Chronic smoking and alcoholism change expression of selective genes in the human prefrontal cortex

Background: Alcoholism is commonly associated with chronic smoking. A number of gene expression profiles of regions within the human mesocorticolimbic system have identified potential alcohol-sensitive genes; however, the influence of smoking on these changes was not taken into account. This study addressed the impact of alcohol and smoking on the expression of 4 genes, previously identified as alcoholism-sensitive. in the human prefrontal cortex (PFC). Methods: mRNA expression of apolipoprotein D, tissue inhibitor of the metalloproteinase 3, high-affinity glial glutamate transporter and midkine, was measured in the PFC of alcoholic Subjects and controls with and without smoking comorbidity using real-time polymerase chain reaction. Results: The results show that alcohol affects transcription of some of these genes. Additionally, smoking has a marked influence on gene expression. Conclusion: This study emphasizes the need for careful case selection in future gene expression studies to delineate the adaptive molecular process associated with smoking and alcohol.

Defective p53 response and apoptosis associated with an ataxia-telangiectasia-like phenotype

Ataxia-telangiectasia mutated (ATM), the protein defective in ataxia-telangiectasia, plays a central role in DNA damage response and signaling to cell cycle checkpoints. We describe here a cell line from a patient with an ataxia-telangiectasia-like clinical phenotype defective in the p53 response to radiation but with normal ATM activation and efficient downstream phosphorylation of other ATM substrates. No mutations were detected in ATM cDNA. A normal level of interaction between p53 and peptidyl-prolyl-isomerase Pin1 suggests that posttranslational modification was intact in these cells but operating at reduced level. Defective p53 stabilization was accompanied by defective induction of p53 effector genes and failure to induce apoptosis in response to DNA-damaging agents. Continued association between p53 and murine double minute-2 (Mdm2) occurred in irradiated ATL2ABR cells in response to DNA damage, and incubation with Mdm2 antagonists, nutlins, increased the stabilization of p53 and its transcriptional activity but failed to induce apoptosis. These results suggest that ATM-dependent stabilization of p53 and induction of apoptosis by radiation involve an additional factor(s) that is defective in ATL2ABR cells.

Distinct physiological mechanisms underlie altered glycinergic synaptic transmission in the murine mutants spastic, spasmodic, and oscillator

Spastic (spa), spasmodic (spd), and oscillator (ot) mice have naturally occurring glycine receptor ( GlyR) mutations, which manifest as motor deficits and an exaggerated startle response. Using whole-cell recording in hypoglossal motoneurons, we compared the physiological mechanisms by which each mutation alters GlyR function. Mean glycinergic miniature IPSC ( mIPSC) amplitude and frequency were dramatically reduced (> 50%) compared with controls for each mutant. mIPSC decay times were unchanged in spa/spa (4.5 +/- 0.3 vs 4.7 +/- 0.2 ms), reduced in spd/spd (2.7 +/- 0.2 vs 4.7 +/- 0.2 ms), and increased in ot/ot (12.3 +/- 1.2 vs 4.8 +/- 0.2 ms). Thus, in spastic, GlyRs are functionally normal but reduced in number, whereas in spasmodic, GlyR kinetics is faster. The oscillator mutation results in complete absence of alpha 1-containing GlyRs; however, some non-alpha 1-containing GlyRs persist at synapses. Fluctuation analysis of membrane current, induced by glycine application to outside-out patches, showed that mean single-channel conductance was increased in spa/spa (64.2 +/- 4.9 vs 36.1 +/- 1.4 pS), but unchanged in spd/spd (32.4 +/- 2.1 vs 35.3 +/- 2.1 pS). GlyR-mediated whole-cell currents in spa/spa exhibited increased picrotoxin sensitivity (27 vs 71% block for 100 mu M), indicating alpha 1 homomeric GlyR expression. The picrotoxin sensitivity of evoked glycinergic IPSCs and conductance of synaptic GlyRs, as determined by nonstationary variance analysis, were identical for spa/spa and controls. Together, these findings show the three mutations disrupt GlyR-mediated inhibition via different physiological mechanisms, and the spastic mutation results in compensatory alpha 1 homomeric GlyRs at extrasynaptic loci.

Emergency department assessment of mild traumatic brain injury and prediction of post-concussion symptoms at one month post injury

Mild traumatic brain injury (mTBI) is a common injury and a significant proportion of those affected report chronic symptoms. This study investigated prediction of post-concussion symptoms using an Emergency Department (ED) assessment that examined neuropsychological and balance deficits and pain severity of 29 concussed individuals. Thirty participants with minor orthopedic injuries and 30 ED visitors were recruited as control subjects. Concussed and orthopedically injured participants were followed up by telephone at one month to assess symptom severity. In the ED, concussed subjects performed worse on some neuropsychological tests and had impaired balance compared to controls. They also reported significantly more post-concussive symptoms at follow-up. Neurocognitive impairment, pain and balance deficits were all significantly correlated with severity of post-concussion symptoms. The findings suggest that a combination of variables assessable in the ED may be useful in predicting which individuals will suffer persistent post-concussion problems.

