Glycosaminoglycans affect the interaction of human plasma kallikrein with plasminogen, factor XII and inhibitors

Human plasma kallikrein, a serine proteinase, plays a key role in intrinsic blood clotting, in the kallikrein-kinin system, and in fibrinolysis. The proteolytic enzymes involved in these processes are usually controlled by specific inhibitors and may be influenced by several factors including glycosaminoglycans, as recently demonstrated by our group. The aim of the present study was to investigate the effect of glycosaminoglycans (30 to 250 µg/ml) on kallikrein activity on plasminogen and factor XII and on the inhibition of kallikrein by the plasma proteins C1-inhibitor and antithrombin. Almost all available glycosaminoglycans (heparin, heparan sulfate, bovine and tuna dermatan sulfate, chondroitin 4- and 6-sulfates) reduced (1.2 to 3.0 times) the catalytic efficiency of kallikrein (in a nanomolar range) on the hydrolysis of plasminogen (0.3 to 1.8 µM) and increased (1.9 to 7.7 times) the enzyme efficiency in factor XII (0.1 to 10 µM) activation. On the other hand, heparin, heparan sulfate, and bovine and tuna dermatan sulfate improved (1.2 to 3.4 times) kallikrein inhibition by antithrombin (1.4 µM), while chondroitin 4- and 6-sulfates reduced it (1.3 times). Heparin and heparan sulfate increased (1.4 times) the enzyme inhibition by the C1-inhibitor (150 nM).

An electronic pressure-meter nociception paw test for rats

The objective of the present investigation was to compare the sensitivity of an electronic nociceptive mechanical paw test with classical mechanical tests to quantify the intensity variation of inflammatory nociception. The electronic pressure-meter test consists of inducing the hindpaw flexion reflex by poking the plantar region with a polypropylene pipette tip adapted to a hand-held force transducer. This method was compared with the classical von Frey filaments test and with the rat paw constant pressure test, a modification of the Randall and Selitto test developed by our group. When comparing the three methods, the electronic pressure-meter and the rat paw constant pressure test, but not the von Frey filaments test, detected time vs treatment interactions in prostaglandin E2 (PGE2)-induced hypernociception. Both methods also detected the PGE2-induced hypernociception in dose- (50-400 ng/paw) and time- (1-4 h) dependent manners, and time vs treatment interactions induced by carrageenin (25-400 µg/paw). Furthermore, the electronic pressure-meter test was more sensitive at early times, whereas the constant pressure test was more sensitive at later times. Moreover, the electronic pressure-meter test detected the dose-dependent antinociceptive effect of local indomethacin (30-300 µg/paw) and dipyrone (80-320 µg/paw) on carrageenin- (200 µg/paw) and PGE2- (100 ng/paw) induced hypernociception, respectively, and also detected the ineffectiveness of indomethacin (300 µg) on the effect of PGE2. Our results show that the electronic pressure-meter provides a sensitive, objective and quantitative mechanical nociceptive test that could be useful to characterize new nociceptive inflammatory mediators and also to evaluate new peripheral analgesic substances.

Air pollution and neonatal deaths in São Paulo, Brazil

Air pollution has been associated with health effects on different age groups. The present study was designed to assess the impact of daily changes in air pollutants (NO2, SO2, CO, O3, and particle matter (PM10)) on total number of daily neonatal deaths (those that occur between the first and the 28th days of life) in São Paulo, from January 1998 to December 2000, since adverse outcomes such as neonatal deaths associated with air pollution in Brazil have not been evaluated before. Generalized additive Poisson regression models were used and nonparametric smooth functions (loess) were adopted to control long-term trend, temperature, humidity, and short-term trends. A linear term was used for holidays. The association between air pollutants and neonatal deaths showed a short time lag. Interquartile range increases in PM10 (23.3 µg/m³) and SO2 (9.2 µg/m³) were associated with increases of 4% (95% CI, 2-6) and 6% (95% CI, 4-8), respectively. Instead of adopting a two-pollutant model we created an index to represent PM10 and SO2 effects. For an interquartile range increase in the index an increase of 6.3% (95% CI, 6.1-6.5) in neonatal deaths was observed. These results agree with previous studies performed by our group showing the deleterious effects of air pollutants during the perinatal period. The method reported here represents an alternative approach to analyze the relationship between highly correlated pollutants and public health problems, reinforcing the idea of the synergic effects of air pollutants in public health.

