Human decidua-derived mesenchymal stromal cells differentiate into hepatic-like cells and form functional three-dimensional structures

Background aimsPreviously, we have shown that human decidua-derived mesenchymal stromal cells (DMSC) are mesenchymal stromal cells (MSC) with a clonal differentiation capacity for the three embryonic layers. The endodermal capacity of DMSC was revealed by differentiation into pulmonary cells. In this study, we examined the hepatic differentiation of DMSC.MethodsDMSC were cultured in hepatic differentiation media or co-cultured with murine liver homogenate and analyzed with phenotypic, molecular and functional tests.Results and ConclusionsDMSC in hepatic differentiation media changed their fibroblast morphology to a hepatocyte-like morphology and later formed a 3-dimensional (3-D) structure or hepatosphere. Moreover, the hepatocyte-like cells and the hepatospheres expressed liver-specific markers such as synthesis of albumin (ALB), hepatocyte growth factor receptor (HGFR), α-fetoprotein (AFP) and cytokeratin-18 (CK-18), and exhibited hepatic functions including glycogen storage capacity and indocyanine green (ICG) uptake/secretion. Human DMSC co-cultured with murine liver tissue homogenate in a non-contact in vitro system showed hepatic differentiation, as evidenced by expression of AFP and ALB genes. The switch in the expression of these two genes resembled liver development. Indeed, the decrease in AFP and increase in ALB expression throughout the co-culture were consistent with the expression pattern observed during normal liver organogenesis in the embryo. Interestingly, AFP and ALB expression was significantly higher when DMSC were co-cultured with injured liver tissue, indicating that DMSC respond differently under normal and pathologic micro-environmental conditions. In conclusion, DMSC-derived hepatospheres and DMSC co-cultured with liver homogenate could be suitable in vitro models for toxicologic, developmental and pre-clinical hepatic regeneration studies.     

Noncellular nonspecific defence mechanisms of fish

Fish tissues and body fluids contain naturally occurring proteins or glycoproteins of non-immunoglobulin (Ig) nature that react with a diverse array of environmental antigens and may confer an undefined degree of natural immunity to fish. They consist of microbial growth inhibitory compounds that include “acute phase” proteins such as transferrins, caeruloplasmin, and metallothionein. Their action is simply to chelate metal ions and deprive bacteria and other parasites of essential inorganic ion sources. Both serum and cellular interferons are found in fish, and this anti-virus protein has been demonstrated mainly in salmonids during viral disease studies. Enzyme-inhibitors (α2-macroglobulin and other α-globulins) thus far detected in fish appear to be antibacterial proteinases, and are involved in the inhibition of extracellular proteases secreted by fish pathogens. Fish also possess a variety of relatively specific lytic molecules that cause cell lysis, and some of these materials are hydrolase enzymes (lysozyme, chitinase, chitobiase) whose main actions are against bacteria and fungi. In addition, mucus contains trypsin-like proteinases which destroy gram-negative bacteria. Nonspecific lysins and agglutinins against erythrocytes and other cellular antigens are found in serum, eggs, and skin mucus. The lysins, including toxins, some of which are bacteriolytic in activity, are, in their mode of action, natural or spontaneous, antibody-independent and noncomplement-mediated. In contrast, specific hemolytic antibodies (Ig), which complex with antigens, bind complement, and cause complement-mediated immune lysis, are reported to exist. The agglutinins are generally reactive toward certain sugar residues on erythrocyte or bacterial cell walls, and in most cases act as lectins or lectin-like molecules. Natural lysins and agglutinins behave in a similar way as antigen-induced antibodies or Igs, but exhibit a high degree of cross-reactions, due to the occurrence of similar carbohydrate determinants on many types of microbial cell surface. As with mammals, both C-type (calcium-dependent) and S-type (thiol-dependent) lectins are present in fish. They more resemble invertebrate lectins than those of higher animals. Fish lectins appear to play antibacterial or antifungal roles and in some instances seem to be involved in egg-sperm fusion, polyspermy prevention, and embryo development. Natural, non-Ig precipitins (e.g. α-precipitin and C-reactive proteins) are found largely, but not exclusively, in fish serum and precipitate with simple monosaccharides or long chain polysaccharides of certain stereochemistry and glycosidic linkages. Their functions remain unknown, but C-reactive protein is induced following stress-induction and exposure to inflammatory agents. Many of the above mentioned “defence” substances are present in skin mucus and possess the capacity to react with potentially infective microorganisms including parasites. Mucus thus acts as an immediate defence barrier to invasion and/or colonisation of pathogens.     

