Patterns of expression of trichocytic and epithelial cytokeratins in mammalian tissues

Cells forming hair and nail material are characterized by the synthesis of members of a particular group of alpha-keratin polypeptides (trichocytic cytokeratins. "T cytokeratins") different from epithelial cytokeratins ("E cytokeratins"). As the precursor cells to trichocytes are derived from fetal epidermal keratinocytes expressing only E cytokeratins, we have studied the patterns of expression of both T and E cytokeratins in developing human hair-and nail-forming tissues of different fetal stages, by immunocytochemistry using antibodies specific for certain T or E cytokeratins and by two-dimensional gel electrophoresis and immunoblotting. In developing hair follicles up to the early bulbous-peg stage (weeks 12-15 of gestational age), only certain E but no T cytokeratins were identified. T cytokeratins were first detected in the late bulbous-peg stage (in week-14 scalp skin) in certain cells of the central part of the hair cone. In hair-producing follicles (weeks 18-25), the lower hair matrix cells were positive for certain E cytokeratins, whereas T cytokeratins appeared in the uppermost portion of the matrix and, most prominently, in the maturing trichocytes. From the late bulbous-peg stage on. E cytokeratin antibody Ks13.1 selectively decorated the inner root sheath. In finger nail "anlagen", T cytokeratins were detected first in week 12 and 13 fetuses, specifically in cells of the lunula region. In more-advanced stages of nail formation, expression of T cytokeratins extended not only to the upper layers of the ventral nail matrix but was also found, albeit more sparsely, in cells of the whole nail-bed epithelium. Throughout these developmental stages, coexpression of T and E cytokeratins was noted in certain cells, including E cytokeratin 19. While in earlier stages E cytokeratins 10/11, characteristic of epidermal-type cornification, were noted in different regions, including the superficial stratum of the nail bed epithelium, they were later restricted to the epithelium of the proximal nail fold. The results show that terminal trichocytic differentiation starts, both in ontogeny and during the steady growth of hairs and nails, in cells expressing E cytokeratins and that coexpression of E and T polypeptides occurs in both kinds of appendages. While in the hair follicle, the change to the exclusive synthesis of T cytokeratins appears to take place relatively abruptly and simply, the development of nail structures from the ventral nail matrix seems to be more gradual and is characterized by more-complex patterns of coexpression of both kinds of cytokeratins.     

Patterns of expression of trichocytic and epithelial cytokeratins in mammalian tissues II. Concomitant and mutually exclusive synthesis of trichocytic and epithelial cytokeratins in diverse human and bovine tissues (hair follicle, nail bed and matrix, lingual papilla, thymic reticulum)

The hair-forming cells (trichocytes) and the mature hair contain four major trichocytic cytokeratins from each of the subfamilies, basic (Hb1-4) and acidic (Ha1-4); these are related - but not identical - to the epithelial cytokeratins. Here we show, by biochemical methods and immunofluorescence microscopy using antibodies specific for either epithelial or trichocyte cytokeratins, that the same set of hair-type cytokeratins, including two newly identified minor components, designated Hax (type I) and Hbx (type II), are also expressed in cells forming nails, in the filiform papillae of the dorsal surface of human and bovine tongue, and, most surprisingly, in some cells of the epithelial reticulum of bovine and human thymus. By double-label immunofluorescence microscopy, we also show that the expression of the two subsets of cytokeratins, i.e., the epithelial and the trichocytic ones, is not necessarily mutually exclusive, but that certain cells of hair follicles, nail matrix and bed, lingual papillae, and the nonlymphoid cell system of the thymus contain both trichocytic and certain epithelial cytokeratins. This indicates that these cells coexpress representatives of both kinds of cytokeratin. Implications of these findings with respect to problems of regulatory control of cytokeratin synthesis in tissue development and differentiation, and the possible functional meaning of the occurrence of trichocytic cytokeratins in such histologically diverse tissues, are discussed.     

