Measurement and initial characterization of leukocyte telomere length in 474,074 participants in UK Biobank.

Leukocyte telomere length (LTL) is a proposed marker of biological age. Here we report the measurement and initial characterization of LTL in 474,074 participants in UK Biobank. We confirm that older age and male sex associate with shorter LTL, with women on average ~7 years younger in 'biological age' than men. Compared to white Europeans, LTL is markedly longer in African and Chinese ancestries. Older paternal age at birth is associated with longer individual LTL. Higher white cell count is associated with shorter LTL, but proportions of white cell subtypes show weaker associations. Age, ethnicity, sex and white cell count explain ~5.5% of LTL variance. Using paired samples from 1,351 participants taken ~5 years apart, we estimate the within-individual variability in LTL and provide a correction factor for this. This resource provides opportunities to investigate determinants and biomedical consequences of variation in LTL.

Pain in knee osteoarthritis is associated with variation in the neurokinin 1/substance P receptor (TACR1) gene.

BACKGROUND: Substance P (SP) is a pain- and inflammation-related neuropeptide which preferentially binds to the neurokinin receptor 1 (NK1 ). SP and NK1 receptors have been implicated in joint pain, inflammation and damage in animal models and human studies of osteoarthritis (OA). The aim of this study was to test if genetic variation at the neurokinin 1 receptor gene (TACR1) is associated with pain in individuals with radiographic knee OA. METHODS: Participants from the Genetics of OA and Lifestyle study were used for the discovery group (n = 1615). Genotype data for six SNPs selected to cover most variation in the TACR1 gene were used to test for an association with symptomatic OA. Replication analysis was performed using data from the Chingford 1000 Women Study, Hertfordshire Cohort Study, Tasmanian Older Adult Cohort Study and the Clearwater OA Study. In total, n = 1715 symptomatic OA and n = 735 asymptomatic OA individuals were analysed. RESULTS: Out of six SNPs tested in the TACR1 gene, one (rs11688000) showed a nominally significant association with a decreased risk of symptomatic OA in the discovery cohort. This was then replicated in four additional cohorts. After adjusting for age, gender, body mass index and radiographic severity, the G (minor) allele at rs11688000 was associated with a decreased risk of symptomatic OA compared to asymptomatic OA cases (p = 9.90 × 10-4 , OR = 0.79 95% 0.68-0.90 after meta-analysis). CONCLUSIONS: This study supports a contribution from the TACR1 gene in human OA pain, supporting further investigation of this gene's function in OA. SIGNIFICANCE: This study contributes to the knowledge of the genetics of painful osteoarthritis, a condition which affects millions of individuals worldwide. Specifically, a contribution from the TACR1 gene to modulating pain sensitivity in osteoarthritis is suggested.

Intergenic mRNAs. Minor gene products or tools of diversity?

The general model of gene expression implies that the units of genetic information, the genes, constitute the basis of the mRNA and protein products detected in living organisms. It is well known that in eukaryotic cells a single gene may give rise to various gene products due to alternative splicing and/or different positions of transcriptional start and termination. However, recent experimental results suggest that in addition to this variation in gene expression, another level of complexity may also exist as evidence for the presence of mRNAs that combine sequence information (exons) from distinct genes is accumulating. Moreover, the mechanisms that allow this production of intergenic mRNAs are starting to unravel and it appears that these follow two general pathways: (a) bypass of transcriptional termination, resulting in the generation of bicistronic pre-mRNAs; and (b) authentic trans-splicing events between pre-mRNAs of distinct genes.

Kinetics of formation of specific styrene oxide adducts in double-stranded DNA.

