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Two-gradient hypothesis of primary embryonic induction.

The "two-gradient" hypothesis of primary embryonic induction was developed by Sulo Toivonen and his school in 1938--1968. The model postulates two inductive principles distributed as gradients in the inductor tissue. The prospective neuroectoderm becomes first uniformly neuralized by one of the factors, and only afterwards segregation is brought about by cells mesodermalized by the other inductor. The short review summarizes the experimental evidence for the "two-gradient" hypothesis.

Animals

Cell contacts between chorda-mesoderm and the overlaying neuroectoderm (presumptive central nervous system) during the period of primary embryonic induction in amphibians.

Using transmission and scanning electron microscopy we were able to show that during primary embryonic induction in amphibians (Triturus alpestris) the interspace between the inducing chorda-mesoderm and the reacting ectoderm (presumptive medullary plate) of mid-gastrula stages is traversed by cell projections starting from cells of both tissue layers. In addition intimate membrane contacts between the main bodies of the ectodermal and chorda-mesodermal cells could be observed. It could be ruled out that cytoplasmic bridges (anastomosis) exist between cells of inducing chorda-mesoderm and reacting ectoderm, which would allow a free transfer of inducing substances without passing through membranes, as Eakin and Lehmann [1] have postulated. The possible role of cell to cell contact for neural induction is emphasized.

Animals

Mechanisms of cell interaction during primary embryonic induction studied in transfilter experiments.

The transmission mechanisms operative at different stages of neutralisation during primary embryonic induction of the newt Triturus vulgaris were studied in experiments employing Nuclepore filters placed between interactive tissue explants. The transmission time of the neuralising effect was determined with 0.2 mum Nuclepore filter. In another series of experiments the transformation of neuralised ectoderm by archenteron roof mesoderm into other parts of the CNS was studied. Although sufficiently long induction times were used no transformation into hindbrain structures could be induced across filters with pore sizes from 0.1 mum to 1.0 mum. However, electron microscopy demonstrated cytoplasmic penetration into 0.6 mum filters at 15 h of induction. The results speak against free long-range diffusion of inductive material at the stage of transformation of the neuralised ectoderm to more caudal parts of CSN and warrant a more detailed structural study of the transmission phenomenon in question.

Animals

Embryonic inductive tissues that cause histologic differentiation of murine mammary carcinoma in vitro.

A murine mammary tumor was cultured in vitro for 14 days, either in direct combination with various embryonic murine inductive tissues or separated by a Millipore filter from these tissues. From 456 test cultures and 269 control cultures of tumor alone, morphologic, histochemical, and autoradiographic evidence for cytodifferentiation was obtained in the tumor after exposure to inductive tissues directly or through the filter. There appeared to be a gradient in potency of the inductive tissues; embryonic mammary mesenchyme was the most active of the tissues tested. Tumor growth was not different from that of controls, however, when the cultured, inductive tissue-exposed neoplasm was returned to the murine host.

Adenocarcinoma

Embryonic induction.

Inductive interactions between tissue components in proximity constitute a universal guiding principle for synchronized development during embryogenesis. Such sequential morphogenetic events involve both specific, determinative "instructions" and less specific, supporting or "permissive" influences acting upon predetermined target cells. Transmission of these intercellular messages may be mediated by diffusible signal substances, by morphogenetically active interfacial materials,or via actual cell contacts. Inductive interactions can be upset experimentally by exposure to various exogenous agents known to be potential teratogens, and several mutant strains of animals are known in which a genetic defect is manifested as a malformation through a defective interactive process. Hence, such inductive interactions should be considered likely targets for both genetic and exogenous factors in teratogenesis.

Abnormalities, Drug-Induced

Microheterogeneities, non-equivalance, and embryonic induction.

The thrust of this report is to stress the importance of microheterogeneities in the microenvironment of differentiating tissues as a possible inducer or regulator of differentiation. During chondrogenesis both qualitative and quantitative changes occur in the proteoglycan population. Using molecular sieve chromatography, these changes can be characterized and used as indices of differentiation. Microheterogeneities of the extracellular matrix may be an example of "non-equivalence" as a regulatory device for differentiation.

Animals

The role of growth factors in embryonic induction in Xenopus laevis.

Establishment of the body pattern in all animals, and especially in vertebrate embryos, depends on cell interactions. During the cleavage and blastula stages in amphibians, signal(s) from the vegetal region induce the equatorial region to become mesoderm. Two types of peptide growth factors have been shown by explant culture experiments to be active in mesoderm induction. First, there are several isoforms of fibroblast growth factor (FGF), including aFGF, bFGF, and hst/kFGF. FGF induces ventral, but not the most dorsal, levels of mesodermal tissue; bFGF and its mRNA, and an FGF receptor and its mRNA, are present in the embryo. Thus, FGF probably has a role in mesoderm induction, but is unlikely to be the sole inducing agent in vivo. Second, members of the transforming growth factor-beta (TGF-beta) family. TGF-beta 2 and TGF-beta 3 are active in induction, but the most powerful inducing factors are the distant relatives of TGF-beta named activin A and activin B, which are capable of inducing all types of mesoderm. An important question relates to the establishment of polarity during the induction of mesoderm. While all regions of the animal hemisphere of frog embryos are competent to respond to activins by mesoderm differentiation, only explants that include cells close to the equator form structures with some organization along dorsoventral and anteroposterior axes. These observations suggest that cells in the blastula animal hemisphere are already polarized to some extent, although inducers are required to make this polarity explicit.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals

Fish swimbladder: an excellent mesodermal inductor in primary embryonic induction.