The complement of native α-keratin polypeptides of hair-forming cells: A subset of eight polypeptides that differ from epithelial cytokeratins

Living hair-forming cells (trichocytes) were obtained from basal portions of human, bovine and ovine hair-follicles, free from contaminations of root-sheath epithelia. Their intermediate filament (IF) cytoskeleton was studied by gel electrophoresis of the native, i.e. non-S-carboxymethylated polypeptides, by peptide-map analysis of the individual components, by reconstitution experiments and by immunological methods. The IF protein complement of trichocytes from all three species is characterized by a very similar set of eight highly conserved alpha-keratin polypeptides, comprising four members of the basic (type II; Mr 56,500-60,000) and four members of the acidic (type I; Mr 41,000-44,000) cytokeratin subfamily. None of these eight trichocyte alpha-keratin polypeptides, which form heterotypic complexes and IF in vivo and in vitro, is identical to any of the epithelial cytokeratins of the same species. All the trichocyte-specific cytokeratins are native polypeptides encoded by different mRNAs, as demonstrated by in vitro translation of hair follicle mRNA. The same polypeptides are also found in mature hairs, although with different patterns of modification. Our study provides the first analysis of the native unmodified alpha-keratin polypeptides of trichocytes and hairs and therefore allows a direct comparison of these with the epithelial cytokeratins and other IF proteins from the same species. These findings indicate that, during fetal hair-follicle formation, the differentiation of trichocytes from epithelial cells involves a complete cessation of the synthesis of epithelial cytokeratins and a marked induction of the synthesis of a complex set of trichocyte-specific cytokeratins.     

Changes in the Pattern of Cytokeratin Polypeptides in Epidermis and Hair Follicles During Skin Development in Human Fetuses

Abstract. The cytokeratin polypeptides of microdissected epidermis and hair follicles from human fetuses (from week 10 of pregnancy until birth) have been analysed by two-dimensional gel electrophoresis. Two-layered epidermis in 10-week fetuses contains major amounts of cytokeratin polypeptides typical of simple epithelia (components Nos. 8, 18, and 19 according to Moll et al. [31]). These cytokeratins are gradually reduced in their relative amounts and eventually disappear in the multilayered epidermis of later stages. At advanced stages of development, cytokeratins characteristic of adult epidermis are detected and finally predominate. These include the large and basic epidermal cytokeratin No. 1 (apparent molecular weight 68,000) which is already present in the three-layered epidermis of 13-week fetuses. Hair follicle germ cells of 13-week fetuses differ from fetal epidermal keratinocytes and show a very simple cytokeratin pattern, dominated by only two major polypeptides (Nos. 5 and 17). More developed hair follicles of 20-week fetuses have established a cytokeratin pattern similar to, but not identical with, that of hair follicles from adult skin. Different staining patterns obtained by indirect immunofluorescence microscopy using cytokeratin antibodies with different specificities suggest that, in three-layered epidermis, different cytokeratin patterns might exist in the specific cell layers. Such a differential location might explain the high complexity of polypeptide components found in fetal skin. Possible contributions of peridermal cytokeratins to this complex pattern of fetal epidermis are discussed.     

A subfamily of relatively large and basic cytokeratin polypeptides as defined by peptide mapping is represented by one or several polypeptides in epithelial cells

Epithelial cells contain a class of intermediate-sized filaments formed by proteins related to epidermal alpha-keratins ('cytokeratins'). Different epithelia can express different combinations of cytokeratin polypeptides widely varying in apparent mol. wt. (40 000-68 000) and isoelectric pH (5.0-8.5). We have separated, by two-dimensional gel electrophoresis, cytokeratin polypeptides from various tissues and cultured cells of man, cow, and rodents and examined their relatedness by tryptic peptide mapping. By this method, a subfamily of closely related cytokeratin polypeptides has been identified which comprises the relatively large (greater than or equal to mol. wt. 52 500 in human cells) and basic (pH greater than or equal to 6.0) polypeptides but not the smaller and acidic cytokeratins. In all species examined, the smallest polypeptide of this subfamily is cytokeratin A, which is widespread in many simple epithelia and is the first cytokeratin expressed during embryogenesis. This cytokeratin polypeptide subfamily is represented by at least one member in all epithelial and carcinoma cells examined, indicating that polypeptides of this subfamily serve an important role as tonofilament constitutents . Diverse stratified epithelia and tumours derived therefrom contain two or more polypeptides of this subfamily, and the patterns of expression in different cell types suggest that some polypeptides of this subfamily are specific for certain routes of epithelial differentiation.     

