Suppression of inflammation by low-dose methotrexate is mediated by adenosine A2A receptor but not A3 receptor activation in thioglycollate-induced peritonitis

Prior studies demonstrate that adenosine, acting at one or more of its receptors, mediates the anti-inflammatory effects of methotrexate in animal models of both acute and chronic inflammation. Both adenosine A_2A and A₃ receptors contribute to the anti-inflammatory effects of methotrexate treatment in the air pouch model of inflammation, and the regulation of inflammation by these two receptors differs at the cellular level. Because different factors may regulate inflammation at different sites we examined the effect of low-dose weekly methotrexate treatment (0.75 mg/kg/week) in a model of acute peritoneal inflammation in adenosine A_2A receptor knockout mice and A₃ receptor knockout mice and their wild-type littermates. Following intraperitoneal injection of thioglycollate there was no significant difference in the number or type of leukocytes, tumor necrosis factor alpha (TNF-α) and IL-10 levels that accumulated in the thioglycollate-induced peritoneal exudates in adenosine A_2A knockout mice or wild-type control mice. In contrast, there were more leukocytes, TNF-α and IL-10 in the exudates of the adenosine A₃ receptor-deficient mice. Low-dose, weekly methotrexate treatment increased the adenosine concentration in the peritoneal exudates of all mice studied, and reduced the leukocyte accumulation in the wild-type mice and A₃ receptor knockout mice but not in the A_2A receptor knockout mice. Methotrexate reduced exudate levels of TNF-α in the wild-type mice and A₃ receptor knockout mice but not the A_2A receptor knockout mice. More strikingly, IL-10, a critical regulator of peritoneal inflammation, was increased in the methotrexate-treated wild-type mice and A₃ knockout mice but decreased in the A_2A knockout mice. Dexamethasone, an agent that suppresses inflammation by a different mechanism, was similarly effective in wild-type mice, A_2A mice and A₃ knockout mice. These findings provide further evidence that adenosine is a potent regulator of inflammation that mediates the anti-inflammatory effects of methotrexate. Moreover, these data provide strong evidence that the anti-inflammatory effects of methotrexate and adenosine are mediated by different receptors in different inflammatory loci, an observation that may explain why inflammatory diseases of some organs but not of other organs respond to methotrexate therapy.     

Liver Clock Protein BMAL1 Promotes de Novo Lipogenesis through Insulin-mTORC2-AKT Signaling

The clock protein BMAL1 (brain and muscle Arnt-like protein 1) participates in circadian regulation of lipid metabolism, but its contribution to insulin AKT-regulated hepatic lipid synthesis is unclear. Here we used both Bmal1^−/− and acute liver-specific Bmal1-depleted mice to study the role of BMAL1 in refeeding-induced de novo lipogenesis in the liver. Both global deficiency and acute hepatic depletion of Bmal1 reduced lipogenic gene expression in the liver upon refeeding. Conversely, Bmal1 overexpression in mouse liver by adenovirus was sufficient to elevate the levels of mRNA of lipogenic enzymes. Bmal1^−/− primary mouse hepatocytes displayed decreased levels of de novo lipogenesis and lipogenic enzymes, supporting the notion that BMAL1 regulates lipid synthesis in hepatocytes in a cell-autonomous manner. Both refed mouse liver and insulin-treated primary mouse hepatocytes showed impaired AKT activation in the case of either Bmal1 deficiency or Bmal1 depletion by adenoviral shRNA. Restoring AKT activity by a constitutively active mutant of AKT nearly normalized de novo lipogenesis in Bmal1^−/− hepatocytes. Finally, Bmal1 deficiency or knockdown decreased the protein abundance of RICTOR, the key component of the mTORC2 complex, without affecting the gene expression of key factors of insulin signaling. Thus, our study uncovered a novel metabolic function of hepatic BMAL1 that promotes de novo lipogenesis via the insulin-mTORC2-AKT signaling during refeeding.     

