Data Resource: population level family justice administrative data with opportunities for data linkage.

INTRODUCTION: Although there has been considerable progress in the use of administrative data for applied health research, the family justice field lags behind. Better use of administrative data are essential to enhance understanding of how the family justice system is working, as well as the characteristics of, and outcomes for, children and families. The Family Justice Data Partnership (FJDP) supports this aim through analyses of core family justice and linked datasets in the SAIL Databank (Secure Anonymised Information Linkage). Cafcass Cymru provide expert advice for children involved in family court proceedings in Wales, ensuring decisions are made in the best interests of the child. We provide an overview of Cafcass Cymru data. We also describe and illustrate linkage to administrative datasets within SAIL. METHODS: Cafcass Cymru data was transferred to SAIL using a standardised approach to provide de-identified data with Anonymised Linking Fields (ALF) for successfully matched records. Three cohorts were created: all individuals involved in family court applications; all individuals with an ALF allowing subsequent health data linkage; and all individuals with a Residential Anonymised Linking Field (RALF) enabling area-level deprivation analysis. RESULTS: Cafcass Cymru application data are available for child protection matters (public law, range 2011-2019, n=12,745), and child arrangement disputes (private law, range 2005-2019, n=52,023). An 80% data linkage match rate was achieved. 40% had hospital admissions within two years pre or post application; 54% had emergency department attendances and 61% had outpatient appointments. Individuals were more likely to reside in deprived areas regardless of law type. CONCLUSION: Cafcass Cymru data can be accessed through the SAIL Databank. The FJDP will continue to enhance research opportunities for all to better understand the family justice system, and outcomes for those involved, such as health and wellbeing for children and family members.

Exploring barriers and solutions in advancing cross-centre population data science.

INTRODUCTION: It is widely acknowledged that population health and administrative data, especially when linked at the individual level, hold great value for research. Cross-centre working between data centres providing access to such data has the potential to further increase this value by effectively expanding the data available for research. However, there is limited published information on how to address the challenges and achieve success. The aim of this paper is to explore perceived barriers and solutions to inform developments in cross-centre working across data centres. METHODS: We carried out a narrative literature review on data sharing and cross centre working. We used a mixed methods approach to assess the opinions of members of the public on cross-centre data sharing, and the views and experiences of among data centre staff connected with the UK Farr Institute for Health Informatics Research. RESULTS: The literature review uncovered a myriad of practical and cultural issues. Our engagement with a public group suggested that cross-centre working involving anonymised data being moved between established centres is considered acceptable. The main themes emerging from discussions with data centre staff were dedicated resourcing, practical issues, information governance and culture. CONCLUSION: In seeking to advance cross-centre working between data centres, we conclude that there is a need for dedicated resourcing, indicators to recognise data reuse, collaboration to solve common issues, and balancing necessary barrier removal with incentivisation. This will require on-going commitment, engagement and an academic culture change.

A Profile of the SAIL Databank on the UK Secure Research Platform.

BACKGROUND: The Secure Anonymised Information Linkage (SAIL) Databank is a national data safe haven of de identified datasets principally about the population of Wales, made available in anonymised form to researchers across the world. It was established to enable the vast arrays of data collected about individuals in the course of health and other public service delivery to be made available to answer important questions that could not otherwise be addressed without prohibitive effort. The SAIL Databank is the bedrock of other funded centres relying on the data for research. APPROACH: SAIL is a data repository surrounded by a suite of physical, technical and procedural control measures embodying a proportionate privacy-by-design governance model, informed by public engagement, to safeguard the data and facilitate data utility. SAIL operates on the UK Secure Research Platform (SeRP), which is a customisable technology and analysis platform. Researchers access anonymised data via this secure research environment, from which results can be released following scrutiny for disclosure risk. SAIL data are being used in multiple research areas to evaluate the impact of health and social exposures and policy interventions. DISCUSSION: Lessons learned and their applications include: managing evolving legislative and regulatory requirements; employing multiple, tiered security mechanisms; working hard to increase analytical capacity efficiency; and developing a multi-faceted programme of public engagement. Further work includes: incorporating new data types; enabling alternative means of data access; and developing further efficiencies across our operations. CONCLUSION: SAIL represents an ongoing programme of work to develop and maintain an extensive, whole population data resource for research. Its privacy-by-design model and UK SeRP technology have received international acclaim, and we continually endeavour to demonstrate trustworthiness to support data provider assurance and public acceptability in data use. We strive for further improvement and continue a mutual learning process with our contemporaries in this rapidly developing field.

