absence of the inhibitory effect of fluid-phase IgG in ROS production when complement is present in the immune complex shows that complement may be important in vivo not only in the production of chemotactic factors for PMN, but also in the next phase of the process, i.e., the generation of ROS.

Evaluation by blue native polyacrylamide electrophoresis colorimetric staining of the effects of physical exercise on the activities of mitochondrial complexes in rat muscle

Blue native polyacrylamide electrophoresis (BN-PAGE) is a technique developed for the analysis of membrane complexes. Combined with histochemical staining, it permits the analysis and quantification of the activities of mitochondrial oxidative phosphorylation enzymes using whole muscle homogenates, without the need to isolate muscle mitochondria. Mitochondrial complex activities were measured by emerging gels in a solution containing all specific substrates for NADH dehydrogenase and cytochrome c oxidase enzymes (complexes I and IV, respectively) and the colored bands obtained were measured by optique densitometry. The objective of the present study was the application of BN-PAGE colorimetric staining for enzymatic characterization of mitochondrial complexes I and IV in rat muscles with different morphological and biochemical properties. We also investigated these activities at different times after acute exercise of rat soleus muscle. Although having fewer mitochondria than oxidative muscles, white gastrocnemius muscle presented a significantly higher activity (26.7 ± 9.5) in terms of complex I/V ratio compared to the red gastrocnemius (3.8 ± 0.65, P < 0.05) and soleus (9.8 ± 0.9, P < 0.001) muscles. Furthermore, the complex IV/V ratio of white gastrocnemius muscle was always significantly higher when compared to the other muscles. Ninety-five minutes of exhaustive physical exercise induced a decrease in complex I/V and complex IV/V ratios after all resting times (0, 3 and 6 h) compared to control (P < 0.05), probably reflecting the oxidative damage due to increasing free radical production in mitochondria. These results demonstrate the possible and useful application of BN-PAGE-histochemical staining to physical exercise studies.

An EDTA-β-cyclodextrin adsorbent for the adsorption of rare earth elements and its application in preconcentration of ultratrace rare earth elements from seawater

The objectives of this work were synthesizing an EDTA-β-CD adsorbent and investigating its adsorption potential and applications in preconcentration of REEs from aqueous phase. The adsorption capacity of EDTA-β-CD was investigated. The adsorption studies were performed by batch techniques both in one- and multi-component systems. The effects of pH, contact time and initial concentration were evaluated. The analytical detection methods and characterization methods were presented. EDTA-β-CD adsorbent was synthesized successfully with high EDTA coverage. The maximum REEs uptake was 0.310 mmol g-1 for La(III), 0.337 mmol g-1 for Ce(III) and 0.353 mmol g-1 for Eu(III), respectively. The kinetics of REEs onto EDTA-β-CD fitted well to pseudo-second-order model and the adsorption rate was affected by intra-particle diffusion. The experimental data of one component studies fitted to Langmuir isotherm model indicating the homogeneous surface of the adsorbent. The extended Sips model was applicable for the isotherm studies in three-component system. The electrostatic interaction, chelation and complexation were all involved in the adsorption mechanism. The preconcentration of RE ions and regeneration of EDTA-β-CD were successful. Overall, EDTA-β-CD is an effective adsorbent in adsorption and preconcentration of REEs.

New nitric oxide donors based on ruthenium complexes

Nitric oxide (NO) donors produce NO-related activity when applied to biological systems. Among its diverse functions, NO has been implicated in vascular smooth muscle relaxation. Despite the great importance of NO in biological systems, its pharmacological and physiological studies have been limited due to its high reactivity and short half-life. In this review we will focus on our recent investigations of nitrosyl ruthenium complexes as NO-delivery agents and their effects on vascular smooth muscle cell relaxation. The high affinity of ruthenium for NO is a marked feature of its chemistry. The main signaling pathway responsible for the vascular relaxation induced by NO involves the activation of soluble guanylyl-cyclase, with subsequent accumulation of cGMP and activation of cGMP-dependent protein kinase. This in turn can activate several proteins such as K+ channels as well as induce vasodilatation by a decrease in cytosolic Ca2+. Oxidative stress and associated oxidative damage are mediators of vascular damage in several cardiovascular diseases, including hypertension. The increased production of the superoxide anion (O2-) by the vascular wall has been observed in different animal models of hypertension. Vascular relaxation to the endogenous NO-related response or to NO released from NO deliverers is impaired in vessels from renal hypertensive (2K-1C) rats. A growing amount of evidence supports the possibility that increased NO inactivation by excess O2- may account for the decreased NO bioavailability and vascular dysfunction in hypertension.

Lipoprotein glomerulopathy: a case report of a rare disease in a brazilian child

Lipoprotein glomerulopathy (LPG) is a rare autosomal recessive glomerulopathy associated with the deposition of lipoprotein thrombi in the capillary lumina due to apoE gene mutations. Abnormal plasma lipoprotein profile and marked increase in serum apoliprotein E (apoE) are characteristic clinical data. The compromised patients can present nephrotic syndrome, hematuria, and progressive renal failure. Herein, the authors present the first described case of LPG in a Brazilian male patient, 11 years, who presented with a steroid-resistant nephrotic syndrome. Renal function was normal. Kidney biopsy showed markedly enlarged glomerulus, with dilated capillary loops and weak eosinophilic lipoprotein thrombi in the capillary lumina. Interstitium, tubules, arteries, and veins showed normal histologic aspect. Genotypic study for the apoE gene showed the presence of the alleles E3 and E4. The diagnosis of LPG was then performed. The patient received lipid-lowering treatment. After 2 years of follow-up, renal function is gradually decreasing, with persisting heavy proteinuria, despite a marked decrease in serum cholesterol and triglycerides levels.