pH titration of native and unfolded ß-trypsin: evaluation of the D D G0 titration and the carboxyl pK values

The stabilizing free energy of ß-trypsin was determined by hydrogen ion titration. In the pH range from 3.0 to 7.0, the change in free energy difference for the stabilization of the native protein relative to the unfolded one (D D G0 titration) was 9.51 ± 0.06 kcal/mol. An isoelectric point of 10.0 was determined, allowing us to calculate the Tanford and Kirkwood electrostatic factor w. This factor presented a nonlinear behavior and indicated more than one type of titratable carboxyl groups in ß-trypsin. In fact, one class of carboxyl group with a pK = 3.91 ± 0.01 and another one with a pK = 4.63 ± 0.03 were also found by hydrogen ion titration of the protein in the folded state

Dynamic Analysis Model of Spherical Roller Bearings with Defects

Rolling element bearings are essential components of rotating machinery. The spherical roller bearing (SRB) is one variant seeing increasing use, because it is self-aligning and can support high loads. It is becoming increasingly important to understand how the SRB responds dynamically under a variety of conditions. This doctoral dissertation introduces a computationally efficient, three-degree-of-freedom, SRB model that was developed to predict the transient dynamic behaviors of a rotor-SRB system. In the model, bearing forces and deflections were calculated as a function of contact deformation and bearing geometry parameters according to nonlinear Hertzian contact theory. The results reveal how some of the more important parameters; such as diametral clearance, the number of rollers, and osculation number; influence ultimate bearing performance. Distributed defects, such as the waviness of the inner and outer ring, and localized defects, such as inner and outer ring defects, are taken into consideration in the proposed model. Simulation results were verified with results obtained by applying the formula for the spherical roller bearing radial deflection and the commercial bearing analysis software. Following model verification, a numerical simulation was carried out successfully for a full rotor-bearing system to demonstrate the application of this newly developed SRB model in a typical real world analysis. Accuracy of the model was verified by comparing measured to predicted behaviors for equivalent systems.

Evidence for the expression of native Mycobacterium tuberculosis phospholipase C: recognition by immune sera and detection of promoter activity

The genome of Mycobacterium tuberculosis H37Rv contains three contiguous genes (plc-a, plc-b and plc-c) which are similar to the Pseudomonas aeruginosa phospholipase C (PLC) genes. Expression of mycobacterial PLC-a and PLC-b in E. coli and M. smegmatis has been reported, whereas expression of the native proteins in M. tuberculosis H37Rv has not been demonstrated. The objective of the present study was to demonstrate that native PLC-a is expressed in M. tuberculosis H37Rv. Sera from mice immunized with recombinant PLC-a expressed in E. coli were used in immunoblots to evaluate PLC-a expression. The immune serum recognized a 49-kDa protein in immunoblots against M. tuberculosis extracts. No bands were visible in M. tuberculosis culture supernatants or extracts from M. avium, M. bovis and M. smegmatis. A 550-bp DNA fragment upstream of plc-a was cloned in the pJEM12 vector and the existence of a functional promoter was evaluated by detection of ß-galactosidase activity. ß-Galactosidase activity was detected in M. smegmatis transformed with recombinant pJEM12 grown in vitro and inside macrophages. The putative promoter was active both in vitro and in vivo, suggesting that expression is constitutive. In conclusion, expression of non-secreted native PLC-a was demonstrated in M. tuberculosis.

