Biomedical subjects

S M Cannata

Publications and source records attributed to S M Cannata.

10 recordsLinked to original sources

PubMed1997

Lens regeneration in larval Xenopus laevis: experimental analysis of the decline in the regenerative capacity during development.

In Xenopus laevis, the capacity to regenerate a new lens from the outer cornea gradually decreases between stages 50 and 58, is almost negligible during the metamorphic climax, and disappears after metamorphosis. The factors responsible for lens transdifferentiation of the outer cornea are produced by the neural retina and are located in the vitreous chamber. This decrease in the regenerative capacity may be due to: (1) a reduction of the inductive power of the retina, (2) a reduction of lens-forming competence of the outer cornea, (3) an inhibition of the lens transdifferentiation process, (4) a combination of these causes. In order to test these hypotheses, fragments of outer cornea or of outer and inner corneas joined together were isolated from early larvae, late larvae and froglets, and implanted into the eye of host larvae during the premetamorphosis or the metamorphic climax. Results from implants of outer cornea into the vitreous chamber showed that the drop in lens regeneration capacity during the metamorphic climax is not due to a decrease in the inductive power of the retinal factor and that the gradual decrease in the regenerative capacity observed between stages 50 and 58 is not related to a substantial diminution in the capacity of outer cornea cells to transdifferentiate into lens fibers. Results from implants of outer and inner corneas joined together showed that in these implants the lens transdifferentiation of the outer cornea was partially inhibited. These findings indicate that the decrease in lens regeneration is mainly due to an inhibition of the lens transdifferentiation process of the outer cornea by the inner cornea. However, even implants of cornea (multilayered epithelium and substantia propria) excised from metamorphosed animals were able to form lens fibers, although to a lesser percentage than that obtained after implantation of fragments of larval outer and inner corneas. Thus, the lens-forming competence in the corneal epithelium is still present to a certain degree even when lens regeneration capacity is lost. Several observations suggest that in the lentectomized eye of late larvae and froglets the mechanical inhibition of lens transdifferentiation process exerted by the inner cornea (or the substantia propria), due to the rapid formation of a connective barrier against the spreading of the retinal factor toward the outer cornea, has a decisive role in maintaining the phenotypic stability of the outer cornea.

Animals↗

PubMed1996

Morphogenesis and differentiation of grafted blastemas formed in vitro from amputated hindlimbs of larval Xenopus laevis.

The present study was designed to test the morphogenetic potency of limb blastemas formed in vitro from amputated limbs of larval Xenopus laevis. Hindlimbs of larvae at stage 55 (according to Nieuwkoop and Faber [1956] Normal Table of Xenopus laevis (Daudin)) were amputated through the tarsalia, excised at the base of the thigh and cultured in Leibovitz's L-15 supplemented with 2% FCS. After 8-10 days, 50% of the cultured limbs formed a conic blastema on the amputation surface. However, on the excision surface no blastema was present. Three different parts (blastema, blastema with the shank region and proximal part of the limb) of the cultured limbs were then grafted to the axial musculature or to the hindlimb of stage 57 host larvae. Results showed that the blastema formed in vitro were true autodifferentiating regeneration blastemas, since they were able to form well-differentiated autopodia not only when grafted with the shank region to a neutral territory (axial musculature) or to the limb territory, but also when transplanted alone to the two environments. The morphological complexity (no. of toes) of the autopodia differentiated from the grafted blastemas was superimposable to that observed in vivo. Moreover, as in vivo, the entire regeneration process was nerve-independent. In fact, the regeneration blastemas, formed in vitro in the complete absence of nerves, could grow and differentiate also when grafted to denervated host limbs. The grafted proximal parts of the cultured limbs never formed a regenerate.

Animals↗

PubMed1995

Acquisition of nerve dependence for the formation of a regeneration blastema in amputated hindlimbs of larval Xenopus laevis: the role of limb innervation and that of limb differentiation.

In larval and adult urodeles and late-stage larval anurans, blastema formation after limb amputation requires an adequate nerve supply. Experimental evidence obtained from aneurogenic limbs indicates that, in urodeles, the acquisition of nerve dependence during embryonic development is due to the "addiction" of limb tissues to factors released by the ingrowing nerves rather than to limb differentiation. The aim of this work was to establish whether, in the toad Xenopus laevis, nerve-dependence for blastema formation after hindlimb amputation, which is acquired gradually during larval development and becomes complete at stage 57 is due to limb innervation or to limb differentiation. Two series of experiments were carried out. In the first series, limb differentiation was inhibited by treating the larvae with an anti-thyroid drug, and innervation was maintained for an interval much longer than that normally required for development from nerve-independent stages to stage 57. In the second series, the limb was caused to differentiate in the absence of nerves by maintaining the limbs denervated. Limb differentiation was often accelerated by treating early-stage larvae with thyroxine or by grafting early-stage limbs onto denervated limbs of late larvae, which, being near metamorphic climax, possessed high levels of circulating thyroid hormones. Results showed that in the first series of experiments the denervated limbs formed regeneration blastemas after amputation, but in the second series they did not. It was therefore concluded that the acquisition of nerve dependence for blastema formation in larval Xenopus laevis is not directly imposed by factors released by the nerve fibers, but is strongly related to differentiation of limb tissues.

