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.

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.

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.

Cell death and the development of limb form and skeletal pattern in normal and wingless (ws) chick embryos

Development ◽  
1973 ◽  
Vol 30 (3) ◽  
pp. 753-772
Author(s):  
J. R. Hinchliffe ◽  
D. A. Ede
Keyword(s):  
Cell Death ◽  
Phenotypic Expression ◽  
Mesenchymal Cell ◽  
Limb Bud ◽  
Death Process ◽  
Electron Microscopic ◽  
Wide Range ◽  
Skeletal Pattern ◽  
Autoradiographic Analysis ◽  
Mutant Phenotypes

The wingless condition resulting from the action of the sex-linked wingless (ws) gene arises from the precocious appearance of cell death in the anterior necrotic zone (ANZ) of the forelimb-bud at stage 19 (3 days) and its progressive extension beyond its normal area during stages 20–23. A similar though less pronounced effect occurs in the hindlimb-bud. Although some wingless hindlimb-buds are normal, others are affected by the precocious appearance of cell death in the ANZ. The ws wingless mutant resembles the different wingless mutant investigated by Zwilling (1956) in that the apical ectodermal ridge (AER) is absent in most ws wing-buds. AER absence could be due to ws mesenchymal cell death interfering with the production of apical ectodermal maintenance factor (AEMF), which Zwilling claims is necessary to maintain the AER which plays an essential role in inducing limb outgrowth. Wingless mutant phenotypes range from birds with rudimentary wings and normal legs through a modal type with forelimbs absent and hindlimbs normal to wingless and legless forms showing a high degree of expressivity. Individual wingless embryos vary in the degree to which the precocious ANZ appearing at 3 days is extended into the limb-bud and the wide range of wingless phenotypic expression is attributed to this variation. Electron microscopic and histochemical analysis of the cell death process in wingless wing-buds revealed the presence of both isolated dead cells and macrophages, which contained intense acid phosphatase activity. These findings are interpreted as showing that isolated dead cells are ingested by neighbouring mesenchymal cells which thus become transformed into macrophages, first ingesting and then digesting further dead cells. A study was made of the origin of the anomalous hindlimb condition, including absence or reduction of the tibia and digits, characteristic of severely affected wingless embryos. Autoradiographic analysis of the pattern of 35SO4 uptake revealed that at stage 24/5 (4½ days) wingless hindlimb-buds which were smaller than normal had a normal prospective fibula region, but that the prospective tibia region was small or absent. Thus the effect of a precocious hindlimb ANZ at stages 19–22 is to reduce or delete the pre-axial prospective tibia at stage 24/5.

The BMP antagonist Gremlin regulates outgrowth, chondrogenesis and programmed cell death in the developing limb

Development ◽  
1999 ◽  
Vol 126 (23) ◽  
pp. 5515-5522 ◽  
Author(s):  
R. Merino ◽  
J. Rodriguez-Leon ◽  
D. Macias ◽  
Y. Ganan ◽  
A.N. Economides ◽  
...  
Keyword(s):  
Cell Death ◽  
Soft Tissue ◽  
Programmed Cell Death ◽  
Limb Development ◽  
Limb Bud ◽  
Central Core ◽  
Limb Outgrowth ◽  
Bmp Antagonists ◽  
And Function ◽  
Bmp Antagonist

In this study, we have analyzed the expression and function of Gremlin in the developing avian limb. Gremlin is a member of the DAN family of BMP antagonists highly conserved through evolution able to bind and block BMP2, BMP4 and BMP7. At early stages of development, gremlin is expressed in the dorsal and ventral mesoderm in a pattern complementary to that of bmp2, bmp4 and bmp7. The maintenance of gremlin expression at these stages is under the control of the AER, ZPA, and BMPs. Exogenous administration of recombinant Gremlin indicates that this protein is involved in the control of limb outgrowth. This function appears to be mediated by the neutralization of BMP function to maintain an active AER, to restrict the extension of the areas of programmed cell death and to confine chondrogenesis to the central core mesenchyme of the bud. At the stages of digit formation, gremlin is expressed in the proximal boundary of the interdigital mesoderm of the chick autopod. The anti-apoptotic influence of exogenous Gremlin, which results in the formation of soft tissue syndactyly in the chick, together with the expression of gremlin in the duck interdigital webs, indicates that Gremlin regulates the regression of the interdigital tissue. At later stages of limb development, gremlin is expressed in association with the differentiating skeletal pieces, muscles and the feather buds. The different expression of Gremlin in relation with other BMP antagonists present in the limb bud, such as Noggin, Chordin and Follistatin indicates that the functions of BMPs are regulated specifically by the different BMP antagonists, acting in a complementary fashion rather than being redundant signals.

scholarly journals Frequent amplification of HDAC genes and efficacy of HDAC inhibitor chidamide and PD-1 blockade combination in soft tissue sarcoma

