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Biomedical subjects

P Ferretti

Publications and source records attributed to P Ferretti.

At least 37 records · Page 2Linked to original sources
PubMed1996

Regeneration of lower and upper jaws in urodeles is differentially affected by retinoic acid.

The vitamin A derivative retinoic acid (RA) is a powerful teratogen which can induce severe craniofacial and limb malformations if administered at certain stages of gestation. In addition this compound has been shown to affect patterning in regenerating systems. A classical example is the induction of supernumerary structures along the proximodistal axis of the regenerating amphibian limb. We have investigated the effect of RA on other regenerating systems, the amphibian lower and upper jaws, both in developing and adult animals. We report here that RA does not induce formation of extra structures either in the lower or in the upper jaw of adult newts under experimental conditions where duplications of the regenerating limb occur. However, RA selectively induces severe malformations in the upper jaw regenerate that resemble those induced in avian and mammalian embryos. Analysis of the expression of the newt retinoic acid receptors RAR alpha and delta in upper and lower jaws showed that RAR alpha was expressed at a significant level in the wound epidermis, but not in blastemal cells, whereas no RAR delta could be detected in the regenerate either by in situ hybridization or by using an anti-RAR delta antibody. Therefore, unlike in the limb, in jaws RAR delta is not up-regulated following amputation, and this difference in expression may be causally related to the different effects induced by RA on jaws and limbs. In order to establish whether retinoids affected regeneration of developing jaws in a similar fashion, their effects were studied in animals whose jaws had been amputated at different developmental stages. Under the experimental conditions used overall growth retardation and head defects were observed in the majority of embryos which had been amputated and treated with retinol palmitate (RP) between stages 26-28 and 38-39. In contrast, patterning of upper jaw regenerates in larvae amputated at stage 26-28 and 38-39. In contrast, patterning of upper jaw regenerates in larvae amputated at stage 45 was not significantly affected by the treatment, although the early phase of regeneration was slower than in controls. The different responses to retinoids of regenerating facial structures in embryos, larvae and adults will be discussed.

Abnormalities, Drug-Induced↗
PubMed1995

Retinoic acid-induced cell death in the wound epidermis of regenerating zebrafish fins.

Previous work has shown that treatment with retinoic acid (RA) can induce malformations in regenerating pectoral and caudal fins. RA-treated regenerates are narrower than unamputated and regenerated control fins because of a decrease in the distance between rays, and either partial or total fusion of some of them. In order to tackle the issue of how RA induces its teratogenic effects on regenerating fins, and which cell types may be specifically affected by RA, we have examined the cellular changes occurring in early regenerates following treatment with retinoids. The work presented here shows for the first time that RA induces significant apoptosis in the wound epidermis, but not in the mesenchyme, of a regenerating appendage, besides inhibiting blastema development as reported in other species. We also show that RA does not retard regeneration by inhibiting accumulation of blastemal cells, but probably by impairing their ability to migrate distal to the amputation plane. This effect is rapidly reversed by discontinuing the treatment, and within 24 hr of removing the drug, blastema development is well advanced. By this time the teratogenic effects induced by RA are already apparent. A correlation between the length of the apical ectodermal ridge (AER) and the number of digits formed has been demonstrated in developing limb buds. We therefore suggest that RA-induced patterning abnormalities in regenerating fins are the consequence of a reduction in the size of the wound epidermis, due to increased cell death, which would affect patterning of the underlying mesenchyme.

Animals↗
PubMed1995

Restoration of normal Hox code and branchial arch morphogenesis after extensive deletion of hindbrain neural crest.

