Palate development in the TGF-beta 3 knockout mouse. Low vacuum scanning electron microscopy reveals changes in the medial edge epithelium.
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
M W Ferguson
Publications and source records attributed to M W Ferguson.
Ontogeny of the skin and the transition from scar-free to scarring phenotype during wound healing in the pouch young of a marsupial, Monodelphis domestica.
The marsupial Monodelphis domestica provides a versatile model for the study of "fetal" wound healing. The pouch young of this species have been used to perform a systematic histological study of wound healing and skin development throughout pouch life; from Pouch Day 0 to Pouch Day 30 with comparisons drawn against adults. We have demonstrated that pouch young heal without macroscopic scars if wounded before Pouch Day 9 and that a transition into a scarring healing phenotype occurs around the ninth pouch day. The inflammatory reaction of wounds and aspects of skin differentiation such as dermal collagen organisation, adipose layer development, hair follicle growth, and epithelial maturation have been documented using simple ranking scales to highlight trends through pouch life. Inflammation was noted to become prominent after Pouch Day 9. There was also a general temporal correlation between the other aspects of differentiation and the transition into a scarring phenotype. It is concluded that although fetal wounds may have different cytokine profiles due to the differences of inflammatory reaction compared to adults, other aspects of skin development play a role in determining the healing phenotype.
Aging↗
The expression of the gene coding for parathyroid hormone-related protein (PTHrP) during tooth development in the rat.
By means of in situ hybridisation studies, it is shown that parathyroid hormone-related protein (PTHrP) mRNA is strongly expressed in the developing enamel organs of rat teeth. In particular, the cervical loop hybridises strongly with the PTHrP probe and expression is maintained at this site throughout life in the permanently erupting incisor teeth. In mature molar teeth, expression is downregulated to low levels and confined to the epithelial cell rests of Malassez and/or cementoblasts which may derive from these. The gene is also expressed at low levels in the tissue overlying the erupting molars and, thereafter, in the junctional epithelia and connective tissue cells of the epithelial attachment on all tooth surfaces. The premise that PTHrP may undergo post-translational processing and that the resultant products could act in different ways raises the possibility of its exerting multiple paracrine actions during tooth development. These could include the control of cell division and local vascular dilation during development.
Animals↗
Transforming growth factor-beta 3 is required for secondary palate fusion.
Mice lacking TGF-beta 3 exhibit an incompletely penetrant failure of the palatal shelves to fuse leading to cleft palate. The defect appears to result from impaired adhesion of the apposing medial edge epithelia of the palatal shelves and subsequent elimination of the mid-line epithelial seam. No craniofacial abnormalities were observed. This result demonstrates that TGF-beta 3 affects palatal shelf fusion by an intrinsic, primary mechanism rather than by effects secondary to craniofacial defects.
Animals↗
Neutralisation of TGF-beta 1 and TGF-beta 2 or exogenous addition of TGF-beta 3 to cutaneous rat wounds reduces scarring.
