Immunomodulatory benefits of cyclosporine A in inflammatory bowel disease

Etiopathogenesis of mucosal inflammation in inflammatory bowel disease remains a complex and enigmatic field; various factors (genetic, environmental and microbial) trigger an event that activates intestinal immune and nonimmune systems culminating in inflammation and tissue injury. Specifically, both innate and adaptive immune systems seem to play important roles in the pathophysiology of this disease. Cyclosporine A represents a macrolide immune modulator with primary inhibitory effects on T helper lymphocyte production of interleukin-2, and other cytokines leading to altered T-lymphocyte and B-lymphocyte function. The diversity of its therapeutic outcome reported in inflammatory bowel disease may be due to the intricate immuno-pathogenic profile of the disease and the variety of the applied dose-dependent courses of therapy. Cyclosporine A exerts additional actions on other components of the inflammatory infiltrate, including neutrophils and mast cells, thereby appearing to be a multi-dynamic therapeutic approach, although with potential drawbacks, that may be applied alone or combined with other immunomodulatory agents in inflammatory bowel disease patients. Because cyclosporine A induces apoptosis of T-lymphocytes responsible for perpetuation of the chronic inflammatory process in the disease with potential tumorigenic effect, it may exert a further inhibitory effect on cancer development in inflammatory bowel disease patients, and can be combined with other relative agents, such as rapamycin, which also promotes T-lymphocyte apoptosis. Therefore, recently established multifactorial action of cyclosporine A in relation to the pathogenesis of the disease can open new horizons for prospective, controlled trials in large cohorts, aiming to emphasize cyclosporine A's potential.     

Gastrointestinal immune system and brain dialogue implicated in neuroinflammatory and neurodegenerative diseases

A common characteristic of the central nervous system (CNS) neurodegenerative disorders is neuroinflammation, marked by augmented numbers of activated and primed microglia, increased inflammatory cytokines and decreased anti-inflammatory molecules. CNS neuroinflammation is a critical component in the progression of several neurodegenerative diseases which sensitize the brain to produce an exaggerated response to immune stimuli in the periphery. Neuroinflammation might initiate from the periphery and peripheral conditions through disrupted blood-brain barrier powerfully influence various brain pathologies. Gastrointestinal tract (GIT) represents a vulnerable area through which pathogens influence the brain and induce CNS neuroinflammation. The pathogens may access the CNS through blood, the nasal olfactory pathways and the GIT. Potential GI pathogens, such as Helicobacter pylori, induce humoral and cellular immune responses that, owing to the sharing of homologous epitopes (molecular mimicry), cross-react with CNS components thereby contributing and possibly perpetuating neural tissue damage. GIT is strictly connected to the CNS and a bi-directional communication exists between them. The brain is involved in regulating the immune and gut system. Conversely, limited attention has been paid on the GIT role in the development and regulation of the CNS autoimmune diseases. The GIT is the primary immune organ with specialized immunoregulatory and anti-inflammatory functions, represented by the gastrointestinal immune system (GIS). This review focuses on the potential GIS and brain dialogue implicated in neurodegenerative diseases. Gaining a better understanding of the relationship between GIS and CNS could provide an insight on the pathogenesis and therapeutic strategies of these disorders.     

The association between Helicobacter pylori infection and insulin resistance: a systematic review

Background: Helicobacter pylori infection has been associated with diverse extradigestive morbidity, including insulin resistance (IR) syndrome. The aim of this systematic review was to summarize the epidemiologic evidence concerning the association between H. pylori infection and IR quantitative indexes. Materials and Methods: A computerized literature search in PubMed electronic databases and Cochrane Central Register of Controlled Trials was performed. Results: Nine studies reporting data on 2120 participants were finally eligible for this systematic review. Seven of them were cross‐sectional studies and two were nonrandomized, open‐label, controlled trials investigating the effect of H. pylori eradication on IR. Homeostatic model of assessment insulin resistance (HOMA‐IR) was used in all studies to quantify IR. There seems to be a trend toward a positive association between H. pylori infection and HOMA‐IR, strengthened by regression analysis in one study. However, there was significant heterogeneity between studies regarding the method(s) of H. pylori infection diagnosis based on and the study populations. The studies for the effect of H. pylori eradication on HOMA‐IR revealed conflicting results. Conclusions: Although data seem to indicate a potential association between H. pylori infection and IR, further studies are needed to strengthen this association and to clarify whether there is a causative link between them. If a causal link is confirmed in the future, this may have a major impact on the pathophysiology and management of IR syndrome, including type 2 diabetes mellitus and nonalcoholic fatty liver disease.     

