Rupture strength of living cell monolayers.

To fulfil their function, epithelial tissues need to sustain mechanical stresses and avoid rupture. Although rupture is usually undesired, it is central to some developmental processes, for example, blastocoel formation. Nonetheless, little is known about tissue rupture because it is a multiscale phenomenon that necessitates comprehension of the interplay between mechanical forces and biological processes at the molecular and cellular scales.

Left ventricle endomyocardial fibrosis: a case report.

Background: Endomyocardial fibrosis is a grim disease. It is the most common restrictive cardiomyopathy worldwide, but the exact etiology and pathogenesis both remain unknown. Endomyocardial fibrosis is recurrently associated with chronic eosinophilia and probable dietary, environmental, and infectious factors, which contribute not only to the onset of the disease (an inflammatory process) but also to its progression and maintenance (endomyocardial damage and scar formation).

Spindle reorientation in response to mechanical stress is an emergent property of the spindle positioning mechanisms.

Proper orientation of the mitotic spindle plays a crucial role in embryos, during tissue development, and in adults, where it functions to dissipate mechanical stress to maintain tissue integrity and homeostasis. While mitotic spindles have been shown to reorient in response to external mechanical stresses, the subcellular cues that mediate spindle reorientation remain unclear. Here, we used a combination of optogenetics and computational modeling to investigate how mitotic spindles respond to inhomogeneous tension within the actomyosin cortex.

Chemical Priming by Isothiocyanates Protects Against Intoxication by Products of the Mustard Oil Bomb.

In Brassicaceae, tissue damage triggers the mustard oil bomb i.e., activates the degradation of glucosinolates by myrosinases leading to a rapid accumulation of isothiocyanates at the site of damage.

SPIN90 associates with mDia1 and the Arp2/3 complex to regulate cortical actin organization.

Cell shape is controlled by the submembranous cortex, an actomyosin network mainly generated by two actin nucleators: the Arp2/3 complex and the formin mDia1. Changes in relative nucleator activity may alter cortical organization, mechanics and cell shape. Here we investigate how nucleation-promoting factors mediate interactions between nucleators.

Reactive electrophilic oxylipins trigger a heat stress-like response through HSFA1 transcription factors.

Abiotic and biotic stresses are often characterized by an induction of reactive electrophile species (RES) such as the jasmonate 12-oxo-phytodienoic acid (OPDA) or the structurally related phytoprostanes. Previously, RES oxylipins have been shown massively to induce heat-shock-response (HSR) genes including HSP101 chaperones. Moreover, jasmonates have been reported to play a role in basal thermotolerance.

Sterol regulatory element binding protein-dependent regulation of lipid synthesis supports cell survival and tumor growth.

Background: Regulation of lipid metabolism via activation of sterol regulatory element binding proteins (SREBPs) has emerged as an important function of the Akt/mTORC1 signaling axis. Although the contribution of dysregulated Akt/mTORC1 signaling to cancer has been investigated extensively and altered lipid metabolism is observed in many tumors, the exact role of SREBPs in the control of biosynthetic processes required for Akt-dependent cell growth and their contribution to tumorigenesis remains unclear.

Results: We first investigated the effects of loss of SREBP function in non-transformed cells.

Antagonism between FOXO and MYC Regulates Cellular Powerhouse.

Alterations in cellular metabolism are a key feature of the transformed phenotype. Enhanced macromolecule synthesis is a prerequisite for rapid proliferation but may also contribute to induction of angiogenesis, metastasis formation, and tumor progression, thereby leading to a poorer clinical outcome. Metabolic adaptations enable cancer cells to survive in suboptimal growth conditions, such as the limited supply of nutrient and oxygen often found in the tumor microenvironment.

FOXO3a regulates reactive oxygen metabolism by inhibiting mitochondrial gene expression.

Forkhead transcription factors of the O class (FOXOs) are important targets of the phosphatidylinositol 3-kinase/Akt pathway, and are key regulators of the cell cycle, apoptosis and response to oxidative stress. FOXOs have been shown to have tumour suppressor function and are important for stem cell maintenance. We have performed a detailed analysis of the transcriptional programme induced in response to Forkhead-box protein O3a (FOXO3a) activation.

A role for the cleaved cytoplasmic domain of E-cadherin in the nucleus.

Cell-cell contacts play a vital role in intracellular signaling, although the molecular mechanisms of these signaling pathways are not fully understood. E-cadherin, an important mediator of cell-cell adhesions, has been shown to be cleaved by gamma-secretase. This cleavage releases a fragment of E-cadherin, E-cadherin C-terminal fragment 2 (E-cad/CTF2), into the cytosol.

p32 is a novel mammalian Lgl binding protein that enhances the activity of protein kinase Czeta and regulates cell polarity.

