Research Publications and Outputs

A prospective cohort study of trends in selfpoisoning, Newcastle, Australia, 1987–2012: plus ça change, plus c’est la même chose

Thu, 01 Jan 2015 00:00:00 GMT

A prospective cohort study of trends in selfpoisoning, Newcastle, Australia, 1987–2012: plus ça change, plus c’est la même chose Buckley, Nicholas Objective: To examine inhospital mortality and morbidity associated with self-poisoning with different drug classes over an extended period. Design, setting and participants: A prospective cohort study over 26 years (1987–2012) with limited follow-up of patients presenting consecutively to a primary and tertiary referral toxicology centre covering Newcastle, Lake Macquarie and Port Stephens, Australia. Main outcome measures: Hospital length of stay, types of drugs ingested, intensive care unit (ICU) admission, requirement for ventilation, inhospital deaths and rates of antidepressant drug use in Australia. Results: Over the study period, there were 17 266 admissions of patients poisoned by 34 342 substances (16 723 drugs available only on prescription). The median length of stay was 16 hours, 12.2% of patients (2101/17 266) were admitted to an ICU, 7.4% (1281/17 266) were ventilated and 78 (0.45%) died in hospital. Patient demographics, social and psychiatric factors remained stable over the 26-year period, but case fatality decreased (from 0.77% [15/1955] to 0.17% [7/4060]) as did ICU admissions (19.2% [376/1955] to 6.9% [280/4060]), ventilation (13.7% [268/1955] to 4.8% [193/4060]) and LOS. The most frequently ingested substances were alcohol, benzodiazepines, paracetamol, antidepressants and antipsychotics. There was a substantial fall in some highly toxic drugs (tricyclic antidepressants, barbiturates, conventional antipsychotics and theophylline), but increases in less toxic selective serotonin reuptake inhibitors, serotonin–noradrenaline reuptake inhibitors and paracetamol. A greater than sixfold increase in community antidepressant use was accompanied by only minor changes in overall and antidepressant selfpoisoning rates. Conclusion: Over two decades, there were decreases in poisonings by many highly toxic drugs which were associated with substantial reductions in morbidity and inhospital deaths. Despite massive increases in the number of antidepressant prescriptions, neither rates of self-harm nor the proportion of antidepressant poisonings increased markedly.

Discontinuities and disruptions in drug dosage guidelines for the paediatric population

Wed, 07 Feb 2018 00:00:00 GMT

Discontinuities and disruptions in drug dosage guidelines for the paediatric population Chitty, Kate M.; Chan, Bosco; Pulanco, Camille L.; Luu, Sonya; Egunsola, Oluwaseun; Buckley, Nicholas A. AIMSThis study investigates paediatric drug dosage guidelines withthe aim of investigating their agreement with body surface area(BSA) scaling principles.METHODSA total of 454 drug dosage guidelines listed in the AMH-CDC 2015 were examined. Data extracted included the administration,frequency and dose per age bracket from 0 to 18 years. Drug treatments were categorized as follows: (1) The same dose rec-ommendation in milligrams per kilogram (mg kg 1) for all age/weights; (2) Change in the mg kg 1dosing according toage/weight; (3) Change in dose in mg according to age/weight; (4) Change from mg kg 1dosing to a dose in mg according toage/weight; (5) The same recommendation for all age/weight groups in mg; or (6) BSA dosing. Example drugs were selected toillustrate dose progression across ages.RESULTSMost drug treatments (63%) have the same mg kg 1dose for all age/weight groups, 14% are dosed in mg kg 1across all ageswith dose changes according to age/weight, 13% were dosed in mg across all ages with dose changes, 10% switched frommg kg 1to a set dose in mg, 4.2% have the same dose in mg for all age and weight groups and 2.2% are dosed according to BSA.CONCLUSIONSPaediatric dosage guidelines are based on weight-based formulas, available dosing formulations and prior patterns of use. Sub-stantial variation from doses predicted by BSA scaling are common, as are large shifts in recommended doses at age thresholds.Further research is required to determine if better outcomes could be achieved by adopting biologically based scaling of paedi-atric doses.

Vasoplegic Shock treated with Methylene Blue complicated by Severe Serotonin Syndrome.

Mon, 13 Nov 2017 00:00:00 GMT

Vasoplegic Shock treated with Methylene Blue complicated by Severe Serotonin Syndrome. Chan, Betty S.; Becker, Therese; Chiew, Angela L.; Abdalla, Ahmed M.; Robertson, Tom A.; Liu, Xin; Roberts, Michael S.; Buckley, Nicholas A. Introduction: Management of severe vasoplegic shock in overdose can be very challenging. We describe a case of severe refractory vasodilatory shock in poisoning where methylene blue (MB) was used with success. Case Report. A 70kg 15-year-old male presented 1.5 hours post ingestion of a large polypharmacy overdose of quetiapine slow release 1.5g, quetiapine immediate release 12g, desvenlafaxine slow release 5.6g, venlafaxine 1050mg, amlodipine 290mg, ramipril 100mg, fluoxetine 560mg, promethazine 500mg and an unknown amount of lithium. He developed severe vasoplegic shock that was resistant to maximal doses of noradrenaline and vasopressin. MB was administered 6.5 hour post ingestion. Within 1 hour there was an improvement in his haemodynamic status and reduction of catecholamine requirements. Twelve hours post ingestion, he developed severe serotonin syndrome that lasted 5 days as a result of interaction between MB, a reversible monoamine oxidase inhibitor, and the antidepressants taken in overdose. MB had a calculated half-life of 38 hours. Conclusion MB is a useful second or third line strategy for severe drug induced vasodilatory shock, and may be potentially life-saving. Conversely, physicians should be aware that it can interact with other drugs and cause life-threatening serotonin syndrome. Lower doses or shorter durations may be wise in patients at risk of this interaction.

Bromoxynil and 2-methyl-4-chlorophenoxyacetic acid (MCPA) poisoning could be a bad combination

Tue, 30 Jan 2018 00:00:00 GMT

Bromoxynil and 2-methyl-4-chlorophenoxyacetic acid (MCPA) poisoning could be a bad combination Chan, Betty S.H.; Angela L., Chiew; Grainger, Sarah; Page, Colin B.; Gault, Alan; Mostafa, Ahmed; Roberts, Michael S.; Buckley, Nicholas A.; Isbister, Geoffrey K.

Acute alcohol co-ingestion and hospital-treated deliberate self-poisoning; is there an effect on subsequent self-harm?

Thu, 01 Mar 2018 00:00:00 GMT

Acute alcohol co-ingestion and hospital-treated deliberate self-poisoning; is there an effect on subsequent self-harm? Borusso, LD; Buckley, Nicholas A.; Kirby, KA; Carter, G; Pilgrim, JL; Chitty, Kate M. The aim of this study was to determine the relationship between alcohol co-ingestion in an index deliberate self-poisoning (DSP) episode with repeated DSP and subsequent suicide. A retrospective cohort study was conducted involving 5,669 consecutive index presentations to a toxicology service following DSP between January 1, 1996, and October 31, 2010. Records were probabilistically matched to National Coronial Information System data to identify subsequent suicide. Index DSPs were categorized on co-ingestion of alcohol, and primary outcomes analyzed were repetition of any DSP, rates of repeated DSP, time to first repeat DSP, and subsequent suicide. Co-ingestion of alcohol occurred in 35.9% of index admissions. There was no difference between those who coingested alcohol (ALC+) and those who did not co-ingest alcohol (ALC ) in terms of proportion of repeat DSP, number of DSP events, or time to first repeat DSP event. Forty-one (1.0%) cases were probabilistically matched to a suicide death; there was no difference in the proportion of suicide between ALC+ and ALC- at 1 or 3 years. There was no significant relationship between the coingestion of alcohol in an index DSP and subsequent repeated DSP or suicide. Clinically, this highlights the importance of mental health assessment of patients that present after DSP, irrespective of alcohol co-ingestion at the time of event.

Co-ingested alcohol and the timing of deliberate self-poisonings

Wed, 01 Aug 2018 00:00:00 GMT

Co-ingested alcohol and the timing of deliberate self-poisonings Chitty, Kate M.; Kirby, Katharine; Osborne, Nicholas J.; Isbister, Geoffrey K.; Buckley, Nicholas A. Objective: Investigating diurnal variation in the timing of suicidal behaviours offers opportunity to better understand its various proximal risk factors. Acute use of alcohol is a potent proximal risk factor for suicidal behaviour, though the nature of this risk is poorly understood. The aim of this study was to compare the diurnal variation in time of poison ingestion between deliberate self-poisonings that involve alcohol versus those that do not. Methods: A retrospective analysis of consecutive presentations to a toxicology service following deliberate self-poisoning, 1996 - 2016. An independent samples Kolmogorov-Smirov test was performed to test the null hypothesis that the diurnal distribution of poison ingestion time was equal across selfpoisonings that did and did not involve alcohol co-ingestion. Presence of circadian rhythmicity was established using cosinor analysis. Results: 11088 deliberate self-poisoning records, for 7467 patients (60·8% females) were included in the analysis. 31·3% of the total records involved alcohol co-ingestion. Distribution of exposure time was significantly different between deliberate self-poisonings that did and did not involve alcohol (p < 0·001). The alcohol co-ingestion group showed a significantly greater prominent peak with poisoning occurring later in the evening (~2000 hours) compared to poisonings that did not involve alcohol (~1800 hours). Conclusion: This study exposed the differential diurnal patterns in deliberate self-poisoning according to the presence of alcohol co-ingestion. This analysis adds to the accumulating evidence that suicidal behaviour that involves alcohol co-ingestion represents a distinct subtype, which may be driven by alcohol consumption patterns in society. This also means that this large proportion of DSPs may not have otherwise have occurred if it were not for alcohol consumption, underscoring the importance of drug and alcohol services for alcohol related self-harm.

The Use of Cannabis and Cannabinoids in Treating Symptoms of Multiple Sclerosis: a Systematic Review of Reviews

Fri, 19 Oct 2018 00:00:00 GMT

The Use of Cannabis and Cannabinoids in Treating Symptoms of Multiple Sclerosis: a Systematic Review of Reviews Nielsen, Suzanne; Germanos, Rad; Weier, Megan; Pollard, John; Degenhardt, Louisa; Hall, Wayne; Buckley, Nicholas A. Purpose of review: Pharmaceutical cannabinoids such as nabiximols, nabilone and dronabinol, and plant-based cannabinoids have been investigated for their therapeutic potential in treating multiple sclerosis (MS) symptoms. This review of reviews aimed to synthesise findings from high-quality systematic reviews that examined the safety and effectiveness of cannabinoids in multiple sclerosis. We examined the outcomes of disability and disability progression, pain, spasticity, bladder function, tremor/ataxia, quality of life and adverse effects. Recent findings: We identified 11 eligible systematic reviews providing data from 32 studies, including 10 moderate- to high-quality RCTs. Five reviews concluded that there was sufficient evidence that cannabinoids may be effective for symptoms of pain and/or spasticity in MS. Few reviews reported conclusions for other symptoms. Summary: Recent high quality reviews find cannabinoids may have modest effects in MS for pain or spasticity. Future research should include studies with non-cannabinoid comparators is this an important gap in the evidence.

