bluuebunny/crossref_metadata_2025 huggingface dataset:
Infections of People with Complement Deficiencies and Patients Who Have Undergone Splenectomy
Authors: Ram Sanjay , Lewis Lisa A. , Rice Peter A.
Published: October 2010

The complement system comprises several fluid-phase and membrane-associated proteins. Under physiological conditions, activation of the fluid-phase components of complement is maintained under tight control and complement activation occurs primarily on surfaces recognized as “nonself” in an attempt to minimize damage to bystander host cells. Membrane complement components act to limit complement activation on host cells or to facilitate uptake of antigens or microbes “tagged” with complement fragments. While this review focuses on the role of complement in infectious diseases, work over the past couple of decades has defined several important functions of complement distinct from that of combating infections. Activation of complement in the fluid phase can occur through the classical, lectin, or alternative pathway. Deficiencies of components of the classical pathway lead to the development of autoimmune disorders and predispose individuals to recurrent respiratory infections and infections caused by encapsulated organisms, includingStreptococcus pneumoniae,Neisseria meningitidis, andHaemophilus influenzae. While no individual with complete mannan-binding lectin (MBL) deficiency has been identified, low MBL levels have been linked to predisposition to, or severity of, several diseases. It appears that MBL may play an important role in children, who have a relatively immature adaptive immune response. C3 is the point at which all complement pathways converge, and complete deficiency of C3 invariably leads to severe infections, including those caused by meningococci and pneumococci. Deficiencies of the alternative and terminal complement pathways result in an almost exclusive predisposition to invasive meningococcal disease. The spleen plays an important role in antigen processing and the production of antibodies. Splenic macrophages are critical in clearing opsonized encapsulated bacteria (such as pneumococci, meningococci, andEscherichia coli) and intraerythrocytic parasites such as those causing malaria and babesiosis, which explains the fulminant nature of these infections in persons with anatomic or functional asplenia. Paramount to the management of patients with complement deficiencies and asplenia is educating patients about their predisposition to infection and the importance of preventive immunizations and seeking prompt medical attention.
Serum Intercellular Adhesion Molecule 1 Variations in Young Children with Acute Otitis Media
Authors: Liu Keyi , Casey Janet , Pichichero Michael
Published: October 2010

Acute otitis media (AOM) is an inflammatory reaction in the middle ear, most often occurring in young children.Streptococcus pneumoniae, nontypeableHaemophilusinfluenzae, andMoraxella catarrhalisare the most common bacteria isolated. Intercellular adhesion molecule 1 (ICAM-1) is involved in the innate immune response to infection by microorganisms, in effective antigen presentation, and in subsequent T-cell activation. Here we prospectively studied levels of serum soluble ICAM-1 (sICAM-1) before, at the time of, and after antimicrobial treatment of AOM in a group of 138 children ages 6 to 30 months. Middle ear fluids were collected by tympanocentesis to identify otopathogens. We found that (i) serum levels of sICAM-1 were significantly higher inS. pneumoniae-, nontypeableH. influenzae-, andM. catarrhalis-infected children than in well children (P< 0.001), confirming that a systemic inflammatory response occurs during AOM; (ii) sICAM-1 levels varied from no elevation (110 ng/ml) to elevation to high levels (maximum, 1,470 ng/ml) among children with AOM; (iii) in paired samples, sICAM-1 levels increased 4- to 20-fold when children developed AOM compared to their sICAM-1 levels before infection; and (iv) the level of sICAM-1 returned to the pre-AOM level at the convalescent stage of AOM after successful antimicrobial therapy. We conclude that AOM often causes a systemic inflammatory reaction, as measured by elevation of the serum sICAM-1 level, and that a high variability in sICAM-1 responses occurs with the presence of otopathogens during AOM.
Development and Characterization of ProtectiveHaemophilus parasuisSubunit Vaccines Based on Native Proteins with Affinity to Porcine Transferrin and Comparison with Other Subunit and Commercial Vaccines
Authors: Frandoloso Rafael , Martínez Sonia , Rodríguez-Ferri Elías F. , García-Iglesias María José , Pérez-Martínez Claudia , Martínez-Fernández Beatriz , Gutiérrez-Martín César B.
Published: October 2010

Haemophilus parasuisis the agent responsible for causing Glässer's disease, which is characterized by fibrinous polyserositis, polyarthritis, and meningitis in pigs. In this study, we have characterized native outer membrane proteins with affinity to porcine transferrin (NPAPT) fromH. parasuisserovar 5, Nagasaki strain. This pool of proteins was used as antigen to developed two vaccine formulations: one was adjuvanted with a mineral oil (Montanide IMS 2215 VG PR), while the other was potentiated with a bacterial neuraminidase fromClostridium perfringens. The potential protective effect conferred by these two vaccines was compared to that afforded by two other vaccines, consisting of recombinant transferrin-binding protein (rTbp) A or B fragments fromH. parasuis, Nagasaki strain, and by a commercially available inactivated vaccine. Five groups of colostrum-deprived piglets immunized with the vaccines described above, one group per each vaccine, and a group of nonvaccinated control animals were challenged intratracheally with a lethal dose (3 × 108CFU) ofH. parasuis, Nagasaki strain. The two vaccines containing rTbps yielded similar results with minimal protection against death, clinical signs, gross and microscopic lesions, andH. parasuisinvasion. In contrast, the two vaccines composed of NPAPT antigen and commercial bacterin resulted in a strong protection against challenge (without deaths and clinical signs), mild histopathological changes, and no recovery ofH. parasuis, thus suggesting their effectiveness in preventing Glässer's disease outbreaks caused by serovar 5.
Detection of Humoral Response Using a Recombinant Heat Shock Protein 70, DnaK, ofMycoplasma haemofelisin Experimentally and Naturally Hemoplasma-Infected Cats
Authors: Barker Emily N. , Helps Chris R. , Heesom Kate J. , Arthur Christopher J. , Peters Iain R. , Hofmann-Lehmann Regina , Tasker Séverine
Published: October 2010

