Development of LCA benchmarks for Austrian torrent control structures

The International Journal of Life Cycle Assessment - This study emerged from a research project that aimed to develop a Life Cycle Assessment (LCA) model for torrent control structures. This...     

Screening LCA of torrent control structures in Austria

Purpose

The purpose of this article is to find a suitable life cycle assessment (LCA) method to quantify the most important environmental burdens caused by construction processes of torrent control structures. To find these environmental burdens, 17 construction projects of the “Austrian Service for Torrent and Avalanche Control” (WLV) were analyzed using the “cradle to gate with options” LCA methodology (CEN, 2013).

Methods

This article explains an LCA methodology for the product stage and the construction process of torrent control structures following existing standards. The iterative approach of LCA methodology (ISO, 2006a) was used to record all important processes of the system and to supplement missing information. The LCA methodology has been developed from existing standards of the construction and product sector. Since the production of some construction materials takes place locally, the generic data, for Austria, was adapted. Wood inherent biogenic carbon and primary energy, used as raw material, are treated as materials inherent properties (CEN, 2014). The contribution of the various processes was reproduced by hotspot.

Results and discussion

Hotspots of the different stages are related to the construction materials used. The emissions and primary energy inputs in the product stage are clearly dominated by concrete and steel. If these two materials are used sparingly, the focus is on machine application and transportation. Depending on the selected scenarios, the smallest share of emissions, in relation to the total result of product and construction stage emitted by transport, is 3% and the maximum share is 69%. The greatest environmental impacts in the construction stage are caused by excavation work and transportation on-site. With an average of 4% in the construction stage, the transport of workers to the construction site cannot be neglected as is done in the building sector.

Conclusions

The conclusion of this study is that existing LCA models can be adapted to protective structures. In contrast to conventional buildings, the construction process and transportation are much more important and cannot be neglected. Shifting the hotspots to these processes requires specific calculation rules for that particular field. There is still a need for research to find a suitable functional unit and to develop a methodology for the use and end of life stage of these structures.

     

Phosphatidylinositol 3′-kinase signalling supports cell height in established epithelial monolayers

Cell–cell interactions influence epithelial morphogenesis through an interplay between cell adhesion, trafficking and the cytoskeleton. These cellular processes are coordinated, often by cell signals found at cell–cell contacts. One such contact-based signal is the phosphatidylinositol 3′-kinase (PI3-kinase; PI3K) pathway. PI3-kinase is best understood for its role in mitogenic signalling, where it regulates cell survival, proliferation and differentiation. Its precise morphogenetic impacts in epithelia are, in contrast, less well-understood. Using phosphoinositide-specific biosensors we confirmed that E-cadherin-based cell–cell contacts are enriched in PIP₃, the principal product of PI3-kinase. We then used pharmacologic inhibitors to assess the morphogenetic impact of PI3-kinase in MDCK and MCF7 monolayers. We found that inhibiting PI3-kinase caused a reduction in epithelial cell height that was reversible upon removal of the drugs. This was not attributable to changes in E-cadherin expression or homophilic adhesion. Nor were there detectable changes in cell polarity. While Myosin II has been implicated in regulating keratinocyte height, we found no effect of PI3-kinase inhibition on apparent Myosin II activity; nor did direct inhibition of Myosin II alter epithelial height. Instead, in pursuing signalling pathways downstream of PI3-kinase we found that blocking Rac signalling, but not mTOR, reduced epithelial cell height, as did PI3-kinase inhibition. Overall, our findings suggest that PI3-kinase exerts a major morphogenetic impact in simple cultured epithelia through preservation of cell height. This is independent of potential effects on adhesion or polarity, but may occur through PI3-kinase-stimulated Rac signaling.     