Impact of reactive oxygen species generation on Helicobacter pylori-related extragastric diseases: a hypothesis

Helicobacter pylori (H. pylori) induces reactive oxygen species (ROS) production that contribute to pathogenesis of a variety of H. pylori-related gastric diseases, as shown in animal and human studies. Helicobacter pylori infection is also associated with variety of systemic extragastric diseases in which H. pylori-related ROS production might also be involved in the pathogenesis of these systemic conditions. We proposed that Hp-related ROS may play a crucial role in the pathophysiology of Hp-related systemic diseases including Alzheimer’s disease, multiple sclerosis, glaucoma and other relative neurodegenerative diseases, thereby suggesting introduction of relative ROS scavengers as therapeutic strategies against these diseases which are among the leading causes of disability and are associated with a large public health global burden. Moreover, we postulated that H. pylori-related ROS might also be involved in the pathogenesis of extragastric common malignancies, thereby suggesting that H. pylori eradication might inhibit the development or delay the progression of aforementioned diseases. However, large-scale future studies are warranted to elucidate the proposed pathophysiological mechanisms, including H. pylori-related ROS, involved in H. pylori-associated systemic and malignant conditions.     

Flexible open conformation of the AP-3 complex explains its role in cargo recruitment at the Golgi

Vesicle formation at endomembranes requires the selective concentration of cargo by coat proteins. Conserved adapter protein complexes at the Golgi (AP-3), the endosome (AP-1), or the plasma membrane (AP-2) with their conserved core domain and flexible ear domains mediate this function. These complexes also rely on the small GTPase Arf1 and/or specific phosphoinositides for membrane binding. The structural details that influence these processes, however, are still poorly understood. Here we present cryo-EM structures of the full-length stable 300 kDa yeast AP-3 complex. The structures reveal that AP-3 adopts an open conformation in solution, comparable to the membrane-bound conformations of AP-1 or AP-2. This open conformation appears to be far more flexible than AP-1 or AP-2, resulting in compact, intermediate, and stretched subconformations. Mass spectrometrical analysis of the cross-linked AP-3 complex further indicates that the ear domains are flexibly attached to the surface of the complex. Using biochemical reconstitution assays, we also show that efficient AP-3 recruitment to the membrane depends primarily on cargo binding. Once bound to cargo, AP-3 clustered and immobilized cargo molecules, as revealed by single-molecule imaging on polymer-supported membranes. We conclude that its flexible open state may enable AP-3 to bind and collect cargo at the Golgi and could thus allow coordinated vesicle formation at the trans-Golgi upon Arf1 activation.