Assessing weight-related quality of life in adolescents

Objective : The development of a new weight‐related measure to assess quality of life in adolescents [Impact of Weight on Quality of Life (IWQOL)‐Kids] is described. Research Methods and Procedures : Using a literature search, clinical experience, and consultation with pediatric clinicians, 73 items were developed, pilot tested, and administered to 642 participants, 11 to 19 years old, recruited from weight loss programs/studies and community samples (mean z‐BMI, 1.5; range, ?1.2 to 3.4; mean age, 14.0; 60% female; 56% white). Participants completed the 73 items and the Pediatric Quality of Life Inventory and were weighed and measured. Results : Four factors (27 items) were identified (physical comfort, body esteem, social life, and family relations), accounting for 71% of the variance. The IWQOL‐Kids demonstrated excellent psychometric properties. Internal consistency coefficients ranged from 0.88 to 0.95 for scales and equaled 0.96 for total score. Convergent validity was demonstrated with strong correlations between IWQOL‐Kids total score and the Pediatric Quality of Life Inventory (r = 0.76, p < 0.0001). Significant differences were found across BMI groups and between clinical and community samples, supporting the sensitivity of this measure. Participants in a weight loss camp demonstrated improved IWQOL‐Kids scores, suggesting responsiveness of the IWQOL‐Kids to weight loss/social support intervention. Discussion : The present study provides preliminary evidence regarding the psychometric properties of the IWQOL‐Kids, a weight‐related quality of life measure for adolescents. Given the rise of obesity in youth, the development of a reliable and valid weight‐related measure of quality of life is timely.     

Adherence to vitamin supplementation following adolescent bariatric surgery

Objective: Adolescents with extreme obesity, who have undergone bariatric surgery, must adhere to many lifestyle and nutritional recommendations, including multivitamin therapy. Little is known about multivitamin adherence following adolescent bariatric surgery. Design and Methods: The present study aims to document self‐reported and electronically‐monitored adherence to multivitamins, determine convergence between self‐report and electronic monitoring adherence for multivitamins, and identify barriers to multivitamin adherence for adolescents who have undergone bariatric surgery. Results: The study used a prospective, longitudinal observational design to assess subjective (self‐reported) and objective (electronic monitors) multivitamin adherence in a cohort of 41 adolescents (Mean age = 17.1 ± 1.5; range = 13‐19) who have undergone bariatric surgery at Cincinnati Children's Hospital Medical Center. Mean adherence as derived from electronic monitoring for the entire 6‐month study period was 29.8% ± 23.9. Self‐reported adherence was significantly higher than electronically monitored adherence across both the 1 and 6‐month assessment points (z = 4.5, P < 0.000 and z = 4.0, P < 0.0001, respectively). Forgetting and difficulty swallowing multivitamins were the two primary barriers identified. While there are no established data regarding best practice for multivitamins following bariatric surgery, high rates of nonadherence to multivitamin therapy were observed in adolescents who had undergone bariatric surgery with forgetting and difficulty swallowing pills as reported barriers to adherence. Conclusion: These high rates of nonadherence to multivitamin therapy should be considered when devising treatment and family education pathways for adolescents considering weight loss surgery.     

Geometrical features of lips using the properties of parabola for recognizing facial expression

Various real-time applications such as Human–Computer Interactions, Psychometric analysis, etc. use facial expressions as one of the important parameters. The researchers have used Action Units (AU) of the face as feature points and its deformation is compared with the reference points on the face to estimate the facial expressions. Among many parts of the face, features from the mouth contribute largely to all the well-known emotions. In this paper, the parabola theory is used to identify and mark various points on the lips. These points are considered as feature points to construct feature vectors. The Latus Rectum, Focal Point, Directrix, Vertex, etc. are also considered to identify the feature points of the lower lips and upper lips. The proposed approach is evaluated on benchmark datasets such as JAFFEE and Cohn–Kanade dataset and it is found that the performance is encouraging in understanding the facial expressions. The results are compared with contemporary methods and found that the proposed approach has given good classification accuracy in recognizing facial expressions.     

Nitric oxide synthase-2 (NOS2) gene polymorphism c.1832C>T (Ser608Leu) associated with nitrosative stress in Fanconi anaemia

Fanconi anemia (FA) occurs due to genomic instability with predisposition to bone marrow failure, phenotypic abnormalities and cancers. Though mutations in 22 genes leading to DNA repair defect have been identified, the cellular factor such as oxidative stress has also shown to be associated with FA. Nitrosative Stress (NS) is biochemically correlated to many oxidative stress related disorders and the NS as a pathological hallmark in FA has been so far overlooked. We carried out the study first time in Indian patients with FA with an objective to understand the role of NS in the pathogenesis of FA. The study was carried out in 70 FA subjects. The FA subjects were diagnosed by chromosomal breakage analysis. Molecular study was carried out by Next Generation Sequencing and Sanger sequencing. The 3-nitrotyrosine [3-NT] levels were estimated through enzyme-linked immuno-sorbent assay (ELISA) and the nitric oxide synthase genes- NOS1 (c.-420-34221G>A (rs1879417), c.-420-10205C>T (rs499776), c.4286+720G>C (rs81631)) and NOS2 (c.1823C>T (p. Ser608Leu) (rs2297518)) polymorphism were studied by direct sequencing. Chromosomal breakage analysis revealed a high frequency of chromosomal breaks (Mean chromosomal breakage—4.13?±?1.5 breaks/metaphase) in 70 FA patients as compared to the control. Molecular studies revealed FANCA (58.34%), FANCG (18.34%) and FANCL (16.6%) complementation groups. The 3-nitrotyrosine [3-NT] levels showed to be significantly (p?<?0.05) elevated in FA subjects when compared to the age match controls. Genotyping of the NOS2 gene c.1823C>T (p. Ser608Leu) (rs2297518), showed statistically significant (P?<?0.05) association with FA. Elevated level of 3-NT is one of the cause of NS and NOS2 gene polymorphism associated with FA is an important target in the treatment regimen.

