Morphological modularity in the vertebral column of Felidae (Mammalia, Carnivora)

BACKGROUND: Previous studies have demonstrated that the clear morphological differences among vertebrae across the presacral column are accompanied by heterogeneous functional signals in vertebral shape. Further, several lines of evidence suggest that the mammalian axial skeleton is a highly modular structure. These include its composition of serial units, a trade-off between high shape variance and strong conservation of vertebral count, and direct association of regions with anterior expression sites of Hox genes. Here we investigate the modular organisation of the presacral vertebral column of modern cats (Felidae, Carnivora, Mammalia) with pairwise comparisons of vertebral shape covariation (i.e. integration) and evaluate our results against hypotheses of developmental and functional modularity. We used three-dimensional geometric morphometrics to quantify vertebral shape and then assessed integration between pairs of vertebrae with phylogenetic two-block partial least square analysis (PLS). RESULTS: Six modules were identified in the pairwise analyses (vertebrae included are designated as ‘C’ for cervical, ‘T’ for thoracic, and ‘L’ for lumbar): an anterior module (C1 to T1); a transitional module situated between the last cervicals and first thoracics (C6 to T2); an anterior to middle thoracic set (T4 to T8); an anticlinal module (T10 and T11); a posterior set composed of the last two thoracics and lumbars (T12 to L7); and a module showing covariation between the cervicals and the posterior set (T12 to L7). These modules reflect shared developmental pathways, ossification timing, and observed ecological shape diversification in living species of felids. CONCLUSIONS: We show here that patterns of shape integration reflect modular organisation of the vertebral column of felids. Whereas this pattern corresponds with hypotheses of developmental and functional regionalisation in the axial skeleton, it does not simply reflect major vertebral regions. This modularity may also have permitted vertebral partitions, specifically in the posterior vertebral column, to be more responsive to selection and achieve higher morphological disparity than other vertebral regions

Shape Covariation (or the Lack Thereof) Between Vertebrae and Other Skeletal Traits in Felids: The Whole is Not Always Greater than the Sum of Parts

Within carnivorans, cats show comparatively little disparity in overall morphology, with species differing mainly in body size. However, detailed shape analyses of individual osteological structures, such as limbs or skulls, have shown that felids display significant morphological differences that correlate with their observed ecological and behavioural ranges. Recently, these shape analyses have been extended to the felid axial skeleton. Results demonstrate a functionally-partitioned vertebral column, with regions varying greatly in level of correlation between shape and ecology. Moreover, a clear distinction is evident between a phylogenetically-constrained neck region and a selection-responsive posterior spine. Here, we test whether this regionalisation of function reflected in vertebral column shape is also translated into varying levels of phenotypic integration between this structure and most other skeletal elements. We accomplish this comparison by performing pairwise tests of integration between vertebral and other osteological units, quantified with 3D geometric morphometric data and analysed both with and without phylogenetic correction. To our knowledge, this is the first study to test for integration across a comprehensive sample of whole-skeleton elements. Our results show that, prior to corrections, strong covariation is present between vertebrae across the vertebral column and all other elements, with the exception of the femur. However, most of these significant correlations disappear after correcting for phylogeny, which is a significant influence on cranial and limb morphology of felids and other carnivorans. Our results thus suggest that the vertebral column of cats displays relative independence from other skeletal elements and may represent several distinct evolutionary morphological modules

Der Effekt von Dexamethason und Triiodthyronin auf primäre bovine Chondrozyten und chondrogen differenzierte mesenchymale Stammzellen