Person-level changes in oxycodone use after the introduction of a tamper-resistant formulation in Australia

Mon, 26 Mar 2018 00:00:00 GMT

Person-level changes in oxycodone use after the introduction of a tamper-resistant formulation in Australia Schaffer, Andrea L; Buckley, Nicholas A.; Degenhardt, Louisa; Larance, Briony; Cairns, Rose; Dobbins, Timothy A; Pearson, Sallie-Anne ABSTRACT BACKGROUND: Australia introduced tamper-resistant controlled-release (CR) oxycodone in April 2014. We quantified the impact of the reformulation on dispensing, switching and poisonings. METHODS: We performed interrupted time-series analyses using populationrepresentative national dispensing data from 2012 to 2016. We measured dispensing of oxycodone CR (≥ 10 mg), discontinuation of use of strong opioids and switching to other strong opioids after the reformulation compared with a historical control period. Similarly, we compared calls about intentional opioid poisoning using data from a regional poisons information centre. RESULTS: After the reformulation, dispensing decreased for 10–30 mg (total level shift –11.1%, 95% confidence interval [CI], –17.2% to –4.6%) and 40–80 mg oxycodone CR (total level shift –31.5%, 95% CI –37.5% to –24.9%) in participants less than 65 years of age but was unchanged in people 65 years of age or older. Compared with the previous year, discontinuation of use of strong opioids did not increase (adjusted hazard ratio [HR] 0.95, 95% CI 0.91 to 1.00), but switching to oxycodone/ naloxone did increase (adjusted HR 1.54, 95% CI 1.32 to 1.79). Switching to morphine varied by age (p < 0.001), and the greatest increase was in participants less than 45 years of age (adjusted HR 4.33, 95% CI 2.13 to 8.80). Participants switching after the reformulation were more likely to be dispensed a tablet strength of 40 mg or more (adjusted odds ratio [OR] 1.40, 95% CI 1.09 to 1.79). Calls for intentional poisoning that involved oxycodone taken orally increased immediately after the reformulation (incidence rate ratio (IRR) 1.31, 95% CI 1.05–1.64), but there was no change for injected oxycodone. INTERPRETATION: The reformulation had a greater impact on opioid access patterns of people less than 65 years of age who were using higher strengths of oxycodone CR. This group has been identified as having an increased risk of problematic opioid use and warrants closer monitoring in clinical practice.

Opioid use and harms associated with a sustained-release tapentadol formulation: a postmarketing study protocol

Fri, 23 Mar 2018 00:00:00 GMT

Opioid use and harms associated with a sustained-release tapentadol formulation: a postmarketing study protocol Peacock, Amy; Larance, Briony; Farrell, Michael; Cairns, Rose; Buckley, Nicholas A.; Degenhardt, Louisa Introduction It has been argued that tapentadol may pharmacologically have lower abuse potential than other pharmaceutical opioids currently available. However, there has been no comprehensive triangulation of data regarding use and harms associated with this formulation. A sustained-release formulation (SRF) of tapentadol (Palexia) was released in Australia in 2011 and listed for public subsidy in 2013. We summarise here the methods of a postmarketing study which will measure postintroduction: (1) population level availability, (2) extramedical use and diversion, (3) attractiveness for extramedical use and (4) associated harms, of tapentadol compared against other pharmaceutical opioids. Methods and analysis We evaluated key sources on pharmaceutical use and harms in Australia. This review indicateddata from four sources that disaggregate pharmaceutical opioid formulations and capture tapentadol SRF could be triangulated. These data sources comprised: (1) national pharmaceutical opioid community sales data from 2011 to 2017, (2) national pharmaceutical opioid poisonings reported to Poison Information Centres (PICs) from 2011 to 2017, (3) number of vendors on online marketplaces listing pharmaceutical opioids for sale and (4) data on pharmaceutical opioid extramedical use, attractiveness and harms from interviews with people who regularly inject drugs in Australia. Ethics and dissemination Ethics approval is not required for use of pharmaceutical sales data. Ethics approval has been obtained for use of national pharmaceutical opioid poisonings reported to PICs (LNR/16/SCHN/44) and for use of online marketplace data and interview data from people who inject drugs (HC12086). Key findings will be published mid-2018 in a peer-reviewed academic journal, and presented at various conferences and professional meetings.

Serum creatinine and cystatin C provide conflicting evidence of acute kidney injury following acute ingestion of potassium permanganate and oxalic acid

Tue, 23 May 2017 00:00:00 GMT

Serum creatinine and cystatin C provide conflicting evidence of acute kidney injury following acute ingestion of potassium permanganate and oxalic acid Wijerathna TM; Gawarammana IB; Dissanayaka DM; Palanagasinghe C; Shihana F; Dassanayaka G; Shahmy S; Endre ZH; Mohamed F; Buckley, Nicholas A. AIM: Acute kidney injury (AKI) is common following deliberate self-poisoning with a combination washing powder containing oxalic acid (H2C2O4) and potassium permanganate (KMnO4). Early and rapid increases in serum creatinine (sCr) follow severe poisoning. We investigated the relationship of these increases with direct nephrotoxicity in an ongoing multicenter prospective cohort study in Sri Lanka exploring AKI following poisoning. METHODS: Multiple measures of change in kidney function were evaluated in 48 consenting patients who had serial sCr and serum cystatin C (sCysC) data available. RESULTS: Thirty-eight (38/48, 79%) patients developed AKI (AKIN criteria). Twenty-eight (58%) had AKIN stage 2 or 3. Initial increases in urine creatinine (uCr) excretion were followed by a substantial loss of renal function. The AKIN stage 2 and 3 (AKIN2/3) group had very rapid rises in sCr (a median of 118% at 24 h and by 400% at 72 h post ingestion). We excluded the possibility that the rapid rise resulted from the assay used or muscle damage. In contrast, the average sCysC increase was 65% by 72 h. CONCLUSIONS: In most AKI, sCysC increases to the same extent but more rapidly than sCr, as sCysC has a shorter half-life. This suggests either a reduction in Cystatin C production or, conversely, that the rapid early rise of sCr results from increased production of creatine and creatinine to meet energy demands following severe oxidative stress mediated by H2C2O4 and KMnO4. Increased early creatinine excretion supports the latter explanation, since creatinine excretion usually decreases transiently in AKIN2/3 from other causes.

Clinical and Pharmacological Investigation of Myotoxicity in Sri Lankan Russell's Viper (Daboia russelii) Envenoming.

Fri, 02 Dec 2016 00:00:00 GMT

Clinical and Pharmacological Investigation of Myotoxicity in Sri Lankan Russell's Viper (Daboia russelii) Envenoming. Silva, A; Johnston, C; Kuruppu, S; Kneisz, D; Maduwage, K; Kleifeld O, O; Smith AI, AI; Siribaddana, S; Buckley, Nicholas A.; Hodgson, WC; Isbister, GK BACKGROUND: Sri Lankan Russell's viper (Daboia russelii) envenoming is reported to cause myotoxicity and neurotoxicity, which are different to the effects of envenoming by most other populations of Russell's vipers. This study aimed to investigate evidence of myotoxicity in Russell's viper envenoming, response to antivenom and the toxins responsible for myotoxicity. METHODOLOGY AND FINDINGS: Clinical features of myotoxicity were assessed in authenticated Russell's viper bite patients admitted to a Sri Lankan teaching hospital. Toxins were isolated using high-performance liquid chromatography. In-vitro myotoxicity of the venom and toxins was investigated in chick biventer nerve-muscle preparations. Of 245 enrolled patients, 177 (72.2%) had local myalgia and 173 (70.6%) had local muscle tenderness. Generalized myalgia and muscle tenderness were present in 35 (14.2%) and 29 (11.8%) patients, respectively. Thirty-seven patients had high (>300 U/l) serum creatine kinase (CK) concentrations in samples 24h post-bite (median: 666 U/l; maximum: 1066 U/l). Peak venom and 24h CK concentrations were not associated (Spearman's correlation; p = 0.48). The 24h CK concentrations differed in patients without myotoxicity (median 58 U/l), compared to those with local (137 U/l) and generalised signs/symptoms of myotoxicity (107 U/l; p = 0.049). Venom caused concentration-dependent inhibition of direct twitches in the chick biventer cervicis nerve-muscle preparation, without completely abolishing direct twitches after 3 h even at 80 μg/ml. Indian polyvalent antivenom did not prevent in-vitro myotoxicity at recommended concentrations. Two phospholipase A2 toxins with molecular weights of 13kDa, U1-viperitoxin-Dr1a (19.2% of venom) and U1-viperitoxin-Dr1b (22.7% of venom), concentration dependently inhibited direct twitches in the chick biventer cervicis nerve-muscle preparation. At 3 μM, U1-viperitoxin-Dr1a abolished twitches, while U1-viperitoxin-Dr1b caused 70% inhibition of twitch force after 3h. Removal of both toxins from whole venom resulted in no in-vitro myotoxicity. CONCLUSION: The study shows that myotoxicity in Sri Lankan Russell's viper envenoming is mild and non-life threatening, and due to two PLA2 toxins with weak myotoxic properties.

Treatments for Latrodectism-A Systematic Review on Their Clinical Effectiveness

Fri, 21 Apr 2017 00:00:00 GMT

Treatments for Latrodectism-A Systematic Review on Their Clinical Effectiveness Ryan, NM; Buckley, Nicholas A.; Graudins, A Latrodectism or envenomation by widow-spiders is common and clinically significant worldwide. Alpha-latrotoxin is the mammalian-specific toxin in the venom that results in toxic effects observed in humans. Symptoms may be incapacitating and include severe pain that can persist for days. The management of mild to moderate latrodectism is primarily supportive while severe cases have variously been treated with intravenous calcium, muscle relaxants, widow-spider antivenom and analgesic opioids. The object of this systematic review is to examine the literature on the clinical effectiveness of past and current treatments for latrodectism. MEDLINE, EMBASE and Google Scholar were searched from 1946 to December 2016 to identify clinical studies on the treatment of latrodectism. Studies older than 40 years and not in English were not reviewed. There were only two full-publications and one abstract of placebo-controlled randomised trials on antivenom use for latrodectism. Another two randomised comparative trials compared the route of administration of antivenom for latrodectism. There were fourteen case series (including two abstracts), fourteen case reports and one letter investigating drug treatments for latrodectism with the majority of these also including antivenom for severe latrodectism. Antivenom with opioid analgesia is often the major treatment reported for latrodectism however; recent high quality evidence has cast doubt on the clinical effectiveness of this combination and suggests that other treatments need to be investigated.

Nephrotoxicity-induced proteinuria increases biomarker diagnostic thresholds in acute kidney injury.

Mon, 03 Apr 2017 00:00:00 GMT

Nephrotoxicity-induced proteinuria increases biomarker diagnostic thresholds in acute kidney injury. Mohamed, F; Buckley, Nicholas A.; Pickering, JW; Wunnapuk, K; Dissanayake, S; Chathuranga, U; Gawarammana, I; Jayamanne, S; Endre, ZH BACKGROUND: Paraquat ingestion is frequently fatal. While biomarkers of kidney damage increase during paraquat-induced acute kidney injury (AKI), significant concurrent proteinuria may alter diagnostic thresholds for diagnosis and prognosis to an unknown extent. This study evaluated the effect of albuminuria on biomarker cutoffs for diagnosis and outcome prediction. METHODS: This was a multi-centre prospective clinical study of patients following acute paraquat self-poisoning in 5 Sri Lankan hospitals. Biomarker concentrations were quantified using ELISA and microbead assays and correlated with urinary albumin. Functional-AKI was defined by the Acute Kidney Injury Network serum creatinine definition and alternatively by a ≥50% increase in serum cystatin C. Albuminuria was defined as albumin-creatinine ratio >30 mg/g. The study outcomes were compared with a retrospective analysis of a pre-clinical study of paraquat-induced nephrotoxicity with appropriate controls. RESULTS: Albuminuria was detected in 34 of 50 patients, and increased with functional-AKI severity. The concentrations of uNGAL, uCysC, uClusterin, uβ2M, and uKIM-1 were higher in albuminuric compared to non-albuminuric patients (p < 0.001). Albuminuria correlated with biomarker concentration (r > 0.6, p < 0.01) and was associated with death (p = 0.006). Optimal biomarker cutoffs for prediction of death were higher in the albuminuric group. Similar outcomes with more detailed analysis were obtained in experimental paraquat nephrotoxicity. CONCLUSION: Albuminuria was associated with paraquat-induced nephrotoxicity and increased excretion of low-molecular weight protein biomarkers. AKI biomarker cutoffs for diagnosis, outcome prediction and AKI stratification increased in the presence of albuminuria. This may lead to over-diagnosis of AKI in conditions independently associated with proteinuria.

