Interdigital tissue chondrogenesis induced by surgical removal of the ectoderm in the embryonic chick leg bud

Development ◽  
1986 ◽  
Vol 94 (1) ◽  
pp. 231-244
Author(s):  
J. M. Hurle ◽  
Y. Gañan
Keyword(s):  
Cell Death ◽  
Vital Staining ◽  
Mesenchymal Cell ◽  
Neutral Red ◽  
Surgical Removal ◽  
Experimental Conditions ◽  
Local Inhibition ◽  
Dorsal Ectoderm ◽  
Interdigital Cell Death ◽  
Proximal Zone

In the present work, we have analysed the possible involvement of ectodermal tissue in the control of interdigital mesenchymal cell death. Two types of experiments were performed in the stages previous to the onset of interdigital cell death: (i) removal of the AER of the interdigit; (ii) removal of the dorsal ectoderm of the interdigit. After the operation embryos were sacrificed at 10–12h intervals and the leg buds were studied by whole-mount cartilage staining, vital staining with neutral red and scanning electron microscopy. Between stages 27 and 30, ridge removal caused a local inhibition of the growth of the interdigit. In a high percentage of the cases, ridge removal at these stages was followed 30–40 h later by the formation of ectopic nodules of cartilage in the interdigit. The incidence of ectopic cartilage formation was maximum at stage 29 (60%). In all cases, cell death took place on schedule although the intensity and extent of necrosis appeared diminished in relation to the intensity of inhibition of interdigital growth and to the presence of interdigital cartilages. Ridge removal at stage 31 did not cause inhibition of the growth of the interdigit and ectopic chondrogenesis was only detected in 3 out of 35 operated embryos. Dorsal ectoderm removal from the proximal zone of the interdigit at stage 29 caused the chondrogenesis of the proximal interdigital mesenchyme in 6 out of 18 operated embryos. The pattern of neutral red vital staining was consistent with these results revealing a partial inhibition of interdigital cell death in the proximal zone of the interdigit. It is proposed that under the present experimental conditions the mesenchymal cells are diverted from the death programme by a primary transformation into cartilage.

Genetic inhibition of mesenchymal cell death and the development of form and skeletal pattern in the limbs of talpid3 (ta3) mutant chick embryos

Development ◽  
1974 ◽  
Vol 31 (3) ◽  
pp. 747-760
Author(s):  
J. R. Hinchliffe ◽  
P. V. Thorogood
Keyword(s):  
Cell Death ◽  
Soft Tissue ◽  
Limb Development ◽  
Chondroitin Sulphate ◽  
Vital Staining ◽  
Mesenchymal Cell ◽  
Equivalent Position ◽  
Skeletal Pattern ◽  
Mesenchyme Cells ◽  
Interdigital Cell Death

Vital staining reveals that in homozygous (ta3/ta3) talpid3 embryos, the areas of mesenchymal cell death which occur regularly in normal limb development are absent or reduced. The necrotic locus in the central mesenchyme (the ‘opaque patch’) which in the normal chick limb reaches maximum development at stages 24 and 25 (4½–5 days) is absent or much reduced in talpid3 fore- and hindlimb-buds. Autoradiographic studies, following application of a 2 h pulse of 40 μCi of 35SO4 to the vitelline circulation, show that normal tibia and fibula incorporate 35SO4 into chondroitin sulphate at stage 24 and more strongly at stage 26 during the process of chondrogenesis. The mesenchyme in the opaque patch region of normal limbs ceases to incorporate 35SO4 into chondroitin sulphate at stage 24. Talpid3 mesenchyme cells in the equivalent position at stages 24 and 26 continue to incorporate 35SO4, remain viable and become chondrogenic. It is suggested that absence or reduction of this central necrotic locus in talpid3 is causally related to the fusion of radius/ulna and (in some cases) of tibia/fibula characteristic of the later stages (28–35) of talpid3 limb development. This evidence supports the hypothesis that cell death in the opaque patch plays a morphogenetic role in separation of radius/ulna and tibia/fibula. The digital plate of stage 32 (7½ days) normal limbs is characterized by massive necrosis of the interdigital tissue. In talpid3 forelimbs of stages 30–35 interdigital necrosis is absent, and there is no regression of the tissue between the digits (‘soft tissue syndactyly’). In talpid3 hindlimbs of stage 30–35 interdigital necrosis is either absent or much reduced, and there is little or no erosion of the soft tissue between the digits. This evidence supports the hypothesis that the morphogenetic role of interdigital cell death is in causing separation of the digits through shaping and remodelling the contours of the digital plate.

Syndactyly induced by Janus Green B in the embryonic chick leg bud: a reexamination

Development ◽  
1984 ◽  
Vol 84 (1) ◽  
pp. 159-175
Author(s):  
M. A. Fernandez-Teran ◽  
J. M. Hurle
Keyword(s):  
Vital Staining ◽  
Inhibitory Effect ◽  
Neutral Red ◽  
Epithelial Tissue ◽  
Structural Modifications ◽  
Janus Green ◽  
Mesenchymal Tissue ◽  
Whole Mount ◽  
Direct Inhibitory Effect ◽  
Interdigital Cell Death

In an attempt to clarify the mechanism of production of the syndactyly induced by Janus Green B (JGB) we have studied the morphology and structural modifications of the chick embryo leg bud after JGB administration by means of (1) neutral red vital staining, (2) whole-mount cartilage staining and (3) light microscopy and transmission and scanning electron microscopy. The results show that the well-known inhibition of interdigital cell death is accompanied by a precocious alteration of the epithelial tissue and especially of the epithelial-mesenchymal interface. 24 h after JGB administration the cells of the AER reduce the number of junctions and the basal ectodermal cells are detached into the mesenchymal tissue in zones in which the basal lamina undergoes disruption. In addition the interdigital mesenchymal cells diverted from the dying program are able to undergo a rapid differentiation into cartilage. It is proposed that the mechanism of production of JGB-induced syndactyly might be due to an alteration of the normal epithelial—mesenchymal interactions rather than to a direct inhibitory effect of the JGB on the dying program.

