Human Interleukin-6 Facilitates Hepatitis B Virus Infection in Vitro and in Vivo

Background and aim. Research on hepatitis B virus (HBV) infection in vivo has been limited due to the absence of a suitable animal model. We have developed a human–mouse radiation chimera in which normal mice, preconditioned by lethal total body irradiation and radioprotected with SCID mouse bone marrow cells, are permissive for engraftment of human hematopoietic cells and solid tissues. This resulting human–mouse model, which comprises three genetically disparate sources of tissue, is therefore termed Trimera. This study was aimed at assessing the effect of human IL-6 on HBV infection in vivo in Trimera mice. Methods. Trimera mice were transplanted with human liver tissue fragments or with HepG2-derived cell lines, which had been previously infected ex vivo with HBV in the presence or absence of human interleukin-6 (hIL-6) and in the presence of anti-IL-6-neutralizing antibodies. Results. HBV sequences appeared in the sera of animals in which the liver tissue was incubated with both HBV and hIL-6 prior to transplantation. A similar result was obtained when a human hepatoblastoma cell line (HepG2), expressing the hIL-6 receptor, was infected ex vivo with HBV in the presence of hIL-6 prior to their injection into spleens of Trimera mice. However, when liver fragments were infected ex vivo and simultaneously treated with neutralizing antibodies against hIL-6 or were incubated with HBV prior to transplantation without hIL-6, the rate of mice positive for HBV DNA in their sera was lower. Human mononuclear cells are also permissive for HBV infection in vitro: in the presence of hIL-6 the infection of these cells is enhanced; and this infection is suppressed by the chimeric protein named Hyper-IL-6, generated by the fusion of hIL-6 to the soluble hIL-6 receptor (sIL-6Rα, gp80). Conclusion. hIL-6 facilitates HBV infection in vitro and in vivo.     

Genetic background effects on dental and other craniofacial abnormalities in homozygous small eye ( Pax6 Sey /Pax6 Sey ) mice

 Small eye (Pax6 ^Sey ) is a semi-dominant mutation affecting development of the eyes, brain and nasal structures. The mutant phenotype arises from defects within the Pax6 gene and several mutant alleles have been identified. A previous study reported that Pax6 ^Sey /Pax6 ^Sey homozygotes, in a random-bred stock, had a median cartilaginous rod-like structure in the nasal region and 80% had supernumerary upper incisor teeth. In this study we show that supernumerary upper incisor teeth and a previously unreported nasal capsule-derived cartilaginous ’spur’ occur in compound heterozygous Pax6 ^Sey-Neu /Pax6 ^Sey and homozygous Pax6 ^Sey /Pax6 ^Sey fetuses from several strains of mice. The frequencies of the abnormal phenotypes were not related to allele type but showed variable penetrance, which was dependent on genetic background. The median nasal cartilaginous rod-like structure was present in all homozygous small eye fetuses. The Pax6 ^Sey /Pax6 ^Sey homozygote may provide insight into the complex gene interactions involved in eye, nasal and craniofacial morphogenesis. Accepted: 28 April 1997     

Use of Fractional Analysis for Evaluation of Liver Structure and Function in Rats In Vivo

Fractional and morphometrical analysis of images obtained by life-time photo- and videorecording of microhemocirculatory changes in the rats in health, experimental cirrhosis, and variants of its treatment was carried out. Differences in the fractional dimensions of the studied organs were significant and correlated with morphological values. This suggests fractional analysis for the diagnosis and prediction of hepatic tissue status in vivo. Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 147, No. 2, pp. 237–240, February, 2009 An erratum to this article can be found at     

Role of Neuronal NMDA and non-NMDA Glutamate Receptors in Medial Vestibular Nucleus in the Regulation of Respiratory Rhythmogenesis in Newborn Rats In Vitro

We studied isolated pontobulbospinal preparations from newborn rat brain. In the early postnatal period, the rostral part of the medial vestibular nucleus produces a potent inhibitory effect on neuronal structures of the bulbar respiratory center via the glutamatergic system. Microinjection of L-glutamate (50 mmol/liter) into the rostral part of the vestibular nucleus completely blocks respiratory rhythmogenesis in 0-1-day-old rat pups and reduced the frequency of generation of inspiratory discharges in 2-3-day-old rats from 8.42±0.68 to 2.68±0.32 min^—1. It was found that the leading role in the mechanism of glutamatergic modulation of the respiratory rhythmogenesis by neurons of the medial vestibular nucleus is played by NMDA and, to a lesser extent, non-NMDA glutamate receptors.     