Die noch junge Disziplin der regenerativen Medizin eröffnet neue Lösungsansätze für die Behandlung großer Knochen- und Knorpeldefekte. Mesenchymale Stammzellen, ebenso wie Gelenk-Chondrozyten, sind mögliche Ausgangszellen für die Herstellung von Knochen oder Knorpel. Die enchondralen Ossifizierung von Knorpel stellt einen physiologischen Prozess der Knochenneubildung dar, reguliert - unter anderem - durch die Hormone Dexamethason und Triiodthyronin. In der vorliegenden Arbeit werden die Einflüsse dieser beiden Hormone auf Gelenkchondrozyten und mesenchymale Stammzellen untersucht, ausgehend von der Hypothese, dass diese Hormone in den Zellen den Prozess der enchondralen Ossifizierung auslösen können, und dass Chondrozyten und chondrogen differenzierte Stammzellen sich in ihrer Antwort auf diesen Hormonstimulus ähnlich verhalten würden. In einem 3D-Zellkultursystem wurden bovine Chondrozyten und chondrogen differenzierte mesenchymale Stammzellen kultiviert und unter definierten Bedingungen dem Einfluss von Triiodthyronin und Dexamethason ausgesetzt. Neben Zellproliferation und Matrix-Biosynthese wurde die mRNA Expression von Kollagen Typ I, II und X in der real-time PCR gemessen, sowie zusätzlich die Aktivität der alkalischen Phosphatase und die Deposition von Kalzium in der Matrix als Zeichen der terminalen Differenzierung der Zellen, welche notwendigerweise der enchondralen Ossifizierung vorangehen muss. Das Alginat-Zellkultursystem bewährte sich in der Anwendung gut, sowohl für die Kultur der primären Chondrozyten als auch für die Differenzierung der mesenchymalen Stammzellen. Dexamethason führte zu einer gesteigerten Produktion von Glykosaminoglykanen. Triiodthyronin steigerte nur in den primären Chondrozyten die Produktion von Kollagen, und induzierte in diesen Zellen Zeichen der terminalen Differenzierung, also einen Anstieg sowohl in der Aktivität der alkalischen Phosphatase, als auch der Kollagen Typ X mRNA. Dexamethason hatte einen vergleichbaren Effekt auf die chondrogen differenzierten mesenchymalen Stammzellen. Die noch immaturen Chondrozyten in dieser Studie zeigten ein Potenzial für eine terminale Differenzierung und enchondrale Ossifizierung, was diese Zellen potentiell für die Verwendung zum Tissue Engineering von Knochen nutzbar macht. Zugleich deutet dies auf einen möglichen pathogenetischen Prozess in der Entstehung der Gelenkarthrose hin. Die chondrogen differenzierten Zellen wurden durch Dexamethason zur terminalen Differenzierung gebracht, was ihren passageren Phänotyp unterstreicht, und als mögliches Modell für die Enchondrale Ossifizierung genutzt werden kann. Andererseits muss diese Limitation bei der Nutzung von Stammzellen zur Knorpelregeneration beachtet werden.The Effect of Dexamethasone and Triiodothyronine on Terminal Differentiation of Primary Bovine Chondrocytes and Chondrogenically Differentiated Mesenchymal Stem Cells The newly evolved field of regenerative medicine is offering solutions in the treatment of bone or cartilage loss and deficiency. Mesenchymal stem cells, as well as articular chondrocytes, are potential cells for the generation of bone or cartilage. The natural mechanism of bone formation is that of endochondral ossification, regulated, among other factors, through the hormones dexamethasone and triiodothyronine. We investigated the effects of these hormones on articular chondrocytes and chondrogenically differentiated mesenchymal stem cells, hypothesizing that these hormones would induce terminal differentiation, with chondrocytes and differentiated stem cells being similar in their response. Using a 3D-alginate cell culture model, bovine chondrocytes and chondrogenically differentiated stem cells were cultured in presence of triiodothyronine or dexamethasone, and cell proliferation and extracellular matrix production were investigated. Collagen mRNA expression was measured by real-time PCR. Col X mRNA and alkaline phosphatase were monitored as markers of terminal differentiation, a prerequisite of endochondral ossification. The alginate culture system worked well, both for the culture of chondrocytes and for the chondrogenic differentiation of mesenchymal stem cells. Dexamethasone led to an increase in glycosaminoglycan production. Triiodothyronine increased the total collagen production only in chondrocytes, where it also induced signs of terminal differentiation, increasing both collagen X mRNA and alkaline phosphatase activity. Dexamethasone induced terminal differentiation in the differentiated stem cells. The immature articular chondrocytes used in this study seem to be able to undergo terminal differentiation, pointing to their possible role in the onset of degenerative osteoarthritis, as well as their potential for a cell source in bone tissue engineering. When chondrocyte-like cells, after their differentiation, can indeed be moved on towards terminal differentiation, they can be used to generate a model of endochondral ossification, but this limitation must be kept in mind when using them in cartilage tissue engineering application.