Protein defects in neuromuscular diseases

Muscular dystrophies are a heterogeneous group of genetically determined progressive disorders of the muscle with a primary or predominant involvement of the pelvic or shoulder girdle musculature. The clinical course is highly variable, ranging from severe congenital forms with rapid progression to milder forms with later onset and a slower course. In recent years, several proteins from the sarcolemmal muscle membrane (dystrophin, sarcoglycans, dysferlin, caveolin-3), from the extracellular matrix (alpha2-laminin, collagen VI), from the sarcomere (telethonin, myotilin, titin, nebulin), from the muscle cytosol (calpain 3, TRIM32), from the nucleus (emerin, lamin A/C, survival motor neuron protein), and from the glycosylation pathway (fukutin, fukutin-related protein) have been identified. Mutations in their respective genes are responsible for different forms of neuromuscular diseases. Protein analysis using Western blotting or immunohistochemistry with specific antibodies is of the utmost importance for the differential diagnosis and elucidation of the physiopathology of each genetic disorder involved. Recent molecular studies have shown clinical inter- and intra-familial variability in several genetic disorders highlighting the importance of other factors in determining phenotypic expression and the role of possible modifying genes and protein interactions. Developmental studies can help elucidate the mechanism of normal muscle formation and thus muscle regeneration. In the last fifteen years, our research has focused on muscle protein expression, localization and possible interactions in patients affected by different forms of muscular dystrophies. The main objective of this review is to summarize the most recent findings in the field and our own contribution.

Congenital upper limb defects in Finland 1993-2005

Background: Interest in limb defects has grown after the thalidomide tragedy in the 1960s. As a result, congenital malformation registries, monitoring changes in birthprevalence and defect patterns, have been established in several countries. However, there are only a few true population based studies on birth prevalence of upper limb defects. The burden of hospital care among these children, specifically in terms of the number of admissions and total time spent in hospital, is also unknown. Aims and Methods: This study is based on information gathered from the Finnish Register of Congenital malformations (FRM) and the Finnish Hospital Discharge Register (FHDR). A total of 417 children born between 1993 and 2005 with an upper limb defect were gathered from the FRM. The upper limb defects were classified using the International Federation of Societies for Surgery of the Hand -classification that enables comparison with previous and future studies. Birth and live birth prevalence, sex and side distribution, frequency of associated anomalies as well as the proportion of perinatal and infant deaths according to the different subtypes were calculated. The number of hospital admissions, days spent in hospital, number and type of surgical operations were collected from the FHDR. Special features of two subgroups, radial ray defects (RRD) and constriction band syndrome (CBS), were explored. Results: Upper limb defects were observed in 417 of 753 342 consecutive births and in 392 of 750 461 live births. Birth prevalence was 5.5 per 10 000 births and 5.2 per 10 000 live births. Multiple anomalies or a known syndrome was found in 250 cases (60%). Perinatal mortality was 139 per 1000 births and infant mortality 135 per 1000 live births (overall Finnish perinatal mortality <5 per 1000 births and infant mortality 3.7 per 1000 live births). Altogether, 138 infants had RRD and 120 (87%) of these had either a known syndrome or multiple major anomalies. The proportion of perinatal deaths in RRD group was 29% (40/138) and infant deaths 35% (43/123). Fifty-one children had CBS in upper limbs. Fifteen of these (29%) had other major anomalies associated with constriction rings. The number of hospital admissions per year of children with congenital upper limb defects was 11-fold and the time spent in hospital 13-fold as compared with the general paediatric population. Conclusions: Birth prevalence of congenital upper limb defects was 5.5 per 10 000 births and 5.2 per 10 000 live births. RRD was especially associated with other major anomalies and high mortality. Nearly one third of the children with CBS also had other major anomalies suggesting different aetiologies inside the group. The annual burden of hospital care of children with congenital upper limb defects was at least 11-fold as compared with the general paediatric population.

Screening of antibacterial extracts from plants native to the Brazilian Amazon Rain Forest and Atlantic Forest

More than 20% of the world's biodiversity is located in Brazilian forests and only a few plant extracts have been evaluated for potential antibacterial activity. In the present study, 705 organic and aqueous extracts of plants obtained from different Amazon Rain Forest and Atlantic Forest plants were screened for antibacterial activity at 100 µg/ml, using a microdilution broth assay against Staphylococcus aureus, Enterococcus faecalis, Pseudomonas aeruginosa and Escherichia coli. One extract, VO581, was active against S. aureus (minimum inhibitory concentration (MIC) = 140 µg/ml and minimal bactericidal concentration (MBC) = 160 µg/ml, organic extract obtained from stems) and two extracts were active against E. faecalis, SM053 (MIC = 80 µg/ml and MBC = 90 µg/ml, organic extract obtained from aerial parts), and MY841 (MIC = 30 µg/ml and MBC = 50 µg/ml, organic extract obtained from stems). The most active fractions are being fractionated to identify their active substances. Higher concentrations of other extracts are currently being evaluated against the same microorganisms.