Hemoplasmas is the trivial name given to a group of erythrocyte-parasitizing bacteria of the genusMycoplasma. Of the feline hemoplasmas,Mycoplasma haemofelisis the most pathogenic, while “CandidatusMycoplasma haemominutum” and “CandidatusMycoplasma turicensis” are less pathogenic. Shotgun libraries of fragmentedM. haemofelisgenomic DNA were constructed, and random colonies were selected for DNA sequencing.In silico-translated amino acid sequences of putative open reading frames were compared to mass spectrometry data fromM. haemofelisprotein spots identified as being immunogenic by two-dimensional gel electrophoresis and Western blotting. Three of the spots matched the predicted sequences of a heat shock protein 70 (DnaK) homolog, elongation factor Ts, and a fragment of phosphoglycerate kinase found during library screening. A full-length copy of theM. haemofelis dnaKgene was cloned intoEscherichia coliand recombinantly expressed. RecombinantM. haemofelisDnaK was purified and then used in Western blotting and an enzyme-linked immunosorbent assay (ELISA) to investigate the humoral immune response during acute infection in cats experimentally infected withM. haemofelis, “Ca.Mycoplasma haemominutum,” or “Ca.Mycoplasma turicensis”. The recombinantM. haemofelisDnaK ELISA also was used to screen clinical samples submitted for hemoplasma PCR testing to a commercial laboratory (n= 254). Experimentally infected cats became seropositive following infection, with a greater and earlier antibody response seen in cats inoculated withM. haemofelisthan those seen in cats inoculated with “Ca.Mycoplasma haemominutum” or “Ca.Mycoplasma turicensis,” by both Western blotting and ELISA. Of the clinical samples, 31.1% had antibodies detected by the ELISA but only 9.8% were positive by PCR for one or more hemoplasmas.
Sensitive Assay for Quantification of Hepatitis B Virus Mutants by Use of a Minor Groove Binder Probe and Peptide Nucleic Acids
Authors: Hige Shuhei , Yamamoto Yoichi , Yoshida Shigeru , Kobayashi Tomoe , Horimoto Hiromasa , Yamamoto Keiko , Sho Takuya , Natsuizaka Mitsuteru , Nakanishi Mitsuru , Chuma Makoto , Asaka Masahiro
Published: October 2010

Lamivudine is the first nucleoside analogue that was shown to have a potent effect on hepatitis B virus (HBV). However, the emergence of mutants resistant or cross-resistant to nucleoside/nucleotide analogues remains a serious problem. Several assays for the detection and quantification of antiviral-resistant mutants have been reported, but it has been difficult to measure the amounts of mutants accurately, especially when the target strain is a minor component of the mixed population. It has been shown that accurate measurement of a minor strain is difficult as long as a matching reaction with a single probe is included in the assay. We developed a new method for the quantification of lamivudine-resistant strains in a mixed-virus population by real-time PCR using minor groove binder probes and peptide nucleic acids, and we achieved a wide and measurable range, from 3 to 10 log
10
copies/ml, and high sensitivity, with a discriminative limit of 0.01% of the predominant strain. The clinical significance of measuring substitutions not only of M204 but also of L180 residues of HBV polymerase was demonstrated by this method. This assay increases the versatility of a sensitive method for the quantification of a single-nucleotide mutation in a heterogeneous population.
Evaluation of Time to Detection of
Mycobacterium tuberculosis
in Broth Culture as a Determinant for End Points in Treatment Trials
Authors: Weiner Marc , Prihoda Thomas J. , Burman William , Johnson John L. , Goldberg Stefan , Padayatchi Nesri , Duran Paula , Engle Melissa , Muzanye Grace , Mugerwa Roy D. , Sturm A. Willem
Published: October 2010

Time to detection of
Mycobacterium tuberculosis
in broth culture was examined for utility as a treatment efficacy end point. Of 146 patients in a phase IIB trial, a decreased mean time to detection was found in 5 with treatment failure. Time to detection in an analysis-of-covariance model was associated with lung cavities, less intensive treatment, and differences in the bactericidal effects of treatment regimens.
Feasibility Study of a Human Papillomavirus E6 Oncoprotein Test for Diagnosis of Cervical Precancer and Cancer
Authors: Schweizer Johannes , Lu Peter S. , Mahoney Charles W. , Berard-Bergery Marthe , Ho Minh , Ramasamy Valli , Silver Jon E. , Bisht Arnima , Labiad Yassine , Peck Roger B. , Lim Jeanette , Jeronimo Jose , Howard Roslyn , Gravitt Patti E. , Castle Philip E.
Published: October 2010

