Expression and Localization of Lung Surfactant Proteins in Human Testis

Background

Surfactant proteins (SPs) have been described in various tissues and fluids including tissues of the nasolacrimal apparatus, airways and digestive tract. Human testis have a glandular function as a part of the reproductive and the endocrine system, but no data are available on SPs in human testis and prostate under healthy and pathologic conditions.

Objective

The aim of the study was the detection and characterization of the surfactant proteins A, B, C and D (SP-A, SP-B, SP-C, SP-D) in human testis. Additionally tissue samples affected by testicular cancer were investigated.

Results

Surfactant proteins A, B, C and D were detected using RT-PCR in healthy testis. By means of Western blot analysis, these SPs were detected at the protein level in normal testis, seminoma and seminal fluid, but not in spermatozoa. Expression of SPs was weaker in seminoma compared to normal testicular tissue. SPs were localized in combination with vimentin immunohistochemically in cells of Sertoli and Leydig.

Conclusion

Surfactant proteins seem to be inherent part of the human testis. By means of physicochemical properties the proteins appear to play a role during immunological and rheological process of the testicular tissue. The presence of SP-B and SP-C in cells of Sertoli correlates with their function of fluid secretion and may support transportation of spermatozoa. In seminoma the expression of all SP''s was generally weaker compared to normal germ cells. This could lead to a reduction of immunomodulatory and rheology processes in the germ cell tumor.     

Modelling transition state analogues and enzyme-inhibitor complexes of zinc-containing class II aldolases and metalloproteases

The zinc complex Tp^*Zn---OH (1:Tp^* = hydrotris-(3-cumyl,5-methyl-pyrazolyl)borate) which is a model of hydrolytic zinc enzymes was reacted with hydroxamic acids and hydroxyketones. Two hydroxamates Tp^*Zn-Hya (2: Hya = acetohydroxamate, 3: Hya = 2-hydroxamato-4-methylpentanoyl-alanyl-glycylamide) and two ketoalcoholates Tp^*Zn-Kea (4: Kea = hydroxyacetonate, 5: Kea = cumoylacetonate) were obtained. Crystal structure determinations of 2, 3 and 5 have revealed distorted ZnN₃O₂ coordinations in each case. The immediate environment of the zinc ion in the hydroxamates closely resembles that in enzyme-(hydroxamate)inhibitor complexes of zinc-containing metalloproteases like collagenases and thermolysin or of class II aldolases like fuculose-1-phoshate aldolase. Like in the enzymes the hydroxamates and the ketoalcoholates can serve as transition state analogues of the enzyme-catalysed reactions.     

Tissue factor coagulant function is enhanced by protein-disulfide isomerase independent of oxidoreductase activity

Protein-disulfide isomerase (PDI) switches tissue factor (TF) from coagulation to signaling by targeting the allosteric Cys186-Cys209 disulfide. Here, we further characterize the interaction of purified PDI with TF. We find that PDI enhances factor VIIa-dependent substrate factor X activation 5-10-fold in the presence of wild-type, oxidized soluble TF but not TF mutants that contain an unpaired Cys186 or Cys209. PDI-accelerated factor Xa generation was blocked by bacitracin but not influenced by inhibition of vicinal thiols, reduction of PDI, changes in redox gradients, or covalent thiol modification of reduced PDI by N-ethylmaleimide or methyl-methanethiosulfonate, which abolished PDI oxidoreductase but not chaperone activity. PDI had no effect on fully active TF on either negatively charged phospholipids or in activating detergent, indicating that PDI selectively acts upon cryptic TF to facilitate ternary complex formation and macromolecular substrate turnover. PDI activation was reduced upon mutation of TF residues in proximity to the macromolecular substrate binding site, consistent with a primary interaction of PDI with TF. PDI enhanced TF coagulant activity on microvesicles shed from cells, suggesting that PDI plays a role as an activating chaperone for circulating cryptic TF.     

A leaf-based regeneration and transformation system for maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.)

