In silico docking studies of Lupeol with MAPK pathway proteins- Raf-1, MEK & ERK.

OBJECTIVE: Lupeol, A triterpenoid found in variety of plants is reported to have beneficial medicinal effects on several ailments. Lupeol is also found to show inhibitory effect on proliferation of breast cancer cells. Metastasis is considered to be a major cause for worldwide deaths related to cancer. Ras related MAPK Signaling Pathway is one of the crucial pathways leading to metastasis. Lupeols binding possibility with Ras is already reported. In present study, Interaction between with downstream proteins of Ras- MAPK pathway, Raf ,MEK ,ERK1/2 and their corresponding domains are studied using STRING Database and their structures are retrieved in PDB Format. Lupeols binding affinity with downstream proteins of these signaling proteins at their interacting domains are analyzed. Here in silico docking approach to identify binding sites of each of these proteins with Lupeol is used. FDA approved standard drug molecule CH5126766 was used as reference ligand. Lupeol shows potent binding at significant sites with extremely high affinity. Since it binds with all the proteins involved in the pathway with high efficiency it is an important compound which can be developed as a therapeutic molecule.

Percutaneous nephrolithotomy in patients on chronic anticoagulant/antiplatelet therapy.

Percutaneous nephrolithotomy (PCNL) is an integral component in the management of large volume renal stone disease either as monotherapy or in combination with shock wave lithotripsy. Stone disease in patients on chronic anticoagulation/antiplatelet therapy, however, poses a difficult scenario. Bleeding is a major concern for any patient undergoing PCNL. We retrospectively analyzed our series of patients with renal calculi who were on chronic anticoagulant therapy and who underwent PCNL. We reviewed the case records of patients undergoing PCNL during the period from January 2005 to December 2011. We analyzed the changes in preoperative and postoperative hemoglobin, serum creatinine, and clotting parameters, as well as intraoperative and postoperative bleeding and thromboembolic complications. During the 5-year study period, a total of 36 patients (30 males and 6 females) with a mean age of 46.33±9.96 years (range, 29-61 years) who were on chronic anticoagulant/antiplatelet therapy underwent PCNL for urolithiasis. The mean size of the stone was 6.40±1.98 cm(2) (range, 2.8-9 cm(2)). The mean operating time was 62.08±10.10 min. The bleeding was successfully managed in all patients and the anticoagulant/antiplatelet agents were restarted after an appropriate duration. The mean rise in serum creatinine at discharge was 0.05±0.03 mg/dl and the mean fall in serum hemoglobin was 1.63±0.77 g/dl. At 3 months after surgery, the stone-free rate was 100%. With careful preoperative care and regulation of anticoagulation/antiplatelet therapy and appropriate intraoperative management, PCNL can be performed safely and successfully in properly selected patients with renal calculi who are on chronic anticoagulant/antiplatelet therapy.

Surgical outcomes of laparoscopic dismembered pyeloplasty in children with giant hydronephrosis secondary to ureteropelvic junction obstruction.

OBJECTIVE: Studies of surgical outcomes after reconstructive surgery for giant hydronephrosis (GH) secondary to ureteropelvic junction (UPJ) obstruction are limited. Over the past two decades, laparoscopic pyeloplasty has gradually replaced open repair in children in several centres. The objective of this study was to assess surgical outcomes of laparoscopic pyeloplasty in children with GH. MATERIALS AND METHODS: Children with unilateral primary UPJ obstruction and GH were prospectively included and underwent laparoscopic pyeloplasty. Postoperative ultrasonography was repeated at 3 and 12 months to assess renal parenchymal thickness, and similarly a renogram was repeated to assess improvement in differential renal function. RESULTS: During the study period 2005-2009, 53 children underwent laparoscopic dismembered pyeloplasty for UPJ obstruction. Of these, 8 children had GH caused by UPJ obstruction. The postoperative differential renal function improved in all of them. The postoperative improvement in renal parenchymal thickness at the end of 12 months was comparable to that of the non-GH group. CONCLUSIONS: At 12 months, surgical outcomes after laparoscopic pyeloplasty for GH were satisfactory. Relief of obstruction allows adequate and comparable nephron sparing.

First Report of Herbacious Hosts for Citrus yellow mosaic badna virus from India.