Factors affecting female reproductive success and the survival of pouch young in the threatened brush-tailed rock-wallaby, Petrogale penicillata

Knowledge of factors affecting the survival of individuals and their reproductive success is essential for threatened species management, but studies assessing these factors are lacking for many threatened rock-wallaby species. In this study we investigated the factors influencing the breeding performance of females and the survival of pouch young in a wild colony of the threatened brush-tailed rock-wallaby. Individuals were trapped between October 2000 and April 2004. More than 50% of the females in the colony were breeding below their full potential and giving birth to only one offspring per year. Most females within the colony bred in synchrony, with a substantial birth peak evident during autumn. Pouch young born in autumn left the pouch during spring and were weaned during summer and autumn when forage was most abundant. Pouch young born during the autumn birth peak or in winter had a substantially higher probability of surviving through to pouch emergence than those born during spring or summer. This study provides demographic parameters that may be used in population models and for comparison with other populations, particularly those that are small and declining. To optimise reproductive success in reintroduction programs, females in good condition and with small pouch young should be released at the end of the wettest season.

Genetics and genomics of osteoclast differentiation: Integrating cell signaling pathways and gene networks

The regulation of osteoclast differentiation in the bone microenvironment is critical for normal bone remodeling, as well as for various human bone diseases. Over the last decade, our knowledge of how osteoclast differentiation occurs has progressed rapidly. We highlight some of the major advances in understanding how cell signaling and transcription are integrated to direct the differentiation of this cell type. These studies used genetic, molecular, and biochemical approaches. Additionally, we summarize data obtained from studies of osteoclast differentiation that used the functional genomic approach of global gene profiling applied to osteoclast differentiation. This genomic data confirms results from studies using the classical experimental approaches and also may suggest new modes by which osteoclast differentiation and function can be modulated. Two conclusions that emerge are that osteoclast differentiation depends on a combination of fairly ubiquitously expressed transcription factors rather than unique osteoclast factors, and that the overlay of cell signaling pathways on this set of transcription factors provides a powerful mechanism to fine tune the differentiation program in response to the local bone microenvironment.

How is leflunomide prescribed and used in Australia? analysis of prescribing and adverse effect reporting

Purpose To evaluate the use of leflunomide in the Australian community since introduction in 2000. Trends in adverse drug reaction (ADR) reporting were also studied. Methods Annual Australian prescription and dispensing statistics were analysed. Drug utilisation was estimated as defined daily doses (DDD)/1000 inhabitants/day. ADR data from the Therapeutic Goods Administration's Adverse Drug Reactions Advisory Committee (ADRAC) national monitoring system were compared with the World Health Organisation (WHO) Vigibase records. Results Leflunomide use in Australia (dispensing data) increased from 0.2 in 2000 to 0.4 DDD/1000 inhabitants/day in 2002. The same overall pattern was observed in the 'authority to prescribe' data. From 2000-2002, prescribing of the starter pack (3 x 100 mg loading dose plus 30 x 20 mg tablets) declined (down 74%); likewise for the 20mg (30 tablets) pack. Gradual increases were noted for the 10 mg (30 tablets) pack (up 40%). Approximately 135 reports, detailing about 370 individual ADR, were generated annually. Gastro-intestinal disorders predominated, accounting for 24% of reactions reported to ADRAC. Skin and appendages disorders constituted 14% of reported reactions. Deaths in leflunomide users were attributed to a combination of haematological and gastro-intestinal complications, but it was not possible to ascertain other medication usage or contributing factors. Trends observed with the ADRAC reports were consistent with the WHO database. Conclusions Leflunomide was the first registered DMARD in Australia in over a decade and its use has increased within the community. The ADR reports might have contributed to Australian rheumatologists gradually abandoning loading patients with high doses of leflunomide in favour of starting therapy at lower doses. Copyright (c) 2006 John Wiley & Sons, Ltd.