Diversity of cytokeratins. Differentiation specific expression of cytokeratin polypeptides in epithelial cells and tissues

Epithelial cells contain a cytoskeletal system of intermediate-sized (7 to 11 nm) filaments formed by proteins related to epidermal keratins (cytokeratins). Cytoskeletal proteins from different epithelial tissues (e.g. epidermis and basaliomas, cornea, tongue, esophagus, liver, intestine, uterus) of various species (man, cow, rat, mouse) as well as from diverse cultured epithelial cells have been analyzed by one and two-dimensional gel electrophoresis. Major cytokeratin polypeptides are identified by immunological cross-reaction and phosphorylated cytokeratins by [³²P]phosphate labeling in vivo.It is shown that different epithelia exhibit different patterns of cytokeratin polypeptides varying in molecular weights (range: 40,000 to 68,000) and electrical charges (isoelectric pH range: 5 to 8.5). Basic cytokeratins, which usually represent the largest cytokeratins in those cells in which they occur, have been found in all stratified squamous epithelia examined, and in a murine keratinocyte line (HEL) but not in hepatocytes and intestinal cells, and in most other cell cultures including HeLa cells. Cell type-specificity of cytokeratin patterns is much more pronounced than species diversity. Anatomically related epithelia can express similar patterns of cytokeratin polypeptides. Carcinomas and cultured epithelial cells often continue to synthesize cytokeratins characteristic of their tissue of origin but may also produce, in addition or alternatively, other cytokeratins. It is concluded: (1) unlike other types of intermediate-sized filaments, cytokeratin filaments are highly heterogeneous in composition and can contain basic polypeptides: (2) structurally indistinguishable filaments of the same class, i.e. cytokeratin filaments, are formed, in different epithelial cells of the same species, by different proteins of the cytokeratin family; (3) vertebrate genomes contain relatively large numbers of different cytokeratin genes which are expressed in programs characteristic of specific routes of epithelial differentiation; (4) individual cytokeratins provide tissue- or cell type-specific markers that are useful in the definition and identification of the relatedness or the origin of epithelial and carcinoma cells.     

Detection of a cytokeratin determinant common to diverse epithelial cells by a broadly cross-reacting monoclonal antibody.

A monoclonal antibody derived from a mouse immunized with bovine epidermal prekeratin has been characterized by its binding to cytoskeletal polypeptides separated by one- or two-dimensional gel electrophoresis and by immunofluorescence microscopy. This antibody (KG 8.13) binds to a determinant present in a large number of human cytokeratin polypeptides, notably some polypeptides (Nos. 1, 5, 6, 7, and 8) of the 'basic cytokeratin subfamily' defined by peptide mapping, as well as a few acidic cytokeratins such as the epidermis-specific cytokeratins Nos. 10 and 11 and the more widespread cytokeratin No. 18. This antibody reacts specifically with a wide variety of epithelial tissues and cultured epithelial cells, in agreement with previous findings that at least one polypeptide of the basic cytokeratin subfamily is present in all normal and neoplastic epithelial cells so far examined. The antibody also reacts with corresponding cytokeratin polypeptides in a broad range of species including man, cow, chick, and amphibia but shows only limited reactivity with only a few rodent cytokeratins. The value of this broad-range monoclonal antibody, which apparently recognizes a stable cytokeratin determinant ubiquitous in human epithelia, for the immunohistochemical identification of epithelia and carcinomas is discussed.     

Cytokeratins in normal lung and lung carcinomas. I. Adenocarcinomas, squamous cell carcinomas and cultured cell lines