Citrus mosaic disease, a potential threat to citrus production throughout India, is currently an important disease in the southern and northeastern states (2). The reported incidence of the disease ranges from 10 to 77% (K. Gopal, G. S. Aparna, M. Sreenivasuluk, K. V. Subbaiah, and A. R. K. Rao, unpublished data). This yellow mosaic disease of citrus is caused by Citrus yellow mosaic badna virus (CMBV), formerly citrus yellow mosaic disease, (CYMD) (1). Host range studies were done to find herbaceous noncitrus host plant species for virus maintenance. The following are the noncitrus plants tested in this study: Arachis hypogaea, Chenopodium amaranticolor, Chenopodium quinoa, Vigna mungo, Macrotyloma uniflorum, Cicer arietinum, Helianthus annuus, Cajanus cajan, V. sinensis, Cyamopsis tetragonoloba, V. radiata, Pisum sativum, Phaseolus vulgaris, Trichosanthes anguina, Nicotiana tabacum (Harrison special), Dolichos lablab, Petunia × hybrida, Gomphrena globosa, Cucumis melo, Cucumis pepo, Glycine max, Sorghum bicolor, Zea mays, and Canna indica. Young leaves with mosaic symptoms were collected from Citrus sinensis Osbeck, Citrus aurantiifolia Osbeck, and Citrus × limonia Osbeck plants, which are being maintained in an insect-proof glasshouse. The leaves were cut into small pieces, transferred to a chilled mortar, and macerated using 0.01 M phosphate buffer, pH 7.0, containing 0.2% (v/v) of 2-mercapto-ethanol at a tissue/buffer ratio of 1 g/9 ml (wt/vol). The extract was filtered and used for inoculation. The above-mentioned noncitrus plants were uniformly dusted with 600-mesh Carborundum and inoculated with sap extract from the citrus species. The plants were kept in an insect-proof glasshouse and observed for 6 weeks for symptom development. Only three hosts, Canna indica, sorghum, and maize produced visible symptoms. Symptoms were observed 14 days postinoculation on C. indica as chlorotic spots, which later developed into a mosaic pattern. Developing young leaves showed severe mosaic with vein banding symptoms. In sorghum and maize, chlorotic streaks were observed on young leaves after 10 days, which developed into dark green streaks in the leaf lamina. All the inoculated hosts were checked using virus double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) and dot blot ELISA using CMBV polyclonal antiserum (Department of Virology, S.V. University, Tirupati, India). In both tests, only the C. indica, sorghum, and maize samples reacted positively. In dot blot ELISA, as little as 100 ng of virus could be detected in C. indica, sorghum, and maize. Virus from all three citrus sources produced the same symptoms on C. indica, sorghum, and maize. To our knowledge, this is the first report of herbaceous hosts of CMBV, which should prove useful as propagation and index hosts for CMBV. References: (1) Y. S. Ahlawat et al. Plant Dis. 80:590, 1996. (2) G. S. Reddy et al. Page 130 in: 3rd Int. Symp. Subtrop. Hortic. Bangalore, 1972.

Selective inhibition of cyclooxygenase-2 by C-phycocyanin, a biliprotein from Spirulina platensis.

We report data from two related assay systems (isolated enzyme assays and whole blood assays) that C-phycocyanin a biliprotein from Spirulina platensis is a selective inhibitor of cyclooxygenase-2 (COX-2) with a very low IC(50) COX-2/IC(50) COX-1 ratio (0.04). The extent of inhibition depends on the period of preincubation of phycocyanin with COX-2, but without any effect on the period of preincubation with COX-1. The IC(50) value obtained for the inhibition of COX-2 by phycocyanin is much lower (180 nM) as compared to those of celecoxib (255 nM) and rofecoxib (401 nM), the well-known selective COX-2 inhibitors. In the human whole blood assay, phycocyanin very efficiently inhibited COX-2 with an IC(50) value of 80 nM. Reduced phycocyanin and phycocyanobilin, the chromophore of phycocyanin are poor inhibitors of COX-2 without COX-2 selectivity. This suggests that apoprotein in phycocyanin plays a key role in the selective inhibition of COX-2. The present study points out that the hepatoprotective, anti-inflammatory, and anti-arthritic properties of phycocyanin reported in the literature may be due, in part, to its selective COX-2 inhibitory property, although its ability to efficiently scavenge free radicals and effectively inhibit lipid peroxidation may also be involved.

Differential formation of octadecadienoic acid and octadecatrienoic acid products in control and injured/infected potato tubers.

