Hemolytic disorders are characterized by hemolysis and are prone to thrombosis. Previously, it has been shown that the RNA released from damaged blood cells activates clotting. However, the nature of RNA released from hemolysis is still elusive. We found that after hemolysis, the red blood cells from both zebrafish and humans release 5.8S rRNA. This RNA activated coagulation in zebrafish and human plasmas. Using both natural and synthetic 5.8S rRNA and its synthetic truncated fragments, we found that the 3'-end 26 nucleotide-long RNA (3'-26 RNA) and its stem-loop secondary structure were necessary and sufficient for clotting activity. Corn trypsin inhibitor (CTI), a coagulation factor XII (FXII) inhibitor blocked 3'-26 RNA-mediated coagulation activation of both zebrafish and human plasma. CTI also inhibited zebrafish coagulation in vivo. 5.8S rRNA monoclonal antibody inhibited both 5.8S rRNA- and 3'-26 RNA-mediated zebrafish coagulation activity. Both 5.8S rRNA and 3'-26 RNA activates normal human plasma but did not activate FXII-deficient human plasma. Taken together, these results suggested that the activation of zebrafish plasma is via FXII-like protein. Since zebrafish has no FXII and hepatocyte growth factor activator (Hgfac) has sequence similarities to FXII, we knocked down the hgfac in adult zebrafish. We found that plasma from this knockdown fish does not respond to 3'-26 RNA. In conclusion, we identified 5.8S rRNA released in hemolysis activates clotting in human and zebrafish plasma. Only 3'-end 26 nucleotides of the 5.8S rRNA is needed for the clotting activity. Furthermore, we showed that fish Hgfac plays a role in 5.8S rRNA-mediated activation of coagulation.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / The Pampa is the newest Brazilian biomes. It is one of the lest studied and the one of the most economically exploited. Among these forms of exploitation, there is the cultivation of great dominated grassy fields with exotic forest species. Researches involving animal and plant species have revealed great biodiversity, however, they are still rare for microorganisms. The aim of the study was to collect, to isolate and to analyze morphologically and molecularly three wood-decaying fungi growing on Pinus sp. in Pampa biome: Pseudomerulius curtisii (Berk.) Redhead & Ginns, Gelatoporia subvermispora (Pilat) Niemelä and Sarcoporia polyspora P. Karsten. It was conducted the morphological characterization and the sequencing of ITS1-5.8S-ITS2 of species allowing their identification and phylogenetic comparisons with the literature. P. curtisii calls attention to the rarity and beauty of the basidiome and morphological similarities with specimens of the Northern Hemisphere and Oceania, but with a genetic distance, and may show the geographic isolation and the new taxon to be described. G. subvermispora, widely studied by the high biotechnology and industrial potential. It has high similarity with morphogenetic U.S. specimens, but with genetic divergence of Eurasian specimens. S. polyspora shows high morphological similarity with the specimens of the northern hemisphere, but there is a lack of sequences available for phylogenetic comparison. These species were registered for the first time in South America. P. curtisii, G. subvermispora and S. polyspora, fungi Basidiomycota show high potential for nutrient cycling and soil formation by the capacity or lignin and cellulose degradation. / O Pampa é o mais novo dos biomas brasileiros, sendo o menos estudado e um dos mais explorados comercialmente. Dentre estas formas de exploração, destaca-se o plantio de grandes áreas com espécies florestais exóticas. Os estudos envolvendo espécies animais e vegetais têm revelado grande biodiversidade, mas ainda são escassos para microrganismos. O objetivo deste trabalho foi coletar, isolar e analisar morfológica e molecularmente três fungos decompositores de madeira de Pinus sp. no bioma Pampa: Pseudomerulius curtisii (Berk.) Redhead & Ginns, Gelatoporia subvermispora (Pilát) Niemelä e Sarcoporia polyspora P. Karsten. Efetuou-se a caracterização morfológica e o sequenciamento da região ITS1-5.8S-ITS2 das espécies, permitindo suas identificações e comparações filogenéticas com a literatura. P. curtisii chama atenção pela raridade e beleza dos basidiomas e similaridade morfológica com os espécimes do Hemisfério Norte e Oceania, mas com uma distância genética, podendo mostrar o isolamento geográfico e novo taxon a ser descrito. G. subvermispora, amplamente estudado pelo elevado potencial biotecnológico e industrial, apresenta alta similaridade morfogenética com espécimes dos EUA, porém com divergência genética dos espécimes da Eurásia. S. polyspora apresenta alta similaridade morfológica com os espécimes do hemisfério Norte, mas há falta de sequências disponíveis para comparação filogenética. Essas espécies foram registradas pela primeira vez na América do Sul. P. curtisii, G. subvermispora and S. polyspora (Basidiomycota) apresentam grande potencial para estudos de ciclagem de nutrientes e formação do solo pela capacidade de degradação de lignina e/ou celulose.

