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Biomedical subjects

I Thesleff

Publications and source records attributed to I Thesleff.

At least 145 records · Page 8Linked to original sources
PubMed1986

Dental papilla cells in culture. Comparison of morphology, growth and collagen synthesis with two other dental-related embryonic mesenchymal cell populations.

The dental papilla is a mesenchymal cell condensation which plays an important regulatory role during tooth development. Dental papilla mesenchymes were enzymatically separated from the dental epithelia from tooth germs of 17-day-old mouse embryos and disaggregated for monolayer culture. These cells were compared with gingival mesenchyme overlying the same tooth germs and with undifferentiated jaw mesenchyme from mandibles of 11-day-old embryos. The dental papilla cells were large and flat with numerous cell processes, whereas the gingival cells resembled typical spindle-shaped fibroblasts and grew to a higher cell density. Although the two mesenchymes differ in their collagen contents in vivo, no differences were detected either in the amount or type of collagen synthesized in vitro. Type I and III collagens were found in the culture media and type V collagen in the cell layer of both cell populations. The mandibular mesenchymal cells of the younger embryos resembled the dental papilla cells in morphology and growth rate. This may reflect retention of undifferentiated embryonic characteristics in the dental papilla. The successful culture of dental papilla cells now enables subsequent studies on the cellular properties related to the unique morphogenetic capabilities of these cells.

Animals↗
PubMed1986

The relationship between the keratocyst antigen (KCA) and keratin.

The relationship of the keratocyst antigen (KCA), the soluble component present in most keratocyst fluids, and keratin, was studied with immunofluorescence microscopy comparing their distribution in developing mouse embryonic teeth and in human ameloblastomas. In these tissues both molecules showed a strong codistribution in epithelial cells. In the embryonic teeth both molecules were present in the stratum intermedium cells between the stellate reticulum cells and ameloblasts, but the secretory ends of the ameloblasts showed fluorescent staining only for keratin. The relationship was further investigated by comparing the physicochemical characteristics of KCA and keratin. Results on immunoblotting and two-dimensional gel electrophoresis showed that KCA existed in keratocyst fluid as a 60-68,000 dalton polypeptide with an isoelectric point of pI 6.8. Immunoblotting analysis of various isolated keratins revealed a typical polypeptide pattern of each keratin when anti-KCA antiserum was used for staining. These findings suggest that KCA and keratin are related molecules and that KCA may be a soluble component of keratin.

Ameloblastoma↗
PubMed1986

Development of mouse embryonic molars in vitro: an attempt to design defined culture conditions allowing mineralization.

Mandibular first molars from 17 day old mouse embryos were cultivated in vitro for 14 days in BGJb-medium supplemented with 20% horse serum and 10% chick embryo extract or in BGJb containing various compounds suggested to have a role in biological mineralization. The incubation atmosphere consisted either of 5% CO2 in air or 50% O2, 45% N2 and 5% CO2. Histologically, explants grown in serum-supplemented medium frequently showed dentin mineralization, the frequency being highest, 90%, in high oxygen partial pressure. In medium without serum, only two out of 43 explants which were cultured in BGJb containing 5 mM Na-beta-glycerophosphate, formed mineralized dentin. The amounts of DNA and calcium, and the activity of alkaline phosphatase in explants after culture showed no direct correlation to mineralization potential. Calcified dentin was always confined to one or two clearly demarcated areas, which were intensely stained by the von Kossa method. It appears that mineralization continues readily when it has started and that it is the initiation of mineralization that is the critical threshold and depends on serum factors.

Alkaline Phosphatase↗
PubMed1986

The cellular origin of fibronectin in the basement membrane zone of developing tooth.

