Salicylates Ameliorate Intestinal Inflammation by Activating Macrophage AMPK.

BACKGROUND: Inflammatory bowel diseases are the most common chronic intestinal inflammatory conditions, and their incidence has shown a dramatic increase in recent decades. Limited efficacy and questionable safety profiles with existing therapies suggest the need for better targeting of therapeutic strategies. Adenosine monophosphate-activated protein kinase (AMPK) is a key regulator of cellular metabolism and has been implicated in intestinal inflammation. Macrophages execute an important role in the generation of intestinal inflammation. Impaired AMPK in macrophages has been shown to be associated with higher production of proinflammatory cytokines; however, the role of macrophage AMPK in intestinal inflammation and the mechanism by which it regulates inflammation remain to be determined. In this study, we investigated the role of AMPK with a specific focus on macrophages in the pathogenesis of intestinal inflammation. METHODS: A dextran sodium sulfate-induced colitis model was used to assess the disease activity index, histological scores, macroscopic scores, and myeloperoxidase level. Proinflammatory cytokines such as tumor necrosis factor-α, interleukin-6, and interleukin-1ß were measured by enzyme-linked immunosorbent assay. Transient transfection of AMPKß1 and LC3-II siRNA in RAW 264.7 cells was performed to elucidate the regulation of autophagy by AMPK. The expression of p-AMPK, AMPK, and autophagy markers (eg, LC3-II, p62, Beclin-1, and Atg-12) was analyzed by Western blot. RESULTS: Genetic deletion of AMPKß1 in macrophages upregulated the production of proinflammatory cytokines, aggravated the severity of dextran sodium sulfate-induced colitis in mice, which was associated with an increased nuclear translocation of nuclear factor-κB, and impaired autophagy both in vitro and in vivo. Notably, the commonly used anti-inflammatory 5-aminosalicylic acid (ie, mesalazine) and sodium salicylate ameliorated dextran sodium sulfate-induced colitis through the activation of macrophage AMPK targeting the ß1 subunit. CONCLUSIONS: Together, these data suggest that the development of therapeutic agents targeting AMPKß1 may be effective in the treatment of intestinal inflammatory conditions including inflammatory bowel disease.

Novel Sequence Features of DNA Repair Genes/Proteins from Deinococcus Species Implicated in Protection from Oxidatively Generated Damage.

Deinococcus species display a high degree of resistance to radiation and desiccation due to their ability to protect critical proteome from oxidatively generated damage; however, the underlying mechanisms are not understood. Comparative analysis of DNA repair proteins reported here has identified 22 conserved signature indels (CSIs) in the proteins UvrA1, UvrC, UvrD, UvsE, MutY, MutM, Nth, RecA, RecD, RecG, RecQ, RecR, RuvC, RadA, PolA, DnaE, LigA, GyrA and GyrB, that are uniquely shared by all/most Deinococcus homologs. Of these CSIs, a 30 amino acid surface-exposed insert in the Deinococcus UvrA1, which distinguishes it from all other UvrA homologs, is of much interest. The uvrA1 gene in Deinococcus also exhibits specific genetic linkage (predicted operonic arrangement) to genes for three other proteins including a novel Deinococcus-specific transmembrane protein (designated dCSP-1) and the proteins DsbA and DsbB, playing central roles in protein disulfide bond formation by oxidation-reduction of CXXC (C represents cysteine, X any other amino acid) motifs. The CXXC motifs provide important targets for oxidation damage and they are present in many DNA repair proteins including five in UvrA, which are part of Zinc-finger elements. A conserved insert specific for Deinococcus is also present in the DsbA protein. Additionally, the uvsE gene in Deinococcus also shows specific linkage to the gene for a membrane-associated protein. To account for these novel observations, a model is proposed where specific interaction of the Deinococcus UvrA1 protein with membrane-bound dCSP-1 enables the UvrA1 to receive electrons from DsbA-DsbB oxido-reductase machinery to ameliorate oxidation damage in the UvrA1 protein.

Comparative analysis of differential network modularity in tissue specific normal and cancer protein interaction networks.

