Low-Input RNA-Sequencing in Patients with Cartilage Lesions, Osteoarthritis, and Healthy Cartilage.

OBJECTIVE: To analyze and compare cartilage samples from 3 groups of patients utilizing low-input RNA-sequencing. DESIGN: Cartilage biopsies were collected from patients in 3 groups (n = 48): Cartilage lesion (CL) patients had at least ICRS grade 2, osteoarthritis (OA) samples were taken from patients undergoing knee replacement, and healthy cartilage (HC) was taken from ACL-reconstruction patients without CLs. RNA was isolated using an optimized protocol. RNA samples were assessed for quality and sequenced with a low-input SmartSeq2 protocol. RESULTS: RNA isolation yielded 48 samples with sufficient quality for sequencing. After quality control, 13 samples in the OA group, 9 in the HC group, and 9 in the CL group were included in the analysis. There was a high degree of co-clustering between the HC and CL groups with only 6 genes significantly up- or downregulated. OA and the combined HC/CL group clustered significantly separate from each other, yielding 659 significantly upregulated and 1,369 downregulated genes. GO-term analysis revealed that genes matched to cartilage and connective tissue development terms. CONCLUSION: The gene expression profiles from the 3 groups suggest that there are no major differences in gene expression between cartilage from knees with a cartilage injury and knees without an apparent cartilage injury. OA cartilage, as expected, showed markedly different gene expression from the other 2 groups. The gene expression profiles resulting from this low-input RNA-sequencing study offer opportunities to discover new pathways not previously recognized that may be explored in future studies.

Molecular cloning of MDA5, phylogenetic analysis of RIG-I-like receptors (RLRs) and differential gene expression of RLRs, interferons and proinflammatory cytokines after in vitro challenge with IPNV, ISAV and SAV in the salmonid cell line TO.

The RIG-I receptors RIG-I, MDA5 and LGP2 are involved in viral recognition, and they have different ligand specificity and recognize different viruses. Activation of RIG-I-like receptors (RLRs) leads to production of cytokines essential for antiviral immunity. In fish, most research has focused on interferons, and less is known about the production of proinflammatory cytokines during viral infections. In this study, we have cloned the full-length MDA5 sequence in Atlantic salmon, and compared it with RIG-I and LGP2. Further, the salmonid cell line TO was infected with three fish pathogenic viruses, infectious pancreatic necrosis virus (IPNV), infectious salmon anaemia virus (ISAV) and salmonid alphavirus (SAV), and differential gene expression (DEG) analyses of RLRs, interferons (IFNa-d) and proinflammatory cytokines (TNF-α1, TNF-α2, IL-1ß, IL-6, IL-12 p40s) were performed. The DEG analyses showed that the responses of proinflammatory cytokines in TO cells infected with IPNV and ISAV were profoundly different from SAV-infected cells. In the two aforementioned, TNF-α1 and TNF-α2 were highly upregulated, while in SAV-infected cells these cytokines were downregulated. Knowledge of virus recognition by the host and the immune responses during infection may help elucidate why and how some viruses can escape the immune system. Such knowledge is useful for the development of immune prophylactic measures.

Amendment of arsenic and chromium polluted soil from wood preservation by iron residues from water treatment.

An iron-rich water treatment residue (WTR) consisting mainly of ferrihydrite was used for immobilization of arsenic and chromium in a soil contaminated by wood preservatives. A leaching batch experiment was conducted using two soils, a highly contaminated soil (1033 mg kg(-1) As and 371 mg kg(-1) Cr) and slightly contaminated soil (22 5mg kg(-1) As and 27 mg kg(-1) Cr). Compared to an untreated reference soil, amendment with 5% WTR reduced leaching in the highly contaminated soil by 91% for Cr and 98% for As. No aging effect was observed after 103 d. In a small field experiment, soil was mixed with 2.5% WTR in situ. Pore water was extracted during 3 years from the amended soil and a control site. Pore water arsenic concentrations in the amended soil were more than two orders of magnitude lower than in the control for the upper samplers. An increased release of arsenic was observed during winter in both fields, mostly in the deepest samplers. This is likely due to the formation of a pseudo-gley because of precipitation surplus. Stabilization of arsenic and chromium contaminated soil using WTR is a promising method but the transformation of ferrihydrite in soil proves a concern in case of waterlogged soils. Still the amendment minimized the leaching of arsenic, even in cases of seasonal releases.

An atomic force microscopy study of the interactions between indolicidin and supported planar bilayers.