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.

Emission, kinetic and magnetic phenomena in rare-earth and transition metal doped ZnSe single crystals

In this work emission, optical, electrical and magnetic properties of the d- and f- elements doped zinc selenide crystals were investigated within a wide temperature range. Doping was performed in various technological processes: during the growth by chemical vapor transport method; by thermal diffusion from the Bi or Zn melt. Concentration of the doping impurity in the crystals was controlled by amount of the dopant in the source material or by its concentration in the doping media. Special interest in the work was paid to the influence of the different concentrations of Cr and Yb impurities on ZnSe crystals’ properties, correlations between observed effects and similarities with the Ni, Mn and Gd dopants are analysed. Possibility of formation of the excitons bound to the doping d-ions was shown. In contrast to this, it was observed that f-elements do not bound excitons, but prevent formation of excitons bound to some uncontrolled impurities. A mechanism of Cr doping impurity interaction with background impurities and zinc selenide structural defects was proposed based on experimental data. An assumption about resonant energy transfer between double charged chromium ions and complexes based on crystals’ vacancy defects was made. A correlation between emission and magnetic properties of the d- ions doped samples was established. Based on this correlation a mechanism explaining the concentration quench of the emission was proposed. It was found that f-ions bind electrically active shallow and deep donor and acceptor states of background impurity to electrically neutral complexes. This may be observed as “purification” of ZnSe crystals by doping with the rare-earth elements, resulting i tendency of the properties of f-ion doped crystals to the properties of intrinsic crystals, but with smaller concentration of uncontrolled native and impurity defects. A possible interpretation of this effect was proposed. It was shown that selenium substituting impurities decrease efficiency of the Yb doping. Based on this experimental results an attempt to determine ytterbium ion surroundings in the crystal lattice was made. It was shown that co-doping of zinc selenide crystals with the d- and f- ions leads to the combination of the impurities influence on the material’s properties. On the basis of obtained data an interaction mechanism of the d- and f-elements co-dopants was proposed. Guided by the model of the ytterbium ion incorporation in the selenide sublattice of the ZnSe crystals, an assumption about stabilization of single charged chromium ions in the zinc sublattice crystal nodes, by means of formation of the local charge compensating clusters, was made.

Congenital ventricular septal defects and prenatal exposure to cyclooxygenase inhibitors

Ventricular septal defects (VSDs) are common congenital abnormalities which have been reported to be associated with maternal fever and various environmental factors. The aim of the present study was to evaluate the effect of prenatal exposure to cyclooxygenase (COX) inhibitors on heart defects. A retrospective statistical analysis was performed using data collected in our laboratory during various teratological studies carried out on albino CRL:(WI)WUBR Wistar strain rats from 1997 to 2004. The observations were compared with concurrent and historic control data, as well as findings from other developmental toxicological studies with selective and nonselective COX-2 inhibitors. Despite the lack of significant differences in the frequency of VSDs between drug-exposed and control groups, statistical analysis by the two-sided Mantel-Haenszel test and historical control data showed a higher incidence of heart defects in offspring exposed to nonselective COX inhibitors (30.06/10,000). Unlike other specific inhibitors, aspirin (46.26/10,000) and ibuprofen (106.95/10,000) significantly increased the incidence of the VSD when compared with various control groups (5.38-19.72/10,000). No significant differences in length or weight were detected between fetuses exposed to COX inhibitors and born with VSD and non-malformed offsprings. However, a statistically significant increase of fetal body length and decrease of body mass index were found in fetuses exposed to COX inhibitors when compared with untreated control. We conclude that prenatal exposure to COX inhibitors, especially aspirin and ibuprofen, increased the incidence of VSDs in rat offspring but was not related to fetal growth retardation.