Multicomponent analysis of junctional movements regulated by myosin II isoforms at the epithelial zonula adherens

The zonula adherens (ZA) of epithelial cells is a site of cell-cell adhesion where cellular forces are exerted and resisted. Increasing evidence indicates that E-cadherin adhesion molecules at the ZA serve to sense force applied on the junctions and coordinate cytoskeletal responses to those forces. Efforts to understand the role that cadherins play in mechanotransduction have been limited by the lack of assays to measure the impact of forces on the ZA. In this study we used 4D imaging of GFP-tagged E-cadherin to analyse the movement of the ZA. Junctions in confluent epithelial monolayers displayed prominent movements oriented orthogonal (perpendicular) to the ZA itself. Two components were identified in these movements: a relatively slow unidirectional (translational) component that could be readily fitted by least-squares regression analysis, upon which were superimposed more rapid oscillatory movements. Myosin IIB was a dominant factor responsible for driving the unilateral translational movements. In contrast, frequency spectrum analysis revealed that depletion of Myosin IIA increased the power of the oscillatory movements. This implies that Myosin IIA may serve to dampen oscillatory movements of the ZA. This extends our recent analysis of Myosin II at the ZA to demonstrate that Myosin IIA and Myosin IIB make distinct contributions to junctional movement at the ZA.     

The topology of phosphatidylglycerol populations is essential for sustaining photosynthetic electron flow activities in thylakoid membranes

The transmembrane distribution of phosphatidylglycerol (PG) was determined in rightside-out (RO) and inside-out vesicles (IO) obtained by fragmentation of spinach thylakoids in a Yeda press, followed by partition in an aqueous dextran-polyethyleneglycol two-phase system. Using the phospholipase A(2) from porcine pancreas to digest selectively PG molecules in the outer monolayer (exposed to the incubation medium) of the membrane, we found the molar outside/inside distribution to be 70/30+/-5 in RO and 40/60+/-3 in IO. The transmembrane distribution of PG in IO was the opposite of that in intact thylakoids (molar ratio 58/42+/-3). The phospholipid population which sustained most of the uncoupled photosystem II electron flow activity was localized in the inner monolayer (exposed to the thylakoid lumen) of both thylakoid and RO membranes. In contrast, the activity in IO membranes was highly dependent on the PG population located in the outer monolayer. This finding brings the first direct demonstration of the dependence of the photosynthetic electron flow activity on the integrity of the inner topological pool of PG in the thylakoid membrane.     

ATP citrate lyase: cloning, heterologous expression and possible implication in root organic acid metabolism and excretion

In order to cope with phosphate deficiency, white lupin produces bottle‐brushed like roots, so‐called cluster or proteoid roots which are specialized in malate and citrate excretion. Young, developing cluster roots mainly excrete malate whereas mature cluster roots mainly release citrate. Mature proteoid roots excrete four to six times more carboxylates compared with juvenile proteoid roots. Using a cDNA‐amplified restriction fragment length polymorphism (AFLP) approach we identified a gene coding for a putative ATP‐citrate lyase (ACL) up‐regulated in young cluster roots. Cloning of the lupin ACL revealed that plant ACL is constituted by two polypeptides (ACLA and ACLB) encoded by two different genes. This contrasts with the animal ACL, constituted of one polypeptide which covers ACLA and ACLB. The ACL function of the two lupin gene products has been demonstrated by heterologous expression in yeast. Both subunits are required for ACL activity. In lupin cluster roots, our results suggest that ACL activity could be responsible for the switch between malate and citrate excretion in the different developmental stages of cluster roots. In primary roots of lupin and maize, ACL activity was positively correlated with malate exudation. These results show that ACL is implicated in root exudation of organic acids and hence plays a novel role in addition to lipid synthesis. Our results suggest that in addition to lipid biosynthesis, in plants, ACL is implicated in malate excretion.     

Effect of dimethyl sulfoxide on lysine production by a mutant ofBacillus subtilis with low homoserine dehydrogenase activity

SomeBacillus subtilis mutants with different levels of homoserine dehydrogenase were described. Strains that do not accumulate methionine have a high homoserine dehydrogenase activity. Low activity was detected in mutants where cell growth was completely inhibited by 0.7 mmol/L methionine. A low concentration of dimethyl sulfoxide had a stimulatory effect on lysine production by the methionine-sensitive mutant ofBacillus subtilis.     

