Molecular basis for the development of diagnostic methods and specific treatment modalities for idiopathic pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is an interstitial lung disease with unknown aetiology and poor prognosis. IPF is characterized by alveolar epithelial damage that leads tissue remodelling and ultimately to the loss of normal lung architecture and function. Treatment has been focused on anti-inflammatory therapies, but due to their poor efficacy new therapeutic modalities are being sought. There is a need for early diagnosis and also for differential diagnostic markers for IPF and other interstitial lung diseases. The study utilized patient material obtained from bronchoalveolar lavage (BAL), diagnostic biopsies or lung transplantation. Human pulmonary fibroblast cell cultures were propagated and asbestos-induced pulmonary fibrosis in mice was used as an experimental animal model of IPF. The possible markers for IPF were scanned by immunohistochemistry, RT-PCR, ELISA and western blot. Matrix metalloproteinases (MMPs) are proteolytic enzymes that participate in tissue remodelling. Microarray studies have introduced potential markers that could serve as additional tools for the assessment of IPF and one of the most promising was MMP 7. MMP-7 protein levels were measured in the BAL fluid of patients with idiopathic interstitial lung diseases or idiopathic cough. MMP-7 was however similarly elevated in the BAL fluid of all these disorders and thus cannot be used as a differential diagnostic marker for IPF. Activation of transforming growth factor (TGF)-ß is considered to be a key element in the progression of IPF. Bone morphogenetic proteins (BMP) are negative regulators of intracellular TGF-ß signalling and BMP-4 signalling is in turn negatively regulated by gremlin. Gremlin was found to be highly upregulated in the IPF lungs and IPF fibroblasts. Gremlin was detected in the thickened IPF parenchyma and endothelium of small capillaries, whereas in non-specific interstitial pneumonia it localized predominantly in the alveolar epithelium. Parenchymal gremlin immunoreactivity might indicate IPF-type interstitial pneumonia. Gremlin mRNA levels were higher in patients with end-stage fibrosis suggesting that gremlin might be a marker for more advanced disease. Characterization of the fibroblastic foci in the IPF lungs showed that immunoreactivity to platelet-derived growth factor (PDGF) receptor-α and PDGF receptor-β was elevated in IPF parenchyma, but the fibroblastic foci showed only minor immunoreactivity to the PDGF receptors or the antioxidant peroxiredoxin II. Ki67 positive cells were also observed predominantly outside the fibroblastic foci, suggesting that the fibroblastic foci may not be composed of actively proliferating cells. When inhibition of profibrotic PDGF-signalling by imatinib mesylate was assessed, imatinib mesylate reduced asbestos-induced pulmonary fibrosis in mice as well as human pulmonary fibroblast migration in vitro but it had no effect on the lung inflammation.

Nanoscale Heterostructures with Molecular-Scale Single-Crystal Metal Wires

Creating nanoscale heterostructures with molecular-scale (<2 nm) metal wires is critical for many applications and remains a challenge. Here, we report the first time synthesis of nanoscale heterostructures with single-crystal molecular-scale Au nanowires attached to different nanostructure substrates. Our method involves the formation of Au nanoparticle seeds by the reduction of rocksalt AuCl nanocubes heterogeneously nucleated on the Substrates and subsequent nanowire growth by oriented attachment of Au nanoparticles from the Solution phase. Nanoscale heterostructures fabricated by such site-specific nucleation and growth are attractive for many applications including nanoelectronic device wiring, catalysis, and sensing.

Fluorescent Fields: electric lighting and the rationalization of the modern corporate workplace

This study investigates the implications of the introduction of electric lighting systems, building technologies, and theories of worker efficiency on the deep spatial and environmental transformations that occurred within the corporate workplace during the twentieth century. Examining the shift from daylighting strategies to largely artificially lit workplace environments, this paper argues that electric lighting significantly contributed to the architectural rationalization of both office work and the modern office environment. Contesting the historical and critical marginalization of lighting within the discourse of the modern built environment, this study calls for a reassessment of the role of artificial lighting in the development of the modern corporate workplace. Keywords: daylighting, fluorescent lighting, rationalization, workplace design

Inferring molecular function: contributions from functional linkages

In the current era of high-throughput sequencing and structure determination, functional annotation has become a bottleneck in biomedical science. Here, we show that automated inference of molecular function using functional linkages among genes increases the accuracy of functional assignments by >= 8% and enriches functional descriptions in >= 34% of top assignments. Furthermore, biochemical literature supports >80% of automated inferences for previously unannotated proteins. These results emphasize the benefit of incorporating functional linkages in protein annotation.

