Ira Ghosh

Some aspects of the Feulgen reaction in situ

SummaryCytological observations combined with studies on absorption spectra of Feulgen stained normal and lipid — extractet HeLa and ehrlich-Lettré mouse ascites cells were performed after fixation of the cells as well in neutral formaldehyde as in Serra fixative. The effects of formaldehyde treatment of the stained cells to substitute all the free amino groups of DNA bond pararosaniline molecules, were also studied. The results obtained by using DNA samples containing 2% protein and relatively free from protein, led to the conclusion that after acid hydrolysis for a short period purines in DNA become splitted and these released aldehydes react with one or two amino groups of pararosaniline, a triphenylmethane dye (according to the arrangement of purines and pyrimidines in the helices). Some protein molecules also take part in the reaction and substitute some of the free amino groups of DNA bound pararosaniline. Peulgen stained cells fixed in Serra fixative show an absorption maximum at 546–550 mμ. Under appropriate conditions, as in cells fixed in formaldehyde, other substances e.g. phospholipids and lipoproteins interfere with the reaction by substituting most of the free amino groups of DNA bound pararosaniline molecules. It has been argued that in histochemical reactions monosubstituted pararosaniline molecules should be coloured and further substitution of free amino groups of pararosaniline, bound in DNA helices, does not change the intensity of the colour, but gives a shift in the wavelength of the absorption spectra.It has been suggested that the differential response of the nucleoli to the Feulgen-reaction, depending on whether the cells were fixed in formaldehyde or in Serra fixative, may be due to the formation of a protecting shield around the finely distributed intranucleolar chromatin strands, when formaldehyde is being used. After this fixation lipoproteins and other lipids, present in a relatively high percentage and closely associated with the intranucleolar chromatin strands, are especially well preserved.Evidences have been put foreward in support of the amino alkylsulfonic acid theory of Rumpf (1935) and Hörmannet al. (1958) whereas the amino sulfinic acid theory to explain the Schiffs reaction (Wieland and Scheuing, 1921) was shown not to be in agreement with our results.

Chromosomal alterations in the evolution of substrains of the Ehrlich-Lettré-mouse ascites tumour

The Ehrlich-Lettré mouse ascites tumour and four of its suhstrains were cytogenetically studied. The detailed karyotypes of the stemlines present in these cell populations have been described. It has been shown that in the evolution of the new substrains, structural and numerical changes in the stemline karyotype of the original stock, are involved. The genetic importance of such somatic cell variation has been discussed.

On the composition of the nucleolus with special reference to its filamentous structure

SummaryDifferent staining procedures, various digestion methods and autoradiographic techniques were employed to study the structure and composition of the nucleolus and of the nucleolonema, after unmasking the latter by adenosine treatment. The presence of DNA, RNA, protein and lipid in these structures has been shown. It has been demonstrated that the filamentous structure within the nucleolus — the nucleolonema— has a core of DNA, around which RNA and protein have accumulated. The structure of the nucleolonema suggests that it is in a highly active state, in synthesizing ribosomal RNA and protein.