Validating the portal population of the United Kingdom Multiple Sclerosis Register.

The UK Multiple Sclerosis Register (UKMSR) is a large cohort study designed to capture 'real world' information about living with multiple sclerosis (MS) in the UK from diverse sources. The primary source of data is directly from people with Multiple Sclerosis (pwMS) captured by longitudinal questionnaires via an internet portal. This population's diagnosis of MS is self-reported and therefore unverified. The second data source is clinical data which is captured from MS Specialist Treatment centres across the UK. This includes a clinically confirmed diagnosis of MS (by Macdonald criteria) for consented patients. A proportion of the internet population have also been consented at their hospital making comparisons possible. This dataset is called the 'linked dataset'. The purpose of this paper is to examine the characteristics of the three datasets: the self-reported portal data, clinical data and linked data, in order to assess the validity of the self-reported portal data. The internet (n = 11,021) and clinical (n = 3,003) populations were studied for key shared characteristics. We found them to be closely matched for mean age at diagnosis (clinical = 37.39, portal = 39.28) and gender ratio (female %, portal = 73.1, clinical = 75.2). The Two Sample Kolmogorov-Smirnov test was for the continuous variables to examine is they were drawn from the same distribution. The null hypothesis was rejected only for age at diagnosis (D = 0.078, p < 0.01). The populations therefore, were drawn from different distributions, as there are more patients with relapsing disease in the clinical cohort. In all other analyses performed, the populations were shown to be drawn from the same distribution. Our analysis has shown that the UKMSR portal population is highly analogous to the entirely clinical (validated) population. This supports the validity of the self-reported diagnosis and therefore that the portal population can be utilised as a viable and valid cohort of people with Multiple Sclerosis for study.

On moving targets and magic bullets: Can the UK lead the way with responsible data linkage for health research?

PURPOSE: To provide an overview of essential elements of good governance of data linkage for health-related research, to consider lessons learned so far and to examine key factors currently impeding the delivery of good governance in this area. Given the considerable hurdles which must be overcome and the changing landscape of health research and data linkage, a principled, proportionate, risk-based approach to governance is advocated. DISCUSSION: In light of the considerable value of data linkage to health and well-being, the United Kingdom aspires to design and deliver good governance in health-related research. A string of projects have been asking: what does good governance look like in data linkage for health research? It is argued here that considerable progress can and must be made in order to develop the UK's contribution to future health and wealth economies, particularly in light of mis-start initiatives such as care.data in NHS England. Discussion centres around lessons learned from previous successful health research initiatives, identifying those governance mechanisms which are essential to achieving good governance. CONCLUSION: This article suggests that a crucial element in any step-increase of research capability will be the adoption of adaptive governance models. These must recognise a range of approaches to delivering safe and effective data linkage, while remaining responsive to public and research user expectations and needs as these shift and change with time and experience. The targets are multiple and constantly moving. There is not--nor should we seek--a single magic bullet in delivering good governance in health research.

A 5-year retrospective report of Gallibacterium anatis and Pasteurella multocida isolates from chickens in Mississippi.

A 5-yr retrospective study (November 2006-December 2011) was conducted to determine the isolation frequency of Pasteurella multocida and Gallibacterium anatis and their antibiograms from chickens submitted to the Mississippi Poultry Research and Diagnostic Laboratory. The number of isolations of G. anatis increased over the last 5 yr in broiler and broiler breeder type chickens. For P. multocida, the number of isolations increased from 2006 to 2010, but decreased through 2011 with all isolations being from boiler breeder type chickens. Gallibacterium anatis demonstrated almost complete resistance to novobiocin, tylosin, lincosamide, and tetracycline antimicrobials with moderate to high sensitivity to sulfonamides, fluoroquinolones, and florfenicol. There was intermediate sensitivity for spectinomycin and erythromycin and variable resistance to ß-lactam and aminoglycoside antimicrobials. In sharp contrast, P. multocida showed moderate to high sensitivity to ß-lactam, novobiocin, and tetracycline antimicrobials, but had antibiograms similar to G. anatis for the other antimicrobials. Sensitivities were determined using minimum inhibitory concentration. This study examines the trends over a 5-yr period of the number of isolates of P. multocida and G. anatis and their sensitivities. These 2 pathogens produce very similar clinical signs and lesions (fowl cholera-like) in breeders despite having extremely antagonistic sensitivity patterns. This study shows the necessity for producers to attempt culture and sensitivity in suspect fowl cholera-like flocks before initiating antimicrobial treatment commonly used with P. multocida for fear that the culprit may actually be the more antimicrobial-resistant G. anatis.

