Functions of Liver Natural Killer Cells Are Dependent on the Severity of Liver Inflammation and Fibrosis in Chronic Hepatitis C

During chronic hepatitis C virus (HCV) infection, the role of intra-hepatic (IH) natural killer (NK) cells is still controversial. To clarify their functions, we investigated anti-viral and cytotoxic activity of NK cells in human fresh liver biopsies. We compared the functions of IH-NK cells in HCV-infected and NASH patients in physiological conditions as well as after stimulation using flow cytometric and immunohistochemical analyses. Interestingly, few IH-NK cells produced anti-viral cytokine IFN-γ in HCV-infected patients similarly as in non-infected individuals. Spontaneous degranulation activity was extremely low in peripheral NK cells compared to IH-NK cells, and was significantly higher in IH-NK cells from HCV-infected patients compared to non-infected individuals. Immunohistochemical analysis revealed that perforin granules were polarized at the apical pole of IH-NK cells. The presence of CD107a and perforin in IH-NK cells demonstrated that NK cells exerted a cytolytic activity at the site of infection. Importantly, IH-NK cell functions from HCV-infected patients were inducible by specific exogenous stimulations. Upon ex vivo K562 target cell stimulations, the number of degranulating NK cells was significantly increased in the pool of IH-NK cells compared to circulating NK cells. Interestingly, after stimulation, the frequency of IFN-γ-producing IH-NK cells in HCV-infected patients was significantly higher at early stage of inflammation whereas the spontaneous IH-NK cell degranulation activity was significantly impaired in patients with highest inflammation and fibrosis Metavir scores. Our study highlights that some IH-NK cells in HCV-infected patients are able to produce INF-γ and degranulate and that those two activities depend on liver environment including the severity of liver injury. Thus, we conclude that critical roles of IH-NK cells have to be taken into account in the course of the liver pathogenesis associated to chronic HCV infection.     

GWAS findings for human iris patterns: associations with variants in genes that influence normal neuronal pattern development

Human iris patterns are highly variable. The origins of this variation are of interest in the study of iris-related eye diseases and forensics, as well as from an embryological developmental perspective, with regard to their possible relationship to fundamental processes of neurodevelopment. We have performed genome-wide association scans on four iris characteristics (crypt frequency, furrow contractions, presence of peripupillary pigmented ring, and number of nevi) in three Australian samples of European descent. Both the discovery (n = 2121) and replication (n = 499 and 73) samples showed evidence for association between (1) crypt frequency and variants in the axonal guidance gene SEMA3A (p = 6.6 × 10⁻¹¹), (2) furrow contractions and variants within the cytoskeleton gene TRAF3IP1 (p = 2.3 × 10⁻¹²), and (3) the pigmented ring and variants in the well-known pigmentation gene SLC24A4 (p = 7.6 × 10⁻²¹). These replicated findings individually accounted for around 1.5%–3% of the variance for these iris characteristics. Because both SEMA3A and TRAFIP1 are implicated in pathways that control neurogenesis, neural migration, and synaptogenesis, we also examined the evidence of enhancement among such genes, finding enrichment for crypts and furrows. These findings suggest that genes involved in normal neuronal pattern development may also influence tissue structures in the human iris.     

YcdB from Escherichia coli reveals a novel class of Tat-dependently translocated hemoproteins

The Tat (twin-arginine translocation) system of Escherichia coli serves to translocate folded proteins across the cytoplasmic membrane. The reasons established so far for the Tat dependence are cytoplasmic cofactor assembly and/or heterodimerization of the respective proteins. We were interested in the reasons for the Tat dependence of novel Tat substrates and focused on two uncharacterized proteins, YcdO and YcdB. Both proteins contain predicted Tat signal sequences. However, we found that only YcdB was indeed Tat-dependently translocated, whereas YcdO was equally well translocated in a Tat-deficient strain. YcdB is a dimeric protein and contains a heme cofactor that was identified to be a high-spin Fe(III)-protoporphyrin IX complex. In contrast to all other periplasmic hemoproteins analyzed so far, heme was assembled into YcdB in the cytoplasm, suggesting that heme assembly could take place prior to translocation. The function of YcdB in the periplasm may be related to a detoxification reaction under specific conditions because YcdB had peroxidase activity at acidic pH, which coincides well with the known acid-induced expression of the gene. The data demonstrate the existence of a class of heme-containing Tat substrates, the first member of which is YcdB.     

Bioaugmentation of microbial communities in laboratory and pilot scale sequencing batch biofilm reactors using the TOL plasmid

The aim of this study was to investigate the effectiveness of bioaugmentation and transfer of plasmid pWWO (TOL plasmid) to mixed microbial populations in pilot and laboratory scale sequencing batch biofilm reactors (SBBRs) treating synthetic wastewater containing benzyl alcohol (BA) as a model xenobiotic. The plasmid donor was a Pseudomonas putida strain chromosomally tagged with the gene for the red fluorescent protein carrying a green fluorescent protein labeled TOL plasmid, which confers degradation capacity for several compounds including toluene and BA. In the pilot scale SBBR donor cells were disappeared 84 h after inoculation while transconjugants were not detected at all. In contrast, both donor and transconjugant cells were detected in the laboratory scale reactor where the ratio of transconjugants to donors fluctuated between 1.9 × 10^?1 and 8.9 × 10^?1 during an experimental period of 32 days. BA degradation rate was enhanced after donor inoculation from 0.98 mg BA/min prior to inoculation to 1.9 mg BA/min on the seventeenth day of operation. Survival of a bioaugmented strain, conjugative plasmid transfer and enhanced BA degradation was demonstrated in the laboratory scale SBBR but not in the pilot scale SBBR.     