Impacts of feeding system and season on milk composition and Cheddar cheese yield in a subtropical environment

Milk obtained from cows on 2 subtropical dairy feeding systems were compared for their suitability for Cheddar cheese manufacture. Cheeses were made in a small-scale cheesemaking plant capable of making 2 blocks ( about 2 kg each) of Cheddar cheese concurrently. Its repeatability was tested over 10 separate cheesemaking days with no significant differences being found between the 2 vats in cheesemaking parameters or cheese characteristics. In the feeding trial, 16 pairs of Holstein - Friesian cows were used in 2 feeding systems (M1, rain-grown tropical grass pastures and oats; and M5, a feedlot, based on maize/barley silage and lucerne hay) over 2 seasons ( spring and autumn corresponding to early and late lactation, respectively). Total dry matter, crude protein (kg/cow. day) and metabolisable energy (MJ/cow.day) intakes were 17, 2.7, and 187 for M1 and 24, 4, 260 for M5, respectively. M5 cows produced higher milk yields and milk with higher protein and casein levels than the M1 cows, but the total solids and fat levels were similar (P > 0.05) for both M1 and M5 cows. The yield and yield efficiency of cheese produced from the 2 feeding systems were also not significantly different. The results suggest that intensive tropical pasture systems can produce milk suitable for Cheddar cheese manufacture when cows are supplemented with a high energy concentrate. Season and stage of lactation had a much greater effect than feeding system on milk and cheesemaking characteristics with autumn ( late lactation) milk having higher protein and fat contents and producing higher cheese yields.

Liver fluke-associated and sporadic cholangiocarcinoma: an immunohistochemical study of bile duct, peribiliary gland and tumour cell phenotypes

Aim: To compare cell phenotypes displayed by cholangiocarcinomas and adjacent bile duct lesions in patients from an area endemic in liver-fluke infestation and those with sporadic cholangiocarcinoma. Methods: 65 fluke-associated and 47 sporadic cholangiocarcinomas and 6 normal livers were studied. Serial paraffin-wax sections were stained immunohistochemically with monoclonal antibodies characterising a Brunner or pyloric gland metaplasia cell phenotype (antigens D10 and 1F6), intestinal goblet cells (antigen 17NM), gastric foveolar apomucin (MUC5AC), a gastrointestinal epithelium cytokeratin (CK20) and the p53 protein. Results: 60% of the 112 cholangiocarcinomas expressed antigen D10, 68% MUC5AC, 33% antigen 17NM and 20% CK20; 37% showed overexpression of p53. When present together in a cholangiocarcinoma, cancer cells expressing D10 were distinct from those displaying 17NM or MUC5AC. Many more fluke-associated cholangiocarcinomas than sporadic cholangiocarcinomas displayed 17NM and p53 expression. Most cases of hyperplastic and dysplastic biliary epithelium expressed D10 strongly. Pyloric gland metaplasia and peribiliary glands displayed D10 and 1F6, with peribiliary gland hyperplasia more evident in the livers with fluke-associated cholangiocarcinoma; goblet cells in intestinal metaplasia stained for 17NM. No notable association of expression between any two antigens (including p53) was found in the cancers. Conclusions: Most cases of dysplastic biliary epithelium and cholangiocarcinoma display a Brunner or pyloric gland cell phenotype and a gastric foveolar cell phenotype. The expression of D10 in hyperplastic and dysplastic epithelium and in cholangiocarcinoma is consistent with a dysplasia-carcinoma sequence. Many more fluke-associated cholangiocarcinomas than sporadic cholangiocarcinoma display an intestinal goblet cell phenotype and overexpress p53, indicating differences in the aetiopathology of the cancers in the two groups of patients.

Neural progenitor number is regulated by nuclear factor-kappa B p65 and p50 subunit-dependent proliferation rather than cell survival

The number of cells generated by a proliferating stem or precursor cell can be influenced both by proliferation and by the degree of cell death/survival of the progeny generated. In this study, the extent to which cell survival controls progenitor number was examined by comparing the growth characteristics of neurosphere cultures derived from mice lacking genes for the death inducing Bcl-2 homologue Hara Kiri (Hrk), apoptosis-associated protein 1 (Apaf1), or the prosurvival nuclear factor-kappa B (NF kappa B) subunits p65, p50, or c-rel. We found no evidence that Hrk or Apaf1, and by inference the mitochondrial cell death pathway, are involved in regulating the number of neurosphere-derived progeny. However, we identified the p65p50 NF kappa B dimer as being required for the normal growth and expansion of neurosphere cultures. Genetic loss of both p65 and p50 NF kappa B subunits resulted in a reduced number of progeny but an increased proportion of neurons. No effect on cell survival was observed. This suggests that the number and fate of neural progenitor cells are more strongly regulated by cell cycle control than survival. (c) 2005 Wiley-Liss, Inc.