Lgl (lethal giant larvae) plays an important role in cell polarity. Atypical protein kinase C (aPKC) binds to and phosphorylates Lgl, and the phosphorylation negatively regulates Lgl activity. In this study, we identify p32 as a novel Lgl binding protein that directly binds to a domain on mammalian Lgl2 (mLgl2), which contains the aPKC phosphorylation site.

Casein kinase 1 is a novel negative regulator of E-cadherin-based cell-cell contacts.

Cadherins are the most crucial membrane proteins for the formation of tight and compact cell-cell contacts. Cadherin-based cell-cell adhesions are dynamically established and/or disrupted during various physiological and pathological processes. However, the molecular mechanisms that regulate cell-cell contacts are not fully understood.

The transcriptional repressor Glis2 is a novel binding partner for p120 catenin.

In epithelial cells, p120 catenin (p120) localizes at cell-cell contacts and regulates adhesive function of the cadherin complex. In addition, p120 has been reported to localize in the nucleus, although the nuclear function of p120 is not fully understood. Here, we report the identification of Gli-similar 2 (Glis2) as a novel binding protein for p120.

TNF revisited: TNF-independent antitumor activity in sera of mice sensitized with Propionibacterium acnes and challenged with lipopolysaccharide.

Sera of mice sensitized with bacteria and subsequently challenged with lipopolysaccharide promote hemorrhagic necrosis of tumors in vivo and display cytotoxic activity against tumor cells in vitro, which has been attributed to the induction of tumor necrosis factor (TNF). Here, we describe the induction of a previously unrecognized antitumor activity in such sera, which is distinct from TNF but displays tumor-specific cytocidal activity in vitro as well as potent tumor-regressing activity in vivo. Biochemical analysis of this activity yielded a molecular mass of approximately 150 kDa, closely resembling a novel tumoricidal factor of murine macrophages (Mphi) termed MTC 170 (Mphi tumor cytotoxin, approximate molecular mass 170 kDa), which we have previously proposed to constitute a major effector pathway for the destruction of tumor cells by activated Mphi.

Involvement of protein kinase A in the induction of arginase in murine bone marrow-derived macrophages.

Arginase is induced in bone marrow-derived macrophages by agents that increase the intracellular concentrations of cAMP (Br-cAMP, prostaglandin E2) and, in their presence, the LPS induced NO synthesis is down regulated. Moreover, interleukin 10 which induces arginase in macrophages is able to increase the cAMP-dependent protein kinase A activity. In contrast, suppressors of NOS synthesis like protein kinase C inhibitors and calmodulin antagonists (W7), or NO activators (A23187) have no effect on the induction of arginase by LPS.

Involvement of reactive oxygen species in phospholipase A2 activation: inhibition of protein tyrosine phosphatases and activation of protein kinases.

Activators of PKC in combination with vanadate induce massive formation of reactive oxygen species. The formation of ROS leads to suppression of protein tyrosine phosphatase activity and consequently to enhanced protein tyrosine phosphorylation. This culminates in the activation of the MAP-K cascade and of PLA2 (Scheme 1).

A role for reactive oxygen species in zymosan and beta-glucan induced protein tyrosine phosphorylation and phospholipase A2 activation in murine macrophages.

Previously we have shown that reactive oxygen species (ROS) formation induced by phorbol ester in association with vanadate is essential for protein tyrosine phosphorylation and phospholipase A2 (PLA2) activation. Here we show that the interaction of beta-glucan particles (glucanp) or zymosan with complement receptor type 3 (CR3) leads, when associated with vanadate, to a cascade of reactions culminating in PLA2 activation. Vanadate + zymosan (or glucanp) markedly enhance protein tyrosine phosphorylation in bone marrow derived macrophages (BMMs), whereas neither of the agents alone has any effect.

Reactive oxygen species mediate phorbol ester-regulated tyrosine phosphorylation and phospholipase A2 activation: potentiation by vanadate.

We have previously shown that vanadate potentiates the activating effect of phorbol ester (TPA) on cellular phospholipase A2 (PLA2) in a pathway dependent on the formation of reactive oxygen species (ROS). Here we evaluate the chain of enzymes (protein kinases and phosphatases) that participate in this process. Treatment of macrophages with vanadate plus TPA led to activation of protein kinase C (PKC) and NADPH oxidase (O2- generation in intact cells), massive cellular protein tyrosine phosphorylation, suppression of protein tyrosine phosphatase (PTP) activity and a sustained activation of protein tyrosine kinase (PTK) and myelin basic protein kinase activity (the latter three enzyme activities were assessed in cell lysates).