Exogenous addition of neutralising antibody to transforming growth factor-beta 1,2 to cutaneous wounds in adult rodents reduces scarring. Three isoforms of transforming growth factor-beta (1, 2 and 3) have been identified in mammals. We investigated the isoform/isoforms of TGF-beta responsible for cutaneous scarring by: (i) reducing specific endogenous TGF-beta isoforms by exogenous injection of isoform specific neutralising antibodies; and (ii) increasing the level of specific TGF-beta isoforms by exogenous infiltration into the wound margins. Exogenous addition of neutralising antibody to TGF-beta 1 plus neutralising antibody to TGF-beta 2 reduced the monocyte and macrophage profile, neovascularisation, fibronectin, collagen III and collagen I deposition in the early stages of wound healing compared to control wounds. Treatment with neutralising antibodies to TGF-betas 1 and 2 markedly improved the architecture of the neodermis to resemble that of normal dermis and reduced scarring while the control wounds healed with scar formation. Exogenous addition of neutralising antibody to TGF-beta 1 alone also reduced the monocyte and macrophage profile, fibronectin, collagen III and collagen I deposition compared to control wounds. However, treatment with neutralising antibody to TGF-beta 1 alone only marginally reduced scarring. By contrast, wounds treated with neutralising antibody to TGF-beta 2 alone did not differ from control wounds. Interestingly, exogenous addition of the TGF-beta 3 peptide also reduced the monocyte and macrophage profile, fibronectin, collagen I and collagen III deposition in the early stages of wound healing and markedly improved the architecture of the neodermis and reduced scarring. By contrast, wounds treated with either TGF-beta 1 or with TGF-beta 2 had more extracellular matrix deposition in the early stages of wound healing but did not differ from control wounds in the final quality of scarring. This study clearly demonstrates isoform specific differences in the role of TGF-betas in wound healing and cutaneous scarring. TGF-beta 1 and TGF-beta 2 are implicated in cutaneous scarring. This study also suggests a novel therapeutic use of exogenous recombinant, TGF-beta 3 as an anti-scarring agent.
Animals↗
The effects of ageing on cutaneous wound healing in mammals.
The dogma that cutaneous wound healing is impaired as a function of age is largely unsubstantiated. This can be attributed to poor experimental design of human studies, the lack of subject characterisation with the exclusion of disease processes, and the study of inappropriate animal models. Structural and functional changes in skin with age have been reported, such as a decrease in dermal thickness, decline in collagen content, a subtle alteration in the glycosaminoglycan profile, and a loss of elasticity, but these reports are subject to the above criticisms in addition to the often-neglected requirement for site specificity. Wound repair can be thought of as a culmination of three major overlapping phases: inflammation, proliferation and remodelling. The inflammatory process has not been studied systematically with respect to age, and despite a reported decline in cellular function and number, there is a confounding increase in the production of specific cytokines involved in the process of repair. The proliferative phase is associated with a loss of cellular responsiveness to specific cytokines with a decline in motility and proliferation; however caution in interpreting these findings is important as, for example, the definition of 'ageing' is used rather loosely with the result that neonatal versus young adult cells are compared instead of young versus old adults. During remodelling, fibronectin and collagen production may increase with age, as may wound contraction; the deposition of elastin has not been assessed and the resulting mechanical properties of the scar are controversial, not least because human in vivo studies have been ignored. The absence of a critical review on the effects of advancing age on wound healing has conspired to permit the perpetuation of the belief that well defined tenets exist. This review aims to redress this imbalance and to highlight the need for well designed research into an increasingly important field.
Aged↗
Mathematical models for growth in alligator (Alligator mississippiensis) embryos developing at different incubation temperatures.
A variety of model-based (growth models) and model-free (cubic splines, exponentials) equations were fitted using weighted-nonlinear least squares regression to embryonic growth data from Alligator mississippiensis eggs incubated at 30 and 33 degrees C. Goodness of fit was estimated using a chi 2 on the sum of squared, weighted residuals, and run and sign tests on the residuals. One of the growth models used (Preece & Baines, 1978) was found to be superior to the classical growth models (exponential, monomolecular, logistic, Gompertz, von Bertalanffy) and gave an adequate fit to all longitudinal measures taken from the embryonic body and embryonic mass. However, measurements taken from the head could not be fitted by growth models but were adequately fitted by weighted least squares cubic splines. Data for the stage of development were best fitted by a sum of 2 exponentials with a transition point. Comparison of the maximum growth rates and parameter values, indicated that the growth data at 30 degrees C could be scaled to 33 degrees C to multiplying the time by a scaling factor of 1.2. This is equivalent to a Q10 of about 1.86 or, after solving the Arrhenius equation, an E++ of 46.9 kJmol-1. This may be interpreted as indicating a common rate-limiting step in development at the 2 temperatures.
