Men's Acceptance of Screening for Prostate Cancer with Prostate-specific Antigen, Magnetic Resonance Imaging, and Prostate Biopsy.

BACKGROUND: A prerequisite before introducing a screening program is that the screening examinations are acceptable to participants. OBJECTIVE: To evaluate the acceptance and bother of prostate cancer screening examinations. DESIGN, SETTING, AND PARTICIPANTS: The randomized population-based GÖTEBORG-2 prostate cancer screening trial invited >37 000 men for prostate-specific antigen (PSA) testing followed by magnetic resonance imaging (MRI) in case of elevated PSA and prostate biopsy (targeted and/or systematic) if indicated. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Participants were asked to fill out a questionnaire and rate the level of bother associated with each examination (PSA, MRI, and prostate biopsy) on a categorical scale ranging from 1 to 5 (1 = "not at all bothersome" and 5 = "very bothersome"), and to rate their willingness to repeat the examinations, by marking an X on a continuous scale ranging from 0 to 10 (0 = "yes, without any hesitation" and 10 = "no, absolutely not"). Wilcoxon signed rank test was used. RESULTS AND LIMITATIONS: Compliance with MRI was 96% (1790/1872), compliance with biopsy was 89% (810/907), and the response rate to the questionnaire was 75% (608/810). Men who underwent all examinations (n = 577) responded that biopsy was more bothersome than PSA test (p < 0.001) and MRI (p < 0.001). High levels of bother (≥4 out of 5) were reported by 2% (12/577) for PSA test, 8% (46/577) for MRI, and 43% (247/577) for biopsy. Men were more willing to repeat MRI than biopsy (p < 0.001), but the difference was small (median 0.2 [interquartile range 0.1-0.6] vs 0.5 [0.1-2.0]). CONCLUSIONS: Biopsies are more bothersome than MRI, but a large majority of men accept to repeat both examinations if necessary. Omitting biopsy for MRI-negative men and shifting to targeted biopsies only will reduce bother for men participating in prostate cancer screening. PATIENT SUMMARY: We asked men how bothersome they found the prostate-specific antigen (PSA) test, magnetic resonance imaging (MRI), and prostate biopsies. Biopsies were more bothersome than PSA and MRI, but most men were willing to repeat all procedures if necessary.

Men's Perception of Being Invited for Prostate Cancer Testing and the Information About Its Pros and Cons-A Survey from Two Population-based Testing Programmes.

Background: There is no national screening programme for prostate cancer in Sweden. Instead, population-based organised prostate cancer testing (OPT) projects are introduced to make information and testing more equal and effective. Objective: To evaluate men's perception of being invited to OPT and of the information in the invitation letter, and whether their perception is influenced by educational level. Design setting and participants: A questionnaire was sent out to men invited to OPT in 2020: 600 50-yr-old men in Region Västra Götaland and 1000 50-, 56-, and 62-yr-old men in Region Skåne. Outcome measurements and statistical analysis: Responses were evaluated on a Likert scale. The chi-square test was used to compare proportions. Results and limitations: A total of 534 men (34%) responded. Almost all considered the OPT concept as very good (84%) or good (13%). Among men not previously undergone a prostate-specific antigen (PSA) test, a larger proportion with nonacademic (53%) than with academic education (41%) responded that the text about disadvantages was very clear (p = 0.03). A similar difference was observed for the text about advantages (68% vs 58%, p = 0.09). There was no association between education and searching for more information elsewhere. The low response rate is the main limitation. Conclusions: Almost all responding men evaluating the invitation letter for OPT were positive about making a personal decision regarding whether or not to have a PSA test. Most were content with the brief information. Men with academic education were somewhat less likely to find the information very clear. This shows a need for further research about how best to describe the advantages and disadvantages of prostate cancer testing. Patient summary: Almost all men who responded to a questionnaire to evaluate the invitation letter for organised prostate cancer testing were positive about the opportunity to make a personal decision regarding whether or not to have a prostate-specific antigen test.

Mapping geographical areas at risk for tick-borne encephalitis (TBE) by analysing bulk tank milk from Swedish dairy cattle herds for the presence of TBE virus-specific antibodies.

