TTLL12 is required for primary ciliary axoneme formation in polarized epithelial cells.

The primary cilium is a critical sensory organelle that is built of axonemal microtubules ensheathed by a ciliary membrane. In polarized epithelial cells, primary cilia reside on the apical surface and must extend these microtubules directly into the extracellular space and remain a stable structure. However, the factors regulating cross-talk between ciliation and cell polarization, as well as, axonemal microtubule growth and stabilization in polarized epithelia are not fully understood. In this study, we find TTLL12, a previously uncharacterized member of the Tubulin Tyrosine Ligase-Like (TTLL) family, localizes to the base of primary cilia and is required for cilia formation in polarized renal epithelial cells. We also show that TTLL12 directly binds to the α/ß-tubulin heterodimer in vitro and regulates microtubule dynamics, stability, and post-translational modifications (PTMs). While all other TTLLs catalyze the addition of glutamate or glycine to microtubule C-terminal tails, TTLL12 uniquely affects tubulin PTMs by promoting both microtubule lysine acetylation and arginine methylation. Together, this work identifies a novel microtubule regulator and provides insight into the requirements for apical extracellular axoneme formation.

Child and adolescent psychiatry training in Australia and New Zealand.

The specialty of Child and Adolescent Psychiatry was formally recognised in the 1930s. The Faculty of Child and Adolescent Psychiatry was established in 1964 in Australia, as a subspecialty in The Royal Australian and New Zealand College of Psychiatrists (RANZCP). The aim of the current article is first to provide a brief summary and overview of the current status of Child and Adolescent Psychiatry (CAP), followed by an outline of the requirements of the Training Program for CAP in Australia and New Zealand. The training required to become a fully qualified child and adolescent psychiatrist in Australia and New Zealand consists of different stages and takes the form of competency-based training. Information relating to assessment types, supervision and research requirements is also described. Accreditation procedures for the training program are stipulated by RANZCP to monitor standards and to ensure consistency within the programs delivered across Australia and New Zealand. Employment opportunities for trainees upon completion of the program are discussed. In summary, this article highlights the requirements of the training programs for CAP in Australia and New Zealand.

Patients with the worst outcomes after paracetamol (acetaminophen)-induced liver failure have an early monocytopenia.

BACKGROUND: Acute liver failure (ALF) is associated with significant morbidity and mortality. Studies have implicated the immune response, especially monocyte/macrophages as being important in dictating outcome. AIM: To investigate changes in the circulating monocytes and other immune cells serially in patients with ALF, relate these with cytokine concentrations, monocyte gene expression and patient outcome. METHODS: In a prospective case-control study in the Scottish Liver Transplant Unit, Royal Infirmary Edinburgh, 35 consecutive patients admitted with paracetamol-induced liver failure (POD-ALF), 10 patients with non-paracetamol causes of ALF and 16 controls were recruited. The peripheral blood monocyte phenotype was analysed by flow cytometry, circulating cytokines quantified by protein array and monocyte gene expression array performed and related to outcome. RESULTS: On admission, patients with worst outcomes after POD-ALF had a significant monocytopenia, characterised by reduced classical and expanded intermediate monocyte population. This was associated with reduced circulating lymphocytes and natural killer cells, peripheral cytokine patterns suggestive of a 'cytokine storm' and increased concentrations of cytokines associated with monocyte egress from the bone marrow. Gene expression array did not differentiate patient outcome. At day 4, there was no significant difference in monocyte, lymphocyte or natural killer cells between survivors and the patients with adverse outcomes. CONCLUSIONS: Severe paracetamol liver failure is associated with profound changes in the peripheral blood compartment, particularly in monocytes, related with worse outcomes. This is not seen in patients with non-paracetamol-induced liver failure. Significant monocytopenia on admission may allow earlier clarification of prognosis, and it highlights a potential target for therapeutic intervention.

Comprehensive microRNA profiling in acetaminophen toxicity identifies novel circulating biomarkers for human liver and kidney injury.