Cell death in the dorsal part of the chick optic cup. Evidence for a new necrotic area

Development ◽  
1984 ◽  
Vol 80 (1) ◽  
pp. 241-249
Author(s):  
Juan A. García-Porrero ◽  
Elvira Colvée ◽  
José L. Ojeda
Keyword(s):  
Cell Death ◽  
Vital Staining ◽  
Neutral Red ◽  
Necrotic Area ◽  
Spatiotemporal Pattern ◽  
Dorsal Part ◽  
Cell Degeneration ◽  
Chick Retina ◽  
Conventional Procedure ◽  
Optic Cup

The spatiotemporal pattern of morphogenetic cell death during the early development of the chick retina was studied by means of the neutral red vital staining and light microscopy. A modification of the conventional procedure of vital staining, which consisted of the injection of the dye into the neural tube lumen, was used for this purpose. In addition to the two areas of cell death known from previous literature, the first located in the ventral part of the optic cup and the second located in the insertion of the optic stalk with the diencephalon, a new area of cell death was described. This third necrotic area was located in the protruding dorsal part of the optic cup rim and was present throughout the stages 15 to 18. The area consisted of dying cells, fragments and phagocytosed cells. We suggest that this dorsal area of cell death could stop the intense dorsal growth of the optic cup and/or reshape the optic cup rim. Moreover, this area may influence the production of cell degeneration in the dorsal part of the invaginating lens placode.

Interdigital cell death during limb development of the turtle and lizard with an interpretation of evolutionary significance

Development ◽  
1977 ◽  
Vol 40 (1) ◽  
pp. 285-289
Author(s):  
John F. Fallon ◽  
Jo Ann Cameron
Keyword(s):  
Cell Death ◽  
Limb Development ◽  
Evolutionary Significance ◽  
Painted Turtles ◽  
Interdigital Cell Death

Cell death accompanies the formation of free digits in birds and mammals. However, in species with webbing between the adult digits, little or no cell death occurs in the prospectively webbed region of the developing interdigit. Cell death does not occur during the formation of free digits in amphibians. In this paper we report that cell death accompanies the formation of the digits in snapping and painted turtles and in the skink (a lizard). We conclude that cell death accompanying the formation of free digits had its origin at the point of amniote emergence during evolution.

scholarly journals Label-free cell based impedance measurements of ZnO nanoparticles—human lung cell interaction: a comparison with MTT, NR, Trypan blue and cloning efficiency assays

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Giuseppina Bozzuto ◽  
Giuseppe D’Avenio ◽  
Maria Condello ◽  
Simona Sennato ◽  
Ezio Battaglione ◽  
...  
Keyword(s):  
Trypan Blue ◽  
A549 Cells ◽  
Cell Interaction ◽  
Neutral Red ◽  
Free Cell ◽  
Mitogenic Activity ◽  
Label Free ◽  
Cloning Efficiency ◽  
Zno Nps ◽  
Experimental Conditions

Abstract Background There is a huge body of literature data on ZnOnanoparticles (ZnO NPs) toxicity. However, the reported results are seen to be increasingly discrepant, and deep comprehension of the ZnO NPs behaviour in relation to the different experimental conditions is still lacking. A recent literature overview emphasizes the screening of the ZnO NPs toxicity with more than one assay, checking the experimental reproducibility also versus time, which is a key factor for the robustness of the results. In this paper we compared high-throughput real-time measurements through Electric Cell-substrate Impedance-Sensing (ECIS®) with endpoint measurements of multiple independent assays. Results ECIS-measurements were compared with traditional cytotoxicity tests such as MTT, Neutral red, Trypan blue, and cloning efficiency assays. ECIS could follow the cell behavior continuously and noninvasively for days, so that certain long-term characteristics of cell proliferation under treatment with ZnO NPs were accessible. This was particularly important in the case of pro-mitogenic activity exerted by low-dose ZnO NPs, an effect not revealed by endpoint independent assays. This result opens new worrisome questions about the potential mitogenic activity exerted by ZnO NPs, or more generally by NPs, on transformed cells. Of importance, impedance curve trends (morphology) allowed to discriminate between different cell death mechanisms (apoptosis vs autophagy) in the absence of specific reagents, as confirmed by cell structural and functional studies by high-resolution microscopy. This could be advantageous in terms of costs and time spent. ZnO NPs-exposed A549 cells showed an unusual pattern of actin and tubulin distribution which might trigger mitotic aberrations leading to genomic instability. Conclusions ZnO NPs toxicity can be determined not only by the intrinsic NPs characteristics, but also by the external conditions like the experimental setting, and this could account for discrepant data from different assays. ECIS has the potential to recapitulate the needs required in the evaluation of nanomaterials by contributing to the reliability of cytotoxicity tests. Moreover, it can overcome some false results and discrepancies in the results obtained by endpoint measurements. Finally, we strongly recommend the comparison of cytotoxicity tests (ECIS, MTT, Trypan Blue, Cloning efficiency) with the ultrastructural cell pathology studies. Graphic Abstract

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