Dysregulation of miR-34a links neuronal development to genetic risk factors for bipolar disorder.

Bipolar disorder (BD) is a heritable neuropsychiatric disorder with largely unknown pathogenesis. Given their prominent role in brain function and disease, we hypothesized that microRNAs (miRNAs) might be of importance for BD. Here we show that levels of miR-34a, which is predicted to target multiple genes implicated as genetic risk factors for BD, are increased in postmortem cerebellar tissue from BD patients, as well as in BD patient-derived neuronal cultures generated by reprogramming of human fibroblasts into induced neurons or into induced pluripotent stem cells (iPSCs) subsequently differentiated into neurons. Of the predicted miR-34a targets, we validated the BD risk genes ankyrin-3 (ANK3) and voltage-dependent L-type calcium channel subunit beta-3 (CACNB3) as direct miR-34a targets. Using human iPSC-derived neuronal progenitor cells, we further show that enhancement of miR-34a expression impairs neuronal differentiation, expression of synaptic proteins and neuronal morphology, whereas reducing endogenous miR-34a expression enhances dendritic elaboration. Taken together, we propose that miR-34a serves as a critical link between multiple etiological factors for BD and its pathogenesis through the regulation of a molecular network essential for neuronal development and synaptogenesis.

Developmental and activity-dependent genomic occupancy profiles of CREB in monkey area V1.

The mammalian neocortex displays significant plastic rearrangement in response to altered sensory input, especially during early postnatal development. It is believed that cyclic AMP-response element-binding (CREB) plays an important role in orchestrating the molecular events that guide neuroplastic change, although the details of its genomic targets during normal postnatal development or in response to sensory deprivation remain unknown. Here, we performed CREB chromatin immunoprecipitation (ChIP) from monkey area V1 tissue and hybridized enriched DNA fragments to promoter microarrays (ChIP chip analysis). Our goal was to determine and categorize the CREB regulon in monkey area V1 at two distinct developmental stages (peak of critical period vs. adulthood) and after 5 days of monocular enucleation (ME) at both ages. Classification of enriched candidates showed that the majority of isolated promoter loci (n = 795) were common to all four conditions. A particularly interesting group of candidates (n = 192) was specific to samples derived from enucleated infant area V1. Gene ontology analysis of CREB targets during early postnatal development showed a subgroup of genes implicated in cytoskeleton-based structural modification. Analysis of messenger RNA expression (quantitative real-time-polymerase chain reaction) of candidate genes showed striking differences in expression profiles between infant and adult area V1 after ME. Our study represents the first extensive genomic analysis of CREB DNA occupancy in monkey neocortex and provides new insight into the multifaceted transcriptional role of CREB in guiding neuroplastic change.

Temporal trends and spatial variability of mercury in four fish species in the Ontario segment of the St. Lawrence River, Canada.

The Massena (New York) and Cornwall (Ontario) region has a long history of Hg discharge into the St. Lawrence River. The objectives of this study were to evaluate if Hg levels have declined in this portion of the river since 1975 and to compare Hg level in fish species upstream and downstream of this area in order to evaluate the anthropogenic contribution to Hg levels in fish. Mercury levels in four fish species were monitored over a 20-year period (1975-1995). A general linear model and an analysis of covariance were used to extract temporal trends and spatial variability, respectively, while correcting the data for fish length. Over time, Hg levels declined in most fish species. In the four regions studied, Hg levels in fish were similar, which suggests that other sources like atmospheric deposition and Hg loading from the Great Lakes may also contribute to the Hg burden in fish in the St. Lawrence River. This indicates that fish, with large home range, are good biomonitors of temporal Hg releases but their ability to avoid point sources makes them less appealing as biomonitors to address spatial variability in Hg releases.

The effects of topiramate and sex hormones on energy balance of male and female rats.