Our objective was to identify microRNA (miRNA) biomarkers of drug-induced liver and kidney injury by profiling the circulating miRNome in patients with acetaminophen overdose. Plasma miRNAs were quantified in age- and sex-matched overdose patients with (N = 27) and without (N = 27) organ injury (APAP-TOX and APAP-no TOX, respectively). Classifier miRNAs were tested in a separate cohort (N = 81). miRNA specificity was determined in non-acetaminophen liver injury and murine models. Sensitivity was tested by stratification of patients at hospital presentation (N = 67). From 1809 miRNAs, 75 were 3-fold or more increased and 46 were 3-fold or more decreased with APAP-TOX. A 16 miRNA classifier model accurately diagnosed APAP-TOX in the test cohort. In humans, the miRNAs with the largest increase (miR-122-5p, miR-885-5p, miR-151a-3p) and the highest rank in the classifier model (miR-382-5p) accurately reported non-acetaminophen liver injury and were unaffected by kidney injury. miR-122-5p was more sensitive than ALT for reporting liver injury at hospital presentation, especially combined with miR-483-3p. A miRNA panel was associated with human kidney dysfunction. In mice, miR-122-5p, miR-151a-3p and miR-382-5p specifically reported APAP toxicity - being unaffected by drug-induced kidney injury. Profiling of acetaminophen toxicity identified multiple miRNAs that report acute liver injury and potential biomarkers of drug-induced kidney injury.

Systematic review: the effects of autologous stem cell therapy for patients with liver disease.

BACKGROUND: As morbidity and mortality from liver disease continues to rise, new strategies are necessary. Liver transplantation is not only an expensive resource committing the patient to lifelong immunosuppression but also suitable donor organs are in short supply. Against this background, autologous stem cell therapy has emerged as a potential treatment option. AIM: To evaluate if it is possible to make a judgement on the safety, feasibility and effect of autologous stem cell therapy for patients with liver disease. METHODS: MEDLINE and EMBASE were searched up until July 2013 to identify studies where autologous stem cell therapy was administered to patients with liver disease. RESULTS: Of 1668 studies identified, 33 were eligible for inclusion evaluating a median sample size of 10 patients for a median follow-up of 6 months. Although there was marked heterogeneity between studies with regards to type, dose and route of delivery of stem cell, the treatment was shown to be safe and feasible largely when a peripheral route of administration was used. Of the studies which also looked at biochemical outcome, statistically significant improvement in liver function tests was seen in 16 studies post-treatment. CONCLUSION: Although autologous stem cell therapy is a much needed possibility in the treatment of liver disease, further robust clinical trials and collaborative protocols are required.

Persistently elevated troponin I in paracetamol hepatotoxicity: association with liver injury, organ failure, and outcome.

CONTEXT: An elevated troponin I (TnI) is associated with a poorer prognosis during critical illness. OBJECTIVE: Our aims were to determine whether significant paracetamol-induced hepatotoxicity was associated with an elevated TnI; if this elevation was persistent and was associated with worse clinical outcomes. MATERIALS AND METHODS: In this retrospective cohort study, the requirement for orthotopic liver transplantation (OLT) or death and/or the development of multiorgan failure (MOF) was evaluated for 48 consecutive patients admitted to the Royal Infirmary of Edinburgh (a university tertiary referral centre) with acute liver injury or acute liver failure secondary to paracetamol overdose. RESULTS: TnI was elevated (≥ 0.05 ng/L) in 13/48 patients (27%). This appeared to be sustained for at least 6 days which has not been shown previously in the context of Acute Liver Injury (ALI). Elevated TnI was strongly associated with MOF, with the requirement for inotropic support being the strongest predictor (p = 0.003, OR 9.00, 95% CI 2.13-37.98). TnI elevations also correlated strongly with Acute Physiology and Chronic Health Evaluation (APACHE) II scores (p = 0.0006, r = 0.482, 95% CI 0.22-0.68) and with interleukin 6 (IL-6) levels (p = 0.0001, r = 0.55, 95% CI 0.29-0.73). Although a raised TnI was associated with a markedly increased risk of death or orthotopic liver transplant (p = 0.005, OR 7.73, 95% CI 1.87-32.05) on univariate analysis, this was primarily seen in the context of MOF (SOFA score p = 0.003, OR 1.23, 95% CI 1.07-1.41) and was not an independent predictor of death. There was no correlation between TnI or outcome with other cardiac biomarkers and markers of cardiovascular risk. DISCUSSION AND CONCLUSION: An elevated TnI in the context of acute liver injury or liver failure following paracetamol overdose is associated with a significantly worse patient outcome but it is not an independent prognostic factor. Further studies should be undertaken to investigate the mechanism behind this elevated troponin association.