Alligators and Crocodiles↗
The distribution of PDGFs and PDGF-receptors during murine secondary palate development.
The distribution of the 2 homodimer isoforms of platelet-derived growth factor (PDGF)-AA and -BB, and their receptors (PDGF alpha-receptor and PDGF beta-receptor) were mapped immunocytochemically during the development of the embryonic mouse secondary palate, from embryonic day (E) 12 to E15. The staining patterns of PDGF-AA/alpha-receptor and PDGF-BB/beta-receptor are different. Generally, the former is present and the latter is sparse/absent during the period of palate development. The localisations of PDGF-AA and alpha-receptor are more intensive in the epithelia than in the mesenchyme, and change temporally during palatal shelf elevation, midline epithelial seam formation and disruption. Before the palatal shelf has elevated (E13), PDGF alpha-receptor is present in the palatal mesenchyme, nasal and medial edge epithelia (MEE), whilst PDGF-AA stains lightly in the palatal epithelia. During palatal midline epithelial seam formation and following its degeneration throughout E14, PDGF-AA and PDGF alpha-receptor were intensively colocalised in the nasal and midline seam epithelia. These findings indicate that the interaction between PDGF-AA and its receptor may participate in palate development, particularly in regulating palatal midline epithelial seam formation and degeneration.
Amino Acid Sequence↗
The effect of suramin on healing adult rodent dermal wounds.
Scarring, leading to impaired function, growth and appearance, is a major problem following many forms of surgery. Fetal wounds, unlike those in the adult, are characterised by a reduced growth factor profile and the absence of scar tissue (Whitby & Ferguson, 1991 a, b). The antiparasitic drug, suramin (a heparin analogue) inhibits binding of various growth factors (e.g. PDGF, bFGF, TGF-beta, EGF, IGF-I, IGF-II) to their receptors in vitro. These growth factors play key roles in wound healing. We attempted to manipulate experimentally their effectiveness in healing adult rat dermal incisional wounds by injecting suramin into the wound margins and comparing the resultant healing with an unmanipulated control wound in the same animal. Immunohistochemical staining demonstrated that, on d 7 and 14 postwounding, the numbers of monocytes/macrophages and blood vessels are markedly increased in suramin treated wounds compared with controls. Extracellular matrix deposition is lower, although very compact in organisation, lacking the usual honeycombed appearance of normal skin. These effects are widespread, being present not only in the wound area, but also in the surrounding tissue. No difference was detected at 70 d postwounding between the scars of suramin-treated and unmanipulated control wounds in the same animals. All such effects are increased slightly through the concentration range of 0.04-40 mg/kg suramin, with no significant change as concentrations greater than 40 mg/kg are applied. This suggests that suramin has marked effects on the early stages of wound healing, which plateau at 40 mg/kg concentration, but has no effect on scar formation.
Administration, Cutaneous↗
Inhibition of TGF-beta 3 (but not TGF-beta 1 or TGF-beta 2) activity prevents normal mouse embryonic palate fusion.
The critical stage of palatal development is the fusion of apposing individual palatal shelves. Palatal shelves, from the day 13 murine embryo, cultured in vitro fuse normally in the absence of exogenous factors. Therefore, some endogenous factor(s) is responsible for the normal fusion process. Prevention of mature TGF-beta 3 activity during a specific time window of development in palate organ cultures, either by antisense oligodeoxynucleotides or neutralizing antibody, resulted in failure of palate fusion. Northern analysis was used to demonstrate that the antisense treatment down-regulated TGF-beta 3 mRNA. Inhibition of TGF-beta 1 or -beta 2 activity (by either antibodies or antisense oligodeoxynucleotides) had no such effect on palate development and palate fusion was normal. These data indicate an isoform specific role for TGF-beta 3 in palatal fusion.
Animals↗
Chromosomal and temperature-dependent sex determination: the search for a conserved mechanism.