Lipoxygenases in plants have been implicated in the activation of defense responses against injury/infection. Pathogen-derived polyunsaturated fatty acids, such as arachidonic acid, eicosapentaenoic acid and their metabolites have been shown to elicit defense responses against pathogen infection in plants. However, not much is known about the role of host-derived fatty acids and their metabolites in plant defense responses. In this study, isolation and characterisation of endogenous lipoxygenase metabolites formed in potato tubers in response to injury/infection was undertaken. While 9-hydroperoxyoctadecadienoic acid (9-HPODE), derived from octadecdienoic acid (linoleic acid) is the major lipoxygenase product formed in control potato tubers, 9-hydroperoxyoctadecatrienoic acid (9-HPOTrE), derived from octadecatrienoic acid (alpha-linolenic acid) is the major lipoxygenase product formed in potato tubers in response to injury or infection with Rhizoctonia bataticola. As a result, the relative ratio of 9-HPODE to 9-HPOTrE showed a shift from 4:1 in control to 1:2 and 1:4.5 in injured and infected potato tubers respectively. From this study, it is proposed that lipoxygenase metabolites of octadecadienoic acid may be involved in physiological responses under control conditions, while octadecatrienoic acid metabolites are mediating the defense responses. This forms the first report on the differential formation of endogenous lipoxygenase products in potato tubers under control and stress conditions.

Effect of septic shock on nitrate, free amino acids, and urea in murine plasma and urine.

OBJECTIVES: Concentration changes of free amino acids, urea and nitrate in plasma and urine were studied for the murine model of septic shock. METHODS: After administration of a bolus dose of bacterial lipopolysaccharide (LPS), concentrations of amino acids and urea in plasma, and urea and nitrate in urine were determined. RESULTS: For individual amino acids four different trends were observed: (1) no change ( e.g., taurine, histidine, phenylalanine, hydroxproline); (2) continuous increase (e.g., aspartate and glutamate); (3) continuous decrease (e.g., threonine, serine, asparagine, proline, methionine, tyrosine); and (4) decrease during the first 4 hours, but return to normal at 8 hours after the LPS treatment (e.g., all the other amino acids). The ratio of phenylalanine to tyrosine was increased to about 2x. In plasma, urea concentration was increased about 3x, but in urine it decreased about 4x. Nitrate levels were increased 3x in urine. CONCLUSION: These early changes in the concentrations of amino acids as well as in the urea and nitrate may be useful as sensitive markers for the early and rapid diagnosis of septic shock.

Application of sodium metaperiodate for the determination of ribavirin in pharmaceutical formulations.

Three simple and sensitive visible spectrophotometric methods (A-C) have been described for the assay of ribavirin either in pure form or in pharmaceutical formulations. They are based on the oxidation of ribavirin with excess sodium metaperiodate and estimating either the products formed (dialdehyde with MBTH, method A; iodate with metol-sulphanilamide, in the presence of Mo(VI) and iodide, method B) or the amount of periodate consumed (celestine blue in the presence of telurium (IV), method C). All of the variables have been optimized and the reaction mechanisms presented. The concentration measurements are reproducible within a R.S.D. of 1.0%. Recoveries are 99.2-101.2%.

Spectrophotometric determination of azathioprine in pharmaceutical formulations.

Four simple and sensitive visible spectrophotometric methods (A-D) have been described for the assay of azathioprine (ATP) either in pure form or in pharmaceutical formulations. Methods A and B are based on the oxidation of ATP with excess N-bromosuccinimide (NBS) or chloramine-T (CAT) and determining the consumed NBS or CAT with a decrease in colour intensity of celestine blue (CB) (method A) or gallocyanine (GC) (method B), respectively. Methods C and D are based on the diazotisation of reduced azathioprine (RATP) with excess nitrous acid and estimating either the consumed nitrous acid (HNO(2)) with cresyl fast violet acetate (CFVA) (method C) or by coupling reaction of the diazonium salt formed with N-1-naphthyl ethylene diamine dihydrochloride (NED) (method D). All of the variables have been optimized and the reactions presented. The concentration measurements are reproducible within a relative standard deviation of 1.0%. Recoveries are 99.2-100.3%.

Determination of felbamate concentration in pediatric samples by high-performance liquid chromatography.