Diagnosis-based and external cause-based criteria to identify adverse drug reactions in hospital ICD-coded data: application to an Australia population-based study

Thu, 27 Apr 2017 00:00:00 GMT

Diagnosis-based and external cause-based criteria to identify adverse drug reactions in hospital ICD-coded data: application to an Australia population-based study Du, W; Pearson, SA; Buckley, Nicholas A.; Day, C; Banks, E OBJECTIVES: External cause International Classification of Diseases (ICD) codes are commonly used to ascertain adverse drug reactions (ADRs) related to hospitalisation. We quantified ascertainment of ADR-related hospitalisation using external cause codes and additional ICD-based hospital diagnosis codes. METHODS: We reviewed the scientific literature to identify different ICD-based criteria for ADR-related hospitalisations, developed algorithms to capture ADRs based on candidate hospital ICD-10 diagnoses and external cause codes (Y40-Y59), and incorporated previously published causality ratings estimating the probability that a specific diagnosis was ADR related. We applied the algorithms to the NSW Admitted Patient Data Collection records of 45 and Up Study participants (2011-2013). RESULTS: Of 493 442 hospitalisations among 267 153 study participants during 2011-2013, 18.8% (n = 92 953) had hospital diagnosis codes that were potentially ADR related; 1.1% (n = 5305) had high/very high-probability ADR-related diagnosis codes (causality ratings: A1 and A2); and 2.0% (n = 10 039) had ADR-related external cause codes. Overall, 2.2% (n = 11 082) of cases were classified as including an ADR-based hospitalisation on either external cause codes or high/very high-probability ADR-related diagnosis codes. Hence, adding high/very high-probability ADR-related hospitalisation codes to standard external cause codes alone (Y40-Y59) increased the number of hospitalisations classified as having an ADR-related diagnosis by 10.4%. Only 6.7% of cases with high-probability ADR-related mental symptoms were captured by external cause codes. CONCLUSION: Selective use of high-probability ADR-related hospital diagnosis codes in addition to external cause codes yielded a modest increase in hospitalised ADR incidence, which is of potential clinical significance. Clinically validated combinations of diagnosis codes could potentially further enhance capture.

A randomized controlled trial of fresh frozen plasma for coagulopathy in Russell's viper (Daboia russelii) envenoming

Thu, 16 Feb 2017 00:00:00 GMT

A randomized controlled trial of fresh frozen plasma for coagulopathy in Russell's viper (Daboia russelii) envenoming Isbister, G.K.; Jayamanne, S.; Mohamed, F.; Dawson, A.H.; Maduwage, K.; Gawarammana, I.; Lalloo, D.G.; de Silva, H.J.; Scorgie, F.E.; Lincz, L.F.; Buckley, Nicholas A. Essentials Russell's viper envenoming is a major health issue in South Asia and causes coagulopathy. We studied the effect of fresh frozen plasma and two antivenom doses on correcting coagulopathy. Fresh frozen plasma did not hasten recovery of coagulopathy. Low-dose antivenom did not worsen coagulopathy. SUMMARY: Background Russell's viper (Daboia russelii) envenoming is a major health issue in South Asia and causes venom-induced consumption coagulopathy (VICC). Objectives To investigate the effects of fresh frozen plasma (FFP) and two antivenom doses in correcting VICC. Methods We undertook an open-label randomized controlled trial in patients with VICC at two Sri Lankan hospitals. Patients with suspected Russell's viper bites and coagulopathy were randomly allocated (1 : 1) to high-dose antivenom (20 vials) or low-dose antivenom (10 vials) plus 4 U of FFP. The primary outcome was the proportion of patients with an International Normalized Ratio (INR) of < 2 at 6 h after antivenom administration. Secondary outcomes included anaphylaxis, major hemorrhage, death, and clotting factor recovery. Results From 214 eligible patients, 141 were randomized: 71 to high-dose antivenom, and 70 to low-dose antivenom/FFP; five had no post-antivenom blood tests. The groups were similar except for a delay of 1 h in antivenom administration for FFP patients. Six hours after antivenom administration, 23 of 69 (33%) patients allocated to high-dose antivenom had an INR of < 2, as compared with 28 of 67 (42%) allocated to low-dose antivenom/FFP (absolute difference 8%; 95% confidence interval - 8% to 25%). Fifteen patients allocated to FFP did not receive it. Severe anaphylaxis occurred equally frequently in each group. One patient given FFP developed transfusion-related acute lung injury. Three deaths occurred in low-dose antivenom/FFP patients, including one intracranial hemorrhage. There was no difference in recovery rates of INR or fibrinogen, but there was more rapid initial recovery of factor V and FX in FFP patients. Conclusion FFP after antivenom administration in patients with Russell's viper bites did not hasten recovery of coagulopathy. Low-dose antivenom/FFP did not worsen VICC, suggesting that low-dose antivenom is sufficient.

Mechanisms underlying early rapid increases in creatinine in paraquat poisoning

Fri, 27 Mar 2015 00:00:00 GMT

Mechanisms underlying early rapid increases in creatinine in paraquat poisoning Mohamed, F.; Endre, Z.; Jayamanne, S.; Pianta, T.; Peake, P.; Palangasinghe, C.; Chathuranga, U.; Jayasekera, K.; Wunnapuk, K.; Shihana, F.; Shahmy, S.; Buckley, Nicholas A. Background Acute kidney injury (AKI) is common after severe paraquat poisoning and usually heralds a fatal outcome. The rapid large increases in serum creatinine (Cr) exceed that which can be explained by creatinine kinetics based on loss of glomerular filtration rate (GFR). Methods and Findings This prospective multi-centre study compared the kinetics of two surrogate markers of GFR, serum creatinine and serum cystatin C (CysC), following paraquat poisoning to understand and assess renal functional loss after paraquat poisoning. Sixty-six acute paraquat poisoning patients admitted to medical units of five hospitals were included. Relative changes in creatinine and CysC were monitored in serial blood and urine samples, and influences of non-renal factors were also studied. Results Forty-eight of 66 patients developed AKI (AKIN criteria), with 37 (56%) developing moderate to severe AKI (AKIN stage 2 or 3). The 37 patients showed rapid increases in creatinine of >100% within 24 hours, >200% within 48 hours and >300% by 72 hours and 17 of the 37 died. CysC concentration increased by 50% at 24 hours in the same 37 patients and then remained constant. The creatinine/CysC ratio increased 8 fold over 72 hours. There was a modest fall in urinary creatinine and serum/urine creatinine ratios and a moderate increase in urinary paraquat during first three days. Conclusion Loss of renal function contributes modestly to the large increases in creatinine following paraquat poisoning. The rapid rise in serum creatinine most probably represents increased production of creatine and creatinine to meet the energy demand following severe oxidative stress. Minor contributions include increased cyclisation of creatine to creatinine because of acidosis and competitive or non-competitive inhibition of creatinine secretion. Creatinine is not a good marker of renal functional loss after paraquat poisoning and renal injury should be evaluated using more specific biomarkers of renal injury.

Kinetic Estimation of GFR Improves Prediction of Dialysis and Recovery after Kidney Transplantation

Mon, 04 May 2015 00:00:00 GMT

Kinetic Estimation of GFR Improves Prediction of Dialysis and Recovery after Kidney Transplantation Pianta, T. J.; Endre, Z. H.; Pickering, J. W.; Buckley, Nicholas A.; Peake, P. W. Background The early prediction of delayed graft function (DGF) would facilitate patient management after kidney transplantation. Methods In a single-centre retrospective analysis, we investigated kinetic estimated GFR under non-steady-state conditions, KeGFR, in prediction of DGF. KeGFRsCr was calculated at 4h, 8h and 12h in 56 recipients of deceased donor kidneys from initial serum creatinine (sCr) concentrations, estimated creatinine production rate, volume of distribution, and the difference between consecutive sCr values. The utility of KeGFRsCr for DGF prediction was compared with, sCr, plasma cystatin C (pCysC), and KeGFRpCysC similarly derived from pCysC concentrations. Results At 4h, the KeGFRsCr area under the receiver operator characteristic curve (AUC) for DGF prediction was 0.69 (95% CI: 0.56–0.83), while sCr was not useful (AUC 0.56, (CI: 0.41–0.72). Integrated discrimination improvement analysis showed that the KeGFRsCr improved a validated clinical prediction model at 4h, 8h, and 12h, increasing the AUC from 0.68 (0.52–0.83) to 0.88 (0.78–0.99) at 12h (p = 0.01). KeGFRpCysC also improved DGF prediction. In contrast, sCr provided no improvement at any time point. Conclusions Calculation of KeGFR from sCr facilitates early prediction of DGF within 4 hours of renal transplantation.

Clinical and Pharmacological Investigation of Myotoxicity in Sri Lankan Russell's Viper (Daboia russelii) Envenoming

Fri, 02 Dec 2016 00:00:00 GMT

Clinical and Pharmacological Investigation of Myotoxicity in Sri Lankan Russell's Viper (Daboia russelii) Envenoming Silva, A.; Johnston, C.; Kuruppu, S.; Kneisz, D.; Maduwage, K.; Kleifeld, O.; Smith, A. I.; Siribaddana, S.; Buckley, Nicholas A.; Hodgson, W. C.; Isbister, G. K. BACKGROUND: Sri Lankan Russell's viper (Daboia russelii) envenoming is reported to cause myotoxicity and neurotoxicity, which are different to the effects of envenoming by most other populations of Russell's vipers. This study aimed to investigate evidence of myotoxicity in Russell's viper envenoming, response to antivenom and the toxins responsible for myotoxicity. METHODOLOGY AND FINDINGS: Clinical features of myotoxicity were assessed in authenticated Russell's viper bite patients admitted to a Sri Lankan teaching hospital. Toxins were isolated using high-performance liquid chromatography. In-vitro myotoxicity of the venom and toxins was investigated in chick biventer nerve-muscle preparations. Of 245 enrolled patients, 177 (72.2%) had local myalgia and 173 (70.6%) had local muscle tenderness. Generalized myalgia and muscle tenderness were present in 35 (14.2%) and 29 (11.8%) patients, respectively. Thirty-seven patients had high (>300 U/l) serum creatine kinase (CK) concentrations in samples 24h post-bite (median: 666 U/l; maximum: 1066 U/l). Peak venom and 24h CK concentrations were not associated (Spearman's correlation; p = 0.48). The 24h CK concentrations differed in patients without myotoxicity (median 58 U/l), compared to those with local (137 U/l) and generalised signs/symptoms of myotoxicity (107 U/l; p = 0.049). Venom caused concentration-dependent inhibition of direct twitches in the chick biventer cervicis nerve-muscle preparation, without completely abolishing direct twitches after 3 h even at 80 μg/ml. Indian polyvalent antivenom did not prevent in-vitro myotoxicity at recommended concentrations. Two phospholipase A2 toxins with molecular weights of 13kDa, U1-viperitoxin-Dr1a (19.2% of venom) and U1-viperitoxin-Dr1b (22.7% of venom), concentration dependently inhibited direct twitches in the chick biventer cervicis nerve-muscle preparation. At 3 μM, U1-viperitoxin-Dr1a abolished twitches, while U1-viperitoxin-Dr1b caused 70% inhibition of twitch force after 3h. Removal of both toxins from whole venom resulted in no in-vitro myotoxicity. CONCLUSION: The study shows that myotoxicity in Sri Lankan Russell's viper envenoming is mild and non-life threatening, and due to two PLA2 toxins with weak myotoxic properties.