In a feasibility study using a prototype, lateral-flow test for human papillomavirus type 16, 18, and/or 45 (HPV16/18/45) E6 oncoproteins, 51 of 75 (68%; 95% confidence interval [95% CI] of 56 to 78%) of HPV16/18/45 DNA-positive specimens from women with a diagnosis of CIN3+ (cervical intraepithelial neoplasia grade 3+ or cervical cancer) tested positive for HPV16/18/45 E6 oncoprotein. None of 16 (95% CI of 0 to 37%) HPV16/18/45 DNA-positive cervical specimens from women with a negative or CIN1 diagnosis tested positive for HPV16/18/45 E6 oncoprotein.
Isolation of a Novel
Orientia
Species (
O. chuto
sp. nov.) from a Patient Infected in Dubai
Authors: Izzard Leonard , Fuller Andrew , Blacksell Stuart D. , Paris Daniel H. , Richards Allen L. , Aukkanit Nuntipa , Nguyen Chelsea , Jiang Ju , Fenwick Stan , Day Nicholas P. J. , Graves Stephen , Stenos John
Published: October 2010

In July 2006, an Australian tourist returning from Dubai, in the United Arab Emirates (UAE), developed acute scrub typhus. Her signs and symptoms included fever, myalgia, headache, rash, and eschar.
Orientia tsutsugamushi
serology demonstrated a 4-fold rise in antibody titers in paired serum collections (1:512 to 1:8,192), with the sera reacting strongest against the Gilliam strain antigen. An
Orientia
species was isolated by the
in vitro
culture of the patient's acute blood taken prior to antibiotic treatment. The gene sequencing of the 16S rRNA gene (
rrs
), partial 56-kDa gene, and the full open reading frame 47-kDa gene was performed, and comparisons of this new
Orientia
sp. isolate to previously characterized strains demonstrated significant sequence diversity. The closest homology to the
rrs
sequence of the new
Orientia
sp. isolate was with three strains of
O. tsutsugamushi
(Ikeda, Kato, and Karp), with a nucleotide sequence similarity of 98.5%. The closest homology to the 47-kDa gene sequence was with
O. tsutsugamushi
strain Gilliam, with a nucleotide similarity of 82.3%, while the closest homology to the 56-kDa gene sequence was with
O. tsutsugamushi
strain TA686, with a nucleotide similarity of 53.1%. The molecular divergence and geographically unique origin lead us to believe that this organism should be considered a novel species. Therefore, we have proposed the name “
Orientia chuto
,” and the prototype strain of this species is strain Dubai, named after the location in which the patient was infected.
Molecular Phylogeny and Proposal of Two New Species of the Emerging Pathogenic FungusSaksenaea
Authors: Alvarez E. , Garcia-Hermoso D. , Sutton D. A. , Cano J. F. , Stchigel A. M. , Hoinard D. , Fothergill A. W. , Rinaldi M. G. , Dromer F. , Guarro J.
Published: October 2010

Saksenaeais a monotypic genus belonging to the orderMucoralesand capable of producing severe human infections. Through a polyphasic study based on analysis of the sequences of the internal transcribed spacer (ITS) region, domains D1 and D2 of the 28S rRNA gene, and the elongation factor 1α (EF-1α) gene, as well as by evaluation of relevant morphological and physiological characteristics of a set of clinical and environmental strains, we have demonstrated thatSaksenaea vasiformisis a complex of species. We propose as new speciesSaksenaea oblongispora, characterized by oblong sporangiospores and unable to grow at 42°C, andSaksenaea erythrospora, characterized by large sporangiophores and sporangia and by ellipsoid sporangiospores, biconcave in the lateral view. Itraconazole, posaconazole, and terbinafine were active against all isolates included in the study, while amphotericin B, voriconazole, and the echinocandins showed low activity.
Genomic Profiling of Tumor Necrosis Factor Alpha (TNF-α) Receptor and Interleukin-1 Receptor Knockout Mice Reveals a Link between TNF-α Signaling and Increased Severity of 1918 Pandemic Influenza Virus Infection
Authors: Belisle Sarah E. , Tisoncik Jennifer R. , Korth Marcus J. , Carter Victoria S. , Proll Sean C. , Swayne David E. , Pantin-Jackwood Mary , Tumpey Terrence M. , Katze Michael G.
Published: October 2010

The influenza pandemic of 1918 to 1919 was one of the worst global pandemics in recent history. The highly pathogenic nature of the 1918 virus is thought to be mediated in part by a dysregulation of the host response, including an exacerbated proinflammatory cytokine response. In the present study, we compared the host transcriptional response to infection with the reconstructed 1918 virus in wild-type, tumor necrosis factor (TNF) receptor-1 knockout (TNFRKO), and interleukin-1 (IL-1) receptor-1 knockout (IL1RKO) mice as a means of further understanding the role of proinflammatory cytokine signaling during the acute response to infection. Despite reported redundancy in the functions of IL-1β and TNF-α, we observed that reducing the signaling capacity of each of these molecules by genetic disruption of their key receptor genes had very different effects on the host response to infection. In TNFRKO mice, we found delayed or decreased expression of genes associated with antiviral and innate immune signaling, complement, coagulation, and negative acute-phase response. In contrast, in IL1RKO mice numerous genes were differentially expressed at 1 day postinoculation, including an increase in the expression of genes that contribute to dendritic and natural killer cell processes and cellular movement, and gene expression profiles remained relatively constant at later time points. We also observed a compensatory increase in TNF-α expression in virus-infected IL1RKO mice. Our data suggest that signaling through the IL-1 receptor is protective, whereas signaling through the TNF-α receptor increases the severity of 1918 virus infection. These findings suggest that manipulation of these pathways may have therapeutic benefit.
Hepatitis C Virus Nonstructural Protein 5A: Biochemical Characterization of a Novel Structural Class of RNA-Binding Proteins
Authors: Hwang Jungwook , Huang Luyun , Cordek Daniel G. , Vaughan Robert , Reynolds Shelley L. , Kihara George , Raney Kevin D. , Kao C. Cheng , Cameron Craig E.
Published: October 2010