Indolicidin, a tryptophane-rich antimicrobial peptide, was used to investigate the interactions with a zwitterionic phosphatidylcholine as a model membrane system. In situ atomic force microscopy in liquid medium and phosphatidylcholine supported planar bilayers enabled the study of the interactions between indolicidin and the lipid membrane in real time. It was evident that indolicidin induced a continuous shrinking and thinning of the supported planar bilayers. The effect of indolicidin was dependent upon the composition and physical properties of the membrane bilayer. The interaction of indolicidine with the membrane could be best and most pronounced seen and studied at the boundary of the gel-fluid-domains. Dye leakage experiments with phosphatidylcholine vesicles encapsulated with calcein revealed that indolicidin induced fluidisation of the lipid membrane leading to dye release. The present study indicates that the mode of action for indolicidin can be best described by a stepwise interaction of the peptide with the membrane. Formation of pores however can not be supported on the basis of our experiments.

Bone marrow mesenchymal stem cells in a hyaluronan scaffold for treatment of an osteochondral defect in a rabbit model.

The purpose of this study was to evaluate the efficiency of using mesenchymal stem cells (MSC) in a hyaluronan scaffold for repair of an osteochondral defect in rabbit knee. Bone marrow was harvested from the posterior iliac crest in 11 New Zealand White rabbits. MSC were isolated and cultured in autologous serum for 28 days and transferred to a hyaluronan scaffold 24 h prior to implantation. A 4 mm diameter and 1.5 mm deep defect was created in the medial femoral condyle of both knees and the scaffold with MSC was implanted in one knee while an empty scaffold was implanted in the contra-lateral knee. After 24 weeks the rabbits were killed and histological sections were subjected to semiquantitative and quantitative evaluation by observers blinded regarding treatment modality. High degree of filling was obtained, but there was no statistically significant difference between the two treatments. However, there was a tendency for a better quality of repair in the MSC treated knees. No hypertrophy was observed by either method. MSC in a hyaluronan scaffold may be a promising treatment approach, but further studies are needed to determine the best combination of scaffold and cells.

Dynamics of reductive TCE dechlorination in two distinct H(2) supply scenarios and at various temperatures.

Anaerobic microbial dechlorination of trichloroethene (TCE) by a mixed, Dehalococcoides containing culture was investigated at different temperatures (4-60 degrees C) using propionate and lactate as a slow- and fast-releasing hydrogen (H(2)) source, respectively. Distinct temperature-dependent dynamics of substrate fermentation and H(2) levels could explain observed patterns of dechlorination. While varying the temperature caused changes in rate, the overall pattern of dechlorination was characteristic of the supplied electron donor. Feeding cultures with a rapidly fermentable substrate such as lactate generally resulted in high H(2) concentrations and fast and complete dechlorination accompanied by rapid methanogenesis. In contrast, low H(2) release rates resulting from fermentation of propionate were associated with 2 to 3-fold longer time frames necessary for complete dechlorination at intermediate temperatures (15-30 degrees C). A lag-phase prior to dechlorination of cis-dichloroethene (cDCE), together with a characteristic build-up of H(2) and methane, was consistently observed at slow H(2) supply. At temperatures of 10 degrees C and lower, the system remained in this lag phase and no dechlorination past cDCE was observed within the experimental time frame. However, when lactate was the substrate, complete dechlorination of TCE occurred within 74 days at 10 degrees C, accompanied by methane production. The choice of fermentable substrate decisively influenced the rate and degree of dechlorination at an electron donor/TCE ratio as high as 666:1. Temperature-dependent H(2) levels resulting from fermentation of different substrates could be satisfactorily explained through thermodynamic calculations of the Gibbs free energy yield assuming a constant metabolic energy threshold of -20 kJ/(mol reaction).

Filtration through nylon membranes negatively affects analysis of arsenic and phosphate by the molybdenum blue method.

Filtering synthetic arsenic- or phosphate-containing solutions (1.5-47.6mumol/L) with nylon syringe filters significantly reduced absorbances (by 6-74%) when analyzed with the colorimetric molybdenum blue method. Filtering the same solutions with cellulose acetate syringe filters yielded no significant differences as compared to unfiltered controls. The detrimental effect of nylon membranes was also observed when pure Milli-Q water was filtered and subsequently spiked with arsenic(III) or phosphate suggesting that some compound(s) eluting from the filter membranes interfere with the color formation in the assay. Consequently, we caution against using nylon filters when filtering water samples for the determination of arsenic or phosphate with the molybdenum blue method.

Tracer test with As(V) under variable redox conditions controlling arsenic transport in the presence of elevated ferrous iron concentrations.