Dexamethasone Modifies Cystatin C-Based Diagnosis of Acute Kidney Injury During Cisplatin-Based Chemotherapy

Fri, 17 Mar 2017 00:00:00 GMT

Dexamethasone Modifies Cystatin C-Based Diagnosis of Acute Kidney Injury During Cisplatin-Based Chemotherapy Pianta, Timothy J.; Pickering, John W.; Succar, Lena; Chin, Melvin; Davidson, Trent; Buckley, Nicholas A.; Mohamed, Fahim; Endre, Zoltan H. BACKGROUND/AIMS: Plasma cystatin C (pCysC) may be superior to serum creatinine (sCr) as a surrogate of GFR. However, the performance of pCysC for diagnosing acute kidney injury (AKI) after cisplatin-based chemotherapy is potentially affected by accompanying corticosteroid anti-emetic therapy and hydration. METHODS: In a prospective observational study pCysC, sCr, urinary kidney injury molecule-1 (KIM-1), and urinary clusterin were measured over 2 weeks in 27 patients given first-cycle chemotherapy. The same variables were measured over 2 weeks in Sprague-Dawley rats given a single intraperitoneal injection of dexamethasone, cisplatin, or both, and in controls. RESULTS: In patients, pCysC increases were greater than sCr 41% vs. 16%, mean paired difference 25% (95% CI: 16-34%)], relative increases were ≥ 50% in 9 patients (35%) for pCysC compared with 2 (8%) for sCr (p = 0.04) and increases in sCr were accompanied by increased KIM-1 and clusterin excretion, but increases in pCysC alone were not. In rats, dexamethasone administration produced dose-dependent increases in pCysC (and augmented cisplatin-induced increases in pCysC), but did not augment histological injury, increases in sCr, or KIM-1 and clusterin excretion. CONCLUSIONS: In the presence of dexamethasone, elevation of pCysC does not reliably diagnose AKI after cisplatin-based chemotherapy.

Hippocampal glutamatergic/NMDA receptor functioning in bipolar disorder: a combined MMN and 1H-MRS study

Sun, 30 Aug 2015 00:00:00 GMT

Hippocampal glutamatergic/NMDA receptor functioning in bipolar disorder: a combined MMN and 1H-MRS study Chitty, Kate M.; Lagopoulos, Jim; Hickie, Ian B.; Hermens, Daniel F. Disturbances in the hippocampal glutamate (Glu)/N-methyl-d-aspartate (NMDA) system have been implicated in the pathophysiology of bipolar disorder (BD). Here we aim to provide a targeted integration of two measures of glutamatergic functioning in BD; the association between mismatch negativity (MMN) and in vivo hippocampal-Glu measured via proton magnetic resonance spectroscopy ((1)H MRS). Participants comprised of 33 patients with BD and 23 matched controls who underwent a two-tone passive, duration deviant MMN paradigm and (1)H MRS. Levels of Glu/creatine (Cr) in the hippocampus were determined. Pearson's correlations were used to determine associations between MMN and Glu/Cr. In controls, MMN amplitude was positively associated with Glu/Cr at the left temporal site. We did not find any significant associations with Glu/Cr and frontocentral MMN nor did we find any significant associations in BD patients. The results provide further insight into the neurophysiology of MMN, with evidence supporting the role of hippocampal-Glu signalling through the NMDA receptor in temporal MMN. Our data also demonstrate that Glu/Cr regulation of MMN is dampened in BD, which may indicate a lack of tightly regulated hippocampal NMDA functioning. These findings provide insight into the underlying basis of glutamatergic transmission disturbances implicated in the disorder.

Neuropsychological and Functional Outcomes in Recent-Onset Major Depression, Bipolar Disorder and Schizophrenia-Spectrum Disorders: A Longitudinal Cohort Study.

Tue, 28 Apr 2015 00:00:00 GMT

Neuropsychological and Functional Outcomes in Recent-Onset Major Depression, Bipolar Disorder and Schizophrenia-Spectrum Disorders: A Longitudinal Cohort Study. Lee, R. S.C.; Hermens, D. F.; Naismith, S. L.; Lagopoulos, J.; Jones, A.; Scott, J.; Chitty, K. M.; White, D.; Robillard, R.; Scott, E. M.; Hickie, I. B. Functional disability is the lead contributor to burden of mental illness. Cognitive deficits frequently limit functional recovery, although whether changes in cognition and disability are longitudinally associated in recent-onset individuals remains unclear. Using a prospective, cohort design, 311 patients were recruited and assessed at baseline. One hundred and sixty-seven patients met eligibility criteria (M=21.5 years old, s.d.=4.8) and returned for follow-up (M=20.6 months later, s.d.=7.8). Two-hundred and thirty participants were included in the final analysis, comprising clinically stable patients with major depression (n=71), bipolar disorder (BD; n=61), schizophrenia-spectrum disorders (n=35) and 63 healthy controls. Neuropsychological functioning and self-rated functional disability were examined using mixed-design, repeated-measures analysis, across diagnoses and cognitive clusters, covarying for relevant confounds. Clinical, neuropsychological and functional changes did not differ between diagnoses (all P>0.05). Three reliable neuropsychological subgroups emerged through cluster analysis, characterized by psychomotor slowing, improved sustained attention, and improved verbal memory. Controlling for diagnosis and changes in residual symptoms, clusters with improved neuropsychological functioning observed greater reductions in functional disability than the psychomotor slowing cluster, which instead demonstrated a worsening in disability (P<0.01). Improved sustained attention was independently associated with greater likelihood of follow-up employment (P<0.01). Diagnosis of BD uniquely predicted both follow-up employment and independent living. Neuropsychological course appears to be independently predictive of subjective and objective functional outcomes. Importantly, cognitive phenotypes may reflect distinct pathophysiologies shared across major psychiatric conditions, and be ideal targets for personalized early intervention.

Alcohol use in bipolar disorder: A neurobiological model to help predict susceptibility, select treatments and attenuate cortical insult.

Tue, 01 Sep 2015 00:00:00 GMT

Alcohol use in bipolar disorder: A neurobiological model to help predict susceptibility, select treatments and attenuate cortical insult. Chitty, Kate M.; Lagopoulos, Jim; Hickie, Ian B.; Hermens, Daniel F. In a series of neurophysiological and neuroimaging studies we investigated the neurobiology related to alcohol use in young people with bipolar disorder. Impairments were identified across frontal and temporal representations of event-related potential and proton magnetic resonance spectroscopy markers; mismatch negativity and in vivo glutathione, respectively. We propose these findings reflect impairments in the N-methyl-D-aspartate receptor and antioxidant capacity. This review seeks to place these findings within the broader literature in the context of two propositions: 1. Pathophysiological impairments in N-methyl-D-aspartate receptor functioning in bipolar disorder contribute to susceptibility toward developing alcohol problems. 2. Alcohol aggravates bipolar disorder neuroprogression via oxidative stress. A neurobiological model that incorporates these propositions is presented, with a focus on the potential for N-methyl-D-aspartate receptor antagonism and glutathione augmentation as potential adjunctive pharmacotherapies to treat the comorbidity. While this review highlights the importance of alcohol monitoring and reduction strategies in the treatment of bipolar disorder, the clinical impact of the proposed model remains limited by the lack of controlled trials of novel pharmacological interventions.

A longitudinal proton magnetic resonance spectroscopy study investigating oxidative stress as a result of alcohol and tobacco use in youth with bipolar disorder

Wed, 01 Apr 2015 00:00:00 GMT

A longitudinal proton magnetic resonance spectroscopy study investigating oxidative stress as a result of alcohol and tobacco use in youth with bipolar disorder Chitty, Kate M.; Lagopoulos, Jim; Hickie, Ian B.; Hermens, Daniel F. Alcohol and tobacco have been suggested to be "aggravating factors" for neuroprogression in bipolar disorder (BD), however the impact of these substances on the underlying neurobiology is limited. Oxidative stress is a key target for research into neuroprogression in BD and in accordance with this model, our previous cross-sectional studies have found that risky alcohol and tobacco use in BD is associated with increased oxidative stress, investigated via in vivo glutathione (GSH) measured by proton magnetic resonance spectroscopy ((1)H-MRS) in the anterior cingulate cortex (ACC). What remains unknown is whether the negative impact on GSH levels can be modified as a result of limiting alcohol and tobacco use. Thirty BD patients were included in the study. (1)H-MRS and tobacco and alcohol measures were conducted at baseline and follow-up assessments (15.5±4.6 months apart). Pearson׳s correlations were performed between percentage change in GSH concentration and changes in alcohol/tobacco use. Regression analyses were then conducted to further explore the significant correlations. An increase in GSH was associated with a decrease in alcohol consumption (r=-0.381, p<0.05) and frequency of tobacco use (-0.367, p=0.05). Change in alcohol consumption, tobacco use and age were significant predictors of change in GSH concentration (F (3, 26)=3.69, p<0.05). Due to the high comorbidity of alcohol and tobacco use in the sample, the individual effects of these substances on GSH levels could not be determined. This study offers longitudinal evidence that changing risky drinking patterns and tobacco use early in the course of BD is associated with improvements in antioxidant capacity, and therefore may be specific targets for early intervention and prevention of neuroprogression in BD.

Prediction of paraquat exposure and toxicity in clinically ill poisoned patients: a model based approach.

Fri, 19 Sep 2014 00:00:00 GMT

Prediction of paraquat exposure and toxicity in clinically ill poisoned patients: a model based approach. Wunnapuk, Klintean; Mohammed, Fahim; Gawarammana, Indika; Liu, Xin; Verbeeck, Roger K.; Buckley, Nicholas A.; Roberts, Michael S.; Musuamba, Flora T. AIMS: Paraquat poisoning is a medical problem in many parts of Asia and the Pacific. The mortality rate is extremely high as there is no effective treatment. We analyzed data collected during an ongoing cohort study on self-poisoning and from a randomized controlled trial assessing the efficacy of immunosuppressive therapy in hospitalized paraquat-intoxicated patients. The aim of this analysis was to characterize the toxicokinetics and toxicodynamics of paraquat in this population. METHODS: A non-linear mixed effects approach was used to perform a toxicokinetic/toxicodynamic population analysis in a cohort of 78 patients. RESULTS: The paraquat plasma concentrations were best fitted by a two compartment toxicokinetic structural model with first order absorption and first order elimination. Changes in renal function were used for the assessment of paraquat toxicodynamics. The estimates of toxicokinetic parameters for the apparent clearance, the apparent volume of distribution and elimination half-life were 1.17 l h(-1) , 2.4 l kg(-1) and 87 h, respectively. Renal function, namely creatinine clearance, was the most significant covariate to explain between patient variability in paraquat clearance.This model suggested that a reduction in paraquat clearance occurred within 24 to 48 h after poison ingestion, and afterwards the clearance was constant over time. The model estimated that a paraquat concentration of 429 μg l(-1) caused 50% of maximum renal toxicity. The immunosuppressive therapy tested during this study was associated with only 8% improvement of renal function. CONCLUSION: The developed models may be useful as prognostic tools to predict patient outcome based on patient characteristics on admission and to assess drug effectiveness during antidote drug development.