Molecular phylogeny and biogeography of langurs and leaf monkeys of South Asia (Primates: Colobinae)

The two recently proposed taxonomies of the langurs and leaf monkeys (Subfamily Colobinae) provide different implications to our understanding of the evolution of Nilgiri and purple-faced langurs. Groves (2001) [Groves, C.P., 2001. Primate Taxonomy. Smithsonian Institute Press, Washington], placed Nilgiri and purple-faced langurs in the genus Trachypithecus, thereby suggesting disjunct distribution of the genus Trachypithecus. [Brandon-Jones, D., Eudey, A.A., Geissmann, T., Groves, C.P., Melnick, D.J., Morales, J.C., Shekelle, M., Stewart, C.-B., 2003. Asian primate classification. Int. J. Primatol. 25, 97–162] placed these langurs in the genus Semnopithecus, which suggests convergence of morphological characters in Nilgiri and purple-faced langurs with Trachypithecus. To test these scenarios, we sequenced and analyzed the mitochondrial cytochrome b gene and two nuclear DNA-encoded genes, lysozyme and protamine P1, from a variety of colobine species. All three markers support the clustering of Nilgiri and purple-faced langurs with Hanuman langur (Semnopithecus), while leaf monkeys of Southeast Asian (Trachypithecus) form a distinct clade. The phylogenetic position of capped and golden leaf monkeys is still unresolved. It is likely that this species group might have evolved due to past hybridization between Semnopithecus and Trachypithecus clades.

Biology and Molecular Markers of Malignant Gonadal Germ Cell Tumors

Germ cell tumors occur both in the gonads of both sexes and in extra-gonadal sites during adoles-cence and early adulthood. Malignant ovarian germ cell tumors are rare neoplasms accounting for less than 5% of all cases of ovarian malignancy. In contrast, testicular cancer is the most common malignancy among young males. Most of patients survive the disease. Prognostic factors of gonadal germ cell tumors include histology, clinical stage, size of the primary tumor and residua, and levels of tumor markers. Germ cell tumors include heterogeneous histological subgroups. The most common subgroup includes germinomas (ovarian dysgerminoma and testicular seminoma); other subgroups are yolk sac tumors, embryonal carcinomas, immature teratomas and mixed tumors. The origin of germ cell tumors is most likely primordial germ cells. Factors behind germ cell tumor development and differentiation are still poorly known. The purpose of this study was to define novel diagnostic and prognostic factors for malignant gonadal germ cell tumors. In addition, the aim was to shed further light into the molecular mechanisms regulating gonadal germ cell tumorigenesis and differentiation by studying the roles of GATA transcription factors, pluripotent factors Oct-3/4 and AP-2γ, and estrogen receptors. This study revealed the prognostic value of CA-125 in malignant ovarian germ cell tumors. In addition advanced age and residual tumor had more adverse outcome. Several novel markers for histological diagnosis were defined. In the fetal development transcription factor GATA-4 was expressed in early fetal gonocytes and in testicular carcinoma precursor cells. In addition, GATA-4 was expressed in both gonadal germinomas, thus it may play a role in the development and differentiation of the germinoma tumor subtype. Pluripotent factors Oct-3/4 and AP-2γ were expressed in dysgerminomas, thus they could be used in the differential diagnosis of the germ cell tumors. Malignant ovarian germ cell tumors expressed estrogen receptors and their co-regulator SNURF. In addition, estrogen receptor expression was up-regulated by estradiol stimulation. Thus, gonadal steroid hormone burst in puberty may play a role in germ cell tumor development in the ovary. This study shed further light in to the molecular pathology of malignant gonadal germ cell tumors. In addition, some novel diagnostic and prognostic factors were defined. This data may be used in the differential diagnosis of germ cell tumor patients.

Melting of an Anchored Bilayer: Molecular Dynamics Simulations of the Structural Transition in (CnH2n+1NH3)(2)PbI4 (n=12, 14, 16, 18)

The thermally driven Structural phase transition in the organic-inorganic hybrid perovskite (CnH2n+1NH3)(2)PbI4 has been investigated using molecular dynamics (MD) simulations. This system consists of positively charged alkyl-amine chains anchored to a rigid negatively charged PbI4 sheet with the chains organized as bilayers with a herringbone arrangement. Atomistic simulations were performed using ail isothermal-isobaric ensemble over a wide temperature range from 65 to 665 K for different alkyl chain lengths, n = 12, 14, 16, and 18. The simulations are able to reproduce the essential Features of the experimental observations of this system, including the existence of a transition, the linear variation of the transition temperature with alkyl chain length, and the expansion of the bilayer thickness at the transition. By use of the distance fluctuation Criteria, it is Shown that the transition is associated With a Melting of the alkyl chains of the anchored bilayer. Ail analysis of the conformation of the alkyl chains shows increased disorder in the form of gauche defects above due melting transition. Simulations also show that the melting transition is characterized by the complete disappearance of all-trans alkyl chains in the anchored bilayer, in agreement with experimental observations. A conformationally disordered chain has a larger effective cross-sectional area, and above due transition a uniformly tilted arrangement of the anchored chains call no longer be Sustained. At the melt the angular distribution of the orientation of the chains are 110 longer uniform; the chains are splayed allowing for increased space for individual chains of the anchored bilayer. This is reflected in a sharp rise in the ratio of the mean head-to-head to tail-to-tail distance of the chains of the bilayer at the transition resulting in in expansion of the bilayer thickness. The present MD simulations provide a simple explanation as to how changes in conformation of individual alkyl-chains gives rise to the observed increase in the interlayer lattice spacing of (CnH2n+1NH3)(2)PbI4 at the melting transition.