OBJECTIVE: The effects of topiramate (TPM) on components of energy balance were tested in male and female rats that were (i) left intact, (ii) castrated or (iii) castrated with replacement therapies consisting of testosterone administration in orchidectomized (OCX) rats and of estradiol or progesterone treatments in ovariectomized (OVX) rats. METHODS: TPM was mixed into the diet and administered at a dose of 60 mg per kg of body weight. Male and female rats were treated for 28 and 35 days, respectively. At the end of the treatment period, variables of energy balance and determinants of lipid and glucose metabolism were assessed. RESULTS: TPM reduced energy and fat gains in both male and female rats either in the absence or in the presence of hormone replacement therapies. In male rats, it also decreased food intake, protein gain and energetic efficiency. In female animals, TPM reduced energetic efficiency while it stimulated lipoprotein lipase activity in brown adipose tissue. TPM also reduced plasma glucose and plasma leptin levels in female rats as well as plasma insulin and liver triglycerides in male animals. As expected, castration and sex hormones also strongly influenced energy balance. In male rats, OCX led to a decrease in energy and protein gains that was blocked by treatment with testosterone. In female rats, OVX caused increases in energy, fat and protein gains that were prevented by treatment with estradiol. CONCLUSION: In female rats, the effects of TPM on fat and energy gains were clearly not influenced by the sex hormone status of the rats. In male animals, there was also no interaction of TPM and the status of sex hormones on energy balance, suggesting that OCX and testosterone minimally interfere with the action of TPM on energy balance. The effects of TPM on energy balance were accounted for by a decrease in energetic efficiency, resulting from an effect exerted by the drug on both energy intake and thermogenesis. The present results also suggest that TPM can enhance insulin sensitivity.

Crystal structure of Staphylococcus aureus tyrosyl-tRNA synthetase in complex with a class of potent and specific inhibitors.

SB-219383 and its analogues are a class of potent and specific inhibitors of bacterial tyrosyl-tRNA synthetases. Crystal structures of these inhibitors have been solved in complex with the tyrosyl-tRNA synthetase from Staphylococcus aureus, the bacterium that is largely responsible for hospital-acquired infections. The full-length enzyme yielded crystals that diffracted to 2.8 A resolution, but a truncated version of the enzyme allowed the resolution to be extended to 2.2 A. These inhibitors not only occupy the known substrate binding sites in unique ways, but also reveal a butyl binding pocket. It was reported that the Bacillus stearothermophilus TyrRS T51P mutant has much increased catalytic activity. The S. aureus enzyme happens to have a proline at position 51. Therefore, our structures may contribute to the understanding of the catalytic mechanism and provide the structural basis for designing novel antimicrobial agents.

Why do young women diet? The roles of body fat, body perception, and body ideal.

BACKGROUND: To assess the relative roles of body fat, body perception, and body ideals as motivations for dieting in college women. METHOD: We compared 45 college women who reported having dieted with 32 who had not, using a novel computerized test of body image called the somatomorphic matrix. RESULTS: As expected, the difference in body fat between subjects' "perceived body" and "ideal body" was significantly greater in dieters than in nondieters (p < .001). Remarkably, however, this difference remained highly significant even after adjusting for the subjects' actual measured body fat (p = .002). Further analysis revealed that this difference persisted, not because dieters had unrealistic ideals of thinness, but because they had distorted perceptions of their fatness. CONCLUSION: Distorted body image perception, a potentially treatable condition, may play an unexpectedly large role in motivating young women to diet.

Canadian and American psychiatrists' attitudes toward dissociative disorders diagnoses.

OBJECTIVE: To compare the opinions of Canadian psychiatrists regarding dissociative disorder diagnoses with those of previously surveyed American psychiatrists. METHOD: We sent a 1-page questionnaire to a stratified representative sample of 550 Canadian psychiatrists. RESULTS: Eighty percent of Canadian psychiatrists responded. Fewer than one-third replied that dissociative amnesia and dissociative identity disorder should be included without reservations in the DSM-IV; fewer than 1 in 7 felt that the validity of these diagnoses was supported by strong scientific evidence. French- and English-speaking Canadians had similar opinions. Overall, Canadians were significantly less accepting than Americans. CONCLUSION: Both Canadian and American psychiatrists show little consensus regarding the diagnostic status or scientific validity of dissociative amnesia and dissociative identity disorder.

Photooxidation of Hg(0) in artificial and natural waters.

The oxidation of volatile aqueous Hg(0) in aquatic systems may be important in reducing fluxes of Hg out of aquatic systems. Here we report the results of laboratory and field experiments designed to identify the parameters that control the photooxidation of Hg(0)(aq) and to assess the possible importance of this process in aquatic systems. The concentrations of elemental and total Hg were measured as a function of time in both artificial and natural waters irradiated with a UV-B lamp. No change in Hg speciation was observed in dark controls, while a significant decrease in Hg(0) was observed in UV-B irradiated artificial solutions containing both chloride ions and benzoquinone. Significant photooxidation rates were also measured in natural samples spiked with Hg(0)(aq); the photooxidation of Hg(0) then follows pseudo first-order kinetics (k = 0.6 h(-1)). These results indicate that the previously observed Hg(II) photoreduction rates in natural waters could represent a net balance between Hg(0) photoreduction and Hg(0) photooxidation. As calculated from Hg(0) photooxidation rates, the dominant Hg(0) sink is likely to be photooxidation rather than volatilization from the water column during summer days.

