Subcellular distribution of the enzymes of the malate-aspartate shuttle in rat heart and effect of experimental cardiac hypertrophy

The effect of experimental cardiac hypertrophy on the enzymes of the malate - aspartate shuttle aspartate aminotransferase (AAT) and malate dehydrogenase (MDH) was studied. ( l ) Aortic constriction in adult rats resulted in 25% cardiac hypertrophy in 2 1/2-3 weeks. Total DNA (mg per heart) did not change. ( 2 ) The proportions of mitochondrial and cytosolic isozymes of AAT and MDH did not change as a result of cardiac h y p e r t r o p h y . About two-thirds of each enzyme occurred in the mitochondrial form and one-third in the cytosolic form. ( 3 ) Total AAT in hypertrophic hearts, in enzyme units per mg DNA, increased by 24% compared to AAT content in the hearts of sham-operated animals . Total MDH did not change. SoIubilized protein increased by 20%. Normal hearts contained 10 times more enzyme units of MDH than of AAT. (4) Cardiac growth stimulation induced in newborn rats did not result in specific changes of either enzyme. It is suggested that true cardiac hypertrophy acts as a specific stimulus for the possibly rate-limiting enzyme AAT of the shuttle.     

Child Behavior Checklist—Mania Scale (CBCL-MS): Development and Evaluation of a Population-Based Screening Scale for Bipolar Disorder

Context

Early identification of Bipolar Disorder (BD) remains poor despite the high levels of disability associated with the disorder.

Objective

We developed and evaluated a new DSM orientated scale for the identification of young people at risk for BD based on the Child Behavior Checklist (CBCL) and compared its performance against the CBCL-Pediatric Bipolar Disorder (CBCL-PBD) and the CBCL-Externalizing Scale, the two most widely used scales.

Methods

The new scale, CBCL-Mania Scale (CBCL-MS), comprises 19 CBCL items that directly correspond to operational criteria for mania. We tested the reliability, longitudinal stability and diagnostic accuracy of the CBCL-MS on data from the TRacking Adolescents'' Individual Lives Survey (TRAILS), a prospective epidemiological cohort study of 2230 Dutch youths assessed with the CBCL at ages 11, 13 and 16. At age 19 lifetime psychiatric diagnoses were ascertained with the Composite International Diagnostic Interview. We compared the predictive ability of the CBCL-MS against the CBCL-Externalising Scale and the CBCL-PBD in the TRAILS sample.

Results

The CBCL-MS had high internal consistency and satisfactory accuracy (area under the curve = 0.64) in this general population sample. Principal Component Analyses, followed by parallel analyses and confirmatory factor analyses, identified four factors corresponding to distractibility/disinhibition, psychosis, increased libido and disrupted sleep. This factor structure remained stable across all assessment ages. Logistic regression analyses showed that the CBCL-MS had significantly higher predictive ability than both the other scales.

Conclusions

Our data demonstrate that the CBCL-MS is a promising screening instrument for BD. The factor structure of the CBCL-MS showed remarkable temporal stability between late childhood and early adulthood suggesting that it maps on to meaningful developmental dimensions of liability to BD.     

A highly sensitive molecular structural probe applied to in situ biosensing of metabolites using PEDOT:PSS

A large amount of research within organic biosensors is dominated by organic electrochemical transistors (OECTs) that use conducting polymers such as poly(3,4-ethylene dioxythiophene) doped with poly(styrenesulfonate) (PEDOT:PSS). Despite the recent advances in OECT-based biosensors, the sensing is solely reliant on the amperometric detection of the bioanalytes. This is typically accompanied by large undesirable parasitic electrical signals from the electroactive components in the electrolyte. Herein, we present the use of in situ resonance Raman spectroscopy to probe subtle molecular structural changes of PEDOT:PSS associated with its doping level. We demonstrate how such doping level changes of PEDOT:PSS can be used, for the first time, on operational OECTs for sensitive and selective metabolite sensing while simultaneously performing amperometric detection of the analyte. We test the sensitivity by molecularly sensing a lowest glucose concentration of 0.02 mM in phosphate-buffered saline solution. By changing the electrolyte to cell culture media, the selectivity of in situ resonance Raman spectroscopy is emphasized as it remains unaffected by other electroactive components in the electrolyte. The application of this molecular structural probe highlights the importance of developing biosensing probes that benefit from high sensitivity of the material's structural and electrical properties while being complimentary with the electronic methods of detection.     