The possible carcinogenicity of styrene is believed to be related to the DNA-binding properties of styrene 7,8-oxide (SO). In order to compare the intrinsic reactivity of the different nucleophilic sites in DNA towards SO and to evaluate the candidates for human biomonitoring we have determined the second-order rate constants and stabilities of several SO-adducts in double-stranded DNA. These include alpha- and beta-isomers of N7-substituted and alphaN(2)-substituted guanines, alpha- and betaN3-substituted and alphaN(6)-substituted adenines as well as betaN3- and alphaN(4)-substituted cytosines. The highest rate constants were found for the spontaneously depurinating N7-guanines being ca. 3-15-fold higher than those for the stable adducts. When the relative proportions of different alkylation products were determined in course of time, after a single addition of SO, the labile N7-guanines and N3-adenines were the major products at early time points. After 144 h of incubation at 37 degrees C, alphaN(6)-SO-adenine and alphaN(2)-SO-guanine as well as betaN3-SO-uracil were the major adducts. Regarding human biomonitoring, the N7-substituted guanines should be one of the main targets because of the high reactivity of the N7-atom of guanine. However, in the case of chronic styrene exposures the chemically more stable DNA adducts may become important.

Intergenic transcripts containing a novel human cytochrome P450 2C exon 1 spliced to sequences from the CYP2C9 gene.

The cytochrome P450 2C (CYP2C) gene locus was found to include a novel exon 1 sequence with high similarity to the canonical exon 1 of CYP2C18. Rapid amplification of cDNA ends (RACE) and PCR amplifications of human liver cDNA revealed the presence of several intergenic species containing the CYP2C18 exon 1-like sequence spliced to different combinations of exonic and intronic sequences from the CYP2C9 gene. One splice variant was found to have an open reading frame starting at the canonical translation initiation codon of the CYP2C18 exon 1-like sequence. Another variant consisted of the nine typical CYP2C9 exons spliced after the CYP2C18 exon 1-like sequence through a segment of CYP2C9 5' flanking sequences. Moreover, analysis of bacterial artificial chromosome (BAC) clones revealed that the CYP2C18 exon 1-like sequence was located in the intergenic region between the CYP2C19 and CYP2C9 genes. The finding that a solitary exon is spliced with sequences from a neighboring gene may be interpreted as representing a general evolutionary mechanism aimed at using the full expression potential of a cell's genomic informational content.

Mutations in the N-terminus of VP5 alter its interaction with the scaffold proteins of herpes simplex virus type 1.

During the assembly process of herpes simplex virus type 1 capsids, there is an essential interaction between the C-terminal tail of the scaffold proteins (22a and 21) and the major capsid protein (VP5). Recent studies of spontaneous revertant viruses that overcome a blocked maturation cleavage site of the scaffold proteins have shown that the N-terminus of VP5 is important for this interaction. One of the revertant viruses, PR7, encodes a second-site mutation at residue 69 of VP5 which unlike wild-type VP5 fails to interact with 22a and thus gives white colonies in the yeast two-hybrid assay. In the present study a small DNA fragment, encoding residues 1 to 85 of wild-type and PR7 VP5, was mutagenized using error-prone PCR. Mutagenized DNA was used in the yeast two-hybrid assay to identify mutations in wild-type VP5 that resulted in loss of 22a binding (white colonies), or in PR7 VP5 that resulted in a gain of function (blue colonies). For the loss of function experiments, using KOS VP5, a row of eight thymidine nucleotides (codons 37-40) resulted in many frameshift mutations, which led us to terminate the study without reaching a statistically significant result. For the PR7 experiment, 30 clones were identified that had single amino acid substitutions, and these mutations were localized to amino acids 27-45 and 63-84 of VP5. The most frequent mutation was a reversion back to wild-type. The next most frequent were E28K and N63S, and these gave the highest beta-galactosidase enzyme activities (indicative of PR7VP5-22a interaction), 30 and 20% of wild-type, respectively. When E28K and N63S were transferred into the wild-type VP5 background, that is, in the absence of the PR7 mutation, they gave rise to different phenotypes. The E28K mutation lost its ability to interact with the scaffold proteins as judged by this assay. Therefore, it may be acting as a compensatory mutation whose phenotype is only expressed in the presence of the original PR7 mutation. However, the N63S mutation in the wild-type VP5 background increased the interaction, as judged by the beta-galactosidase activity, by a factor of 9 relative to when the PR7 mutation was present. Even more surprising, in the absence of the PR7 mutation the enzyme activity was still greater, by a factor of 2, than that observed for wild-type VP5. This study provides further evidence that the N-terminus of VP5 is in intimate association with the C-terminus of the scaffold proteins.