Analysis of five streptokinase formulations using the euglobulin lysis test and the plasminogen activation assay

Streptokinase, a 47-kDa protein isolated and secreted by most group A, C and G ß-hemolytic streptococci, interacts with and activates human protein plasminogen to form an active complex capable of converting other plasminogen molecules to plasmin. Our objective was to compare five streptokinase formulations commercially available in Brazil in terms of their activity in the in vitro tests of euglobulin clot formation and of the hydrolysis of the plasmin-specific substrate S-2251™. Euglobulin lysis time was determined using a 96-well microtiter plate. Initially, human thrombin (10 IU/ml) and streptokinase were placed in individual wells, clot formation was initiated by the addition of plasma euglobulin, and turbidity was measured at 340 nm every 30 s. In the second assay, plasminogen activation was measured using the plasmin-specific substrate S-2251™. Streptase™ was used as the reference formulation because it presented the strongest fibrinolytic activity in the euglobulin lysis test. The Unitinase™ and Solustrep™ formulations were the weakest, showing about 50% activity compared to the reference formulation. All streptokinases tested activated plasminogen but significant differences were observed. In terms of total S-2251™ activity per vial, Streptase™ (75.7 ± 5.0 units) and Streptonase™ (94.7 ± 4.6 units) had the highest activity, while Unitinase™ (31.0 ± 2.4 units) and Strek™ (32.9 ± 3.3 units) had the weakest activity. Solustrep™ (53.3 ± 2.7 units) presented intermediate activity. The variations among the different formulations for both euglobulin lysis test and chromogenic substrate hydrolysis correlated with the SDS-PAGE densitometric results for the amount of 47-kDa protein. These data show that the commercially available clinical streptokinase formulations vary significantly in their in vitro activity. Whether these differences have clinical implications needs to be investigated.

The effect of an aerobic training program on the electrical remodeling of the heart: high-frequency components of the signal-averaged electrocardiogram are predictors of the maximal aerobic power

Increased heart rate variability (HRV) and high-frequency content of the terminal region of the ventricular activation of signal-averaged ECG (SAECG) have been reported in athletes. The present study investigates HRV and SAECG parameters as predictors of maximal aerobic power (VO2max) in athletes. HRV, SAECG and VO2max were determined in 18 high-performance long-distance (25 ± 6 years; 17 males) runners 24 h after a training session. Clinical visits, ECG and VO2max determination were scheduled for all athletes during thew training period. A group of 18 untrained healthy volunteers matched for age, gender, and body surface area was included as controls. SAECG was acquired in the resting supine position for 15 min and processed to extract average RR interval (Mean-RR) and root mean squared standard deviation (RMSSD) of the difference of two consecutive normal RR intervals. SAECG variables analyzed in the vector magnitude with 40-250 Hz band-pass bi-directional filtering were: total and 40-µV terminal (LAS40) duration of ventricular activation, RMS voltage of total (RMST) and of the 40-ms terminal region of ventricular activation. Linear and multivariate stepwise logistic regressions oriented by inter-group comparisons were adjusted in significant variables in order to predict VO2max, with a P < 0.05 considered to be significant. VO2max correlated significantly (P < 0.05) with RMST (r = 0.77), Mean-RR (r = 0.62), RMSSD (r = 0.47), and LAS40 (r = -0.39). RMST was the independent predictor of VO2max. In athletes, HRV and high-frequency components of the SAECG correlate with VO2max and the high-frequency content of SAECG is an independent predictor of VO2max.

Plasma amino acids in pregnancy, placental intervillous space and preterm newborn infants

Plasma amino acid levels have never been studied in the placental intervillous space of preterm gestations. Our objective was to determine the possible relationship between plasma amino acids of maternal venous blood (M), of the placental intervillous space (PIVS) and of the umbilical vein (UV) of preterm newborn infants. Plasma amino acid levels were analyzed by ion-exchange chromatography in M from 14 parturients and in the PIVS and UV of their preterm newborn infants. Mean gestational age was 34 ± 2 weeks, weight = 1827 ± 510 g, and all newborns were considered adequate for gestational age. The mean Apgar score was 8 and 9 at the first and fifth minutes. Plasma amino acid values were significantly lower in M than in PIVS (166%), except for aminobutyric acid. On average, plasma amino acid levels were significantly higher in UV than in M (107%) and were closer to PIVS than to M values, except for cystine and aminobutyric acid (P < 0.05). Comparison of the mean plasma amino acid concentrations in the UV of preterm to those of term newborn infants previously studied by our group showed no significant difference, except for proline (P < 0.05), preterm > term. These data suggest that the mechanisms of active amino acid transport are centralized in the syncytiotrophoblast, with their passage to the fetus being an active bidirectional process with asymmetric efflux. PIVS could be a reserve amino acid space for the protection of the fetal compartment from inadequate maternal amino acid variations.