Hepatitis C virus (HCV) nonstructural protein 5A (NS5A) exhibits a preference for G/U-rich RNA
in vitro
. Biological analysis of the NS5A RNA-binding activity and its target sites in the genome will be facilitated by a description of the NS5A-RNA complex. We demonstrate that the C-4 carbonyl of the uracil base and, by inference, the C-6 carbonyl of the guanine base interact with NS5A. U-rich RNA of 5 to 6 nucleotides (nt) is sufficient for high-affinity binding to NS5A. The minimal RNA-binding domain of NS5A consists of residues 2005 to 2221 (referred to as domain I-plus). This region of the protein includes the amino-terminal domain I as well as the subsequent linker that separates domains I and II. This linker region is the site of adaptive mutations. U-rich RNA-binding activity is not observed for an NS5A derivative containing only residues 2194 to 2419 (domains II and III). Mass spectrometric analysis of an NS5A-poly(rU) complex identified domains I and II as sites for interaction with RNA. Dimerization of NS5A was demonstrated by glutaraldehyde cross-linking. This dimerization is likely mediated by domain I-plus, as dimers of this protein are trapped by cross-linking. Dimers of the domain II-III protein are not observed. The monomer-dimer equilibrium of NS5A shifts in favor of dimer when U-rich RNA is present but not when A-rich RNA is present, consistent with an NS5A dimer being the RNA-binding-competent form of the protein. These data provide a molecular perspective of the NS5A-RNA complex and suggest possible mechanisms for regulation of HCV and cellular gene expression.
Antibody to the E3 Glycoprotein Protects Mice against Lethal Venezuelan Equine Encephalitis Virus Infection
Authors: Parker Michael D. , Buckley Marilyn J. , Melanson Vanessa R. , Glass Pamela J. , Norwood David , Hart Mary Kate
Published: October 2010

Six monoclonal antibodies were isolated that exhibited specificity for a furin cleavage site deletion mutant (V3526) of Venezuelan equine encephalitis virus (VEEV). These antibodies comprise a single competition group and bound the E3 glycoprotein of VEEV subtype I viruses but failed to bind the E3 glycoprotein of other alphaviruses. These antibodies neutralized V3526 virus infectivity but did not neutralize the parental strain of Trinidad donkey (TrD) VEEV. However, the E3-specific antibodies did inhibit the production of virus from VEEV TrD-infected cells. In addition, passive immunization of mice demonstrated that antibody to the E3 glycoprotein provided protection against lethal VEEV TrD challenge. This is the first recognition of a protective epitope in the E3 glycoprotein. Furthermore, these results indicate that E3 plays a critical role late in the morphogenesis of progeny virus after E3 appears on the surfaces of infected cells.
cis
-Acting Sequences That Contribute to Synthesis of Minus-Strand DNA Are Not Conserved between Hepadnaviruses
Authors: Maguire Megan L. , Loeb Daniel D.
Published: October 2010

Hepadnaviruses are DNA viruses that are found in several mammalian and avian species. These viruses replicate their genome through reverse transcription of an RNA intermediate termed pregenomic RNA (pgRNA). pgRNA is reverse transcribed by the viral polymerase into a minus-strand DNA, followed by synthesis of the plus-strand DNA. There are multiple
cis
-acting sequences that contribute to the synthesis of minus-strand DNA for human hepatitis B virus (HBV). Less is known about the
cis
-acting sequences of avian hepadnaviruses that contribute to synthesis of minus-strand DNA. To identify
cis
-acting sequences of duck hepatitis B virus (DHBV) and heron hepatitis B virus (HHBV), we analyzed variants containing 200-nucleotide (nt) deletions. Most variants of DHBV synthesized minus-strand DNA to 50 to 100% of the wild-type (WT) level, while two variants synthesized less than 50%. For HHBV, most variants synthesized minus-strand DNA to less than 50% the WT level. These results differ from those for HBV, where most of the genome can be removed with little consequence. HBV contains a sequence, φ, that contributes to the synthesis of minus-strand DNA. It has been proposed that DHBV has an analogous sequence. We determined that the proposed φ sequence of DHBV does not contribute to the synthesis of minus-strand DNA. Finally, we found that the DR2 sequence present in all hepadnaviruses is important for synthesis of minus-strand DNA in both DHBV and HHBV but not in HBV. These differences in
cis
-acting sequences suggest that the individual hepadnaviruses have evolved differences in their mechanisms for synthesizing minus-strand DNA, more so than for other steps in replication.
Efficient Activation of the Severe Acute Respiratory Syndrome Coronavirus Spike Protein by the Transmembrane Protease TMPRSS2
Authors: Matsuyama Shutoku , Nagata Noriyo , Shirato Kazuya , Kawase Miyuki , Takeda Makoto , Taguchi Fumihiro
Published: October 2010