Urban legend versus rural reality: patients' experience of attendance at accident and emergency departments in west Wales.

OBJECTIVES: To investigate why and how patients decide to attend accident and emergency (A&E) departments, and to assess their satisfaction with the experience, in a predominantly rural west Wales population. METHODS: This was a semi-structured follow up telephone interview of patients who walked in to A&E in one of four general hospitals in west Wales and were triaged as Manchester Triage score 4 or 5. Patients were recruited by nurses during the period July-November 2002. The study sample consisted of 176 male and 145 female patients, mean (SD) age 36.6 (20.0) years. The main outcome measure was a quantitative and qualitative description of the recalled experiences of A&E attenders, the circumstances of their attendance, and their satisfaction with the experience. RESULTS: Of the study sample, 78% attended with injury or illnesses of recent origin, and 50% with actual or presumed musculoskeletal injury, 73% of which were sustained within 10 miles of home. Travel to hospital was by private transport for 86%, average distance 7.4 miles. The majority (90%) were registered with a local GP, but 32% felt A&E was the obvious choice, and a further 44% considered their GP inaccessible to their needs. Patients' reasons for seeking health care at A&E were similar to those described in an English urban study. Waiting times were rarely excessive; 80% left within 2 hours, and patient satisfaction was generally high. Among the 87 patients (27%) who reported a less satisfactory experience, 48 (55%) of these complained of dismissive attitudes of doctors. CONCLUSIONS: Anecdotal accounts of abuse of A&E services and unreasonable patient expectations gain the status of "urban legends" within the medical profession. Among the predominantly settled rural population in west Wales, there is little evidence of unreasonable patient expectations, and most patients report high satisfaction levels. Patients' bad experiences most frequently arise from a dismissive attitude on the part of medical staff. These attitudes are often consequent on an A&E culture that views some patients' attendances as less appropriate than others.

Randomised controlled trial comparing an acute paediatric hospital at home scheme with conventional hospital care.

AIMS: To assess the clinical effectiveness of a paediatric hospital at home service compared to conventional hospital care. METHODS: A total of 399 children suffering from breathing difficulty (n = 202), diarrhoea and vomiting (n = 125), or fever (n = 72) were randomised to Hospital at Home or in-patient paediatric care. Main outcome measures were: comparative clinical effectiveness as measured by readmission rate within three months (used as a proxy for parental coping with illness); and length of stay/care and comparative satisfaction of both patients and carers. RESULTS: Clinical effectiveness of both services was not significantly different. Length of care was one day longer in the Hospital at Home group; however, most parents and children preferred home care. CONCLUSIONS: Hospital at Home is a clinically acceptable form of care for these groups of acute paediatric illness. Readmission rates within three months failed to show any advantage in terms of parental coping. Parents and patients expressed a strong preference for hospital at home.

Effects of daily dermal application of DEET and epermethrin, alone and in combination, on sensorimotor performance, blood-brain barrier, and blood-testis barrier in rats.

DEET and permethrin were implicated in the development of illnesses in some veterans of the Persian Gulf War. This study was designed to investigate the effects of daily dermal application of these chemicals, alone or in combination, on the permeability of the blood-brain barrier (BBB) and blood-testes barrier (BTB) and on sensorimotor performance in male Sprague-Dawley rats. Groups of five rats were treated with a dermal daily dose of 4, 40, or 400 mg/kg DEET in ethanol or 0.013, 0.13, or 1.3 mg/kg permethrin in ethanol for 60 d. A group of 10 rats received a daily dermal dose of ethanol and served as controls. BBB permeability was assessed by injection of an iv dose of the quaternary ammonium compound [3H]hexamethonium iodide. While permethrin produced no effect on BBB permeability, DEET alone caused a decrease in BBB permeability in brainstem. A combination of DEET and permethrin significantly decreased the BBB permeability in the cortex. BTB permeability was decreased by treatment with DEET alone and in combination with permethrin. The same animals underwent a battery of functional behavior tests 30, 45, and 60 d after exposure to evaluate their sensorimotor abilities. All treatments caused a significant decline in sensorimotor performance in a dose- and time-dependent manner. These results show that daily dermal exposure to DEET, alone or in combination with permethrin, decreased BBB permeability in certain brain regions, and impaired sensorimotor performance.