To study transport and reactions of arsenic under field conditions, a small-scale tracer test was performed in an anoxic, iron-reducing zone of a sandy aquifer at the USGS research site on Cape Cod, Massachusetts, USA. For four weeks, a stream of groundwater with added As(V) (6.7 muM) and bromide (1.6 mM), was injected in order to observe the reduction of As(V) to As(III). Breakthrough of bromide (Br(-)), As(V), and As(III) as well as additional parameters characterizing the geochemical conditions was observed at various locations downstream of the injection well over a period of 104 days. After a short lag period, nitrate and dissolved oxygen from the injectate oxidized ferrous iron and As(V) became bound to the freshly formed hydrous iron oxides. Approximately one week after terminating the injection, anoxic conditions had been reestablished and increases in As(III) concentrations were observed within 1 m of the injection. During the observation period, As(III) and As(V) were transported to a distance of 4.5 m downgradient indicating significant retardation by sorption processes for both species. Sediment assays as well as elevated concentrations of hydrogen reflected the presence of As(V) reducing microorganisms. Thus, microbial As(V) reduction was thought to be one major process driving the release of As(III) during the tracer test in the Cape Cod aquifer.

A salmonid cell line (TO) for production of infectious salmon anaemia virus (ISAV).

A new cell line designated TO which provides a high yield of infectious salmon anaemia virus (ISAV) has been established. The cells originate from head kidney leukocytes isolated from Atlantic salmon and grow well at 20 degrees C in EMEM with 5% CO2 and without CO2 supplement in HMEM. The cells have at present been passed more than 150 times and no changes in morphology, growth or virus production have been observed. The virus infection results in cytopathic effects (CPE) within 9 d, and the virus titre obtained from centrifuged and filtrated cell lysates, measured as TCID50, was about 10(9.1) ml(-1). The virus isolated from lysates of infected cells by a sucrose gradient provided purified ISAV when examined by silver stained SDS-PAGE. Salmon injected with diluted virus supernatant showed mortalities, hematocrit values and clinical signs in accordance with infectious salmon anaemia.

The biphasic expression of IMP/Vg1-RBP is conserved between vertebrates and Drosophila.

The human IGF-II mRNA-binding proteins (IMPs) 1-3, and their Xenopus homologue Vg1 RNA-binding protein (Vg1-RBP) are RNA-binding proteins implicated in mRNA localization and translational control in vertebrate development. We have sequenced the Drosophila homologue (dIMP) of these genes, and examined its expression pattern in Drosophila embryos by in situ hybridization. The study shows that dIMP exhibits a biphasic expression pattern. In the early stages of development, a maternal pool of dIMP mRNA is evenly distributed in the embryo and degraded by the end of stage 4. Expression reappears in the developing central nervous system, where dIMP is expressed throughout neurogenesis. In addition, dIMP is present in the pole cells.

Stability of dental health behavior: a 3-year prospective cohort study of 15-, 16- and 18-year-old Norwegian adolescents.

UNLABELLED: There is a need for a descriptive epidemiology of patterns of dental health behavior through adolescence. OBJECTIVES: The purpose of this study was to assess the tracking (degree of stability) of several categories of self-reported dental health behavior in adolescence over a 3-year period. METHODS: In 1992, a representative sample of 970 15-year-old adolescents of Hordaland county, Norway, completed questionnaires under supervision at school. Postal follow-up studies 1 and 3 years later provided 709 and 781 answers, respectively. The present analyses included 581 adolescents who participated on all three occasions and 670 adolescents who replied in 1992 and 1995. Paired sample t-tests, cross-tabulations and Pearson's correlation coefficients were used to assess stability. For use in the cross-tabulations all variables were transformed into binary categories (active and inactive). RESULTS: A substantial proportion of adolescents remained consistently in the same behavioral category for all 3 survey years. Toothbrushing (80%) and use of dental floss (64%) were the most stable activities, while consumption of sugared mineral water (53%), intake of chocolate/sweets (52%) and use of F-rinse (54%) were less stable. However, Pearson's correlations between corresponding behaviors assessed in 1992 and 1995 were moderate and varied from 0.59 (toothbrushing) to 0.22 (F-rinse) (P<0.001). CONCLUSION: The results do not conclusively indicate a pattern of tracking. The tendency is, however, clear enough to support the assumption that dental health behavior continues during adolescence into adulthood.

Two-year longitudinal study of alcohol expectancies and drinking among Norwegian adolescents.