Scientist-friendly policies for non-native English-speaking authors: timely and welcome

That English is the lingua franca of today's science is an indisputable fact. Publication in English in international journals is a pre-requisite for a research paper to gain visibility in academia. However, English proficiency appears to be taken for granted in the scientific community, though this language can be a hurdle for a number of authors, particularly from non-native English-speaking countries. The influence of English proficiency on the publication output of Brazilian authors has never been assessed. We report our preliminary data on the relationship between the English proficiency of 51,223 researchers registered in the CNPq database and their publication output in international journals. We have found that publication rates are higher for those authors with good command of English, particularly written English. Although our research is still underway and our results are preliminary, they suggest that the correlation between written English proficiency and research productivity should not be underestimated. We also present the comments of some Brazilian scientists with high publication records on the relevance of communication skills to the scientific enterprise.

Proteoliposomes as matrix vesicles' biomimetics to study the initiation of skeletal mineralization

During the process of endochondral bone formation, chondrocytes and osteoblasts mineralize their extracellular matrix by promoting the formation of hydroxyapatite (HA) seed crystals in the sheltered interior of membrane-limited matrix vesicles (MVs). Ion transporters control the availability of phosphate and calcium needed for HA deposition. The lipidic microenvironment in which MV-associated enzymes and transporters function plays a crucial physiological role and must be taken into account when attempting to elucidate their interplay during the initiation of biomineralization. In this short mini-review, we discuss the potential use of proteoliposome systems as chondrocyte- and osteoblast-derived MVs biomimetics, as a means of reconstituting a phospholipid microenvironment in a manner that recapitulates the native functional MV microenvironment. Such a system can be used to elucidate the interplay of MV enzymes during catalysis of biomineralization substrates and in modulating in vitro calcification. As such, the enzymatic defects associated with disease-causing mutations in MV enzymes could be studied in an artificial vesicular environment that better mimics their in vivo biological milieu. These artificial systems could also be used for the screening of small molecule compounds able to modulate the activity of MV enzymes for potential therapeutic uses. Such a nanovesicular system could also prove useful for the repair/treatment of craniofacial and other skeletal defects and to facilitate the mineralization of titanium-based tooth implants.

Nichesourcing for the benefit of linguistic research and native speakers

The National Library of Finland is implementing the Digitization Project of Kindred Languages in 2012–16. Within the project we will digitize materials in the Uralic languages as well as develop tools to support linguistic research and citizen science. Through this project, researchers will gain access to new corpora 329 and to which all users will have open access regardless of their place of residence. Our objective is to make sure that the new corpora are made available for the open and interactive use of both the academic community and the language societies as a whole. The project seeks to digitize and publish approximately 1200 monograph titles and more than 100 newspapers titles in various Uralic languages. The digitization will be completed by the early of 2015, when the Fenno-Ugrica collection would contain around 200 000 pages of editable text. The researchers cannot spend so much time with the material that they could retrieve a satisfactory amount of edited words, so the participation of a crowd in editing work is needed. Often the targets in crowdsourcing have been split into several microtasks that do not require any special skills from the anonymous people, a faceless crowd. This way of crowdsourcing may produce quantitative results, but from the research’s point of view, there is a danger that the needs of linguistic research are not necessarily met. Also, the number of pages is too high to deal with. The remarkable downside is the lack of shared goal or social affinity. There is no reward in traditional methods of crowdsourcing. Nichesourcing is a specific type of crowdsourcing where tasks are distributed amongst a small crowd of citizen scientists (communities). Although communities provide smaller pools to draw resources, their specific richness in skill is suited for the complex tasks with high-quality product expectations found in nichesourcing. Communities have purpose, identity and their regular interactions engenders social trust and reputation. These communities can correspond to research more precisely. Instead of repetitive and rather trivial tasks, we are trying to utilize the knowledge and skills of citizen scientists to provide qualitative results. Some selection must be made, since we are not aiming to correct all 200,000 pages which we have digitized, but give such assignments to citizen scientists that would precisely fill the gaps in linguistic research. A typical task would editing and collecting the words in such fields of vocabularies, where the researchers do require more information. For instance, there’s a lack of Hill Mari words in anatomy. We have digitized the books in medicine and we could try to track the words related to human organs by assigning the citizen scientists to edit and collect words with OCR editor. From the nichesourcing’s perspective, it is essential that the altruism plays a central role, when the language communities involve. Upon the nichesourcing, our goal is to reach a certain level of interplay, where the language communities would benefit on the results. For instance, the corrected words in Ingrian will be added onto the online dictionary, which is made freely available for the public and the society can benefit too. This objective of interplay can be understood as an aspiration to support the endangered languages and the maintenance of lingual diversity, but also as a servant of “two masters”, the research and the society.