Locomotor and sensorimotor performance deficit in rats following exposure to pyridostigmine bromide, DEET, and permethrin, alone and in combination.

Since their return from Persian Gulf War (PGW), many veterans have complained of symptoms including muscle and joint pain, ataxia, chronic fatigue, headache, and difficulty with concentration. The causes of the symptoms remain unknown. Because these veterans were exposed to a combination of chemicals including pyridostigmine bromide (PB), DEET, and permethrin, we investigated the effects of these agents, alone and in combination, on the sensorimotor behavior and central cholinergic system of rats. Male Sprague-Dawley rats (200-250 gm) were treated with DEET (40 mg/kg, dermal) or permethrin (0.13 mg/kg, dermal), alone and in combination with PB (1.3 mg/kg, oral, last 15 days only), for 45 days. Sensorimotor ability was assessed by a battery of behavioral tests that included beam-walk score, beam-walk time, incline plane performance, and forepaw grip on days 30 and 45 following the treatment. On day 45 the animals were sacrificed, and plasma and CNS cholinesterase, and brain choline acetyl transferase, muscarinic and nicotinic acetylcholine receptors were evaluated. Animals treated with PB, alone or in combination with DEET and permethrin, showed a significant deficit in beam-walk score as well as beam-walk time as compared with controls. Treatment with either DEET or permethrin, alone or in combination with each other, did not have a significant effect on beam-walk score. All chemicals, alone or in combination, resulted in a significant impairment in incline plane testing on days 30 and 45 following treatment. Treatment with PB, DEET, or permethrin alone did not have any inhibitory effect on plasma or brain cholinesterase activities, except that PB alone caused moderate inhibition in midbrain acetylcholinesterase (AChE) activity. Treatment with permethrin alone caused significant increase in cortical and cerebellar AChE activity. A combination of DEET and permethrin or PB and DEET led to significant decrease in AChE activity in brainstem and midbrain and brainstem, respectively. A significant decrease in brainstem AChE activity was observed following combined exposure to PB and permethrin. Coexposure with PB, DEET, and permethrin resulted in significant inhibition in AChE in brainstem and midbrain. No effect was observed on choline acetyl transferase activity in brainstem or cortex, except combined exposure to PB, DEET, and permethrin caused a slight but significant increase in cortical choline acetyltransferase activity. Treatment with PB, DEET, and permethrin alone caused a significant increase in ligand binding for m2 muscarinic acetylcholine receptor (mAChR) in the cortex. Coexposure to PB, DEET, and permethrin did not have any effect over that of PB-induced increase in ligand binding. There was no significant change in ligand binding for nicotinic acetylcholine receptor (nAChR) associated with treatment with the chemical alone; a combination of PB and DEET or coexposure with PB, DEET, and permethrin caused a significant increase in nAChR ligand binding in the cortex. Thus, these results suggest that exposure to physiologically relevant doses of PB, DEET, and permethrin, alone or in combination, leads to neurobehavioral deficits and region-specific alterations in AChE and acetylcholine receptors.

Acute sarin exposure causes differential regulation of choline acetyltransferase, acetylcholinesterase, and acetylcholine receptors in the central nervous system of the rat.