The effect of anaesthetic agents on induction, recovery and patient preferences in adult day case surgery: a 7-day follow-up randomized controlled trial.

BACKGROUND AND OBJECTIVE: To compare induction, pre- and post-discharge recovery characteristics and patient preferences between four anaesthetic regimens in adult day-surgery. METHODS: Randomized controlled trial. In all, 1158 adults assigned to: propofol induction and maintenance, propofol induction with isoflurane/N2O, or sevoflurane/N2O maintenance, or sevoflurane/N2O alone. We prospectively recorded induction and pre-discharge recovery characteristics, collected 7-day post-discharge recovery characteristics using patient diaries and patient preferences by telephone follow-up. RESULTS: Recruitment rate was 73%--of the 425 refusals, 226 were not willing to risk a volatile induction. During induction, excitatory movements and breath holding were more common with sevoflurane only (P < 0.01). Injection pain and hiccup were more common with propofol induction (P < 0.01). In the recovery room and the postoperative ward, both nausea and vomiting were more common with sevoflurane only (P < 0.01). This difference disappeared within 48 h. There was no difference between groups in the mental state on awakening, recovery time, time to discharge or overnight admissions; then was also no difference in pain between the four groups for each of the seven postoperative days (P < 0.01), nor any differences in concentration or forgetfulness. Patients took 6.5 days (95% CI: 6.0-7.0, n = 693) to resume normal activities. Patients who received sevoflurane only were more likely to recall an unpleasant induction and least likely to want the same induction method again (P < 0.01). CONCLUSION: Differences in outcome between the four regimens are transient; sevoflurane is not an ideal sole agent for adult day case anaesthesia and, in this setting, patients base their preferences for future anaesthetics on the method of induction.

Modulation of antigen presentation by autoreactive B cell clones specific for GAD65 from a type I diabetic patient.

We used a GAD65-specific human B-T cell line cognate system in vitro to investigate the modulation of GAD65 presentation by autoantibody, assessed in a proliferation assay. Generally, if the T cell determinant overlaps or resides within the antibody epitope, effects of presentation are blunted while if they are distant can lead to potent presentation. For three different autoreactive B-T cell line cognate pairs, the modulation of GAD65 presentation followed the mode of overlapping or distant epitopes with resultant potent or undetectable presentation. However, other cognate pairs elicited variability in this pattern of presentation. Notably, one B cell line, DPC, whose antibody epitope did not overlap with the T cell determinants, was consistently poor in presenting GAD65. Using the fluorescent dye Alexa Fluor 647 conjugated to GAD65 to study receptor-mediated antigen endocytosis showed that all the antigen-specific B cell clones were efficient in intracellular accumulation of the antigen. Additionally, multicolour immunofluorescence microscopy showed that the internalized GAD65/surface IgG complexes were rapidly targeted to a perinuclear compartment in all GAD-specific B cell clones. This analysis also demonstrated that HLA-DM expression was reduced strongly in DPC compared to the stimulatory B cell clones. Thus the capability of antigen-specific B cells to capture and present antigen to human T cell lines is dependent on the spatial relationship of B and T cell epitopes as well other factors which contribute to the efficiency of presentation.

An initiative to reform senior house officer training in histopathology.

Senior house officer schools in histopathology have been established in Leeds, Leicester and Southampton, each training six new recruits each year. Nine hours of protected teaching time is provided each week giving a ratio of apprenticeship learning to formal teaching of 3:1. Evaluations have been very positive. Much of this success is attributed to careful planning and adequate funding. This may be a useful model for other specialties to follow.

Clinical and economic choices in anaesthesia for day surgery: a prospective randomised controlled trial.

