Enhanced polyploidy by glutaraldehyde in cultured HL60 leukemic cells.

Glutaraldehyde 10(-4) M weakened cell proliferation of HL60 cultured cells and enhanced the appearance of giant polyploid cells, up to 32.5% after 6 days. The size and structure of these cells, the quantitative changes in their DNA content with respect to diploid ones demonstrate their polyploid nature, which may be corrected by the occurrence of pluripolar mitoses. However the slowing down of cell proliferation is not enough to orient the cells towards differentiation. Maturation of polyploid cells may be stimulated by retinoic acid and dexamethasone as for diploid ones. Several possible mechanisms of polyploidy are discussed. Except the possibility that the cells may directly fuse, the mechanisms which are considered, may involve a preprophase inhibition, a mitotic arrest at metaphase or a reduction of asters which may result in a defect in cytokinesis, the latter followed by secondary fusion of nuclei.

[Thyroid incorporation of radioactive iodine in neotenic tritons (Triturus helveticus Raz.). In vivo studies by high resolution radioautography. Comparison of iodine incorporation in adults and larvae]. / Incorporation thyroïdienne de l'iode radioactif chez des tritons néoténiques (Triturus helveticus Raz.). Etude in vivo et par radioautographie à haute résolution. Comparaison avec l'incorporation d'iode des adultes et des larves.

In vivo kinetics of radioiodine incorporation have been established for 70 newts, collected from Triturus helveticus Raz. populations where spontaneous neoteny occurs. Fifty of these newts which appeared to present four degrees of neoteny (more or less metamorphosed), have been studied. Main results obtained were: 1) The intensity of the iodine incorporation varies according to the degree of neoteny. The higher is this one, the lower is the intensity. 2) In general, iodine uptake rates are more or less similar to these of adults, but more often the evolution of the 125I uptake approaches a plateau-curve. From thyroid radioautographies on some animals (17 neotenic and 2 adult newts) for which in vivo iodine kinetics have been examined and on 9 normal larvae in premetamorphosis or in metamorphosis, we have noticed in those neotenic thyroids, a very small number of silver grains on apical vesicles and colloidal lumina when the degree of neoteny is the highest. This number of marked apical vesicles and colloid is more important in the neotenics which have lost some larval characters. Silver grains on colloid droplets indicating thyroid hormone excretion are inexistent in the most larval neotenics, more numerous in most metamorphosed neotenics. But they always are less numerous than in metamorphosing larvae. For a few animals, the fixation curve is very high, probably because of a stimulation of thyroid function, induced by the test and not because of a goitrous tendancy, as radioautographic observation of their thyroid shows. Thyroid function and its relation with the origin of this total and accidental neoteny are discussed.

Inhibition of spindle assembly by colchicine and of cell cycle by cycloheximide. Comparison and combination of their cytological action on cleavage mitoses.

In cleavage mitoses colchicine (2 x 10(-3) M) does not interrupt the cycle of chromatin and its action is exerted on all the mitotic phases. Cycloheximide (10(-3) M) prevent normal condensation of the chromosomes and breakdown of the nuclear envelope; it brings on various nuclear abnormalities, particularly irregularities in chromosome condensation. At 10(-4) M, cycloheximide allows some mitoses, often abnormal ones, but anaphase is inhibited. The action of colchicine, more efficient upon the spindle, obscures the action of cycloheximide when both substances are associated. In return, cycloheximide imposes its action upon chromosome condensation, so that the structure of the nuclei treated by both substances is determined by cycloheximide.

Nuclear and mitotic abnormalities produced by cycloheximide in the newt egg during cleavage and their relationship to the cell cycle.

In cleaving newt eggs short-lasting treatments with millimolar quantities of cycloheximide (1/2, 1/3, 1/6 cycle) produce, after 6 h recovery, nuclear and mitotic abnormalities which are less drastic, but similar to those produced by treatments lasting 6 h. Special disturbances of the spindle and asters are different from those produced by spindle inhibitors (reduction in size of the asters and of the number of poles by nucleus, usual suppression of the spindle, failure of cytokinesis, separation of asters from nuclei). But the structure of nuclei is different according to the time when the treatment started (telophasic when it begun at anaphase or telophase, prophasic when begun at prophase). The difference in percentage of these two types of nuceli is highly significant and independent of the time passed after the beginning of the treatment. Each nucleus has to complete the whole cycle before the effect can be stopped.

Transformation, remodeling and dissociation of achromatic fibres by butyric acid in mitoses of cleaving eggs. Effects of associated antitubulin substances.