Eukaryotic cells have membrane-enclosed organelles, which carry out specialized functions, including compartmentalized biochemical reactions, metabolic channeling, and regulated signaling, inside a single cell. The transport of proteins, lipids, and other molecules between these organelles is mediated largely by small vesicular carriers that bud off at a donor compartment and fuse with the target membrane to deliver their cargo. The generation of these vesicles has been subject to extensive studies and has led to the identification of numerous coat proteins that are required for their formation at different sites (1, 2). Coat proteins can be monomers, but in most cases, they consist of several proteins, which form a heteromeric complex.Heterotetrameric adapter protein (AP) complexes are required at several endomembranes for cargo binding. Five well-conserved AP-complexes with differing functions have been identified in mammalian cells, named AP-1–AP-5, of which three (AP-1–AP-3) are conserved from yeast to human (3, 4). The three conserved adapter complexes function at different membranes along the endomembrane system. AP-1 is required for cargo transport between the Golgi and the endosome, AP-2 is required for cargo recognition and transport between the plasma membrane and the early endosome. Finally, AP-3 functions between the trans Golgi and the vacuole in yeast, whereas mammalian AP-3 localizes to a tubular endosomal compartment, in addition to or instead of the TGN (2, 5, 6).Each of the complexes consists of four different subunits: two large adaptins (named α−ζ and β1-5 respectively), a medium-sized subunit (μ1-5), and a small subunit (σ1-5). While μ- and σ-subunits together with the N-termini of the large adaptins build the membrane-binding core of the complex, the C-termini of both adaptins contain the ear domains, which are connected via flexible linkers (2). The recruitment of these complexes to membranes is not entirely conserved. They all require cargo binding, yet AP-1 binds Arf1-GTP with the γ and β1 subunit and phosphatidylinositol-4-phosphate (PI4P) via a proposed conserved site on its γ-subunit (7, 8). AP-2, on the other hand, interacts with PI(4,5)P₂ at the plasma membrane via its α, β2, and μ2 subunits (9, 10, 11).Several studies have uncovered how AP-3 functions in cargo sorting in yeast. AP-3 recognizes cargo at the Golgi via two sorting motifs in the cytosolic segments of membrane proteins: a Yxxφ sorting motif, as found in yeast in the SNARE Nyv1 or the Yck3 casein kinase, which binds to a site in μ3, as shown for mammalian AP-3, which is similar to μ2 in AP-2 (12, 13, 14), and dileucine motifs as found in the yeast SNARE Vam3 or the alkaline phosphatase Pho8, potentially also at a site comparable to AP-1 and AP-2 (15, 16). Unlike AP-1 and AP-2-coated vesicles, which depend on clathrin for their formation (2, 17), AP-3 vesicle formation in yeast does not require clathrin or the HOPS subunit Vps41 (18), yet Vps41 is required at the vacuole to bind AP-3 vesicles prior to fusion (19, 20, 21, 22). Studies in metazoan cells revealed that Vps41 and AP-3 function in regulated secretion (23, 24, 25), and AP-3 is required for biogenesis of lysosome-related organelles (26). This suggests that the AP-3 complex has features that are quite different from AP-1 and AP-2 complexes, which cooperate with clathrin in vesicle formation (2).Among the three conserved AP complexes, the function of the AP-3 complex is the least understood. Arf1 is necessary for efficient AP-3 vesicle generation in mammalian cells and shows a direct interaction with the β3 and δ subunits of AP-3 (27, 28). In addition, in vitro experiments on mammalian AP-3 using liposomes or enriched Golgi membranes suggest Arf1 as an important factor in AP-3 recruitment, whereas acidic lipids do not have a major effect, in contrast to what was found for AP-1 and AP-2 (7, 11, 29, 30). Another study showed that membrane recruitment of AP-3 depends on the recognition of sorting signals in cargo tails and PI3P (31), similar to AP-1 recruitment via cargo tails, Arf1 and PI4P (32).However, since AP-1 and AP-3 are both recruited to the trans-Golgi network (TGN) in yeast (33), the mechanism of their recruitment likely differs. Even though Arf1 is required, yeast AP-3 seems to be present at the TGN before the arrival of the Arf1 guanine nucleotide exchange factor (GEF) Sec7 (33). This implies the necessity for additional factors at the TGN and a distinct mechanism to allow for spatial and temporal separation of AP-1 and AP-3 recruitment to membranes. Structural data on mammalian AP-1 and AP-2 “core” complexes without the hinge and ear domains of their large subunits revealed that both exist in at least two very defined conformational states: a “closed” cytosolic state, where the cargo-binding sites are buried within the complex, and an “open” state, where the same sites are available to bind cargo (7, 8, 10, 34, 35). Binding of Arf1 to AP-1 or PI(4,5)P₂ in case of AP-2 induces a conformational change in the complexes that enables them to bind cargo molecules carrying a conserved acidic di-Leucine or a Tyrosine-based motif, as for all three AP complexes in yeast (8, 34). Additional conformational states and intermediates have been reported for both, mammalian AP-1 and AP-2 complex. AP-1, for example, can be hijacked by the human immunodeficiency virus-1 (HIV-1) proteins viral protein u (Vpu) and negative factor (Nef), resulting in a hyper-open conformation of AP-1 (36, 37).An emerging model over the past years has suggested that APs have several binding sites that allow for the stabilization of membrane binding and the open conformation of the complexes, but there are initial interactions required that dictate their recruitment to the target membrane. Although these interaction sites for mammalian AP-1 and AP-2 have been identified in great detail based on interaction analyses and structural studies (8, 10, 11, 35, 36, 38, 39), structural data for AP-3 is largely missing. The C-terminal part of the μ-subunit of mammalian AP-3 has been crystallized together with a Yxxφ motif-containing a cargo peptide, which revealed a similar fold and cargo-binding site as shown for AP-1 and AP-2 (14). However, positively charged binding surfaces required for PIP-interaction were not well conserved. Although the “trunk” segment of AP-1 and AP-2 is known quite well by now, information on hinge and ear domains in context of these complexes is largely missing. Crystal structures of the isolated ear domains of α-, γ- and β2-adaptin have been published (40, 41, 42), and a study on mammalian AP-3 suggested a direct interaction between δ-ear and δ3 that interfered with Arf1-binding (43). Furthermore, during tethering of AP-3 vesicles with the yeast vacuole, the δ−subunit Apl5 of the yeast AP-3 complex binds to the Vps41 subunit of the HOPS complex as a prerequisite of fusion (18, 19, 21, 22).In this study, we applied single particle electron cryo-microscopy (cryo-EM) to analyze the purified full-length AP-3 complex from yeast and unraveled the factors required for AP-3 recruitment to membranes by biochemical reconstitution. Our data reveal that a surprisingly flexible AP-3 complex requires a combination of cargo, PI4P, and Arf1 for membrane binding, which explains its function in selective cargo sorting at the Golgi.     

When should cuckolded males care for extra-pair offspring?

In socially monogamous species with bi-parental care, males suffer reduced reproductive success if their mate engages in extra-pair copulations (EPCs). One might therefore expect that males should refuse to care for a brood if they can detect that an EPC has occurred. Here, we use a game-theory model to study male brood care in the face of EPCs in a cooperatively breeding species in which offspring help to raise their (half-) siblings in their parents' next breeding attempt. We show that under certain conditions males are selected to care even for broods completely unrelated to themselves. This counterintuitive result arises through a form of pseudo-reciprocity, whereby surviving extra-pair offspring, when helping to rear their younger half-siblings, can more than compensate for the cost incurred by the male that raised them. We argue that similar effects may not be limited to cooperative breeders, but may arise in various contexts in which cooperation between (half-) siblings occurs.