Second-site mutations encoding residues 34 and 78 of the major capsid protein (VP5) of herpes simplex virus type 1 are important for overcoming a blocked maturation cleavage site of the capsid scaffold proteins.

During assembly of the herpes simplex type 1 capsid, the major capsid protein VP5 interacts with the C-terminal residues of the scaffold proteins encoded by UL26 and UL26.5. Subsequent to capsid assembly the scaffold proteins are cleaved at the maturation site by a serine protease also encoded by UL26, thereby enabling the bulk of the scaffold proteins to be released from the capsid. Previously, a mutant virus (KUL26-610/611) was isolated in which this maturation cleavage site was blocked by replacing the Ala/Ser at the 610/611 cleavage site by Glu/Phe. This mutation was lethal and required a transformed cell line expressing wild-type UL26 gene products for growth. Although the mutation was lethal, spontaneous reversions occurred at a high frequency. Previously, a small number of revertants were isolated and all were found to have second-site mutations in VP5. The purpose of the present study was to do a comprehensive determination of the sites altered in VP5 by the second-site mutations. To do this, an additional 25 independent spontaneous revertants were characterized. Seven of the 25 arose by GC --> GT changes in codon 78, giving rise to an alanine to valine substitution. Four were the result of base changes at codon 34 but two different amino acids were produced as the changes were at different positions in the codon. Two mutations were detected at position 41 and mutations that occurred once were found at codons 69 and 80. Thus, 15 of the 25 second-site mutants were localized to codons 34 to 80 of VP5, which contains 1374 amino acids. The remaining 10 revertants had codon changes at nine different sites, of which the most N-terminal was altered at codon 187 and the most C-terminal at codon 1317. As noted in the much smaller study a preponderance of the second-site mutants in VP5 were altered in codons at the extreme N-terminus of VP5. It is especially noteworthy that 11 out of 25 of the mutations occurred at codons 34 and 78. As expected, all of the revertants isolated were shown to retain the original KUL26-610/611 mutation, and the scaffold proteins remain uncleaved. All showed decreased retention of VP24 in the B capsids compared to the wild-type KOS, but more than the KUL26-610/611 parental virus. The revertants all had decreased growth rates of 2 to 18% compared to that of KOS and showed varying degrees of sensitivity when grown at 39.5 degrees C. The mutations in VP5 of three of the previously isolated viruses (PR5, PR6, and PR7) were transferred into a wild-type background, i.e., a virus encoding wild-type UL26 and UL26.5 gene products. All replicated in nonpermissive (Vero) cells and cleaved scaffold proteins. PR5 and PR6 in the wild-type background gave wild-type burst sizes and gave C-capsids that retained VP24 at approximately wild-type levels. The third revertant, PR7, in the wild-type background showed only a twofold increase of burst size (to 20% of wild-type) and the capsids showed little or no increase of VP24 retention. Therefore, the second-site mutations of PR7 (R69C) by itself had a negative effect on virus replication. By contrast the temperature sensitivity of PR6 and PR7 remained unchanged in the wild-type background. Thus the temperature sensitivity of PR6 and PR7 resides in VP5 independently of the mutation in the UL26 cleavage site.

Treating pancreatitis in the acute care setting.

Acute pancreatitis is associated with significant complications and mortality. This article describes the pathophysiology and complications of, and treatments for acute pancreatitis, and the nurse's role in caring for a patient in the acute care setting.

Outcome of cardiopulmonary resuscitation and predictors of resuscitation status in an urban community teaching hospital.

Medical records of all expired patients as well as all patients designated on billing logs as having received cardiopulmonary resuscitation (CPR) during a 6-month period were reviewed. Patients were considered to have been 'coded' if they were found unresponsive and if the advanced cardiac life support (ACLS) protocol of the American Heart Association (AHA) was subsequently initiated. Of 105 patients who received CPR, 98 died during their hospital stay. Of the seven remaining patients, four had undergone coronary by-pass graft surgery, one was discharged in a persistent vegetative state, one died during an admission 2 months later, and one patient was transferred to another institution where he died. Various factors were studied in an effort to determine how patients on whom resuscitation was attempted differed from those who died without ever having received CPR. Patients who underwent CPR at least once during their hospitalization were more likely to have had cardiac diagnoses on admission (P < 0.001), to have been postoperative (P = 0.02), to have been admitted to a monitored bed on admission (P < 0.001) to have received more days of intensive care (P < 0.001) and to have received more specialist consultations (P = 0.004). Patients not receiving CPR were more likely to have had a primary diagnosis of neoplastic disease (P < 0.001), stroke or intracranial hemorrhage (P = 0.02) or dementia (P < 0.001). Age, race, or gender did not differ significantly between the two groups.