To test the hypothesis that the same (or similar) genes may be involved in chromosomal sex determination and temperature dependent sex determination we investigated the expression of the mammalian testis determining gene SRY in the American alligator (Alligator mississippiensis), a species with temperature dependent sex. We have cloned Alligator SRY-related genes (A-Srys) which constitute three major gene classes with high degrees of sequence homology (45-87%) to mammalian SRY and related genes. Expression analysis and cellular localisation studies using staged embryonic material from male and female incubation temperatures are underway to assess the role of A-Sry genes in temperature dependent sex determination.
Alligators and Crocodiles↗
The karyotype of Alligator mississippiensis, and chromosomal mapping of the ZFY/X homologue, Zfc.
Comparative mapping studies of X-linked genes in mammals have provided insights into the evolution of the X chromosome. Many reptiles including the American alligator, Alligator mississippiensis, do not appear to possess heteromorphic sex chromosomes, and sex is determined by the incubation temperature of the egg during embryonic development. Mapping of homologues of mammalian X-linked genes in reptiles could lead to a greater understanding of the evolution of vertebrate sex chromosomes. One of the genes used in the mammalian mapping studies was ZFX, an X-linked copy of the human ZFY gene which was originally isolated as a candidate for the mammalian testis-determining factor (TDF). ZFX is X-linked in eutherians, but maps to two autosomal locations in marsupials and monotremes, close to other genes associated with the eutherian X. The alligator homologue of the ZFY/ZFX genes, Zfc, has been isolated and described previously. A detailed karyotype of A. mississippiensis is presented, together with chromosomal in situ hybridisation data localising the Zfc gene to chromosome 3. Further chromosomal mapping studies using eutherian X-linked genes may reveal conserved chromosomal regions in the alligator that have become part of the eutherian X chromosome during evolution.
Alligators and Crocodiles↗
Craniofacial malformations: towards a molecular understanding.
Explore the source record for details and available documents.
Abnormalities, Multiple↗
Skin wound healing: transforming growth factor beta antagonists decrease scarring and improve quality.
Explore the source record for details and available documents.
Animals↗
Adult skin wounds in the fetal environment heal with scar formation.
OBJECTIVE: This study investigated the influence of the fetal environment on the healing characteristics of adult skin. SUMMARY BACKGROUND DATA: The remarkable ability of the fetus to heal without scarring is poorly understood. The unique qualities of fetal wound healing may be caused by the fetal environment, the fetal tissues, or a combination of both. There are numerous differences between the prenatal and postnatal environments that may play a role in the unique fetal response to injury. METHODS: Full-thickness adult sheep skin was transplanted onto the backs of 60-day-gestation fetal lambs (term, 145 days of gestation). The adult skin grafts were thus perfused by fetal blood and bathed in amniotic fluid. Previous work has demonstrated that, before midgestation, fetal lambs do not reject allogenic skin grafts. Forty days later (100 days of gestation), incisional wounds were made on both the adult skin graft and the adjacent fetal skin. The wounds were harvested 14 days postwounding and analyzed by both light microscopy and immunohistochemical testing using antibodies to collagen types I, III, and VI. RESULTS: The wounds in the adult skin grafts healed with scar formation. This observation contrasts strongly with the scarless healing of the incisional fetal skin wounds. CONCLUSIONS: This study suggests that scarless fetal skin healing properties are intrinsic to fetal skin and are not primarily the result of the fetal environment.
Amniotic Fluid↗
Effects of cytokines on gingival fibroblasts in vitro are modulated by the extracellular matrix.