BACKGROUND: The vector-borne human viral zoonosis tick-borne encephalitis (TBE) is of growing concern in Sweden. The area where TBE is considered endemic has expanded, with an increasing geographical distribution of Ixodes ricinus as the tick vector and a rising number of reported TBE cases in humans. Efforts to map TBE risk areas have been carried out by sentinel monitoring, mainly based on individual sampling and analysis of wild and domestic animals, as well as ticks, for tick-borne encephalitis virus (TBEV). However, the interpretation of the geographical distribution has been hampered by the patchy and focal nature of TBEV occurrence. This study presents TBEV surveillance data based on antibody analysis of bulk tank milk collected from dairy herds located throughout Sweden before (May) and after (November) the vector season. A commercial TBEV antibody ELISA was modified and evaluated for use in this study. RESULTS: The initial comparative TBEV antibody analysis revealed a good correlation between milk and serum antibody levels from individually sampled cows. Also, the TBEV-antibody levels for the mean-herd serum showed good comparability with TBEV antibody levels from bulk tank milk, thus indicating good predictability of seroprevalence when analysing bulk tank milk from a herd. Analyses of bulk tank milk samples collected from 616 herds in May and 560 herds in November showed a geographical distribution of TBEV seropositive herds that was largely consistent with reported human TBE cases. A few TBEV-reactive herds were also found outside known locations of human TBE cases. CONCLUSION: Serological examination of bulk tank milk from dairy cattle herds may be a useful sentinel surveillance method to identify geographical presence of TBEV. In contrast to individual sampling this method allows a large number of animals to be monitored. TBEV seropositive herds were mainly found in coastal areas of southern Sweden similar to human TBE cases. However, some antibody-reactive herds were found outside known TBE areas at the time of the study.

MTH1 inhibition eradicates cancer by preventing sanitation of the dNTP pool.

Cancers have dysfunctional redox regulation resulting in reactive oxygen species production, damaging both DNA and free dNTPs. The MTH1 protein sanitizes oxidized dNTP pools to prevent incorporation of damaged bases during DNA replication. Although MTH1 is non-essential in normal cells, we show that cancer cells require MTH1 activity to avoid incorporation of oxidized dNTPs, resulting in DNA damage and cell death. We validate MTH1 as an anticancer target in vivo and describe small molecules TH287 and TH588 as first-in-class nudix hydrolase family inhibitors that potently and selectively engage and inhibit the MTH1 protein in cells. Protein co-crystal structures demonstrate that the inhibitors bind in the active site of MTH1. The inhibitors cause incorporation of oxidized dNTPs in cancer cells, leading to DNA damage, cytotoxicity and therapeutic responses in patient-derived mouse xenografts. This study exemplifies the non-oncogene addiction concept for anticancer treatment and validates MTH1 as being cancer phenotypic lethal.

Structural insight into DNA binding and oligomerization of the multifunctional Cox protein of bacteriophage P2.

The Cox protein from bacteriophage P2 is a small multifunctional DNA-binding protein. It is involved in site-specific recombination leading to P2 prophage excision and functions as a transcriptional repressor of the P2 Pc promoter. Furthermore, it transcriptionally activates the unrelated, defective prophage P4 that depends on phage P2 late gene products for lytic growth. In this article, we have investigated the structural determinants to understand how P2 Cox performs these different functions. We have solved the structure of P2 Cox to 2.4 Å resolution. Interestingly, P2 Cox crystallized in a continuous oligomeric spiral with its DNA-binding helix and wing positioned outwards. The extended C-terminal part of P2 Cox is largely responsible for the oligomerization in the structure. The spacing between the repeating DNA-binding elements along the helical P2 Cox filament is consistent with DNA binding along the filament. Functional analyses of alanine mutants in P2 Cox argue for the importance of key residues for protein function. We here present the first structure from the Cox protein family and, together with previous biochemical observations, propose that P2 Cox achieves its various functions by specific binding of DNA while wrapping the DNA around its helical oligomer.

Organellar oligopeptidase (OOP) provides a complementary pathway for targeting peptide degradation in mitochondria and chloroplasts.

