PubMed Health⌕ Search

Biomedical subjects

S Filoni

Publications and source records attributed to S Filoni.

At least 19 recordsLinked to original sources
PubMed1999

Nerve-independence of limb regeneration in larval Xenopus laevis is related to the presence of mitogenic factors in early limb tissues.

Early limbs of larval Xenopus laevis can form a regeneration blastema in the absence of nerves. The nerve-independence could be due to the synthesis of neurotrophic-like factors by the limb bud cells. To test this hypothesis, two series of experiments were performed. Series A: the right hindlimbs of stage 57 larvae (acc. to Nieuwkoop and Faber. 1956. Normal table of Xenopus laevis [Daudin]. Amsterdam: North-Holland Pub. Co.), which are nerve-dependent for regeneration, were amputated through the tarsalia. The regenerating limbs were submitted to: sham denervation; denervation; denervation and implantation of a fragment of an early limb, or a late limb, or a spinal cord. Series B: froglets were subjected to amputation of both forelimbs. The cone blastemas were transplanted into denervated hindlimbs of stage 57 larvae, together with a fragment of an early or a late limb. The results in series A showed that the implantation of early limb tissue into the denervated blastema maintained cell proliferation at levels similar to those observed after the implantation of a spinal cord fragment or in sham denervated blastemas. However, the implantation of late limb tissues were ineffective. The results of series B showed that the implantation of early limb tissue, but not of late limb tissue prevented the inhibition of cell proliferation and the regression of denervated limb blastemas of juveniles. These results indicate that the nerve-independence is related to the synthesis of diffusible mitogenic neurotrophic-like factors in early limb tissues, and that nerve-dependence is established when differentiated cells of late limb tissues stop producing these factors.

Amputation, Surgical↗
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↗
PubMed1996

Effect of ammodytin L from the venom of Vipera ammodytes on Xenopus laevis differentiated muscle fibres and regenerating limbs.

Ammodytin L is a non-catalytic, phospholipase-like snake venom toxin from Vipera ammodytes, which shows a cytotoxic activity on differentiated myotubes when tested in vitro. In the range of concentrations in which ammodytin L induced necrosis of myogenic cells in culture, other cell types (erythrocytes, platelets, fibroblasts) did not appear to be affected. To test the in vivo toxicity and the effective cytolytic specificity of ammodytin L we have followed the morphological changes in muscle tissue of Xenopus laevis limbs after intramuscular toxin injection. Only muscular cells were affected by ammodytin L, and the toxin did not induce any morphological change in other cell types. Further evidence of the muscle-specific action of the toxin was obtained from experiments carried out using the Xenopus kidney cell line B3.2 in culture. Ammodytin L was unable to affect parameters of cell viability such as lactate dehydrogenase leakage, [3H]thymidine incorporation, growth curves and morphological changes. Moreover, direct ammodytin L application to cultured regenerative limbs did not provoke alterations in undifferentiated myoblasts. These data suggest that ammodytin L, like other phospholipase-like toxins, exerts its toxicity by selectively damaging differentiated muscle fibres.

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↗
PubMed1993

Persistence of a high degree of structural organization of the telencephalon in brain homotransplants in adult Triturus carnifex (Urodele Amphibians).

Brains of adult Urodele Amphibians (Triturus carnifex Laur.) were homoplastically transplanted in conditions of complete morpho-functional isolation and fixed 100 and 365 days after the operation. Results show that in the post-telencephalic regions structural organization disappear in all cases, while, in some cases the telencephalon retain a fairly complex structural organization even after one year. The heterogeneous nature of the histological patterns observed may be attributed to the different size of the matrix areas, besides the different degree of immune response of the host to the graft. In the telencephalic district, where the matrix areas are particularly well developed, undifferentiated elements of these areas would be capable of opposing the degenerative phenomena in the transplants by neoformation of neurons. However, in post-telencephalic districts, where the matrix areas are less developed or absent, it would not be possible to make up for neuronal degeneration and these regions disappear or are formed only by some undifferentiated elements outside the surviving ependyma portions.

Animals↗
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↗
PubMed1992

Degenerative and regenerative phenomena in brain heterotopic homoplastic transplants of adult Triturus carnifex (Urodele Amphibians).