Therapeutic ultrasound for chronic total coronary artery occlusions.

This article focuses on the use of catheter-delivered ultrasound as a device for treatment of coronary arterial total occlusion (CTO). The standard treatment of CTO is reviewed and serves as the basis for the need for new treatment options, such as catheter-delivered ultrasound (SONICROSS). The results of FDA-approved Phase I and Phase II clinical trials using SONICROSS for refractory CTO are described. The Phase II trials indicate that the SONICROSS-facilitated guidewire passage in 71% of CTOs are otherwise refractory to guidewire passage. However, at present, the SONICROSS catheter system is limited in its ability to be delivered reliably to the CTO because of catheter size and trackability.

Influence of topiramate in the regulation of energy balance.

Topiramate (TPM) is a novel neurotherapeutic agent currently indicated for the treatment of epilepsy and undergoing development for other central nervous system indications including neuropathic pain, bipolar disorder, and migraine prophylaxis. TPM is synthesized from D-fructose and contains a sulfamate moiety that is essential for its pharmacologic activity. TPM has been observed to significantly reduce body weight in patients treated for seizure, which has prompted the realization of preclinical studies to characterize the effects of TPM in the regulation of energy balance. Studies carried out in various strains of rats have provided good evidence for the ability of TPM to blunt energy deposition. Body composition analyses from rat trials have demonstrated that TPM inhibits fat deposition while reducing the activity of lipoprotein lipase (LPL) in various white adipose tissue depots. High doses of TPM (likely above the therapeutic dose range) have also been observed to reduce protein gain without catabolic effects. Although TPM cannot be described as a potent anorectic agent, it seems to have the ability to reduce food intake; significant reductions in food intake have been observed in female obese (fa/fa) Zucker rats and in female Wistar rats. TPM can also reduce energy deposition in the absence of alterations in food intake. This effect has been clearly emphasized in female lean (Fa/?) Zucker rats. In female Sprague-Dawley rats, TPM also increased energy expenditure and it has been observed to increase LPL activity in brown adipose tissue, which could indicate that TPM has the ability to enhance regulatory thermogenesis. In addition, TPM stimulates LPL activity in skeletal muscles, further emphasizing its potential to promote substrate oxidation. The mechanisms whereby TPM affects the regulation of energy balance have yet to be understood. TPM represents an antiepileptic drug (AED) with complex biochemical/pharmacologic actions. Its negative effects on energy deposition cannot be readily predicted from these actions, as AEDs are generally expected to stimulate body weight gain. Recent data, obtained from investigations aimed at assessing the effects of TPM on neuropeptidergic systems involved in the regulation of energy balance, have failed to demonstrate any significant effects of TPM on the neuropeptide Y and proopiomelanocortin systems. In conclusion, it is clear that TPM can reduce fat deposition by either reducing food intake or stimulating energy expenditure. The mechanisms whereby an AED such as TPM controls food intake and energy expenditure remains to be delineated. Copyright1999 ASCRS and ESCRS

Effects of the estrogen antagonist EM-652.HCl on energy balance and lipid metabolism in ovariectomized rats.

OBJECTIVE: The estrogen antagonist EM-652.HCl behaves as a highly potent and pure antiestrogen in human breast and uterine cancer cells. Because of its pure antiestrogenic activity in these cells, and because its prodrug, EM-800, reduces bone loss and decreases serum cholesterol and triglycerides in the rat, EM-652.HCl can be classified as a pure selective estrogen receptor modulator (SERM). This study was conducted to assess the ability of EM-652.HCl to prevent obesity and abnormalities of lipid metabolism induced by ovariectomy in a rat model. DESIGN: Female rats were left intact or ovariectomized (OVX), and OVX rats were treated with placebo, estradiol (E2), or EM-652.HCl for 20 days. At the end of the treatment period, parameters of energy balance and determinants of lipid metabolism were assessed. RESULTS: As expected, OVX increased energy intake, which in turn was accompanied by an increased energy, fat and protein gain and higher food efficiency. OVX also increased the triglyceride content of the liver and produced hypercholesterolemia and hyperinsulinemia. The weight of representative white adipose depots was higher in OVX than in intact rats. Lipoprotein lipase activity was higher in white adipose tissues of OVX rats than in those of intact animals, whereas its activity was lower in oxidative tissues (brown adipose and soleus muscle). Replacement therapy with a physiological dose of E2 prevented most of the abnormalities in energy and lipid metabolism brought about by OVX, although its orexigenic effect was only partially corrected. In contrast, treatment of OVX rats with EM-652. HCl completely abolished OVX-induced obesity and its related abnormalities in lipid metabolism and glucose/insulin homeostasis. CONCLUSION: These findings demonstrate that EM-652.HCl can be considered as an effective agent to prevent OVX-induced obesity. The present study also shows that EM-652.HCl reduces cardiovascular risk factors associated with obesity such as hyperlipidemia and insulin resistance.