Phagocytic Activity of Macrophages against Liposomes with Conjugates of Oxidized Dextrans and Isonicotinic Acid Hydrazide during Modeling of Phagocytosis Disturbances In Vitro

We studied phagocytic activity of macrophages against molecular-liposome hybrid compositions consisting of liposomes (diameter 200–450 nm) containing oxidized dextrans with a molecular weight of 35 or 60 kDa conjugated with the basic antituberculosis preparation isonicotinic acid hydrazide (dextrazides) during modeling of various disturbances of endocytosis function of phagocytic cells in vitro. Preincubation of macrophages with trypsin, colchicine, or sodium azide did not change the parameters of adhesion of molecular-liposome hybrid compositions to macrophages. It was found that preincubation of cells with colchicine or sodium azide reduced parameters of phagocytosis of the molecular-liposome hybrid compositions; this reduction did not depend on the molecular weight of dextrans entering the composition of the molecular-liposome hybrid compositions.     

分子标记及其在生物遗传多样性研究中的应用

遗传标记(源于DNA序列差异的特征)可以通过很多不同的途径显示出来。随着生化与分子生物技术的发展完善,可以从分子水平上直接显示它的变化-分子标记。分子标记技术具有可靠性,高效性, 因而被广泛应用于多种研究领域。分子标记通常包括同工酶(或蛋白质)和DNA标记。本文综述生化和DNA标记(蛋白/酶,RFLP, RAPD, AFLP, SSR,SNP, SSLP等)技术的原理、发展及应用。     

Analysis of the p16INK4, p14ARF, p15, TP53, and MDM2 Genes and Their Prognostic Implications in Osteosarcoma and Ewing Sarcoma

We examined alterations of the p16INK4, p14ARF, p15, TP53, and MDM2 genes in 30 osteosarcomas and 24 Ewing sarcomas. Among 21 osteosarcomas and 24 Ewing sarcomas, p16INK4, p14ARF, and p15 abnormalities were found in 4 (19%), 2 (9%), and 3 (14%) osteosarcomas, respectively, and in 4 (17%), 3 (13%), and 4 (17%) Ewing sarcomas, respectively. The alterations of p16INK4, p14ARF, and p15 included homozygous deletions spanning all 3 genes, methylation of p16INK4 or p15, and a nonsense mutation of p16INK4, which simultaneously caused a missense mutation of p14ARF. Alterations of TP53 were found in 15 (50%) of 30 osteosarcomas and 1 (3%) of 24 Ewing sarcomas. None of the sarcomas showed MDM2 amplification. While TP53 abnormalities were far more frequent in osteosarcoma than in Ewing sarcoma, alterations of p16INK4, p14ARF, and p15 were present at similar frequencies in the two types of sarcoma. The event-free survival (EFS) was worse in Ewing sarcoma patients with p16INK4 and p14ARF mutation/deletion than in those without the mutation/deletion (P = 0.019), and EFS was worse in osteosarcoma patients with TP53 alterations than in those without TP53 alterations (P = 0.048). The different incidence of TP53 abnormalities in the 2 types of sarcoma may reflect differences of the molecular processes through which the 2 types of tumor develop.     

Influences of polymorphic variants of DRD2 and SLC6A3 genes, and their combinations on smoking in Polish population

Background  

Polymorphisms in dopaminergic genes may influence cigarette smoking by their potential impact on dopamine reward pathway function. A1 allele of DRD2 gene is associated with a reduced dopamine D2 receptor density, and it has been hypothesised that A1 carriers are more vulnerable to smoking. In turn, the 9-repeat allele of dopamine transporter gene (SLC6A3) has been associated with a substantial reduction in dopamine transporter, what might result in the higher level of dopamine in the synaptic cleft, and thereby protective role of this allele from smoking. In the present study we investigated whether polymorphic variants of DRD2 and SLC6A3 genes and their combinations are associated with the smoking habit in the Polish population.     

Interferon β2/B-Cell Stimulating Factor 2/Interleukin 6 Affects Glycosylation of Acute Phase Proteins in Human Hepatoma Cell Lines

Undefined monocyte-derived cytokines have previously been shown to affect glycan processing in glycoproteins secreted by human hepatoma cell lines. Hep 3B cells, when incubated with the cytokine interferon beta 2/B-cell stimulating factor 2/interleukin 6, secreted forms of alpha 1-protease inhibitor, ceruloplasmin, and alpha-fetoprotein with increased reactivity with concanavalin A (Con A) while incubation of Hep G2 cells with this cytokine led to secretion of forms of these proteins with decreased reactivity with Con A, reflecting changes in their oligosaccharide chains. The difference in response of these two transformed cell lines to this cytokine undoubtedly reflects differences in their intracellular glycan processing mechanisms. Changes in glycosylation patterns were dissociated from changes in rate of synthesis: this cytokine caused increased synthesis of alpha 1-protease inhibitor and ceruloplasmin, and decreased synthesis of alpha-fetoprotein in both cell lines.     