Household water efficiency strategies in Cornwall, SW of England

Mon, 01 Jun 2015 00:00:00 GMT

Household water efficiency strategies in Cornwall, SW of England Sharpe, Richard A.; Osborne, Nicholas J.; Skerratt, Glynn Demand-side measures are thought to be a sustainable approach to meeting the future supply-demand balance. We assess the uptake of domestic demand-side measures and potential factors that may promote the uptake of water efficiency devices. Fifty-one face-to-face questionnaires were used to collect demographic and household characteristics data. We use descriptive statistics and univariate models to assess factors promoting water efficiency. Fifty-one adult participants aged between 30 and 64 years provided data on water consumption and efficiency. Participants investigating water saving solutions and homeowners were more likely to utilise water efficiency devices. Targeted factors shown to promote consumer up-take of water efficiency measures along with strategies utilising low-cost efficiency devices provide a cost-effective means to reduce water consumption.

Kidney damage biomarkers detect acute kidney injury but only functional markers predict mortality after paraquat ingestion

Tue, 09 Jun 2015 00:00:00 GMT

Kidney damage biomarkers detect acute kidney injury but only functional markers predict mortality after paraquat ingestion Mohamed, Fahim; Buckley, Nicholas A.; Jayamanne, Shaluka; Pickering, John W.; Peake, Philip; Palangasinghe, Chathura; Wijerathna, Thilini; Ratnayake, Indira; Shihana, Fathima; Endre, Zoltan H. Acute kidney injury (AKI) is common following paraquat ingestion. The diagnostic performance of injury biomarkers was investigated in serial blood and urine samples from patients from 5 Sri Lankan hospitals. Functional AKI was diagnosed using serum creatinine (sCr) or serum cystatin C (sCysC). The 95th centile in healthy subjects defined the urinary biomarker cutoffs for diagnosing structural AKI. 50 poisoned patients provided 2 or more specimens, 76% developed functional AKI [AKIN stage 1 (n=12), 2 (n=7) or 3 (n=19)]; 19/26 patients with AKIN stage 2/3 also had functional AKI by sCysC criteria (≥50% increase). Urinary cystatin C (uCysC), clusterin (uClu) and NGAL (uNGAL) increased within 24h of ingestion compared with NoAKI patients and healthy controls. Each biomarker demonstrated moderate diagnostic utility [AUC-ROC: uCysC 0.79, uNGAL 0.79, uClu 0.68] for diagnosis of functional AKI at 16h. Death occurred only in subjects with functional AKI. Structural biomarker-based definitions detected more AKI than did sCr or sCysC, but did not independently predict death. Renal injury biomarkers did not add clinical value to patients who died rapidly due to multi-organ failure. Use of injury biomarkers within 16-24h may guide early intervention for reno-protection in less severe paraquat poisoning.

Fuel poverty increases risk of mould contamination, regardless of adult risk perception & ventilation in social housing properties

Mon, 01 Jun 2015 00:00:00 GMT

Fuel poverty increases risk of mould contamination, regardless of adult risk perception & ventilation in social housing properties Sharpe, Richard A.; Thornton, Christopher R.; Nikolaou, Vasilis; Osborne, Nicholas J. INTRODUCTION: Fuel poverty affects 2.4 million UK homes leading to poor hygrothermal conditions and risk of mould and house dust mite contaminations, which in turn increases risk of asthma exacerbation. For the first time we assess how fuel poverty, occupants' risk perception and use of mechanical ventilation mediate the risk of mould contamination in social housing. METHODS: Postal questionnaires were sent to 3867 social housing properties to collect adult risk perception, and demographic and environmental information on occupants. Participant details were linked to data pertaining to the individual properties. Multiple logistic regression was used to calculate odds ratios and confidence intervals while allowing for clustering of individuals coming from the same housing estate. We used Structured Equation Modelling and Goodness of Fit analysis in mediation analyses to examine the role of fuel poverty, risk perception, use of ventilation and energy efficiency. RESULTS: Eighteen percent of our target social housing populations (671 households) were included into our study. High risk perception (score of 8-10) was associated with reduced risk of mould contamination in the bedrooms of children (OR 0.5 95% CI; 0.3-0.9) and adults (OR 0.4 95% CI; 0.3-0.7). High risk perception of living with inadequate heating and ventilation reduced the risk of mould contamination (OR 0.5 95% CI; 0.3-0.8 and OR 0.5 95% CI; 0.3-0.7, respectively). Participants living with inadequate heating and not heating due to the cost of fuel had an increased risk of mould contamination (OR 3.4 95% CI; 2.0-5.8 and OR 2.2 95% CI; 1.5-3.2, respectively). Increased risk perception and use of extractor fans did not mediate the association between fuel poverty behaviours and increased risk of mould contamination. DISCUSSION: Fuel poverty behaviours increased the risk of mould contamination, which corresponds with existing literature. For the first time we used mediation analysis to assess how this association maybe modified by occupant behaviours. Increased risk perception and use of extractor fans did not modify the association between fuel poverty and mould contamination. This suggests that fuel poor populations may not benefit from energy efficiency interventions due to ineffective heating and ventilation practices of those occupants residing participating households. Our findings may be modified by a complex interaction between occupant behaviours and the built environment. We found that participant age, occupancy, SES, pets, drying washing indoors, geographic location, architectural design/age of the property, levels of insulation and type of heating regulated risk of mould contamination. CONCLUSION: Fuel poverty behaviours affected around a third of participating households and represent a risk factor for increased exposures to damp and mouldy conditions, regardless of adult risk perception, heating and ventilation practices. This requires multidisciplinary approach to assess the complex interaction between occupant behaviours, risk perception, the built environment and the effective use of heating and ventilation practices. STUDY IMPLICATIONS: Our findings have implications for housing policies and future housing interventions. Effective communication strategies focusing on awareness and perception of risk may help address indoor air quality issues. This must be supported by improved household energy efficiency with the provision of more effective heating and ventilation strategies, specifically to help alleviate those suffering from fuel poverty.

Functional Analysis of Novel Polymorphisms in the Human SLCO1A2 Gene that Encodes the Transporter OATP1A2

Tue, 06 Aug 2013 00:00:00 GMT

Functional Analysis of Novel Polymorphisms in the Human SLCO1A2 Gene that Encodes the Transporter OATP1A2 Zhou, Fanfan; Zheng, Jian; Zhu, Ling; Jodal, Andreas; Cui, Pei H.; Wong, Mark; Gurney, Howard; Church, W. Bret; Murray, Michael The solute carrier organic anion transporting polypeptide 1A2 (OATP1A2, SLCO1A2) is implicated in the cellular influx of a number of drugs. We identified five novel single nucleotide polymorphisms (SNPs) in coding exons of the SLCO1A2 gene in a cohort of subjects: G550A, G553A, G673A, A775C, and G862A, that encoded the OATP1A2 variants E184K, D185N, V255I, T259P, and D288N, respectively. The function and expression of these variant transporters were assessed in HEK-293 cells. We found that the novel variants, E184K, D185N, T259P, and D288N, were associated with impaired estrone-3-sulfate, imatinib, and methotrexate transport (∼20-50% of wild-type control); function was retained by OATP1A2-V255I. From biotinylation assays, the decreased function of these variants was due, at least in part, to impaired plasma membrane expression. The four loss-of-function variants were studied further using mutagenesis to produce variants that encode residues with different charges or steric properties. From immunoblotting, the replacement of negatively charged residues at amino acid positions 184 and 185 impaired membrane expression, while either a positive or negative charge at residue 288 supported the correct membrane targeting of OATP1A2. Replacement of T259 with bulky residues disrupted transporter stability. From molecular models, E184, D185, and D288 were located near several charged residues such that intramolecular ionic interactions may stabilize the transporter structure. Individuals who carry these novel SNPs in the SLCO1A2 gene may be at risk from impaired efficacy or enhanced toxicity during treatment with drugs that are substrates for OATP1A2.

PDZK1 and NHERF1 Regulate the Function of Human Organic Anion Transporting Polypeptide 1A2 (OATP1A2) by Modulating Its Subcellular Trafficking and Stability

Fri, 11 Apr 2014 00:00:00 GMT

PDZK1 and NHERF1 Regulate the Function of Human Organic Anion Transporting Polypeptide 1A2 (OATP1A2) by Modulating Its Subcellular Trafficking and Stability Zheng, Jian; Chan, Ting; Cheung, Florence Shin Gee; Zhu, Ling; Murray, Michael; Zhou, Fanfan The human organic anion transporting polypeptide 1A2 (OATP1A2) is an important membrane protein that mediates the cellular influx of various substances including drugs. Previous studies have shown that PDZ-domain containing proteins, especially PDZK1 and NHERF1, regulate the function of related membrane transporters in other mammalian species. This study investigated the role of PDZK1 and NHERF1 in the regulation of OATP1A2 in an in vitro cell model. Transporter function and protein expression were assessed in OATP1A2-transfected HEK-293 cells that co-expressed PDZK1 or NHERF1. Substrate (estrone-3-sulfate) uptake by OATP1A2 was significantly increased to ,1.6- (PDZK1) and ,1.8- (NHERF1) fold of control; this was dependent on the putative PDZ-binding domain within the C-terminus of OATP1A2. The functional increase of OATP1A2 following PDZK1 or NHERF1 over-expression was associated with increased transporter expression at the plasma membrane and in the whole cell, and was reflected by an increase in the apparent maximal velocity of estrone-3-sulfate uptake (Vmax: 138.964.1 (PDZK1) and 181.4616.7 (NHERF1) versus 55.563.2 pmol*(mg*4 min)21 in control; P,0.01). Co-immunoprecipitation analysis indicated that the regulatory actions of PDZK1 and NHERF1 were mediated by direct interaction with OATP1A2 protein. In further experiments PDZK1 and NHERF1 modulated OATP1A2 expression by decreasing its internalization in a clathrin-dependent (but caveolin-independent) manner. Additionally, PDZK1 and NHERF1 enhanced the stability of OATP1A2 protein in HEK-293 cells. The present findings indicated that PDZK1 and NHERF1 regulate the transport function of OATP1A2 by modulating protein internalization via a clathrin-dependent pathway and by enhancing protein stability.

NEDD9 regulates 3D migratory activity independent of the Rac1 morphology switch in glioma and neuroblastoma.

Sat, 01 Feb 2014 00:00:00 GMT

NEDD9 regulates 3D migratory activity independent of the Rac1 morphology switch in glioma and neuroblastoma. Zhong, Jessie; Bach, Cuc T.; Shum, Michael S.Y.; O'Neill, Geraldine M. Metastasizing tumor cells must transmigrate the dense extracellular matrix that surrounds most organs. The use of three-dimensional (3D) collagen gels has revealed that many cancer cells can switch between different modes of invasion that are characterized by distinct morphologies (e.g., rounded vs. elongated). The adhesion protein NEDD9 has the potential to regulate the switch between elongated and rounded morphologies; therefore, its role was interrogated in the invasion switch of glioblastoma and neuroblastoma tumors that similarly derive from populations of neural crest cells. Interestingly, siRNA-mediated depletion of NEDD9 failed to induce cell rounding in glioma or neuroblastoma cells, contrasting the effects that have been described in other tumor model systems. Given that Rac1 GTPase has been suggested to mediate the switch between elongated and rounded invasion, the functionality of the Rac1 morphology switch was evaluated in the glioma and neuroblastoma cells. Using both dominant-negative Rac1 and Rac1-specific siRNA, the presence of this morphologic switch was confirmed in the neuroblastoma, but not in the glioma cells. However, in the absence of a morphologic change following NEDD9 depletion, a significant decrease in the cellular migration rate was observed. Thus, the data reveal that NEDD9 can regulate 3D migration speed independent of the Rac1 morphology switch. IMPLICATIONS: NEDD9 targeting is therapeutically viable as it does not stimulate adaptive changes in glioma and neuroblastoma invasion.