The distribution of the severe acute respiratory syndrome coronavirus (SARS-CoV) receptor, an angiotensin-converting enzyme 2 (ACE2), does not strictly correlate with SARS-CoV cell tropism in lungs; therefore, other cellular factors have been predicted to be required for activation of virus infection. In the present study, we identified transmembrane protease serine 2 (TMPRSS2), whose expression does correlate with SARS-CoV infection in the upper lobe of the lung. In Vero cells expressing TMPRSS2, large syncytia were induced by SARS-CoV infection. Further, the lysosome-tropic reagents failed to inhibit, whereas the heptad repeat peptide efficiently inhibited viral entry into cells, suggesting that TMPRSS2 affects the S protein at the cell surface and induces virus-plasma membrane fusion. On the other hand, production of virus in TMPRSS2-expressing cells did not result in S-protein cleavage or increased infectivity of the resulting virus. Thus, TMPRSS2 affects the entry of virus but not other phases of virus replication. We hypothesized that the spatial orientation of TMPRSS2 vis-a-vis S protein is a key mechanism underling this phenomenon. To test this, the TMPRSS2 and S proteins were expressed in cells labeled with fluorescent probes of different colors, and the cell-cell fusion between these cells was tested. Results indicate that TMPRSS2 needs to be expressed in the opposing (target) cell membrane to activate S protein rather than in the producer cell, as found for influenza A virus and metapneumoviruses. This is the first report of TMPRSS2 being required in the target cell for activation of a viral fusion protein but not for the S protein synthesized in and transported to the surface of cells. Our findings suggest that the TMPRSS2 expressed in lung tissues may be a determinant of viral tropism and pathogenicity at the initial site of SARS-CoV infection.
Association of Potent Human Antiviral Cytidine Deaminases with 7SL RNA and Viral RNP in HIV-1 Virions
Authors: Zhang Wenyan , Du Juan , Yu Kevin , Wang Tao , Yong Xiong , Yu Xiao-Fang
Published: October 2010

7SL RNA promotes the formation of the signal recognition particle that targets secretory and membrane proteins to the endoplasmic reticulum. 7SL RNA is also selectively packaged by many retroviruses, including HIV-1. Here, we demonstrate that 7SL RNA is an integral component of the viral ribonucleoprotein (RNP) complex containing Gag, viral genomic RNA, and

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. Only the potent anti-HIV-1 cytidine deaminases can bind to 7SL RNA and target to HIV-1 RNP. A conserved motif in the amino-terminal region of A3G is important for 7SL RNA interaction. The weak anti-HIV-1 A3C did not interact with 7SL RNA and failed to target to viral RNPs, despite efficient virion packaging. However, a chimeric construct of A3C plus the 7SL-binding amino terminus of A3G did target to viral RNPs and showed enhanced anti-HIV-1 activity. 7SL RNA binding is a conserved feature of human anti-HIV-1 cytidine deaminases. Thus, potent anti-HIV-1 cytidine deaminases have evolved to possess a unique RNA-binding ability for precise HIV-1 targeting and viral inhibition.
Beyond the neglect of psychological consequences: post-traumatic stress disorder increases the non-fatal burden of injury by more than 50%
Authors: Haagsma Juanita A , Polinder Suzanne , Toet Hidde , Panneman Martien , Havelaar Arie H , Bonsel Gouke J , van Beeck Ed F
Published: October 2010

Background

Psychological consequences such as post-traumatic stress disorder (PTSD) are currently neglected in burden-of-injury calculations.

Aim

To assess the disease burden of PTSD due to unintentional injury and compare this health loss with physical injury consequences.

Methods

From literature sources, the prevalence of PTSD at four follow-up periods (<3 months, 3–6 months, 7–12 months and >12 months) was estimated. The uncertainty of the estimated PTSD prevalence was modelled by a Bayesian approach. The prevalence rates were then linked to national data on unintentional injury, disability weights and duration to estimate the incidence and disability-adjusted life years (DALY) resulting from PTSD in addition to physical injury consequences.

Results

The data suggest that PTSD prevalence among injury victims decreases over time. The average PTSD prevalence at <3 months follow-up was 21% (90% credibility interval (CI) 17% to 24%) for patients presenting at the emergency department and 30% (90% CI 27% to 33%) for patients who were hospitalised, tapering down after 12 months to 4% (90% CI 3% to 5%) and 6% (90% CI 4% to 10%), respectively. These estimates translate into 191 000 (90% CI 161 000 to 222 000) cases of PTSD per year in the Dutch population (1.2%) due to unintentional injury. Including PTSD increases the non-fatal burden of disease of unintentional injuries by 53% (from 116 000 to 178 000 DALYs (90% CI 150 000 to 217 000)).

Conclusions

Ignoring PTSD in burden-of-injury studies results in a considerable underestimation of the burden of injury. This may affect resource allocation and the identification of important prevention priorities.
Chromatin Regulatory Mechanisms in Pluripotency
Authors: Lessard Julie A. , Crabtree Gerald R.
Published: October 2010

Stem cells of all types are characterized by a stable, heritable state permissive of multiple developmental pathways. The past five years have seen remarkable advances in understanding these heritable states and the ways that they are initiated or terminated. Transcription factors that bind directly to DNA and have sufficiency roles have been most easy to investigate and, perhaps for this reason, are most solidly implicated in pluripotency. In addition, large complexes of ATP-dependent chromatin-remodeling and histone-modification enzymes that have specialized functions have also been implicated by genetic studies in initiating and/or maintaining pluripotency or multipotency. Several of these ATP-dependent remodeling complexes play non-redundant roles, and the esBAF complex facilitates reprogramming of induced pluripotent stem cells. The recent finding that virtually all histone modifications can be rapidly reversed and are often highly dynamic has raised new questions about how histone modifications come to play a role in the steady state of pluripotency. Another surprise from genetic studies has been the frequency with which the global effects of mutations in chromatin regulators can be largely reversed by a single target gene. These genetic studies help define the arena for future mechanistic studies that might be helpful to harness pluripotency for therapeutic goals.
Plant Nuclear Hormone Receptors: A Role for Small Molecules in Protein-Protein Interactions
Authors: Lumba Shelley , Cutler Sean , McCourt Peter
Published: October 2010