Differential modulation of the effects of lipopolysaccharide on macrophages by a major outer membrane protein of Proteus mirabilis.

We previously showed that a major protein isolated from purified cell walls of Proteus mirabilis (39-kDa protein) is a strong modulator of the specific immune responses to LPS from this bacterium in mice. When mixed with LPS before immunization, this protein enhances T cell-dependent, IgG antibody-producing cell responses specific for LPS. Furthermore, complexes of the 39-kDa protein with LPS drastically inhibit the production of oxygen radicals by murine macrophages activated with LPS, as measured in a chemiluminescence assay.

A fast and sensitive method for detection of phospholipid-binding proteins on nitrocellulose membranes.

We describe a sensitive new method for specific detection and quantitation of phospholipid-binding proteins using two purified lipocortins as model proteins. The method consists of blotting proteins to nitrocellulose membranes followed by incubation with radiolabeled phospholipids and autoradiographic detection of bound phospholipids. It allows specific detection of phospholipid binding proteins to a threshold of about 100 ng/spot and their quantitation up to several micrograms as well as rapid analysis of specific phospholipid binding properties of individual proteins.

Reactive oxygen species are involved in the activation of cellular phospholipase A2.

Vanadate (V) potentiated (4- to 10-fold) the activation of cellular phospholipase A2 (PLA2) induced by H2O2 (H), a phorbol ester (T), a Ca(2+)-ionophore (A) and opsonized zymosan in macrophages. V+H induced in intact cells the activation and translocation of PLA2 and protein kinase C (PKC) to the plasma membrane. V+H and V+T+A induced strong chemiluminescence (CL) which was abrogated by a specific NADPH oxidase inhibitor diphenylene iodonium (DPI).

Characterization and partial purification of a high molecular weight tumoricidal activity secreted by murine bone marrow macrophages.

Cultured murine bone marrow-derived macrophage (BMM phi) can be induced to secrete tumoricidal activity in vitro when activated with recombinant IFN-gamma and bacterial lipopolysaccharide (LPS). We have analyzed this activity for tumor specificity, relationship to tumor necrosis factor-alpha (TNF-alpha), serine proteases, and reactive nitrogen intermediates, and partially purified this activity by high pressure liquid chromatography. Cytolytic activity was recovered in conditioned culture supernatants of serum-free cultivated BMM phi treated with a combination of IFN-gamma and LPS but was not inducible by either stimulant alone.

Human macrophages secrete a tumoricidal activity distinct from tumour necrosis factor-alpha and reactive nitrogen intermediates.

Human macrophages, differentiated in vitro from blood monocytes, can be induced to secrete tumouricidal activity when activated by combined treatment with recombinant interferon gamma and bacterial lipopolysaccharide. We have analysed conditioned culture supernatants of activated human monocytes and in vitro differentiated macrophages cultivated under serum-free conditions for cytolytic activity against a TNF alpha-insensitive human tumour cell line and characterized this activity with respect to its relationship to TNF alpha and reactive nitrogen intermediates. Cytolytic activity was recovered in the high molecular weight fraction of culture supernatants conditioned by terminally differentiated macrophages, whereas conditioned culture supernatants of freshly isolated blood monocytes, processed under identical conditions, were devoid of significant cytolytic activity.

Synergistic effect of magnesium and calcium ions in the activation of phospholipase A2 of liver macrophages.

In cell-free extracts of rat liver macrophages (Kupffer cells) phospholipase A2 was found to be strongly activated at free Ca2+ concentrations from 100 nM to 1 microM in the presence of 4 mM free Mg2+. This is within the range of intracellular free Ca2+ reported for basal and various stimulated conditions, respectively. Ca2+ alone increased phospholipase A2 activity at high Ca2+ concentrations (1 mM) whereas Mg2+ alone had only little stimulatory effect.

Arachidonic acid release by basophilic leukemia cells and macrophages stimulated by Ca2+ ionophores, antigen and diacylglycerol: essential role for protein kinase C and prevention by glucocorticosteroids.

The role of protein kinase C in phospholipase A2 (PLA2) activation in rat basophilic leukemia cells (RBL-2H3) and macrophages was investigated. 12-O-Tetradecanoyl phorbol 13-acetate (TPA) doubled ionomycin-induced PLA2 activity, assessed by [3H]arachidonate release. Protein kinase C inhibitors, staurosporine and K252a (100 nM) or H-7 (15 micrograms/ml) inhibited ionomycin-stimulation of PLA2 activity by 62, 75 and 80%, respectively.