SIRT1 redistribution on chromatin promotes genomic stability but alters gene expression during aging.

Genomic instability and alterations in gene expression are hallmarks of eukaryotic aging. The yeast histone deacetylase Sir2 silences transcription and stabilizes repetitive DNA, but during aging or in response to a DNA break, the Sir complex relocalizes to sites of genomic instability, resulting in the desilencing of genes that cause sterility, a characteristic of yeast aging. Using embryonic stem cells, we show that mammalian Sir2, SIRT1, represses repetitive DNA and a functionally diverse set of genes across the mouse genome. In response to DNA damage, SIRT1 dissociates from these loci and relocalizes to DNA breaks to promote repair, resulting in transcriptional changes that parallel those in the aging mouse brain. Increased SIRT1 expression promotes survival in a mouse model of genomic instability and suppresses age-dependent transcriptional changes. Thus, DNA damage-induced redistribution of SIRT1 and other chromatin-modifying proteins may be a conserved mechanism of aging in eukaryotes.

Blending organic material with municipal solid waste incinerator bottom ash to promote in-situ carbonation in road base.

The use of municipal solid waste incinerator bottom ash for road-base construction is an accepted practice in Europe and Asia, and of growing interest in the US. It is common practice to cure bottom ash by stockpiling it for several weeks before using it in this application. The curing process exposes the bottom ash to atmospheric carbon dioxide, which promotes carbonation, lowering its pH (making it less alkaline), and making many heavy metals less soluble. While this process makes bottom ash a more environmentally acceptable material, it takes time and requires additional handling. This article investigates a concept to facilitate carbonation of bottom ash in its compacted state, potentially eliminating the stockpile curing process. It is demonstrated here that blending a small amount of organic material with bottom ash will accelerate carbonation and lower pH in compacted samples by providing a carbon source for bacteria to produce carbon dioxide. Different quantities of biosolids (1%, 2%, 3%, and 5% by mass) were added to compacted bottom ash samples to examine the effect of organic materials on carbonation, and results were compared with a compacted control bottom ash sample. The pH of the control bottom ash sample decreased from 12.07 to 9.78 after 63 days, while the pH of the sample containing 5% biosolids decreased from 11.70 to 9.74 in only 7 days and to 8.18 after 63 days. Physical testing was conducted to examine suitability for beneficial use. The results indicate that bottom ash containing less than 3% biosolids met minimum bearing strength requirements for road base.

Establishment of a hepatocyte-kupffer cell coculture model for assessment of proinflammatory cytokine effects on metabolizing enzymes and drug transporters.

Elevated levels of proinflammatory cytokines associated with infection and inflammation can modulate cytochrome P450 enzymes, leading to potential disease-drug interactions and altered small-molecule drug disposition. We established a human-derived hepatocyte-Kupffer cell (Hep:KC) coculture model to assess the indirect cytokine impact on hepatocytes through stimulation of KC-mediated cytokine release and compared this model with hepatocytes alone. Characterization of Hep:KC cocultures showed an inflammation response after treatment with lipopolysaccharide and interleukin (IL)-6 (indicated by secretion of various cytokines). Additionally, IL-6 exposure upregulated acute-phase proteins (C-reactive protein, alpha-1-acid glycoprotein, and serum amyloid A2) and downregulated CYP3A4. Compared with hepatocytes alone, Hep:KC cocultures showed enhanced IL-1ß-mediated effects but less impact from both IL-2 and IL-23. Hep:KC cocultures treated with IL-1ß exhibited a higher release of proinflammatory cytokines, an increased upregulation of acute-phase proteins, and a larger extent of metabolic enzyme and transporter suppression. IC50 values for IL-1ß-mediated CYP3A4 suppression were lower in Hep:KC cocultures (98.0-144 pg/ml) compared with hepatocytes alone (IC50 > 5000 pg/ml). Cytochrome suppression was preventable by blocking IL-1ß interaction with IL-1R1 using an antagonist cytokine or an anti-IL-1ß antibody. Unlike IL-1ß, IL-6-mediated effects were comparable between hepatocyte monocultures and Hep:KC cocultures. IL-2 and IL-23 caused a negligible inflammation response and a minimal inhibition of CYP3A4. In both hepatocyte monocultures and Hep:KC cocultures, IL-2RB and IL-23R were undetectable, whereas IL-6R and IL-1R1 levels were higher in Hep:KC cocultures. In summary, compared with hepatocyte monocultures, the Hep:KC coculture system is a more robust in vitro model for studying the impact of proinflammatory cytokines on metabolic enzymes.

Bioactivation and toxicity of acetaminophen in a rat hepatocyte micropatterned coculture system.

We have recently shown that primary rat hepatocytes organized in micropatterned cocultures with murine embryonic fibroblasts (HepatoPac™) maintain high levels of liver functions for at least 4 weeks. In this study, rat HepatoPac was assessed for its utility to study chemical bioactivation and associated hepatocellular toxicity. Treatment of HepatoPac cultures with acetaminophen (APAP) over a range of concentrations (0-15 mM) was initiated at 1, 2, 3, or 4 weeks followed by the assessment of morphological and functional endpoints. Consistent and reproducible concentration-dependent effects on hepatocyte structure, viability, and basic functions were observed over the 4-week period, and were exacerbated by depleting glutathione using buthionine sulfoximine or inducing CYP3A using dexamethasone, presumably due to increased reactive metabolite-induced stress and adduct formation. In conclusion, the results from this study demonstrate that rat HepatoPac represents a structurally and functionally stable hepatic model system to assess the long-term effects of bioactivated compounds.

