Determination of the content of FAD cofactor and NAD(P)H-oxidase complexes in mouse splenocytes and Lewis carcinoma cells under conditions of apoptosis by confocal microscopy method

Capa

Citar

Texto integral

Acesso aberto Acesso aberto
Acesso é fechado Acesso está concedido
Acesso é fechado Acesso é pago ou somente para assinantes

Resumo

In this work, using fluorescence and confocal microscopy, we studied the content of the cofactor FAD and enzymatic NAD(P)H-oxidase complexes (with fluorophores AnnexinV-FITC, 7-AAD (7-aminoactinomycin D), EtBr) under conditions of apoptosis caused by sodium anphene with hydrogen peroxide in healthy mouse splenocytes and Lewis carcinoma tumor cells. The use of fluorescence microscopy allows observing and quantifying the apoptotic effect of sodium anphen and hydrogen peroxide, and visualization of metabolic changes in the cell, including increased fluorescence of FAD in tumor cells and NAD(P)H-oxidase complexes in splenocytes. The data obtained indicate the possibility of using sodium anphen in combination with hydrogen peroxide as an antitumor drug acting on certain types of cells.

Texto integral

Acesso é fechado

Sobre autores

E. Mil

Emanuel Institute of Biochemical Physics Russian Academy of Sciences

Email: albantovaaa@mail.ru
Rússia, Moscow

A. Albantova

Emanuel Institute of Biochemical Physics Russian Academy of Sciences

Autor responsável pela correspondência
Email: albantovaaa@mail.ru
Rússia, Moscow

L. Matienko

Emanuel Institute of Biochemical Physics Russian Academy of Sciences

Email: albantovaaa@mail.ru
Rússia, Moscow

A. Goloshchapov

Emanuel Institute of Biochemical Physics Russian Academy of Sciences

Email: albantovaaa@mail.ru
Rússia, Moscow

M. Korovin

Emanuel Institute of Biochemical Physics Russian Academy of Sciences

Email: albantovaaa@mail.ru
Rússia, Moscow

V. Kuvyrkova

Emanuel Institute of Biochemical Physics Russian Academy of Sciences

Email: albantovaaa@mail.ru
Rússia, Moscow

Bibliografia

  1. N.V. Beloborodova, General Reanimatology 15, 62 (2019); doi::10.15360/1813-9779-2019-6-62-79
  2. V.I. Binyukov, E.M. Mil, L.I. Matienko, et al., Micro (MDPI) 3, 382 (2023); htpp//doi.org/10.3390/micro3020026
  3. L.I. Matienko, E.M. Mil, V.I Binyukov, Russ. J. Phys. Chem. B 14, 559 (2020); https://doi.org/10.1134/S1990793120030227
  4. J.R. Mcintosh, J. Cell Biol., 153, 25 (2001); https://doi.org/10.1083/jcb.153.6.F25
  5. N.S. Zakharov, A.N. Popova, Yu.A. Zakharov, et al., Russ. J. Phys. Chem. B 16, 780 (2022); doi::10.1134/s1990793122040170
  6. V.A. Tkachuk, P.A. Tyurin-Kuzmin, V.V. Belousov, et al., Biological membranes, 29, 21 (2012) (in Russian).
  7. I.F. Rusina, T.L. Veprintsev, R.F. Vasiliev, Russ. J. Phys. Chem. B 16, 50 (2022); https://doi.org/10.1134/S1990793122010274
  8. N.Yu. Gerasimov, O.V. Nevrova, I.V. Zhigacheva, et al., Russ. J. Phys. Chem. B 17, 135 (2023); doi::10.1134/S1990793123010049
  9. R.A. Sadykov, S.L. Khursan, A.A. Sukhanov, et al., Russ. J. Phys. Chem. B 17, 1251 (2023); doi::10.1134/S1990793123060209
  10. E.M. Mil, V.I. Binyukov, V.N. Erokhin, et al., Cytology 62, 503 (2020) (in Russian); doi::10.31857/S0041377120070032
  11. V.N. Erokhin, A.V. Krementsova, V.A. Semenov, et al., Bull. Russ. Acad. Sci., Biochemistry No 5, 583 (2020) (in Russian).
  12. E.M. Mil, V.I. Binyukov, V.N. Erokhin, Dokl. Chem. 482, 598 (2018) (in Russian).
  13. W. Becker, J. Microscopy 247, 119 (2012); doi::10.1111/j.1365-2818.2012.03618.x
  14. I.N. Druzhkova, M.M. Lukina, V.V. Dudenkova, et al., Cell Cycle 15, 1257 (2016); doi::10.1080/15384101.2016.1160974
  15. N.J. Clifton, Methods in molecular biology 412, 273 (2007); doi::10.1007/978-1-59745-467-4_18
  16. K. Rokutan, T. Kawahara, Y. Kuwano, et al., Antioxid. Redox. Signal 8, 1573 (2006); doi::10.1089/ars.2006.8.1573
  17. M.W. Ma, Wang J., Zhang Q., et al., Molecular Neurodegeneration No 12, 7 (2017); https://doi.org/10.1186/s13024-017-0150-7
  18. M.M. Lukina, V.V. Dudenkova, N.I. Ignatova, et al., Biochim. Biophys. Acta - Genetic Subj. 1862, 1693 (2018); doi::10.1016/j.bbagen.2018.04.021
  19. А.S. Babkina, General Reanimatology 15, 50 (2019); https://doi.org/10.15360/1813-9779-2019-6-50-61
  20. A. Vermot, I. Petit-Härtlein, S.M.E. Smith, et al., Antioxidants (Basel) 10, 890 (2021); doi::10.3390/antiox10060890
  21. K. Bedard, K.H. Krause, Physiol. Rev. 87, 245 (2007); doi::10.1152/physrev.00044.2005

Arquivos suplementares

Arquivos suplementares
Ação
1. JATS XML
Baixar
2. Fig. 1. FAD fluorescence in mouse splenocytes: single (left) and multiple fluorescence (right) in cells during apoptosis.

Baixar (124KB)
Metadados
3. Fig. 2. Micrographs of splenocytes during apoptosis induced by sodium anfen (10-4 M) and a 6-fold magnified image of the cell (granules of NAD(P)H-oxidase enzyme complexes NOX are visible on the membrane). Confocal microscope with 100x magnification (fluorophore – Annexin V-FITC).

Baixar (204KB)
Metadados
4. Fig. 3. Diagram of the content of apoptotic cells in the splenocyte culture (fluorophore – AnnexinV-FITC) – 1, the number of non-viable cells (fluorophore EtBr) – 2. Changes in the cell content under the influence of hydrogen peroxide (5 μM), sodium anfen (10-4 M), and their combined effect.

Baixar (17KB)
Metadados
5. Fig. 4. Percentage of cells with FAD-1 fluorescence and immunofluorescence of granules of NAD(P)H-oxidase complexes-2 in a splenocyte culture under the influence of hydrogen peroxide (5 μM), sodium anfen (10-4 M) and their combined effect.

Baixar (65KB)
Metadados
6. Fig. 5. Micrographs of a neutrophil in a splenocyte culture, which is in the stage of NETosis – the release of a DNA network surrounded by NAD(P)H-oxidase complexes – (fluorophore – AnnexinV-FITC (a) and 7AAD (b)). Confocal microscope with 100x magnification.

Baixar (171KB)
Metadados

Declaração de direitos autorais © Russian Academy of Sciences, 2024