Efficient methods for in vitro propagation, regeneration, and transformation of plants are of pivotal importance to both basic and applied research. While being the world’s major food crops, cereals are among the most difficult-to-handle plants in tissue culture which severely limits genetic engineering approaches. In maize, immature zygotic embryos provide the predominantly used material for establishing regeneration-competent cell or callus cultures for genetic transformation experiments. The procedures involved are demanding, laborious and time consuming and depend on greenhouse facilities. We have developed a novel tissue culture and plant regeneration system that uses maize leaf tissue and thus is independent of zygotic embryos and greenhouse facilities. We report here: (i) a protocol for the efficient induction of regeneration-competent callus from maize leaves in the dark, (ii) a protocol for inducing highly regenerable callus in the light, and (iii) the use of leaf-derived callus for the generation of stably transformed maize plants.     

Contained metabolic engineering in tomatoes by expression of carotenoid biosynthesis genes from the plastid genome

Applications of chloroplast engineering in agriculture and biotechnology will depend critically on success in extending the crop range of chloroplast transformation, and on the feasibility of expressing transgenes in edible organs (such as tubers and fruits), which often are not green and thus are much less active in chloroplast gene expression. We have improved a recently developed chloroplast-transformation system for tomato plants and applied it to engineering one of the central metabolic pathways in fruits: carotenoid biosynthesis. We report that plastid expression of a bacterial lycopene beta-cyclase gene results in herbicide resistance and triggers conversion of lycopene, the main storage carotenoid of tomatoes, to beta-carotene, resulting in fourfold enhanced pro-vitamin A content of the fruits. Our results demonstrate the feasibility of engineering nutritionally important biochemical pathways in non-green plastids by transformation of the chloroplast genome.     

Petrosal and inner ear anatomy and allometry amongst specimens referred to Litopterna (Placentalia)

New isolated petrosals from the Itaboraí beds of Brazil (late Palaeocene or early Eocene) are here described and referred to the early diverging litoptern Miguelsoria parayirunhor, based on phylogenetic, size, and abundance arguments. Both the external and internal anatomy of these specimens were investigated, which for the first time document many details of the auditory region of a Palaeogene litoptern. Our cladistic analysis, which included our new observations, failed to recover a monophyletic Litopterna but did not exclude it. A constrained analysis for the monophyly of this order showed that several features such as a (sub)quadrangular and anteroposteriorly elongated tensor tympani fossa and a large notch in the vicinity of the external opening of the cochlear canaliculus may constitute synapomorphies for Litopterna. The evolution of several other auditory characters amongst Litopterna is discussed and the relative dimensions of the inner ear and surrounding petrosal in the group were also investigated. This allowed detection of negative allometry of the bony labyrinth within the petrosal, which was confirmed by measurements and regression analysis across a larger sample of placental mammals. This scaling effect probably has an important influence on several characters of the bony labyrinth and petrosal, amongst which are the length of the vestibular aqueduct and cochlear canaliculus. It demonstrates that many aspects of the morphological variation of the bony labyrinth need to be thoroughly investigated before being incorporated into phylogenetic analyses. © 2015 The Linnean Society of London     

Biolistic co-transformation of the nuclear and plastid genomes

Particle gun-mediated (so-called 'biolistic') transformation represents a universal genetic transformation technology that is widely applied in nearly all groups of organisms. The mechanism of how accelerated DNA-coated particles, after their entry into the cell, deliver the foreign DNA to the target compartment is not known. Here we have studied this process in plants by performing co-transformation experiments with vectors targeted to two different cellular compartments, the nucleus and the plastids (chloroplasts). We find that coating of particles with both plastid and nuclear transformation vectors can result in co-transformation of chloroplasts and the nucleus. In contrast, mixing of particles coated individually with the vectors does not produce co-transformed plants. Our data suggest that a single DNA-coated particle can transform more than one compartment of the plant cell, opening up the possibility to generate doubly transgenic plants in one step. Importantly, co-transformation can also be obtained in the absence of selection, thus providing a method to produce marker-free transgenic genomes. In addition, our findings raise the possibility of occasional inadvertent co-transformation of two genomes and, therefore, have important implications for the molecular characterization and regulation of transgenic plants.     