Plant hormones are a group of chemically diverse small molecules that direct processes ranging from growth and development to biotic and abiotic stress responses. Surprisingly, genome analyses suggest that classic animal nuclear hormone receptor homologs do not exist in plants. It now appears that plants have co-opted several protein families to perceive hormones within the nucleus. In one solution to the problem, the hormones auxin and jasmonate (JA) act as “molecular glue” that promotes protein-protein interactions between receptor F-boxes and downstream corepressor targets. In another solution, gibberellins (GAs) bind and elicit a conformational change in a novel soluble receptor family related to hormone-sensitive lipases. Abscisic acid (ABA), like GA, also acts through an allosteric mechanism involving a START-domain protein. The molecular identification of plant nuclear hormone receptors will allow comparisons with animal nuclear receptors and testing of fundamental questions about hormone function in plant development and evolution.
Control of Mitotic Spindle Length
Authors: Goshima Gohta , Scholey Jonathan M.
Published: October 2010

The mitotic spindle accurately segregates genetic instructions by moving chromosomes to spindle poles (anaphase A) and separating the poles (anaphase B) so that, in general, the chromosomes and poles are positioned near the centers of the nascent daughter cell products of each cell division. Because the size of different types of dividing cells, and thus the spacing of their daughter cell centers, can vary significantly, the length of the metaphase or postanaphase B spindle often scales with cell size. However, significant exceptions to this scaling rule occur, revealing the existence of cell size–independent, spindle-associated mechanisms of spindle length control. The control of spindle length reflects the action of mitotic force-generating mechanisms, and its study may illuminate general principles by which cells regulate the size of internal structures. Here we review molecules and mechanisms that control spindle length, how these mechanisms are deployed in different systems, and some quantitative models that describe the control of spindle length.
Assembly of Fibronectin Extracellular Matrix
Authors: Singh Purva , Carraher Cara , Schwarzbauer Jean E.
Published: October 2010

In the process of matrix assembly, multivalent extracellular matrix (ECM) proteins are induced to self-associate and to interact with other ECM proteins to form fibrillar networks. Matrix assembly is usually initiated by ECM glycoproteins binding to cell surface receptors, such as fibronectin (FN) dimers binding to α5β1 integrin. Receptor binding stimulates FN self-association mediated by the N-terminal assembly domain and organizes the actin cytoskeleton to promote cell contractility. FN conformational changes expose additional binding sites that participate in fibril formation and in conversion of fibrils into a stabilized, insoluble form. Once assembled, the FN matrix impacts tissue organization by contributing to the assembly of other ECM proteins. Here, we describe the major steps, molecular interactions, and cellular mechanisms involved in assembling FN dimers into fibrillar matrix while highlighting important issues and major questions that require further investigation.
Modulation of Host Cell Function byLegionella pneumophilaType IV Effectors
Authors: Hubber Andree , Roy Craig R.
Published: October 2010

Macrophages and protozoa ingest bacteria by phagocytosis and destroy these microbes using a conserved pathway that mediates fusion of the phagosome with lysosomes. To survive within phagocytic host cells, bacterial pathogens have evolved a variety of strategies to avoid fusion with lysosomes. A virulence strategy used by the intracellular pathogen Legionella pneumophila is to manipulate host cellular processes using bacterial proteins that are delivered into the cytosolic compartment of the host cell by a specialized secretion system called Dot/Icm. The proteins delivered by the Dot/Icm system target host factors that play evolutionarily conserved roles in controlling membrane transport in eukaryotic cells, which enables L. pneumophila to create an endoplasmic reticulum-like vacuole that supports intracellular replication in both protozoan and mammalian host cells. This review focuses on intracellular trafficking of L. pneumophila and describes how bacterial proteins contribute to modulation of host processes required for survival within host cells.
Regulatory Mechanisms for Specification and Patterning of Plant Vascular Tissues
Authors: Caño-Delgado Ana , Lee Ji-Young , Demura Taku
Published: October 2010

Plant vascular tissues, the conduits of water, nutrients, and small molecules, play important roles in plant growth and development. Vascular tissues have allowed plants to successfully adapt to various environmental conditions since they evolved 450 Mya. The majority of plant biomass, an important source of renewable energy, comes from the xylem of the vascular tissues. Efforts have been made to identify the underlying mechanisms of cell specification and patterning of plant vascular tissues and their proliferation. The formation of the plant vascular system is a complex process that integrates signaling and gene regulation at transcriptional and posttranscriptional levels. Recently, a wealth of molecular genetic studies and the advent of cell biology and genomic tools have enabled important progress toward understanding its underlying mechanisms. Here, we provide a comprehensive review of the cell and developmental processes of plant vascular tissue and resources recently available for studying them that will enable the discovery of new ways to develop sustainable energy using plant biomass.
A Decade of Systems Biology
Authors: Chuang Han-Yu , Hofree Matan , Ideker Trey
Published: October 2010

