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F ree radicals, and the counteraction of COX-2/Nrf2/ARE(s) in lung cancer (A tentative study with a lcohol /acetaldehy

F ree radicals, and the counteraction of COX-2/Nrf2/ARE(s) in lung cancer (A tentative study with a lcohol /acetaldehyde in A549 cells). Cheng Luo , Ph. D University of Tartu. COX-2/Nrf2/ARE pathway. EFOX molecules by LC-MS.

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F ree radicals, and the counteraction of COX-2/Nrf2/ARE(s) in lung cancer (A tentative study with a lcohol /acetaldehy

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  1. Free radicals, and the counteraction of COX-2/Nrf2/ARE(s) in lung cancer(A tentative study with alcohol /acetaldehyde in A549 cells) Cheng Luo, Ph. D University of Tartu

  2. COX-2/Nrf2/ARE pathway

  3. EFOX molecules by LC-MS Thanks to modern LC (HPLC)-MS, so EFOX molecules can be discovered!!! LC-MS is able to detect the trace amount of small molecules in a very short time. LCMS-2020 Mass Spectrometer Systems Delivering the ultimate in measurement speed and sensitivity, the compact LCMS-2020 mass spectrometer offers faster measurements and higher detection sensitivity for quicker and more accurate detection of trace impurities, environmental pollutants, and other contaminants. When combined with a Prominence ultra-fast LC (UFLC/UFLCXR), you’ll achieve the ultimate in separation performance and productivity. http://en.wikipedia.org/wiki/Mass_spectrometry

  4. Tp-1 peptide:  15d-PGJ2 modification site is in the sequence on Cys-35 but not Cys-32.  Takahiro Shibata et al  Thioredoxin as a Molecular Target of Cyclopentenone Prostaglandins, JBC 2003

  5. PGJ2 Electrophilic molecules are endogenously generated and are causally involved in many pathophysiological effects. Prostaglandin D2 (PGD2), a major cyclooxygenase product in a variety of tissues, readily undergoes dehydration to yield the cyclopentenone-type PGs of the J2-series such as 15-deoxy-Δ12,14-PGJ2 (15d-PGJ2). 15d-PGJ2 is an electrophile, which can covalently react via the Michael addition reaction with nucleophiles, such as the free sulfhydryls of glutathione and cysteine residues in cellular proteins that play an important role in the control of the redox cell-signaling pathways. Covalent binding of 15d-PGJ2 to cellular proteins may be one of the mechanisms by which 15d-PGJ2 induces a cellular response involved in most of the pathophysiological effects associated with inflammation. In the present perspective, we provide a comprehensive summary of 15d-PGJ2 as an electrophilic mediator of cellular responses.

  6. PGD2  apoptosis

  7. Antioxidant-apoptosis of A549

  8. More sophisticated approaches to personal medicine

  9. Cell Biology International (2011) Immediate Publication, doi:10.1042/CBI20100707 Induction of apoptosis in non-small cell lung carcinoma A549 cells by PGD2 metabolite, 15d-PGJ2 Jun Jie Wang and Oi Tong Mak, paulmak@ukn.edu.tw

  10. The free-radical theory of aging (FRTA) states that organisms age because cells accumulate free radical damage over time. A free radical is any atom or molecule that has a single unpaired electron in an outer shell. 

  11. What are free radicals?

  12. Free Radicals Radicals (often referred to as free radicals) are atoms, molecules, or ions with unpaired electrons on an open shell configuration. Free radicals may have positive, negative, or zero charge. With some exceptions, the unpaired electrons cause radicals to be highly chemically reactive. Radicals, if allowed to run free in the body, are believed to be involved in degenerative diseases, senescence and cancers. UV Cl2 Cl + Cl 2Cl·→ Cl2