Barnfattigdom, en begreppsfråga? - En diskursanalys av barnfattigdomsdebatten

This paper is about the debate on child poverty in Sweden. The study is designed as a discourse analysis of editorials written after an audit program on Swedish TV, Uppdrag Granskning (Mission Review). The program was a critic for statistics and campaigns that child rights organizations have come out with current poor children in Sweden. The review was whether these data were incorrect or not and the reporter wanted to show that there were no poor children in Sweden. Our study deals with how various writers on the leader pages have reacted to the program and their position in the debate about how child poverty looks like in Sweden. We have, on the basis of discourse analysis, social constructionism and political ideologies analyzed how these writers write about poverty, how the terms are used and what is different, and is common to the different sides in the debate. We have analyzed the constructions that form the basis of how writers write. We have come to the conclusion that what it's different in the debate is how to define who is poor and needs help from the society. Much of the debate is also about trust in the children's rights organizations and if their statistic can be trusted. Something in common that we could find in our articles was that everyone seemed to agree that society has a responsibility to ensure that no child should have to be poor in Sweden and that children in these situations are described as victims

Cleavage of intron from the standard or non-standard position of the precursor tRNA by the splicing endonuclease of Aeropyrum pernix, a hyper-thermophilic Crenarchaeon, involves a novel RNA recognition site in the Crenarchaea specific loop

In Crenarchaea, several tRNA genes are predicted to express precursor-tRNAs (pre-tRNAs) with canonical or non-canonical introns at various positions. We initially focused on the tRNAThr species of hyperthermophilic crenarchaeon, Aeropyrum pernix (APE) and found that in the living APE cells three tRNAThr species were transcribed and subsequently matured to functional tRNAs. During maturation, introns in two of them were cleaved from standard and non-standard positions. Biochemical studies revealed that the APE splicing endonuclease (APE-EndA) removed both types of introns, including the non-canonical introns, without any nucleotide modification. To clarify the underlying reasons for broad substrate specificity of APE-EndA, we determined the crystal structure of wild-type APE-EndA and subsequently compared its structure with that of Archaeaoglobus fulgidus (AFU)-EndA, which has narrow substrate specificity. Remarkably, structural comparison revealed that APE-EndA possesses a Crenarchaea specific loop (CSL). Introduction of CSL into AFU-EndA enhanced its intron-cleaving activity irrespective of the position or motif of the intron. Thus, our biochemical and crystallographic analyses of the chimera-EndA demonstrated that the CSL is responsible for the broad substrate specificity of APE-EndA. Furthermore, mutagenesis studies revealed that Lys44 in CSL functions as the RNA recognition site

Estudio anatómico e histoquímico del tallo y las hojas de Jasminum azoricum L.

El presente estudio tuvo como objetivo caracterizar la anatomía e histoquímica del tallo y las hojas de Jasminum azoricum L. Para ello, se realizaron preparaciones histológicas semipermanentes que contenían cortes transversales de tallo y hoja, además de cortes paradérmicos de la lámina foliar, analizados por microscopía óptica. También se realizaron pruebas histoquímicas para verificar la ubicación de acumulación de constituyentes químicos en secciones transversales de la hoja. Las caracterizaciones anatómicas aportaron importantes datos para distinguir las especies y el estudio histoquímico reveló los sitios de síntesis y/o almacenamiento de los metabolitos que poseen importantes propiedades medicinales.The present study aimed to characterize the anatomy and histochemistry of the stem and leaves of Jasminum azoricum L. Thus, semi-permanent slides were prepared containing transverse sections of the stem and leaf, in addition to paradermal sections of the leaf blade, analyzed by optical microscopy. Histochemical tests were also carried out in order to verify the site of accumulation of chemical constituents in cross-sections of the leaf blade. The anatomical characterizations allowed the identification of important characteristics to distinguish the species and the histochemistry revealed the sites of synthesis and/or storage of the metabolites. The results found help to identify the species studied and point out possible metabolites that have considerable medicinal properties

Drivers and constraints of shape evolution in the vertebral column of felids (Felidae, Carnivora)