2021 ◽  
Vol 9 (2) ◽  
pp. e001696
Author(s):  
Yi Que ◽  
Xiao-Long Zhang ◽  
Ze-Xian Liu ◽  
Jing-Jing Zhao ◽  
Qiu-Zhong Pan ◽  
...  
Keyword(s):  
Cell Death ◽  
Soft Tissue ◽  
Soft Tissue Sarcoma ◽  
Programmed Cell Death ◽  
Gene Family ◽  
Murine Model ◽  
Immune Checkpoint ◽  
Drug Target ◽  
Class I ◽  
Mechanistic Study

BackgroundThe advent of immune checkpoint therapy has been a tremendous advance in cancer treatment. However, the responses are still insufficient in patients with soft tissue sarcoma (STS). We aimed to identify rational combinations to increase the response to immune checkpoint therapy and improve survival.MethodsWhole-exome sequencing (WES) was performed in 11 patients with liposarcoma. Somatic copy number alterations (SCNAs) were analyzed at the gene level to identify obvious amplification patterns in drug-target genes. The expression and prognostic value of class I histone deacetylases (HDACs) was evaluated in 49 patients with sarcoma in our center and confirmed in 263 sarcoma samples from The Tumor Cancer Genome Atlas (TCGA) database. Q-PCR, flow cytometry and RNA-seq were performed to determine the correlations between class I HDACs, chidamide and PD-L1 in vitro and in vivo. The efficacy of combining chidamide with PD-1 blockade was explored in an immunocompetent murine model and a small cohort of patients with advanced sarcoma. Western blot, ChIP assay and dual luciferase assessment were applied in the mechanistic study.ResultsThe HDAC gene family was frequently amplified in STS. SCNAs in the HDAC gene family were extensively amplified in 8 of 11 (73%) patients with liposarcoma, based on a drug-target gene set, and we verified amplification in 76.65% (197/257) of cases by analyzing TCGA sarcoma cohort. Class I HDAC expression is associated with a poor prognosis for patients with STS, and its inhibition is responsible for promoting apoptosis and upregulating of programmed cell death ligand 1 (PD-L1). The HDAC class I inhibitor chidamide significantly increases PD-L1 expression, increased the infiltration of CD8+ T cells and reduced the number of MDSCs in the tumor microenvironment. The combination of chidamide with an anti-PD-1 antibody significantly promotes tumor regression and improves survival in a murine model. Moreover, chidamide combined with the anti-PD-1 antibody toripalimab is effective in patients with advanced and metastatic sarcoma, and the side effects are tolerable. Mechanistically, chidamide increases histone acetylation at the PD-L1 gene through the activation of the transcriptional factor STAT1.ConclusionsThe combination of chidamide and anti-programmed cell death 1 (PD-1) therapy represents a potentially important strategy for STS.

Limb deformity proteins: role in mesodermal induction of the apical ectodermal ridge

Development ◽  
1997 ◽  
Vol 124 (1) ◽  
pp. 133-139 ◽  
Author(s):  
J. Kuhlman ◽  
L. Niswander
Keyword(s):  
Sonic Hedgehog ◽  
Limb Development ◽  
Apical Ectodermal Ridge ◽  
Primary Defect ◽  
Limb Deformity ◽  
Normal Morphology ◽  
Fibroblast Growth Factor 8 ◽  
Skeletal Pattern ◽  
Two Phases ◽  
And Function

During early limb development, distal tip ectoderm is induced by the underlying mesenchyme to form the apical ectodermal ridge. Subsequent limb growth and patterning depend on reciprocal signaling between the mesenchyme and ridge. Mice that are homozygous for mutations at the limb deformity (ld) locus do not form a proper ridge and the anteroposterior axis of the limb is shortened. Skeletal analyses reveal shortened limbs that involve loss and fusion of distal bones and digits, defects in both anteroposterior and proximodistal patterning. Using molecular markers and mouse-chick chimeras we examined the ridge-mesenchymal interactions to determine the origin of the ld patterning defects. In the ld ridge, fibroblast growth factor 8 (Fgf8) RNA is decreased and Fgf4 RNA is not detected. In the ld mesenchyme, Sonic hedgehog (Shh), Evx1 and Wnt5a expression is decreased. In chimeras between ld ectoderm and wild-type mesenchyme, a ridge of normal morphology and function is restored, Fgf8 and Shh are expressed normally, Fgf4 is induced and a normal skeletal pattern arises. These results suggest that the ld mesenchyme is unable to induce the formation of a completely functional ridge. This primary defect causes a disruption of ridge function and subsequently leads to the patterning defects observed in ld limbs. We propose a model in which ridge induction requires at least two phases: an early competence phase, which includes induction of Fgf8 expression, and a later differentiation phase in which Fgf4 is induced and a morphological ridge is formed. Ld proteins appear to act during the differentiation phase.

scholarly journals Cdc42 is required for chondrogenesis and interdigital programmed cell death during limb development

2012 ◽  
Vol 129 (1-4) ◽  
pp. 38-50 ◽  
Author(s):  
Ryo Aizawa ◽  
Atsushi Yamada ◽  
Dai Suzuki ◽  
Tadahiro Iimura ◽  
Hidetoshi Kassai ◽  
...  
Keyword(s):  
Cell Death ◽  
Programmed Cell Death ◽  
Limb Development
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