Swimbladder of the crucian carp, Carassius auratus, was found to be better as a vegatalizing tissue than other tissues, such as guinea-pig bone marrow, when presumptive ectoderm of Triturus gastrulae was used as reacting tissue. Swimbladder usually induced assemblies of highly organized mesodermal tissues, such as notochord, somites and pronephric tubules, some of which were covered by mesodermal epithelium without any epidermal covering. A special character of the effect of swimbladder was the rather frequent induction of solid balls of undifferentiated cells, which were identified as mesodermal or mesodermal and probably endodermal. These findings show that swimbladder has a strong and fast spreading vegetalizing effect on the responding presumptive ectoderm.

Air Sacs

Embryonic lens induction: shedding light on vertebrate tissue determination.

The principle of embryonic induction was defined by early studies of lens determination, and because of the relative simplicity of the developing lens and its interaction with presumptive retinal tissue it has been a favored system for examining mechanisms of induction. Recent studies have led to substantial alterations of the classic model for this process, introducing several elements that significantly refine our view of vertebrate tissue determination.

Animals

Interaction between growth factors and retinoic acid in the induction of kidney tubulogenesis in tissue culture.

Kidney tubulogenesis is the initial step in renal organogenesis. The precise molecular determinants of this pattern formation are presently unknown, although soluble factors, such as growth factors, and insoluble factors, such as extracellular matrix molecules, most likely play fundamental roles in this process. To define the molecular determinants of renal proximal tubule morphogenesis, primary cultures of rabbit renal proximal tubule cells in hormonally defined, serum-free media were treated with transforming growth factor-beta 1 (TGF-beta 1), epidermal growth factor (EGF), and the retinoid, all trans-retinoic acid (RA), singly or in combination. Utilizing phase contrast and light and transmission electron microscopy, the simultaneous administration of TGF-beta 1 (10 ng/ml), EGF (1 nM), and RA (0.1 nM) transformed a confluent monolayer of renal proximal tubule cells within 5 to 6 days into three-dimensional cell aggregates containing lumens within the interior of the cell clusters. The lumens were bordered by tubule cells possessing a polarized epithelial cell phenotype with extensive microvilli formation and tight junctional complexes along the luminal border. All three factors were necessary and sufficient to induce this phenotypic transformation. Further studies demonstrated that RA promoted the deposition of the A and B1 chains of laminin, a cell attachment protein of the basement membrane, in a small subset of proximal tubule cells in culture, as deduced by indirect immunofluorescent microscopy. Additional studies demonstrated that soluble purified laminin fully substituted for RA in this system to promote renal tubulogenesis when combined with TGF-beta 1 and EGF. These results demonstrate that the growth factors, TGF-beta 1 and EGF, and the retinoid, RA, promote tubulogenesis in adult renal proximal tubule cells in tissue culture in a manner reminiscent of inductive embryonic kidney morphogenesis. These observations define a coordinated interplay between growth factors and retinoids to induce pattern formation and morphogenesis. Furthermore, the demonstration of RA-induced laminin deposition as a critical event in this morphogenic process identifies laminin as a possible target protein for RA to act as a morphogen.

Animals

Induction of tubules in rat metanephrogenic mesenchyme in the absence of an inductive tissue.

Differentiation of metanephrogenic mesenchyme to renal tubular epithelium requires induction by the ureteric bud in vivo or any of several embryonic tissues in vitro. In an effort to eliminate the tissue requirement in embryonic induction, extracellular matrices and soluble factors were analyzed individually or in combination for their ability to stimulate tubulogenesis in uninduced metanephrogenic mesenchyme from 13-gestation-day rat embryos. These evaluations have established that pituitary extract and epidermal growth factor (EGF) in concert with a matrix can promote morphogenesis of mesenchymal rudiments in culture. While type I collagen, laminin, or fibronectin matrices all promoted tubulogenesis in the presence of pituitary extract and EGF, type IV collagen proved the most effective. Under these conditions, tubules were induced in 23/24 mesenchymal rudiments by 9 days in culture. Mesenchyme was not induced prior to explanation since it formed no tubules when cultured in a medium that allowed tubulogenesis in intact embryonic kidneys. Preliminary characterization of the undefined factor in pituitary extract was consistent with a protein of molecular weight greater than 100,000 but less than 300,000. When uninduced metanephrogenic mesenchyme from mouse was used instead of rat tissue, a similar pattern of morphogenesis was not observed, suggesting that the described medium is inappropriate for promoting differentiation in mouse or, less likely, that different mechanisms mediate differentiation in rat and mouse. These studies show that embryonic induction can occur in explanted rat renal mesenchyme in an appropriate environment and does not require the presence of an inductive tissue.

Animals