Lifestyle Modifies the Relationship Between Body Composition and Adrenergic Receptor Genetic Polymorphisms, ADRB2, ADRB3 and ADRA2B: A Secondary Analysis of a Randomized Controlled Trial of Physical Activity Among Postmenopausal Women

Genetic variations in the adrenergic receptor (ADR) have been associated with body composition in cross-sectional studies. Recent findings suggest that ADR variants may also modify body composition response to lifestyle. We assessed the role of ADR variants in body composition response to 12 months of resistance training versus control in previously sedentary postmenopausal women. Randomized trial completers were genotyped for A2B ^Glu9/12 by fragment length analysis, and B2 ^Gln27Glu and B3 ^Trp64Arg by TaqMan (n = 148, 54% hormone therapy users). Associations between genotypes and body composition, by dual energy X-ray absorptiometry, were analyzed using univariate models. There was no main effect of individual genes on change in body composition, however, gene × exercise interactions were observed for A2B ^Glu9/12 and B2 ^Gln27Glu on change in lean soft tissue (LST, p = 0.02); exercisers on the A2B ^Glu9− background gained LST compared to a loss among controls over 12 months (p < 0.05), with no significant intervention effect on the A2B ^Glu9+ background. Similarly, there was a significant LST gain with exercise on the B2 ^Glu27+ background compared to loss among controls and no intervention effect on the B2 ^Glu27− background. A non-significant association between total body fat (TBF) and B3 ^Trp64Arg persisted among sedentary controls only when intervention groups were separated (%TBF gain with B3 ^Arg64+ carriage, p = 0.03); exercisers lost TBF regardless of genotype. In summary, effect modification by lifestyle was demonstrated on ADRA2B, B2, and B3 genetic backgrounds. Individuals with certain ADR genotypes may be more vulnerable to adverse changes in body composition with sedentary behavior, thus these candidate genes warrant further study.     

Molecular detection and identification of Ehrlichia and Anaplasma species in ixodid ticks

A polymerase chain reaction (PCR) assay followed by partial sequencing of the 16S ribosomal RNA gene was performed for the presence of Ehrlichia and/or Anaplasma. A total of 242 ixodid ticks were collected from domestic ruminants and their shelters, as well as humans, and their individual salivary glands were dissected out for DNA. From the 242 ticks analyzed, six (2.47%), comprising three Hyalomma anatolicum anatolicum, one Rhipicephalus bursa, and two Rhipicephalus sanguineus, were positive. Of these sequenced samples directly obtained from the PCR products, three sequences from H. a. anatolicum were identical to that of the gene of Ehrlichia spp. strains. One sequence identified in R. bursa was closely related to Anaplasma platys. The remaining two sequences detected in R. sanguineus were similar to that of the gene of Anaplasma ovis. The study presented here provides preliminary data regarding the presence of rickettsial pathogens in ticks in Turkey. Nucleotide sequence data reported in this paper are available in GenBank, EMBL, and DDBJ databases under accession numbers EU191227–EU191231 and EU191233.     

Comparative Analysis of Bone Marrow-derived Mast Cell Differentiation in C57BL/6 and BALB/c Mice

Allergic diseases have increased in the last three decades. Mast cells play a critical role in allergic diseases along with allergen-specific immunoglobulin E (IgE). Following mast cell degranulation elicited by ligation of the IgE-FcεRI receptor complex with allergen, allergic reactions are followed by various symptoms such as vascular hyperpermeability, mucous secretion, itching, sneezing, wheezing, rashes, fever, and anaphylactic shock. Susceptibility or inclination to allergy varies depending on individual genetic traits and living environment, and it has long been believed that such an inclination is determined by an immunologic balance of T helper cell types.

Mouse strains also have different susceptibilities to allergy. Similar to T helper cells and macrophages, it is not known whether mast cells can also be divided into two different types between mouse strains. In this study, we prepared bone marrow-derived mast cells from BALB/c and C57BL/6 mice and examined their cellular properties. Cellular response to IL-3 and the process of mast cell differentiation from bone marrow cells were different on the basis of cell surface marker molecules. BALB/c-derived cells more efficiently exhibited degranulation than did C57BL/6-derived cells following both calcium ionophore and receptor crosslinking.

These functional differences persisted even after a longer cell culture for 8 weeks, suggesting a difference in cell-autonomous characteristics. These results support the concept that mast cells also have different cell types dependent on their genetic background.