Regulation of scytophycin accumulation in cultures of Scytonema ocellatum. I. Physical factors

Scytonema ocellatum (Cyanobacteria or Cyanophyta) produces macrolide antibiotics of the scytophycin family that are antifungal, cytostatic agents and act by disrupting actin microfilaments. Scytophycin accumulation paralleled vegetative growth. Tolytoxin continued to accumulate throughout the growth cycle, whereas 6-hydroxy-7-O-methylscytophycin E (HMSE) and 19-O-demethylscytophycin C (DMSC) reached plateau levels prior to cessation of growth, suggesting a logical biosynthetic pathway of DMSC → ? → HMSE → tolytoxin. A rapid decrease in scytophycin content observed in newly inoculated cultures suggests that the scytophycins are continuously metabolized. The optimal temperature for production was 25°C. Continuous illumination at an intensity of at least 25 μml photons m^?2 s^?1 was required for maximum yield. Growth and metabolite production were optimal in the pH range of 8.0 to 8.5.     

Costs to host defence and the persistence of parasitic cuckoos.

Raising genetically unrelated young is maladaptive, yet brood parasitism is widespread in birds. In several systems, hosts can evolve near-perfect defences against the parasite (discrimination and rejection of unlike eggs), making it difficult to understand how the parasite continues to exist. This study demonstrates costs to host defences (e.g. rejection of one's own eggs) such that once the parasite goes extinct on a particular host species, defence mechanisms are selectively disadvantageous. The consequent loss of host defences, and potential for re-exploitation of the host by the parasite, can explain the continued persistence of avian brood parasites. The results provide one general explanation for coexistence of parasites and their hosts.     

Doxycycline Does Not Influence Established Abdominal Aortic Aneurysms in Angiotensin II-Infused Mice

Background

There is no proven medical approach to attenuating expansion and rupture of abdominal aortic aneurysms (AAAs). One approach that is currently being investigated is the use of doxycycline. Despite being primarily used as an antimicrobial drug, doxycycline has been proposed to function in reducing AAA expansion. Doxycycline is effective in reducing the formation in the most commonly used mouse models of AAAs when administered prior to the initiation of the disease. The purpose of the current study was to determine the effects of doxycycline on established AAAs when it was administered at a dose that produces therapeutic serum concentrations.

Methods and Results

LDL receptor −/− male mice fed a saturated-fat supplemented diet were infused with AngII (1,000 ng/kg/min) via mini-osmotic pumps for 28 days. Upon verification of AAA formation by noninvasive high frequency ultrasonography, mice were stratified based on aortic lumen diameters, and continuously infused with AngII while also administered either vehicle or doxycycline (100 mg/kg/day) in drinking water for 56 days. Administration of doxycycline led to serum drug concentrations of 2.3±0.6 µg/ml. Doxycycline administration had no effect on serum cholesterol concentrations and systolic blood pressures. Doxycycline administration did not prevent progressive aortic dilation as determined by temporal measurements of lumen dimensions using high frequency ultrasound. This lack of effect on AAA regression and progression was confirmed at the termination of the study by ex vivo measurements of maximal width of suprarenal aortas and AAA volumes. Also, doxycycline did not reduce AAA rupture. Medial and adventitial remodeling was not overtly changed by doxycycline as determined by immunostaining and histological staining.

Conclusions

Doxycycline administration did not influence AngII-induced AAA progression and aortic rupture when administered to mice with established AAAs.     

Function and therapeutic potential of host defence peptides.