  13. Internal oxidation and antioxidation

  14. Alcohol metabolism http://hamsnetwork.org/metabolism/

  15. AldehydeDehydrogenase (ALDH2) Deficiency

  16. Alcohol - Cancer - Nrf2/ARE

  17. ADH and ALDH

  18. ADH and ALDH (gene network)

  19. Ethanol induce oxidative DNA damage in vitro by comet assay DNA damage by Co60 (or by acetaldehyde, or by UV)

  20. Detection of DNA damaging in A549 cells by comet assay (a) Representative micrograph of fluorescent DNA stain of control cells, showing undamaged and supercoiled DNA remaining within the nuclear cell membrane. (b) Representative micrograph of fluorescent DNA stain of PA|[plus]|DMTU-exposed cells, showing mild degree of denatured DNA fragments migrating out from cell. (c) Representative micrograph of fluorescent DNA stain of PA-exposed cells, showing denatured DNA fragments migrating out from cell in a long comet tail. (d) Quantification of podocytes with long comet tails. Following exposure to PA, at 2, 6, and 24h, there was a statistically significant increase in the number of cells with long comet tails compared with time-matched controls. DMTU significantly lessened PA-induced DNA damage at time points 2 and 6|[thinsp]|h. |[ast]|P<0.05, for PA versus PA|[plus]|DMTU. DNA breaker: puromycinaminonucleoside  (PA), DMTU: a radical scavenger

  21. Significant molecular networks of modulated miRNAs affected by formaldehyde exposure.

  22. Most alcohol diseases are associated to DNA damage, or DNA-protein crosslink FrédéricLangevin, et al Fancd2 counteracts the toxic effects of naturally produced aldehydes in mice Nature 475, 53–58 (07 July 2011)  Reactive aldehydes are common carcinogens. They are also by-products of several metabolic pathways and, without enzymatic catabolism, may accumulate and cause DNA damage. Ethanol, which is metabolised to acetaldehyde, is both carcinogenic and teratogenic in humans. Here we find that the Fanconianaemia DNA repair pathway counteracts acetaldehyde-induced genotoxicity in mice. Our results show that the acetaldehyde-catabolising enzyme Aldh2 is essential for the development of Fancd2−/−embryos. Nevertheless, acetaldehyde-catabolism-competent mothers (Aldh2+/−) can support the development of double-mutant (Aldh2−/−Fancd2−/−) mice. However, these embryos are unusually sensitive to ethanol exposureinutero, and ethanol consumption by postnatal double-deficient mice rapidly precipitates bone marrow failure. Lastly, Aldh2−/−Fancd2−/− mice spontaneously develop acute leukaemia. Acetaldehyde-mediated DNA damage may critically contribute to the genesis of fetal alcohol syndrome in fetuses, as well as to abnormal development, haematopoietic failure and cancer predisposition in Fanconianaemia patients.

  23. Status of KEAP1 and NRF2 Is Altered in Cancer Cells (A) Immunoblot showing increased nuclear localization of NRF2 in nuclear extracts (NE) from cancer cells. Cancer cells showed lower levels of KEAP1 (~69 kDa) and higher levels of NRF2 (~110 kDa) in total protein lysates (TP). NIVT and KIVT indicate NRF2 and KEAP1 in vitro transcribed/translated product, respectively. (B and C) Quantification of NRF2 and KEAP1 protein in immunoblots. For band densitometry, bands in nuclear extract blot (B) were normalized to Lamin B1, and those in total protein (C) were normalized to GAPDH. (D) Heat map showing relative expression of KEAP1, NRF2, and NRF2-dependent genes by real-time RT-PCR. Raw data for the heat maps are presented in Table S

  24. KEAP1 Sequence Alterations in Lung-Tumor-Derived Cell Lines and Patients

  25. Depletion of alcohol (acetaldehyde) L-cysteine + acetaldehyde -- 2-methylthiaszolidine-4-carboxylic acid (L-MTCA) (L-MTCA), which is composed of L-Cysteine (one half) and ACETALDEHYDE (one half).1968-1970, L-MTCA was used as a 'cough inhibitor'.