Role of dynamin in elongated cell migration in a 3D matrix.

Sun, 01 Mar 2015 00:00:00 GMT

Role of dynamin in elongated cell migration in a 3D matrix. Lees, Justin G.; Gorgani, NN; Ammit, AJ; McCluskey, A; Robinson, PJ; O'Neill, GM The use of 3-dimensional (3D) collagen gels has yielded new insights into the migratory behaviour of cancer cells. While the large GTPase dynamin has emerged as an important regulator of cancer cell migration and invasion under 2D conditions, its role in 3D migration is unclear. We have used a potent dynamin modulator, a bis-tyrphostin derivative, Ryngo® 1-23, to investigate the role of dynamin in 3D migration in 3 different cell lines. The compound specifically inhibits persistent, elongated 3D migration in U87MG and SMA-560 cells. Treated U87MG cells adopt a rounded morphology that is not due to apoptosis, loss of matrix metalloprotease activity or inhibition of clathrin-mediated endocytosis. Given that Ryngo 1-23 is known to regulate dynamin oligomerisation and actin dynamics at the leading edge, we analysed actin filament distribution. Ryngo 1-23 induced a switch in actin filament organization in 3D cultures resulting in the generation of multiple short actin-rich microspikes. Correlated with the change in actin filament distribution, cells displayed reduced collagen gel contraction. Since acto-myosin force transmission to the extra-cellular matrix underpins persistent, elongated migration, our results suggest that Ryngo 1-23 modulates this process in 3D migration via dynamin-mediated regulation of acto-myosin force transmission to the extra-cellular matrix.

Tropomyosin Regulates Cell Migration during Skin Wound Healing

Fri, 01 May 2015 00:00:00 GMT

Tropomyosin Regulates Cell Migration during Skin Wound Healing Lees, Justin G.; Ching, Yu Wooi; Adams, Damian H.; Bach, Cuc T.T.; Samuel, Michael S.; Kee, Anthony J.; Hardeman, Edna C.; Gunning, Peter; Cowin, Allison J.; O’Neill, Geraldine M. Precise orchestration of actin polymer into filaments with distinct characteristics of stability, bundling, and branching underpins cell migration. A key regulator of actin filament specialization is the tropomyosin family of actin-associating proteins. This multi-isoform family of proteins assemble into polymers that lie in the major groove of polymerized actin filaments, which in turn determine the association of molecules that control actin filament organization. This suggests that tropomyosins may be important regulators of actin function during physiological processes dependent on cell migration, such as wound healing. We have therefore analyzed the requirement for tropomyosin isoform expression in a mouse model of cutaneous wound healing. We find that mice in which the 9D exon from the TPM3/γTm tropomyosin gene is deleted (γ9D -/-) exhibit a more rapid wound-healing response 7 days after wounding compared with wild-type mice. Accelerated wound healing was not associated with increased cell proliferation, matrix remodeling, or epidermal abnormalities, but with increased cell migration. Rac GTPase activity and paxillin phosphorylation are elevated in cells from γ9D -/- mice, suggesting the activation of paxillin/Rac signaling. Collectively, our data reveal that tropomyosin isoform expression has an important role in temporal regulation of cell migration during wound healing.

Lipid analogues as potential drugs for the regulation of mitochondrial cell death

Fri, 28 Mar 2014 00:00:00 GMT

Lipid analogues as potential drugs for the regulation of mitochondrial cell death Murray, Michael; Dyari, Herryawan Ryadi Eziwar; Allison, Sarah E; Rawling, Tristan The mitochondrion plays an important role in the production of energy as ATP, the regulation of cell viability and apoptosis, and the biosynthesis of major structural and regulatory molecules, such as lipids. During ATP production, reactive oxygen species are generated that alter the intracellular redox state and activate apoptosis. Mitochondrial dysfunction is a well-recognized component of the pathogenesis of diseases such as cancer. Understanding mitochondrial function, and how this is dysregulated in disease, offers the opportunity for the development of drug molecules to specifically target such defects. Altered energy metabolism in cancer, in which ATP production occurs largely by glycolysis, rather than by oxidative phosphorylation, is attributable in part to the up-regulation of cell survival signalling cascades. These pathways also regulate the balance between pro- and anti-apoptotic factors that may determine the rate of cell death and proliferation. A number of anti-cancer drugs have been developed that target these factors and one of the most promising groups of agents in this regard are the lipid-based molecules that act directly or indirectly at the mitochondrion. These molecules have emerged in part from an understanding of the mitochondrial actions of naturally occurring fatty acids. Some of these agents have already entered clinical trials because they specifically target known mitochondrial defects in the cancer cell.

Effects of 8 weeks of CPAP on lipid-based oxidative markers in obstructive sleep apnea: a randomized trial

Mon, 22 Dec 2014 00:00:00 GMT

Effects of 8 weeks of CPAP on lipid-based oxidative markers in obstructive sleep apnea: a randomized trial Sivam, S.; Witting, P. K.; Hoyos, C. M.; Yee, B. J.; Grunstein, R. R; Phillips, C. L Dyslipidaemia and increased oxidative stress have been reported in severe obstructive sleep apnea, and both may be related to the development of cardiovascular disease. We have previously shown in a randomized crossover study in patients with moderate to severe obstructive sleep apnea that therapeutic continuous positive airway pressure treatment for 8 weeks improved postprandial triglycerides and total cholesterol when compared with sham continuous positive airway pressure. From this study we have now compared the effect of 8 weeks of therapeutic continuous positive airway pressure and sham continuous positive airway pressure on oxidative lipid damage and plasma lipophilic antioxidant levels. Unesterified cholesterol, esterified unsaturated fatty acids (cholesteryl linoleate: C18:2; and cholesteryl arachidonate: C20:4; the major unsaturated and oxidizable lipids in low-density lipoproteins), their corresponding oxidized products [cholesteryl ester-derived lipid hydroperoxides and hydroxides (CE-O(O)H)] and antioxidant vitamin E were assessed at 20:30 hours before sleep, and at 06:00 and 08:30 hours after sleep. Amongst the 29 patients completing the study, three had incomplete or missing [CE-O(O)H] data. The mean apnea -hypopnoea index, age and body mass index were 38 per hour, 49 years and 32 kg m(-2) , respectively. No differences in lipid-based oxidative markers or lipophilic antioxidant levels were observed between the continuous positive airway pressure and sham continuous positive airway pressure arms at any of the three time-points [unesterified cholesterol 0.01 mm, P > 0.05; cholesteryl linoleate: C18:2 0.05 mm, P > 0.05; cholesteryl arachidonate: C20:4 0.02 mm, P = 0.05; CE-O(O)H 2.5 nm, P > 0.05; and lipid-soluble antioxidant vitamin E 0.03 μm, P > 0.05]. In this study, accumulating CE-O(O)H, a marker of lipid oxidation, does not appear to play a role in oxidative stress in obstructive sleep apnea.

Occupy tissue: the movement in cancer metastasis

Sat, 01 Sep 2012 00:00:00 GMT

Occupy tissue: the movement in cancer metastasis Bradbury, Peta; Fabry, Ben; O’Neill, Geraldine M. The critical role of migration and invasion in cancer metastasis warrants new therapeutic approaches targeting the machinery regulating cell migration and invasion. While 2-dimensional (2D) models have helped identify a range of adhesion molecules, cytoskeletal components and regulators that are potentially important for cell migration, the use of models that better mimic the 3-dimensional (3D) environment has yielded new insights into the physiology of cell movement. For example, studying cells in 3D models has revealed that invading cancer cells may switch between heterogeneous invasion modes and thus evade pharmacological inhibition of invasion. Here we summarize published data in which the role of cell adhesion molecules in 2D vs. 3D migration have been directly compared and discuss mechanisms that regulate migration speed and persistence in 2D and 3D. Finally we discuss limits of 3D culture models to recapitulate the in vivo situation.

Src Kinase Determines the Dynamic Exchange of the Docking Protein NEDD9 (Neural Precursor Cell Expressed Developmentally Down-regulated Gene 9) at Focal Adhesions

Fri, 05 Sep 2014 00:00:00 GMT

Src Kinase Determines the Dynamic Exchange of the Docking Protein NEDD9 (Neural Precursor Cell Expressed Developmentally Down-regulated Gene 9) at Focal Adhesions Bradbury, Peta; Bach, Cuc T.; Paul, Andre; O’Neil, Geraldine M. Dynamic exchange of molecules between the cytoplasm and integrin-based focal adhesions provides a rapid response system for modulating cell adhesion. Increased residency time of molecules that regulate adhesion turnover contributes to adhesion stability, ultimately determining migration speed across two-dimensional surfaces. In the present study we test the role of Src kinase in regulating dynamic exchange of the focal adhesion protein NEDD9/HEF1/Cas-L. Using either chemical inhibition or fibroblasts genetically null for Src together with fluorescence recovery after photobleaching (FRAP), we find that Src significantly reduces NEDD9 exchange at focal adhesions. Analysis of NEDD9 mutant constructs with the two major Src-interacting domains disabled revealed the greatest effects were due to the NEDD9 SH2 binding domain. This correlated with a significant change in two-dimensional migratory speed. Given the emerging role of NEDD9 as a regulator of focal adhesion stability, the time of NEDD9 association at the focal adhesions is key in modulating rates of migration and invasion. Our study suggests that Src kinase activity determines NEDD9 exchange at focal adhesions and may similarly modulate other focal adhesion-targeted Src substrates to regulate cell migration.

PP2A phosphatase suppresses function of the mesenchymal invasion regulator NEDD9

Wed, 01 Feb 2012 00:00:00 GMT

PP2A phosphatase suppresses function of the mesenchymal invasion regulator NEDD9 Bradbury, Peta; Mahmassani, Maha; Zhong, Jessie; Turner, Kylie; Paul, Andre; Verrills, Nicole M.; O'Neill, Geraldine M. The mesenchymal mode of cancer cell invasion characterized by active adhesion turnover and a polarized actin cytoskeleton, is critically regulated by the adaptor protein NEDD9/HEF1/Cas-L. While it is known that NEDD9 is subject to extensive phosphorylation modification, the molecules that determine NEDD9 phosphorylation to stimulate adhesion turnover and mesenchymal cell morphologies are currently unknown. Earlier studies have suggested that the serine/threonine phosphatase PP2A regulates interconversion between a low molecular mass NEDD9 phosphoform and higher molecular mass phosphoforms. However, previous studies have used chemical inhibitors to block PP2A activity. In the present study we therefore aimed to specifically inhibit PP2A activity via siRNA and dominant negative approaches to investigate the effect of PP2A on interconversion between 115 kDa and 105 kDa NEDD9 and determine the functional consequence of PP2A activity for NEDD9 function. Strikingly, we find that while the phosphatase inhibitor Calyculin A indeed abrogates detachment-induced dephosphorylation of the 115 kDa NEDD9 phosphoform, PP2A depletion does not inhibit 115 kDa to 105 kDa interconversion. Our data suggest instead that PP2A targets discrete NEDD9 phosphorylation modifications separate to the events that mediate interconversion between the two forms. Functionally, PP2A depletion increases NEDD9 mediated cell spreading and mutation of S369 in the serine-rich region of NEDD9 to aspartate mimics this effect. Importantly, mutation of S369 to alanine abrogates the ability of dominant negative PP2A to increase NEDD9-mediated cell spreading. Collectively, our data reveal that the tumour suppressor PP2A may act via S369 to regulated NEDD9-mediated cell spreading.