Among the derivatives of the cephalic neural crest is the ectomesenchyme which subsequently constitutes most of the craniofacial skeleton. There is evidence to suggest that the skeletogenic fate of the hindbrain neural crest is specified before emigration from the neural tube and that Antennapedia class Hox genes are involved in that process. To explore the putative causal link between Hox expression and craniofacial morphology, we produced a specific series of bilateral crest deletions in chick embryos and assessed branchial arch morphology, Hox gene expression, and patterning of skeletal structures in the postoperative embryo. Surprisingly, we found that deletion of the bulk of the rhombencephalic crest and substantial portions of the dorsal rhombencephalon did not prevent normal branchial arch morphogenesis and normal patterns of Hox gene (-A3 and -B4) expression 48 h after operation. Neural crest-like cells have been identified on crest migration pathways at the level of the original ablation, further confirming that ablated cephalic neural crest is replaced by regeneration from the cut edge of the neuroepithelium. Furthermore, in such embryos ectomesenchyme from regenerated crest is able to form a facial skeleton in which the mandible and hyoid apparatus are normal in size and organization. These findings demonstrate that the cranial neuroepithelium has more extensive regenerative capacities than was previously thought, which has important implications for investigations of craniofacial development.

Animals↗
PubMed1995

Caudal fin regeneration in wild type and long-fin mutant zebrafish is affected by retinoic acid.

Zebrafish (Danio rerio) represents an ideal experimental model to tackle fundamental issues concerned with organogenesis during development and regeneration of complex body structures. We discuss here the development of the skeleton in zebrafish caudal fins, their regenerative ability in wild type and long-fin mutant adult fish, and how retinoic acid (RA), which induces duplications along the proximodistal axis in regenerating limbs, affects regeneration of the caudal fin. The dorsal and ventral lobes of zebrafish caudal fins are apparently symmetrical along the dorsoventral axis, but all of the skeletal elements and most of the soft tissues of both lobes originate from the ventral part of the embryo, as demonstrated by whole-mount staining of developing fish. Analysis of caudal fin regenerates in wild type adults does not reveal any difference in the regenerative ability of the two lobes, and in the length of the regenerate in comparison with the amputated part. In contrast, in the long-fin mutant the regenerated caudal fin is always somehow defective in that the original asymmetry in the length of the two lobes observed in this mutant is not reproduced in the regenerate. Furthermore, in the majority of the batches studied the regenerate is much smaller in size than the amputated part. This suggests that this mutant may be valuable to further our understanding of the mechanisms underlying growth control and patterning during regeneration. Finally, we show that the regenerating caudal fin is sensitive to RA-treatment, and clear teratogenic effects on the dorso-ventral axis are observed under many of the experimental conditions investigated both in wild type and long-fin mutants.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Glutathione metabolism in Crohn's disease.

A statistically significant decrease of glutathione (GSH) and an increase of GSH disulfide (GSSG) both in healthy and ill ileum of patients with Crohn's disease in comparison with the controls (without this pathology) is demonstrated. However, the lowering of these levels was more remarkable in ill ileum in which high levels of GSSG were detected, too. These alterations may be in part explained by the changes obtained in GSH-related enzyme levels. Finally, considering the results that others and we obtained by studies on GSH oral absorption in rat intestine, an oral therapy of GSH in Crohn's disease is suggested.

Administration, Oral↗
PubMed1994

Regenerative capability of upper and lower jaws in the newt.

