Polymorphonuclear leukocyte migration through human amnion membrane

A new in vitro model has been developed for studying migration of human polymorphonuclear leukocytes (PMN) through living native cellular and matrix barriers. Human amnion membrane consists of a single layer of epithelium bound to a continuous basement membrane interfacing an avascular collagenous stroma. Living amnion was placed in plastic chambers with separate compartments on each side of the membrane. PMN were introduced on the epithelial side of the amnion, and a Millipore filter (Millipore Corp., Bedford, Mass.) was placed against the stromal side. In response to N-formylmethionyl-leucyl- phenylanlanine (FMLP) chemoattractant, PMN penetrated the full thickness of the amnion and were collected and counted on the filter. The rate of PMN traversal of the amnion was dependent on the concentration of FMLP (optimal at 10(-8)M) as well as the slope of the FMLP gradient across the amnion. The route of PMN migration was studied by transmission electron microscopy. PMN first attached to the epithelial surface, then infiltrated between intercellular junctions. PMN migrated around or through tight junction and hemidesmosome attachments. The PMN then penetrated the basement membrane and migrated through the dense collagenous stroma. The present amnion migration system has characteristics of the in vivo inflammatory state not described in any previous method for monitoring PMN migration in vitro. Prior methods have not used native epithelium, whole basement membrane, or collagenous stroma. PMN penetration of these barriers occurs in the normal inflammatory response and probably involves biochemical mechanisms not required for simple migration through the pores of an artificial filter. The amnion system can be useful for future biochemical and morphological studies of PMN penetration of these barriers and possible repair processes that may follow.     

Vergleichende Untersuchungen zum Kontaktverhalten verschiedener Arten der Gattung Tilapia (Cichlidae,Pisces) und ihrer Bastarde

• 1 Tilapia tholloni (Substratbrüter), T. nilotica (weiblicher Maulbrüter) und T. heudeloti macrocephala (männlicher Maulbrüter) wurden künstlich erbrütet und ihre angeborenen Kontaktreaktionen in standardisierten Attrappenversuchen untersucht.
• 2 Das Kontaktverhalten muß während einer kritischen Phase (bei T. nilotica unter den angegebenen Versuchsbedingungen bis ungefähr zum 21. Tag nach dem Ablaichen) aktiviert werden, wenn es längere Zeit andauern soll (= Reaktionsphase). Die Reaktionsphase kann bei T. nilotica mehrere Wochen dauern. In ihr nimmt die Reaktionsstärke (Anzahl und Dauer der Kontakte) zunächst rasch zu, erreicht ein Maximum und nimmt dann allmählich wieder ab. Anstieg, Maximum und Abnahme sind an bestimmte Entwicklungsabschnitte gebunden, weitgehend unabhängig davon, ob die Tiere zuvor schon Kontaktverhalten geäußert haben oder nicht. Die Kontaktreaktionen unterscheiden sich u. a. durch die Dauer der Kontakte: tholloni = 0,7 Sek., nilotica = 86,5 Sek., heudeloti m. = 1,5 Sek. je Tag und Tier (Maximalwerte bei bestimmten, für alle Arten gleichen Versuchsbedingungen).
• 3 Von der Aufzuchttemperatur hängt es ab, in welchem Entwicklungsabschnitt die Reaktionsphase liegt. Das Reaktionsmaximum junger T. nilotica lag bei 24° C am 9., bei 29° C am Tag nach der Eiablage.
• 4 Geblendete nilotica-Jungfische zeigten nur zu Beginn der Reaktionsphase schwaches Kontaktverhalten, normale Tiere äußerten gegenüber einer durchsichtigen Glasattrappe abgeschwächtes, nur kurze Zeit dauerndes Kontaktverhalten. Das Kontaktverhalten wird durch mechanische (Strömung) und optische Reize ausgelöter und gesteuert.
• 5 Junge Maulbrüter aus kleinen Eiern erreichten eine längere Kontaktdauer als solche aus größeren. Nach künstlicher Reduktion der Dottermenge um 10–20%) erhöhte sich die Kontaktdauer bei jungen nilotica um 23,1%, die Zahl der Kontakte nahm gleichzeitig um 7,6% ab. Die ♀♀ der substratbrütenden T. mariae legen große Eier, die Jungen zeigten gegenüber Attrappen intensive Kontaktreaktionen. Es wird die Frage diskutiert, ob das Erreichen einer bestimmten Eigröße eine Voraussetzung für das Entstehen des Kontaktverhaltens gewesen sein könnte.
• 6 Die Kurzkontakte junger T. tholloni werden aus ihrer Orientierungsreaktion abgeleitet und als Vorstufe des Kontaktverhaltens gedeutet.
• 7 7. Mehrere Tilapia-Arten wurden künstlich gekreuzt. Bei Verwendung von tholloni-Sperma wiesen die Bastarde eine erhöhte Sterblichkeit auf. Aus der Kreuzung T. tholloni ♀ ～ T. nilotica ♂ (Substratbrüter ～ weiblicher Maulbrüter) gingen nur ♀♀ hervor. Die Kreuzung T. heudeloti macrocephala ～ T. nilotica (männlicher ～ weiblicher Maulbrüter) erbrachte fertile F₁- und F₂-Generationen sowie alle vier möglichen Rückkreuzungen. Bei der Vererbung des Kontaktverhaltens (gemessen an der Dauer der Kontakte) scheinen relativ wenig Erbfaktoren mitzuwirken, T. heudeloti m. erwies sich gegenüber nilotica als praevalent. Ein Teil der Bastarde aus der Kreuzung T. tholloni ♀ ～ T. nilotica ♂ (F₁) (Substratbrüter ～ weiblicher Maulbrüter) lag auf der Merkmalsskala zwischen den Ausgangsarten, der Rest verteilte sich sowohl auf den Bereich von tholloni als auch auf den Bereich von nilotica.