Topiramate reduces energy and fat gains in lean (Fa/?) and obese (fa/fa) Zucker rats.

OBJECTIVE: This study examined the effects of topiramate (TPM), a novel neurotherapeutic agent reported to reduce body weight in humans, on the components of energy balance in female Zucker rats. RESEARCH METHODS AND PROCEDURES: A 2 x 3 factorial experiment was performed in which two cohorts of Zucker rats differing in their phenotype (phenotype: lean, Fa/?; obese, fa/fa) were each divided into three groups defined by the dose of TPM administered (dose: TPM 0, vehicle; TPM 15, 15 mg/kg; TPM 60, 60 mg/kg). RESULTS: The reduction in body weight gain induced by TPM in both lean and obese rats reflected a decrease in total body energy gain, which was more evident in obese than in lean rats. Whereas TPM administration did not influence the intake of digestible energy in lean rats, it induced a reduction in food intake in obese animals. In lean, but not in obese rats, apparent energy expenditure (as calculated by the difference between energy intake and energy gain) was higher in rats treated with TPM than in animals administered the vehicle. The low dose of TPM decreased fat gain (with emphasis on subcutaneous fat) without affecting protein gain, whereas the high dose of the drug induced a reduction in both fat and protein gains. The effects of TPM on muscle and fat depot weights were representative of the global effects of TPM on whole body fat and protein gains. The calculated energetic efficiency (energy gain/energy intake) was decreased in both lean and obese rats after TPM treatment. TPM dose independently reduced hyperinsulinemia of obese rats, but it did not alter insulinemia of lean animals. DISCUSSION: The present results provide sound evidence for the ability of TPM to reduce fat and energy gains through reducing energetic efficiency in both lean and obese Zucker rats.

HLS7, a hemopoietic lineage switch gene homologous to the leukemia-inducing gene MLF1.

Hemopoietic lineage switching occurs when leukemic cells, apparently committed to one lineage, change and display the phenotype of another pathway. cDNA representational difference analysis was used to identify myeloid-specific genes that may be associated with an erythroid to myeloid lineage switch involving the murine J2E erythroleukemic cell line. One of the genes isolated (HLS7) is homologous to the novel human oncogene myeloid leukemia factor 1 (MLF1) involved in the t(3;5)(q25.1;q34) translocation associated with acute myeloid leukemia. Enforced expression of HLS7 in J2E cells induced a monoblastoid phenotype, thereby recapitulating the spontaneous erythroid to myeloid lineage switch. HLS7 also inhibited erythropoietin- or chemically-induced differentiation of erythroleukemic cell lines and suppressed development of erythropoietin-responsive colonies in semi-solid culture. However, intracellular signaling activated by erythropoietin was not impeded by ectopic expression of HLS7. In contrast, HLS7 promoted maturation of M1 monoblastoid cells and increased myeloid colony formation in vitro. These data show that HLS7 can influence erythroid/myeloid lineage switching and the development of normal hemopoietic cells.

Polarized human immunodeficiency virus budding in lymphocytes involves a tyrosine-based signal and favors cell-to-cell viral transmission.

Maturation and release of human immunodeficiency virus type 1 (HIV-1) is targeted at the pseudopod of infected mononuclear cells. However, the intracellular mechanism or targeting signals leading to this polarized viral maturation are yet to be identified. We have recently demonstrated the presence of a functional YXXL motif for specific targeting of HIV-1 virions to the basolateral membrane surface in polarized epithelial Madin-Darby canine kidney cells (MDCK). Site-directed mutagenesis was used to demonstrate that the membrane-proximal tyrosine in the intracytoplasmic tail of the HIV-1 transmembrane glycoprotein (gp41) is an essential component of this signal. In the present study, immunolocalization of viral budding allowed us to establish that this tyrosine-based signal is involved in determining the exact site of viral release at the surface of infected mononuclear cells. Substitution of the critical tyrosine residue was also shown to increase the amount of envelope glycoprotein at the cell surface, supporting previous suggestions that the tyrosine-based motif can promote endocytosis. Although alteration of the dual polarization-endocytosis motif did not affect the infectivity of cell-free virus, it could play a key role in cell-to-cell viral transmission. Accordingly, chronically infected lymphocytes showed a reduced ability to transmit the mutant virus to a cocultivated cell line. Overall, our data indicate that the YXXL targeting motif of HIV is active in various cell types and could play an important role in viral propagation; this may constitute an alternative target for HIV therapeutics and vaccine development.