Both mitochondria and chloroplasts contain distinct proteolytic systems for precursor protein processing catalyzed by the mitochondrial and stromal processing peptidases and for the degradation of targeting peptides catalyzed by presequence protease. Here, we have identified and characterized a component of the organellar proteolytic systems in Arabidopsis thaliana, the organellar oligopeptidase, OOP (At5g65620). OOP belongs to the M3A family of peptide-degrading metalloproteases. Using two independent in vivo methods, we show that the protease is dually localized to mitochondria and chloroplasts. Furthermore, we localized the OPP homolog At5g10540 to the cytosol. Analysis of peptide degradation by OOP revealed substrate size restriction from 8 to 23 aa residues. Short mitochondrial targeting peptides (presequence of the ribosomal protein L29 and presequence of 1-aminocyclopropane-1-carboxylic acid deaminase 1) and N- and C-terminal fragments derived from the presequence of the ATPase beta subunit ranging in size from 11 to 20 aa could be degraded. MS analysis showed that OOP does not exhibit a strict cleavage pattern but shows a weak preference for hydrophobic residues (F/L) at the P1 position. The crystal structures of OOP, at 1.8-1.9 Å, exhibit an ellipsoidal shape consisting of two major domains enclosing the catalytic cavity of 3,000 Å(3). The structural and biochemical data suggest that the protein undergoes conformational changes to allow peptide binding and proteolysis. Our results demonstrate the complementary role of OOP in targeting-peptide degradation in mitochondria and chloroplasts.

Crystal structures of botulinum neurotoxin DC in complex with its protein receptors synaptotagmin I and II.

Botulinum neurotoxins (BoNTs) can cause paralysis at exceptionally low concentrations and include seven serotypes (BoNT/A-G). The chimeric BoNT/DC toxin has a receptor binding domain similar to the same region in BoNT/C. However, BoNT/DC does not share protein receptor with BoNT/C. Instead, it shares synaptotagmin (Syt) I and II as receptors with BoNT/B, despite their low sequence similarity. Here, we present the crystal structures of the binding domain of BoNT/DC in complex with the recognition domains of its protein receptors, Syt-I and Syt-II. The structures reveal that BoNT/DC possesses a Syt binding site, distinct from the established Syt-II binding site in BoNT/B. Structure-based mutagenesis further shows that hydrophobic interactions play a key role in Syt binding. The structures suggest that the BoNT/DC ganglioside binding sites are independent of the protein receptor binding site. Our results reveal the remarkable versatility in the receptor recognition of the BoNTs.

Crystal structure of human MTH1 and the 8-oxo-dGMP product complex.

MTH1 hydrolyzes oxidized nucleotide triphosphates, thereby preventing them from being incorporated into DNA. We here present the structures of human MTH1 (1.9Å) and its complex with the product 8-oxo-dGMP (1.8Å). Unexpectedly MTH1 binds the nucleotide in the anti conformation with no direct interaction between the 8-oxo group and the protein. We suggest that the specificity depends on the stabilization of an enol tautomer of the 8-oxo form of dGTP. The binding of the product induces no major structural changes. The structures reveal the mode of nucleotide binding in MTH1 and provide the structural basis for inhibitor design.

Structural determination of functional domains in early B-cell factor (EBF) family of transcription factors reveals similarities to Rel DNA-binding proteins and a novel dimerization motif.

The early B-cell factor (EBF) transcription factors are central regulators of development in several organs and tissues. This protein family shows low sequence similarity to other protein families, which is why structural information for the functional domains of these proteins is crucial to understand their biochemical features. We have used a modular approach to determine the crystal structures of the structured domains in the EBF family. The DNA binding domain reveals a striking resemblance to the DNA binding domains of the Rel homology superfamily of transcription factors but contains a unique zinc binding structure, termed zinc knuckle. Further the EBF proteins contain an IPT/TIG domain and an atypical helix-loop-helix domain with a novel type of dimerization motif. The data presented here provide insights into unique structural features of the EBF proteins and open possibilities for detailed molecular investigations of this important transcription factor family.