AIMS: The potential importance of alcohol outcome expectancies in the initiation and maintenance of drinking has been supported by studies showing that these expectancies are present before drinking begins, and that they predict drinking both cross-sectionally and longitudinally. Although initiation of drinking behavior may be influenced by expectancy, subsequent drinking experience may modify expectations. We used structural modeling techniques to investigate the relative influence of expectancy and drinking in a three-wave longitudinal study of Norwegian adolescents. DESIGN: Survey incorporating self-administered questionnaires. SETTING: Twenty-two schools in Hordaland County on the west coast of Norway. PARTICIPANTS: Nine hundred and twenty-four seventh-grade students; 45.7% female. MEASUREMENTS: Alcohol use (frequency, quantity, drunkenness); the Norwegian version of the Alcohol Expectancy Questionnaire for adolescents. FINDINGS: Among students who were already drinkers upon entry into the study, expectations of positive social effects of alcohol predicted drinking longitudinally. Among those who began drinking during the study, these social expectancies predicted drinking initiation, but drinking also influenced subsequent expectancy in the early stages of drinking. CONCLUSIONS: These results support a reciprocal relationship of drinking to positive expectancy, highlighting the importance both of expectancies on influencing drinking, and of early drinking experiences on the development of positive expectancies.

Noncoital sexual interactions and problem behaviour among young adolescents: the Norwegian Longitudinal Health Behaviour Study.

This paper focuses on whether early onset of noncoital sexual interactions is best considered in the same paradigm as adolescent problem behaviour. Among adolescents (n = 927) followed longitudinally from age 13 to 15, the interrelationships between noncoital sexual interactions and various problem and conventional behaviours were examined. Early onset of noncoital sexual interactions was related to problem behaviour, and the higher ordered the noncoital sexual interaction was, the more strongly it correlated with measures of problem behaviour. This study provides support for the usefulness of the notion of an underlying factor of proneness to problem behaviours, which is relevant when studying developmental change in terms of accumulation of early noncoital sexual experience. It is suggested that in the construction of countermeasures to prevent the negative consequences of early sexual activities among adolescents, it may be useful to target broader correlates of behaviour.

Adolescents' Knowledge about HIV/AIDS: One Dimensional or Two Dimensional? A Confirmatory Factor Analysis.

This article sets out to study the dimensionality of adolescents' knowledge of HIV transmission by contrasting a two-factor and a one-factor model using confirmatory factor analysis (LISREL). The empirical data stem from a longitudinal study on health behaviour in Norway. This study uses cross-sectional data of the same subjects in 1990 with mean age 13.3 (n = 927) and 1992 ( n = 974). The respondents were confronted with 12 items describing different types of risky situations and asked whether HIV could be transmitted in these situations. The two-factor model provided a significantly better fit to the empirical data than did the one-factor model, suggesting that knowledge of how HIV is transmitted is distinct from knowledge of how HIV is not transmitted. Several explanations for this difference are suggested, and implications for constructing health- education messages are discussed.

The effect of parental dental health behavior on that of their adolescent offspring.

This study was designed to investigate the effect of parental dental health behavior on that of their adolescent offspring. The data stemmed from The Norwegian Longitudinal Health Behavior Study performed in 1993 in the County of Hordaland in Norway and comprised separate questionnaires for both parents and a 16-year-old child in 436 family units. A compound measure of parental dental health behavior was applied in logistic regression analyses. The results showed that there were statistically significant associations of use of dental floss, tooth brushing, and drinking of non-sugared mineral water among parents and their adolescent offspring. No significant gender interaction in the association between same- or different-sex parent-adolescent dyads was observed. These findings indicate that parents function as social models for their offspring well into the adolescent period with regard to several dental health behaviors.

Predicting 13-year-olds' drinking using parents' self-reported alcohol use and restrictiveness compared with offspring's perception.

This study examined whether offspring's perception of parental frequency of alcohol use and restrictiveness towards offspring's alcohol use could be used as a substitute for parental self-reports. Offspring's perception were compared with parents' self-reported alcohol use and restrictiveness in ability to predict offspring's own alcohol use. Respondents were 924 offspring, 642 fathers and 729 mothers. Correlations between fathers' and mothers' self-reported frequency of drinking alcohol and offspring's report of parental frequency of drinking alcohol ranged from 0.55 to 0.70. Fathers' and mothers' self-reported alcohol use and restrictiveness explained less than two per cent of the variance in their offspring's alcohol use in regression analyses, while offspring's perception of parental drinking frequency and restrictiveness explained about eight per cent of the variance in offspring's alcohol use. These results represent a challenge to studies of young adolescents which interpret offspring's reports of parents' alcohol use as a surrogate measure of parents' self-reported alcohol use.