The cellular source of fibronectin in the dental epitheliomesenchymal interface was studied in interspecies combinations of mouse and quail tissue. Species-specific fibronectin antibodies were produced by immunizing rabbits with purified mouse or chicken fibronectin and by absorbing both antisera with purified heterologous fibronectin and insoluble tissue extract. The absorbed antisera to mouse and chicken fibronectin showed fluorescent staining only in mouse and chicken tissue sections, respectively, but not vice versa. When the mouse mesenchymal dental papilla was combined and cultured either with the mouse enamel organ or with the quail pharyngeal epithelium, mesenchymal cell differentiation was initiated and typical alignment of mesenchymal cells along the basement membrane was seen. Examination with transmission electron microscope revealed a typical bilaminar basal lamina with adherent fibrillar matrix on its mesenchymal aspect. Immunofluorescent localization of fibronectin with the mouse-specific fibronectin antiserum showed a brilliant staining in the mesenchymal tissue and in the basement membrane zone. When the chicken-specific fibronectin antiserum was used, no staining was detected in either tissue recombinations. We have suggested earlier that fibronectin in the dental basement membrane plays an important role during the differentiation of mesenchymal cells into odontoblasts. The present study demonstrates that fibronectin in the basement membrane of the developing tooth is produced exclusively by the differentiating mesenchymal cells.

Animals↗
PubMed1985

Control of kidney differentiation by soluble factors secreted by the embryonic liver and the yolk sac.

Since transferrin is necessary for the differentiation of the embryonic kidney in organ culture, we have suggested that the component is a growth factor for in vivo development as well. In the present study we demonstrate that transferrin is present in the serum of 11-day-old mouse embryos, at the time when kidney differentiation starts. We have also tested whether various embryonic tissues can replace transferrin as stimulators of the differentiation and proliferation of the metanephric mesenchyme. We used a transfilter model system where nephrogenic mesenchymes are cultured with spinal cord, a known inductor of kidney tubules. The embryonic liver could not replace the spinal cord as an inducer of tubular differentiation. However, when the kidney mesenchymes were cultured together with both the spinal cord and the liver, the mesenchymes proliferated and differentiated also in the absence of exogenous transferrin. In such cocultures the spinal cord had to be in close contact with the mesenchyme while the embryonic liver could be located several cell layers apart. The liver-mediated stimulation of proliferation of the induced mesenchyme could be inhibited by anti-transferrin antibodies. Immunoprecipitation and immunoblotting with these antibodies of the liver-conditioned medium demonstrated that the 11-day mouse liver produces transferrin. Other potential mitogens produced by liver cells, alpha-fetoprotein, or multiplication stimulating activity, did not in any way stimulate the proliferation of induced mesenchymes. These studies suggest that the mitogen in the liver medium is transferrin. This is supported by data which show that another embryonic transferring producer, the visceral yolk sac, can replace the effect of the liver, whereas a tissue not producing transferrin, the salivary mesenchyme, cannot. In conclusion, an essential function of the inducer is to make the mesenchyme responsive to transferrin. The liver and the yolk sac stimulate early kidney differentiation by producing the soluble factor, transferrin, but they are ineffective as inductors of the transferrin responsiveness.

Amniotic Fluid↗
PubMed1985

Epidermal growth factor inhibits morphogenesis and cell differentiation in cultured mouse embryonic teeth.

Although local epithelial-mesenchymal tissue interactions which are presumably mediated by extracellular matrix molecules are important regulators of tooth morphogenesis and differentiation, our studies have indicated that these developmental processes also depend on circulating molecules. The iron-carrying serum protein transferrin is necessary for the early morphogenesis of mouse tooth in organ culture (A-M. Partanen, I. Thesleff, and P. Ekblom, 1984, Differentiation 27, 59-66). In the present study we have examined the effects of other growth factors on mouse tooth germs grown in a chemically defined medium containing transferrin. Fibroblast growth factor and platelet derived growth factor had no detectable effects but epidermal growth factor (EGF) inhibited dramatically the morphogenesis of teeth, and prevented odontoblast and ameloblast cell differentiation. EGF stimulated cell proliferation in the explants measured as [3H]thymidine incorporation in DNA. However, when the distribution of dividing cells was visualized in autoradiographs, it was observed that cell proliferation was stimulated in the dental epithelium but was inhibited in the dental mesenchyme. The inhibition of cell proliferation in the dental mesenchyme apparently caused the inhibition of morphogenesis. We do not know whether the dental epithelium or mesenchyme was the primary target for the action of EGF in the inhibition of morphogenesis. It is, however, apparent that the response of the dental mesenchymal cells to EGF (inhibition of proliferation) is regulated by their local environment, since EGF enhanced proliferation when these cells were disaggregated and cultured as monolayers. This indicates that the organ culture system where the various embryonic cell lineages are maintained in their original environment corresponds better to the in vivo situation when the roles of exogenous growth factors during development are examined.

