Open AccessAutomated absorption image cytometry of electromagnetically exposed frog erythrocytes
Author(s) -
Chiabrera A.,
Viviani R.,
Parodi G.,
Vernazza G.,
Hinsenkamp M.,
Pilla A. A.,
Ryaby J.,
Beltrame F.,
Grattarola M.,
Nicolini C.
Publication year - 1980
Publication title -
cytometry
Language(s) - English
Resource type - Journals
eISSN - 1097-0320
pISSN - 0196-4763
DOI - 10.1002/cyto.990010110
Subject(s) - acridine orange , chromatin , fluorescence , flow cytometry , biophysics , dna , biology , microbiology and biotechnology , chemistry , biochemistry , optics , physics , apoptosis
Frog erythrocytes dedifferentiate in vitro when exposed to a suitable electromagnetic field in a chemically potentiated microenvironment. These electrochemical stimuli are able to induce a chromatin decondensation, as previously shown by measuring the chromatin accessible sites for acridine orange intercalation by means of flow cytometry (acridine orange green fluorescence). Automated absorption image cytometry of Feulgen stained smeared erythrocytes has been performed to further elucidate the above processes. After measuring or computing the area A of the nucleus, which is related to its volume, the nucleic integrated optical density D, which is related to the DNA content, the average optical density D A = D/A, which is related to chromatin conformation, and the accessible chromatin sites S A = D 2/3 A 1/3 , the following results have been obtained: ( a ) electromagnetically exposed cells progress to stages which correspond to values of D equal to those of controls, in the potentiating solution, whereas A increases, so that D A is smaller in exposed erythrocytes than in control ones, confirming that chromatin decondensation is occurring as an early dedifferentiation step. ( b ) By using the electromagnetic signal that is most effective in promoting dedifferentiation, erythrocytes further progress toward more advanced stages, which correspond to larger values of both D and A than in controls, i.e. , to larger DNA content. ( c ) In all cases, the histograms of S A are in agreement with those previously obtained by flow microfluorometry of chromatin conformation (acridine orange green fluorescence). Finally, flow microfluorometric measurements of acridine orange red fluorescence give an increase of RNA content for case ( a ), as compared with controls, and for case ( b ) as compared with ( a ). These results point out that frog erythrocytes can be electromagnetically reactivated, resuming both RNA and DNA syntheses after initial chromatin decondensation.