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Miodrag Stojkovic

Miodrag Stojkovic. Nis, 20/03/2008. Struktura DNK. Nis, 20/03/2008. Dr. Watson. Nis, 20/03/2008. Organizacija genetskog materijala. Nis, 20/03/2008. Microarray chip. Nis, 20/03/2008. Molekularna karakterizacija. Nis, 20/03/2008. Diferencijacija. Nis, 20/03/2008. Terminologija.

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Miodrag Stojkovic

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  1. Miodrag Stojkovic Nis, 20/03/2008

  2. Struktura DNK Nis, 20/03/2008

  3. Dr. Watson Nis, 20/03/2008

  4. Organizacija genetskog materijala Nis, 20/03/2008

  5. Microarray chip Nis, 20/03/2008

  6. Molekularna karakterizacija Nis, 20/03/2008

  7. Diferencijacija Nis, 20/03/2008

  8. Terminologija • Oplodjena jajna celija je totipotentna, daje sve i razlicite tipove celija ljudskog organizma. • Pluripotentna maticna celijatakodje daje sve tipove celija organizma osim onih koje su potrebne da se razviju u kompletni fetus. • Maticna celija koja daje ne sve ali vise vrsta celija je multipotentna. Nis, 20/03/2008

  9. Istorija • 60-tih proslog veka, naucnici otkrivaju da kostana srz sardzi najmanje dva tipa maticnih celija: i) hematopoetske, stvaraju sve tipove krvnih celija ii) kostana srz (stromalne celije) koje daju kosti, hrskavicu, masno i fibrozno vezivno tkivo. • 90-tih naucnici prihvataju da i odrasli mozak sadrzi maticne celije koje imaju mogucnost da stvaraju tri celijska tipa: astrocite, oligodendrocite, i neurone. • Krv iz pupcane vrpce, celije iz placente, fetusa, tkiva I organa …adultne maticne celije • 1998 izolovane prve linije ljudskih embrionalnih maticnih celija Nis, 20/03/2008

  10. Kljucni izvori maticnih celija su ljudsko telo i rani embrioni Nis, 20/03/2008

  11. Adultne maticne celije Nis, 20/03/2008

  12. Rani ljudski razvoj Nis, 20/03/2008

  13. Preimplantaciona genetska diagnostika Nis, 20/03/2008

  14. Derivacija ljudskih embrionalnih maticnih celija Nis, 20/03/2008

  15. Embrioni koji su stali u razvoju… Nis, 20/03/2008

  16. …kao izvor maticnih celija Nis, 20/03/2008

  17. Karakterizacija ljudskih embrionalnih maticnih celija Nis, 20/03/2008

  18. Slicnosti i razlike izmedju embrionalnih i adultnih maticnih celija • Embrionalne su pluripotentne. • Adultne su multipotentne. • Veliki broh embrionalnih maticnih celija je lako cuvati pod in vitro uslovima. • Adultne su retke u zrelom tkivu a metod za njihovo ekspandiranje je ogranicen. • Adultne koje poticu od bolesnika mogu se vratiti istom bolesniku bez imunosupresije. • Embrionalne bi bile odbacene bez imunosupresije. • Imaju razlicite markere. Nis, 20/03/2008

  19. Osobine Sve maticne celije, bez obzira na njihovo poreklo imaju tri zajednicke osobine: • Dele se i daju maticne celije za vrlo dugi period (neke ne stare); • Nisu specijalizovane; • Daju specijalizovane celije. Nis, 20/03/2008

  20. Zasto nam maticne celije trebaju? • Studiranje razvojne biologije i poremecaja vezanih za nju • Otkrivanje novih polipeptida, faktora rasta i diferencijacija • Model za ljudske bolesti pod in vitro uslovima koristeci: • Otrivanje novih lekova i putem toksikologije • Celijska terapija • Inzenjering tkiva i organa • Kraj upotrebe imunosupresije kod transplantacije? Nis, 20/03/2008

  21. Mehanizmi koji drze misje i ljudske embrionalne maticne celije u pluripotentnom stanju Nis, 20/03/2008

  22. A B Normalised Ratio (Target/GAPDH) NANOG 1 0.8 hES-NCL1 hES-NCL1 H1 H1 NT2-SP12 NT2-SP12 0.6 0.4 0.2 0 C D B NT2-SP12 hES-NCL N N G G NANOG GAPDH Manipulacija gena putem RNK interferencije kod maticnih celija A EC ES Normalised Ratio (Target/GAPDH) NANOGP1 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 Nis, 20/03/2008

