Non-neoplastic portal vein thrombosis in HCV cirrhosis : the weight of inflammation on a fragile hemostatic balance

The historical paradigm of spontaneous bleeding tendency of cirrhosis as consequence of an intrinsic coagulopathy has faded in the last years (1). This theory has been first challenged by milestones studies of Tripodi et al. demonstrating normal thrombin generation by plasma from cirrhotic patients and, consequently, the need of more comprehensive coagulation tests to evaluate hemostatic changes in advanced chronic liver disease (2). This is coherent with the clinical observation of thrombotic events occurring among cirrhotic patients, despite altered prothrombin- and partial thromboplastin-time, which are inadequate to evaluate the profound changes in balance between pro- and anti-coagulant factors alongside the disruption of liver function (3,4). Moreover, plasma from patients with cirrhosis shows a pro-coagulant imbalance alongside disease severity, as expressed by the resistance to the action of thrombomodulin, a strong anti-coagulant factor (5)

Coagulation, Microenvironment and Liver Fibrosis

Fibrosis is the main consequence of any kind of chronic liver damage. Coagulation and thrombin generation are crucial in the physiological response to tissue injury; however, the inappropriate and uncontrolled activation of coagulation cascade may lead to fibrosis development due to the involvement of several cellular types and biochemical pathways in response to thrombin generation. In the liver, hepatic stellate cells and sinusoidal endothelial cells orchestrate fibrogenic response to chronic damage. Thrombin interacts with these cytotypes mainly through protease-activated receptors (PARs), which are expressed by endothelium, platelets and hepatic stellate cells. This review focuses on the impact of coagulation in liver fibrogenesis, describes receptors and pathways involved and explores the potential antifibrotic properties of drugs active in hemostasis in studies with cells, animal models of liver damage and humans

Multifaceted SlyD from Helicobacter pylori: implication in [NiFe] hydrogenase maturation

SlyD belongs to the FK506-binding protein (FKBP) family with both peptidylprolyl isomerase (PPIase) and chaperone activities, and is considered to be a ubiquitous cytosolic protein-folding facilitator in bacteria. It possesses a histidine- and cysteine-rich C-terminus binding to selected divalent metal ions (e.g., Ni2+, Zn2+), which is important for its involvement in the maturation processes of metalloenzymes. We have determined the solution structure of C-terminus-truncated SlyD from Helicobacter pylori (HpSlyDΔC). HpSlyDΔC folds into two well-separated, orientation-independent domains: the PPIase-active FKBP domain and the chaperone-active insert-in-flap (IF) domain. The FKBP domain consists of a four-stranded antiparallel β-sheet with an α-helix on one side, whereas the IF domain folds into a four-stranded antiparallel β-sheet accompanied by a short α-helix. Intact H. pylori SlyD binds both Ni2+ and Zn2+, with dissociation constants of 2.74 and 3.79 μM respectively. Intriguingly, binding of Ni2+ instead of Zn2+ induces protein conformational changes around the active sites of the FKBP domain, implicating a regulatory role of nickel. The twin-arginine translocation (Tat) signal peptide from the small subunit of [NiFe] hydrogenase (HydA) binds the protein at the IF domain. Nickel binding and the recognition of the Tat signal peptide by the protein suggest that SlyD participates in [NiFe] hydrogenase maturation processes

Long-Term IGF-I Exposure Decreases Autophagy and Cell Viability

A reduction in IGF-I signaling has been found to increase lifespan in multiple organisms despite the fact that IGF-I is a trophic factor for many cell types and has been found to have protective effects against multiple forms of damage in acute settings. The increase in longevity seen in response to reduced IGF-I signaling suggests that there may be differences between the acute and chronic impact of IGF-I signaling. We have examined the possibility that long-term stimulation with IGF-I may have a negative impact at the cellular level using quiescent human fibroblasts. We find that fibroblast cells exposed to IGF-I for 14 days have reduced long-term viability as judged by colony forming assays, which is accompanied by an accumulation of senescent cells. In addition we observe an accumulation of cells with depolarized mitochondria and a reduction in autophagy in the long-term IGF-I treated cultures. An examination of mice with reduced IGF-I levels reveals evidence of enhanced autophagy and fibroblast cells derived from these mice have a larger mitochondrial mass relative to controls indicating that changes in mitochondrial turnover occurs in animals with reduced IGF-I. The results indicate that chronic IGF-I stimulation leads to mitochondrial dysfunction and reduced cell viability

Outer membrane protein folding from an energy landscape perspective

The cell envelope is essential for the survival of Gram-negative bacteria. This specialised membrane is densely packed with outer membrane proteins (OMPs), which perform a variety of functions. How OMPs fold into this crowded environment remains an open question. Here, we review current knowledge about OFMP folding mechanisms in vitro and discuss how the need to fold to a stable native state has shaped their folding energy landscapes. We also highlight the role of chaperones and the β-barrel assembly machinery (BAM) in assisting OMP folding in vivo and discuss proposed mechanisms by which this fascinating machinery may catalyse OMP folding

LRRK2 at the interface of autophagosomes, endosomes and lysosomes

Over the past 20 years, substantial progress has been made in identifying the underlying genetics of Parkinson’s disease (PD). Of the known genes, LRRK2 is a major genetic contributor to PD. However, the exact function of LRRK2 remains to be elucidated. In this review, we discuss how familial forms of PD have led us to hypothesize that alterations in endomembrane trafficking play a role in the pathobiology of PD. We will discuss the major observations that have been made to elucidate the role of LRRK2 in particular, including LRRK2 animal models and high-throughput proteomics approaches. Taken together, these studies strongly support a role of LRRK2 in vesicular dynamics. We also propose that targeting these pathways may not only be beneficial for developing therapeutics for LRRK2-driven PD, but also for other familial and sporadic cases

Effects of Periplasmic Chaperones and Membrane Thickness on BamA-Catalyzed Outer-Membrane Protein Folding

The biogenesis of outer-membrane proteins (OMPs) in gram-negative bacteria involves delivery by periplasmic chaperones to the β-barrel assembly machinery (BAM), which catalyzes OMP insertion into the outer membrane. Here, we examine the effects of membrane thickness, the Escherichia coli periplasmic chaperones Skp and SurA, and BamA, the central subunit of the BAM complex, on the folding kinetics of a model OMP (tOmpA) using fluorescence spectroscopy, native mass spectrometry, and molecular dynamics simulations. We show that prefolded BamA promotes the release of tOmpA from Skp despite the nM affinity of the Skp:tOmpA complex. This activity is located in the BamA β-barrel domain, but is greater when full-length BamA is present, indicating that both the β-barrel and polypeptide transport-associated (POTRA) domains are required for maximal activity. By contrast, SurA is unable to release tOmpA from Skp, providing direct evidence against a sequential chaperone model. By varying lipid acyl chain length in synthetic liposomes we show that BamA has a greater catalytic effect on tOmpA folding in thicker bilayers, suggesting that BAM catalysis involves lowering of the kinetic barrier imposed by the hydrophobic thickness of the membrane. Consistent with this, molecular dynamics simulations reveal that increases in membrane thinning/disorder by the transmembrane domain of BamA is greatest in thicker bilayers. Finally, we demonstrate that cross-linking of the BamA barrel does not affect tOmpA folding kinetics in 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) liposomes, suggesting that lateral gating of the BamA barrel and/or hybrid barrel formation is not required, at least for the assembly of a small 8-stranded OMP in vitro