Variation of stemline karyotype in a HeLa cell line

100 karyotypes of a HeLa cell line (modal number 69) were studied in details. It was observed that the stemline cells of this “triploid” somatic cell population showed high degree of chromosomal polymorphism. It has been discussed that the stemline cells have number of extra chromosomes (or parts of chromosomes) which are not essential for the genetic integrity of this cell population. A loss of some of these chromosomes (or their parts) does not induce any appreciable change in the genetic make-up of these cells. The broken chromosomes lose their identity and are grouped together with the non-homologues, when the Denver system of classification is followed. However, it has been argued that in absence of any other acceptable system for classification, the Denver system can still be employed for analyzing human chromosomes both at diploid and abnormal heteroploid level. It has been held in contrast to the pseudo-stemline concept that these cells in general, have the essential genes in common and are responsible for the genetic make-up of this cell line and constitute together the stemline of this somatic cell population. Eine detaillierte Analyse der Karyogramme von hundert Zellen einer Unterlinie des Stammes HeLa wurde durchgeführt. Die Zellen der Stammlinie dieser triploiden Population zeigen ein hohes Maß an Chromosomenpolymorphismus. Es wird diskutiert, ob die Zellen der Stammlinie eine bestimmte Zahl von Extrachromosomen der Chromosomenteilen besitzen, die für die genetische Unversehrtheit der Unterlinie unwesentlich sind. Ein Verlust einiger dieser Chromosomen oder Chromosomenteile bedeutete danach keine entscheidende Veränderung der genetischen Beschaffenheit der Zellen. Wenn man dem Denver-System der Klassifizierung der menschlichen Chromosomen folgt, verlieren die zerbrochenen Chromosomen ihre Identität und gehören dann zur Gruppe der nicht-homologen Chromosomen. Es wurde nachgewiesen, daß wegen Fehlens eines geeigneten Systems zur Klassifizierung der Chromosomen das Denver-System sowohl für diploide als auch heteroploide anomale Zellen benutzt werden kann. Im Gegensatz zu dem “pseudostemline concept” wird angenommen, daß die Zellen die wesentlichen Gene gemeinsam besitzen, die für die genetische Beschaffenheit verantwortlich sind. Derart ausgestattete Zellen bilden dann die Stammlinie der Population.

Karyological studies on two HeLa lines

Zwei Unterlinien des menschlichen Tumorstammes HeLa, welche mit verschiedenen Methoden in vitro gezüchtet werden und verschiedene Wachstumsgeschwindigkeit aufweisen, wurden karyologisch untersucht. Bei der sog. “monolayer” Linie wurde in der Stammlinie eine Chromosomenzahl von 69 festgestellt, welche jedoch in ihrer Zusammensetzung nicht einem tripoloiden Chromosomensatz entspricht. In der Stammlinie der sog. “clot” Linie ließ sich eine Chromosomenzahl von 74 nachweisen. Es wurde in der Diskussion besprochen, daß die Unterschiede im Wachstumsverhalten beider Linien nicht einfach auf die verschiedene Anzahl von Chromosomen in beiden Linien zurückgeführt werden sollten, sondern mit den Unterschieden in der Zusammenstellung der Chromosomen verschiedener Typen zusammenhängen. Es wurde betont, daß die Charakterisierung einer Stammlinie nach der Zahl ihrer Chromosomen als unzureichend angesehen werden muß und daß eine detailliertere Analyse der Stammzellen-Karyogramme (nach dem Denver-System) notwendig erscheint. Two HeLa cell lines grown under different conditions were studied karyologically. The “monolayer” line was found to posses a stemline chromosome number of 69 (but not typically triploid in chromosomal characteristics), whereas the “clot” line had 74 chromosomes in its stemline cells. In the discussion it was decided that the differences in growth behaviour of these two lines should not be attributed to mere numerical differences of their stemline karyotypes, but should be related to the differences in the nature of their stemline karyogrammes. It was stressed that a more detailed characterization of the stemline karyogramme (following the Denver-system) is necessary.

Endokrine Organe · Endocrine Organs · Glandes endocrines

We have previously reported the utility of the histochemical method for the oxidative enzyme α-glycerophosphate dehydrogenase in depicting certain cells of the anterior pituitary in the rat. Extension of these studies to more than 50 human pituitary glands reveals similar selective anterior lobe enzymic staining in cells which may be thyrotrophs. In addition, cells running along pituitary stalk in the human have been demonstrated to have high complements of this enzyme. These are the cells of the pars tuberalis. Of particular interest is the clear cut finding that these positively stained cells enter into and merge with the basophils of the coronal portion of anterior lobe at their junction with the pars intermedia. They do not enter posterior lobe as does the stalk. Since these cells accompany stalk along much of its course from hypothalamus to hypophysis, it is suggested that they may serve as an anatomical link between brain, on the one hand, and the non-nervous portion of the hypophysis, on the other.