The various epithelial cells of the lower respiratory tract and the carcinomas derived from them differ markedly in their differentiation characteristics. Using immunofluorescence microscopy and two-dimensional gel electrophoresis of cytoskeletal proteins from microdissected tissues we have considered whether cytokeratin polypeptides can serve as markers of cell differentiation in epithelia from various parts of the human and bovine lower respiratory tract. In addition , we have compared these protein patterns with those found in the two commonest types of human lung carcinoma and in several cultured lung carcinoma cell lines. By immunofluorescence microscopy, broad spectrum antibodies to cytokeratins stain all epithelial cells of the respiratory tract, including basal, ciliated, goblet, and alveolar cells as well as all tumor cells of adenocarcinomas and squamous cell carcinomas. However, in contrast, selective cytokeratin antibodies reveal cell type-related differences. Basal cells of the bronchial epithelium react with antibodies raised against a specific epidermal keratin polypeptide but not with antibodies derived from cytokeratins characteristic of simple epithelia. When examined by two-dimensional gel electrophoresis, the alveolar cells of human lung show cytokeratin polypeptides typical of simple epithelia (nos. 7, 8, 18 and 19) whereas the bronchial epithelium expresses, in addition, basic cytokeratins (no. 5, small amounts of no. 6) as well as the acidic polypeptides nos. 15 and 17. Bovine alveolar cells also differ from cells of the tracheal epithelium by the absence of a basic cytokeratin polypeptide. All adenocarcinomas of the lung reveal a "simple-epithelium-type" cytokeratin pattern (nos. 7, 8, 18 and 19). In contrast, squamous cell carcinomas of the lung contain an unusual complexity of cytokeratins. We have consistently found polypeptides nos. 5, 6, 8, 13, 17, 18 and 19 and, in some cases, variable amounts of cytokeratins nos. 4, 14 and 15. Several established cell lines derived from human lung carcinomas (SK-LU-1, Calu -1, SK-MES-1 and A-549) show a uniform pattern of cytokeratin polypeptides (nos. 7, 8, 18 and 19), similar to that found in adenocarcinomas. In addition, vimentin filaments are produced in all the cell lines examined, except for SK-LU-1.(ABSTRACT TRUNCATED AT 400 WORDS)     

Different keratin polypeptides in epidermis and other epithelia of human skin: a specific cytokeratin of molecular weight 46,000 in epithelia of the pilosebaceous tract and basal cell epitheliomas

Cytokeratin polypeptides of human epidermis, of epithelia microdissected from various zones of the pilosebaceous tract (outer root-sheath of hair follicle, sebaceous gland), and of eccrine sweat-glands have been separated by one- and two-dimensional gel electrophoresis and characterized by binding of cytokeratin antibodies and by peptide mapping. The epithelium of the pilosebaceous tract has three major keratin polypeptides in common with interfollicular epidermis (two basic components of mol wts 58,000 and 56,000 and one acidic polypeptide of mol wt 50,000); however, it lacks basic keratin polypeptides in the mol wt range of 64,000-68,000 and two acidic keratin-polypeptides of mol wts 56,000 and 56,500 and contains an additional characteristic acidic cytokeratin of mol wt 46,000. Another cytokeratin polypeptide of mol wt 48,000 that is prominent in hair-follicle epithelium is also found in nonfollicular epidermis of foot sole. Both epidermis and pilosebaceous tract are different from eccrine sweat-gland epithelium, which also contains two major cytokeratins of mol wts 52,500 and 54,000 (isoelectric at pH 5.8-6.1) and a more acidic cytokeratin of mol wt 40,000. A striking similarity between the cytokeratins of human basal-cell epitheliomas and those of the pilosebaceous tract has been found: all three major cytokeratins (mol wts 58,000; 50,000; 46,000) of the tumor cells are also expressed in hair-follicle epithelium. The cytokeratin of mol wt 46,000, which is the most prominent acidic cytokeratin in this tumor, is related, by immunological and peptide map criteria, to the acidic keratin-polypeptides of mol wts 48,000 and 50,000, but represents a distinct keratin that is also found in other human tumor cells such as in solid adamantinomas and in cultured HeLa cells. The results show that the various epithelia present in skin, albeit in physical and ontogenic continuity, can be distinguished by their specific cytokeratin-polypeptide patterns and that the cytoskeleton of basal-cell epitheliomas is related to that of cells of the pilosebaceous tract.     

