HCV core protein localizes in the nuclei of nonparenchymal liver cells from chronically HCV-infected patients

Understanding the mechanism of hepatitis C virus (HCV) pathogenesis is an important part of HCV research. Recent experimental evidence suggests that the HCV core protein (HCcAg) has numerous functional activities. These properties suggest that HCcAg, in concert with cellular factors, may contribute to pathogenesis during persistent HCV infection. HCV is capable of infecting cells other than hepatocytes. Although the extrahepatic cellular tropism of HCV may play a role in the pathophysiology of this infection, the precise biological significance of the presence of HCV components in different liver cell types presently remains to be established. In this study, HCcAg was detected in nonparenchymal liver cells of six patients out of eight positive for serum HCV RNA. Immunostaining with anti-HCcAg mAbs revealed the presence of this protein in different liver cell types such as lymphocytes, Kupffer, polymorphonuclear, pit, endothelial, stellate, and fibroblast-like cells. Interestingly, HCcAg was immunolabeled not only in the cytoplasm but also in the nucleus of these cells. Remarkably, HCcAg co-localized with large lipid droplets present in stellate cells and with collagen fibers in the extracellular matrix. Moreover, HCcAg was immunolabeled in bile canaliculus suggesting the involvement of the biliary system in the pathobiology of HCV. Data suggest that nonparenchymal liver cells may constitute a reservoir for HCV replication. Besides, HCcAg may contribute to modulate immune function and fibrosis in the liver as well as steatosis.     

Analysis of the stability of the spermadhesin PSP-I/PSP-II heterodimer. Effects of Zn2+ and acidic pH

Spermadhesins are a family of 12-16 kDa proteins with a single CUB domain. PSP-I and PSP-II, the most abundant boar spermadhesins, are present in seminal plasma as a noncovalent heterodimer. Dimerization markedly affects the binding ability of the subunits. Notably, heparin and mannose 6-phosphate binding abilities of PSP-II are abolished, indicating that the corresponding binding sites may be located at (or near) the dimer interface. Pursuing the hypothesis that cryptic binding sites in PSP-I/PSP-II may be exposed in specific physiological environments, we examined the influence of Zn2+ and acidic pH on the heterodimer stability. According to near-UV CD spectra, the core native fold is preserved in the presence of physiological concentrations of Zn2+, a cation unusually abundant in boar seminal plasma. However, the thermostability of the heterodimer decreases significantly, as observed by CD and differential scanning calorimetry. The effect is Zn2+-specific and is reversed by EDTA. Destabilization is also observed at acidic pH. Gel filtration analysis using radioiodinated PSP-I/PSP-II reveals that dissociation of the heterodimer at low (nanomolar) protein concentrations is promoted by both Zn2+ and acidic pH. Although the integrity of the heterodimer in seminal plasma seems to be guaranteed by its high concentration, dissociation may be facilitated in the female genital tract because of dilution of the protein in the intraluminal fluids of the cervix and the uterus, and the acidic fluid of the uterotubal junction. Such a mechanism may be relevant in the regulation of uterine immune reactions.     

Characterization of liposomal tacrolimus in lung surfactant-like phospholipids and evaluation of its immunosuppressive activity

Tacrolimus (FK506) is a hydrophobic immunosuppressive agent that rapidly penetrates the plasmatic membrane and inhibits the signal transduction cascade of T lymphocytes. The objective of this study was the characterization of liposomal FK506 with surfactant-like phospholipids to be administered intratracheally after lung transplantation or in inflammatory lung diseases. We evaluated the optimal incorporation of FK506 in dipalmitoylphosphatidylcholine (DPPC) and DPPC/1-palmitoyl-2-oleoylphosphatidylglycerol (POPG) monolayers and bilayers and the effects of FK506 on the physical properties of DPPC and DPPC/POPG (8:2 w/w) vesicles. In addition, we assessed the immunosuppressive effects of surfactant-like phospholipid vesicles containing different amounts of FK506 on T-cell proliferation and interleukin 2 production. From surface pressure measurements of FK506/DPPC and FK506/DPPC/POPG mixed monolayers, we determined that FK506 was embedded into these monolayers up to an FK506 concentration of about 0.4 mol %. Beyond this concentration, FK506 was not quantitatively incorporated into the monolayer, suggesting possible concentration-dependent aggregation of tacrolimus. The incorporation of FK506 into DPPC monolayers, at concentrations 相似文献   