Survival rates in a small hibernator,the edible dormouse: a comparison across Europe

Understanding how local environmental factors lead to temporal variability of vital rates and to plasticity of life history tactics is one of the central questions in population ecology. We used long‐term capture‐recapture data from five populations of a small hibernating rodent, the edible dormouse Glis glis, collected over a large geographical range across Europe, to determine and analyze both seasonal patterns of local survival and their relation to reproductive activity. In all populations studied, survival was lowest in early summer, higher in late summer and highest during hibernation in winter. In reproductive years survival was always lower than in non‐reproductive years, and females had higher survival rates than males. Very high survival rates during winter indicate that edible dormice rarely die from starvation due to insufficient energy reserves during the hibernation period. Increased mortality in early summer was most likely caused by high predation risk and unmet energy demands. Those effects have probably an even stronger impact in reproductive years, in which dormice were more active. Although these patterns could be found in all areas, there were also considerable differences in average survival rates, with resulting differences in mean lifetime reproductive success between populations. Our results suggest that edible dormice have adapted their life history strategies to maximize lifetime reproductive success depending on the area specific frequency of seeding events of trees producing energy‐rich seeds.     

Hibernation is associated with increased survival and the evolution of slow life histories among mammals

Survival probability is predicted to underlie the evolution of life histories along a slow-fast continuum. Hibernation allows a diverse range of small mammals to exhibit seasonal dormancy, which might increase survival and consequently be associated with relatively slow life histories. We used phylogenetically informed GLS models to test for an effect of hibernation on seasonal and annual survival, and on key attributes of life histories among mammals. Monthly survival was in most cases higher during hibernation compared with the active season, probably because inactivity minimizes predation. Hibernators also have approximately 15 per cent higher annual survival than similar sized non-hibernating species. As predicted, we found an effect of hibernation on the relationships between life history attributes and body mass: small hibernating mammals generally have longer maximum life spans (50% greater for a 50 g species), reproduce at slower rates, mature at older ages and have longer generation times compared with similar-sized non-hibernators. In accordance with evolutionary theories, however, hibernating species do not have longer life spans than non-hibernators with similar survival rates, nor do they have lower reproductive rates than non-hibernators with similar maximum life spans. Thus, our combined results suggest that (i) hibernation is associated with high rates of overwinter and annual survival, and (ii) an increase in survival in hibernating species is linked with the coevolution of traits indicative of relatively slow life histories.     

Diet-independent remodeling of cellular membranes precedes seasonally changing body temperature in a hibernator

Polyunsaturated fatty acids (PUFA) have a multitude of health effects. Their incorporation into membrane phospholipids (PL) is generally believed to depend directly on dietary influx. PL influence transmembrane protein activity and thus can compensate temperature effects; e.g. PL n-6 PUFA are thought to stabilize heart function at low body temperature (T(b)), whereas long chain (>C18) n-3 PUFA may boost oxidative capacity. We found substantial remodeling of membranes in free-living alpine marmots which was largely independent of direct dietary supply. Organ PL n-6 PUFA and n-6 to n-3 ratios were highest at onset and end of hibernation after rapid increases during a brief transitional period prior to hibernation. In contrast, longer chain PL n-3 PUFA content was low at end of summer but maximal at end of hibernation. After termination of hibernation in spring, these changes in PL composition were rapidly reversed. Our results demonstrate selective trafficking of PUFA within the body, probably governed by a circannual endogenous rhythm, as hibernating marmots were in winter burrows isolated for seven months from food and external cues signaling the approaching spring. High concentrations of PL n-6 PUFA throughout hibernation are in line with their hypothesized function of boosting SERCA 2a activity at low T(b). Furthermore, we found increasing rate of rewarming from torpor during winter indicating increasing oxidative capacity that could be explained by the accumulation of long-chain PL n-3 PUFA. It may serve to minimize the time necessary for rewarming despite the increasing temperature range to be covered, because rewarming is a period of highest metabolic rate and hence production of reactive oxygen species. Considering the importance of PUFA for health our results may have important biomedical implications, as seasonal changes of T(b) and associated remodeling of membranes are not restricted to hibernators but presumably common among endothermic organisms.