The brains of adult Urodele Amphibians (Triturus carnifex Laur.) were homoplastically transplanted in conditions of complete morpho-functional isolation. Despite the heterogeneous nature of the histological aspects observed, the results point to the following general pattern for the fate of the brain transplants: 1) rooting phase (about day 5 after the operation) as indicated by degenerative patterns affecting the various encephalic districts and mainly involving the more peripheral neurons and the nerve fibers, and to a lesser extent the periventricular gray and the ependyma; 2) phase of rejection by host (from day 10 until end of experiment on day 30) as indicated by lymphocytic infiltration, the presence of macrophages and pyknoses, and the formation of a connective tissue capsule surrounding the transplant; 3) regenerative phase (from about day 10) as indicated by the onset of mitotic activity affecting elements of the ependymal epithelium and the periventricular gray, particularly in the telencephalic district. By the end of the experiment, in most of the transplants examined, degenerative processes were found to have prevailed over the regenerative phenomena. However, in some cases, and particularly in one of them, a distinctly higher degree of structural organization of neurons and nerve fibers can be observed at the telencephalic level. It may be postulated that, whenever a greater degree of compatibility exists between the host and the transplanted organ, it is possible, even in conditions of complete morpho-functional isolation, for the brain to express regenerative power through the ependyma and the matrix zones which have already been identified by some Authors in telencephalic periventricular areas or scattered through the mesencephalic gray matter.

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↗
PubMed1991

Transient expression of glial-fibrillary acidic protein (GFAP) in the ependyma of the regenerating spinal cord in adult newts.

In the homeotherms the presence of glial-fibrillary acidic protein (GFAP) and that of neurofilaments (NF) are used as specific molecular markers associated with intermediate filaments (IF) of the glial and neuronal elements respectively. On this basis a comparison was made between trends in the immunohistochemical response to these markers of the ependyma of a both a normal and a regenerating spinal cord in Triturus carnifex (Urodele Amphibian) adults, whose high spinal cord regenerative capacity is due to the ependymal epithelium cells. Under normal conditions, the ependymal cells are vimentin positive and negative for GFAP and the IF, respectively, while GFAP and NF positivity is found in the grey and white matter. During regeneration, by about day 10, vimentin positivity begins to decrease in the ependymal epithelium of the caudal stump and GFAP positive elements appear (subsequently, this was observed also in the ependymal epithelium of the cephalic stump). High GFAP positivity is found also in the newly formed apical ampullae. From day 20 to after day 30 these patterns develop both in the stumps and in the regenerating spinal cord. NF positivity is always restricted to the grey matter alone and to the axonal processes of the white matter. At about day 60, by which time the regenerative processes may be considered as having terminated and the spinal cord has virtually regained its original structure, the immunohistochemical features typical of the rest state are restored in the ependymal epithelium.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Effect of denervation on hindlimb regeneration in Xenopus laevis larvae.

Xenopus laevis larvae at stages 51-57, according to Nieuwkoop and Faber, were subjected to amputation of the right hindlimb or of both limbs at the thigh or the tarsal level, as well as to somatic denervation of the right limb. Larvae at the same stage having undergone amputation of the right limb or of both limbs and sham denervation of the right limb were used as controls. In experimental series I a single denervation of the right limb was performed at the time of amputation. In experimental series II repeated denervations were performed (before, during and after amputation). Results show that in larvae at stages 51-53 subjected to limb amputation at the proximal level (thigh) even repeated denervation of the right limb did not prevent regeneration, although giving rise to various degrees of hypotrophy. In stage-55 larvae partial inhibition of the regenerative process in the right limb was clearly visible only after repeated denervations and amputation at the proximal level. After amputation at the distal level (tarsalia) the regenerative process in the right limb underwent no significant delay with respect to the controls, although the regenerated right limb was hypotrophic. In stage-57 larvae even a single denervation at the time of amputation was enough to inhibit regeneration of the right limb after either proximal or distal amputation. Therefore, in Xenopus laevis larvae, nerve-dependence for hindlimb regeneration takes place proximodistally as the nerve fibers grow in the limb and it gradually undergoes a process of proximodistal differentiation.(ABSTRACT TRUNCATED AT 250 WORDS)

Amputation, Surgical↗
PubMed1989

Autoimmunity and central nervous system regeneration in urodele amphibians.

Spinal cord lesion in newt is followed by complete cell and fibre regeneration. Previous observations showed that the presence of a granuloma due to a foreign substance (talcum) in the vicinity of the cut spinal cord slows down and/or prevents regeneration. The present experiments, while confirming previous evidence, show that, in animals with a paraspinal granuloma or a subcutaneous granuloma containing an autoplastic and homoplastic spinal cord implant, immunocomplexes appear on the cut ends as has been observed by the same authors in animals in which spinal cord regeneration does not occur (Mammals). The authors discuss the results in view of their theory of the autoimmune nature of the absence of axonal regeneration.

Animals↗