Cationic host defence (antimicrobial) peptides are an important component of the innate immune systems of a wide variety of plants, animals, and bacteria. Although most of these compounds have direct antimicrobial activities under specific conditions, a greater appreciation for the diversity of functions of these molecules is beginning to develop in the field. In addition to their directly antimicrobial activities, they also have a broad spectrum of activity on the host immune system, with both pro-inflammatory and anti-inflammatory effects being invoked. Increasingly sophisticated approaches to understand the role of host defence peptides in modulating innate immunity are already serving to guide the development of novel therapeutics.     

Macrophage lectins in host defence

Macrophage lectins contribute to host defence by a variety of mechanisms. The best characterised, mannose receptor (MR) and complement receptor three (CR3), are both able to mediate phagocytosis of pathogenic microbes and induce intracellular killing mechanisms. The regulation of the effector functions induced via MR is complex, and may involve both host and microbial factors. Therefore, MR is likely to play a dynamic role in the response to infection; it may act as a classical pattern recognition receptor in phagocytosis, whereas other poorly characterised factors may make a more decisive contribution to its function in physiologic settings. In contrast, the lectin site of CR3 appears to lack host-derived ligands and may be a true pattern recognition receptor. Further studies are required to evaluate the roles of other macrophage lectins in recognition of and responses to microbes.     

Development of a method for the quantitative evaluation of microorganism sensitivity to antibiotics utilizing disks. The determination of the minimal doxycycline concentration that depresses microorganism growth using disks containing this antibiotic]

A quantitative method for estimation of microbial sensitivity to doxycycline with the use of discs containing 10 gamma of the antibiotic was developed. The antibiotic concentrations in the agar were determined at a distance equal to the radius of the growth inhibition zone with the help of a curve of the dependence of the logarithm of the doxycycline concentration in agar at the period of the average critical time of the zone formation equal to 5 hours and the distance from the disc center. The antibiotic concentration in the agar at the zone edges was almost the same as the MIC of doxycycline against the test-cultures determined with the method of serial dilutions in agar.     

Disruption of neutrophil migration in a conditional transgenic model: evidence for CXCR2 desensitization in vivo.

We developed transgenic mice conditionally expressing the neutrophil chemoattracting chemokine KC and the beta-galactosidase gene in multiple tissues. In these transgenic mice, doxycycline treatment induced a strong up-regulation in the expression of KC in several tissues, including heart, liver, kidney, skin, and skeletal muscle. Expression of KC within these tissues led to a rapid and substantial increase in the serum levels of KC (serum KC levels were higher than 200 ng/ml 24 h after treatment). Accordingly, beta-galactosidase expression was also detected after injection of doxycycline and was highest in skeletal muscle, pancreas, and liver. Surprisingly, despite expression of KC in multiple tissues, no neutrophil infiltration was observed in any of the tissues examined, including skin. Doxycycline treatment of nontransgenic mice grafted with transgenic skin caused dense neutrophilic infiltration of the grafts, but not the surrounding host skin, indicating that the KC produced in transgenic tissues was biologically active. In separate experiments, neutrophil migration toward a localized source of recombinant KC was impaired in animals overexpressing KC but was normal in response to other neutrophil chemoattractants. Analysis of transgenic neutrophils revealed that high concentrations of KC in transgenic blood had no influence on L-selectin cell surface expression but caused desensitization of the receptor for KC, CXCR2. These results confirm the neutrophil chemoattractant properties of KC and provide a mechanistic explanation for the paradoxical lack of leukocyte infiltration observed in the presence of elevated concentrations of this chemokine.     