Do adolescents and parents report each other's physical activity accurately?

In research concerning socialization into sports there is a need to assess children's reports of parental physical activity. Reports by young adolescents and their parents of each other's leisure-time physical activity and related issues were compared with self-reported measures. A total of 755 family units in western Norway, consisting of 755 13-year-olds, mean age 13.3 +/- 0.3 years (425 boys, 330 girls, 644 fathers and 732 mothers) completed a self-administered physical activity questionnaire. Pearson's between self-reports by parents and the reports by their children varied between 0.56 (P < 0.001) and 0.07 (nonsignificant). Pearson's r between off-springs' self-reported physical activity and reports by their parents varied between 0.41 and 0.55 (P < 0.001). The results support cautious use of young adolescents' reports of parental physical activity.

Organization of HIV/AIDS related attitudes and predictors of attitudes.

The purpose of this study was to investigate the psychological organization of attitudes towards public measures instituted to combat AIDS; secondly, to study the cognitive information about HIV transmission upon which these attitudes are based; and thirdly, to study whether education plans and gender also predicted negative attitudes. The study subjects were 229 students attending a Norwegian high-school (median age = 19), and the data were collected using a self-administered questionnaire. Confirmatory factor analysis confirmed the existence of two separate attitudinal dimensions: "Restrictive" and "Supportive". The hypothesis was confirmed that beliefs about uncontrollable contact predicted "Restrictive" attitudes, while beliefs about controllable contracts did not. Educational plans, gender and "Uncontrolable contact" beliefs each predicted "Restrictive" attitudes. This suggests that beliefs and attitudes have instrumental as well as symbolic bases. Implications for public health strategies are discussed.

[The national plan for development projects in disease prevention and health promotion. Background, organization and progress 1989-91]. / Samlet plan for utviklingsprosjekter innen det sykdomsforebyggende og helsefremmende arbeid. Bakgrunn, organisasjon og fremdrift 1989-91.

A five-year national programme for development projects in disease prevention and health promotion in Norwegian municipalities was implemented in 1989. Major goals were to stimulate the municipal health services to increase their health promotion activities and to gain experience of alternative organizational developments, such as inter-municipal and cross-sectoral collaborative health promotion efforts. During the first three years, 43% of Norway's municipalities applied for grants and 26% received them. The smaller municipalities applied less often and received fewer grants than the larger municipalities did. In all, a total of NOK 61 million was granted for 258 projects covering a wide range of individual and environmental health promotion areas. So far, the programme seems to have generated renewed interest for health promotion work in Norwegian municipalities.

Structural analysis of botulinum neurotoxin types A and E in aqueous and nonpolar solvents by Fourier transform infrared, second derivative UV absorption, and circular dichroic spectroscopies.

Two pharmacologically similar but antigenetically distinct botulinum neurotoxins, types A and E with a 1000-fold difference in their toxicity, were examined for nonpolar solvent-induced changes in secondary structures and polypeptide foldings to understand their structural differences and their comparative responsiveness/susceptibility to solvent perturbation. Analysis of far UV circular dichroic spectra in aqueous buffer for types A and E neurotoxins yielded the following: the alpha-helix contents were 27 and 20%; the beta-sheets were 36 and 44%, the beta-turns were 6.0 and 0%, and the random coils were 31 and 36%, respectively. Fourier transform infrared spectra, obtained by using attenuated total reflection technique, indicated high content of alpha-helix and beta-pleated sheet structures for both neurotoxins as judged by strong bands at 1651 and 1633 cm-1 in the amide I frequency region and bands at 1314 and 1245 cm-1 in the amide III frequency region. The peak height ratio of 1314 and 1245 cm-1 bands, suggests that the type A neurotoxin has slightly higher alpha-helical content than the type E neurotoxin. These observations are consistent with the secondary structures estimated from far UV circular dichroic spectra. Fourier transform infrared spectra of the neurotoxins, exposed to methanol, showed sharp increases of the 1651 cm-1 band and a significant increase in the height of the 1314 cm-1 band, suggesting increases in the alpha-helical contents of the proteins. The changes were more in the type A than in the type E neurotoxin. The changes were reversible upon reexposure of the proteins to the aqueous buffer. Second derivative absorption spectroscopy demonstrated that methanol also induced changes in the degree of Tyr exposure to solvent. The results are discussed in terms of structural differences between the single and dichain neurotoxins and in terms of their mode of action.