Magnetization studies of polystyrene/multiwall carbon nanotube composite films

Lappeenranta University of Technology School of Technology Technical Physics Evgenii Zhukov MAGNETIZATION STUDIES OF POLYSTYRENE/MULTIWALL CARBON NANOTUBE COMPOSITE FILMS Master’s thesis 2015 55 pages, 41 pictures, 9 Tables. Examiners: Professor Erkki Lähderanta D.Sc. Ivan Zakharchuk Keywords: polystyrene, multi-walled carbon nanotubes, MWCNT, composite, magnetization, SQUID. In this thesis magnetic properties of polystyrene/multiwall carbon nanotube (MWCNT) composites are investigated with Quantum Design SQUID magnetometer (MPMS XL). The surface of the composite films is studied via BRUKER Multimode 8 Atomic Force Microscope, as well. The polystyrene/MWCNT composites have been prepared by the group of professor Okotrub (Physics Chemistry of Nanomaterials laboratory, Nikolaev Institute of Inorganic Chemistry, Russia). The composite films have been prepared by solution processing and stretching method. The approximate length and inner diameter of the MWCNTs used in fabrication are 260 μm and 10 nm, respectively. The content of MWCNTs is 1 and 2.5 contents percent (wt%) for studied samples. The stretching of the samples is 30% for samples with 1 and 2.5 wt% content, and one sample with 1 wt% loading of MWCNTs is 100% stretched. MWCNTs aligned perpendicular to a silicon substrate are used as a reference sample. The magnetization field dependencies of the samples exhibit hysteresis behavior. The values of saturation magnetization of composite films are much less compared to that of the reference sample. The saturation magnetization coercitivity field value drops with decrease of MWCNT content. At high magnetic fields strong presence of diamagnetism is observed. Measurements in magnetic field parallel and perpendicular to the composite plate display anisotropy with respect to the direction of stretching. Temperature dependences of magnetization for all samples display difference between zero-field cooled and field-cooled curves of magnetization. This divergence confirms the presence of magnetic interactions in the material. The atomic force microscopy study of the composites’ surfaces revealed that they are relatively smooth and the nanotubes are aligned with the axis of stretching to some extent. 

Prokaryotic expression of a novel mouse pro-apoptosis protein PNAS-4 and application of its polyclonal antibodies

Mouse PNAS-4 (mPNAS-4) has 96% identity with human PNAS-4 (hPNAS-4) in primary sequence and has been reported to be involved in the apoptotic response to DNA damage. However, there have been no studies reported of the biological functions of mPNAS-4. In studies conducted by our group (unpublished data), it was interesting to note that overexpression of mPNAS-4 promoted apoptotic death in Lewis lung carcinoma cells (LL2) and colon carcinoma cells (CT26) of mice both in vitro and in vivo. In our studies, mPNAS-4 was cloned into the pGEX-6P-1 vector with GST tag at N-terminal in Escherichia coli strain BL21(DE3). The soluble and insoluble expression of recombinant protein mPNAS-4 (rmPNAS-4) was temperature-dependent. The majority of rmPNAS-4 was insoluble at 37°C, while it was almost exclusively expressed in soluble form at 20°C. The soluble rmPNAS-4 was purified by one-step affinity purification, using a glutathione Sepharose 4B column. The rmPNAS-4 protein was further identified by electrospray ionization-mass spectrometry analysis. The search parameters of the parent and fragment mass error tolerance were set at 0.1 and 0.05 kDa, respectively, and the sequence coverage of search result was 28%. The purified rmPNAS-4 was further used as immunogen to raise polyclonal antibodies in New Zealand white rabbit, which were suitable to detect both the recombinant and the endogenous mPNAS-4 in mouse brain tissue and LL2 cells after immunoblotting and/or immunostaining. The purified rmPNAS-4 and our prepared anti-mPNAS-4 polyclonal antibodies may provide useful tools for future biological function studies for mPNAS.