Myeloperoxidase in human neutrophil host defence

Human neutrophils represent the predominant leucocyte in circulation and the first responder to infection. Concurrent with ingestion of microorganisms, neutrophils activate and assemble the NADPH oxidase at the phagosome, thereby generating superoxide anion and hydrogen peroxide. Concomitantly, granules release their contents into the phagosome, where the antimicrobial proteins and enzymes synergize with oxidants to create an environment toxic to the captured microbe. The most rapid and complete antimicrobial action by human neutrophils against many organisms relies on the combined efforts of the azurophilic granule protein myeloperoxidase and hydrogen peroxide from the NADPH oxidase to oxidize chloride, thereby generating hypochlorous acid and a host of downstream reaction products. Although individual components of the neutrophil antimicrobial response exhibit specific activities in isolation, the situation in the environment of the phagosome is far more complicated, a consequence of multiple and complex interactions among oxidants, proteins and their by‐products. In most cases, the cooperative interactions among the phagosomal contents, both from the host and the microbe, culminate in loss of viability of the ingested organism.     

Candida albicans morphogenesis and host defence: discriminating invasion from colonization

Candida albicans is a common fungal pathogen of humans that colonizes the skin and mucosal surfaces of most healthy individuals. Until recently, little was known about the mechanisms by which mucosal antifungal defences tolerate colonizing C. albicans but react strongly when hyphae of the same microorganism attempt to invade tissue. In this Review, we describe the properties of yeast cells and hyphae that are relevant to their interaction with the host, and the immunological mechanisms that differentially recognize colonizing versus invading C. albicans.     

Interaction between Doxycycline and Barbiturates

In a cross-over study of five hospitalized patients the half life of doxycycline was significantly shortened after 10 days'' treatment with phenobarbitone. In five patients on continuous barbiturate therapy the half life of doxycycline was even shorter. Barbiturates or other agents inducing drug metabolism should be used cautiously in combination with doxycycline, since this might result in therapeutically inadequate serum concentrations of the antibiotic.     

Importance of microbial defence systems to bile salts and mechanisms of serum cholesterol reduction

An important feature of the intestinal microbiota, particularly in the case of administered probiotic microorganisms, is their resistance to conditions in the gastrointestinal tract, particularly tolerance to and growth in the presence of bile salts. Bacteria can use several defence mechanisms against bile, including special transport mechanisms, the synthesis of various types of surface proteins and fatty acids or the production of exopolysaccharides. The ability to enzymatically hydrolyse bile salts occurs in a variety of bacteria. Choloylglycine hydrolase (EC 3.5.1.24), a bile salt hydrolase, is a constitutive intracellular enzyme responsible for the hydrolysis of an amide bond between glycine or taurine and the steroid nucleus of bile acids. Its presence was demonstrated in specific microorganisms from several bacterial genera (Lactobacillus spp., Bifidobacterium spp., Clostridium spp., Bacteroides spp.). Occurrence and gene arrangement encoding this enzyme are highly variable in probiotic microorganisms. Bile salt hydrolase activity may provide the possibility to use the released amino acids by bacteria as sources of carbon and nitrogen, to facilitate detoxification of bile or to support the incorporation of cholesterol into the cell wall. Deconjugation of bile salts may be directly related to a lowering of serum cholesterol levels, from which conjugated bile salts are synthesized de novo. Furthermore, the ability of microorganisms to assimilate or to bind ingested cholesterol to the cell wall or to eliminate it by co-precipitation with released cholic acid was also documented. Some intestinal microflora produce cholesterol reductase that catalyses the conversion of cholesterol to insoluble coprostanol, which is subsequently excreted in faeces, thereby also reducing the amount of exogenous cholesterol.     

A remote sensing approach to estimating harvestable kelp biomass

Regulations of the Alaska Department of Fish and Game require that all fisheries in the state have a harvest management plan. In southeast Alaska two species of floating kelps, Nereocystis luetkeana and Alaria fistulosa, have been commercially harvested since 1992 for use as agrochemicals by the Alaska Kelp Company. However, there is currently no harvest management plan for this fishery. The lack of a formalized management plan is one factor that has kept the kelp industry from expanding in the state. We have employed an aerial digital multispectral imaging system (DMSC) calibrated with ground truthing for performing such an assessment. The system can be flown at varying altitudes to achieve spatial resolutions ranging from 0.5 to 2 m. Rapid ground truthing techniques were developed using morphometric measurements to predict biomass. Analysis of the DMSC imagery showed that good correlations could be developed between the multispectral imagery and kelp biomass estimates collected at the ground-truth sites. Repeatable estimates of kelp bed area derived from the multispectral imagery could be made at varying tidal levels. However, broad scale maps of kelp biomass suitable for estimating harvest rates could not be made at different tide levels. Multispectral imagery suitable for this purpose must be collected at a standard tidal level.     