Molecular characterization and expression of the stratification-related cytokeratins 4 and 15

A number of human cytokeratins are expressed during the development of stratified epithelia from one-layered polar epithelia and continue to be expressed in several adult epithelial tissues. For studies of the regulation of the synthesis of stratification-related cytokeratins in internal tissues, we have prepared cDNA and genomic clones encoding cytokeratin 4, as a representative of the basic (type II) cytokeratin subfamily and cytokeratin 15, as representative of the acidic (type I) subfamily, and determined their nucleotide sequences. The specific expression of mRNAs encoding these two polypeptides in certain stratified tissues and cultured cell lines is demonstrated by Northern blot hybridization. Hybridization in situ with antisense riboprobes and/or synthetic oligonucleotides shows the presence of cytokeratin 15 mRNA in all layers of esophagus, whereas cytokeratin 4 mRNA tends to be suprabasally enriched, although to degrees varying in different regions. We conclude that the expression of the genes encoding these stratification-related cytokeratins starts already in the basal cell layer and does not depend on vertical differentiation and detachment from the basal lamina. Our results also show that simple epithelial and stratification-related cytokeratins can be coexpressed in basal cell layers of certain stratified epithelia such as esophagus. Implications of these findings for epithelial differentiation and the formation of squamous cell carcinomas are discussed.     

Monoclonal antibodies to human cytokeratins: application to various epithelial and mesothelial cells

Immunohistological analysis of human tissue using monoclonal antibodies against cytokeratins, which are confined to cells of epithelial origin, is a valuable technique. Using human epidermal keratins as antigen, we prepared monoclonal antibodies against cytokeratins (ZK1, ZK7, ZK61 and ZK99) and against a desmosomal protein (ZK31). Immunohistochemical staining of human skin sections using these antibodies showed a specific reaction with the epidermis: ZK1 stained the entire epidermis, ZK7 only the basal layer, ZK61 and ZK99 the suprabasal layers, and ZK31 the cellular interfaces. In order to test for antibody specificity, immunoblots with human epidermal and amnion epithelial cytokeratin polypeptides, as well as immunofluorescence microscopy of simple epithelia (glandular and simple columnar epithelia) were performed. ZK1, ZK61 and ZK99 reacted preferentially with cytokeratin polypeptides of stratified squamous epithelia and ZK7 recognized cytokeratins of stratified and simple epithelia. When the ZK antibodies were tested on mesothelial cells in pleural effusions, only ZK7 reacted with these cells. Biochemical analysis of cytokeratin accumulation in cells of primary and long-term cultures indicated that the cytokeratin pattern of mesothelial cells was quite unstable, while that of amnion epithelial cells showed only minor quantitative changes. The use of these antibodies to determine the epithelial origin of cells present in pleural effusions is proposed.     

Cell type heterogeneity of cytokeratin expression in complex epithelia and carcinomas as demonstrated by monoclonal antibodies specific for cytokeratins nos. 4 and 13

Three monoclonal antibodies, 1C7, 2D7 and 6B10, directed against cytokeratins of human esophagus were isolated and characterized by one- and two-dimensional gel electrophoresis and by immunohistochemical staining on sections of human epithelial tissues. In immunoblot experiments, antibodies of clones 1C7 (IgG2a) and 2D7 (IgG2b) react only with cytokeratin no. 13 of the acidic (type I) subfamily of cytokeratin polypeptides (Mr 54000; pI 5.1); antibodies of clone 6B10 (IgG1) detect only cytokeratin no. 4 (Mr 59000; pI 7.3) of the basic (type II) cytokeratin subfamily and allows the detection of this protein and possible degradation products at high sensitivity. In immunohistochemical staining all three antibodies stain non-cornifying squamous epithelium (e.g., tongue, esophagus, anus) and transitional epithelium of the bladder. Antibodies of clone 6B10 also stain cells in certain ciliated pseudostratified epithelia and ductal epithelia of various exocrine glands. These monoclonal antibodies are the first examples of antibodies specific for individual cytokeratin polypeptides characteristic of certain complex epithelia. They allow the identification of distinct minor populations of cells present in certain complex and glandular epithelia and in tumors derived therefrom which hitherto have not been distinguished. The possible reasons for the occurrence of cell type heterogeneity of cytokeratin expression in complex epithelia and in some carcinomas are discussed.     

