A monoclonal antibody against mouse oocyte cytoskeleton recognizing cytokeratin-type filaments

Development ◽  
1985 ◽  
Vol 90 (1) ◽  
pp. 197-209
Author(s):  
E. Lehtonen
Keyword(s):  
Epithelial Cells ◽  
Immunofluorescence Microscopy ◽  
Staining Pattern ◽  
Blastocyst Stage ◽  
Tissue Sections ◽  
Mouse Oocytes ◽  
Thick Filaments ◽  
Trophectoderm Cells ◽  
Early Embryos ◽  
Cytokeratin Filaments

Monoclonal antibodies were raised to detergent-extracted cytoskeleton preparations of mouse oocytes. In immunofluorescence microscopy, one of the antibodies, OCS-1, localizes exclusively to epithelial cells in frozen tissue sections, including various simple and stratified epithelia. The antibody decorates a keratin-type of fibrillax, vinblastine-resistant network in various cultured, epithelial-type cells, but not in myoid or fibroblastoid cells. In mouse oocytes and cleavage-stage embryos, the OCS-1 antibody gives a diffuse, spotty staining pattern. In blastocyst-stage embryos, the antibody reveals a keratin-type filamentous organization in the trophectoderm cells. In immunoelectron microscopy, the OCS-1 antibody decorates 10 nm-thick filaments, often identifiable as desmosome-attached tonofilaments, in detergent-treated trophectoderm cells. The antigen(s) recognized by the OCS-1 antibody is apparently present in, or closely associated with, cytokeratin filaments. In addition to mouse oocytes and early embryos, a wide variety of epithelial cells in various species seem to share this antigen(s). The present results suggest that at the early stages, the cytokeratin-related antigen(s) defined by the OCS-1 antibody are stored in a non-fibrillar form which is then converted into a fibrillar network at the blastocyst stage. A pre-existing supply of cytokeratin-related protein may be essential for the development of the blastocyst.

Disruption of the cytokeratin filament network in the preimplantation mouse embryo

Development ◽  
1988 ◽  
Vol 104 (2) ◽  
pp. 219-234
Author(s):  
J.A. Emerson
Keyword(s):  
Mouse Embryo ◽  
Blastocyst Stage ◽  
Cell Stage ◽  
Trophectoderm Cells ◽  
Preimplantation Mouse Embryos ◽  
Preimplantation Mouse Embryo ◽  
Preimplantation Mouse ◽  
Morula Stage ◽  
Cytokeratin Filaments

The distribution of the cytokeratin network in the intact preimplantation mouse embryo and the role of cytokeratin filaments in trophectoderm differentiation were investigated by means of whole-mount indirect immunofluorescence microscopy and microinjection of anti-cytokeratin antibody. Assembled cytokeratin filaments were detected in some blastomeres as early as the compacted 8-cell stage. The incidence and organization of cytokeratin filaments increased during the morula stage, although individual blastomeres varied in their content of assembled filaments. At the blastocyst stage, each trophectoderm cell contained an intricate network of cytokeratin filaments, and examination of sectioned blastocysts confirmed that extensive arrays of cytokeratin filaments were restricted to cells of the trophectoderm. Microinjection of anticytokeratin antibody into individual mural trophectoderm cells of expanded blastocysts resulted in a dramatic rearrangement of the cytokeratin network in these cells. Moreover, antibody injection into 2-cell embryos inhibited assembly of the cytokeratin network during the next two days of development. Despite this disruption of cytokeratin assembly, the injected embryos compacted and developed into blastocysts with normal morphology and nuclear numbers. These results suggest that formation of an elaborate cytokeratin network in preimplantation mouse embryos is unnecessary for the initial stages of trophectoderm differentiation resulting in blastocyst formation.

Cytoskeletal involvement in the modulation of cell-cell junctions by the protein kinase inhibitor H-7

1994 ◽  
Vol 107 (3) ◽  
pp. 683-692 ◽  
Author(s):  
S. Citi ◽  
T. Volberg ◽  
A.D. Bershadsky ◽  
N. Denisenko ◽  
B. Geiger
Keyword(s):  
Protein Kinase ◽  
Epithelial Cells ◽  
Immunofluorescence Microscopy ◽  
Kinase Inhibitor ◽  
Cytochalasin D ◽  
Extracellular Calcium ◽  
Protein Kinase Inhibitor ◽  
Madin Darby Canine Kidney ◽  
Darby Canine Kidney ◽  
Canine Kidney