     

Homopolygalacturonan Molecular Size in Plant Cell Wall Matrices Via Paramagnetic Ion and Nitroxyl Amide Dipolar Spin-Spin Interactions

Mn²⁺, Cu²⁺, and nitroxyl amines have been shown to bond to plant homopolygalacturonan matrices in a spatially sequential fashion. As a consequence of this special form of cooperativity the lattice constant (κ), determined from Van Vleck's second moment relationship, approaches 1 only when the average number of dipolar interactions per spin approaches 1 (e.g., an array of dimers). Assuming that one paramagnetic ion or nitroxyl amide pair is bonded per polymer block within the matrix when κ = 1, the anionic ligand's average degree of polymerization ([unk]) can be estimated from the concentration of bonded paramagnetic dimers (e.g., [1/χ]_κ~1 = [unk]; χ is the mole fraction of bonded paramagnetic dimers). We have utilized this technique to estimate the average molecular size of homopolygalacturonan blocks in intact higher plant cortical cell walls ([unk] ~83), Nitella cell walls ([unk] ~27) and a commercially available galacturonic acid polymer ([unk] ~35). The [unk] determined from both the intact cortical cell wall lattice and the polygalacturonan were similar to literature values; these findings argue that the electron paramagnetic resonance, (EPR) dipolar spin-spin interaction technique reported herein is a valid approach for estimating molecular size in plant cell walls.     

Using functional groups to investigate community response to environmental changes: two grassland case studies

Functional groups (FG) are an useful generalization to investigate environmental change effects on biotic communities. Assigning species to FGs is a contextual task and carries an arbitrary element, regardless of whether the grouping is obtained a priori or by sophisticated numerical methods. Using two grassland community case studies, we show that even simple FG allocation based on growth form, architecture and longevity (plants and mosses), or foraging characteristics (above-ground invertebrates) can be useful to increase our understanding of community processes. For example, the sensitivity of organisms to climate change increases with trophic rank and is higher in disturbed than in undisturbed communities. Complexity of interaction webs (in terms of web connectance), however, is larger in undisturbed than in disturbed communities. A significant and important relationship is likely to exist between anthropogenic disturbances, community complexity and the ecosystem effects of climate change. Trophic interactions may be disrupted much easier by climate changes in disturbed than in undisturbed communities where complexity may be buffering these effects.     

Development of Candida albicans hyphae in different growth media--variations in growth rates, cell dimensions and timing of morphogenetic events

In six liquid culture media, all of which stimulated Candida albicans to grow in the hyphal form, the rates of hyphal extension and increase in cellular ATP concentration, hyphal diameters, times of evagination of hyphae, times of septum formation and positions of septa in the hyphae appeared to vary independently. There were no discernible associations between properties such as length or volume of hyphal compartments at the time of septation and temporal parameters of hyphal growth. The results suggest that growth environment influences many of the processes contributing to hyphal development, but that these processes are not necessarily interrelated.     