A mM solution of butyric acid transforms the achromatic apparatus of segmentation mitoses: asters are reduced or suppressed, cytokinesis is inhibited before mitosis, spindle fibers are broken and fan out progressively in the equatoria; region becoming radially oriented instead of axially. The results are either: 1) hollow fibrillar systems, 2) radiating spindle systems, apparently pluripolar, 3) dissociated fibrillar systems, depending on the phase of the mitotic cycle in which the butyric acid acted. Colchicine 2.5 X 10-3 M and vincaleukoblastine 2 X 10-4 M cause the depolarized nuclei and mitoses to migrate towards the animal cortex and the furrows, if they are administered before the abnormal systems produced by butyric acid are fully formed. Once formed, these systems are usually unchanged by administration of these drugs. Thus their formation appears to depend on microtubule assembly, since colchicine and VLB can prevent it.

[Glutaraldehyde and formaldehyde as fixative and antimitotic agents (author's transl)]. / Le glutaraldéhyde et le formaldéhyde, agents fixateurs et antimitotiques

Glutaraldehyde and formaldehyde produce a fixation effect at higher concentrations and an antimitotic one at lower concentrations. The arrested mitosis is characterized by immobilization of chromosomes in the equatorial plane, disappearance of all the spindle and astral fibers and "blocking" of centrospheres. It is a quinoline-like effect. The blocked centropheres persist at telophase or telo-prophase unlike the normal ones, which dissociate. This phenomenon is interpreted as a consequence of the storage of precursor material of microtubules in a more or less structural form around the centriole.

Progressive disassembly of the mitotic spindle into two halves at anaphase and assembly of an astral neo-spindle during cleavage mitosis in the newt embryo.

In the second cleavage mitosis of newt embryos at anaphase, disassembly of mitotic spindle fibres, mediated by the 'plus' ends of their microtubules, results in equatorial division into two half-spindles. Yolk-filled cytoplasm invades the area free of fibres and is itself penetrated by antiparallel fibres from asters, which have become nucleated at opposite poles. These fibres immediately supplant the mitotic spindle; they participate in the lengthening of the interpolar distance and in the positioning of the newly formed daughter nuclei. The mechanisms of these successive phenomena must be considered in order to understand the complexities of the cell cycle in early mitosis.

Methods of fixing, sectioning and staining amphibian eggs for cytological study.

A combination of methods for fixation (sublimate, cobalt nitrate, formaldehyde, acetic acid in water), inclusion (celloidin dissolved in methyl salicylate, paraffin-paraplast) and staining was used to make serial sections easy, to avoid clefts and to give a good picture of segmentation mitoses, as well as a good contrast of yolk and cytoplasmic components. Four methods of staining were used concerning the Urodele eggs: Safranin-methyl blue-orange G, safranin-picro-blue black naphthol (Curtis), safranin-violet crystal-orange G (Flemming) and Feulgen-methyl blue-orange G. The achromatic apparatus of the normal segmentation mitoses is clearly delineated and the relationships between astral fibers and yolk are different at metaphase and anaphase. By these methods, particularly suitable for demonstration of nuclei, cytoplasm and achromatic apparatus, the cleaving egg may be used as a test for the inhibition of achromatic apparatus and chromosome damage by antimitotic substances. The contrast between vitelline cytoplasm and cytoplasmic non-vitelline abnormal fibrillar systems, produced by transformation of astral and diastematic fibres, is made particularly evident by these methods of staining.

[Trifluralin, an inhibitor of the achromatic apparatus which modifies the chromosomes]. / Un inhibiteur de l'appareil achromatique qui altère les chromosomes, la trifluraline.

Trifluralin weakens or destroys polarity in segmentation mitoses of Triturus helveticus Raz. and Pleurodeles waltlii Michah. Previously it inhibits the cytokinesis, by reducing the activity of asters which are necessary to it. It produces chromosome abnormalities, specially involving their condensation, and makes appear discontinuities at prophase. The local thickenings of metaphase chromosomes predominate at heterochromatic regions. The action is stronger in animal than in vegetative half and this difference is highly significant. The stoppage of Amphibian eggs development states the problem of the possible action of trifluralin on the aquatic fauna.

Action of short-term cycloheximide administration associated with colchicine on nuclear and chromosome structure in cleaving Newt eggs. Reappearance of chromomeres?

Association of short-term cycloheximide (CHM) administration (1/3, 1/2 or 1 cycle) with long-term treatment by colchicine (Co) confirms the results obtained by CHM alone under similar conditions, namely the striking relationship between the starting-time of treatment and the type of nuclei obtained (centred telophasic or non-centred prophasic). This relationship is highly significant. Various nuclear structures were observed: they correspond to a special evolution of the abnormal nuclei during telophase, telo-prophase and prophase which is very different from that in untreated ones. A difference in evolution of the central and peripheral parts of the same nucleus explains their centred structure. On the contrary, this difference disappears at prophase. The association CHM-Co favours the unveiling of beads which are interpreted as prophasic chromomeres. These beads are of different size, spacing and form according to the moment of the prophasic evolution attained. Electron microscopy study confirms the strong dispersion of chromatin by CHM and the beaded appearance of chromosomes by CHM and Co. Simultaneously, many dense bodies are accumulated in the centrospheres, as after quinoline and glutaraldehyde administration. The mechanisms of production of centred nuclei and of the beaded appearance of chromosomes are discussed.