The molecular characterization of twenty six Theileria spp. isolates and one C. felis isolate were done on the small subunit ribosomal RNA (SSU rRNA) gene, the 5.8S gene, and the two internal transcribed spacer regions using gDNA. The SSU rRNA gene is increasingly accepted as a widely used marker for characterization, taxonomic classification, and phylogenetic analysis and this gene has been sequenced from a variety of different organisms, resulting in a large database for sequence comparisons (Chae et al. 1998; Chae et al., 1999 a,b,c; Stockham et al., 2000; Cossio-Bayugar et al., 2002; Gubbels et al., 2000). The genomic region consists of the internal transcribed spacer 1 (ITS 1), the 5.8S gene, and internal transcribed spacer 2 (ITS 2) (ITS 1-5.8S-ITS 2 gene region) and separates the SSU rRNA gene from the large subunit ribosomal RNA gene. The 5.8S rRNA gene is highly conserved in size and nucleotide sequence, is relatively constant in molecular weight, and has an average chain length of approximately 160 nucleotides and has proven useful in dividing subgenera of Gyrodactylus ((Nazar, 1984; Zietara et al., 2002). Pairwise comparisons were done between the clones of an individual isolate and among the clones of the different isolates. Phylogenetic trees were made from the resulting sequences. This study shows that different SSU rRNA genes may be associated with ITS 1-5.8S-ITS 2 gene regions of distinct sequence in the same isolate. This study also demonstrates that considerable ITS 1-5.8S-ITS 2 gene region sequence variation may exist within a species. This may be useful for subspeciation designation, or may simply reflect considerable variation within the population. This study shows that the ITS 1-5.8S-ITS 2 gene region may be a useful molecular marker for the taxonomy of Theileria spp.

This study applies molecular and morphological approaches addressing the identification of morphologically similar larval stages (cercariae) of Plagiorchis spp. (Digenea: Plagiorchiidae) parasitising lymnaeid snail populations in the freshwater ecosystems of central Europe. Five morphologically homogeneous and genetically distinct lineages of Plagiorchis spp. were identified via matching molecular data for the mitochondrial cox1 gene with detailed morphometric data. Phylogenetic and comparative sequence analyses using partial 28S rDNA and ITS1-5.8S-ITS2 sequences allowed molecular identification of three species (P. elegans, P. maculosus and P. koreanus) via matching sequences from larval and adult digenean stages. A key for the identification of the cercariae of Plagiorchis spp. parasitising lymnaeid populations in central Europe is provided.