Variations among cell lines in the synthesis of sphingolipids in de novo and recycling pathways

There are several pathways for the incorporation of sugars into glycosphingolipids (GSL). Sugars can be added to ceramide that contains sphinganine (dihydrosphingosine) synthesized de novo (pathway 1), to ceramide synthesized from sphingoid bases produced by hydrolysis of sphingolipids (pathway 2), and into GSL recycling from the endosomal pathway through the Golgi (pathway 3). We reported previously the surprising observation that SW13 cells, a human adrenal carcinoma cell line, synthesize most of their GSL in pathway 2. We now present data on the synthesis of GSL in four additional cell lines. Approximately 90% of sugar incorporation took place in pathway 2, and 10% or less in pathway 1, in human foreskin fibroblasts and NB41A3 neuroblastoma cells. In contrast, approximately 50-90% of sugar incorporation took place in pathway 1 in C2C12 myoblasts. The C2C12 cells divide more rapidly and synthesize 10-14 times as much GSL as the other three cell lines. In C6 glioma cells, approximately 30% of sugar incorporation occurred in pathway 1 and 60% in pathway 2. There was no relation between the utilization of pathways for GSL and sphingomyelin synthesis in foreskin fibroblasts and C2C12 cells. In both cells pathways 1 and 2 each accounted for 50% of incorporation of choline into sphingomyelin. In five of the six cell lines that we have studied, most GSL synthesis takes place in pathway 2. We suggest that when the need for synthesis is relatively low, as in slowly dividing cells, GSL are synthesized predominantly from sphingoid bases salvaged from the hydrolytic pathway. When cells are dividing more rapidly, the need for increased synthesis is met by upregulating the de novo pathway.      

Growth and macromolecular content of the dimorphic fungus Aureobasidium pullulans and the effect of hydroxyurea and other inhibitors

The growth kinetics and the macromolecular content of the yeast and ethanol-induced hyphal forms of Aureobasidium pullulans were studied. During the morphological transition from yeasts to hyphae, both the protein and RNA content decreased significantly, the mycelial form containing only 76% of the amount of protein in the yeasts, and 38% of the RNA. The DNA was the only component tested whose level increased during the transition. Among several compounds inhibiting macromolecular synthesis, only hydroxyurea showed a remarkable effect on the morphology of A. pullulans, inducing the mycelial morphology. The macromolecular composition of hydroxyurea-treated cultures changed with time in a way similar to that of the ethanol-Tween 80-ammonia medium, and to that of carbon-starved cultures, without ethanol or glucose.     

Vasorelaxant activity of phthalazinones and related compounds

Several series of dihydrostilbenamide, imidazo[2,1-a]isoindole, pyrimido[2,1-a]isoindole and phthalazinone derivatives were obtained and their vasorelaxant activity was measured on isolated rat aorta rings pre-contracted with phenylephrine (10(-5)M). Some phthalazinones attained, practically, the total relaxation of the organ at micromolar concentrations. For the most potent compound 9h (EC(50)=0.43microM) the affinities for alpha(1A), alpha(1B) and alpha(1D) adrenergic sub-receptors were determined.     

Parallels,nonparallels, and plasticity in population differentiation of threespine stickleback within a lake

The frequent occurrence of parallel phenotypic divergence in similar habitats is often evoked when emphasizing the role of ecology in adaptive radiation and speciation. However, because phenotypic plasticity can contribute to the observed pattern of divergence, confirmation of divergence at loci underlying phenotypic traits is important for confirming adaptive divergence. In the present study, we examine parallel morphological, neutral, and potentially adaptive genetic divergence of threespine stickleback inhabiting different habitats within a lake. Three genetic clusters best explained the neutral genetic structure within the lake; however, morphological differences were only weakly connected to genetic clusters and there was considerable phenotypic variation within clusters. Among the factors that could contribute to the observed pattern of morphological and genetic divergence are phenotypic plasticity, selective mortality of hybrids, and habitat choice based on morphology. Several loci are identified as outliers indicating divergent selection between the morphs and some parallels in morphological and adaptive genetic divergence are found in stickleback spawning at two lava sites. However, neutral genetic structure indicates considerable genetic connectivity among the two lava sites, and the parallels in morphology may therefore represent selective distribution of phenotypes rather than parallel divergence. © 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 98 , 803–813.