2-methyl-4-chlorophenoxyacetic acid and bromoxynil herbicide death.

Wed, 15 Apr 2015 00:00:00 GMT

2-methyl-4-chlorophenoxyacetic acid and bromoxynil herbicide death. Berling, I; Buckley, Nicholas A.; Downes, M.; Isbister, GK. CASE REPORT: We report a fatal case of a 37 year old gentleman who ingested a MCPA/bromoxynil co-formulation herbicide. Although clinically well on initial examination, our patient declined dramatically over his 18 h admission with increasing CO2 production, hyperthermia and metabolic derangement to eventually die from cardiac asystole 20 h post ingestion. Two hours after ingestion the MCPA concentration was 83.9 μg/mL and bromoxynil concentration was 137 μg/mL. DISCUSSION: The patients' mechanism of death appeared to be uncoupling of oxidative phosphorylation, excess CO2 production and hyperthermia. There is limited knowledge on the acute toxicity of these herbicides, in particular bromoxynil, and this case highlights the relentless progression of severe toxicity in humans.

A decade of Australian methotrexate dosing errors

Mon, 06 Jun 2016 00:00:00 GMT

A decade of Australian methotrexate dosing errors Cairns, R; Brown, J; Robinson, J; Lynch, AM; Wylie, C; Buckley, Nicholas A. "OBJECTIVE: Accidental daily dosing of methotrexate can result in life-threatening toxicity. We investigated methotrexate dosing errors reported to the National Coronial Information System (NCIS), the Therapeutic Goods Administration Database of Adverse Event Notifications (TGA DAEN) and Australian Poisons Information Centres (PICs). DESIGN AND SETTING: A retrospective review of coronial cases in the NCIS (2000-2014), and of reports to the TGA DAEN (2004-2014) and Australian PICs (2004-2015). Cases were included if dosing errors were accidental, with evidence of daily dosing on at least 3 consecutive days. MAIN OUTCOME MEASURES: Events per year, dose, consecutive days of methotrexate administration, reasons for the error, clinical features. RESULTS: Twenty-two deaths linked with methotrexate were identified in the NCIS, including seven cases in which erroneous daily dosing was documented. Methotrexate medication error was listed in ten cases in the DAEN, including two deaths. Australian PIC databases contained 92 cases, with a worrying increase seen during 2014-2015. Reasons for the errors included patient misunderstanding and incorrect packaging of dosette packs by pharmacists. The recorded clinical effects of daily dosage were consistent with those previously reported for methotrexate toxicity. CONCLUSION: Dosing errors with methotrexate can be lethal and continue to occur despite a number of safety initiatives in the past decade. Further strategies to reduce these preventable harms need to be implemented and evaluated. Recent suggestions include further changes in packet size, mandatory weekly dosing labelling on packaging, improving education, and including alerts in prescribing and dispensing software."

Tropomyosin Tm5NM1 spatially restricts src kinase activity through perturbation of Rab11 vesicle trafficking

Mon, 01 Dec 2014 00:00:00 GMT

Tropomyosin Tm5NM1 spatially restricts src kinase activity through perturbation of Rab11 vesicle trafficking Bach, Cuc T.; Murray, Rachael Z.; Owen, Dylan; Gaus, Kat; O'Neill, Geraldine M. In order for cells to stop moving, they must synchronously stabilize actin filaments and their associated focal adhesions. How these two structures are coordinated in time and space is not known. We show here that the actin association protein Tm5NM1, which induces stable actin filaments, concurrently suppresses the trafficking of focal-adhesion-regulatory molecules. Using combinations of fluorescent biosensors and fluorescence recovery after photobleaching (FRAP), we demonstrate that Tm5NM1 reduces the level of delivery of Src kinase to focal adhesions, resulting in reduced phosphorylation of adhesion-resident Src substrates. Live imaging of Rab11-positive recycling endosomes that carry Src to focal adhesions reveals disruption of this pathway. We propose that tropomyosin synchronizes adhesion dynamics with the cytoskeleton by regulating actin-dependent trafficking of essential focal-adhesion molecules.

Osteoblasts mediate the adverse effects of glucocorticoids on fuel metabolism

Wed, 24 Oct 2012 00:00:00 GMT

Osteoblasts mediate the adverse effects of glucocorticoids on fuel metabolism Brennan-Speranza, Tara C.; Henneicke, Holger; Gasparini, Sylvia J.; Blankenstein, Katharina I.; Heinevetter, Uta; Cogger, Victoria C.; Svistounov, Dmitri; Zhang, Yaqing; Cooney, Gregory J.; Buttgereit, Frank; Dunstanl, Colin R.; Gundberg, Caren; Zhou, Hong; Seibel, Markus J. Long-term glucocorticoid treatment is associated with numerous adverse outcomes, including weight gain, insulin resistance, and diabetes; however, the pathogenesis of these side effects remains obscure. Glucocorticoids also suppress osteoblast function, including osteocalcin synthesis. Osteocalcin is an osteoblast-specific peptide that is reported to be involved in normal murine fuel metabolism. We now demonstrate that osteoblasts play a pivotal role in the pathogenesis of glucocorticoid-induced dysmetabolism. Osteoblast-targeted disruption of glucocorticoid signaling significantly attenuated the suppression of osteocalcin synthesis and prevented the development of insulin resistance, glucose intolerance, and abnormal weight gain in corticosterone-treated mice. Nearly identical effects were observed in glucocorticoid-treated animals following heterotopic (hepatic) expression of both carboxylated and uncarboxylated osteocalcin through gene therapy, which additionally led to a reduction in hepatic lipid deposition and improved phosphorylation of the insulin receptor. These data suggest that the effects of exogenous high-dose glucocorticoids on insulin target tissues and systemic energy metabolism are mediated, at least in part, through the skeleton.

NEDD9 Stabilizes Focal Adhesions, Increases Binding to the Extra-Cellular Matrix and Differentially Effects 2D versus 3D Cell Migration

Wed, 11 Apr 2012 00:00:00 GMT

NEDD9 Stabilizes Focal Adhesions, Increases Binding to the Extra-Cellular Matrix and Differentially Effects 2D versus 3D Cell Migration Zhong, J; Baquiran, JB; Bonakdar, N; Lees, Justin G.; Ching, YW; Pugacheva, E; Fabry, B; O'Neill, GM The speed of cell migration on 2-dimensional (2D) surfaces is determined by the rate of assembly and disassembly of clustered integrin receptors known as focal adhesions. Different modes of cell migration that have been described in 3D environments are distinguished by their dependence on integrin-mediated interactions with the extra-cellular matrix. In particular, the mesenchymal invasion mode is the most dependent on focal adhesion dynamics. The focal adhesion protein NEDD9 is a key signalling intermediary in mesenchymal cell migration, however whether NEDD9 plays a role in regulating focal adhesion dynamics has not previously been reported. As NEDD9 effects on 2D migration speed appear to depend on the cell type examined, in the present study we have used mouse embryo fibroblasts (MEFs) from mice in which the NEDD9 gene has been depleted (NEDD9 −/− MEFs). This allows comparison with effects of other focal adhesion proteins that have previously been demonstrated using MEFs. We show that focal adhesion disassembly rates are increased in the absence of NEDD9 expression and this is correlated with increased paxillin phosphorylation at focal adhesions. NEDD9−/− MEFs have increased rates of migration on 2D surfaces, but conversely, migration of these cells is significantly reduced in 3D collagen gels. Importantly we show that myosin light chain kinase is activated in 3D in the absence of NEDD9 and is conversely inhibited in 2D cultures. Measurement of adhesion strength reveals that NEDD9−/− MEFs have decreased adhesion to fibronectin, despite upregulated α5β1 fibronectin receptor expression. We find that β1 integrin activation is significantly suppressed in the NEDD9−/−, suggesting that in the absence of NEDD9 there is decreased integrin receptor activation. Collectively our data suggest that NEDD9 may promote 3D cell migration by slowing focal adhesion disassembly, promoting integrin receptor activation and increasing adhesion force to the ECM.

Zirconium Ions Up-Regulate the BMP/SMAD Signaling Pathway and Promote the Proliferation and Differentiation of Human Osteoblasts

Tue, 20 Jan 2015 00:00:00 GMT

Zirconium Ions Up-Regulate the BMP/SMAD Signaling Pathway and Promote the Proliferation and Differentiation of Human Osteoblasts Chen, Yongjuan; Roohani-Esfahani, Seyed-Iman; Lu, ZuFu; Zreiqat, Hala; Dunstan, Colin R. Zirconium (Zr) is an element commonly used in dental and orthopedic implants either as zirconia (ZrO2) or in metal alloys. It can also be incorporated into calcium silicate-based ceramics. However, the effects of in vitro culture of human osteoblasts (HOBs) with soluble ionic forms of Zr have not been determined. In this study, primary culture of human osteoblasts was conducted in the presence of medium containing either ZrCl4 or Zirconium (IV) oxynitrate (ZrO(NO3)2) at concentrations of 0, 5, 50 and 500 μM, and osteoblast proliferation, differentiation and calcium deposition were assessed. Incubation of human osteoblast cultures with Zr ions increased the proliferation of human osteoblasts and also gene expression of genetic markers of osteoblast differentiation. In 21 and 28 day cultures, Zr ions at concentrations of 50 and 500 μM increased the deposition of calcium phosphate. In addition, the gene expression of BMP2 and BMP receptors was increased in response to culture with Zr ions and this was associated with increased phosphorylation of SMAD1/5. Moreover, Noggin suppressed osteogenic gene expression in HOBs co-treated with Zr ions. In conclusion, Zr ions appear able to induce both the proliferation and the differentiation of primary human osteoblasts. This is associated with up-regulation of BMP2 expression and activation of BMP signaling suggesting this action is, at least in part, mediated by BMP signaling.

Tyrosine Y189 in the Substrate Domain of the Adhesion Docking Protein NEDD9 Is Conserved with p130Cas Y253 and Regulates NEDD9-Mediated Migration and Focal Adhesion Dynamics

Tue, 09 Jul 2013 00:00:00 GMT

Tyrosine Y189 in the Substrate Domain of the Adhesion Docking Protein NEDD9 Is Conserved with p130Cas Y253 and Regulates NEDD9-Mediated Migration and Focal Adhesion Dynamics Baquiran, Jaime B.; Bradbury, Peta; O’Neill, Geraldine M. The focal adhesion docking protein NEDD9/HEF1/Cas-L regulates cell migration and cancer invasion. NEDD9 is a member of the Cas family of proteins that share conserved overall protein-protein interaction domain structure, including a substrate domain that is characterized by extensive tyrosine (Y) phosphorylation. Previous studies have suggested that phosphorylation of Y253 in the substrate domain of the Cas family protein p130Cas is specifically required for p130Cas function in cell migration. While it is clear that tyrosine phosphorylation of the NEDD9 substrate domain is similarly required for the regulation of cell motility, whether individual NEDD9 tyrosine residues have discrete function in regulating motility has not previously been reported. In the present study we have used a global sequence alignment of Cas family proteins to identify a putative NEDD9 equivalent of p130Cas Y253. We find that NEDD9 Y189 aligns with p130Cas Y253 and that it is conserved among NEDD9 vertebrate orthologues. Expression of NEDD9 in which Y189 is mutated to phenylalanine results in increased rates of cell migration and is correlated with increased disassembly of GFP.NEDD9 focal adhesions. Conversely, mutation to Y189D significantly inhibits cell migration. Our previous data has suggested that NEDD9 stabilizes focal adhesions and the present data therefore suggests that phosphorylation of Y189 NEDD9 is required for this function. These findings indicate that the individual tyrosine residues of the NEDD9 substrate domain may serve discrete functional roles. Given the important role of this protein in promoting cancer invasion, greater understanding of the function of the individual tyrosine residues is important for the future design of approaches to target NEDD9 to arrest cancer cell invasion.

