1. ByCristianFranco CaasSara Correa Prez Plasma proteomic profile of sulfur mustard exposed lung diseases patients using 2D gel electrophoresis Molecular Biology2012 Hossein Mehrani1, Mostafa Ghanei, Jafar Aslani and Zahra Tabatabaei

2. General Information 3. Introduction Sulfur mustard is a chemical warfare that affect human health Chronic bronchitis, lung fibrosis, corneal ulcer disease Clinical manifestations in lung disorders due to sulfur mustard are different Mustard lung is not responsive to corticosteroids Proteomic can identify and quantify novel proteins in plasma 4. Introduction Biomarkers of the presence or severity of disease states Proteomics pattern in bronchoalveolar lavage (BAL) fluid of SM exposed patients Proteins whose expression is up or down regulated compared to healthy controls Looking new markers in the plasma, using ethanol and 2-dimensional gel Understand nature of long term effects of mustard gas 5. Proteomic

Proteins

Structures

Functions

6. Electrophoresis - 2D gel

Electrophoresis was defined as the movement of charged particles in an electric field.

Its a biochemical separation method analytical, useful for visualization and separation of protein preparations in small.

It has the advantage that helps the analyst to determine quickly how many proteins exist in a particular preparation or degree of purity of a specific protein preparation.

7. Electrophoresis - 2D gel

2-dimensional gel possess properties such as:

Transparency

Elasticity

Porosity

Controllable

Compatibility with a variety of chemical compounds that makes an excellent support means for electrophoretic separations.

Disponible en: http://protocolsonline.com/proteomics/two-dimensional-gel-electrophoresis/ Feb 2012

Electrophoresis allows the experimental determination of properties such as isoelectric point and molecular weight (MW) of a protein.

8. Sulfurd Mustard and Lung Diseases

Sulfur mustard (SM)

Chemical warfare agent.

Remains a threat to human health.

Some complications of mustard gas exposure are:

Chronic bronchitis

Interstitial lung diseases

Chronic obstructive lung disease

Lung fibrosis

Bronchiectasis

Bronchiolitis obliterans (BO).

Disponible en: http://noticias.lainformacion.com/ politica/armas-biologicas-y-quimicas/fuerzas [on line] Feb 2012 9. Objective The aim of this study was to identify protein expression signature or biomarkers that reflect chronic lung damages induced by SM exposure Disponible en: http://www.3plusplus.net/2010/11/how-to-objective-missions-in-5th.html [on -line] Feb 2012 10. Materiales y Mtodos 11. Pacientes

20 de sexo masculino con graves condiciones de enfermedad pulmonar

20 de sexo masculino sanos

Control

Inclusin

Exposicin, de SM

Dx, de enfermedad pulmonar crnica debido al gas mostaza.

Exclusin

Antecedente de una enfermedad crnica

Labectoma

Tratamiento con corticosteroides(2meses antes)

12. Recoleccin de muestras Tomado de: 1. http://www.gbo.com/es/index_1611.php 2.http://sobredeporteymas.blogspot.com/, 2. http://labgeminis.com/plantilla.php?id_sub_seccion=1513-4 http://www.auxilab.es/es/catalogo/vidrio_otros_tubos_Tubo-de-ensayo-borosilicato-20x200-mm.aspx5. http://www.roche-applied-science.com/sis/complete/index .jsp?id=com_0200006. http://laclasedelizpi.blogspot.com/2009/11/para-estudiar-el-clima-de-un-lugar-es.html [On-line] Febrero 2012 -80C 8-10am 4C->10 min HUPOCombinado de inhibicin de la proteasa 13. Preparacin de muestras 4 C Disponible en: 1. http://naturasaludonline.com/2. http://www.insbal.com/productos.php?categoria=Centrifugas 3. http://www.um.es/molecula/prot.htm 4.labotienda.com/es/ catalogo/espectrofotometria_espectrofotometros.aspx, 5. www.directindustry.es/prod/eppendorf/extremos-de-pipetas-22548-449221.htm6. http://www.gram.rs/ser/ etanol_70.htm [On-line] Febrero 2012 1 2 3 4 5 6 14. Preparacin de muestras 4 C Disponible en: 7. http://www.infofarma.es/suscripcion_rss/detalle/-/asset_publisher/Ta2g/content/distribucion-comoda-de-agua-ultrapura-para-laboratorios-de-investigacion /8. http://www.yarethquimicos.uuuq.com/agitador_vortex_gran_bio_pv-1.htm 9. http://doc20.comunidadcoomeva.com/blog/.html10.www.directindustry.es/prod/eppendorf/extremos-de-pipetas-22548-449221.htm 12. http://brumasenlanoche.blogspot.com/ On-line] Febrero 2012 7 8 9 10 11 12 15. SDS PageSeparacin las protenas de acuerdo a su movilidad electrofortica, dependiendo desu carga, forma y tamao. Electroforesisengeldepoliacrilamidacon dodecil-sulfato sdico Disponible en: http://en.wikipedia.org/wiki/SDS-PAGE [on-line] Feb 2012 16. ELECTROFORESISENGELDEPOLIACRILAMIDACON DODECILSULFATO SDICO