Lecithin/cholesterol acyltransferase modulates diet-induced hepatic deposition of triglycerides in mice

Lecithin/cholesterol acyltransferase (LCAT) is responsible for the esterification of the free cholesterol of plasma lipoproteins. Here, we investigated the involvement of LCAT in mechanisms associated with diet-induced hepatic triglyceride accumulation in mice. LCAT-deficient (LCAT^?/?) and control C57BL/6 mice were placed on a Western-type diet (17.3% protein, 48.5% carbohydrate, 21.2% fat, 0.2% cholesterol, 4.5 kcal/g) for 24 weeks, then histopathological and biochemical analyses were performed. We report that, in our experimental setup, male LCAT^?/? mice are characterized by increased diet-induced hepatic triglyceride deposition and impaired hepatic histology and architecture. Mechanistic analyses indicated that LCAT deficiency was associated with enhanced intestinal absorption of dietary triglycerides (3.6±0.5 mg/dl per minute for LCAT^?/? vs. 2.0±0.7 mg/dl per minute for C57BL/6 mice; P<.05), accelerated clearance of postprandial triglycerides and a reduced rate of hepatic very low density lipoprotein triglyceride secretion (9.8±1.1 mg/dl per minute for LCAT^?/? vs. 12.5±1.3 mg/dl per minute for C57BL/6 mice, P<.05). No statistical difference in the average daily food consumption between mouse strains was observed. Adenovirus-mediated gene transfer of LCAT in LCAT^?/? mice that were fed a Western-type diet for 12 weeks resulted in a significant reduction in hepatic triglyceride content (121.2±5.9 mg/g for control infected mice vs. 95.1±5.8 mg/g for mice infected with Ad-LCAT, P<.05) and a great improvement of hepatic histology and architecture. Our data extend the current knowledge on the functions of LCAT, indicating that LCAT activity is an important modulator of processes associated with diet-induced hepatic lipid deposition.     

Conformational study of new amphipathic alpha-helical peptide models of apoA-I as potential atheroprotective agents

Aiming at contributing to the development of potential atheroprotective agents, we report on the concept and design of two peptide models, which mimic the amphipathic helices of apoA-I and incorporate Met into their sequences to validate its role as oxidant scavenger: Ac-ESK(Palm)KELSKSW(10)SEM(13)LKEK(Palm)SKS-NH(2) (model 1 [W(10), M(13)]) and Ac-ESK(Palm)KELSKSM(10)SEW(13)LKEK(Palm)SKS-NH(2) (model 2 [M(10), W(13)]). Hydrophobic residues of both models cover about the half of the surface, while the positively and negatively charged residues constitute two separate clusters on the hydrophilic face. Palmitoyl groups were introduced into the Lys-N(epsilon)H(2) groups at positions 3 and 17 to contribute to the amphipathic character of the peptides and stabilize the nonpolar face of the helix. Conformational study by the combined application of 2D-NMR and molecular dynamics simulations, CD, FTIR, and fluorescence spectroscopy revealed that model 1 adopts helical conformation and Met is well exposed to the microenvironment. Model 2 that derives from model 1 by exchanging W(10) (model 1) with M(10) and M(13) (model 1) with W(13) also displays helical characteristics, while Met is rather shielded. Oxidation experiments indicated that model 1 exhibits a 2-fold more potent antioxidant activity towards LDL oxidation, compared to model 2, confirming the role of Met, when is devoid of steric hindrances, as oxidant scavenger for the protection of LDL.     

The forgotten serine. A critical role for Ser-2035.42 in ligand binding to and activation of the beta 2-adrenergic receptor

Previous work in the beta(2)-adrenergic receptor demonstrated critical interactions between Ser-204 and Ser-207 in the fifth membrane-spanning segment and the meta-OH and para-OH, respectively, of catecholamine agonists (Strader, C. D., Candelore, M. R., Hill, W. S., Sigal, I. S., and Dixon, R. A. (1989) J. Biol. Chem. 264, 13572-13578). Using the substituted cysteine accessibility method in the beta(2)-adrenergic receptor, we have found that in addition to Ser-204 and Ser-207, Ser-203 is also accessible on the surface of the binding-site crevice and is occluded by bound agonist. Mutation of Ser-203 to Ala, Val, or Cys reduced the binding affinity and adenylyl cyclase-activating potency of agonists containing a meta-OH, whereas their affinities and potencies were largely preserved by mutation of Ser-203 to Thr, which maintained an OH at this position. Thus both Ser-203 and Ser-204 appear to interact with the meta-OH of catecholamines, perhaps through a bifurcated H bond. Furthermore, the removal of the OH at position 203 led to a significant loss of affinity of antagonists with nitrogen in their heterocyclic ring structure. The greatest effect was seen with pindolol, a partial agonist, suggesting that a H bond between the heterocyclic ring and Ser-203 may play a role in partial agonism. In contrast, the affinities of antagonists such as propranolol or alprenolol, which have cyclic structures without H-bonding capability, were unaltered after mutation of Ser-203.     