Crystal structure of the catalytic domain of human PARP2 in complex with PARP inhibitor ABT-888.

Poly-ADP-ribose polymerases (PARPs) catalyze transfer of ADP-ribose from NAD(+) to specific residues in their substrate proteins or to growing ADP-ribose chains. PARP activity is involved in processes such as chromatin remodeling, transcription control, and DNA repair. Inhibitors of PARP activity may be useful in cancer therapy. PARP2 is the family member that is most similar to PARP1, and the two can act together as heterodimers. We used X-ray crystallography to determine two structures of the catalytic domain of human PARP2: the complexes with PARP inhibitors 3-aminobenzamide and ABT-888. These results contribute to our understanding of structural features and compound properties that can be employed to develop selective inhibitors of human ADP-ribosyltransferases.

Virulotyping and antimicrobial resistance typing of Salmonella enterica serovars relevant to human health in Europe.

The combination of virulence gene and antimicrobial resistance gene typing using DNA arrays is a recently developed genomics-based approach to bacterial molecular epidemiology. We have now applied this technology to 523 Salmonella enterica subsp. enterica strains collected from various host sources and public health and veterinary institutes across nine European countries. The strain set included the five predominant Salmonella serovars isolated in Europe (Enteritidis, Typhimurium, Infantis, Virchow, and Hadar). Initially, these strains were screened for 10 potential virulence factors (avrA, ssaQ, mgtC, siiD, sopB, gipA, sodC1, sopE1, spvC, and bcfC) by polymerase chain reaction. The results indicated that only 14 profiles comprising these genes (virulotypes) were observed throughout Europe. Moreover, most of these virulotypes were restricted to only one (n = 9) or two (n = 4) serovars. The data also indicated that the virulotype did not vary significantly with host source or geographical location. Subsequently, a representative subset of 77 strains was investigated using a microarray designed to detect 102 virulence and 49 resistance determinants. The results confirmed and extended the previous observations using the virulo-polymerase chain reaction screen. Strains belonging to the same serovar grouped together, indicating that the broader virulence-associated gene complement corresponded with the serovar. There were, however, some differences in the virulence gene profiles between strains belonging to an individual serovar. This variation occurred primarily within those virulence genes that were prophage encoded, in fimbrial clusters or in the virulence plasmid. It seems likely that such changes enable Salmonella to adapt to different environmental conditions, which might be reflected in serovar-specific ecology. In this strain subset a number of resistance genes were detected and were serovar restricted to a varying degree. Once again the profiles of those genes encoding resistance were similar or the same for each serovar in all hosts and countries investigated.

Identification of nine sequence types of the 16S rRNA genes of Campylobacter jejuni subsp. jejuni isolated from broilers.

BACKGROUND: Campylobacter is the most commonly reported bacterial cause of enteritis in humans in the EU Member States and other industrialized countries. One significant source of infection is broilers and consumption of undercooked broiler meat. Campylobacter jejuni is the Campylobacter sp. predominantly found in infected humans and colonized broilers. Sequence analysis of the 16S rRNA gene is very useful for identification of bacteria to genus and species level. The objectives in this study were to determine the degree of intraspecific variation in the 16S rRNA genes of C. jejuni and C. coli and to determine whether the 16S rRNA sequence types correlated with genotypes generated by PFGE analysis of SmaI restricted genomic DNA of the strains. METHODS: The 16S rRNA genes of 45 strains of C. jejuni and two C. coli strains isolated from broilers were sequenced and compared with 16S rRNA sequences retrieved from the Ribosomal Database Project or GenBank. The strains were also genotyped by PFGE after digestion with SmaI. RESULTS: Sequence analyses of the 16S rRNA genes revealed nine sequence types of the Campylobacter strains and the similarities between the different sequence types were in the range 99.6-99.9%. The number of nucleotide substitutions varied between one and six among the nine 16S rRNA sequence types. One of the nine 16S rRNA sequence profiles was common to 12 of the strains from our study and two of these were identified as Campylobacter coli by PCR/REA. The other 10 strains were identified as Campylobacter jejuni. Five of the nine sequence types were also found among the Campylobacter sequences deposited in GenBank. The three 16S rRNA genes in the analysed strains were identical within each individual strain for all 47 strains. CONCLUSION: C. jejuni and C. coli seem to lack polymorphisms in their 16S rRNA gene, but phylogenetic analysis based on 16S rRNA sequences was not always sufficient for differentiation between C. jejuni and C. coli. The strains were grouped in two major clusters according to 16S rRNA, one cluster with only C. jejuni and the other with both C. jejuni and C. coli. Genotyping of the 47 strains by PFGE after digestion with SmaI resulted in 22 subtypes. A potential correlation was found between the SmaI profiles and the 16S rRNA sequences, as a certain SmaI type only appeared in one of the two major phylogenetic groups.