μ -Opioid receptor activation and noradrenaline transport inhibition by tapentadol in rat single locus coeruleus neurons.

Thu, 01 Jan 2015 00:00:00 GMT

μ -Opioid receptor activation and noradrenaline transport inhibition by tapentadol in rat single locus coeruleus neurons. Sadeghi, Mahsa; Tzschentke, Thomas M; Christie, MacDonald J BACKGROUND AND PURPOSE: Tapentadol is a novel analgesic that combines moderate μ-opioid receptor agonism and noradrenaline reuptake inhibition in a single molecule. Both mechanisms of action are involved in producing analgesia; however, the potency and efficacy of tapentadol in individual neurons has not been characterized. EXPERIMENTAL APPROACH: Whole-cell patch-clamp recordings of G-protein-coupled inwardly rectifying K(+) (KIR 3.x) currents were made from rat locus coeruleus neurons in brain slices to investigate the potency and relative efficacy of tapentadol and compare its intrinsic activity with other clinically used opioids. KEY RESULTS: Tapentadol showed agonist activity at μ receptors and was approximately six times less potent than morphine with respect to KIR 3.x current modulation. The intrinsic activity of tapentadol was lower than [Met]enkephalin, morphine and oxycodone, but higher than buprenorphine and pentazocine. Tapentadol inhibited the noradrenaline transporter (NAT) with potency similar to that at μ receptors. The interaction between these two mechanisms of action was additive in individual LC neurons. CONCLUSIONS AND IMPLICATIONS: Tapentadol displays similar potency for both µ receptor activation and NAT inhibition in functioning neurons. The intrinsic activity of tapentadol at the μ receptor lies between that of buprenorphine and oxycodone, potentially explaining the favourable profile of side effects, related to μ receptors. LINKED ARTICLES: This article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2.

High-voltage activated calcium current subtypes in mouse DRG neurons adapt in a subpopulation-specific manner after nerve injury.

Sun, 01 Mar 2015 00:00:00 GMT

High-voltage activated calcium current subtypes in mouse DRG neurons adapt in a subpopulation-specific manner after nerve injury. Murali, S.; Napier, I.; Mohammadi, S.; Alewood, P.; Lewis, R.; Christie, M. Changes in ion channel function and expression are characteristic of neuropathic pain. Voltage-gated calcium channels (VGCCs) are integral for neurotransmission and membrane excitability, but relatively little is known about changes in their expression after nerve injury. In this study, we investigate whether peripheral nerve ligation is followed by changes in the density and proportion of high-voltage-activated (HVA) VGCC current subtypes in dorsal root ganglion (DRG) neurons, the contribution of presynaptic N-type calcium channels in evoked excitatory postsynaptic currents (EPSCs) recorded from dorsal horn neurons in the spinal cord, and the changes in expression of mRNA encoding VGCC subunits in DRG neurons. Using C57BL/6 mice [8- to 11-wk-old males (n = 91)] for partial sciatic nerve ligation or sham surgery, we performed whole cell patch-clamp recordings on isolated DRG neurons and dorsal horn neurons and measured the expression of all VGCC subunits with RT-PCR in DRG neurons. After nerve injury, the density of P/Q-type current was reduced overall in DRG neurons. There was an increase in the percentage of N-type and a decrease in that of P/Q-type current in medium- to large-diameter neurons. No changes were found in the contribution of presynaptic N-type calcium channels in evoked EPSCs recorded from dorsal horn neurons. The α2δ-1 subunit was upregulated by 1.7-fold and γ-3, γ-2, and β-4 subunits were all downregulated 1.7-fold in injured neurons compared with sham-operated neurons. This comprehensive characterization of HVA VGCC subtypes in mouse DRG neurons after nerve injury revealed changes in N- and P/Q-type current proportions only in medium- to large-diameter neurons.

A Continuous, Fluorescence-based Assay of µ-Opioid Receptor Activation in AtT-20 Cells

Fri, 01 Mar 2013 00:00:00 GMT

A Continuous, Fluorescence-based Assay of µ-Opioid Receptor Activation in AtT-20 Cells Knapman, Alisa; Santiago, Marina; Du, Yan Ping; Bennallack, Philip R.; Christie, Macdonald J.; Connor, Mark Opioids are widely prescribed analgesics, but their use is limited due to development of tolerance and addiction, as well as high variability in individual response. The development of improved opioid analgesics requires high-throughput functional assays to assess large numbers of potential opioid ligands. In this study, we assessed the ability of a proprietary "no-wash" fluorescent membrane potential dye to act as a reporter of µ-opioid receptor (MOR) activation and desensitization via activation of G-protein-coupled inwardly rectifying potassium channels. AtT-20 cells stably expressing mouse MOR were assayed in 96-well plates using the Molecular Devices FLIPR membrane potential dye. Dye emission intensity decreased upon membrane hyperpolarization. Fluorescence decreased in a concentration-dependent manner upon application of a range of opioid ligands to the cells, with high-efficacy agonists producing a decrease of 35% to 40% in total fluorescence. The maximum effect of morphine faded in the continued presence of agonist, reflecting receptor desensitization. The effects of opioids were prevented by prior treatment with pertussis toxin and blocked by naloxone. We have demonstrated this assay to be an effective method for assessing ligand signaling at MOR, which may potentially be scaled up as an additional high-throughput screening technique for characterizing novel opioid ligands.

Nucleus accumbens D2- and D1-receptor expressing medium spiny neurons are selectively activated by morphine withdrawal and acute morphine, respectively.

Fri, 01 Jun 2012 00:00:00 GMT

Nucleus accumbens D2- and D1-receptor expressing medium spiny neurons are selectively activated by morphine withdrawal and acute morphine, respectively. Enoksson, T.; Bertran-Gonzalez, J.; Christie, M.J. Opioids are effective analgesic agents but serious adverse effects such as tolerance and withdrawal contribute to opioid dependence and limit their use. Opioid withdrawal involves numerous brain regions and includes suppression of dopamine release and activation of neurons in the ventral striatum. By contrast, acute opioids increase dopamine release. Like withdrawal, acute opioids also activate neurons in the ventral striatum, suggesting that different populations of ventral striatal neurons may be activated by withdrawal and acute opioid actions. Here, immunofluorescence for the activity-related immediate-early gene, c-Fos, was examined in transgenic reporter mouse lines by confocal microscopy to study the specific populations of ventral striatal neurons activated by morphine withdrawal and acute morphine. After chronic morphine, naloxone-precipitated withdrawal strongly increased expression of c-Fos immunoreactivity, predominantly in D2-receptor (D2R) medium-sized spiny neurons (MSNs) of the nucleus accumbens (NAc) core and shell regions. By contrast, a single injection of morphine exclusively activated c-Fos immunoreactivity in D1-receptor expressing (D1R) MSNs of the core and shell of the NAc. These results reveal a striking segregation of neuronal responses occurring in the two populations of MSNs of the NAc in response to morphine withdrawal and acute morphine.

Glutamate transporter dysfunction associated with nerve injury-induced pain in mice

Sun, 01 Jan 2012 00:00:00 GMT

Glutamate transporter dysfunction associated with nerve injury-induced pain in mice Napier, Ian A.; Mohammadi, Sarasa A.; Christie, MacDonald J. Dysfunction at glutamatergic synapses has been proposed as a mechanism in the development of neuropathic pain. Here we sought to determine whether peripheral nerve injury-induced neuropathic pain results in functional changes to primary afferent synapses. Signs of neuropathic pain as well as an induction of glial fibrillary acidic protein in immunostained spinal cord sections 4 days after partial ligation of the sciatic nerve indicated the induction of neuropathic pain. We found that following nerve injury, no discernable change to kinetics of dl-α-amino-3-hydroxy-5-methylisoxazole-propionic acid (AMPA) or N-methyl-d-aspartate receptor (NMDAR)-mediated evoked excitatory postsynaptic currents (eEPSCs) could be observed in dorsal horn (lamina I/II) neurons compared with those of naïve mice. However, we did find that nerve injury was accompanied by slowed decay of the early phase of eEPSCs in the presence of glutamate transporter inhibition by the competitive nontransportable inhibitor dl-threo-β-benzyloxyaspartic acid (TBOA). Concomitantly, expression patterns for the two major glutamate transporters in the spinal cord, excitatory amino acid transporters (EAAT) 1 and EAAT2, were found to be reduced at this time (4 days postinjury). We then sought to directly determine whether nerve injury results in glutamate spillover to NMDARs at dorsal horn synapses. By employing the use-dependent NMDAR blocker (±)MK-801 to block subsynaptic receptors, we found that although TBOA-induced spillover to extrasynaptic receptors trended to increased activation of these receptors after nerve injury, this was not significant compared with naïve mice. Together, these results suggest the development of neuropathic pain involves subtle changes to glutamate transporter expression and function that could contribute to neuropathic pain during excessive synaptic activity.

A novel mechanism of inhibition of high-voltage activated calcium channels by a-conotoxins contributes to relief of nerve injury-induced neuropathic pain

Sun, 01 Mar 2015 00:00:00 GMT

A novel mechanism of inhibition of high-voltage activated calcium channels by a-conotoxins contributes to relief of nerve injury-induced neuropathic pain Klimis, Harry; Adams, D. J; Callaghan, B; Nevin, S; Alewood, P. F; Vaughan, C. W.; Mozar, C. A; Christie, M. J. α-Conotoxins that are thought to act as antagonists of nicotinic acetylcholine receptors (nAChRs) containing α3-subunits are efficacious in several preclinical models of chronic pain. Potent interactions of Vc1.1 with other targets have suggested that the pain-relieving actions of α-conotoxins might be mediated by either α9α10 nAChRs or a novel GABA(B) receptor-mediated inhibition of N-type calcium channels. Here we establish that three α-conotoxins, Vc1.1, AuIB and MII have distinct selectivity profiles for these three potential targets. Their potencies after intramuscular administration were then determined for reversal of allodynia produced by partial nerve ligation in rats. Vc1.1, which potently inhibits α9α10 nAChRs and GABA(B)/Ca(2+) channels but weakly blocks α3β2 and α3β4 nAChRs, produced potent, long-lasting reversal of allodynia that were prevented by pre-treatment with the GABA(B) receptor antagonist, SCH50911. α-Conotoxin AuIB, a weak α3β4 nAChR antagonist, inhibited GABA(B)/Ca(2+) channels but did not act on α9α10 nAChRs. AuIB also produced reversal of allodynia. These findings suggest that GABA(B) receptor-dependent inhibition of N-type Ca(2+) channels can mediate the sustained anti-allodynic actions of some α-conotoxins. However, MII, a potent α3β2 nAChR antagonist but inactive on α9α10 and α3β4 nAChRs and GABA(B)/Ca(2+) channels, was demonstrated to have short-acting anti-allodynic action. This suggests that α3β2 nAChRs may also contribute to reversal of allodynia. Together, these findings suggest that inhibition of α9α10 nAChR is neither necessary nor sufficient for relief of allodynia and establish that α-conotoxins selective for GABA(B) receptor-dependent inhibition of N-type Ca(2+) channels relieve allodynia, and could therefore be developed to manage chronic pain.