The regenerating amphibian jaw represents an important model for studying pattern formation and the mechanisms underlying regeneration of facial structures. We have studied regeneration of upper and lower jaws in the urodele amphibian, Notophthalmus viridescens, using whole mount preparations stained for bone and cartilage, scanning electron microscopy and immunocytochemistry to further characterize these regenerating systems. In addition, we have investigated whether lower jaws of adults and larvae display similar regenerative ability. Although in adult animals the original shape of both the lower and upper jaws is rather faithfully reproduced following amputation, and the teeth and oral mucosa with its specialized sensory organs fully regenerate, significant differences in the regenerative ability of the various skeletal elements are observed. In fact, only tooth-bearing skeletal elements ossify, while the other elements of the regenerated skeleton remain cartilaginous for as long as 5 months after amputation. In contrast, a regenerated lower jaw in the larva is indistinguishable from an unamputated one at the same stage of development. Interestingly, regenerating adult jaws form directly bicuspid teeth, which are the type of teeth normally found in the adult, rather than the monocuspid teeth characteristic of larval jaws, indicating that jaw regeneration is not a recapitulation of development, in that an adult jaw blastema directly regenerates an adult jaw. Finally, we have studied the expression of tissue specific markers in normal and regenerating upper and lower jaws to establish whether the blastemal cells, which will form the missing part of the jaw, express any of these markers of the differentiated state, or are undifferentiated as suggested by their morphological appearance. Under our experimental conditions, no expression of markers of the differentiated state, such as those for muscle, cartilage and glands is detectable in early regenerates. On the contrary, the mesenchymal marker 22/31, whose expression in normal jaws is restricted to dermal fibroblasts and the dental pulp, is expressed in at least a half of the blastemal cells. The significance of these observations in relation to the origin of blastemal cells in the jaw will be discussed.

Animals↗
PubMed1992

Heads and tales: recent advances in craniofacial development.

In the last few years, our understanding of the complex series of events governing craniofacial morphogenesis has significantly increased. A variety of experimental approaches, such as cell lineage marking, monoclonal antibodies, organ culture and recombinant DNA techniques, have been fundamental to this progress. The neural crest contribution to the head and face of the embryo has been mapped, the significance and mechanisms of epithelial-mesenchymal interactions are better understood, and a number of regulatory molecules, which are likely to be causally involved in mediating the morphogenesis of the different craniofacial regions, have been identified. Although the information presently available on the molecules involved is far from complete, and the experimental strategies need further refinement, the existence of specific combinatorial gene-codes (ie the homeobox-containing gene code) has become evident. These findings are proving invaluable to our understanding of the mechanisms underlying craniofacial development, and of certain syndromes affecting craniofacial structures in humans.

Animals↗
PubMed1991

Cell origin and identity in limb regeneration and development.

The occurrence of limb regeneration in adult urodele amphibians raises fundamental questions about the relationship between development and regeneration. The use of monoclonal antibodies as cell markers has provided clear evidence that blastemal cells, the progenitor cells of the regenerate, are not the same as limb bud cells in the embryo. For one of these antibodies the distinction has been traced to the relationship between limb regeneration and the nerve supply. Innervation of the limb bud during development appears to establish nerve-dependent growth control for regeneration. The cell markers have also contributed to the problem of how the blastemal cells arise after amputation, although several important questions remain to be answered.

Amputation, Surgical↗
PubMed1991

A newt type II keratin restricted to normal and regenerating limbs and tails is responsive to retinoic acid.

In order to understand the molecular mechanisms underlying the regenerative ability of the urodele limb, it is important to identify regeneration-associated proteins and to study their regulation. We have recently shown that the anti-cytokeratin monoclonal antibody LP1K reacts strongly with newt blastemal cells, while its reactivity is restricted in normal limbs. By screening a cDNA expression library from the newt blastema with LP1K, we have identified cDNA clones coding for a type II keratin (NvKII) expressed both in the mesenchyme and the specialized wound epithelium of the blastema. While the rod domain of the protein is highly conserved, the homology between NvKII and mammalian type II keratins drops markedly at the N- and C-terminal regions. The expression of this keratin was analysed by Northern blotting and RNAase protection analysis of various newt tissues, and appears to be organ specific, since it is restricted to normal and regenerating limbs and tails. In particular, we have investigated the expression of this keratin mRNA in normal and regenerating limbs. The transcript is barely detectable in the proximal portion of the normal limb, but its level is high in the distal one. After amputation, NvKII mRNA is expressed both in proximal and distal blastemas, although at higher levels distally, indicating that this keratin is regeneration associated. The NvKII transcript is detectable both in mesenchyme and in the wound epithelium of the regenerate, while no transcript is detectable in normal epidermis. The level of NvKII mRNA is markedly down-regulated both in normal and regenerating limbs following intraperitoneal injection with retinoic acid, a putative endogenous morphogen in limb regeneration.