     

Weight-specific health-related quality of life in adolescents with extreme obesity

The objectives of this multisite study were to: (i) examine differences by gender and race on generic and weight- specific health-related quality of life (HRQOL) in adolescents with extreme obesity (BMI > or = 40 kg/m(2)) and (ii) explore HRQOL differences based on treatment pursued (behavioral vs. bariatric surgery). Study participants included 145 obese adolescents (mean age = 15.3 years; 68% female; 46% black; mean BMI = 50.6) referred to pediatric weight management programs. Participants completed generic (PedsQL) and weight-specific (Impact of Weight on Quality of Life-Kids (IWQOL-Kids)) HRQOL measures. Generic and weight-specific measures indicated global (e.g., all domains) HRQOL impairment and significant differences by race. Physical, emotional, and social scores of the PedsQL (Ps < 0.01) and the physical comfort and body esteem scores of the IWQOL-Kids (Ps < 0.001) were significantly higher for black compared to white adolescents with extreme obesity. Extremely obese adolescents pursuing bariatric surgery reported similar HRQOL to adolescents pursuing behavioral treatment (n = 30 matched pairs). HRQOL did not differ for extremely obese adolescents based on type of treatment sought, but race/ethnicity should be considered when characterizing these youth. Although racial differences in adolescent body image/esteem have been reported, it is unknown why black adolescents with extreme obesity would report less impact of weight on their physical functioning. Overall, these data suggest that HRQOL is not homogenous in adolescents with extreme obesity.     

Expression of the neural stem cell markers NG2 and L1 in human angiomyolipoma: are angiomyolipomas neoplasms of stem cells?

Angiomyolipomas are benign tumors of the kidney which express phenotypes of smooth muscle, fat, and melanocytes. These tumors appear with increased frequency in the autosomal dominant disorder tuberous sclerosis and are the leading cause of morbidity in adults with tuberous sclerosis. While benign, these tumors are capable of provoking life threatening hemorrhage and replacement of the kidney parenchyma, resulting in renal failure. The histogenesis of these tumors is currently unclear, although currently, we believe these tumors arise from "perivascular epithelioid cells" of which no normal counterpart has been convincingly demonstrated. Recently, stem cell precursors have been recognized that can give rise to smooth muscle and melanocytes. These precursors have been shown to express the neural stem cell marker NG2 and L1. In order to determine whether angiomyolipomas, which exhibit smooth muscle and melanocytic phenotypes, express NG2 and L1, we performed immunocytochemistry on a cell line derived from a human angiomyolipoma, and found that these cells are uniformly positive. Immunohistochemistry of human angiomyolipoma specimens revealed uniform staining of tumor cells, while renal cell carcinomas revealed positivity only of angiogenic vessels. These results support a novel histogenesis of angiomyolipoma as a defect in differentiation of stem cell precursors.     

Process optimization and characterization of poloxamer solid dispersions of a poorly water-soluble drug

The objective of the present investigation was to improve the dissolution rate of Rofecoxib (RXB), a poorly water-soluble drug by solid dispersion technique using a water-soluble carrier, Poloxamer 188 (PXM). The melting method was used to prepare solid dispersions. A 3² full factorial design approach was used for optimization wherein the temperature to which the melt-drug mixture cooled (X ₁) and the drug-to-polymer ratio (X ₂) were selected as independent variables and the time required for 90% drug dissolution (t₉₀) was selected as the dependent variable. Multiple linear regression analysis revealed that for obtaining higher dissolution of RXB from PXM solid dispersions, a low level ofX ₁ and a high level ofX ₂ were suitable. The differential scanning calorimetry and x-ray diffraction studies demonstrated that enhanced dissolution of RXB from solid dispersion might be due to a decrease in the crystallinity of RXB and PXM and dissolution of RXB in molten PXM during solid dispersion preparation. In conclusion, dissolution enhancement of RXB was obtained by preparing its solid dispersions in PXM using melting technique. The use of a factorial design approach helped in identifying the critical factors in the preparation and formulation of solid dispersion. Published: April 13, 2007