A mouse model for in vivo tracking of the major dust mite allergen Der p 2 after inhalation.

Inhaled environmental antigens, i.e. allergens, cause allergic symptoms in millions of patients worldwide. As little is known about the fate of an allergen upon inhalation, we addressed this issue for a major dust mite allergen, Der p 2. First, a model for Der p 2-sensitization was established in C57BL/6 J mice, in which sensitized mice mounted a Der p 2-specific IgE-response with eosinophilic lung inflammation after allergen challenge in the airways. In this model, we applied recombinant Der p 2 carrying a novel C-terminal tetrapeptide Sel-tag enabling labelling with the gamma-emitting radionuclide 75Se at a single selenocysteine residue ([75Se]Der p 2). In vivo tracking of intratracheally administered [75Se]Der p 2 using whole-body autoradiography revealed that [75Se]Der p 2-derived radioactivity persisted in the lungs of sensitized mice as long as 48 h. Radioactivity was also detected in kidneys, liver and in enlarged lung-associated lymph nodes. Interestingly, a larger proportion of radioactivity was found in the lungs of sensitized compared with nonsensitized mice 24 h after intratracheal instillation of [75Se]Der p 2. A radioactive protein corresponding to intact Der p 2 could only be detected in the lungs, whereas [75Se]Der p 2-derived radioactivity was recovered in known selenoproteins both in lung and other organs. Hence, using the recently developed Sel-tag method in a mouse model for Der p 2-sensitization, we could track the fate of an inhaled allergen in vivo. Based upon our findings, we conclude that the inflammatory state of the lung influences the rate of metabolism and clearance of Der p 2. Thus, an allergic response to the inhaled allergen may lead to prolonged retention of Der p 2 in the lung.

Increased levels of hypoxia-sensitive proteins in allergic airway inflammation.

In this study we investigated the alterations in protein levels that are induced by allergic eosinophilic lung inflammation. Lung tissue eosinophilia and sequestration of inflammatory cells in airspaces were provoked by systemic sensitization with ovalbumin followed by repeated inhalation challenge with aerosolized ovalbumin. Proteome alterations in lung tissue and bronchoalveolar lavage fluid, respectively, were examined by two-dimensional gel electrophoresis followed by identification of proteins by mass spectrometry. Several proteins were markedly increased in inflamed tissue. In particular, several proteins that are known to be associated with hypoxia were elevated, for example, glycolytic enzymes, glucose-regulated protein 78 kD, prolyl-4-hydroxylase, peroxiredoxin 1, and arginase. Out of the identified proteins, Ym2 displayed the clearest increase, present at high levels in animals with lung eosinophilia, while being undetectable in control subjects. Furthermore, the levels of cathepsin S were markedly increased in inflamed tissue. Taken together, this study identifies a number of marker proteins associated with the pathogenesis of allergic lung inflammation and indicates a link between allergic airway inflammation and induction of hypoxia-related gene products.

Oxidised LDL decreases VEGFR-1 expression in human monocyte-derived macrophages.

