Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 13 de 13
Filter
Add more filters










Publication year range
1.
J Cell Sci ; 104 ( Pt 3): 727-40, 1993 Mar.
Article in English | MEDLINE | ID: mdl-7686169

ABSTRACT

Rabbit esophageal epithelium, a parakeratinized stratified epithelium, synthesizes as one of its major differentiation products a keratin pair consisting of a basic K4 (59 kDa) and an acidic K13 (41 kDa) keratin. Although immunohistochemical staining data suggest that in esophageal epithelia of some other species these two keratins are suprabasally located, antigenic masking of the epitopes in the basal cells has not been ruled out. Using several well-characterized monoclonal antibodies including AE8, which specifically recognizes K13, coupled with biochemical analysis of keratins of basal and suprabasal cells isolated from confluent rabbit esophageal epithelial culture, we have obtained direct evidence that K4 and K13 keratins are largely absent in the undifferentiated basal cells, but are present in large amounts in suprabasal cells. We also show that in the cornified cell layers that are formed during the terminal stage of esophageal epithelial differentiation, K4 and K13 keratins become disulfide-crosslinked to form three different dimers. Two of them (110 kDa and 100 kDa) are heterodimers and consist of equimolar amounts of K4 and K13; they presumably represent isomers crosslinked via different cysteine residues. The third dimer (90 kDa) was found to be a homodimer of the acidic K13 keratin. Trypsinization experiment established that at least some of the disulfide crosslinks in the K4/K13 heterodimer must involve cysteine residues residing in the trypsin-resistant rod domains of keratins. Air-oxidation of in vitro reconstituted filaments reproduced the two heterodimers, which most likely involve the crosslinking between type I and type II keratins of different coiled coils. The formation of these disulfide-crosslinked keratin dimers, instead of higher molecular mass oligomers or polymers as occurring in the epidermis and hair, may contribute to the formation of cornified cells with a physical stability and rigidity that are optimal for esophageal function. Our data also suggest that interactions involved in the formation of homodimers, thought to be metastable and unimportant during the initial step of filament assembly (i.e. tetramer formation), may actually play an important role in stabilizing a higher order structure in mature keratin filaments.


Subject(s)
Disulfides/chemistry , Esophagus/metabolism , Keratins/chemistry , Animals , Antibodies, Monoclonal , Biopolymers , Cell Differentiation/physiology , Cells, Cultured , Epithelial Cells , Epithelium/metabolism , Esophagus/cytology , Immunoblotting , Keratins/analysis , Keratins/biosynthesis , Oxidation-Reduction , Rabbits
2.
Differentiation ; 42(2): 103-10, 1989 Dec.
Article in English | MEDLINE | ID: mdl-2483836

ABSTRACT

Cultured rabbit corneal epithelial cells undergo three distinct stages of growth and differentiation characterized by the sequential appearance of K5/K14 keratin markers for basal keratinocytes, K6/K16 keratin markers for "hyperproliferative" keratinocytes, and K3/K12 keratin markers for corneal-type differentiation. Analyses of [35S]methionine-labeled, newly synthesized keratins revealed that K6/K16 are synthesized only briefly when the cells undergo exponential growth, and their synthesis is suppressed when the cells reach confluence and switch to synthesizing K3/K12. Transient synthesis of K6/K16 was also observed in vivo during corneal epithelial regeneration. Although K6/K16 expression in general correlates well with cellular growth, drug-induced inhibition of corneal epithelial growth and related data on human epidermal keratinocytes indicate that these two events are dissociable. These results establish clearly for the first time a reciprocal relationship, on a protein level, between the synthesis of K6/K16 and a differentiation-related keratin pair, K3/K12. Such a relationship strongly suggests a competitive mechanism controlling the synthesis of these two major classes of keratins in the suprabasal compartment. Our results also indicate that although hyperproliferation is usually accompanied by K6/K16 expression, the reverse is not always true. Taken together, the data suggest that K6/K16 are synthesized, perhaps by default, as an alternative suprabasal keratin pair under conditions that are nonpermissive for keratinocytes to express their normal, differentiation-related keratin pairs.


Subject(s)
Cornea/physiology , Keratins/biosynthesis , Animals , Cell Differentiation , Cell Division , Cornea/cytology , Cornea/metabolism , Humans , Rabbits , Regeneration , Skin/cytology , Skin/metabolism , Skin Physiological Phenomena
3.
Exp Cell Res ; 181(1): 141-58, 1989 Mar.
Article in English | MEDLINE | ID: mdl-2465162

