ФЕНОМЕН "СТАЦИОНАРНОГО СТАРЕНИЯ" КЛЕТОЧНЫХ КУЛЬТУР: ТРИДЦАТЬ ЛЕТ СПУСТЯ А.Н.Хохлов

1. ФЕНОМЕН "СТАЦИОНАРНОГО СТАРЕНИЯ" КЛЕТОЧНЫХ КУЛЬТУР: ТРИДЦАТЬ ЛЕТ СПУСТЯ А.Н.Хохлов Сектор эволюционной цитогеронтологии, биологический факультет МГУ имени М.В.Ломоносова, Москва, Россия khokhlov@mail.bio.msu.ru

3. Cell Senescence

4. In culture experiments а la Hayflick, researchers estimate the replicative capacity of cells by counting the number of times a culture can be split before the cells stop dividing. Trypsin is added to free the cells from the extracellular matrix, and cultures are grown until cells carpet the floor of the flasks. This protocol spawned the legendary Hayflick limit (from Hopkin, 2001).

5. Hayflick’s phenomenon(IOW - “cell senescence”; unfortunately, last years this term, in my opinion, has rather often been used incorrectly by those who relate it to so called DNA damage response) – limited normal cell proliferation in vitro and in vivo (Population Doubling Level limit) • From the first to the last passage of “senescing” cells they change in the same way as cells of aging multicellular organism • We cannot explain why we age just with the help of the Hayflick’s phenomenon (say, the most important organs – brain and heart – contain a lot of postmitotic cells like neurons or cardiomyocytes) • The Hayflick’s phenomenon is defined by a set of various correlations only However, now we know how the Hayflick’s phenomenon is realized (thanks to Alexei Olovnikov!)

13. Хохлов А.Н., Чиркова Е.Ю., Ушаков В.Л. Использование стационарных культур для изучения механизмов клеточного старения В сб.: Первый съезд Белорус. общества геронтологов и гериатров. Тез. докл. Минск, 1983, 188-189

17. Influence on this stage by geroprotector or geropromoter modulates the rate of aging only temporarily Influence on this stage allows to find out whether the factor investigated is geroprotector or geropromoter

18. Cell aging is the accumulation in a cell population of various types of damage identical to the damage arising in senescing multicellular organism

19. Schematic representation of cultured cell “stationary phase aging” phenomenon Cell density (relative units) Time after subcultivation (days)

20. Survival curve of Chinese hamster stationary cell culture (solid line – experimental points; dotted line – approximation by the Gompertz equation; straight line – mortality rate changes with time)

25. ?

28. Telomeres and Telomerase

29. «Age» changes shown to really occur in stationary cell cultures Accumulation of DNA breaks and DNA-protein cross-links Accumulation of 8-oxo-2’-deoxyguanosine in DNA Inhibition of poly-ADP-ribosylation of chromatin proteins DNA demethylation Changes of spontaneous sister chromatid exchange level Plasma membrane changes Nuclear structure modulations Decrease of the rate of mitogen-stimulated cell cycling and of cell colony-forming ability

30. «Stationary phase aging» experiments have been already carried out on: Normal and transformed human and animal cells Plant cells Bacteria Mycoplasmas Yeasts

31. Bacteria

33. Yeasts

34. 2012

42. Thanks for your attention and patience, sincerely yours,AN Khokhlov

44. If you want to make God laugh – describe to Him your theory of aging

45. Naturally, anybody from the audience has the right to object to me. And I can certainly agree with him. But then we'll both be wrong - .