BACKGROUND: Large scale understanding of complex and dynamic alterations in cellular and subcellular levels during cancer in contrast to normal condition has facilitated the emergence of sophisticated systemic approaches like network biology in recent times. As most biological networks show modular properties, the analysis of differential modularity between normal and cancer protein interaction networks can be a good way to understand cancer more significantly. Two aspects of biological network modularity e.g. detection of molecular complexes (potential modules or clusters) and identification of crucial nodes forming the overlapping modules have been considered in this regard. METHODS: In the current study, the computational analysis of previously published protein interaction networks (PINs) has been conducted to identify the molecular complexes and crucial nodes of the networks. Protein molecules involved in ten major cancer signal transduction pathways were used to construct the networks based on expression data of five tissues e.g. bone, breast, colon, kidney and liver in both normal and cancer conditions. MCODE (molecular complex detection) and ModuLand methods have been used to identify the molecular complexes and crucial nodes of the networks respectively. RESULTS: In case of all tissues, cancer PINs show higher level of clustering (formation of molecular complexes) than the normal ones. In contrast, lower level modular overlapping is found in cancer PINs than the normal ones. Thus a proposition can be made regarding the formation of some giant nodes in the cancer networks with very high degree and resulting in reduced overlapping among the network modules though the predicted molecular complex numbers are higher in cancer conditions. CONCLUSION: The study predicts some major molecular complexes that might act as the important regulators in cancer progression. The crucial nodes identified in this study can be potential drug targets to combat cancer.

Effect of using silorane-based resin composite for restoring conservative cavities on the changes in cuspal deflection.

OBJECTIVE: To investigate the effect of using two resin-composite materials for restoring conservative mesio-occluso-distal (MOD) cavities on the changes (incremental and cumulative) in cuspal deflection. METHODS: Forty extracted sound human maxillary second premolars were subjected to standardized MOD cavity preparation and then divided into two groups (n=20). The first group of teeth was restored with Filtek Z250 (3M ESPE, St Paul, MN, USA), and Filtek P90 (3M ESPE, St Paul, MN, USA) was used in the second group. Incremental cuspal deflection was calculated by measuring the intercuspal distance between the indexed cusp tips before the restoration and at five-minute intervals up to 30 minutes using a stereomicroscope connected to a digital camera. Cumulative cuspal deflection for both materials was also calculated. RESULTS: Comparing the incremental cuspal deflection of the tested groups at each time interval, it was found that there was no significant difference immediately after curing and at five, 15, 20, and 25 minutes. However, a significant difference was recorded at 10 and 30 minutes. For the cumulative cuspal deflection, Filtek P90 showed significantly lower deflection values than Filtek Z250 only after five minutes. CONCLUSIONS: Incremental cuspal deflections of both materials over the tested intervals were almost comparable. However, after five minutes of curing, silorane-based resin composite surpassed the methacrylate-based resin composite in controlling the cumulative cuspal deflection.

Cervical cancer screening among Sudanese women.

BACKGROUND: Worldwide, cervical cancer is one of the three most common female cancers and accounts for over 370,000 new cases each year ( nearly 10% of all cancers). In Sudan, invasive cervical cancers is a leading cause of cancer death among women. METHODS: We conducted a community-based survey of Sudanese women living in Khartoum, from 2003 to 2008. Indicators of cervical cancer screening participation were examined : at least one previous Pap smear and Pap testing in the last 1 year. In Khartoum more than 30% of the ethnic Sudanese women live in the central and northern parts of the city. RESULTS: The overall estimated response rate was 32%, and the cooperation rate was 42%. Our study sample for this analysis included 256 women. Nearly one-half (35%) of the respondents had never had a Pap test, and only 65% been screened recently. Factors independently associated with cervical cancer screening use included marital status, housing type, and age. CONCLUSION: Our findings confirm low levels of cervical cancer screening among Sudanese women. Culturally and linguistically appropriate Pap testing intervention programs for less acculturated Sudanese women should be developed, implemented, and evaluated.

Salsalate intoxication and ramifications of utilizing nonspecific analytical methods in estimating quantitative blood concentrations.