Comparison of three ELISA protocols to measure antibody responses elicited against serogroup C meningococcal polysaccharide in mouse, monkey and human sera

In this study we compared the following ELISA protocols to measure antibody levels against serogroup C meningococcal polysaccharide: a traditional protocol using poly-L-Lysine mixed with the polysaccharide as coating antigen, a second protocol coating with a mixture of methylated human serum albumin with the C polysaccharide, a modified protocol coating with derivatized polysaccharide and a modification to the last one, specifically without adding ammonium thiocyanate to the sample buffer. Serum bactericidal activity of mouse, monkey and human sera were measured and correlation coefficients were calculated. For all serum types the modified ELISA protocol showed the highest correlation coefficients while the traditional protocol showed the lower ones. We demonstrated that the traditional protocol measures non-specific antibodies to the C polysaccharide, because no differences were detected between pre-immune and post-immune human sera (P>0.05), while the modified protocol detected the highest difference (P<0.01).     

The aureolic acid family of antitumor compounds: structure, mode of action, biosynthesis, and novel derivatives

Members of the aureolic acid family are tricyclic polyketides with antitumor activity which are produced by different streptomycete species. These members are glycosylated compounds with two oligosaccharide chains of variable sugar length. They interact with the DNA minor groove in high-GC-content regions in a nonintercalative way and with a requirement for magnesium ions. Mithramycin and chromomycins are the most representative members of the family, mithramycin being used as a chemotherapeutic agent for the treatment of several cancer diseases. For chromomycin and durhamycin A, antiviral activity has also been reported. The biosynthesis gene clusters for mithramycin and chromomycin A₃ have been studied in detail by gene sequencing, insertional inactivation, and gene expression. Most of the biosynthetic intermediates in these pathways have been isolated and characterized. Some of these compounds showed an increase in antitumor activity in comparison with the parent compounds. A common step in the biosynthesis of all members of the family is the formation of the tetracyclic intermediate premithramycinone. Further biosynthetic steps (glycosylation, methylations, acylations) proceed through tetracyclic intermediates which are finally converted into tricyclic compounds by the action of a monooxygenase, a key event for the biological activity. Heterologous expression of biosynthetic genes from other aromatic polyketide pathways in the mithramycin producer (or some mutants) led to the isolation of novel hybrid compounds.Felipe Lombó and Nuria Menéndez have equally contribute to this work.     

Can occupancy patterns be used to predict distributions in widely separated geographic regions?

Occupancy models, that describe the presence and absence patterns of a species in a given area, are increasingly being used to predict the occurrence of the species in unsurveyed sites, as an aid to conservation planning. In this paper, we consider whether conclusions about local distributions derived from one landscape can be extrapolated to others. We found that habitat patchiness influenced the distribution and abundance of the host-specific moth Wheeleria spilodactylus in a similar way in two landscapes widely separated geographically. In both geographic regions, the spatial location (positive effect of connectivity), and quantity of resource (positive effect of host plant density) increased the likelihood that the moth would be present, consistent with the expectations of metapopulation dynamics. Though some biological attributes of the species appeared to be slightly different, including population density and the timing of the life cycle (phenology), occupancy patterns in one landscape accurately predict occupancy in the other landscape. Our results suggest that it maybe possible to make predictions from one landscape to another, even when the landscapes are widely separated.     