Morphological studies of the skull and limbs of tetrapods are common in the literature. Nonetheless, the vertebral column has been comparatively neglected, and research shows a bias towards developmental and genetic approaches. Still, these studies have highlighted the unusual uniformity in vertebral count across mammals, unlike the great variation in vertebral numbers observed in other tetrapod clades. This meristic constraint has been suggested to drive higher regionalisation in the mammalian axial skeleton, with adaptation to discrete niches happening primarily through modification of vertebral form rather than changes in numbers. Living species of the mammalian family Felidae are an ideal group for vertebral studies as all taxa present the same count of 27 presacral vertebrae but vary in ecological specialisations and body mass. In this thesis, I explore the morphological evolution of the presacral vertebral column by, first, investigating ecological and phylogenetic influences on presacral vertebral shape, and then, examining patterns of vertebral trait covariation with an evolutionary developmental perspective. My results show clear regionalisation of vertebral column shape and function. Specifically, a highly integrated region between the diaphragmatic vertebra and the last lumbar (i.e., T10 – L7) shows the highest levels of ecological 6 specialization, and potentially higher evolvability, contrasting with a phylogenetically conserved neck region. I found strong support for a widespread two-module model of intravertebral shape based on developmental origins of vertebral components, and this analysis also provided an empirical example of phenotypic integration promoting higher morphological disparity. Exceptions to this model are at boundaries of large vertebral modules and suggest functional overprinting of developmental patterns. Further, I demonstrated the presence of modularity at the organismal level, with decoupling of the vertebral column as a whole from other skeletal structures. Combined, the work presented in this thesis demonstrates that axial evolution across Felidae reflects both developmental constraints and functional specialisation by concentrating shape change within distinct evolutionary modules. This thesis provides a foundation for further study of vertebral columns combining both functional and developmental perspectives

Interference activity of a minimal Type I CRISPR-Cas system from Shewanella putrefaciens

Type I CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-Cas (CRISPR-associated) systems exist in bacterial and archaeal organisms and provide immunity against foreign DNA. The Cas protein content of the DNA interference complexes (termed Cascade) varies between different CRISPR-Cas subtypes. A minimal variant of the Type I-F system was identified in proteobacterial species including Shewanella putrefaciens CN-32. This variant lacks a large subunit (Csy1),Csy2 and Csy3 and contains two unclassified cas genes. The genome of S. putrefaciens CN-32 contains only five Cas proteins (Cas1, Cas3, Cas6f, Cas1821 and Cas1822) and a single CRISPR array with 81 spacers. RNA-Seq analyses revealed the transcription of this array and the maturation of crRNAs (CRISPR RNAs). Interference assays based on plasmid conjugation demonstrated that this CRISPR-Cas system is active in vivo and that activity is dependent on the recognition of the dinucleotide GG PAM (Protospacer Adjacent Motif) sequence and crRNA abundance. The deletion of cas1821 and cas1822 reduced the cellular crRNA pool. Recombinant Cas1821 was shown to form helical filaments bound to RNA molecules, which suggests its role as the Cascade backbone protein. A Cascade complex was isolated which contained multiple Cas1821 copies, Cas1822, Cas6f and mature crRNAs

Interference activity of a minimal Type I CRISPR-Cas system from Shewanella putrefaciens

Type I CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-Cas (CRISPR-associated) systems exist in bacterial and archaeal organisms and provide immunity against foreign DNA. The Cas protein content of the DNA interference complexes (termed Cascade) varies between different CRISPR-Cas subtypes. A minimal variant of the Type I-F system was identified in proteobacterial species including Shewanella putrefaciens CN-32. This variant lacks a large subunit (Csy1),Csy2 and Csy3 and contains two unclassified cas genes. The genome of S. putrefaciens CN-32 contains only five Cas proteins (Cas1, Cas3, Cas6f, Cas1821 and Cas1822) and a single CRISPR array with 81 spacers. RNA-Seq analyses revealed the transcription of this array and the maturation of crRNAs (CRISPR RNAs). Interference assays based on plasmid conjugation demonstrated that this CRISPR-Cas system is active in vivo and that activity is dependent on the recognition of the dinucleotide GG PAM (Protospacer Adjacent Motif) sequence and crRNA abundance. The deletion of cas1821 and cas1822 reduced the cellular crRNA pool. Recombinant Cas1821 was shown to form helical filaments bound to RNA molecules, which suggests its role as the Cascade backbone protein. A Cascade complex was isolated which contained multiple Cas1821 copies, Cas1822, Cas6f and mature crRNAs