We compared the cost-effectiveness of general anaesthetic agents in adult and paediatric day surgery populations. We randomly assigned 1063 adult and 322 paediatric elective patients to one of four (adult) or two (paediatric) anaesthesia groups. Total costs were calculated from individual patient resource use to 7 days post discharge. Incremental cost-effectiveness ratios were expressed as cost per episode of postoperative nausea and vomiting (PONV) avoided. In adults, variable secondary care costs were higher for propofol induction and propofol maintenance (propofol/propofol; p < 0.01) than other groups and lower in propofol induction and isoflurane maintenance (propofol/isoflurane; p < 0.01). In both studies, predischarge PONV was higher if sevoflurane/sevoflurane (p < 0.01) was used compared with use of propofol for induction. In both studies, there was no difference in postdischarge outcomes at Day 7. Sevoflurane/sevoflurane was more costly with higher PONV rates in both studies. In adults, the cost per extra episode of PONV avoided was pound 296 (propofol/propofol vs. propofol/ sevoflurane) and pound 333 (propofol/sevoflurane vs. propofol/isoflurane).

Anaesthesia for day case surgery: a survey of paediatric clinical practice in the UK.

BACKGROUND AND OBJECTIVE: In October 2000, we conducted a national postal survey of day case consultant anaesthetists in the UK to explore the range and variation in practice of anaesthetizing a patient for day case surgery (paediatrics, urology and orthopaedics). This paper reports the findings of this national survey of paediatric day case anaesthetic practice carried out as part of a major two-centre randomized controlled trial designed to investigate the costs and outcome of several anaesthetic techniques during day care surgery in paediatric and adult patients (cost-effectiveness study of anaesthesia in day case surgery). METHODS: The survey used a structured postal questionnaire and collected data on the duration of surgical procedure; the use of premedication; the anaesthetic agents used for induction and maintenance; the fresh gas flow rates used for general anaesthesia; the use of antiemetics; and the administration of local anaesthesia and analgesia. RESULTS: The overall response rate for the survey was 74 and 63% for the paediatric section of the survey. Respondents indicated that 19% used premedication, 63% used propofol for induction, 54% used isoflurane for maintenance, 24% used prophylactic antiemetics and 85%, used a laryngeal mask. The findings of this national survey are discussed and compared with published evidence. CONCLUSIONS: This survey identifies the variation in clinical practice in paediatric day surgery anaesthesia in the UK.

Propofol and halothane versus sevoflurane in paediatric day-case surgery: induction and recovery characteristics.

BACKGROUND: The aim of this study was to compare the induction and recovery characteristics associated with propofol induction and halothane maintenance with sevoflurane anaesthesia in paediatric day surgery. METHODS: In total, 322 children were assigned randomly to i.v. propofol induction and halothane/nitrous oxide maintenance or sevoflurane/nitrous oxide alone. The patients' age, sex, and type of surgery were recorded, as were the times required for anaesthetic induction, maintenance, recovery and time to discharge home. Postoperative nausea and vomiting, and the incidence of adverse events during induction and recovery were also noted. RESULTS: No significant differences were detected in age, sex, type of surgery performed or intraoperative opioid administration. Excitatory movement was more common during induction with sevoflurane. The mean time required for induction with propofol was 3.1 min compared with 5 min in the sevoflurane group (P<0.001). The recovery time was shorter in the sevoflurane group compared with propofol/halothane (23.2 vs 26.4 min, P<0.002). The incidence of delirium in recovery was greater in the sevoflurane group (P<0.001). There was no difference between groups in the time spent on the postoperative ward before discharge home. On the postoperative ward the incidence of both nausea and vomiting was significantly higher in the sevoflurane group (P=0.034). Five children were admitted to hospital overnight, none for anaesthetic reasons. CONCLUSIONS: The increased incidence of adverse events during induction, postoperative nausea and vomiting and postoperative delirium in the sevoflurane group suggests that sevoflurane is not ideal as a sole agent for paediatric day case anaesthesia.

Myelination of the human auditory nerve: different time courses for Schwann cell and glial myelin.