Nutritional support of pulmonary patients: a multidisciplinary approach.

Pulmonary disease affects and is affected by the nutritional status of the patient. The relationships between nutrition, medications, complications, and the course of pulmonary disease itself are multifaceted and are the focus of concern for the nutritional support team. Nutritional support of patients with pulmonary disease demands the expertise of a multidisciplinary team in monitoring the nutritional status of patients, appropriately selecting feeding solutions and routines, selecting and administering medications, and planning and implementing nursing interventions. Recognition of the importance of the nutritional component of care for patients with pulmonary disease is the focus of this article.

The status of cancer cluster investigations undertaken by state health departments.

A survey of state health departments indicates the number of cancer cluster reports received is associated with the size of the state, the presence of a population-based tumor registry, and the existence of a centralized system for response. Cancer cluster investigations, have generally been unproductive in terms of etiologic discoveries yet they may have important benefits in terms of public education, allaying public anxiety about environmental concerns and engendering good will toward government agencies.

Linker-insertion nonsense and restriction-site deletion mutations of the gB glycoprotein gene of herpes simplex virus type 1.

To study the effects of missense, nonsense, and deletion mutations of the gB glycoprotein gene of herpes simplex virus type 1, a gB-transformed cell line was isolated that, after virus infection, would express sufficient quantities of gB from the cellular chromosome to complement temperature-sensitive gB mutants. The transformed cell line was then used as a permissive cell to transfer two gB mutations from plasmid to viral DNA. One of the mutants, K082, harbored an HpaI linker insertion that introduced one new amino acid and a chain terminator codon within amino acid residue 43. The other mutant contained a 969-base-pair deletion in a part of the gene that includes the membrane-spanning region; a correspondingly shorter gB polypeptide was detected by sodium dodecyl sulfate-gel electrophoresis after immunoprecipitation of infected-cell extracts with four pooled monoclonal antibodies. No polypeptide was observed from K082-infected cells. The shortened gB polypeptide was efficiently processed and secreted into the growth medium. Each of the four monoclonal antibodies precipitated full-length gB, and three of the four precipitated the shortened polypeptide. Enveloped virus particles could be purified after infection of nonpermissive cells with either mutant virus. Virus particles appeared to possess normal polypeptide and glycopeptide profiles except for the absence of gB. Therefore, the presence of gB is not essential for viral assembly, including envelopment. Recombinants in virus stocks grown on the gB-transformed cells occurred at frequencies on the order of 10(-7) to 10(-5), compared with a frequency of approximately 10(-2) in mixed infections with the two mutants.

The Meaning in Life Scale: determining the reliability and validity of a measure.

The Meaning in Life (ML) Scale and Uniscale were developed to assess the sense of purpose, beliefs, and faith of patients in hospice and rehabilitative programs. Specialists have called for such instruments as meaning in life is not adequately measured by quality of life measures. The reliability and validity of the measures were tested with 257 English and French patients in long term care facilities in Montreal. The internal consistency of the responses to the 15 items in the ML Scale and the stability of the measures over a two week period were at acceptable levels. With respect to construct validity, the direction and magnitude of the correlation of the measures with those of subjective well-being, social support, pain, activities of daily living, quality of life, and social desirability were generally as predicted. Further research is required to determine the utility of the ML Scale and Uniscale in clinical research.

Silver-coated nylon fiber as an antibacterial agent.