Chemical characteristics of pequi fruits (Caryocar brasiliense Camb.) native of three municipalities in the State of Goiás - Brazil

In order to assure that the use of cerrado fruits occur in a sustainable way, studies to investigate their characteristics are extremely relevant. In this context, the present study aims to describe some chemical parameters of pequi fruits picked in three municipalities in southwestern Goiás State (Jataí, Rio Verde, and Serranópolis). In each city, two populations of pequi trees - pequizeiros, denominated areas, were selected. In each area, eight trees were selected for the fruit to be picked. The contents of phosphorus, potassium, calcium, magnesium, nitrogen, zinc, and ether extract were determined in the samples. The results demonstrate differences between the chemical characteristics studied for the fruits picked in different areas, which does not seem to vary in a significant way. Comparing the contents obtained in the present study with those required as human daily supply, further studies are recommended aiming at using the pequi fruit as a complementary alternative source of magnesium, manganese, and copper.

Mineral characterization of native fruits from the southern region of Brazil

Although the greatest variety of Brazilian flora is in the Amazon region, the Southern region of Brazil also has an estimated number of at least 5,000 species of vascular native plants. These species have been neglected as potential food sources, remaining unknown and under-utilized and limiting the potential variety in the diet of Brazilians and other peoples. Therefore the aim of this study was to characterize the mineral composition and content present in seven native fruit species of Southern Brazil using inductively coupled plasma optical emission spectrometry (ICP-OES). The essential element concentrations in the fruit samples were higher or similar to the values reported for traditional fruits. The araticum-do-mato fruit samples had high concentrations of the elements Ca, K, and Cu, and trace elements such as Pb and Sr. Mandacaru-de-três-quinas had predominance of Ba, Bi, and Ga, and the essential elements Mg and Mn. Uvaia and guabiroba had the highest levels of Al and Cr, but uvaia had high levels of Fe and Zn. The pindo palm had high amounts of Cd and Ni, and the yellow guava had high concentrations of Na, while red guava had high levels of Co.

Reconstruction of cranial bone defects with fiber-reinforced composite–bioactive glass implants

A cranial bone defect may result after an operative treatment of trauma, infection, vascular insult, or tumor. New biomaterials for cranial bone defect reconstructions are needed for example to mimic the biomechanical properties and structure of cranial bone. A novel glass fiber-reinforced composite implant with bioactive glass particulates (FRC–BG, fiber-reinforced composite–bioactive glass) has osteointegrative potential in a preclinical setting. The aim of the first and second study was to investigate the functionality of a FRC–BG implant in the reconstruction of cranial bone defects. During the years 2007–2014, a prospective clinical trial was conducted in two tertiary level academic institutions (Turku University Hospital and Oulu University Hospital) to evaluate the treatment outcome in 35 patients that underwent a FRC–BG cranioplasty. The treatment outcome was good both in adult and pediatric patients. A number of conventional complications related to cranioplasty were observed. In the third study, a retrospective outcome evaluation of 100 cranioplasty procedures performed in Turku University Hospital between years 2002–2012 was conducted. The experimental fourth study was conducted to test the load-bearing capacity and fracture behavior of FRC–BG implants under static loading. The interconnective bars in the implant structure markedly increased the load-bearing capacity of the implant. A loading test did not demonstrate any protrusions of glass fibers or fiber cut. The fracture type was buckling and delamination. In this study, a postoperative complication requiring a reoperation or removal of the cranioplasty material was observed in one out of five cranioplasty patients. The treatment outcomes of cranioplasty performed with different synthetic materials did not show significant difference when compared with autograft. The FRC–BG implant was demonstrated to be safe and biocompatible biomaterial for large cranial bone defect reconstructions in adult and pediatric patients.