Amputation, Surgical↗

PubMed1995

The inhibition of cell proliferation by mitomycin C does not prevent transdifferentiation of outer cornea into lens in larval Xenopus laevis.

The aim of the present work is to evaluate the relationship between cell proliferation and transdifferentiation (TS) of the outer cornea into lens in larval Xenopus laevis. Data obtained from corneal fragments treated with Mitomycin C (MMC) (0.1 mg/ml, 50 min) and implanted into the vitreous chamber (MMC/v ch) were compared with those obtained from untreated corneal fragments implanted into the vitreous chamber (contr/v ch) or between outer and inner corneas (contr/o c). Results demonstrated that in contr/v ch implants, which transdifferentiated into lenses or lentoid bodies in 88% of cases, the mitotic index (MI) showed a sharp increase during the period of lens vesicle formation (3 days) and became very low when the formation of lens fibres was under way (7 days). In contro c implants, which did not undergo any lens forming transformations, the MI remained unchanged in comparison to time O. In MMC/v ch implants, the inhibition of the mitotic activity was 100% up to the third day after implantation. On the fifth and seventh days, scant mitotic activity was observed in some cases, but the MI was much lower than the MI of contr/o c implants. The MMC/v ch implants transdifferentiated into lentoid bodies in 26% of cases. The lentoid bodies were much smaller than those observed in control implants, but they reacted positively with the lens antibodies at the same time after implantation as controls. Even the complete inhibition of proliferation due to stronger MMC treatments (e.g. 0.15 mg/ml, 50 min) did not prevent lens TS.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

Differences in the decrease in regenerative capacity of various brain regions of Xenopus laevis are related to differences in the undifferentiated cell populations.

The extent of the undifferentiated cell population in normal and regenerating brains of larvae and metamorphosed individuals of Xenopus laevis has been analyzed by means of an immunocytochemical method and mitotic index determinations. Results show that the decrease in regenerative capacity of the brain during larval development and after metamorphosis is in relation with the gradual reduction of the population of undifferentiated cells and that the different regenerative capacities of the various brain districts are related to quantitative and qualitative differences in this cell population. While in the early larval stages the cell population formed of actively cycling cells is very large and widespread, in late larval stages and after metamorphosis these cells localize in some encephalic areas (matrix zones). This localization occurs later in the telencephalon than in the rhombencephalon and in mesencephalon. The less conspicuous decrement in the regenerative capacity of the telencephalon than of other encephalic districts of froglets, particularly the mesencephalon, is related to the presence of a larger number of actively cycling cells together with a rather large number of undifferentiated cells which are in a temporary quiescent state from which they may re-enter the actively cycling state in response to proliferation promoting factors.

Animals↗

PubMed1995

Medullary and gangliar regeneration after unilateral removal of a segment of spinal cord of the trunk and corresponding ganglion in adult newts.

Regeneration of the spinal cord, segmental nerves and sensory ganglia takes place after tail amputation in the newt. Many histological and immunocytochemical observations provide evidence that the ependymal tube is the source not only of new neurons and glial cells in the spinal cord, but also of some cells that go on to participate in the formation of the spinal ganglia of the regenerating tail. In previous experiments involving the removal of the spinal ganglia of the trunk, no regeneration was observed and it is thought that the trunk region differs from the tail region with regard to the ability to regenerate sensory ganglia. However, in these experiments the spinal cord of the trunk was not damaged. In the present work involving adult newts (Triturus carnifex Laur.), unilateral ablation of a segment of the spinal cord of the trunk in addition to removal of a corresponding spinal ganglion was performed. In these experimental conditions, regeneration of a rudimentary spinal ganglion near the regenerated side of the spinal cord segment was observed in several cases. Histological observations carried out 2, 4, 6 and 13 months after the operation support the view that some cells migrating from the lateroventral part of the regenerating side of the spinal cord via the developing ventral root could participate in the formation of the rudimentary spinal ganglion.

Amputation, Surgical↗

PubMed1992

Regenerative responses in cultured hindlimb stumps of larval Xenopus laevis.