The protein kinase inhibitor H-7 has been shown to block junction dissociation induced by low extracellular calcium in Madin Darby canine kidney epithelial cells (S. Citi, J. Cell Biol. (1992) 117, 169–178). To understand the basis of this effect, we have examined how H-7 affects the organization of junctions and the actin cytoskeleton in different types of epithelial cells in culture. Immunofluorescence microscopy showed that H-7 confers Ca2+ independence on cultured epithelial lens cells, which lack tight junctions and desmosomes but have microfilament-associated adherens junctions. In these cells, H-7 did not protect N-cadherin from trypsin digestion at low extracellular calcium, suggesting that H-7 does not stabilize the ‘active’ cadherin conformation. In cultured Madin Darby canine kidney cells, H-7 partially prevented the fall in transepithelial resistance induced by cytochalasin D, either alone or in conjunction with calcium chelators. Double-immunofluorescence microscopy showed that H-7 inhibits both the fragmentation of labeling for the tight junction protein cingulin and the condensation of actin into cytoplasmic foci induced by cytochalasin D. Taken together, these observations indicate that H-7 inhibits junction dissociation by affecting the contractility of the adherens junction-associated microfilaments following treatment with calcium chelators or cytochalasin D.

Synthesis and phosphorylation of uvomorulin during mouse early development

Development ◽  
1992 ◽  
Vol 115 (1) ◽  
pp. 313-318 ◽  
Author(s):  
M. Sefton ◽  
M.H. Johnson ◽  
L. Clayton
Keyword(s):  
Cell Adhesion ◽  
Early Development ◽  
Adhesion Molecule ◽  
Cell Adhesion Molecule ◽  
Blastocyst Stage ◽  
Embryonic Stage ◽  
Cell Stage ◽  
Mouse Oocytes ◽  
Maternal Mrna ◽  
Embryonic Genome

The cell adhesion molecule, uvomorulin, is synthesised in both the 135 × 10(3) M(r) precursor and 120 × 10(3) M(r) mature forms on maternal mRNA templates in unfertilized and newly fertilized mouse oocytes. Synthesis on maternal message ceases during the 2-cell stage to resume later on mRNA encoded presumptively by the embryonic genome. Uvomorulin is detectable by immunoblotting at all stages upto the blastocyst stage, but shows variations in its total amount and processing with embryonic stage. Whilst only trace levels of phosphorylated uvomorulin are detectable in early and late 4-cell embryos, uvomorulin in 8-cell embryos is phosphorylated.

Sialodacryoadenitis Virus-associated Lesions in the Lower Respiratory Tract of Rats

1986 ◽  
Vol 23 (3) ◽  
pp. 278-286 ◽  
Author(s):  
Z. W. Wojcinski ◽  
D. H. Percy
Keyword(s):  
Epithelial Cells ◽  
Respiratory Tract ◽  
Lower Respiratory Tract ◽  
Viral Antigen ◽  
Wistar Rats ◽  
Immunofluorescence Microscopy ◽  
Sendai Virus ◽  
Post Inoculation ◽  
Mycoplasma Pulmonis ◽  
Associated Lesions

Eight- to 10-week-old outbred Wistar rats were inoculated intranasally with 1029 medium mouse lethal infective doses of sialodacryoadenitis (SDA) virus. Sham inoculated control rats and challenged rats were killed at 1 day intervals for the first 8 days, then on days 10, 12, 14, and 20. Typical lesions associated with SDA were seen microscopically in the salivary and lacrimal glands of inoculated rats. In addition, laryngitis, tracheitis, bronchitis, bronchiolitis, and multifocal alveolitis were present during the acute stages of the disease. Viral antigen was demonstrated in epithelial cells lining airways by immunofluorescence microscopy. SDA virus was recovered from the lower respiratory tract from days 2 to 6 post-inoculation (PI). Serum antibodies to SDA virus, but not to Sendai virus or Mycoplasma pulmonis were present in rats tested at day 20 PI. These findings demonstrate that during the acute stages of the disease, significant lesions do occur in the lower respiratory tract of SDA virus-infected rats.

scholarly journals Involvement of R-cadherin in the early stage of glomerulogenesis.