Poliacrilamida

Gel->bajo tamao molecular (30 min2-DEAnlisis del sobrenadantes y precipitado de etanol recogido. SDS Page

• MALDI-TOF

• Tcnica de ionizacin suave utilizada en espectrometra de masas.

• Anlisis debiomolculasfrgiles

Buffer [ ] Tris-HCl (pH 6,8),2% (w / v) de SDS Azul de bromofenol0,1% (w / v Glicerol 10% (v / v)Buffer [ ] Urea 7 M tiourea 1 MTris, (pH 7,5) 20 mM 18. Resultados 19. Figure 1 :SDS-PAGE de protenas plasmticas fraccionadas con diferentes concentraciones(v/v) de etanol.U,Protenas plasmticas antes de la precipitacin S, Sobrenadante de etanol P, Precipitado de etanol 20. Figure 2 Cada gel muestra control saludable;Precipitado de etanol, (B)Sobrenadante de etanol, (C) Sobrenadante de etanol, Pacientes con enfermedad pulmonar severa Puntosnumerados indicanlosMADITOFMS / MS deprotenas analizadas Patrones2-DEde protenas plasmticas de pacientes sanos y graves con enfermedades pulmonares expuestos a SM . 21.

• Each gel shows healthy control;

• Ethanol precipitate

• Ethanol supernatant

• (C) severe lung diseases patient; ethanol supernatant

• Numbered spots indicate the MADI TOF MS/MS analysed proteins as listed in Table 3.

Figure2 22. Discussion 23. Discussion Author Comment Agree Sapey E. andStockley RA. SerumAmyloid A1is an acute-phase protein that is induced, like CRP, by inflammatory mediators, including IL-6, IL-1b, and TNF-a, that rises in acute exacerbation of COPD. Yes Arredouani et al. Theydescribed the role ofhaptoglobinaffecting the immune system, showing that haptoglobin directly affects T-cells and suppress T-helper-cells through down-regulation ofcytokine production. Yes Mehrani H. et at. In a study of BAL fluid proteomics patterns in SM exposed patients, thehaptoglobinisoforms were significantly elevated in moderate and severe lung disease patients compared to mild and healthy controls Yes Nishiokaet al. The serum concentration ofhaptoglobinwas decreased during acute exacerbation in asthmatic children. No 24. Conclusions 25. Conclussion 26. Conclussion 27. Anexos 28. Plasma proteomic profile of sulfur mustard exposed lung diseases patients using 2D gel electrophoresis 29. Plasma proteomic profile of sulfur mustard exposed lung diseases patients using 2D gel electrophoresis 30. References 31.

• Mehrani H, Ghanei M, Aslani J, Tabatabaei Z.Plasma proteomic profile of sulfur mustard exposed lung diseases patients using 2-dimensional gel electrophoresis. Clinical Proteomics. 2011; 8 (2)

• Hernndez G. Mtodos de caracterizacin de protenas:Electroforesis en gel de poliacrilamid. Disponible en: http://www.medvet.una.ac.cr/carrera/mva505_Practica3. pdf.P.35-42

References 32. Thanks Disponible en: http://suu-construyendounafamilia.blogsp ot.com/2012_01 _01_archive.html [on-line] Feb 2012