Morphine infusions into the rostrolateral periaqueductal gray affect maternal behaviors

It is well established that morphine inhibits maternal behaviors. Previous studies by our group have shown activation of the rostrolateral periaqueductal gray (rlPAG) upon inhibition-intended subcutaneous injections of morphine. In this context, we demonstrated that a single naloxone infusion into the rlPAG, following this opioid-induced inhibition, reactivated maternal behaviors. Since these data were obtained by using peripheral morphine injections, the present study was designed to test whether morphine injected directly into the rlPAG would affect maternal behaviors. Our hypothesis that morphine acting through the rlPAG would disrupt maternal behaviors was confirmed with a local infusion of morphine. The mothers showed shorter latency for locomotor behavior to explore the home cage (P = 0.049). Inhibition was especially evident regarding retrieving (P = 0.002), nest building (P = 0.05) and full maternal behavior (P = 0.023). These results support the view that opioidergic transmission plays a behaviorally meaningful inhibitory role in the rostrolateral PAG.

Elementtikoon vaikutus väsymismenetelmien tarkkuuteen

Kandidaatintyössä tutkittiin kolmen hitsatun palstalevyliitoksen väsymiskestävyyttä äärellisten elementtien menetelmällä. Tutkimuksen pääpaino oli 2D-mallin antamien tulosten suppenevuudessa kolmella eri elementtikoolla ja kolmessa eri mittasuhdetapauksessa. Tutkimustuloksissa havaittiin noin 5 % muutos harvimman ja tiheimmän elementtiverkon tulosten välillä. Tutkimuksessa havaittiin myös että muutos harvimman ja tiheimmän elementtiverkon tuloksissa pieneni hiukan mitä suurempia kappaleet ja niiden hitsausliitokset olivat.

The nature experiences of wilderness recreation leaders : throwing a stone

Through this descriptive exploratory study, the ways that wilderness recreation leaders experience nature are illuminated, deconstructing the assumed environmental benefits of and practices used in outdoor recreation (Haluza-Delay, 2001). This study also offers a foundation for advancing an environmental ethic among wilderness recreation leaders, participants, and organizations. With the continued degradation of and threats to natural environments, and the rising popularity of outdoor recreation participation, the outdoor recreation professional can be a leader in promoting human reconnections to the Earth (Henderson, 1999). Leaders of outdoor recreation experiences play an important role in encouraging these revived relationships to natural settings and can contribute to the necessary environmental consciousness shift needed within Western society (Hanna, 1995; Jordan, 1996). The purpose of this research was to describe the lived-experience in nature of wilderness recreation leaders. Specifically, a phenomenological method of inquiry was used to describe the meaning of nature, the connections and relationships to nature, and the behaviours and emotions experienced in nature by a group of wilderness canoe trip leaders employed by a residential summer camp. In addition to the implications of this research, achieving this outcome provides a rich descriptive understanding of wilderness leaders' experiences—a basis from which to extend future research endeavours and programmatic practices that promote effective environmental outcomes of outdoor recreation participation. Each of the five study participants was employed in the summer of 2003 by an Ontario residential summer camp organization that sponsors extended wilderness river canoe trips for youth. Two in-depth and semi-structured interviews were performed with each participant, asking them to reflect on the canoe trip that they led for the summer camp organization during 2003. Phenomenological data was analyzed according to Colaizzi's (1978) thematic analysis process. Consistent with van Manen's (1997) emphasis on phenomenological writing, the final result presents the essence of the nature experiences of wilderness recreation leaders in the format of a narrative description. This narrative piece is the culmination of this research effort. Throughout the journey, however, various foundations within the outdoor recreation field, such as minimum impact principles, environmentally responsible behaviours, anthropocentric and ecocentric worldviews, and effective leadership are deconstructed and discussed.

Assessing evaluative measures for Internet-based courseware

Although there is a consensus in th~ literature on the many uses of the Internet in education, as well as the unique features of the Internet for presenting facts and information, there is no consensus on a standardized method for evaluating Internetbased courseware. Educators rarely have the opportunity to participate in the development of Internet-based courseware, yet they are encouraged to use the technology in their learning environments. This creates a need for summative evaluation methods for Internet-based health courseware. The purpose ofthis study was to assess evaluative measures for Internet-based courseware. Specifically, two entities were evaluated within the study: a) the outcome of the Internet-based courseware, and b) the Internet-based courseware itself. To this end, the Web site www.bodymatters.com was evaluated using two different approaches by two different cohorts. The first approach was a performance appraisal by a group of endusers. A positive, statistically significant change in the students performance was observed due to the intervention ofthe Web site. The second approach was a productoriented evaluation ofthe Web site with the use of a criterion-based checklist and an open-ended comments section. The findings indicate that a summative, criterion-based evaluation is best completed by a multidisciplinary team. The findi~gs also indicated that the two different cohorts reported different product-oriented appraisals of the Web site. The current research confirmed previous research that found that experts returning a poor evaluation of a Web site did not have a relationship to whether or not the end-users performance improved due to the intervention of the Web site.