Monocyte infiltration followed by differentiation into macrophages and accumulation of oxidised LDL (oxLDL) comprise early stages of atherosclerosis. Vascular endothelial growth factor (VEGF), which is upregulated by oxLDL, may contribute to atherogenesis through monocyte recruitment, increased vascular permeability and promotion of intraplaque vessels. The VEGF receptor-1 (VEGFR-1/Flt-1) mediates monocyte migration towards VEGF and regulates the levels of available VEGF through ligand-entrapment. In this study we investigated the effect of oxLDL on VEGFR-1 expression in human monocyte-derived macrophages. mRNA expression was estimated using RT-PCR, protein secretion was measured with ELISA and the amount of membrane-bound VEGFR-1 was analysed using flow cytometry analysis. Binding of transcription factors to the promoter was studied with EMSA. Incubation with oxLDL decreased VEGFR-1 mRNA expression in a time- and dose-dependent manner, followed by a subsequent decrease in protein secretion of VEGFR-1 and a reduction of the amount of receptor expressed on the cell surface. Furthermore, the PPARgamma agonists 9-hydroxy-(S)-10,12-octadecadienoic acid (9-HODE) and darglitazone also decreased VEGFR-1 mRNA expression. Incubation of macrophages with oxLDL or 9-HODE decreased binding of the transcription factor AP-1 (c-jun/ATF-2) to the VEGFR-1 promoter. Together, these data suggest that oxLDL decreases VEGFR-1 expression in macrophages, probably through a PPARgamma-dependent reduction in the AP-1 transcriptional activity. This implies that oxLDL has effects on the biological availability of VEGF, besides its direct effect on VEGF expression.

gammadelta T cells contribute to the systemic immunoglobulin E response and local B-cell reactivity in allergic eosinophilic airway inflammation.

Allergic airway inflammation induced in mice is T-cell dependent and recruitment of eosinophils to airspaces requires both alphabeta and gammadelta T cells. From previous studies it is evident that alphabeta T cells are essential for the allergic T helper type 2 (Th2)-like response, while the mechanistic contribution of gammadelta T cells is still unclear. In this study, we have investigated the role of gammadelta T cells in allergic airway eosinophilia induced by ovalbumin hypersensitivity. By comparing the responsiveness to sensitizing allergen of wild-type mice with that of T-cell receptor gammadelta knockout mice (TCRgammadelta KO) we demonstrated that mice lacking gammadelta T cells are defective in the systemic ovalbumin-specific immunoglobulin E (IgE) response. Furthermore, after aerosol challenge with allergen, gammadelta T-cell deficient mice exhibited a significantly decreased migration of B cells and natural killer cells to airways and reduced levels of allergen-specific IgG and IgA in bronchoalveolar lavage fluid. The role for B cells in the airway inflammation was indicated by the impaired ability of mice lacking functional B cells to evoke an eosinophilic response. The diminished eosinophilia in TCRgammadelta KO mice could not be explained by a defective Th2 activation since these mice displayed a normal IgG response in serum and an unaffected IG2b/IgG1 ratio in airways. Analysis of immunoregulatory cytokines in isolated lung tissue, thoracic lymph nodes and spleen further supported the notion that these mice are able to evoke a sufficient activation of T helper cells and that gammadelta T cells are not required for maintaining the Th2 profile. These results indicate that gammadelta T cells contribute to allergic airway inflammation by pathways separate from classical Th2 immune activation.

Accumulation and elimination of 16 polycyclic aromatic compounds in the earthworm (Eisenia fetida).

Accumulation and elimination of different polycyclic aromatic compounds (PACs) were studied in earthworms (Eisenia fetida) exposed to contaminated soil from an old gasworks site. In total, 12 polycyclic aromatic hydrocarbons (PAHs), two N- and S-heterocyclic PACs, and two PAC-quinones were included in the study. Peak-shaped accumulation curves were found for many of the compounds. After 19 d of exposure, the ratio between concentrations in worm lipids and soil organic matter was 0.02 on average. The half-lives of the PACs were relatively long, between 2 and 11 d. The elimination rate constants, k2, correlated both with literature-derived octanol-water partition coefficients (Kow) for PAHs (r2 = 0.93) and the computed polarizability (r2 = 0.88) of all the compounds. The elimination rate constants of PAHs are comparable to those of PCBs found in earlier studies, and the linear regression coefficient, r2, of k2 against Kow for PAHs and PCBs together was 0.93.