Animals↗
PubMed1985

The role of transferrin receptors and iron delivery in mouse embryonic morphogenesis.

The iron-carrying serum protein transferrin is required for the proliferation and differentiation of embryonic tissues in culture. We studied the expression and role of transferrin receptors in two model systems using a monoclonal antibody against the transferrin receptor of mice. The addition of 20-100 micrograms/ml antibody to a chemically defined culture medium containing transferrin (10 micrograms/ml) inhibited morphogenesis and cell proliferation in kidneys and teeth. However, the antibody did not inhibit development when iron was delivered to the cells by a lipophilic iron chelator i.e., by-passing the receptor-mediated pathway. Hence, the binding of the receptor antibody to the receptor apparently did not affect cell proliferation, and the antibody was not toxic to the tissues. Our results suggest that the antibody to the transferrin receptor inhibits development by blocking the normal endocytotic route of iron delivery. Cells derived from embryonic kidneys and teeth expressed the transferrin receptor when cultured as monolayers. However, using immunofluorescent techniques, we were unable to detect the receptor in frozen tissue sections. It is possible that the seeding of cells in monolayer cultures affects the expression of the transferrin receptor, since it is known that all types of cells require transferrin for continued proliferation in culture. Organ-cultured kidney mesenchymal cells are not initially responsive to transferrin, but they acquire responsiveness as a consequence of an inductive tissue interaction. Although it remains unknown as to whether the acquisition of transferrin responsiveness is directly related to the expression of transferrin receptors, our results suggest that transferrin and its receptors play a role in embryonic morphogenesis.

Animals↗
PubMed1984

A lipophilic iron chelator can replace transferrin as a stimulator of cell proliferation and differentiation.

Of the different growth supplements used in chemically defined media, only transferrin is required for differentiation of tubules in the embryonic mouse metanephros. Since transferrin is an iron-carrying protein, we asked whether iron is crucial for tubulogenesis. Differentiation of metanephric tubules both in whole embryonic kidneys and in a transfilter system was studied. The tissues were grown in chemically defined media containing transferrin, apotransferrin, the metal-chelator complex ferric pyridoxal isonicotinoyl hydrazone (FePIH), and excesses of ferric ion. Although we found that apotransferrin was not as effective as iron-loaded transferrin in promoting proliferation in the differentiating kidneys, excess ferric ion at up to 100 microM, five times the normal serum concentration, could not promote differentiation or proliferation. However, iron coupled to the nonphysiological, lipophilic iron chelator, pyridoxal isonicotinoyl hydrazone, to form FePIH, could sustain levels of cell proliferation and tubulogenesis similar to those attained by transferrin. Thus, the role of transferrin in cell proliferation during tubulogenesis is solely to provide iron. Since FePIH apparently bypasses the receptor-mediated route of iron intake, the use of FePIH as a tool for investigating cell proliferation and its regulation is suggested.

Animals↗
PubMed1984

Transferrin is required for early tooth morphogenesis.

The role of circulating molecules during early tooth morphogenesis was studied in organ cultures of mouse embryonic molar-tooth germs. Special attention was focused on the effect of transferrin and insulin, which are necessary for the growth of most cells in culture. The requirement of serum factors for tooth morphogenesis was shown to diminish as the developmental stage advances from the bud stage in day-13 embryos to the cap stage at day 15. The day-15 teeth underwent morphogenesis and cell differentiation in unsupplemented basal culture medium, but the addition of transferrin (50 micrograms/ml) was necessary for the morphogenesis of day-14 tooth germs. We demonstrated, by using transferrin-depleted serum, that transferrin is also necessary for the morphogenesis of day-13 tooth germs. However, some still-unidentified serum components are also required for the morphogenesis of the bud-stage day-13 teeth. These factors apparently do not include insulin, since it was shown to inhibit tooth development. Analysis of the DNA content of tooth germs cultured in various culture media showed that the ability of transferrin to support tooth morphogenesis correlated with a stimulation of growth. The results support our earlier suggestions that transferrin functions as a fetal growth factor. The availability of the transferrin-containing chemically defined medium facilitates studies on the roles of other growth factors during tooth development.