Amino Acid Sequence↗
PubMed1990

The monoclonal antibody 22/18 recognizes a conformational change in an intermediate filament of the newt, Notophthalmus viridescens, during limb regeneration.

Previous work has shown that the monoclonal antibody 22/18 identifies progenitor cells (blastemal cells) which depend on the nerve for their division in the early stages of limb regeneration in the newt, Notophthalmus viridescens. This antibody also reacts with cultured cells derived from the newt limb, and the intensity of immunoreactivity appears related to cell density and differentiation into myotubes. We report here that the monoclonal antibody 22/18 recognizes a polypeptide (22/18 antigen) which is intracellular and filamentous. Double staining of cells with 22/18 monoclonal antibody and antibodies against various cytoskeletal components indicates that the epitope is expressed on an intermediate filament component. Although this antibody is specific for blastemal cells in cryostat sections of the regenerating limb, its reactivity on immunoblots is not confined to this tissue. The 22/18 antigen is differentially affected by aldehyde fixatives distinguished by the spacing of their reactive groups. While formaldehyde fixation impairs detection of the antigen, ethylene glycol-bis[succinic acid n-hydroxysuccinimide ester] reveals the antigen in sections of normal and regenerating limbs in a distribution that is consistent with the one obtained from immunoblots. We suggest that the 22/18 monoclonal antibody detects a change in protein conformation, probably related to changes in the physiological state of the cell, that occurs transiently during regeneration and possibly during development.

Animals↗
PubMed1989

Transient expression of simple epithelial keratins by mesenchymal cells of regenerating newt limb.

Structural proteins of the intermediate filament family are an early indicator of differentiation before organogenesis becomes apparent. Keratin intermediate filaments are characteristically expressed only by epithelial and not by mesenchymal cells. Here we show, using monoclonal antibodies, a transient expression of the keratin pair 8 and 18 in a population of mesenchymal cells in the regenerating newt limb, specifically in the undifferentiated progenitor cells (blastemal cells) which give rise to the new tissues. These keratins are also expressed in cultured limb cells that can differentiate into muscle. In contrast no reactivity with anti-keratin 8 and 18 antibodies was observed in the newt limb bud at an early stage of development, indicating a molecular difference between the developing and regenerating limb. The molecular weights of the newt proteins detected by these antibodies are very similar to those of human keratins 8 and 18, further supporting the immunocytochemical evidence that the newt homologs of these keratins are expressed in blastemal cells. This is the first demonstration of keratin expression in mesenchymal progenitor cells in an adult animal.

Animals↗
PubMed1988

Culture of newt cells from different tissues and their expression of a regeneration-associated antigen.

We have established culture conditions for cells from normal limb, early limb regenerate (blastema), heart, and liver of the newt Notophthalmus viridescens. Whereas heart and liver cells had a relatively short life in culture, limb cells have shown no sign of senescence over more than 1 year in culture. Cultured cells from all these tissues express to differing extents the regeneration-associated antigen 22/18. The antigen is intracellular and filamentous, and its expression appears to be regulated by culture density. Furthermore, 22/18 antigen is turned off in limb and blastemal cultures following differentiation into muscle, as also occurs in vivo.

Animals↗
PubMed1986

An immunohistochemical study of synaptogenesis in the electric organ of Torpedo marmorata by use of antisera to vesicular and presynaptic plasma membrane components.