The goal of this study was to trace the development of myelin in the human auditory nerve. To do this, we used the Woelcke iron-hematoxylin technique to stain myelin sheaths in sections from fetal temporal bones and brain stems. In the cochlea, aggregations of Schwann cells are seen in the modiolus and along the spiral lamina by the 15th fetal week. By the 22nd fetal week, dense arrays of stained Schwann cells are present on auditory nerve axons within the temporal bone. By the 24th fetal week, light myelin sheaths extend up to, but not beyond, the glial junction. Myelin sheaths are not present in the auditory nerve central to the glial junction until the 26th fetal week or later. These results demonstrate a gap of several weeks between the onset of Schwann cell myelination distally and glial myelination proximally. The period between these two events may represent the time of final maturation of the organ of Corti.

Phenotypic and functional analysis of T-cell recovery after anti-CD3 immunotoxin treatment for tolerance induction in rhesus macaques.

T-cell reduction utilizing specific antibody has been widely used in human transplantation, and is a cornerstone of several tolerance induction strategies in nonhuman primates. We have established a population of long-term tolerant rhesus macaques induced with an anti-CD3epsilon immunotoxin (IT). This treatment effects transient, specific and profound ablation of T cells in blood and lymphoid tissues. In most instances the IT was used in combination with the NF-kappaB inhibitor, 15-Deoxyspergualin. This 2-week long protocol produces a "window of opportunity" for tolerization in which the animal exhibits an enduring quiescent state of unresponsiveness to the allograft, all accomplished without maintenance immunosuppressive drugs. During this induction period, the treated immune system bears some resemblance to that of the neonate, in that T cell numbers are abnormally low and antigen presentation by dendritic cells is precluded by an arrest in their NF-kappaB dependant maturation. In addition, IL-4 production is prominent during and after the tolerance induction interval. For this study we focused on measuring the monkey's ability to repopulate T cells with particular emphasis on the memory T-cell phenotype. Three "memory" phenotypes were utilized; CD3(+)CD45RO(+), CD3(+)CRTH2(+), and CD3(+)CD4(+)CD8(+). All three phenotypes exhibited different patterns of recovery, all of which included transient bursts in their numbers during repopulation. We also estimated thymic activity after T-cell ablation with the use of a newly-described RTE or recent thymic émigré phenotype (a naïve CD8(+)CD103(+) T cell). This marker revealed production of RTE cells including supranormal levels at approximately 6 months post-transplant, implicating thymic function in the repopulation of T-cells. Finally, we measured antibody responses to a panel of antigens (vaccines, environmental antigen, and foreign proteins) that indicated there was no apparent loss of immunologic function during or after the tolerance induction period. Results of studies of T-cell receptor repertoire expression suggest preservation of the pretreatment repertoire, which is consistent with rapid recovery of immune competence to the test antigens. Taken together, these results suggest that while aggressive, this tolerance induction protocol does not appear to incur a prolonged immunologically-compromised state, if at all.

Treatment of conjunctival and corneal intraepithelial neoplasia with topical interferon alpha-2b.

OBJECTIVE: To evaluate the role of topical interferon alfa-2b (IFNalpha2b) in the treatment of conjunctival and corneal intraepithelial neoplasia (CIN). DESIGN: Noncomparative case series. PARTICIPANTS: Five patients with histologically proven CIN or recurrences of proven CIN were studied prospectively. INTERVENTION: After histologic confirmation, patients were given topical recombinant IFNalpha2b (INTRON A, Schering Plough, Kenilworth, NJ) 1 million IU/ml four times a day. Patients were continued on interferon until complete resolution of the tumor had occurred. MAIN OUTCOME MEASURES: Patients were followed clinically and photographically for evidence of tumor resolution. RESULTS: All patients had complete resolution of the CIN lesion on IFNalpha2b. The mean time to clinical resolution was 11.6 weeks (range, 4-22 weeks). The mean follow-up was 17.6 months (range, 7-28 months). One patient had a clinical recurrence of his corneal CIN 1 year after tumor resolution. This patient was retreated, resulting in clinical resolution within 6 weeks, and has been tumor free for 8 months of follow-up.