Changes in the distribution of intermediate filament proteins and collagen IV in fetal and adult human pancreas. I. Localization of cytokeratin polypeptides

The expression patterns of individual cytokeratin polypeptides in foetal and adult human pancreatic tissues were examined using monoclonal antibodies. We demonstrated that human pancreatic epithelia in early stages of development (14 weeks of gestation) contain cytokeratins 7, 8, 18 and 19, which are typical of simple epithelia, as well as cytokeratin 4 and 17, which are characteristic of stratified epithelia. In the pancreatic ducts, most of these cytokeratins appeared to be expressed together. Cytokeratins 1, 5, 10, 13, 16 and 20 were not detectable. In contrast, the pancreatic parenchyma was only positive for cytokeratins 8 and 18, except a transient expression of cytokeratins 7 and 19 in pancreatic islets and acinar cells during the foetal development. A focal cytokeratin 7 staining of single acinar cells was seen in newborn and in adult islets. In the stromal tissue, vascular smooth muscle cells were partly reactive with cytokeratin 8 and 18 specific antibodies. The results are discussed in the light of differentiation-dependent changes in the expression of individual cytokeratin polypeptides in developing epithelia.     

Expression of simple epithelial type cytokeratins in stratified epithelia as detected by immunolocalization and hybridization in situ

Multi-layered ("stratified") epithelia differ from one-layered ("simple") polar epithelia by various architectural and functional properties as well as by their cytoskeletal complements, notably a set of cytokeratins characteristic of stratified tissue. The simple epithelial cytokeratins 8 and 18 have so far not been detected in any stratified epithelium. Using specific monoclonal antibodies we have noted, in several but not all samples of stratified epithelia, including esophagus, tongue, exocervix, and vagina, positive immunocytochemical reactions for cytokeratins 8, 18, and 19 which in some regions were selective for the basal cell layer(s) but extended into suprabasal layers in others. In situ hybridization with different probes (riboprobes, synthetic oligonucleotides) for mRNAs of cytokeratin 8 on esophageal epithelium has shown, in extended regions, relatively strong reactivity for cytokeratin 8 mRNA in the basal cell layer. In contrast, probes to cytokeratin 18 have shown much weaker hybridization which, however, was rather evenly spread over basal and suprabasal strata. These results, which emphasize the importance of in situ hybridization in studies of gene expression in complex tissues, show that the genes encoding simple epithelial cytokeratins can be expressed in stratified epithelia. This suggests that continual expression of genes coding for simple epithelial cytokeratins is compatible with the formation of squamous stratified tissues and can occur, at least in basal cell layers, simultaneously with the synthesis of certain stratification-related cytokeratins. We also emphasize differences of expression and immunoreactivity of these cytokeratins between different samples and in different regions of the same stratified epithelium and discuss the results in relation to changes of cytokeratin expression during fetal development of stratified epithelia, in response to environmental factors and during the formation of squamous cell carcinomas.     

Cytokeratin cytoskeleton in the differentiating ovarian follicle of the lizard Podarcis sicula Raf

By immunoblotting and immunocytochemical techniques, we characterized the cytokeratins previously localized by us in the previtellogenic ovarian follicle of Podarcis sicula. Our results show that these cytokeratins correspond to those expressed in the monolayered epithelia. In fact, the immunoblotting analysis showed that the NCL-5D3 antibody, specific for human low molecular weight cytokeratins expressed in monolayered epithelia, reacted with the cytokeratins extracted both from the ovary and from the monolayered intestinal mucosa of Podarcis sicula. Furthermore, this antibody, in this reptile as in humans, clearly immunolabeled sections of corresponding tissues. The organization of the cytokeratin cytoskeleton in the main steps of the ovarian follicle differentiation was also clarified. The reported observations suggest that in Podarcis sicula, the cytokeratin cytoskeleton is absent in the early oocytes. It first appears in the growing oocytes as a thin cortical layer in concomitance with its becoming visible also in the enlarging follicle cells. In the larger follicles, this cytoskeleton appears well organized in intermediate cells and in particular in fully differentiated pyriform cells. In both these cells a cytokeratin network connects the cytoplasm to the oocyte cortex through intercellular bridges. At the end of the previtellogenic oocyte growth, the intense immunolabeling of the apex in the regressing pyriform cells suggests that the cytokeratin, as other cytoplasmic components, may be transferred from these follicle cells to the oocyte. At the end of the oocyte growth, in the larger vitellogenic oocytes surrounded by a monolayer of follicle cells, the cytokeratin constitutes a heavily immunolabeled cortical layer thicker than in the previous stages. Mol. Reprod. Dev. 48:536–542, 1997. © 1997 Wiley-Liss, Inc.     