Acute neurotoxic effects of sarin (O:-isopropylmethylphosphonoflouridate) in male Sprague-Dawley rats were studied. The animals were treated with intramuscular (im) injections of either 1 x LD(50) (100 microg/kg), and sacrificed at 0. 5, 1, 3, 6, 15, or 20 h after treatment, or with im injections of either 0.01, 0.1, 0.5, or 1 x LD(50) and sacrificed 15 h after treatment. Plasma butyrylcholinesterase (BChE) and brain regional acetylcholinesterase (AChE) were inhibited (45-55%) by 30 min after the LD(50) dose. BChE in the plasma and AChE in cortex, brainstem, midbrain, and cerebellum remained inhibited for up to 20 h following a single LD(50) treatment. No inhibition in plasma BChE activity was observed 20 h after treatment with doses lower than the LD(50) dose. Midbrain and brainstem seem to be most responsive to sarin treatment at lower doses, as these regions exhibited inhibition (approximately 49% and 10%, respectively) in AChE activity following 0.1 x LD(50) treatment, after 20 h. Choline acetyltransferase (ChAT) activity was increased in cortex, brainstem, and midbrain 6 h after LD(50) treatment, and the elevated enzyme activity persisted up to 20 h after treatment. Cortex ChAT activity was significantly increased following a 0.1 x LD(50) dose, whereas brainstem and midbrain did not show any effect at lower doses. Treatment with an LD(50) dose caused a biphasic response in cortical nicotinic acetylcholine receptor (nAChR) and muscarinic acetylcholine receptor (m2-mAChR) ligand binding, using [(3)H]cytisine and [(3)H]AFDX-384 as ligands for nAChR and mAChR, respectively. Decreases at 1 and 3 h and consistent increases at 6, 15, and 20 h in nAChR and m2-mAChR were observed following a single LD(50) dose. The increase in nAChR ligand binding densities was much more pronounced than in mAChR. These results suggest that a single exposure of sarin, ranging from 0.1 to 1 x LD(50), modulates the cholinergic pathways differently and thereby causes dysregulation in excitatory neurotransmission.

Subchronic effects following a single sarin exposure on blood-brain and blood-testes barrier permeability, acetylcholinesterase, and acetylcholine receptors in the central nervous system of rat: a dose-response study.

Subchronic neurotoxic effects of sarin (O-isopropyl methylphosphonofluoridate) treatment at various doses in male Sprague Dawley rats were studied. The animals were treated with a single intramuscular (im) injection of 0.01, 0.1, 0.5, or 1 x LD50 (100 microg/kg). The animals were maintained for 90 d thereafter. [3H]Hexamethonium iodide was used to monitor the changes in blood-brain barrier (BBB) permeability in cortex, brainstem, midbrain, and cerebellum. Brainstem exhibited a significant decrease (approximately 58% of control) in uptake of [3H]hexamethonium iodide at 1 x LD50 dose. No significant changes were observed in BBB permeability in cortex, midbrain, and cerebellum at any dose. Plasma butyrylcholinesterase (BChE) activity remained unchanged, reflecting recovery of the enzyme activity from the initial inhibition following single exposure of 1 x LD50 sarin. Acetylcholinesterase (AChE) activity in the cortex remained inhibited (approximately 29%), whereas in the brainstem there was an increase (approximately 20%) at 1 x LD50 dose of sarin. The m2-selective muscarinic acetylcholine receptor (m2-mAChR) ligand binding was inhibited significantly at 1 x LD50 in the cortex, whereas brainstem showed significantly increased (approximately 45%) ligand binding at 1 x LD50 dose. Nicotinic acetylcholine receptor (nAChR), on the other hand, showed a biphasic response in ligand binding in the cortex with a decrease (approximately 30%) at 0.01 x LD50 but an increase (approximately 40%) at 1 x LD5O. Brainstem did not show any significant change in nAChR ligand binding. These results suggest that single exposure of sarin could lead to changes that may play an important role in neuropathological abnormalities in the central nervous system.

Gln368STOP myocilin mutation in families with late-onset primary open-angle glaucoma.