Parasitism, host immune defence and dispersal

Host-parasite interactions have been hypothesized to affect the evolution of dispersal by providing a possibility for hosts to escape debilitating parasites, and by influencing the level of local adaptation. We used a comparative approach to investigate the relationship between a component of host immune function (which reflects the evolutionary history of parasite-induced natural selection) and dispersal in birds. We used a sample of 46 species of birds for which we had obtained field estimates of T-cell response for nestlings, mainly from our own field studies in Denmark and Spain. Bird species with longer natal, but not with longer breeding dispersal distances had a stronger mean T-cell-mediated immune response in nestlings than species with short dispersal distances. That was also the case when controlling for the potentially confounding effect of migration from breeding to wintering area, which is known from previous studies to be positively associated with dispersal distance. These relationships held even when controlling for similarity among species because of common ancestry. Avian hosts with a larger number of different breeding habitats had weaker mean T-cell-mediated immune responses than habitat specialists. This relationship held even when controlling for similarity among species because of common ancestry. Therefore, T-cell-mediated immunity is an important predictor of evolutionary changes in dispersal ability among common European birds.     

Restoration of host defense mechanisms in man by levamisole

Levamisole, a broad-spectrum anthelmintic, increased the delayed hypersensitivity skin reaction to tuberculin and to DNCB in about one third of the patients. Levamisole and the racemate tetramisole temporarily increased serum antibody titres in response to vaccination against influenza.     

Flow cytometry as a tool to quantify oyster defence mechanisms

The fast growing oyster aquaculture industry is greatly hindered by Perkinsus marinus and Haplosporidium nelsoni which can kill up to 80% of the production. The relationship between parasites and oyster defence mechanisms is unclear. Two defence mechanisms of the Eastern Oyster (Crassostrea virginica) were quantified at the single cell level utilising flow cytometry. Phagocytosis was measured using fluorescent beads. Respiratory burst activity was quantified as the H2O2-specific increase in dichlorofluorescein-associated fluorescence upon stimulation. These two assays distinguished three populations of haemocytes (granulocytes, hyalinocytes and intermediate cells) with unique functional characteristics. Granulocytes were most active at phagocytosis and H2O2 production while hyalinocytes were relatively inactive. The intermediate cells had moderate phagocytic and respiratory burst activity. Flow cytometry can rapidly, accurately and directly quantify the morphology and function of a large number of individual cells, and will lead to a better understanding of the bivalve immune system.     

Effect of doxycycline on leukocyte adhesion in a multifactorial experiment]

The multifactorial analysis was applied to study the effect of doxycycline on leukocyte adhesion to a plastic within wide ranges of the values of experimental factors such as doxycycline concentrations and magnesium and calcium ions. The data were computer-processed and second-order equations characterizing the influence of the factors on adhesion were derived. It was shown that the effect of the ions on adhesion was non-single-valued; accurate quantitative models describing the ion effect on adhesion were developed when the concentration of doxycycline was changed.     

Unifying themes in host defence effector polypeptides

It is said that nature is the greatest innovator, yet molecular conservation can be equally powerful. One key requirement for the survival of any host is its ability to defend against infection, predation and competition. Recent discoveries, including the presence of a multidimensional structural signature, have revealed a previously unforeseen structural and functional congruence among host defence effector molecules spanning all kingdoms of life. Antimicrobial peptides, kinocidins, polypeptide venoms and other molecules that were once thought to be distinct in form and function now appear to be members of an ancient family of host defence effectors. These molecules probably descended from archetype predecessors that emerged during the beginning of life on earth. Understanding how nature has sustained these host defence molecules with a potent efficacy in the face of dynamic microbial evolution should provide new opportunities to prevent or treat life-threatening infections.