Creativity and the working artist/teacher : the relationships

The purpose of this qualitative study was to understand the relationships between creativity and the working artist/teacher employed by an art college. The topic emerged from my job as an instructor at The Ontario College of Art which was used as the primary data resource and provided the highest caliber of professionals to chose from. Existent data were used to facilitate the study generated by the research of Cawelti, Rappaport, and Wood (1992). The data were generated by a group of 5 faculty members from The University of Northern Iowa, recognized for their expertise in the arts (a painter, a poet, a sculptor, a novelist, and a photographer). They were asked to respond to the following statement: "In as much detail as you like, list the things that you did, thought, or felt the last time you created an artistic product. II Cawelti, Rappaport, and Wood (1992) produced three models of the creative process, each building on the previous, with the resultant third,being in my opinion, an excellent illustration (text/visual) of the creative process. Model three (Appendix D) presented a "multi-dimensional view of the creative process: time, space, observatility, and consciousnessll (p. 90). Model three utilized a visual mapping device along the bottom of the page linked to text segments above. Both the visual and the text were interrelated so that they harmonized into a comprehensive "picture." The parti'cipants of this qualitative study were asked to consider model three from their professional perspective as artist/teachers. The interpretive sciences directed the methodology. The hermeneutic circle of continuous reflection from the whole to the part and back to the whole was an important aspect of the data analyses. Four members of the Foundation Department at The Ontario College of Art were the key participants. A series of conversational interviews was the primary source of data collection, this was augmented by observation, fie,ldnotes, and follow up telephone interviews. Transcripts of interviews were returned to participants for reflection and the telephone was used to discuss any additional -points raised. Analysis consisted of coding and organizing data according to emerging themes. These themes formed the basis for the narrative stories. The text of the narrative stories were given back to each participant for further comment. Revisions were made until both the researcher and the participants felt that the stories reflected reality. The resultant whole was critiqued from the researcher's perspective. The significance of this study was discussed as it pertains to the working artist/teacher and areas in need of further study are pointed out.

Fungal biotransformation of morphine alkaloids

The purpose of the study was to determine the ability of certain fungi to biotransform morphine alkaloids into medicinally relevant intermediates. Fungal strains screened for their ability to affect biotransformation of morphine alkaloids include Cunninghamella echinulata, Helicostylum pirijorme, Pycnoporus sanguinea, Pycnoporus cinnabarina, Curvularia lunata and Sporotrichum sulfurescens. The research demonstrated that Cunninghamella echinulata N-demethylated thebaine, hydrocodone, codeine, oripavine and oxycodone into corresponding nor-compounds in varying yields. The study further focused on the characterization of the enzyme responsible for the biotransformation of thebaine into northebaine by Cunninghamella echinulata. The study clearly showed that incubation of the fungal culture with thebaine over a period of 48 hours was required to activate the biotransformation process. The biotransformation studies with [14C] labeled thebaine showed that Ndemethylation by Cunningham ella echinulata does not involve O-demethylation followed by methyl group transfer as suggested in previous studies.

Ridley College papers, 1983, 1985-1986, 1989-1990

Ridley College was conceived in 1888 by a group of Anglican clergy and laymen eager to establish a school for boys in Ontario that emphasized strong academic and religious values. The school was originally known as Bishop Ridley College, in tribute to Nicholas Ridley, a 16th century English churchman who was martyred during the Protestant Reformation for refusing to renounce his Anglican faith. The first facility was the stately and spacious Springbank Sanatorium; shortly thereafter, construction was begun across the old Welland Canal on a lower school for boys age 5 to 13 on the present-day campus site. The name “Springbank” stems from the name of the hotel constructed in 1864 by Dr. Theophilus Mack on Yates Street. Fortuitously, the directors of what would become Ridley College were looking to found a new boys’ school. The sale of the building was completed in 1888 and Ridley began operations in September 1889. In October 1903, the Springbank building complex was consumed by fire forcing the school to move across the canal to its modern western campus. The Ridley campus grew dramatically during the 1920's, and new buildings and facilities were added in each of the following decades. The school became co-educational in 1973; just over a dozen girls enrolled in the inaugural year, while today almost half of Ridley's students are girls. Adapted from: http://www.ridleycollege.com/podium/default.aspx?t=125335 (March 22, 2011)