PURPOSE: To examine families ascertained for late-onset primary open-angle glaucoma (POAG) to determine mutations in the gene coding for myocilin. METHODS: The diagnosis of late-onset POAG was defined as age at diagnosis more than 35 years, intraocular pressure (IOP) 22 mm Hg or more in both eyes or 19 mm Hg or more while the patient was taking two glaucoma medications, glaucomatous optic neuropathy in both eyes, and visual field loss consistent with optic nerve damage in at least one eye of the proband. Two of three criteria were required in other family members. DNA from all families was screened for polymorphisms in myocilin using single-strand conformation polymorphism analysis. All polymorphisms were sequenced for mutations. RESULTS: Eighty-three affected people in 29 families with late-onset POAG were screened for mutations. Three mutations, two novel missense (Thr377Met and Glu352Lys) and one nonsense (Gln368STOP), were identified. The missense mutations did not segregate with the disease phenotype in these families. The nonsense mutation was found in 3 of 29 unrelated families with POAG. All affected family members and 8 of 12 in whom glaucoma was suspected had the Gln368STOP mutation. All people with this mutation had elevated IOP, and 78% had POAG by age 70. CONCLUSIONS: Three mutations were identified in the gene coding for myocilin in families with late-onset POAG. Of these, the Gln368STOP mutation was highly associated with the development of glaucoma. All people with this mutation had glaucoma or elevated IOP by age 70. In the United States, the Gln368STOP mutation in myocilin is strongly associated with the development of late-onset POAG. However, factors in addition to the presence of this mutation seem to play a role in the development of ocular hypertension and glaucoma in these families.

Adult-onset primary open angle glaucoma does not localize to chromosome 2cen-q13 in North American families.

Glaucoma is one of the leading causes of irreversible blindness in the world and is characterized by elevated intraocular pressure, optic nerve atrophy, and progressive visual field loss. Primary open angle glaucoma (POAG) is the most common subtype of glaucoma in the United States. Recently, Stoilova and coworkers [Genomics 1996;36:142-150] identified a locus for POAG on chromosome 2 (2cen-q13) in families primarily located in the United Kingdom. We examined families with POAG identified within the US for linkage to the 2cen-q13 locus. A total of 18 families with POAG were used in the analysis. Of 77 family members, 46 were classified as affected and 31 were either glaucoma suspects or considered normal. Eight highly polymorphic and informative markers flanking and distributed throughout the region were used. Parametric lod score analysis was performed using both a dominant and recessive low penetrance or 'affecteds-only' model. Multipoint affected sibpair exclusion mapping was also performed. Lod score (both models) and sibpair analysis excluded linkage of the POAG phenotype to the 2cen-q13 region in these families. These data suggest that the chromosome 2cen-q13 locus does not explain a substantial amount of genetic variation in familial POAG.

Cytotoxicity of tocopherols and their quinones in drug-sensitive and multidrug-resistant leukemia cells.

Cytotoxicities of tocopherols (alpha-T, gamma-T, delta-t), their para (alpha-TQ, gamma-TQ, delta-TQ)- and ortho (Tocored)-quinone oxidation products, the synthetic quinone analog of gamma-TQ containing a methyl group substituted for the phytyl side-chain (TMCQ) and the synthetic quinone analog of Tocored containing a methyl group substituted for the phytyl side-chain (PR) were measured in acute lymphoblastic leukemia cell lines that are drug-sensitive (CEM) and multidrug-resistant (CEM/VLB100). Among tocopherols, only delta-T exhibited cytotoxicity. Among para quinones, alpha-TQ showed no cytotoxicity, while gamma-TQ and delta-TQ were highly cytotoxic in both CEM and CEM/VLB100 cell lines (LD50 < 10 muM). delta-TQ and gamma-TQ were more cytotoxic than the widely studied chemotherapeutic agent doxorubicin, which also showed selective cytotoxicity to CEM cells. The orthoquinone Tocored was less cytotoxic than doxorubicin in drug-sensitive cells but more cytotoxic than doxorubicin in multidrug-resistant cells. Cytotoxicity was not a function of the phytyl side-chain since both TMCQ and PR were cytotoxic in leukemia cells. Cytotoxic para and ortho quinones were electrophiles that formed adducts with nucleophilic thiol groups in glutathione and 2-mercaptoethanol. Cytotoxicity was enhanced when the glutathione pool was depleted by preincubation with buthionine-[S,R]-sulfoximine, but cytotoxicity was diminished by the addition of N-acetylcysteine to cultures. alpha-T also diminished the cytotoxicity of para- and orthoquinones. Buthionine-[S,R]-sulfoximine did not block the inhibitory effect of either N-acetylcysteine or alpha-T, showing that these agents did not act solely by maintaining the glutathione pool as an essential antioxidant system. In conclusion, tocopherylquinones represent a new class of alkylating electrophilic quinones that function as highly cytotoxic agents and escape multidrug resistance in acute lymphoblastic leukemia cell lines.

Molecular analysis of EMS-induced frizzled mutations in Drosophila melanogaster.