Cytological characteristics and classification of spindle inhibitors according to their effects on segmentation mitoses.

The effects of spindle inhibitors and of protein synthesis inhibitors on segmentation mitoses allow us to classify them into six groups : 1. Colchicine type : destruction of the whole achromatic apparatus and centrospheres without storing of dense bodies; 2. Quinoline type : same effect on the achromatic apparatus, but blocked centrospheres with accumulation of dense bodies; 3. Chloralhydrate type : Incomplete destruction of achromatic apparatus, spindle residue which maintains the chromosomes in a star shape, inactive centrospheres sequestered by the reticulum, but without accumulation of dense bodies; 4. Phenylurethane type : Incomplete and reversible action, which leads to easy production of pluripolar mitoses; 5. Carboxylic acid type : dissociation of the spindle, sometimes with blocking of the centrosphere, together with profound chromosome changes without primitive breaks; the intensity and quality of their action is related to the number of carbon atoms in the acid considered; 6. Protein synthesis inhibitor type : (cycloheximide, pederin) characterized by a stop of the nuclear cycle at telo-prophase when the action is sufficient, chromosome abnormalities, sometimes, reduced to strings of beads, and freeing of asters; at weaker concentrations mitosis is possible, but the congression of chromosomes at the equator is abnormal because of functional disturbance of the kinetochores. The nature and grading of these effects, their association (or non - association) to chromosome damage, the soundness of the spindle when only the chromosomes are affected (nitrogen mustard) make this one of the tests which gives the most specific data about the action of antimitotic substances.

[Coating of centrospheres by reticulum cisternae and formation of spindle residues in segmentation mitosis under the action of an antitubulin substance: nocodazole]. / Enveloppement des centrosphères par des citernes de réticulum et formation de résidus fusoriaux dans les mitoses de segmentation sous l'influence d'une substance antitubuline, le nocodazole.

When cleaving eggs are treated by nocodazole the reticulum cisternae coat the centrospheres instead of the chromosomes, as normally occurs at the late anaphase. At the same time the spindle is reduced to a common mass or spindle remnant, constituted of unorientated and fragmented microtubules which embed the kinetochores, i. e. the star configuration of the chromosomes. This action is quite similar to that of chloralhydrate but is different from that of colchicine.

[Mechanism of the blocking of the centrosphere by substances similar to quinoline. Its prevention with spindle inhibitors similar to colchicine]. / Le mécanisme du blocage de la centrosphère par des substances semblables à la quinoline. Sa prévention par des inhibiteurs du fuseau semblables à la colchicine

A colchicine-like substance, vincaleukoblastine (2.10-4M, 3 to 6 hours), prevents the blocking of the centrosphere which would be produced by quinoline or by a similar substance (glutaraldehyde), to the extent of the inhibition of polarity by this colchicine-like substance. This is interpreted as due to the fact that each molecule of colchicine-like substance binds to a single subunit (or heterodimer) of tubulin and consequently prevents the constitution of storage structures. It is supposed that on the contrary quinoline-like substances bind to several subunits of tubulin and that they can stabilize the storage structures, binding to them in several points. The ultrastructural study demonstrates that when the centrosphere is blocked by quinoline, there is accumulation, hypertrophy and permanence of the "dense bodies" later than prophase; sometimes they are fused in a large "dense mass". The "dense bodies", and perhaps the "striated bodies", may therefore be interpreted as constituting morphologically the storage structures, which enlarge when all the astral and spindle microtubules completely disappear by action of a quinoline-like substance.

[Round cells of human sperm]. / Les "cellules rondes" du sperme humain.

Many round cells are found whose number may become conspicuous. These cells may originate as well from germinal as from somatic lines. Electron microscopy is the only way to distinguish the two kinds of origin. In some specimens germinal cells predominate (spermattids, plurinuclear cells), in others there are many phagocytic cells which may ingest a great number of spermatozoa and this phenomeon may stimulate an azoospermy by failure of spermatogenesis. The study of human sperm (or testicular biopsies) by electron microscopy is needed to obtain the necessary information in order to orient diagnosis, prognosis and therapy.