The regenerative capacity of larval Xenopus laevis hindlimbs amputated through the tarsalia at different stages of development and explanted in vitro was tested. In the first experimental series hindlimb stumps from stage 53, 54, 55, and 57 larvae (according to Nieuwkoop and Faber, '56) were cultured in Leibovitz's L-15 medium supplemented with 10% FCS, and 0.04 U of insulin and 10(-8) mg of L-thyroxine per ml of medium. Results showed that the distal part of the limb stumps from stages 53, 54, and 55 formed a regeneration blastema composed of proliferating mesenchymal cells beneath a typical apical cap. No blastema was formed in the proximal part of the stump. In limb stumps from stage 57, a regeneration blastema did not form either in the proximal or in the distal part of the stump. In a second experimental series, hindlimb stumps from stage 55 larvae, denervated 5 days prior to amputation in order to eliminate any residual neurotrophic factor, were cultured in a simplified L-15 medium containing 2% FCS and lacking insulin and thyroxine. Results showed that also in these experimental conditions the stumps from stage 55 formed a conical regeneration blastema at the distal tip. The blastema cells duplicated their own DNA and divided. At the proximal extremity no regeneration blastema was formed. In the same culture medium, the stumps of larvae at stage 57 did not form a regeneration blastema.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Spinal cord and ganglia regeneration in larval Xenopus laevis following unilateral ablation.

The experiments were carried out on larvae of Xenopus laevis at stage 48 (acc. to Nieuwkoop and Faber, 1956). Two different kinds of experiments were performed. Experiment I: Unilateral ablation of either a brachial or lumbar segment of the spinal cord and simultaneous removal of the related ganglia. Experiment II: Simple unilateral removal of either brachial or lumbar spinal ganglia. The results obtained in Experiment I show that not only an extensive restitution of the ablated spinal cord does take place, but the regeneration of spinal ganglia may also occur following migration of neural elements from the regenerating spinal cord. The medullary neuroblasts leave the spinal cord along two paths: i) through projections of the gray matter, probably due to the lack of an effective glia limitans; ii) through the motor fibers leaving the spinal cord to form the ventral roots. The first path is followed occasionally while the second is the one usually used when the ventral roots are present. Data based on animals injected with 5-bromodeoxyuridine and sacrificed at fixed intervals, suggest that ganglion precursors, as well as the medullary neurons and glia, originate in the ependyma. This conclusion is supported by the results of Experiment II which demonstrate that when the spinal cord is left intact no discrete groups of ganglion cells and/or glial cells are formed.

Animals↗

PubMed1991

The influence of denervation on grafted hindlimb regeneration of larval Xenopus laevis.

The aim of the present research is to ascertain whether in larval Xenopus laevis nerve-independence for the regeneration of early stage limbs and nerve-dependence of late stage limbs observed in a previous work (Filoni and Paglialunga, '90) is related to extrinsic (systemic) factors or to intrinsic changes taking place in the limb cells themselves during development. In this paper the regenerative capacity of early and late stage hindlimbs under the same extrinsic conditions, insofar as both are grafted onto the denervated hindlimbs of host larvae at the same developmental stage, is studied. All the grafted limbs are amputated after the host larvae have reached stage 57-58 (according to Nieuwkoop and Faber, '56). In experiment I, the grafted limb is amputated at stage 52, at the thigh level; in experiment II, the grafted limb is amputated at stage 54-55, at the tarsalia level; in experiment III the grafted limb is amputated at stage 57, at the tarsalia level. In all three experiments, together with the grafted limb, also the host limb is amputated at the tarsalia level. The results show that while grafted limbs amputated at stages 52 and 54-55 regenerate in the absence of nerves, grafted limbs amputated at stage 57 cannot. The failure of late stage grafted limbs to regenerate cannot be explained in terms of an immune-type inhibiting reaction since it has been observed also in denervated autografted limbs and in the host limbs. Since all the grafted limbs are in the same environmental conditions, the results show that in larval Xenopus laevis nerve-independence for regeneration of early stage limbs and nerve-dependence of late stage limbs are not related to factors extrinsic to the limb but to intrinsic changes taking place in the limb cells themselves during development.

Animals↗

PubMed1991

Lens formation from the cornea following implantation into hindlimbs of larval Xenopus laevis: the influence of limb innervation and extent of differentiation.

Corneal fragments of larval Xenopus laevis at stage 48 (according to Nieuwkoop and Faber, '56), were implanted into sham denervated unamputated hindlimbs, denervated unamputated hindlimbs, amputated and sham denervated hindlimbs, and amputated and denervated hindlimbs of larvae at stages 52 and 57. The results show that unamputated limbs at stage 52, either innervated or denervated, manifest a weak capacity to promote the first lens-forming transformations of the outer cornea. This capacity is absent in both limb types at stage 57. After amputation, limbs of both early and late stages form a regenerative blastema and support lens formation from the outer cornea. Denervation of early stage limbs has no appreciable effect on blastema formation and lens-forming transformation of corneal implants. However, denervation of late stage limbs inhibits both processes. These results indicate that the limb tissues of the early stage limbs contain non-neural inductive factors at a low level and that after limb amputation and blastema formation the level of these factors becomes high enough to promote lens formation from implanted cornea, even after denervation. In contrast, the limb tissues of late stage limbs do not contain a suitable level of non-neural inductive factors.

Animals↗