The frizzled (fz) gene of Drosophila is essential for the development of normal tissue polarity in the adult cuticle of Drosophila. In fz mutants the parallel array of hairs and bristles that decorate the cuticle is disrupted. Previous studies have shown the fz encodes a membrane protein with seven putative transmembrane domains, and that it has a complex role in the development of tissue polarity, as there exist both cell-autonomous and cell nonautonomous alleles. We have now examined a larger number of alleles and found that 15 of 19 alleles display cell nonautonomy. We have examined these and other alleles by Western blot analysis and found that most fz mutations result in altered amounts of Fz protein, and many also result in a Fz protein that migrates aberrantly in SDS-PAGE. We have sequenced a subset of these alleles. Cell nonautonomous fz alleles were found to be associated with mutations that altered amino acids in all regions of the Fz protein. Notably, the four cell-autonomous mutations were all in a proline residue located in the presumptive first cytoplasmic loop of the protein. We have also cloned and sequenced the fz gene from D. virilis. Conceptual translation of the D. virilis open reading frame indicates that the Fz protein is unusually well conserved. Indeed, in the putative cytoplasmic domains the Fz proteins of the two species are identical.

Antioxidant and cytotoxic tocopheryl quinones in normal and cancer cells.

We found previously that [d]-alpha-tocopherol (alpha-T) and [d]-gamma-tocopherol (gamma-T) are lipid antioxidants (thiobarbituric acid test) in model systems containing arachidonic acid (AA), cumene hydroperoxide, and Fe3+ and in smooth muscle cell (SMC) cultures challenged with AA. We now show that [d]-alpha-tocopherylquinone (alpha-TQ), [d]-delta-tocopherylquinone (delta-TQ), and [d]-gamma-tocopherylquinone (gamma-TQ) are antioxidants at low concentrations and prooxidants at high concentrations in the model system. Prooxidant activity is greater with gamma-TQ than either alpha-TQ or delta-TQ. Low concentrations of alpha-TQ, delta-TQ, and gamma-TQ are also antioxidants in SMC cultures challenged with AA. Unlike alpha-TQ, partially substituted gamma-TQ and glutathione (GSH) form a Michael adduct which has been purified and characterized. We found previously that alpha-T, gamma-T, and alpha-TQ are mitogenic in SMC. We now report that both delta-TQ and gamma-TQ but not alpha-TQ show concentration-dependent cytotoxicity (changes in morphology, propidium iodide stain) in SMC cultures. Cytotoxicity is greater with gamma-TQ than delta-TQ. An acute lymphoblastic leukemia (ALL) cell line shows greater chemosensitivity (MTT and Neutral Red assays) to gamma-TQ than to either doxorubicin (DOX) or vinblastine (VLB). An ALL cell line resistant to both DOX and VLB retains the same chemosensitivity to gamma-TQ as the drug-sensitive ALL cell line. ALL cell lines are unaffected by either alpha-TQ or the GSH Michael adduct of gamma-TQ. These data show that partially substituted tocopheryl quinones capable of forming Michael adducts are potential chemotherapeutic agents for multidrug-resistant cancer cells.

Evidence of two pathways for the metabolism of phenol by Aspergillus fumigatus.

Aspergillus fumigatus (ATCC 28282), a thermotolerant fungus, has been shown to be capable of growth on phenol as the sole carbon and energy source. During growth of the organism on phenol, catechol and hydroquinone accumulated transiently in the medium; cells grown on phenol oxidised these compounds without a lag period. Two different routes operating simultaneously, leading to different ring-fission substrates, are proposed for the metabolism of phenol. In one route, phenol undergoes ortho-hydroxylation to give catechol, which is then cleaved by an intradiol mechanism leading to 3-oxoadipate. In the other route, phenol is hydroxylated in the para-position to produce hydroquinone, which is then converted into 1,2,4-trihydroxybenzene for ring fission by ortho-cleavage to give maleylacetate. Cell-free extracts of phenol-grown mycelia were found to contain enzymic activities for the proposed steps. Two ring-fission dioxygenases, one active towards 1,2,4-trihydroxybenzene, but not catechol, and one active towards both ring-fission substrates, were separated by FPLC. Succinate-grown mycelia did not oxidise any of the intermediates until a clear lag period had elapsed and did not contain any of the enzymic activities for phenol metabolism.