Genetic skeletal disorders of the fetus and infant: Pathologic and molecular findings in a series of 41 cases

BACKGROUND: Genetic skeletal disorders of the fetus and infant are a large group of genetic disorders, comprising the groups formerly assigned as skeletal dysplasias (osteochondrodysplasias), dysostoses, and malformation syndromes with a skeletal component. Genetic skeletal disorders may be prenatally detected by ultrasonography or result in intrauterine or early postnatal death, constituting one difficult diagnostic field met by the pathologist who performs the perinatal autopsy. METHODS: In this retrospective study, we have gathered radiologic, physical, histopathologic, and molecular data regarding 41 cases of genetic skeletal disorders diagnosed among 1980 fetal and perinatal autopsies over a 10‐year period. RESULTS: Our series of cases were classified according to the 2006 Nosology and Classification of Genetic Skeletal Disorders. The overall frequency of genetic skeletal disorders was 1:48 autopsies. The FGFR3 group and osteogenesis imperfecta type 2 were the more frequently encountered disorders. The mean gestational age at autopsy was 21.9 weeks (range, 12–37 weeks). A final diagnosis was obtained in 95% of cases. Genetic skeletal disorders were detected by prenatal ultrasound in 90% of cases, with a correct typing of the disorder achieved in only 34%. Molecular analysis was confirmative in 5 cases. CONCLUSIONS: The central role of the perinatal pathologist in collaboration with specialized services is essential for the correct interpretation of the radiologic, physical, and histopathologic findings, to accurately classify specific types of genetic skeletal disorders and enable genetic counseling. Birth Defects Research (Part A), 2009. © 2009 Wiley‐Liss, Inc.     

Effects of the new adiponectin paralogous protein CTRP-3 and of LPS on cytokine release from monocytes of patients with type 2 diabetes mellitus

Aims/Hypothesis: It was the aim to investigate the hypothesis that the new C1q/TNF-family member CTRP-3 (C1q/TNF-related protein-3) acts anti-inflammatory in human monocytes from healthy controls and patients with type 2 diabetes mellitus (T2D). Methods: Monocytes were isolated from 20 healthy controls and 30 patients with T2D. IL-6 and TNF concentrations were measured by ELISA. CTRP-3 was expressed in insect cells and used for stimulation experiments. Results: Basal IL-6 and TNF were not different in control and in T2D monocytes. LPS-stimulation (1 μg/ml) significantly (p < 0.001) increased IL-6 and TNF in the supernatants of control and in T2D monocytes to a similar extent. CTRP-3 (1 μg/ml) significantly (p = 0.03) inhibited LPS-induced IL-6 in control monocytes but not in T2D monocytes. TNF upon co-stimulation with LPS and CTRP-3 was significantly (p = 0.012) lower in control than in T2D monocytes. LPS-induced TNF concentration was significantly and positively correlated with serum total cholesterol and LDL cholesterol in T2D patients. Conclusions: CTRP-3 inhibits LPS-induced IL-6 and TNF release. This anti-inflammatory effect is lost in T2D. Serum cholesterol concentration affects the pro-inflammatory potential of LPS to induce TNF release from T2D monocytes in the presence or absence of CTRP-3. CTRP-3 might partly account for the pro-inflammatory state in T2D.     

E-LDL upregulates TOSO expression and enhances the survival of human macrophages

Uptake of modified lipoproteins by macrophages causes foam cell formation and promotes atherosclerosis. Atherogenic lipoproteins are cytotoxic and induce cell death under certain conditions but may also enhance macrophage survival. Macrophages treated with enzymatically modified LDL (E-LDL) were subjected to GeneChip analysis and the antiapoptotic gene TOSO was found induced. TOSO mRNA is upregulated and apoptosis is reduced in E-LDL but not in oxidized LDL (Ox-LDL) loaded macrophages. FLIP(L) abundance was suggested to mediate the antiapoptotic properties of TOSO; however, FLIP(L) was not changed. Ox-LDL is internalized predominantly by scavenger receptors such as CD36 while E-LDL particles are preferentially internalized by Fc- and complement-receptor dependent phagocytosis and internalization of phagobeads by macrophages upregulates TOSO. In COS-7 cells however, phagocytotic activity was not affected by TOSO. These data indicate that E-LDL-generated foam cells are protected from cell death most likely through the expression of TOSO by a FLIP(L) independent mechanism.