A blend of nylon fiber and silver-coated nylon fiber (the latter known as X-static) was used in these experiments. This fiber was bactericidal when bacteria were exposed to it directly or to an extract derived from its prior incubation in salt solution. At ambient temperatures, a rapid exponential decrease of survival occurred, usually after a delay of approximately 1 h. The rate of killing (decrease of survival) increased with an increase in X-static percentage of the fiber blend, temperature of fiber extraction, concentration of Tris buffer present during extraction, and temperature at which bacteria were exposed to the extract. When bacteria were exposed to the extract at 37 degrees C as opposed to ambient temperature, there was no delay in onset of killing. Escherichia coli was generally the indicator organism tested, but comparable results were also found for Pseudomonas, Klebsiella, Staphylococcus, and Streptococcus species. The rate of killing increased with increasing silver ion concentration of the fiber extract, as determined through atomic absorption spectrophotometry. The rate of killing was greater and the onset was earlier with an extract containing silver ions from fiber than with a salt solution containing the same concentration of silver ions from silver nitrate. Studies of the kinetics of ion release suggested that X-static may be an effective, sustained-release antibacterial agent.

Expression in bacteria of gB-glycoprotein-coding sequences of Herpes simplex virus type 2.

A plasmid with an insert that encodes the glycoprotein B(gB) gene of Herpes simplex virus type 2 (HSV-2) has been isolated. DNA sequences coding for a portion of the HSV-2 gB peptide were cloned into a bacterial lacZ alpha expression vector and used to transform Escherichia coli. Upon induction of lacZpo-promoted transcription, some of the bacteria became filamentous and produced inclusion bodies containing a large amount of a 65-kDal peptide that was shown to be precipitated by broad-spectrum antibodies to HSV-2 and HSV-1. The HSV-2 insert of one of these clones specifies amino acid residues corresponding to 135 through 629 of the gB of HSV-1 [Bzik et al., Virology 133 (1984) 301-314].

The isolation and characterization of mutants of herpes simplex virus type 1 that induce cell fusion.

Six cell fusion-causing syn mutants were isolated from the KOS (syn-101 to syn-106) and three from the HFEM (syn-109) strains of herpes simplex virus type 1 (HSV-1). The mutants were studied by complementation and recombination with syn-20 (a syncytial mutant of KOS) and ts-B5 (a syncytial mutant of HFEM). Some studies also employed MP, a syncytium-inducing strain isolated from the non-syncytial parent, mP. Complementation and recombination of syn-20 and ts-B5 indicated that these two mutants were altered in two different virus genes. The recombination frequency between syn-20 and ts-B5 was very similar to that observed between MP and ts-B5, indicating the syn-20 and MP may represent alterations in the same virus gene. syn-101, syn-103, syn-104 and syn-105 were tentatively assigned to the syn-20 complementation group, while syn-107 and syn-109 were tentatively assigned to the ts-B5 complementation group, syn-106 and syn-108 were excluded from the ts-B5 group. syn-102 could not be excluded from either complementation group. syn-101 induced markedly less fusion at 38 degrees C relative to 34 degrees C. At 34 degrees C the patterns of syn-101-infected cell peptides and glycopeptides, examined by SDS-gel electrophoresis, were normal, but at 38 degrees C the amount of glycopeptide gC was particularly reduced. syn-102 produced decreased amounts of glycoproteins, and a non-glycosylated peptide, probably ICP6, was absent from extracts infected with syn-106.

Kinetics of cell fusion induced by a syncytia-producing mutant of herpes simplex virus type I.

We have isolated a number of plaque-morphology mutants from a strain of herpes simplex virus type I which, unlike the wild type, cause extensive cell fusion during a productive viral infection. After the onset of fusion, there is an exponential decrease in the number of single cells as a function of time after infection. At a multiplicity of infection (MOI) of 3.8 plaque-forming units per cell, fusion begins 5.3 h after infection with the number of single cells decreasing to 10% of the original number 10.2 h after infection. As the MOI is gradually increased from 0.4 to 8, the onset of fusion occurs earlier during infection. However, when the MOI is increased from 8 to 86, the onset of fusion does not occur any earlier. The rate of fusion is independent of the MOI for an MOI greater than 1. The rate of fusion varies linearly with initial cell density up to 3.5 X 10(4) cells/cm2 and is independent of initial cell density at higher cell concentrations. To assay cell fusion we have developed a smiple quantitative assay using a Coulter counter to measure the number of single cells as a function of time after infection. Data obtained using a Coulter counter are similar to those obtained with a microscope assay.