In a case involving poisoning with salsalate, a salicylate analogue, the initial blood concentration of the parent drug was underestimated because of the low cross-reactivity of the immunoassay used for the analysis. This resulted in unnecessary additional clinical and laboratory evaluations to find the cause of metabolic acidosis exhibited by the patient. Additional findings led to the conclusion that parent salsalate contributed to patient toxicity. For testing purposes, all salicylate compounds should not necessarily be classified into one salicylic acid category, and there must be detailed communication between the caregiver and the laboratory regarding the specific drug history.

Liquid-chromatographic monitoring of cytosine arabinoside and its metabolite, uracil arabinoside, in serum.

This is a "high-performance" liquid-chromatographic method for quantifying the antileukemic drug cytosine arabinoside (cytarabine; 1-beta-D-arabinofuranosylcytosine; Ara-C), with a structural analog, 5-methylcytidine, as the internal standard. We used a C18 reversed-phase column and ammonium acetate (0.5 mol/L, pH 6.5) as the mobile phase, monitoring the column effluent at 280 nm. Tetrahydrouridine was present in the sample-collection tubes to inhibit conversion of cytosine arabinoside to uracil arabinoside. The standard curve is linear to 100 mg/L. Analytical recovery is 98%. Coefficients of variation for within-run and between-run imprecision were 2.0% and 4.3% at 20 mg/L and 2.7% and 2.7% at 80 mg/L, respectively. Assay sensitivity was limited by the amount of endogenous material in each patient's serum, making assay of a pre-infusion sample necessary for accurate calculations. In a trial patient population, the assay was shown to have potential for the detection of toxic concentrations in patients receiving high doses of Ara-C.

Comparison of Abbott TDx fluorescence polarization immunoassay, Sandoz radioimmunoassay, and high-performance liquid chromatography methods for the assay of serum cyclosporine.

We have had the opportunity to compare the new FPIA method for the measurement of serum Cs to established assays. The technique used a precipitation step prior to the fluorescence polarization measurement. We compared serum HPLC and RIA to the FPIA procedure. The within run coefficients of variation were 7.2%, 9.5%, and 4%, respectively. Between run CVs were 8.0%, 12.2%, and 3.8%. The correlation coefficient for HPLC and both of the immunoassays was less than 70%, showing the influence of the different specificities of the techniques. Medical centers that have based patient care on the HPLC assay that measures only parent drug will have difficulty using an immunoassay that measures a combination of parent and metabolites. There was a good correlation (R2 = 0.93) between the two immunoassays indicating that those currently using the serum RIA for monitoring could, through careful correlation studies in their patient population, use the FPIA technique. The regression equation was as follows: serum FPIA = 1.27 serum RIA + 1.9. This indicates the higher bias of the FPIA measurements. The advantages of the FPIA assay are that 20 assays could be done in less than one hour. This is in contrast to the longer turnaround time of the standard Sandoz RIA procedure. The technical competence required to perform the assay is less than that needed to perform the current RIA procedure. The assay can be recommended for replacement of the serum RIA; however, a correlation of levels with clinical experience is necessary in view of the difference in values between RIA and FPIA.

Pitfalls and errors in drug monitoring: analytical aspects.

Drug analyses, whether for purposes of evaluating new drug products or as part of clinical monitoring programs, have become frequently requested tests in both clinical and industrial laboratories. From the time of phlebotomy, throughout the transportation, processing and storage of the specimen, during the actual analytical procedure and throughout the clerical reporting of the data, certain potential errors and limitations in the system exist which should be fully recognized for proper evaluation of the laboratory data.

Liquid-chromatographic determination of amoxapine and 8-hydroxyamoxapine in human serum.

We describe a liquid-chromatographic procedure for amoxapine and 8-hydroxyamoxapine, its active metabolite, in serum. We used a mu-Bondapak C18 reversed-phase column and a mobile phase of acetonitrile/water (74/26 by vol) plus 26 microL of n-butylamine per liter. The compounds were measured at 254 nm, with 8-methoxyloxapine as internal standard. Necessary pre-analysis purification consisted of adsorbing the drug from serum onto extraction columns, eluting with 1-butanol/hexane (1/5 by vol), re-extracting into aqueous acid, and from that re-extracting again into the elution-solvent mixture. We prefer this procedure for monitoring both therapeutic and toxic concentrations of amoxapine, because parent drug and metabolite are measured separately.