  23. RNK interferencija Nis, 20/03/2008

  24. RNK interferencija Nis, 20/03/2008

  25. NT2-SP12 hES-NCL1 N N G G GATA6 GAPDH hES-NCL1 hES-NCL1 hES-NCL1 hES-NCL1 hES-NCL1 H1 H1 H1 H1 H1 NT2-SP12 NT2-SP12 NT2-SP12 NT2-SP12 NT2-SP12 GATA6 GFP siRNA Normalised Ratio (Target/PBGD) Normalised Ratio (Target/GAPDH) GATA4 TERT 7 0.35 6 0.3 5 0.25 NANOG siRNA 4 0.2 3 0.15 0.1 2 0.05 1 0 0 hES-NCL1 H1 NT2-SP12 Normalised Ratio (Target/GAPDH) Normalised Ratio (Target/GAPDH) GATA6 AFP 0.25 0.5 0.2 0.4 0.15 0.3 0.1 0.2 0.05 0.1 0 0 Normalised Ratio (Target/GAPDH) Normalised Ratio (Target/GAPDH) LAMININ B1 GATA2 2.5 0.9 2 0.7 1.5 0.5 1 0.3 0.5 0.1 0 0 hES-NCL1 H1 NT2-SP12 Normalised Ratio (Target/GAPDH) Normalised Ratio (Target/GAPDH) CDX2 NODAL 0.35 2 0.3 1.5 0.25 0.2 1 0.15 0.1 0.5 0.05 0 0 hES-NCL1 H1 NT2-SP12 Regulacija NANOG-a AFP GFP siRNA NANOG siRNA Nis, 20/03/2008

  26. Maticne celije i regenerativna medicina • Lista cekanja za transplantaciju organa je porasla triputa dok je u isto vreme lista davaoca organa vrlo kratka • Veliki broj degenerativnih oboljenja • Razvoj novih lekova • Resenje: adultne i embrionalne maticne celije Nis, 20/03/2008

  27. Neuronalne celije Nis, 20/03/2008

  28. Tretman Nis, 20/03/2008

  29. Tretman Nis, 20/03/2008

  30. Diferencirane embrionalne i celije sa neuronalnim karateristikama Tuj1 Glia Neur. Tuj1 A2B5 Glut Nis, 20/03/2008

  31. +30mV A B -60mV -40mV C D 20pA F Transmitter sensitivity -20mV VH E GABA (100M) GABA/Bic (100M) +4-AP (2mM) +60mV VH Fig. 1. Functional characterization of an animal-free stem cell line (n=35). A: Voltage-gated currents evoked by a family of depolarizing steps in a neuron-like stem cell (5-7 days old), showing an early inward current typical of voltage-gated Na+ channels and a delayed outward current of voltage-gated K+ channels, both necessary for obtain action potentials. B: Current-to-voltage plot of the currents showed in A for the inward and outward currents wich fits with the behaviour of the K+ and Na+ channels from an excitable cell (KCl in the pipette). C: Action potentials evoked by the injection of 20pA to the cell in the current-clamp configuration (KCl in the pipette). D: Na+ current (n=24) evoked by a depolarizing step to –20mV (VH: -70mV). D: K+ current (n=35) evoked in a different neuron by a depolarizing step to +60mV. The current was blocked by 4-AP by the 756% (n=10). E: Whole-cell currents evoked by GABA 100M (n=28 from 35). Bicuculine (n=10) was employed at the same concentration to block the currents. Nis, 20/03/2008

  32. Srcane misicne celije iz plasticne solje Nis, 20/03/2008

  33. Srcane misicne celije iz embrionalnih maticnih celija Nis, 20/03/2008

  34. Maticne celije Nis, 20/03/2008

  35. 12 10 8 Vertical displacement μm 6 4 2 0 10 sec 12 10 8 Vertical displacement μm 6 Contraction (vertical displacement) 4 2 Calcium dynamics 1 mM bile acid 10 sec Srcane celije i novi lekovi scanning ion conductance microscopy Nis, 20/03/2008

  36. Maticne celije diferencirane u celije krvi Nis, 20/03/2008

  37. Roznjace iz embrionalnih maticnih celija Nis, 20/03/2008

  38. Oplodnja, kloniranje,maticne celije, epigenetika Nis, 20/03/2008

  39. Organi kloniranih zivotinja jetra srce bubreg kosti Nis, 20/03/2008

  40. Derivacija maticnih celija iz kloniranih embriona Nis, 20/03/2008

  41. Klonirani embrioni Nis, 20/03/2008

  42. Derivation of patient-specific stem cells

  43. Strategije za dobijanje pluripotentnih maticnih celija NT pluripotent ESC enucleated human oocyte donor cell NT-zygot NT-blastocyst Cell Fusion pluripotent ESC donor cell pluripotent SC somatic cell Genetic Manipulation c-MYC, KLF4, OCT-3/4, SOX2 OCT-3/4, SOX2, NANOG, LIN28 somatic cell defined factors iPS Nis, 20/03/2008

  44. Genome-Wide Scan for Seven Human Diseases

  45. Buducnost vec pocela:Srcane celije iz celije kozeSve celije iz celije kozeNasledne bolesti u plasticnoj solji Celije po meri Nis, 20/03/2008

  46. Med. fakultet Kragujevac

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