Animals↗
PubMed1984

Distribution of keratin and laminin in ameloblastoma. Comparison with developing tooth and epidermoid carcinoma.

The nature of the tumor cells in 5 cases of ameloblastomas was studied by immunohistochemistry, and the findings were compared with developing mouse and human teeth as well as with 5 cases of carcinomas in the oral region. The antigens investigated were keratin, an intracellular cytoskeletal protein typical of epithelial cells, and laminin, an extracellular matrix protein found in basement membranes. Our results show that keratin is expressed by all types of epithelial cells in ameloblastomas as well as in the epidermoid carcinomas, and developing teeth. The epithelial, keratin-positive tumor islands in the ameloblastomas were surrounded by a continuous line of laminin, in a pattern similar to that seen in developing tooth. Laminin was seen also around the epidermoid carcinomas but large areas devoid of laminin were constantly seen between the stroma and the neoplastic epithelium. This indicates a lack of proper basement membrane formation by the malignant epidermoid carcinomas. This may be due either to a diminished production or an increased degradation of basement membrane proteins by the carcinoma cells. Our results are in line with suggestions that ameloblastomas are derived from odontogenic epithelial cells. Immunostaining for keratin does not distinguish between carcinomas and the ameloblastomas. However, visualization of basement membrane proteins such as laminin can apparently be used in the differential diagnosis between ameloblastomas and carcinomas.

Ameloblastoma↗
PubMed1984

Role of transferrin in branching morphogenesis, growth and differentiation of the embryonic kidney.

Our previous work has suggested that transferrin is an important serum component for differentiation of the kidney. In this study we have analysed more closely the response of cultured mouse embryonic kidney to exogenous transferrin and the dependence of kidney tubule induction on transferrin. Our results show that transferrin causes a dose-dependent increase in cell proliferation in the differentiating kidney mesenchyme, but no stimulation of cell proliferation in the inductor tissue used, the embryonic spinal cord. In cultures of whole kidney rudiments a remarkable increase in the amounts of DNA and protein are caused by transferrin but not by other serum components present in a transferrin-depleted serum. The morphology of the explants was similar when cultured in the presence of human serum and in the transferrin-depleted serum supplemented with transferrin. In transferrin-containing chemically-defined medium the explants flattened and spread out, but the morphology of the kidney tubules was similar as in explants cultured in the presence of serum. Examination of the cultured explants by electron microscopy showed that in all transferrin-containing culture media the mesenchymal cells had differentiated into kidney tubules consisting of epithelial cells lined by a basement membrane. The experiments with the transferrin-depleted serum demonstrate that the main mitogen for kidney development is transferrin, and that other serum factors are mainly required for maintenance of tissue compactness. Our earlier studies have shown that exogenous transferrin is not needed for certain changes preceding overt tubule formation in the kidney mesenchyme, and we suggested that transferrin responsiveness is acquired during the induction of kidney mesenchyme. Our present results do not contradict the postulate, although they demonstrate that the acquisition of the responsiveness is more complicated than previously thought. When the mesenchyme is exposed to inductor tissue for 24 h without transferrin, and then subcultured without the inductor in the presence of transferrin, morphogenesis fails and there is no proliferation of the mesenchyme. The experiment shows that the inductor, the mesenchyme and transferrin must all three be simultaneously present for the acquisition of the transferrin responsiveness. Other experiments show that the induced mesenchyme can be a direct target tissue, since it can proliferate in response to transferrin also in the absence of the inductor. It is evident that the inductor is required for the acquisition of the responsiveness, as suggested.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1983

Distribution of the transferrin receptor in normal human fibroblasts and fibrosarcoma cells.

Transferrin is required for proliferation of most cells in culture. This effect is presumably mediated by the binding of transferrin to its receptor, a surface glycoprotein which is preferentially expressed by actively growing cells. Here we show that normal human fibroblasts cultured in serum, and other media containing transferrin express transferrin receptors in a distinctly non-random way; punctate foci of the receptor were seen only at the leading lamellae of the cells, whereas cells grown without serum, or in transferrin-depleted serum showed a random distribution of the receptor. In contrast, malignant fibrosarcoma cells showed the receptor uniformly throughout the cell surface in all media tested, including those containing transferrin. The data suggest that the ligand causes a directional lateral movement of the receptor in normal but not in malignant cells. Application of the receptor antibody caused a rapid internalization of the receptor in both cell types.