Neighborly relations: cadherins and mechanotransduction

Cell–cell adhesions are sites where cells experience and resist tugging forces. It has long been postulated, but not directly tested, that cadherin adhesion molecules may serve in mechanotransduction at cell–cell contacts. In this issue, Le Duc et al. (2010. J. Cell Biol. doi: 10.1083/jcb.201001149) provide direct evidence that E-cadherin participates in a mechanosensing pathway that regulates the actomyosin cytoskeleton to modulate cell stiffness in response to pulling force.All of the cells in our body experience force: from the shear stress of blood flow experienced by the vascular endothelium to the tugging of other cells in skeletal muscle. Accordingly, cellular mechanisms exist to preserve tissue integrity by resisting this play of forces. Characteristically, these mechanisms involve adhesion receptors that are mechanically coupled to the cytoskeleton. However, these apparatuses do not simply support passive resistance. Instead, there has been great recent interest in the concept that adhesion receptors contribute to cell signaling pathways, which sense the magnitude of force exerted on cells and trigger cellular responses to those forces (Vogel and Sheetz, 2006). This notion is best established for integrin cell–matrix adhesion molecules in which well-characterized signaling pathways are clearly involved in mechanotransduction, which modifies focal adhesion size in response to force (Balaban et al., 2001) and may ultimately affect processes that range from stem cell differentiation (Engler et al., 2006) to tumor cell progression (Levental et al., 2009).At cell–cell contacts, classical cadherin adhesion molecules play major roles in morphogenesis and in the maintenance of tissue integrity. A role for cadherins in mechanotransduction has often been suspected (Schwartz and DeSimone, 2008) but not directly tested. One challenge in dissecting this problem is to distinguish responses principally elicited by the cadherin from juxtacrine events that occur when adhesion systems bring native cell surfaces into contact with one another. In this issue, Le Duc et al. circumvent this problem by using recombinant cadherin ligands, which contain the entire adhesive ectodomain, to test the capacity for a classical cadherin to participate in mechanosensing. The authors allowed magnetic beads coated with recombinant E-cadherin ectodomains to adhere to the dorsal surfaces of cultured cells. Classical cadherins engage in homophilic interactions via their ectodomains, and ligation of cellular cadherins by these immobilized ligands is a commonly used approach to generate adhesive contacts through E-cadherin alone. They used an oscillating magnetic field to twist the beads, thereby applying shear forces onto the sites of adhesion. By measuring the displacement of the beads in response to twisting stimuli, they could calculate changes in the local stiffness of the adhesive contact of each bead.Strikingly, they found that these adhesive contacts between the cadherin-coated beads and the cells stiffened in response to repetitive twisting force. The magnitude of stiffening increased with the magnitude of the applied force, which is evidence for the existence of a mechanism that could apparently measure the applied force and calibrate a proportionate cellular response. The use of E-cadherin as the ligand for homophilic engagement implied that the cellular cadherin was key to the force-sensing apparatus. This was further substantiated by the demonstration that the stiffening response did not occur when cadherin function was disrupted by removing extracellular calcium or adding a function-blocking antibody. Moreover, stiffening could not be elicited by beads coated with cadherin antibodies, suggesting that a native ligand was required rather than simple binding to the cellular cadherin ectodomain. Moreover, cell stiffening required an intact actomyosin cytoskeleton, implying that it reflected a cellular mechanical response to applied force. Overall, these findings indicate that E-cadherin engaged in homophilic interactions can serve to sense force and trigger a cellular response that involves the actin cytoskeleton, classical hallmarks of a mechanotransduction pathway (Fig. 1).Open in a separate windowFigure 1.E-cadherin mechanotransduction. Forces acting on surface E-cadherin molecules activate mechanosensing processes that lead to proportionate mechanical responses from cells. (1) In this model, E-cadherin engaged in homophilic adhesive interactions acts as a surface receptor for forces that tug on cells. (2) This induces an intracellular signaling cascade, which includes events such as alterations in protein conformation (notably α-catenin) and recruitment of proteins such as vinculin. (3 and 4) The subsequent mechanical response involves the actomyosin cytoskeleton (3), which can alter adhesion stiffness (4) by diverse processes such as changes in cortical organization and contractility. One potential outcome is that this cellular response will be felt as a pulling force by the neighboring cell that initiated the cascade, leading to cooperative interactions between the cells.What do we know of the molecular players in this E-cadherin–activated mechanotransduction pathway? A comprehensive answer to this question must ultimately encompass the signal transduction pathways that are activated by mechanical