Systems biology provides a framework for assembling models of biological systems from systematic measurements. Since the field was first introduced a decade ago, considerable progress has been made in technologies for global cell measurement and in computational analyses of these data to map and model cell function. It has also greatly expanded into the translational sciences, with approaches pioneered in yeast now being applied to elucidate human development and disease. Here, we review the state of the field with a focus on four emerging applications of systems biology that are likely to be of particular importance during the decade to follow: (a) pathway-based biomarkers, (b) global genetic interaction maps, (c) systems approaches to identify disease genes, and (d) stem cell systems biology. We also cover recent advances in software tools that allow biologists to explore system-wide models and to formulate new hypotheses. The applications and methods covered in this review provide a set of prime exemplars useful to cell and developmental biologists wishing to apply systems approaches to areas of interest.
Membrane Fusion: Five Lipids, Four SNAREs, Three Chaperones, Two Nucleotides, and a Rab, All Dancing in a Ring on Yeast Vacuoles
Authors: Wickner William
Published: October 2010

Although fusion mechanisms are highly conserved in evolution and among organelles of the exocytic and endocytic pathways, yeast vacuole homotypic fusion offers unique technical advantages: excellent genetics, clear organelle cytology, in vitro colorimetric fusion assays, and reconstitution of fusion from all-pure components, including a Rab GTPase, HOPS (homotypic fusion and vacuole protein sorting complex), four SNAREs [soluble N-ethylmaleimide-sensitive factor (NSF) attachment receptors] that snare (bind) each other, SNARE-complex disassembly chaperones, and vacuolar lipids. Vacuole fusion studies offer paradigms of the interdependence of lipids and fusion proteins to assemble a fusion microdomain, distinct lipid functions, SNARE complex proofreading through the synergy between HOPS and the SNARE disassembly chaperones, and the role of each fusion protein in promoting radical bilayer restructuring for fusion without lysis.
Mechanical Integration of Actin and Adhesion Dynamics in Cell Migration
Authors: Gardel Margaret L. , Schneider Ian C. , Aratyn-Schaus, Yvonne , Waterman Clare M.
Published: October 2010

Directed cell migration is a physical process that requires dramatic changes in cell shape and adhesion to the extracellular matrix. For efficient movement, these processes must be spatiotemporally coordinated. To a large degree, the morphological changes and physical forces that occur during migration are generated by a dynamic filamentous actin (F-actin) cytoskeleton. Adhesion is regulated by dynamic assemblies of structural and signaling proteins that couple the F-actin cytoskeleton to the extracellular matrix. Here, we review current knowledge of the dynamic organization of the F-actin cytoskeleton in cell migration and the regulation of focal adhesion assembly and disassembly with an emphasis on how mechanical and biochemical signaling between these two systems regulate the coordination of physical processes in cell migration.
Rolling Cell Adhesion
Authors: McEver Rodger P. , Zhu Cheng
Published: October 2010

Rolling adhesion on vascular surfaces is the first step in recruiting circulating leukocytes, hematopoietic progenitors, or platelets to specific organs or to sites of infection or injury. Rolling requires the rapid yet balanced formation and dissociation of adhesive bonds in the challenging environment of blood flow. This review explores how structurally distinct adhesion receptors interact through mechanically regulated kinetics with their ligands to meet these challenges. Remarkably, increasing force applied to adhesive bonds first prolongs their lifetimes (catch bonds) and then shortens their lifetimes (slip bonds). Catch bonds mediate the counterintuitive phenomenon of flow-enhanced rolling adhesion. Force-regulated disruptions of receptor interdomain or intradomain interactions remote from the ligand-binding surface generate catch bonds. Adhesion receptor dimerization, clustering in membrane domains, and interactions with the cytoskeleton modulate the forces applied to bonds. Both inside-out and outside-in cell signals regulate these processes.
Assembling Neural Crest Regulatory Circuits into a Gene Regulatory Network
Authors: Betancur Paola , Bronner-Fraser Marianne , Sauka-Spengler Tatjana
Published: October 2010

The neural crest is a multipotent stem cell–like population that gives rise to a wide range of derivatives in the vertebrate embryo including elements of the craniofacial skeleton and peripheral nervous system as well as melanocytes. The neural crest forms in a series of regulatory steps that include induction and specification of the prospective neural crest territory–neural plate border, specification of bona fide neural crest progenitors, and differentiation into diverse derivatives. These individual processes during neural crest ontogeny are controlled by regulatory circuits that can be assembled into a hierarchical gene regulatory network (GRN). Here we present an overview of the GRN that orchestrates the formation of cranial neural crest cells. Formulation of this network relies on information largely inferred from gene perturbation studies performed in several vertebrate model organisms. Our representation of the cranial neural crest GRN also includes information about direct regulatory interactions obtained from the cis-regulatory analyses performed to date, which increases the resolution of the architectural circuitry within the network.
Tethering Factors as Organizers of Intracellular Vesicular Traffic
Authors: Yu I-Mei , Hughson Frederick M.
Published: October 2010

Intracellular trafficking entails the budding, transport, tethering, and fusion of transport vesicles and other membrane carriers. Here we review recent progress toward a mechanistic understanding of vesicle tethering. The known tethering factors are large complexes important for one or more intracellular trafficking pathways and are capable of interacting directly with many of the other principal components of the cellular trafficking machinery. Our review emphasizes recent developments in the in vitro reconstitution of vesicle tethering and the structural characterization of multisubunit tethering factors. The combination of these and other approaches has led to exciting progress toward understanding how these essential nanomachines work.
Trafficking to the Ciliary Membrane: How to Get Across the Periciliary Diffusion Barrier?
Authors: Nachury Maxence V. , Seeley E. Scott , Jin Hua
Published: October 2010