A simple and rapid procedure to determine felbamate (2-phenyl-1,3-propanediol dicarbamate) concentrations in human plasma/serum by high-performance liquid chromatography is described. The method employs a high-performance liquid chromatography unit equipped with a C18 reverse-phase cartridge (3-microliters particle diameter, 3.2 x 40 mm), an acetonitrile/water gradient, and detection at 210 nm. The sample is deproteinized with acetonitrile containing internal standard (2-methyl-2-phenyl-1,3-propanediol dicarbamate), and the resulting supernatant, after diluting 1:1 with water, is injected onto the column. The felbamate and internal standard are eluted with a linear gradient of 0% to 22% acetonitrile for 11 minutes at a flow rate of 0.8 ml/minute. Under these conditions, felbamate and the internal standard are eluted at 9.2 +/- 0.03 and 10.8 +/- 0.03 minutes, respectively. The assay is linear from 10 to 400 microgram/ml. It is highly reproducible; at 100 micrograms/ml felbamate, within-day and between-day coefficients of variation are less than 0.5% and 4.3%, respectively. Recovery is > or = 95%. No interferences from other common antiepileptic drugs and analgesics are observed. Advantages of this method include simple and fast sample preparation; use of a gradient to eliminate interferences; and use of a cartridge column, which is economical, provides good resolution, allows rapid equilibration and elution, and operates at low back pressures. The method requires samples of only 100 microliters and is ideal for pediatric samples.

Analysis of 24-hour ECG in patients with panic disorder.

Previous evidence suggests an increased cardiovascular morbidity in patients with panic disorder. In this study, we compared 24-hour ECG in patients with panic disorder (n = 22; age: 36.1 +/- 7.6 years) and healthy controls (n = 21; age: 34.6 +/- 10.0 years). The QTc intervals during the day or night were not significantly different between patients and controls. Ventricular ectopic beats were also not significantly different between the two groups. These results do not suggest any overt cardiac arrhythmias in this age group of patients with panic disorder.

Trends and effects of cigarette smoking among girls and women in the United States, 1965-1993.

Each year, cigarette smoking causes more than 140,000 deaths among women in the United States. Here, we describe smoking trends among girls and women, including women of reproductive age and pregnant women. We also provide data regarding the prevalence of indicators of nicotine dependence among women in the United States. The data were derived from the National Health Interview Survey, High School Seniors Survey, National Household Survey on Drug Abuse, and Teenage Attitudes and Practices Survey. The prevalence of smoking among women overall is now declining at a rate comparable to that of men, and women are attempting cessation and maintaining abstinence at the same rate as men. However, smoking prevalence among women in certain demographic groups such as American Indians and Alaska Natives is high. Although the prevalence of smoking increased among young women (particularly women of lower educational attainment) in the early 1980s, more recent surveys show it is declining. Smoking prevalence among young black and Hispanic women is decreasing, but progress in decreasing smoking prevalence among young white women is slow. Young women appear to be as nicotine dependent as older women, and light smokers of all ages report indicators of nicotine dependence.

Rapid quantitation of hemoglobin S by isoelectric focusing.

The feasibility of isoelectric focusing (IEF) to determine hemoglobin S (HbS) at a faster turn-around-time and to resolve the HbS and hemoglobin A (HbA) in presence of high concentrations of hemoglobin F (HbF) is evaluated. The IEF procedure is faster, and the results can be obtained in less than 45 minutes. The resulting data are comparable to gel electrophoresis. It is a superior procedure in resolving both HbS and HbA in the presence of high HbF and, therefore, a desirable technique to use for infants and children. Further, IEF is simpler than the gel electrophoresis, relatively inexpensive, easily adaptable for routine use, and suitable for "stat" conditions.

Isolation and sequence determination of a cDNA clone for rat peroxisomal urate oxidase: liver-specific expression in the rat.

Urate oxidase (UOxase; urate:oxygen oxidoreductase, EC 1.7.3.3), which catalyzes the oxidation of uric acid to allantoin, is present in most mammals but is absent in humans and certain primates. A cDNA clone for UOxase containing an insert of 1.3 kilobases (kb) was isolated from a lambda gt11 cDNA library prepared from rat liver mRNA. This recombinant clone with a 1283-nucleotide insert has sequence for 97% of the coding region together with 401 nucleotides of the 3'-untranslated region of the mRNA. The identity of UOxase cDNA clone was verified by analyzing the fusion protein, immunocytochemical localization with epitope-selected antibody, and hybrid-select translation analysis and by comparing sequences of four CNBr-cleaved peptides of the protein. Blot analysis revealed that the probe hybridizes to a single 1.5-kb mRNA species in the rat liver and a transplantable hepatocellular carcinoma. No UOxase mRNA was detected in 11 nonhepatic tissues of rat, suggesting tissue specificity of expression of this UOxase gene. Blot analysis of RNA from livers of rats treated with a peroxisome proliferator showed 2- to 3-fold increase in UOxase mRNA content, whereas the fatty acyl-CoA oxidase mRNA increased over 30-fold. Southern blot analysis of restriction enzyme digests of rat DNA suggests that there is a single copy of UOxase gene. Analysis of human genomic DNA revealed restriction fragments that are homologous to rat UOxase cDNA, although no UOxase mRNA was detected in human liver.