[The effect of formamide on the achromatic apparatus and the chromosomes of dividing pleurodele eggs: an electron microscope study]. / Action de la formamide sur l'appareil achromatique et les chromosomes dans des oeufs de pleurodèle en segmentation: étude au microscope électronique

Electron microscope studies of blastulae of Pleurodeles waltlii Michah., treated with formamide (0,5 M for 17 hours), show various abnormalities concerning centrospheres, chromosomes and microtubules as well. At the prophase stage, the centrospheres may display at their outer region an accumulation of dense bodies, which shows a temporary blocking of these centrospheres. At metaphase the poles of pluripolar mitoses appear more or less separated from the material which contains the chromosomes of the equatorial region. Each of these poles contains an unique centriole and therefore it is considered as resulting of the subdivision of a normal anaphasic pole. Astral and spindle microtubules are very reduced in number and more or less shortened. The chromosomes are without any recognizable kinetochcre, they show various degrees of condensation. All these electron microscope observations confirm the light microscope ones, and we may conclude that formamide has a double action on microtubules formation and on the ultrastructure of chromosomes, the latter perhaps by a direct action on DNA protein complexes.

Ultrastructure of segmentation mitoses in cleaving newt eggs after blocking of centrospheres by quinoline.

In quinoline treated blastomeres of Triturus helveticus Raz. and Pleurodeles waltlii Michah., microtubules disappear and around the centrioles markedly enlarged dense bodies accompanied by striated bodies are accumulated, from prophase beyond metaphase. It is concluded that these bodies in untreated cells play a role in the formation of the spindle microtubules. - During metaphase the chromosome are smooth-surfaced and show a pronounced tendency to stickiness. During ana-telophase they become surrounded by nuclear membranes and form caryomeres. - Qinoline does not interfere with centriole replication, but prevents the separation if diplosomes that have been already formed, if it acts before that separation. - Some centrioles exhibit different degrees of ultrastructural disarrangement.

[Ultrastructural characteristics of human spermatozoa with elongated head (author's transl)]. / Particularités ultrastructurales des spermatozoïdes humains à tête allongée.

In most spermatozoa with elongated head persists an abundant complex membranous system between the post-acromial zone and the post-nuclear region. This system has a periodical structure, the apparently spherical unit of which is 120 A in diameter; it develops between the two sheaths of the nuclear envelope and seems to be continuous with the innermost one. This system can be seen in spermatids during normal spermiogenesis and in most cases it seems to be eliminated either with cytoplasmic droplet or by swelling into the nucleus. Now, a comparable but permanent structure is located all around the head of the normal spermatozoon at the height of the posterior part of the post-acrosomial sheath. This latter system is organized under plasma membrane and only visible on the ultrathin sections made with an appropriate incidence. For some authors it might contribute to maintain the integrity of the head and that of the neck of the spermatozoon and so it might be comparable to a system of septal desmosomes. For other authors periodic structures might favour adherence and fusion of spermatozoon with ovocyte during fertilisation. The particular membranous system that we describe persists often in the spermatozoon with elongated head, by its exuberance it can trouble or delay the fusion of the two gametes at the moment of fertilisation, which might explain the sterility of men bearing this anomaly.

The fine structure of meningiomas: an attempted classification.

We examined 23 meningiomas by electron microscopy. In each case it was possible to distinguish certain cells with epithelial features (desmosomes, microfliaments, interdigitating extensions) and others with fibroblastic features (collagen fibers). Others cells of transitional form were also seen. The proportion of these cellular types is variable, making it possible to classify meningiomas into seven types of progressing gradually from a purely epithelial type to a purely fibroblastic one.-We found no important ultrastructural abnormalities in the cells. These case reports confirm the uniqueness of meningiomas, which are composed of variously shaped cells but have their origin from a single cellular type. This has double potentiality for fibroblastic and epithelial differentiation.

[Cell coat modifications in a series of gliomas as a function of malignancy]. / Modifications du cell coat dans la série des gliomes en fonction de la malilgnité.

We have compared the cell coat of benign gliomas and of malignant gliomas, by Ruthenium Red staining in vitro and by Thiéry's cytochemical reaction in situ. The thickness of the cell coat is not significantly increased by malignancy on the free surfaces of glioma cells. But in malignant gliomas only the cell coat disappears on the intercellular contact zones. These modifications are related to cell fusion characteristics of malignant transformation.

[Effects of propham and chlorpropham on the embryonic development of various vertebrates]. / Effet du propham et du chloropropham sur le développement embryonnaire de quelques vertébrés.

The effect of propham and chlorpropham on Amphibians embryonic development changes with doses as well as species. The action of C.I.P.C. is always determinative. The egg poisoned in albumen involves important chick embryo malformations, but the development is stopped by the yolky vesicle poisoning. The intraperitoneal injection of C.I.P.C., strongly disturbs fetus physiology in Mouse.