Cytokeratin polypeptide patterns of different epithelia of the human male urogenital tract: immunofluorescence and gel electrophoretic studies

Intermediate filament proteins of normal epithelia of the human and the bovine male urogenital tract and of certain human renal and bladder carcinomas have been studied by immunofluorescence microscopy and by two-dimensional gel electrophoresis of cytoskeletal fractions from microdissected tissue samples. The patterns of expression of cytokeratin polypeptides differ in the various epithelia. Filaments of a cytokeratin nature have been identified in all true epithelial cells of the male urogenital tract, including renal tubules and rete testis. Simple epithelia of renal tubules and collecting ducts of kidney, as well as rete testis, express only cytokeratin polypeptides nos. 7, 8, 18, and 19. In contrast, the transitional epithelia of renal pelvis, ureter, bladder, and proximal urethra contain, in addition to those polypeptides, cytokeratin no. 13 and small amounts of nos. 4 and 5. Most epithelia lining the human male reproductive tract, including those in the epididymis, ductus deferens, prostate gland, and seminal vesicle, synthesize cytokeratin no. 5 in addition to cytokeratins nos. 7, 8, 18, and 19 (cytokeratin no. 7 had not been detected in the prostate gland). Cytokeratin no. 17 has also been identified, but in very low amounts, in seminal vesicle and epididymis. The cytokeratin patterns of the urethra correspond to the gradual transition of the pseudostratified epithelium of the pars spongiosa (cytokeratins nos. 4, 5, 6, 13, 14, 15, and 19) to the stratified squamous epithelium of the fossa navicularis (cytokeratins nos. 5, 6, 10/11, 13, 15, and 19, and minor amounts of nos. 1 and 14). The noncornified stratified squamous epithelium of the glans penis synthesizes cytokeratin nos. 1, 5, 6, 10/11, 13, 14, 15, and 19. In immunofluorescence microscopy, selective cytokeratin antibodies reveal differential staining of different groups or layers of cells in several epithelia that may relate to the specific expression of cytokeratin polypeptides. Human renal cell carcinomas show a simple cytokeratin pattern consisting of cytokeratins nos. 8, 18, and 19, whereas transitional cell carcinomas of the bladder reveal additional cytokeratins such as nos. 5, 7, 13, and 17 in various proportions. The results shows that the wide spectrum of histological differentiation of the diverse epithelia present in the male urogenital tract is accompanied by pronounced changes in the expression of cytokeratin polypeptides and suggest that tumors from different regions of the urogenital tract may be distinguished by their cytokeratin complements.     

Changes in the distribution of intermediate filament proteins and collagen IV in fetal and adult human pancreas

Summary The expression patterns of individual cytokeratin polypeptides in foetal and adult human pancreatic tissues were examined using monoclonal antibodies. We demonstrated that human pancreatic epithelia in early stages of development (14 weeks of gestation) contain cytokeratins 7, 8, 18 and 19, which are typical of simple epithelia, as well as cytokeratin 4 and 17, which are characteristic of stratified epithelia. In the pancreatic ducts, most of these cytokeratins appeared to be expressed together. Cytokeratins 1, 5, 10, 13, 16 and 20 were not detectable. In contrast, the pancreatic parenchyma was only positive for cytokeratins 8 and 18, except a transient expression of cytokeratins 7 and 19 in pancreatic islets and acinar cells during the foetal development. A focal cytokeratin 7 staining of single acinar cells was seen in newborn and in adult islets. In the stromal tissue, vascular smooth muscle cells were partly reactive with cytokeratin 8 and 18 specific antibodies. The results are discussed in the light of differentiation-dependent changes in the expression of individual cytokeratin polypeptides in developing epithelia.     