Evolutionary origin of human and primate malarias: evidence from the circumsporozoite protein gene

We have analyzed the conserved regions of the gene coding for the circumsporozoite protein (CSP) in 12 species of Plasmodium, the malaria parasite. The closest evolutionary relative of P. falciparum, the agent of malignant human malaria, is P. reichenowi, a chimpanzee parasite. This is consistent with the hypothesis that P. falciparum is an ancient human parasite, associated with humans since the divergence of the hominids from their closest hominoid relatives. Three other human Plasmodium species are each genetically indistinguishable from species parasitic to nonhuman primates; that is, for the DNA sequences included in our analysis, the differences between species are not greater than the differences between strains of the human species. The human P. malariae is indistinguishable from P. brasilianum, and P. vivax is indistinguishable from P. simium; P. brasilianum and P. simium are parasitic to New World monkeys. The human P. vivax-like is indistinguishable from P. simiovale, a parasite of Old World macaques. We conjecture that P. malariae, P. vivax, and P. vivax-like are evolutionarily recent human parasites, the first two at least acquired only within the last several thousand years, and perhaps within the last few hundred years, after the expansion of human populations in South America following the European colonizations. We estimate the rate of evolution of the conserved regions of the CSP gene as 2.46 x 10(-9) per site per year. The divergence between the P. falciparum and P. reichenowi lineages is accordingly dated 8.9 Myr ago. The divergence between the three lineages leading to the human parasites is very ancient, about 100 Myr old between P. malariae and P. vivax (and P. vivax-like) and about 165 Myr old between P. falciparum and the other two.      

On stiffness of scaffolds for bone tissue engineering—a numerical study

Tissue scaffolds are typically designed and fabricated to match native bone properties. However, it is unclear if this would lead to the best tissue ingrowth outcome within the scaffold as neo-tissue keeps changing the stiffness of entire construct. This paper presents a numerical method to address this issue for design optimization and assessment of tissue scaffolds. The elasticity tensors of two different types of bones are weighted by different multipliers before being used as the targets in scaffold design. A cost function regarding the difference between the effective elasticity tensor, calculated by the homogenization technique, and the target tensor, is minimized by using topology optimization procedure. It is found that different stiffnesses can lead to different remodeling results. The comparison confirms that bone remodeling is at its best when the scaffold elastic tensor matches or is slightly higher than the elastic properties of the host bone.     

In vitro and in vivo antimuscarinic effects of (-)cis 2,3-dihydro-3-(4-methylpiperazinylmethyl)-2-phenyl-1,5 benzothiazepin-4-(5H)one HCl (BTM-1086) in guinea pig peripheral tissues

The potency and selectivity of (-)cis-2,3-dihydro-3-(4-methylpiperazinylmethyl)-2-phenyl-1,5 benzothiazepin-4-(5H)one HCl (BTM-1086) for muscarinic receptor subtypes was compared in functional assay systems, in guinea pig peripheral tissues, to known reference drugs: atropine (nonselective), pirenzepine (M1), AF-DX 116 (M2) and HHSiD (M3). Like atropine, BTM-1086 was a potent, nonselective, competitive muscarinic antagonist with no detectable antispasmodic activity in urinary bladder or ileal muscle. In vivo, in the guinea pig cystometrogram, BTM-1086 depressed intravesical bladder pressure (PvesP) with the same efficacy and potency as oxybutynin, a drug used clinically for the treatment of urinary incontinence. The pharmacological profile of BTM-1086, however, suggests that it may not be suitable for development for bladder dysfunction disorders.     

Isolation of proteins assembled in lipid body membranes during fat mobilization in cucumber cotyledons

Lipid bodies from fat-mobilizing cotyledons of cucumber and other Cucurbitaceae were investigated. Proteins and glycoproteins were analyzed by electrophoresis and then used to characterize the lipid body membrane at different stages of cell development. Contaminations by other membranes or organelles were ruled out by comparing the main constituents from the endoplasmic reticulum, cytosol, glyoxysomes and protein bodies with the pattern of the lipid body membrane, considering both the prevalent peptides and the dominating glycoproteins. Among the proteins of lipid body membranes in ripening and germinating cotyledons, a 90-kDa peptide was found as unique marker of lipid bodies at the stage of fat mobilization. The 90-kDa protein was purified, and antibodies against it were raised in rabbits. By means of immunoprecipitation and electrophoretic analysis it was demonstrated that the synthesis of the 90-kDa form located in lipid bodies shows a transient increase and subsequent decline, with maximal values being observed at day 3 of germination. At this stage, the rate of de novo synthesis was compared considering lipid body proteins and other organellar proteins. The 90-kDa protein appeared as the lipid body constituent that is synthesized and assembled in the organelle by far at the highest rates.