State programs for medical diagnosis of child abuse and neglect: case studies of five established or fledgling programs.

OBJECTIVE: To describe the programs for medical diagnosis of child abuse and neglect in three states and efforts to establish state-wide programs in two states. To describe common themes and issues that emerged related to the establishment and maintenance of these programs. METHODS: Five states were selected as case studies to represent a range of experience and type of function embodied in programs that address medical diagnosis of child abuse and neglect. Individuals knowledgeable about the programs or efforts to establish state-wide programs in their home states described these in detail. Inductive analysis was used to identify themes and issues that emerged across the states studied. FINDINGS: Themes emerged in three general areas: funding, services, and training. Findings related to funding were: 1) State funding was vital for initiation of statewide programs; 2) Alliances with other groups with parallel interests were successfully used to garner support for child abuse programs; 3) Services needed to be adequately reimbursed to be sustained; 4) Political climate often affected funding. With regard to services we found: 1) There was no optimal way to organize services, but rather many ways that worked well; 2) It was critical to address local service needs; 3) Provision of standardized quality services was essential. With regard to training: 1) Professional training was an integral part of all statewide programs; 2) New technologies, including televideo, have been explored and implemented to assist in training in statewide programs. CONCLUSIONS: Each state has taken a unique approach to programs for the medical diagnosis of child abuse and neglect. However, there are commonalities, particularly among the states that have been successful in establishing and maintaining comprehensive services and/or training.

The identification of a novel T cell activation state controlled by a diabetogenic gene.

The cyclin-dependent kinase inhibitor p27(kip) regulates the cell cycle at the G(1)-S phase restriction point. S phase entry and cell cycle commitment in peripheral T cells requires p27(kip) degradation, normally initiated by the receipt of costimulatory signals such as those provided by B7.1 or IL-2. We have previously reported that T cells from BioBreeding (BB)-diabetes-prone (DP) rats exhibit decreased costimulatory requirements for activation and cell cycle entry. In the present study, we find that peripheral T cell subsets from BB-DP rats demonstrate activation-like characteristics, including significantly reduced levels of p27(kip) as well as increased levels of proliferating cell nuclear Ag (PCNA). Since our previous studies have established that expression of extracellular activation markers are relatively low in unmanipulated peripheral BB-DP T cells; this p27(low) PCNA(high) phenotype represents a novel activation state. Analyses of T cell subsets from congenic rats demonstrate that this phenotype segregates with the lyp diabetogenic locus and that the p27(low) PCNA(high) phenotype is T cell specific. This p27(low) PCNA(high) phenotype is not seen in medullary thymocytes, but appears abruptly in the recent thymic emigrant population, suggesting that the lyp locus does not act directly on cell cycle regulators but rather alters the interaction between T cells and the peripheral environment. These results provide a biochemical basis for costimulation-independent activation and suggest a mechanism whereby a diabetes susceptibility gene contributes to disease development.

Cytoarchitectural and axonal maturation in human auditory cortex.

This study followed the maturation of human auditory cortex from the beginning of the second trimester of gestation to young adulthood. Histological and immunohistochemical techniques were used to trace the development of a laminar cytoarchitecture and an adult pattern of axonal neurofilament expression. From the 16th fetal week to the 4th postnatal month, the cortex progresses from a marginal layer and an undifferentiated cortical plate to incipient lamination. Between the 22nd fetal week and the 4th postnatal month, a two-tiered band of neurofilament-immunoreactive axons develops in layer I, but subsequent to the 4th month, the number of immunopositive axons in this layer is greatly reduced. Between the middle of the first year of life and age 3 years, the laminar pattern of cytoarchitecture becomes fully mature and a network of immunostained axons develops in layers VI, V, IV, and IlIc. This axonal plexus in the deep cortical layers continues to increase in density until age 5. Beginning at 5 years of age, a network of neurofilament-positive axons develops in the superficial layers IIIb, IIIa, and II, and by 11-12 years of age, overall axonal density is equivalent to that seen in young adulthood. This extended time span of axonal maturation has implications for the emergence of auditory cortical function.