Structures and molecular-dynamics studies of three active-site mutants of bovine pancreatic phospholipase A(2)

Phospholipase A(2) hydrolyzes phospholipids at the sn-2 position to cleave the fatty-acid ester bond of L-glycerophospholipids. The catalytic dyad (Asp99 and His48) along with a nucleophilic water molecule is responsible for enzyme hydrolysis. Furthermore, the residue Asp49 in the calcium-binding loop is essential for controlling the binding of the calcium ion and the catalytic action of phospholipase A2. To elucidate the structural role of His48 and Asp49, the crystal structures of three active-site single mutants H48N, D49N and D49K have been determined at 1.9 angstrom resolution. Although the catalytically important calcium ion is present in the H48N mutant, the crystal structure shows that proton transfer is not possible from the catalytic water to the mutated residue. In the case of the Asp49 mutants, no calcium ion was found in the active site. However, the tertiary structures of the three active-site mutants are similar to that of the trigonal recombinant enzyme. Molecular-dynamics simulation studies provide a good explanation for the crystallographic results.

X-ray and neutron scattering studies on some nanoscale structures in molecular biology

Scattering of X-rays and neutrons has been applied to the study of nanostructures with interesting biological functions. The systems studied were the protein calmodulin and its complexes, bacterial virus bacteriophage phi6, and the photosynthetic antenna complex from green sulfur bacteria, chlorosome. Information gathered using various structure determination methods has been combined to the low resolution information obtained from solution scattering. Conformational changes in calmodulin-ligand complex were studied by combining the directional information obtained from residual dipole couplings in nuclear magnetic resonance to the size information obtained from small-angle X-ray scattering from solution. The locations of non-structural protein components in a model of bacteriophage phi6, based mainly on electron microscopy, were determined by neutron scattering, deuterium labeling and contrast variation. New data are presented on the structure of the photosynthetic antenna complex of green sulfur bacteria and filamentous anoxygenic phototrophs, also known as the chlorosome. The X-ray scattering and electron cryomicroscopy results from this system are interpreted in the context of a new structural model detailed in the third paper of this dissertation. The model is found to be consistent with the results obtained from various chlorosome containing bacteria. The effect of carotenoid synthesis on the chlorosome structure and self-assembly are studied by carotenoid extraction, biosynthesis inhibition and genetic manipulation of the enzymes involved in carotenoid biosynthesis. Carotenoid composition and content are found to have a marked effect on the structural parameters and morphology of chlorosomes.

On gravitational fields of stars in f(R) gravity theories

Einstein's general relativity is a classical theory of gravitation: it is a postulate on the coupling between the four-dimensional, continuos spacetime and the matter fields in the universe, and it yields their dynamical evolution. It is believed that general relativity must be replaced by a quantum theory of gravity at least at extremely high energies of the early universe and at regions of strong curvature of spacetime, cf. black holes. Various attempts to quantize gravity, including conceptually new models such as string theory, have suggested that modification to general relativity might show up even at lower energy scales. On the other hand, also the late time acceleration of the expansion of the universe, known as the dark energy problem, might originate from new gravitational physics. Thus, although there has been no direct experimental evidence contradicting general relativity so far - on the contrary, it has passed a variety of observational tests - it is a question worth asking, why should the effective theory of gravity be of the exact form of general relativity? If general relativity is modified, how do the predictions of the theory change? Furthermore, how far can we go with the changes before we are face with contradictions with the experiments? Along with the changes, could there be new phenomena, which we could measure to find hints of the form of the quantum theory of gravity? This thesis is on a class of modified gravity theories called f(R) models, and in particular on the effects of changing the theory of gravity on stellar solutions. It is discussed how experimental constraints from the measurements in the Solar System restrict the form of f(R) theories. Moreover, it is shown that models, which do not differ from general relativity at the weak field scale of the Solar System, can produce very different predictions for dense stars like neutron stars. Due to the nature of f(R) models, the role of independent connection of the spacetime is emphasized throughout the thesis.