Neuronal voltage-gated sodium channel subtypes: Key roles in inflammatory and neuropathic pain

Voltage-gated sodium channels (VGSCs) play an important role in neuronal excitability. Regulation of VGSC activity is a complex phenomenon that occurs at multiple levels in the cell, including transcriptional regulation, post-translational modification and membrane insertion and retrieval. Multiple VGSC subtypes exist that vary in their biophysical and pharmacological properties and tissue distribution. Any alteration of the VGSC subtype profile of a neuron or the mechanisms that regulate VGSC activity can cause significant changes in neuronal excitability. Inflammatory and neuropathic pain states are characterised by alterations in VGSC subtype composition and activity in sensory neurons. This review focuses on the VGSC subtypes involved in such pain states. (c) 2006 Elsevier Ltd. All rights reserved.

Identification and molecular modeling of a novel, plant-like, human purple acid phosphatase

Purple acid phosphatases are a family of binuclear metallohydrolases that have been identified in plants, animals and fungi. Only one isoform of similar to 35 kDa has been isolated from animals, where it is associated with bone resorption and microbial killing through its phosphatase activity, and hydroxyl radical production, respectively. Using the sensitive PSI-BLAST search method, sequences representing new purple acid phosphatase-like proteins have been identified in mammals, insects and nematodes. These new putative isoforms are closely related to the similar to 55 kDa purple acid phosphatase characterized from plants. Secondary structure prediction of the new human isoform further confirms its similarity to a purple acid phosphatase from the red kidney bean. A structural model for the human enzyme was constructed based on the red kidney bean purple acid phosphatase structure. This model shows that the catalytic centre observed in other purple acid phosphatases is also present in this new isoform. These observations suggest that the sequences identified in this study represent a novel subfamily of plant-like purple acid phosphatases in animals and humans. (c) 2006 Elsevier B.V. All rights reserved.

Imaging, anatomical, and molecular analysis of callosal formation in the developing human fetal brain

A complex set of axonal guidance mechanisms are utilized by axons to locate and innervate their targets. In the developing mouse forebrain, we previously described several midline glial populations as well as various guidance molecules that regulate the formation of the corpus callosum. Since agenesis of the corpus callosum is associated with over 50 different human congenital syndromes, we wanted to investigate whether these same mechanisms also operate during human callosal development. Here we analyze midline glial and commissural development in human fetal brains ranging from 13 to 20 weeks of gestation using both diffusion tensor magnetic resonance imaging and immunohistochemistry. Through our combined radiological and histological studies, we demonstrate the morphological development of multiple forebrain commissures/decussations, including the corpus callosum, anterior commissure, hippocampal commissure, and the optic chiasm. Histological analyses demonstrated that all the midline glial populations previously described in mouse, as well as structures analogous to the subcallosal sling and cingulate pioneering axons, that mediate callosal axon guidance in mouse, are also present during human brain development. Finally, by Northern blot analysis, we have identified that molecules involved in mouse callosal development, including Slit, Robo, Netrin1, DCC, Nfia, Emx1, and GAP-43, are all expressed in human fetal brain. These data suggest that similar mechanisms and molecules required for midline commissure formation operate during both mouse and human brain development. Thus, the mouse is an excellent model system for studying normal and pathological commissural formation in human brain development. (c) 2006 Wiley-Liss, Inc.

Nucleolar localization of aprataxin is dependent on interaction with nucleolin and on active ribosomal DNA transcription

The APTX gene, mutated in patients with the neurological disorder ataxia with oculomotor apraxia type 1 (AOA1), encodes a novel protein aprataxin. We describe here, the interaction and interdependence between aprataxin and several nucleolar proteins, including nucleolin, nucleophosmin and upstream binding factor-1 (UBF-1), involved in ribosomal RNA (rRNA) synthesis and cellular stress signalling. Interaction between aprataxin and nucleolin occurred through their respective N-terminal regions. In AOA1 cells lacking aprataxin, the stability of nucleolin was significantly reduced. On the other hand, down-regulation of nucleolin by RNA interference did not affect aprataxin protein levels but abolished its nucleolar localization suggesting that the interaction with nucleolin is involved in its nucleolar targeting. GFP-aprataxin fusion protein co-localized with nucleolin, nucleophosmin and UBF-1 in nucleoli and inhibition of ribosomal DNA transcription altered the distribution of aprataxin in the nucleolus, suggesting that the nature of the nucleolar localization of aprataxin is also dependent on ongoing rRNA synthesis. In vivo rRNA synthesis analysis showed only a minor decrease in AOA1 cells when compared with controls cells. These results demonstrate a cross-dependence between aprataxin and nucleolin in the nucleolus and while aprataxin does not appear to be directly involved in rRNA synthesis its nucleolar localization is dependent on this synthesis.