The primary cilium organizes numerous signal transduction cascades, and an understanding of signaling receptor trafficking to cilia is now emerging. A defining feature of cilia is the periciliary diffusion barrier that separates the ciliary and plasma membranes. Although lateral transport through this barrier may take place, polarized exocytosis to the base of the cilium has been the prevailing model for delivering membrane proteins to cilia. Key players for this polarized exocytosis model include the GTPases Rab8 and Rab11, the exocyst, and possibly the intraflagellar tranport machinery. In turn, the sorting of membrane proteins to cilia critically relies on the recognition of ciliary targeting signals by sorting machines such as the BBSome coat complex or the GTPase Arf4. Finally, some proteins need to exit from cilia, and ubiquitination may regulate this step. The stage is now set to dissect the interplay between signaling and regulated trafficking to and from cilia.
Identification and functional implication of a Rho kinase-dependent moesin-EBP50 interaction in noradrenaline-stimulated artery
Authors: Baeyens Nicolas , Horman Sandrine , Vertommen Didier , Rider Mark , Morel Nicole
Published: October 2010

Ezrin, radixin, and moesin (ERM) proteins are known to be substrates of Rho kinase (ROCK), a key player in vascular smooth muscle regulation. Their function in arteries remains to be elucidated. The objective of the present study was to investigate ERM phosphorylation and function in rat aorta and mesenteric artery and the influence of ERM-binding phosphoprotein 50 (EBP50), a scaffold partner of ERM proteins in several cell types. In isolated arteries, ERM proteins are phosphorylated by PKC and ROCK with different kinetics after either agonist stimulation or KCl-induced depolarization. Immunoprecipitation of EBP50 in noradrenaline-stimulated arteries allowed identification of its interaction with moesin and several other proteins involved in cytoskeleton regulation. This interaction was inhibited by Y27632, a ROCK inhibitor. Moesin or EBP50 depletion after small interfering RNA transfection by reverse permeabilization in intact mesenteric arteries both potentiated the contractility in response to agonist stimulation without any effect on contractile response induced by high KCl. This effect was preserved in ionomycin-permeabilized arteries. These results indicate that, in agonist-stimulated arteries, the activation of ROCK leads to the binding of moesin to EBP50, which interacts with several components of the cytoskeleton, resulting in a decrease in the contractile response.
Regulation of CFTR chloride channel macroscopic conductance by extracellular bicarbonate
Authors: Li Man-Song , Holstead Ryan G. , Wang Wuyang , Linsdell Paul
Published: October 2010

The CFTR contributes to Cl−and HCO3−transport across epithelial cell apical membranes. The extracellular face of CFTR is exposed to varying concentrations of Cl−and HCO3−in epithelial tissues, and there is evidence that CFTR is sensitive to changes in extracellular anion concentrations. Here we present functional evidence that extracellular Cl−and HCO3−regulate anion conduction in open CFTR channels. Using cell-attached and inside-out patch-clamp recordings from constitutively active mutant E1371Q-CFTR channels, we show that voltage-dependent inhibition of CFTR currents in intact cells is significantly stronger when the extracellular solution contains HCO3−than when it contains Cl−. This difference appears to reflect differences in the ability of extracellular HCO3−and Cl−to interact with and repel intracellular blocking anions from the pore. Strong block by endogenous cytosolic anions leading to reduced CFTR channel currents in intact cells occurs at physiologically relevant HCO3−concentrations and membrane potentials and can result in up to ∼50% inhibition of current amplitude. We propose that channel block by cytosolic anions is a previously unrecognized, physiologically relevant mechanism of channel regulation that confers on CFTR channels sensitivity to different anions in the extracellular fluid. We further suggest that this anion sensitivity represents a feedback mechanism by which CFTR-dependent anion secretion could be regulated by the composition of the secretions themselves. Implications for the mechanism and regulation of CFTR-dependent secretion in epithelial tissues are discussed.
Telomerase immortalization of principal cells from mouse collecting duct
Authors: Steele Stacy L. , Wu Yongren , Kolb Robert J. , Gooz Monika , Haycraft Courtney J. , Keyser Kent T. , Guay-Woodford Lisa , Yao Hai , Bell P. Darwin
Published: October 2010

Recently, the use of overexpression of telomerase reverse transcriptase (TERT) has led to the generation of immortalized human cell lines. However, this cell immortalization approach has not been reported in well-differentiated mouse cells, such as renal epithelial cells. We sought to establish and then characterize a mouse collecting duct cell line, using ectopic expression of mTERT. Isolated primary cortical collecting duct (CCD) cell lines were transduced with mouse (m)TERT, using a lentiviral vector. mTERT-negative cells did not survive blasticidin selection, whereas mTERT-immortalized cells proliferated in selection media for over 40 subpassages. mTERT messenger RNA and telomerase activity was elevated in these cells, compared with an SV40-immortalized cell line. Flow cytometry with Dolichos biflorus agglutinin was used to select the CCD principal cells, and we designated this cell line mTERT-CCD. Cells were well differentiated and exhibited morphological characteristics typically found in renal epithelial cells, such as tight junction formation, microvilli, and primary cilia. Further characterization using standard immunofluorescence revealed abundant expression of aquaporin-2 and the vasopressin type 2 receptor. mTERT-CCD cells exhibited cAMP-stimulated/benzamil-inhibited whole cell currents. Whole cell patch-clamp currents were also enhanced after a 6-day treatment with aldosterone. In conclusion, we have successfully used mTERT to immortalize mouse collecting duct cells that retain the basic in vivo phenotypic characteristics of collecting duct cells. This technique should be valuable in generating cell lines from genetically engineered mouse models.