Pair formation and promiscuity of cytokeratins: formation in vitro of heterotypic complexes and intermediate-sized filaments by homologous and heterologous recombinations of purified polypeptides

Cytokeratins are expressed in different types of epithelial cells in certain combinations of polypeptides of the acidic (type I) and basic (type II) subfamilies, showing "expression pairs." We have examined in vitro the ability of purified and denatured cytokeratin polypeptides of human, bovine, and rat origin to form the characteristic heterotypic subunit complexes, as determined by various electrophoretic techniques and chemical cross-linking, and, subsequently, intermediate-sized filaments (IFs), as shown by electron microscopy. We have found that all of the diverse type I cytokeratin polypeptides examined can form complexes and IFs when allowed to react with equimolar amounts of any of the type II polypeptides. Examples of successful subunit complex and IF formation in vitro include combinations of polypeptides that have never been found to occur in the same cell type in vivo, such as between epidermal cytokeratins and those from simple epithelia, and also heterologous combinations between cytokeratins from different species. The reconstituted complexes and IFs show stability properties, as determined by gradual "melting" and reassociation, that are similar to those of comparable native combinations or characteristic for the specific new pair combination. The results show that cytokeratin complex and IF formation in vitro requires the pairing of one representative of each the type I and type II subfamilies into the heterotypic tetramer but that there is no structural incompatibility between any of the members of the two subfamilies. These findings suggest that the co-expression of specific pair combinations observed in vivo has other reasons than general structural requirements for IF formation and probably rather reflects the selection of certain regulatory programs of expression during cell differentiation. Moreover, the fact that certain cytokeratin polypeptide pairs that readily form complexes in vitro and coexist in the same cells in vivo nevertheless show preferential, if not exclusive, partner relationships in the living cell points to the importance of differences of stabilities among cytokeratin complexes and/or the existence of extracytokeratinous factors involved in the specific formation of certain cytokeratin pairs.     

Expression of cytokeratin polypeptides during development of the rat inner ear.

The expression of cytokeratin polypeptides in the different epithelia of the developing inner ear of the rat from 12 days post conception to 20 days after birth was analysed immunohistochemically, using a panel of monoclonal antibodies. Throughout the development of the complex epithelial lining of the inner ear originating from the otocyst epithelium, only cytokeratins which are typical of simple epithelia were expressed. Cytokeratins 8, 18, and 19 were detectable shortly after the formation of the otocyst from the ectoderm (12 dpc), whereas cytokeratin 7 expression was delayed and first appeared in the vestibular portion and subsequently in the developing cochlear duct. During the development of the different types of specialized cells, differentiation-dependent modulation of the cytokeratin expression patterns was observed. In the mature inner ear, the specialized cell types displayed a function-related cytokeratin expression profile, both in the cochlear and vestibular portion. Cytokeratin expression in the flat epithelium of the vestibular portion suggests a more complex composition of this epithelium than has been established from routine morphology. Remarkably, the cochlear sensory cells were apparently devoid of cytokeratins, but no final conclusion could be drawn on the presence of cytokeratins in the sensory cells of the vestibular portion, because of the difficulty to delineate the cell borders between sensory cells and supporting cells.     

Cytokeratin expression in human thymus: immunohistochemical mapping

Cytokeratin expression in normal postnatal human thymus was studied immunohistochemically by using monoclonal antibodies against various cytokeratin polypeptides. An attempt was made to characterize cell populations giving rise to the cornified structures of Hassal's corpuscles. Monoclonal antibody KB-37, a marker of squamous epithelium basal cells, was applied to distinguish the earliest cells capable of undergoing squamous differentiation. Parts of the subcapsular epithelium were extensively stained with this reagent. This epithelium, like the basal layer of certain squamous epithelia, exibited a high incidence of cytokeratins 13 and 14, and pronounced expression of cytokeratin 19. Simple epithelium cytokeratins 8, 18, and 19 were present in the cortex. Scattered cells reacted with KB-37 antibody. All stellate epithelial cells in the medulla were positive for cytokeratin 19. Most of the medullar epithelial cells were positive for cytokeratins 13, 14 and 17 of complex epithelium, in contrast to the cortex, where only a few cells were positive for these cytokeratins. A significant proportion of the medullar cells was positive for KB-37 antigen. Cytokeratins 8 and 18 were expressed in single cells and in groups of cells surrounding Hassal's corpuscles. The outermost cells of these corpuscles were positive for cytokeratin 19 and KB-37. In the peripheral parts of Hassal's corpuscles, simple epithelium cytokeratins 7, 8, 18, and cytokeratins 4, 13, 14, and 17, characteristic of stratified nonkeratinizing epithelia, were coexpressed with keratinization-specific cytokeratins 10/11. The inner parts of the swirls were uniformly positive for cytokeratins was reduced.