Synaptogenesis has been studied in the electric organ of embryonic Torpedo marmorata by use of two antisera directed against components of synaptic vesicles (anti-SV) and presynaptic plasma membranes (ap-anti-TSM), respectively. The anti-SV serum was previously shown to recognize a proteoglycan specific for synaptic vesicles. The ap-anti-TSM serum was raised to plasma membranes of synaptosomes derived from the electromotor nerve terminals and affinity-purified on electric-organ gangliosides. The vesicular antigen was first detectable at the 81-mm stage of development, which is 1-2 weeks earlier than the formation of morphologically mature presynaptic terminals, but is coincident with a rise in choline acetyltransferase levels and the ability of the electric organ to generate discharges. The gangliosidic antigen recognized by the ap-anti-TSM was first detectable on the ventral electrocyte surface at the 93-mm stage of development. This indicates that specific carbohydrate epitopes, not present on the growth cones, are expressed during maturation of the nerve terminal. The nerve terminal components recognized by these sera arose pari passu with neurite coverage of the ventral surface of the electrocyte, reaching a maximum in the adult. In contrast, postsynaptic aggregates of acetylcholine receptor, rendered visible with rhodamine-labeled alpha-bungarotoxin, arose previous to the presynaptic antigens, reaching a maximum surface density at 110 mm and then declining in the adult.

Animals↗
PubMed1986

Putative cholinergic-specific gangliosides in guinea pig forebrain.

The nature of the cholinergic-specific antigen Chol-1 recognized by an antiserum raised against Torpedo cholinergic electromotor synaptosomal plasma membranes was investigated in guinea pig forebrain to establish whether it has a gangliosidic nature in guinea pig as in Torpedo. Gangliosides extracted from guinea pig forebrain and extensively purified to eliminate peptide contaminants were effective in inhibiting the selective lysis of the cholinergic subpopulation of cortical synaptosomes induced by the antiserum. Neuraminidase, protease, alkali, and heat treatment did not impair the inhibitory activity of gangliosides. Whereas the antiserum recognized many gangliosides from Torpedo electric organ, the immunostaining of guinea pig forebrain gangliosides separated on TLC showed only two immunopositive bands migrating close to GT1b and GQ. After affinity purification on Torpedo electric organ gangliosides the activity of the antiserum in inducing complement-mediated lysis was increased and it still recognized the two ganglioside bands on TLC. These results strongly suggest the existence of two polysialogangliosides bearing antigenic determinants specific for the cholinergic neurons.

Animals↗
PubMed1986

Inhibition of gastrointestinal transit by morphine in rats results primarily from direct drug action on gut opioid sites.

The local intestinal component of the constipating action of morphine was assessed through several integrated approaches in an in vivo animal model. The doses of systemically administered morphine reducing to 50% of drug-free controls (ID50) the small intestinal transit of a charcoal meal fed by gavage to overnight fasted rats were 0.04 and 3.8 mg/kg i.p. and p.o. and 0.5 mg/kg either s.c. or i.v., respectively. Transit inhibition with any of these morphine doses occurred within 10 min and was still measurable 20, 30, 45 and 240 min after i.p., i.v., s.c. and p.o. administration, respectively. Morphine given by any of these systemic routes did not reduce significantly transit in rats receiving the putative peripheral antagonist quaternary naloxone (1 mg/kg i.p., 5 min before morphine) that, unlike naloxone, failed to reverse transit inhibition (to about 50% of drug-free controls) induced by 0.08 mg/kg i.c.v. of morphine. Radioassay of thin-layer chromatograms of extracts of central nervous system, plasma, small intestine and small intestinal longitudinal muscle of rats given tritium-labeled morphine and also tested for gastrointestinal transit, showed that morphine concentrations in the longitudinal muscle (with attached myenteric plexus) after i.v., i.p. and s.c. injection fell within a relatively narrow range and were consistent with the appropriate transit scores. Morphine levels in the central nervous system of the same rats were lower than in any other tissue assayed, presented considerable differences depending on administration routes and did not correlate at all with the corresponding intestinal effects. Morphine administered directly into the rat cerebral ventricles effectively inhibits gastrointestinal transit through an opioid-sensitive central nervous system-located action site.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