Smad7 attenuates TGF-β-mediated aging-related hypofunction of submandibular glands

Submandibular glands have essential functions in taste, mastication, swallowing, and digestion. Submandibular gland hypofunction is prevalent in the elderly, impairing the patients’ quality of life. Current clinical treatment strategies have not decelerated or reversed the pathological process of submandibular gland hypofunction. Therefore, novel restoration strategies should be explored. However, studies on the mechanism of aging-related submandibular gland hypofunction remain very limited. The role of the TGF-β/Smad pathway in fibrosis has been studied in other organs. Therefore, this study aimed to elucidate the role of TGF-β/Smad signaling in the aging-related submandibular gland hypofunction. The results showed that Smad7 knockout in mice decreased the salivary flow rate. H&E, Masson trichrome, and immunohistochemistry staining of MCP-1 and α-SMA showed that Smad7 knockout in mice resulted in lymphocytic infiltration, acinar cell atrophy, and interstitial fibrosis. The Western blotting of collagen I and III also confirmed extensive fibrosis. We then found that Smad7 depletion resulted in the TGF-β-mediated fibrosis via mir-21, mir-29, and np_5318, and NFκB-driven inflammation activation. This study confirmed the inhibitory role of Smad7 in the aging-related submandibular gland hypofunction. Therefore, it provided a promising treatment target for aging-related dysfunction and sialadenitis of submandibular gland.     

Comparative Study of Alkaline, Saline, and Mixed Saline–Alkaline Stresses with Regard to Their Effects on Growth, Nutrient Accumulation, and Root Morphology of Lotus tenuis

Both saline and alkaline conditions frequently coexist in nature; however, little is known about the effects of alkaline and salt?Calkaline stresses on plants. We performed pot experiments with four treatments, control without salt addition and three stress conditions??neutral, alkaline, and mixed salt?Calkaline??to determine their effects on growth, nutrient accumulation and root architecture in the glycophytic species Lotus tenuis. Neutral and alkaline salts produced a similar detrimental effect on L. tenuis growth, whereas the effect of their combination was synergistic. Neutral salt addition, alone or mixed with NaHCO₃, led to significant leaf Na⁺ build up and reduced K⁺ concentration. In contrast, in plants treated with NaHCO₃ only, Na⁺ levels and the Na⁺/K⁺ ratio remained relatively unchanged. Proline accumulation was not affected by the high pH in the absence of NaCl, but it was raised by the neutral salt and mixed treatments. The total root length was reduced by the addition of NaCl alone, whereas it was not affected by alkalinity, regardless of the presence of NaCl. The topological trend showed that alkalinity alone or mixed with NaCl turned the root more herringbone compared with control roots, whereas no significant change in this index was observed in the treatment with the neutral salt only. The pattern of morphological changes in L. tenuis root architecture after the alkaline treatment (in the absence of NaCl) was similar to that found in the mixed salt?Calkaline treatment and different from that observed in neutral salt. A unique root morphological response to the mixed salt?Calkaline stress was the reduction in the ratio between xylem vessels and root cross-sectional areas.     

The role of capsid maturation on adenovirus priming for sequential uncoating

Adenovirus assembly concludes with proteolytic processing of several capsid and core proteins. Immature virions containing precursor proteins lack infectivity because they cannot properly uncoat, becoming trapped in early endosomes. Structural studies have shown that precursors increase the network of interactions maintaining virion integrity. Using different biophysical techniques to analyze capsid disruption in vitro, we show that immature virions are more stable than the mature ones under a variety of stress conditions and that maturation primes adenovirus for highly cooperative DNA release. Cryoelectron tomography reveals that under mildly acidic conditions mimicking the early endosome, mature virions release pentons and peripheral core contents. At higher stress levels, both mature and immature capsids crack open. The virus core is completely released from cracked capsids in mature virions, but it remains connected to shell fragments in the immature particle. The extra stability of immature adenovirus does not equate with greater rigidity, because in nanoindentation assays immature virions exhibit greater elasticity than the mature particles. Our results have implications for the role of proteolytic maturation in adenovirus assembly and uncoating. Precursor proteins favor assembly by establishing stable interactions with the appropriate curvature and preventing premature ejection of contents by tightly sealing the capsid vertices. Upon maturation, core organization is looser, particularly at the periphery, and interactions preserving capsid curvature are weakened. The capsid becomes brittle, and pentons are more easily released. Based on these results, we hypothesize that changes in core compaction during maturation may increase capsid internal pressure to trigger proper uncoating of adenovirus.