Antibodies, Monoclonal↗
PubMed1983

Exogenous fibronectin is not required for organogenesis in vitro.

The biological effect of plasma fibronectin on the differentiation of embryonic mouse kidney and tooth was studied in organ cultures. Transferrin (50 micrograms/ml) was a strong mitogen for kidney cells, whereas the addition of soluble fibronectin (50 to 250 micrograms/ml) had no detectable effect on differentiation or proliferation. The same serum-free, transferrin-containing medium did not support tooth differentiation. However, fibronectin was not a necessary serum component because fibronectin-free serum supported tooth development. It was demonstrated with antibodies specific for human fibronectin that the exogenously added human fibronectin at 50 micrograms/ml did not become incorporated to the cultured organs. Only minimal incorporation to the kidney basement membrane area was observed when fibronectin concentration was 250 micrograms/ml. The mesenchymal stroma and the basement membranes of the kidney and tooth rudiments cultured in fibronectin-free media stained intensely with conventional fibronectin antibodies, indicating endogenous production of fibronectin. Outgrowing epithelial cells from isolated kidney tubules produced fibronectin as well as laminin. The results suggest that the fibronectin found in the stroma and basement membranes is an endogenous product of the developing tissues and that plasma fibronectin is not required for in vitro organogenesis. The results also indicate that it is difficult to study the effect of fibronectin on morphogenetic processes because it may not penetrate the organ explants in vitro.

Animals↗
PubMed1983

Transferrin as a fetal growth factor: acquisition of responsiveness related to embryonic induction.

Differentiation of the metanephric mesenchyme, which is triggered by an inductive tissue interaction, has been shown to proceed in a chemically defined medium containing transferrin. Here, we report that neither transferrin-depleted serum nor a chemically defined medium devoid of transferrin promote differentiation and that activity can be restored by the addition of transferrin. It thus appears that we have identified the serum factor required for kidney differentiation. Transferrin seems to affect differentiation by stimulating cell proliferation. We show by using an organ-culture model system that only mesenchymes induced to differentiate by the 24-hr tissue interaction respond to transferrin by proliferation and differentiation, whereas uninduced mesenchymes remain unresponsive. The inductor tissue used is not responsive to transferrin. Thus, the data suggest that the short-range cell-mediated tissue interaction acts by making the nephrogenic mesenchyme responsive to the long-range mediator, which is transferrin. Transferrin is suggested to be an important circulating growth factor required for proliferation during embryogenesis.

Animals↗
PubMed1983

Inhibition of morphogenesis and stimulation of vascular proliferation in embryonic tooth cultures by a sarcoma growth factor preparation.

Sarcoma growth factor (SGF) induces proliferation and anchorage-independent growth of nonmalignant cells. It competes with epidermal growth factor (EGF) for the EGF-receptors at the cell surface. SGF-like factors have recently been isolated from embryos, suggesting that SGFs may represent embryonic forms of EGF. Therefore, we have tested whether SGF preparations affect organogenesis and differentiation of cultured embryonic tissues. The embryonic tooth rudiments were cultivated in the presence of SGF and EGF. Stimulation of vascularization was seen in both of these organ cultures. Therefore, we propose that endothelial cells may be target cells for SGF, and SGF may be involved in the control of vascularization during embryogenesis. SGF and also, to a certain extent, EGF profoundly inhibited morphogenesis and differentiation of the tooth germ, with concomitant stimulation of vascularization. Analysis of cell proliferation revealed that some cell types of the tooth germ did not respond to SGF by proliferation, while a stimulation by EGF was observed. Nevertheless, tooth morphogenesis was also slightly inhibited by EGF, suggesting that growth factors which enhance proliferation do not necessarily stimulate morphogenesis and differentiation. Since the SGF preparations contain several factors, the effects observed could be due to the action of one or more factors.

Animals↗