Fibroblast function in gingival tissue is thought to be regulated by the local cellular environment--both the extracellular matrix and soluble factors. In an attempt to artificially re-create this situation fibroblasts have been cultured within 3-dimensional collagen gels in an environment more physiologically comparable to connective tissue. Using such a model we investigated the effects of the extracellular matrix on gingival fibroblast growth and synthetic activity and on the cellular responsiveness to 4 soluble factors--epidermal growth factor (EGF), platelet-derived growth factor (PDGF), transforming growth factor-beta (TGF-beta 1) and interleukin-1 beta (IL-1 beta). Fibroblasts cultured within collagen gels showed similar growth rates, an increased production of collagen but reduced levels of hyaluronan synthesis in comparison to cells in monolayer culture. Cellular responsiveness to soluble mediators was also modulated by the collagen matrix, with a generalised reduction in response by cells embedded within the matrix. The stimulatory effects of EGF and PDGF on cell growth in monolayer over a 14-day period were only found during the initial stages of culture within gels. Similarly the stimulation of matrix production by cells induced by TGF-beta 1 on plastic was reduced or even negated when cells were cultured in collagen gels. On plastic IL-1 beta significantly stimulated cell growth but had no effect on either collagen or hyaluronan production by fibroblasts. In gel cultures, this cytokine had no effect on cell proliferation, but significantly inhibited both collagen and hyaluronan synthesis.(ABSTRACT TRUNCATED AT 250 WORDS)
Adult↗
Neutralising antibody to TGF-beta 1,2 reduces cutaneous scarring in adult rodents.
Scarring is a major cause of many clinical problems. Scar tissue interferes with growth, impairs function and is aesthetically unpleasant. However, scarring does not appear to be a problem of embryonic life. Embryonic wounds heal with a lower inflammatory and angiogenic response and have a different growth factor profile compared to adult wounds. We have used neutralising antibody to transforming growth factor-beta 1,2 (TGF-beta 1,2) to alter the growth factor profile of cutaneous wounds in adult rodents and studied the effect on scar tissue formation. This paper extends our preliminary report that neutralising antibody to TGF-beta reduces cutaneous scarring in adult rodents. To be effective, the neutralising antibody to TGF-beta needs to be administered at the time of wounding or soon thereafter. The antiscarring effects of this neutralising antibody to TGF-beta were dose dependent. Exogenous addition of neutralising antibody to TGF-beta to incisional wounds reduced the inflammatory and angiogenic responses and reduced the extracellular matrix deposition in the early stages of wound healing without reducing the tensile strength of the wounds. Importantly, the architecture of the neodermis of wounds treated with neutralising antibody to TGF-beta resembled more closely that of normal dermis compared to the unmanipulated control wounds, which healed with an abnormal neodermal architecture resulting in obvious scarring. This study suggests a novel therapeutic approach to reducing scarring in post-natal life.
Animals↗
Sialylation of terminal saccharides of glycoconjugates expressed by murine molar tooth germs developing in vitro and in vivo.
During development of the mammalian tooth germ the pattern of terminal saccharides of glycoconjugates changes, with many structures losing lectin reactivity in a consistent pattern. This study investigated whether the epitopes are lost or become masked by terminal sialylation, using a combination of neuraminidase treatment of sections and sialic acid-reactive lectins. The results suggested that most of the terminal galactosamine and fucose sites in the epithelial enamel organ were removed during morphogenesis. Conversely, during condensation of the dental mesenchyme, masked peanut agglutinin (PNA)-reactive galactose epitopes appeared. During differentiation and organisation of the mesenchyme into odontoblasts and a subodontoblastic layer the PNA-reactive sites became masked again. These regions also specifically expressed sialylated glucosamine. However, at the proliferating epithelial cervical loop galactose sites appeared to be masked. This was more pronounced during in vitro development when abnormal expression of PNA-reactive sites was found at the cervical loop. Additionally, fucosylated sites persisted in the enamel organ, further indicating that the expression of terminal saccharides was disrupted during development in the organ culture system. These data suggest that loss of terminal galactose and galactosamine is related to differentiation of the cells. However, whether this loss occurs by removal or sialic acid masking is not dependent either on the origin of the cells or the epitope being lost.
Animals↗