Abbreviations: Ab: antibody; BMMC: bone marrow-derived mast cell; DNP: dinitrophenyl; FACS: fluorescence-activated cell sorter; FCS: fetal calf serum; FITC: fluorescein isothiocyanate; FSC: forward scatter; HRP: horseradish peroxidase; HSA: human serum albumin; Ig: immunoglobulin; IL: interleukin; MIP-2: macrophage inflammatory protein-2; MCP: mast cell protease; PE: phycoerythrin; PerCP: Peridinin chlorophyll protein complex; SNP: single nucleotide polymorphisms; SSC: side scatter; Th: T helper; TNF-α: tumor necrosis factor alpha     

Multilocus Microsatellite Markers for Molecular Typing of Candida glabrata: Application to Analysis of Genetic Relationships between Bloodstream and Digestive System Isolates

Candida glabrata has emerged as the second most common etiologic agent, after Candida albicans, of superficial and invasive candidiasis in adults. Strain typing is essential for epidemiological investigation, but easy-to-use and reliable typing methods are still lacking. We report the use of a multilocus microsatellite typing method with a set of eight markers on a panel of 180 strains, including 136 blood isolates from hospitalized patients and 34 digestive tract isolates from nonhospitalized patients. A total of 44 different alleles were observed, generating 87 distinct genotypes. In addition to perfect reproducibility, typing ability, and stability, the method had a discriminatory power calculated at 0.97 when all 8 markers were associated, making it suitable for tracing strains. In addition, it is shown that digestive tract isolates differed from blood culture isolates by exhibiting a higher genotypic diversity associated with different allelic frequencies and preferentially did not group in clonal complexes (CCs). The demonstration of the occurrence of microevolution in digestive strains supports the idea that C. glabrata can be a persistent commensal of the human gut.During the last decade, incidence of invasive candidiasis with non-albicans Candida species has increased (40, 46, 48). Candida glabrata is the most significant species that has emerged and now regularly ranks number two, after C. albicans, as the etiologic agent of superficial and invasive candidiasis occurring in adults (17, 27). Reasons for this change in species distribution remain uncertain but may be partially due to the natural resistance of C. glabrata to azole derivates, widely used since the 1980s (6, 10, 26). This lower susceptibility of C. glabrata to azoles (2, 23, 38) and delayed initiation of therapy due to delayed diagnosis (2, 20, 35) may explain in part why the prognosis for C. glabrata candidemia is worse than that for C. albicans candidemia (7, 18, 41, 51).Currently, despite increasing clinical concern, the epidemiology of C. glabrata remains poorly known compared to that of C. albicans. While C. glabrata is considered a commensal of the human digestive tract, its natural reservoir is still uncertain. Possible transmission between patients has been suggested (43), and clusters of invasive infections have been reported (4, 36). An efficient and easy-to-use molecular typing method which would allow tracing strains would also allow better understanding of the spread of this species, notably in a hospital context. Up to now, the most reliable method for C. glabrata typing was Southern blotting with moderately repeated probes (3, 32). However, this method requires the use of radioactive elements, and the sequences of the probes are not available. Sequence-based methods such as multilocus sequence typing (MLST) and multilocus microsatellite analysis have been recently proposed (15, 19, 21), but in all cases one could hope for better discriminatory power.A microsatellite-based typing method using a new set of eight markers has been set up for population genetic analysis of C. glabrata (14). In this work, we positively evaluate this method, which has a discriminatory power calculated at 0.97, for tracing C. glabrata strains. The results of typing a large panel of both blood culture and digestive tract isolates suggest (i) that C. glabrata can undergo microevolution within the gut, supporting a persistent commensal life cycle of C. glabrata, and (ii) that digestive and bloodstream isolates exhibit genetic diversity preferentially induced by polymorphism in some loci.     

Detection and characterization of a ST6 clone of vanB2-Enterococcus faecalis from three different hospitals in Spain

Thirteen vancomycin-resistant and teicoplanin-susceptible Enterococcus faecalis isolates were recovered from unrelated patients in three Spanish hospitals from November 2009 to December 2010. All isolates carried the vanB2 gene, showed indistinguishable or closely-related PFGE patterns and were ascribed to the sequence type ST6 (included into the high-risk clonal-complex CC2). They showed a multiresistance phenotype (erythromycin, tetracycline, ciprofloxacin and high-level-resistance to streptomycin, gentamicin and kanamycin) and harboured the aac(6’)-aph(2”), ant(6)-Ia, and tet(M)+/−tet(L) genes. All isolates produced gelatinase and harboured the gelE gene, but not the esp or hyl genes. The inclusion of the vanB2 gene into the Tn5382 transposon was demonstrated in one isolate. Clonal dissemination of vanB2-containing the E. faecalis strain is demonstrated.