Exploring Chronic Drug Effects on Microengineered Human Liver Cultures Using Global Gene Expression Profiling.

Global gene expression profiling is useful for elucidating a drug's mechanism of action on the liver; however, such profiling in rats is not very sensitive for predicting human drug-induced liver injury, while dedifferentiated monolayers of primary human hepatocytes (PHHs) do not permit chronic drug treatment. In contrast, micropatterned cocultures (MPCCs) containing PHH colonies and 3T3-J2 fibroblasts maintain a stable liver phenotype for 4-6 weeks. Here, we used MPCCs to test the hypothesis that global gene expression patterns in stable PHHs can be used to distinguish clinical hepatotoxic drugs from their non-liver-toxic analogs and understand the mechanism of action prior to the onset of overt hepatotoxicity. We found that MPCCs treated with the clinical hepatotoxic/non-liver-toxic pair, troglitazone/rosiglitazone, at each drug's reported and non-toxic Cmax (maximum concentration in human plasma) for 1, 7, and 14 days displayed a total of 12, 269, and 628 differentially expressed genes, respectively, relative to the vehicle-treated control. Troglitazone modulated >75% of transcripts across pathways such as fatty acid and drug metabolism, oxidative stress, inflammatory response, and complement/coagulation cascades. Escalating rosiglitazone's dose to that of troglitazone's Cmax increased modulated transcripts relative to the lower dose; however, over half the identified transcripts were still exclusively modulated by troglitazone. Last, other hepatotoxins (nefazodone, ibufenac, and tolcapone) also induced a greater number of differentially expressed genes in MPCCs than their non-liver-toxic analogs (buspirone, ibuprofen, and entacapone) following 7 days of treatment. In conclusion, MPCCs allow evaluation of time- and dose-dependent gene expression patterns in PHHs treated chronically with analog drugs.

Construction material properties of slag from the high temperature arc gasification of municipal solid waste.

Slag from the high temperature arc gasification (HTAG) of municipal solid waste (MSW) was tested to evaluate its material properties with respect to use as a construction aggregate. These data were compared to previously compiled values for waste to energy bottom ash, the most commonly produced and beneficially used thermal treatment residue. The slag was tested using gradations representative of a base course and a course aggregate. Los Angeles (LA) abrasion testing demonstrated that the HTAG slag had a high resistance to fracture with a measured LA loss of 24%. Soundness testing indicated a low potential for reactivity and good weathering resistance with a mean soundness loss of 3.14%. The modified Proctor compaction testing found the slag to possess a maximum dry density (24.04kN/m(3)) greater than conventionally used aggregates and WTE BA. The LBR tests demonstrated a substantial bearing capacity (>200). Mineralogical analysis of the HTAG suggested the potential for self cementing character which supports the elevated LBR results. Preliminary material characterization of the HTAG slag establishes potential for beneficial use; larger and longer term studies focusing on the material's possibility for swelling and performance at the field scale level are needed.

MTAP Deletions in Cancer Create Vulnerability to Targeting of the MAT2A/PRMT5/RIOK1 Axis.

Homozygous deletions of p16/CDKN2A are prevalent in cancer, and these mutations commonly involve co-deletion of adjacent genes, including methylthioadenosine phosphorylase (MTAP). Here, we used shRNA screening and identified the metabolic enzyme, methionine adenosyltransferase II alpha (MAT2A), and the arginine methyltransferase, PRMT5, as vulnerable enzymes in cells with MTAP deletion. Metabolomic and biochemical studies revealed a mechanistic basis for this synthetic lethality. The MTAP substrate methylthioadenosine (MTA) accumulates upon MTAP loss. Biochemical profiling of a methyltransferase enzyme panel revealed that MTA is a potent and selective inhibitor of PRMT5. MTAP-deleted cells have reduced PRMT5 methylation activity and increased sensitivity to PRMT5 depletion. MAT2A produces the PRMT5 substrate S-adenosylmethionine (SAM), and MAT2A depletion reduces growth and PRMT5 methylation activity selectively in MTAP-deleted cells. Furthermore, this vulnerability extends to PRMT5 co-complex proteins such as RIOK1. Thus, the unique biochemical features of PRMT5 create an axis of targets vulnerable in CDKN2A/MTAP-deleted cancers.

Medicine and spirituality: a simple path to restore compassion in medicine.

Medical science has achieved impressive accomplishments in the diagnosis and treatment of human disease. However, the emphasis on science and technology has created a generation of physicians who find it difficult to relate to their patients about their suffering. Time constraints and economic pressures also add to the challenge of giving meaningful time to patients. Patients want to talk to their physician about their concerns, but surveys indicate that this is not being accomplished. Medical educators are developing curricula to teach how care can be given compassionately. This article reviews the importance of addressing spiritual care in medicine. Spirituality is defined and the spiritual history is explained. Research on the role of spirituality in health care is also reviewed. The role of the physician as a healer, attending to mind, body and spirit is encouraged.