ABSTRACT

The dorsal surfaces of mammalian tongues are covered with numerous projections known as filiform papillae whose morphology varies in different species. Using a panel of monoclonal antibodies to keratins as probes, we have established that, in both human and mouse, the interpapillary epithelia express mainly the "esophageal-type" keratins, while the papillary epithelia express "skin-type" keratins as well as some keratins reacting with a monoclonal antibody (AE13) to hair keratins. The AE13-reactive proteins of the mouse were found to be very similar to those of authentic mouse hair keratins. However, the corresponding protein of human tongue appears to be different from all known human keratins. This protein has a MW of 51K; it is relatively acidic; it is sulfhydryl-rich, as revealed by iodoacetic acid-induced charge and apparent size shift; it shares an epitope with all the known acidic human hair keratins; and it is associated with keratin fibrils in vivo. This protein may therefore be regarded as a novel type I "hard" keratin. These data establish that mammalian dorsal tongue epithelia can be divided into at least three compartments that undergo mainly "esophageal-", "skin-" and "hair"-types of differentiation. Different keratin filaments, e.g., those of the esophageal- and hair-types, exhibit strikingly different degrees of lateral aggregation, which can potentially account for the different physical strength and rigidity of various cellular compartments. Our data also suggest the possibility that variations in papillary structure in human and mouse may arise from different spatial arrangements of specific keratinocytes, and/or from the expression of specialized hair-related keratins.


Subject(s)
Epidermis/analysis , Esophagus/analysis , Hair/analysis , Keratins/analysis , Tongue/analysis , Animals , Cell Differentiation , Epidermal Cells , Epithelium/analysis , Esophagus/cytology , Hair/cytology , Humans , Hydrogen-Ion Concentration , Keratins/biosynthesis , Keratins/immunology , Mice , Molecular Weight , Tongue/cytology , Tongue/metabolism
4.
J Autism Dev Disord ; 17(3): 315-32, 1987 Sep.
Article in English | MEDLINE | ID: mdl-3654485

ABSTRACT

This study investigated the effects of two levels of teacher intrusion upon the behavior of elementary age children with autism and nonhandicapped peers during dyadic play interactions occurring in two special education classrooms. High versus low levels of teacher intrusion were contrasted in a mixed between- and within-subjects design counterbalanced for order across the two conditions. There were few differences in behavior across the two conditions, though the low-intrusion condition was associated with higher levels of toy contact, appropriate and inappropriate play, and lower levels of spontaneous verbalizations by the students with autism. There was no difference in the occurrence of excess behavior by condition. Results are discussed with respect to future investigations of effective teacher mediation to prepare children for positive peer interactions.


Subject(s)
Autistic Disorder/psychology , Peer Group , Social Behavior , Child , Education of Intellectually Disabled , Female , Humans , Male , Professional-Patient Relations , Research Design
6.
Differentiation ; 34(1): 60-7, 1987.
Article in English | MEDLINE | ID: mdl-2440750

ABSTRACT

We have previously shown that a basic 64-kilodalton (no. 3 in the catalog of Moll et al.) and an acidic 55-kilodalton (no. 12) keratin are characteristic of suprabasal cell layers in cultured rabbit corneal epithelial colonies, and therefore may be regarded as markers for an advanced stage of corneal epithelial differentiation. Moreover, using an AE5 mouse monoclonal antibody, we showed that the 64-kilodalton keratin marker is expressed suprabasally in limbal epithelium but uniformly (basal layer included) in central corneal epithelium, suggesting that corneal basal cells are in a more differentiated state than limbal basal cells. In conjunction with previous data implicating the centripetal migration of corneal epithelial cells, our data support a model of corneal epithelial maturation in which corneal epithelial stem cells are located in the limbus, the transitional zone between the cornea and conjunctiva. In the present study, we analyzed the expression of the 64-kilodalton keratin in developing human corneal epithelium by immunohistochemical staining. At 8 weeks of gestation, the presumptive corneal epithelium is composed of a single layer of cuboidal cells with an overlying periderm; neither of these cell layers is AE5 positive. At 12-13 weeks of gestation, some superficial cells of the three- to four-layered epithelium become AE5 positive, providing the earliest sign of overt corneal epithelial differentiation. At 36 weeks, although the epithelium is morphologically mature (four to six layers), AE5 produces a suprabasal staining pattern, this being in contrast to the adult epithelium which exhibits uniform staining.(ABSTRACT TRUNCATED AT 250 WORDS)


Subject(s)
Cornea/growth & development , Keratins/biosynthesis , Aging , Antigens, Surface/analysis , Cornea/embryology , Cornea/ultrastructure , Epithelial Cells , Epithelium/metabolism , Female , Fetus , Fluorescent Antibody Technique , Humans , Infant , Infant, Newborn , Keratins/analysis , Microscopy, Electron , Molecular Weight , Pregnancy
7.
J Cell Biol ; 103(1): 49-62, 1986 Jul.
Article in English | MEDLINE | ID: mdl-2424919