  26. First medicine to combat Carcinogenic acetaldehyde

  27. Jogurteista ja lasten hedelmäsoseista löytyi korkeita pitoisuuksia asetaldehydiä • Turun Sanomat 7.2 2010 05:00:11 – Tutkin nykyään laboratoriossa, mitä lastenlapseni syövät. Viili on turvallisempi kuin jogurtit, professori Mikko Salapuro sanoo. ProfessoriMikko Salaspuroon huolissaan suomalaisten elintarvikkeiden asetaldehydipitoisuuksista. Maailman terveysjärjestön WHO:n alainen kansainvälinen syöväntutkimuslaitos luokitteli asetaldehydin viime lokakuussa ihmisille syöpää aiheuttavaksi aineeksi alkoholin juomisen yhteydessä. Se nousi ylimpään riskiluokkaan, johon kuuluvat myös tupakka ja asbesti. Suun ja suoliston bakteerit ja hiivat muuttavat alkoholin syöpää aiheuttavaksi asetaldehydiksi. Ainetta on myös alkoholijuomissa ja tupakassa. Lisäksi asetaldehydiä löytyy useista tavallisista elintarvikkeista. Sitä muodostuu käymisellä valmistettuihin tuotteisiin, kuten maitovalmisteisiin, soijaruokiin, säilöttyihin vihanneksiin, etikkaan, kotikaljaan ja simaan. Asetaldehydiä käytetään myös elintarvikkeiden aromiaineena. Ainetta lisätään esimerkiksi hedelmämehuihin, virvoitusjuomiin, jälkiruokiin ja maitovalmisteisiin. Pitoisuuksia ei tarvitse ilmoittaa tuoteselosteessa. Asetaldehydiä muodostuu myös sisäsyntyisesti joihinkin hedelmiin, kuten omenoihin, päärynöihin ja marjoihin. Sitä käytetään myös marjojen ja hedelmien säilöntäaineena. Mikko Salapuron työryhmä on tutkinut laboratoriossa Suomessa myytävien elintarvikkeiden asetaldehydipitoisuuksia, koska niistä on toistaiseksi hyvin vähän tietoa. Testien mukaan jogurttien keskimääräinen asetaldehydipitoisuus ylittää riskirajan kahdeksankertaisesti. Vastaavia pitoisuuksia Salaspuro on mitannut useista hedelmämehuista ja soijakastikkeista. – Jopa vauvojen hedelmäsoseista löytyy samoja pitoisuuksia kuin jogurteista. Pitoisuus ylittää aikuisillekin sallitun rajan. Lapsilla painoon suhteutettu ylitys voi olla jopa 70-kertainen, Salaspuro kertoo. Lisää aiheesta sunnuntain Turun Sanomissa.

  28. Ville J. Salaspuro, Jaana M. Hietala, Martti L. Marvola, Mikko P. Salaspuro, Eliminating Carcinogenic Acetaldehyde By Cysteine From Saliva During Smoking. Cancer Epidemiol Biomarkers Prev January 2006 15; 146

  29. Ascorbic acid, L-cysteine, VB1, L-MTCA, etc.

  30. Ethanol is firstoxidised to ethanal then ethanoic acid. Note that while different enzymes can be used in the first step, only aldehyde (ethanal) dehydrogenase (ALDH) is used in the second

  31. Alcohol influence miRNA regulation through aldehyde CT1 CT2 CT3 FA1 FA2 FA3 Heat map of 89 formaldehyde-modulated miRNAs. Data were mean standardized, and hierarchical clustering was performed. Blue indicates relative low expression, and red indicates relative high expression. Abbreviations: FA, formaldehyde-treated samples; CT, control samples. Julia E. Rager, ET AL Epigenetic Changes Induced by Air Toxics: Formaldehyde Exposure Alters miRNA Expression Profiles in Human Lung Cells volume 119 | number 4 | April 2011 • Environmental Health Perspectives

  32. Keap1 Schematic of the Conserved Domain Structure of Human KEAP1 Protein The protein consists of an N-terminal region (amino acids 1–60), a BTB domain (amino acids 61–179), a central IVR (amino acids 180–314), a double-glycine-rich region comprising six Kelch motifs (amino acids 315–359, 361–410, 412–457, 459–504, 506–551, and 553–598), and a C-terminal domain (amino acids 599–624). The frequency of mutations detected within each domain is indicated below. Amino acid positions of the mutations are listed in 

  33. PCR

  34. No difference in age, COX-1, COX-2 COX-2

  35. Nrf2 (NFE2L2) Nrf2

  36. Keap1-COX-2

  37. KEAP1-oncogenes

  38. NEF2L2

  39. Cell respiration, C-metabolism, N

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