The crystal structure of human procathepsin K.

Cathepsin K is a cysteine protease present in human osteoclasts that plays an important role in bone resorption. Cathepsin K is synthesized as an inactive proenzyme and activated under conditions of low pH. Autoproteolytic processing of the N-terminal 99 amino acid propeptide produces the active, mature form of cathepsin K. It is presumed that the activation of procathepsin K in vivo occurs in the bone resorption pit, which has a low-pH environment. We have determined the structure of human procathepsin K at 2.8 A resolution. The structure of the mature enzyme domain within procathepsin K is virtually identical to that of mature cathepsin K. The fold of the propeptide of procathepsin K is similar to that observed in procathepsins B and L despite differences in length and sequence. A portion of the propeptide occupies the active site cleft of cathepsin K. Hydrophobic interactions, salt bridges, and hydrogen-bonding interactions are observed in the structure of the propeptide and between the propeptide and the mature enzyme of procathepsin K. These interactions suggest an explanation for the stability of the proenzyme. The structure of procathepsin K contributes to an understanding of the molecular basis of inhibition by the propeptide portion of the molecule and activation of this important member of the cysteine protease family.

Use of papain as a model for the structure-based design of cathepsin K inhibitors: crystal structures of two papain-inhibitor complexes demonstrate binding to S'-subsites.

Papain has been used as a surrogate enzyme in a drug design effort to obtain potent and selective inhibitors of cathepsin K, a new member of the papain superfamily of cysteine proteases that is selectively and highly expressed in osteoclasts and is implicated in bone resorption. Here we report the crystal structures of two papain-inhibitor complexes and the rational design of novel cathepsin K inhibitors. Unlike previously known crystal structures of papain-inhibitor complexes, our papain structures show ligand binding extending deep within the S'-subsites. The two inhibitor complexes, carbobenzyloxyleucinyl-leucinyl-leucinal and carbobenzyloxy-L-leucinyl-L-leucinyl methoxymethyl ketone, were refined to 2.2- and 2.5-A resolution with R-factors of 0.190 and 0. 217, respectively. The S'-subsite interactions with the inhibitors are dominated by an aromatic-aromatic stacking and an oxygen-aromatic ring edge interaction. The knowledge of S'-subsite interactions led to a design strategy for an inhibitor spanning both subsites and yielded a novel, symmetric inhibitor selective for cathepsin K. Simultaneous exploitation of both S- and S'-sites provides a general strategy for the design of cysteine protease inhibitors having high specificity to their target enzymes.

Attention-deficit hyperactivity disorder subtypes and comorbid disruptive behaviour disorders in a child and adolescent mental health clinic.

OBJECTIVE: To assess demographic characteristics and patterns of comorbid disruptive behavior disorders (oppositional defiant disorder [ODD] or conduct disorder [CD]) in subtypes of attention-deficit hyperactivity disorder (ADHD). METHOD: One hundred youths consecutively referred to a community child and adolescent mental health clinic and subsequently diagnosed with ADHD by the Diagnostic and Statistical Manual of Mental Disorders (DSM-IV) criteria were evaluated. The diagnosis was made by a child psychiatrist and was based on information from physicians, parents, teachers, and diagnostic interviews with the youth and their parents. RESULTS: The major findings were: 1) ADHD combined (C) type was diagnosed in 78% of the subjects, while 15% had inattentive (1) type and 7% had hyperactive-impulsive (HI) type; and 2) patterns of comorbid disruptive behavioural disorders significantly differed among subtypes. Specifically, subjects with the I type showed lower rates of comorbid ODD than those with the C type (33% and 85%; P < 0.001) and HI type (33% and 100%; P = 0.005); subjects with the HI type displayed a higher prevalence of CD than those with the I type (57% and 0%; P = 0.005) and C type (57% and 8%; P = 0.003). These results should be considered tentative because the reliability of the diagnostic procedures was not formally assessed and the number of subjects in the I and HI groups was small. CONCLUSION: ADHD subtypes showed significant differences in the distribution of comorbid disruptive behaviour disorders. These results support the utility of ADHD subtypes but should be replicated with a larger sample of I and HI type subjects using more rigorous diagnostic methods.