ABSTRACT

In this paper we present keratin expression data that lend strong support to a model of corneal epithelial maturation in which the stem cells are located in the limbus, the transitional zone between cornea and conjunctiva. Using a new monoclonal antibody, AE5, which is highly specific for a 64,000-mol-wt corneal keratin, designated RK3, we demonstrate that this keratin is localized in all cell layers of rabbit corneal epithelium, but only in the suprabasal layers of the limbal epithelium. Analysis of cultured corneal keratinocytes showed that they express sequentially three major keratin pairs. Early cultures consisting of a monolayer of "basal" cells express mainly the 50/58K keratins, exponentially growing cells synthesize additional 48/56K keratins, and postconfluent, heavily stratified cultures begin to express the 55/64K corneal keratins. Cell separation experiments showed that basal cells isolated from postconfluent cultures contain predominantly the 50/58K pair, whereas suprabasal cells contain additional 55/64K and 48/56K pairs. Basal cells of the older, postconfluent cultures, however, can become AE5 positive, indicating that suprabasal location is not a prerequisite for the expression of the 64K keratin. Taken together, these results suggest that the acidic 55K and basic 64K keratins represent markers for an advanced stage of corneal epithelial differentiation. The fact that epithelial basal cells of central cornea but not those of the limbus possess the 64K keratin therefore indicates that corneal basal cells are in a more differentiated state than limbal basal cells. These findings, coupled with the known centripetal migration of corneal epithelial cells, strongly suggest that corneal epithelial stem cells are located in the limbus, and that corneal basal cells correspond to "transient amplifying cells" in the scheme of "stem cells----transient amplifying cells----terminally differentiated cells."


Subject(s)
Cornea/cytology , Keratins/metabolism , Animals , Antibodies, Monoclonal , Cell Differentiation , Cell Movement , Cells, Cultured , Epithelial Cells , Fluorescent Antibody Technique , Isoelectric Point , Keratins/genetics , Keratins/immunology , Mitosis , Molecular Weight , Rabbits , Stem Cells/cytology
10.
Differentiation ; 28(1): 30-5, 1984.
Article in English | MEDLINE | ID: mdl-6083891

ABSTRACT

Recent data have indicated that specific keratin molecules can provide useful markers for studying different types and stages of epithelial differentiation. To utilize these protein markers, however, it is important to establish the keratin nature of the molecules and identify unambiguously the individual keratin species. In this paper, we show that this can be done relatively easily by one- and two-dimensional gel electrophoresis combined with immunoblotting using three monoclonal antibodies (aIF, AE1, and AE3). The aIF antibody has previously been shown to crossreact with all classes of intermediate-filament proteins. Using one- and two-dimensional immunoblotting, we establish that this antibody recognizes all known epithelial keratins of human and rabbit, although the reaction is relatively strong for the larger, basic keratins and is relatively weak for some of the smaller, acidic keratins. In contrast, AE1 and AE3 monoclonal antibodies have previously been shown to be highly specific for the acidic and basic subfamilies of the keratins, respectively. The combined use of the broadly reacting aIF antibody and the subfamily-specific AE1 and AE3 monoclonal antikeratin antibodies should facilitate the immunological definition, identification, and classification of mammalian epithelial keratins.


Subject(s)
Cornea/analysis , Keratins/analysis , Skin/analysis , Animals , Antibodies, Monoclonal , Antigen-Antibody Complex , Cells, Cultured , Electrophoresis, Polyacrylamide Gel , Epithelium/analysis , Epitopes/analysis , Humans , Rabbits , Species Specificity
12.
J Cell Biol ; 85(3): 695-702, 1980 Jun.
Article in English | MEDLINE | ID: mdl-7190148

ABSTRACT

A model has been devised to study the in vitro formation of desmonsomes. The model is based on the differential labeling of two subpopulations of a desmosome-forming human cancer line (C4I). The labeled subpopulations are dispersed, preincubated separately on a shaking water bath for 24 h to allow the internalization of desmosome fragments and the repair of the cell surface, and then mixed, and allowed to aggregate. Aliquots of the mixed suspension are fixed at various intervals. The time between mixing and fixation represents the maximum age of any junction between dissimilarly labeled cells. The beginnings of desmosome formation were observed within a few minutes after the beginning of aggregation. Close apposition of cell membranes was seen immediately after mixing, followed within 15 min by the appearance of a submembrane density in one or both of the interacting cells. Intracytoplasmic filament formation takes place at between 15 and 30 min. Desmosome formation is complete by 90 min. The process is accompanied by a progressive widening of the extracellular space and the desification and organization of the extracellular material and the submembrane plaques.


Subject(s)
Cell Adhesion , Desmosomes/ultrastructure , Cell Differentiation , Cell Line , Cell Membrane/ultrastructure , Cells, Cultured , Cytoskeleton/ultrastructure , Humans , Microscopy, Electron , Models, Biological
SELECTION OF CITATIONS
SEARCH DETAIL
...