stimulation of E-cadherin and the elicited downstream cytoskeletal responses. Many different kinds of signaling events are implicated in other forms of mechanosensing, including the Src tyrosine kinase and ion channels (Vogel and Sheetz, 2006), which can be found at cell–cell contacts (Wang et al., 2006). Another molecular paradigm involves alterations in protein conformation in response to applied force, which thereby reveals novel sites for posttranslational modification or protein binding (del Rio et al., 2009). In this regard, a recent study identifies an apparently cryptic site in α-catenin that is sensitive to the cellular force generator myosin II (Yonemura et al., 2010). The study found that junctional staining with a monoclonal antibody directed to the central region of α-catenin was abolished in cells treated with the myosin II inhibitor blebbistatin, although α-catenin protein remained at cell–cell contacts. Notably, the epitope for this monoclonal antibody resides close to the region of α-catenin that can directly bind the actin regulator vinculin. Both Le Duc et al. (2010) and Yonemura et al. (2010) show that the recruitment of vinculin to cell–cell junctions is blebbistatin sensitive. Moreover, Le Duc et al. (2010) demonstrate that cellular stiffening in response to twisting force is reduced in vinculin-deficient cells. This suggests the attractive hypothesis that transmission of force to α-catenin that is incorporated into the E-cadherin complex may alter its conformation and capacity to interact with binding partners such as vinculin. This notion warrants more detailed analysis; however, if experience with integrin mechanotransduction is any guide (Vogel and Sheetz, 2006; Schwartz and DeSimone, 2008), force-induced conformational change in proteins such as α-catenin are likely to be but one part of a more complex network of signal transduction mechanisms.The force-dependent recruitment of vinculin also provides a potential mechanism to coordinate a cytoskeletal response to E-cadherin mechanosensing. Although long known to concentrate at the zonula adherens (as well for its better-known localization in focal adhesions), the precise role that vinculin plays in cell–cell interactions remains enigmatic. Nonetheless, depletion of vinculin reduces cell–cell adhesion and disrupts the integrity of epithelial cell–cell junctions (Peng et al., 2010). Vinculin depletion also perturbs the junctional actin cytoskeleton (Maddugoda et al., 2007), and vinculin has the capacity to bind actin filaments and diverse actin regulators, thereby influencing both filament bundling and dynamics (Le Clainche et al., 2010). However, vinculin is unlikely to be the sole mediator of the cytoskeletal response. Many cytoskeletal regulators act at E-cadherin cell–cell junctions to control actin filament dynamics and organization. A particularly interesting case is nonmuscle myosin II, which was implicated as the dominant cellular force generator in recent studies (Le Duc et al., 2010; Yonemura et al., 2010). However, the contribution of myosin II is likely to be complex, as myosin II is also necessary for the cytoskeletal response to force (Le Duc et al., 2010). Moreover, there is emerging evidence for both contractile and noncontractile functions for myosin II (Choi et al., 2008). Also, the myosin II A and B isoforms can have distinct contributions to E-cadherin clustering and apical actin regulation (Smutny et al., 2010). Thus, myosin II may have several contributions to cadherin mechanotransduction.Finally, what functions might be served by cadherin-based mechanosensing? One possibility is that local stiffening, and perhaps the cytoskeletal response more broadly, might provide a mechanism to strengthen adhesions against potentially disruptive forces. This notion is supported by a recent study by Liu et al. (2010), who analyzed forces at the contacts between pairs of cells grown on micropatterned substrata. Force vectors oriented approximately perpendicular to the cell–cell contacts could be extracted from their data, and strikingly, the authors identified a linear relationship between the magnitude of the forces and the size of the contacts. This appeared to reflect the coordinated action of Rho- and Rac-based signaling pathways. They proposed that force-dependent growth of adhesions may be a mechanism to reduce stress at the contacts and thus preserve their integrity. In addition, it is interesting to consider the possibility that mechanosensing through E-cadherin could provide a mechanism for cells to assess the mechanical properties of their neighboring cells. Cells appear to use integrin-based mechanosensing to assess the stiffness of their surrounding matrix (Schwartz and DeSimone, 2008; Levental et al., 2009). Their ability to use myosin II–based contractility to pull on adhesion sites is likely critical for cells to assess stiffness of their surroundings. However, an important difference between cell–matrix and cell–cell mechanosensing is that although in the former case the environment is passive, in the latter case, it is active (i.e., neighboring cells can pull back). Is the mechanical response of a neighboring cell an important parameter in cadherin-based cell–cell recognition? Clearly, then, the new work of Le Duc et al. (2010) opens many new avenues for understanding the role of mechanosensing in cadherin biology and tissue organization.