A simple method for the separation of triacylglycerols from fatty acids released in lipase assays.

A simple procedure for the partition of triacylglycerols from albumin-bound fatty acids is described. This procedure is based on the ability of fumed silicon dioxide to remove emulsified triacylglycerols from aqueous media. The method was developed for the assay of lipoprotein lipase activity but it may be used for the assay of other lipases.

A neoplasm with pancreatic and hepatocellular differentiation presenting with subcutaneous fat necrosis.

A neoplasm demonstrating both pancreatic and hepatic phenotypes is described. The tumor, from a 53-year-old woman with the syndrome of subcutaneous fat necrosis and arthropathy, was studied histologically, immunohistochemically, ultrastructurally, and biochemically. The clinical features of this case can be explained by the production of large amounts of lipase by the tumor. The hepatocellular properties of the tumor included characteristic morphology and the synthesis of catalase. The pancreatic properties of the tumor included the production of pancreatic lipase. This neoplasm would appear to be analogous to animal models in which the transdifferentiation of pancreatic acinar cells and hepatocytes has been demonstrated. Although the bulk of the tumor was present in the liver, the authors believe the tumor arose from the pancreas. The distinction between differentiation and site of origin of tumors is discussed.

Peroxisome proliferator-binding protein: identification and partial characterization of nafenopin-, clofibric acid-, and ciprofibrate-binding proteins from rat liver.

Peroxisome proliferators (PP) induce a highly predictable pleiotropic response in rat and mouse liver that is characterized by hepatomegaly, increase in peroxisome number in hepatocytes, and induction of certain peroxisomal enzymes. The PP-binding protein (PPbP) was purified from rat liver cytosol by a two-step procedure involving affinity chromatography and ion-exchange chromatography. Three PP, nafenopin and its structural analogs clofibric acid and ciprofibrate, were used as affinity ligands and eluting agents. This procedure yields a major protein with an apparent Mr of 70,000 on NaDodSO4/PAGE in the presence of reducing agent and Mr 140,000 (Mr 140,000-160,000) on gel filtration and polyacrylamide gradient gel electrophoresis under nondenaturing conditions, indicating that the active protein is a dimer. This protein has an acidic pI of 4.2 under nondenaturing conditions, which rises to 5.6 under denaturing conditions. The isolation of the same Mr 70,000 protein with three different, but structurally related, agents as affinity ligands and the immunological identity of the isolated proteins constitute strong evidence that this protein is the PPbP capable of recognizing PP that are structurally related to clofibrate. The PPbP probably plays an important role in the regulation of PP-induced pleiotropic response.

Purification, properties, and immunocytochemical localization of human liver peroxisomal enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase.

A molecular understanding of genetic disease in which peroxisomal functions are impaired depends on analysis of the structure of normal and mutant enzymes of peroxisomes. We report experiments describing the isolation, characterization, and immunocytochemical localization of enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase bifunctional enzyme (PBE) of the peroxisomal fatty acid beta-oxidation system from normal human liver and compared it with that of rat liver enzyme. The human enzyme, purified approximately equal to 2300-fold by ion-exchange chromatography, is homogeneous as judged by NaDodSO4/PAGE. This PBE is localized exclusively in the matrix of peroxisomes in liver cells by the protein A/gold immunocytochemical method. The human PBE is similar to rat enzyme in size (Mr, approximately equal to 79,000), isoelectric point (pI, 9.8), pH optima, molecular structure as observed by rotary shadowing, and peptide pattern on NaDodSO4/PAGE after proteolytic digestion with Staphylococcus aureus V8 protease. The human and rat enzymes differed in their immunological properties by having partial identity with each other; this is reflected in their slightly dissimilar composition of the amino acids aspartic acid, threonine, glutamic acid, tyrosine, and glycine. COOH-terminal amino acid were similar for both the enzymes: -Gly-Ser-Leu-Ile-COOH. These results suggest that the human and rat liver PBE may be different in their amino acid sequences at their antigenic sites.