Det har forskats mycket på olika signalvägar i det mänskliga genomet, trotts detta finns det många frågetecken som kvarstår. Denna uppsats undersöker några av dem. Syfte: Undersöka förändringar i genuttryck och mRNA-nivåer för hypertrofi- (MRF4) translations- (5.8S & 18S) och proteolysreglerande gener (MuRF1 & GDF-8) efter en 10 veckor lång styrketräningsperiod hos kvinnor och män. Frågeställningar: (1) Finns det en förändring i total mängd RNA före och efter en 10 veckors styrketräningsintervention. (2) Finns det en förändring i uttryck av MRF4, 5.8S, 18S, MuRF1 samt GDF-8 efter en 10 veckors styrketräningsintervention. (3) Finns det en könsskillnad i förändringen av total mängd RNA samt aktivering av MRF4, 5.8S, 18S, MuRF1 och GDF-8 efter en 10 veckors styrketräningsintervention. Metod: Urvalet för analysen bestod av 16 otränade försökspersoner varav 8 var män och 8 var kvinnor. Försökspersonerna utförde unilateral styrketräning av nedre extremiteten under 10 veckor, under 2 av dessa veckor utfördes ocklusionsträning.  Träningsperiodiseringen var vågformig (70-90% av 1RM, 5-12 rep, 3 ggr/vecka). Muskelbiopsier togs i det arbetande benet före träningsperiodens start samt 3-7 dagar efter träningsperiodens avslut. Genuttryck analyserades med qPCR. Resultat: Det fanns ingen signifikant skillnad i förändring mellan män och kvinnors totala RNA eller genuttryck. Total RNA ökade signifikant (p<0,01) med 19,2 %. Kvinnorna hade en signifikant ökning (P<0,05) av RNA på 27,6 % medan männen hade en signifikant ökning (p<0,05) på 14 %. MRF4 hade en signifikant (P>0,05) procentuell ökning i genuttryck med 55,7 % och kvinnor för sig hade en signifikant (P>0,05) ökning på 64 %. GDF-8 ökade signifikant (P>0,05) med 55,5 % medan GAPDH ökade signifikant (P>0,05) för båda könen tillsammans med 70,6 % och för män med 87,8 %. MuRF1 och 5.8S hade inga signifikanta förändringar i genuttryck. Slutsats: Det verkar som att både män och kvinnor får en liknande procentuell förändring av total RNA och mRNA genuttryck 3-7 dagar efter en 10 veckors hypertrofistyrd styrketräningsperiod. För att mäta genuttryck av translationsgenen MRF4 verkar 3-7 dagar efter en 10 veckors styrketräningsperiod vara en tidpunkt då det fortfarande pågår hypertrofi av skelettmuskulaturen.  Av de proteolysreglerande generna GDF-8 och MuRF1 sågs en uppreglering av GDF-8 vilket skulle kunna vara ett tecken på att hypertrofin börjar hämmas. Ett oväntat fynd var att GAPDH visade sig vara olämplig som kontrollgen vid en styrketräningsintervention på 10 veckor och att 18S var väldigt stabil. Detta kan betyda att GAPDH inte skall användas vid längre styrketräningsinterventioner. / There have been much research on signaling pathways in the human genome, but there still remain many questions. This paper examines some of them. Aim: Investigate changes in gene expression and mRNA levels of hypertrophy (MRF4), translation (5.8S & 18S) and proteolysis regulating genes (GDF-8) after a 10-week strength training period in men and women. Research questions: (1) Is there a change in the total amount of RNA before and after a 10-week strength training intervention. (2) Is there a change in the expression of MRF4, 5.8S, 18S, Murf1 and GDF-8 after 10 weeks of strength training. (3) Is there a gender difference in the change of total RNA and the expression of MRF4, 5.8S, Murf1 and GDF-8 after a 10-week long strength training intervention. Method: The sample for analysis consisted of 16 untrained subjects, of whom 8 were men and 8 were women. The subjects performed unilateral resistance training of lower extremities for 10 weeks, during two of these weeks blood flow restriction training were performed. The training was undulating (70-90% of 1RM, 5-12 cord, 3 times / week). Muscle biopsies were taken from the working leg before the start and 3-7 days after the training period. Gene expression was analyzed by qPCR. Results: There was no significant gender difference in total RNA or gene expression. Total RNA was significantly increased (p <0.01) with 19.2 %. The women had a significant increase (P <0.05) of RNA at 27.6 %, while the men had a significant increase (p <0.05) at 14 %. MRF4 had a significant (P> 0.05) percentage increase in gene expression by 55.7 %, and women had a significant (P> 0.05) increase of 64 %. GDF-8 increased significantly (P> 0.05) with 55.5 %, while GAPDH increased significantly (P> 0.05) for both sexes with 70.6 % and for men with 87.8 %. Murf1 and 5.8S had no significant changes in gene expression. Conclusions: It seems that both men and women experience a similar percentage difference of total RNA and mRNA gene expression 3-7 days after a 10 weeks long strength training period. To measure the gene expression of MRF4 3-7 days after a 10-week weight-training period seems to be a time when there still is a anabolic responses in the skeletal muscle. Of the proteolysis regulating genes GDF-8 and Murf1 there was an upregulation of GDF-8, which could be a sign that the inhibition of hypertrophy started. An unexpected finding is that GAPDH was found to be unsuitable as a control gene at a strength training intervention at 10 weeks and rRNA 18S was very stable, which could mean that GAPDH should not be used as control gene in longer strength training studies.