1998 ◽  
Vol 9 (7) ◽  
pp. 1234-1241 ◽  
Author(s):  
S Goto ◽  
E Yaoita ◽  
H Matsunami ◽  
D Kondo ◽  
T Yamamoto ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Upper Limb ◽  
Immunofluorescence Microscopy ◽  
Early Stage ◽  
Capillary Loop ◽  
Tubular Cells ◽  
Glomerular Epithelial Cells ◽  
Ureteric Buds ◽  
Parietal Epithelial Cells ◽  
E Cadherin

The earliest commitment to the formation of glomeruli is recognizable in S-shaped bodies. Although cell-cell adhesion seems likely to play a crucial role in this process, how glomerular epithelial cells segregate from the other parts of the nephron is unknown. In this study, immunofluorescence microscopy and monoclonal antibodies specific for mouse R-, E-, P- and N-cadherins were used to examine which of these adhesion molecules are involved in glomerulogenesis of the mouse kidney. Weak R-cadherin staining was first found in the vesicle stage, becoming restricted to glomerular visceral epithelial cells (VEC) during the S-shaped body stage. The intensity of this staining became stronger in the capillary loop stage, whereas parietal epithelial cells (PEC) and tubular cells did not stain. In the maturing stage, VEC gradually lost their staining for R-cadherin. E-cadherin was detected in ureteric buds and the upper limb of S-shaped bodies. From the capillary loop to the maturing stage, anti-E-cadherin stained epithelial cells in all tubule segments, but no label was seen in VEC or PEC. P-cadherin was also stained in the ureteric buds and in the upper limb of S-shaped bodies. N-Cadherin was weakly stained in cells at the vesicle stage, but thereafter staining of N-cadherin was not detected at any stage of glomerular formation. Immunoelectron microscopy of differentiating VEC was performed using antibodies specific to alpha-catenin, which is associated with cadherin. Subsequently, immunogold particles identifying alpha-catenin were localized on junctions between primary processes of VEC. These findings indicate that R-cadherin is uniquely expressed in differentiating VEC, suggesting an important role in the early stages of glomerulogenesis.

Suppression of chromosome condensation during meiotic maturation induces parthenogenetic development of mouse oocytes

Development ◽  
1988 ◽  
Vol 104 (1) ◽  
pp. 97-103 ◽  
Author(s):  
H.J. Clarke ◽  
J. Rossant ◽  
Y. Masui
Keyword(s):  
Protein Synthesis ◽  
Blastocyst Stage ◽  
Chromosome Condensation ◽  
Meiotic Maturation ◽  
Dibutyryl Cyclic Amp ◽  
Developmental Arrest ◽  
Mouse Oocytes ◽  
Parthenogenetic Development ◽  
No Activation ◽  
Metaphase I

Mouse oocytes at metaphase I were treated with puromycin, which caused the chromosomes to become decondensed within an interphase nucleus. When the oocytes were allowed to resume protein synthesis, they returned to metaphase within 8–10 h and neither synthesized DNA nor cleaved, indicating that they had not been parthenogenetically activated by the puromycin treatment. However, when dibutyryl cyclic AMP was added to the medium after protein synthesis resumed, the oocytes remained in interphase. These oocytes maintained in interphase began DNA synthesis beginning 20 h after puromycin withdrawal, even though no activation stimulus had been given to them. After transfer to the oviducts of foster mothers, the oocytes could develop to the blastocyst stage. These results indicate that oocytes whose chromosomes were decondensed by puromycin treatment at metaphase I could begin parthenogenetic development in the absence of an activating stimulus, provided that they were prevented from returning to metaphase. In contrast, when the puromycin-treated oocytes were allowed to return to metaphase, they became developmentally arrested at the end of maturation. This suggests that the mechanism responsible for the developmental arrest of mature oocytes at metaphase II depends on cytoplasmic conditions that cause chromosome condensation to the metaphase state.

Localization of cytoskeletal proteins in preimplantation mouse embryos

Development ◽  
1980 ◽  
Vol 55 (1) ◽  
pp. 211-225
Author(s):  
E. Lehtonen ◽  
R. A. Badley
Keyword(s):  
Blastocyst Stage ◽  
Cytoskeletal Proteins ◽  
Mouse Embryos ◽  
Trophectoderm Cells ◽  
Apparent Concentration ◽  
Preimplantation Mouse ◽  
Early Mouse ◽  
Filament Protein

The immunofluorescence technique was used to detect the presence and distribution of actin, alpha-actinin, tubulin and 10 nm filament protein in early mouse embryos. Actin and alpha-actinin stainings showed a distinct concentration to a peripheral layer in the cleavage-stage blastomeres and in trophectoderm cells. Dots of fluorescence appeared in this cortical staining pattern. The distribution of tubulin staining in the blastomere cytoplasm was relatively even with apparent concentration at the perinuclear region and frequently at wide intercellular contact areas. 10 nm filament protein was distributed evenly in the blastomere cytoplasm without cortical concentration of the label. At the blastocyst stage, the trophectoderm cells in blastocyst outgrowths in vitro developed well organized cytoskeletons including both microfilament, microtubule and 10 nm filament elements. Comparable structures were not observed in blastocysts in vivo, or in late hatched blastocysts cultured in suspension. The morphogenetic significance of the observations is discussed.

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