Search PubMed⌕ Search

Biomedical subjects

B K Hall

Publications and source records attributed to B K Hall.

At least 37 records · Page 2Linked to original sources

Neural crest potential for tooth development in a urodele amphibian: developmental and evolutionary significance.

Tooth development in urodele amphibians occurs from a restricted region of anterior cranial neural crest. An in vitro culture system was used to test the odontogenic potential of more caudal regions of neural crest, including an "intermediate region" of neural folds which has never previously been tested for either fate or potential. Explants of different axial levels of neural crest with stomodaeal ectoderm and endoderm demonstrated that odontogenic potential extends not only further caudally than the axial level fated to produce teeth, but also beyond that with potential to produce cartilage. Our results show that chondrogenic potential is found only within the most rostral portion of the intermediate region, but that odontogenic potential extends to its most caudal limit. This separation of skeletogenic cell lineages in the neural crest necessitates a reevaluation of the designations of "cranial" and "trunk" and a reconsideration of the evolutionary implications of developmentally distinct crest-derived mesenchyme populations. The proposal that odontogenic potential extends into the trunk neural crest may be explained as conserved from a phylogenetically older, more extensive skeletogenic ability which produced the exoskeleton of more basal vertebrates.

Ambystoma mexicanum↗

Chondrogenic cell differentiation from membrane bone periostea.

Most craniofacial membrane bones are derived from neural crest (NC) cells. Interaction between NC cells and epithelium, and cellular condensation, are two major events that lead NC cells to become osteoblasts that deposit membrane bone. Unlike endochondral bone, membrane bone formation is not preceded by cartilage formation in normal development. However, chondrogenic potential in membrane bone is evidenced by several cartilage-associated phenomena in vivo. Furthermore, in vitro, periosteal cells of some membrane bones express cartilage phenotype gene products and even differentiate into chondrocytes. Hence, membrane bone periosteal cells can undergo chondrogenic differentiation. The precursor of chondrogenic cells in membrane bone is not clear: chondrocytes were proposed to arise from unipotential chondroprogenitor cells, bi- or multipotential progenitor cells, or differentiated osteogenic cells. There is experimental support for each, but studies on clonal and cell cultures provided more support for a common precursor of both chondro- and osteogenic cells. Moreover, in periostea, chondrogenesis probably arises from a differentiated cell type. Membrane bone formation in periostea may include a transient cell stage that is able to undergo both osteo- and chondrogenesis. Osteogenesis would be the normal pathway, but chondrogenesis can be evoked in certain microenvironments. It is not known whether microenvironmental factors trigger chondrogenesis through a universal molecular mechanism, nor is the molecule that triggers chondrogenesis known. Expression of neural cell adhesion molecule (NCAM) is down-regulated during commitment of periostal cells for secondary chondrogenesis, suggesting a possible regulatory role for NCAM in the alternative differentiation pathways of periosteal cells.

Animals↗

Stage-specific expression patterns of alkaline phosphatase during development of the first arch skeleton in inbred C57BL/6 mouse embryos.

Timing and pattern of expression of alkaline phosphatase was examined during early differentiation of the 1st arch skeleton in inbred C57BL/6 mice. Embryos were recovered between 10 and 18 d of gestation and staged using a detailed staging table of craniofacial development prior to histochemical examination. Expression of alkaline phosphatase is initiated at stage 20.2 in the plasma membrane of mesenchymal cells in the distal region of the first arch. Expression is strongest in osteoid (unmineralised bone matrix) and presumptive periosteum at stage 21.32. Mineralisation begins at stage E23. Expression is present in the mineralised bone matrix. Secondary cartilages form in the condylar and angular processes by stage M24. The cartilaginous cells and surrounding cells in the processes are all alkaline phosphatase-positive and surrounded by the common periosteum, suggesting that progenitor cells of the processes, dentary ramus and secondary cartilages all originate from a common pool. Nonhypertrophied chondrocytes of Meckel's cartilage express alkaline phosphatase at stage M23. Expression in these chondrocytes is preceded by the expression in their adjacent perichondrium. This is true of chondrocytes in all other cranial cartilages examined. 3-D reconstruction of expression in Meckel's cartilage also revealed that the chondrocytes of Meckel's cartilage which express alkaline phosphatase and the matrix of which undergoes mineralisation are those surrounded by the alkaline phosphatase-positive dentary ramus. By stage 25, coincident with mineralisation in the distal section of Meckel's cartilage, most chondrocytes are strongly positive. The perichondria of malleus and incus cartilages express alkaline phosphatase at stage M24. Nonhypertrophied chondrocytes along these perichondria also express alkaline phosphatase. Superficial and deep cells in the dental laminae of incisor and 1st molar teeth become alkaline phosphatase-positive at the bud stage, stages 21.16 and 21.32, respectively. Dental papillae are negative until stage M24 when alkaline phosphatase expression begins in the dental papillae and follicles of the incisor teeth and the dental follicles of the 1st molar teeth. The dental papillae of the 1st molar teeth express alkaline phosphatase at stage 25. Expression in the dental papillae and follicles appears to coincide with cellular differentiation of follicle from papilla. The presumptive squamosal, ectotympanic and gonial membrane bones, lingual oral epithelial cells connected to the dental laminae of the incisor teeth, hair follicle papillae and sheath and surrounding dermis all express alkaline phosphatase in a stage-specific manner.

Alkaline Phosphatase↗

The in vivo and in vitro effects of bone morphogenetic protein-2 on the development of the chick mandible.

During embryonic development, neural crest derived mesenchymal (ectomesenchymal) cells in the chick mandible give rise to cartilage and membrane bone. Signaling molecules involved in the development of the mandible are less understood. To examine whether BMP-2 is involved in morphogenesis and growth of the mandible in vivo, agarose beads, loaded with BMP-2 at concentrations of 5 to 150 ng/microliter were implanted into the mandible at HH stage 22 and embryos were maintained in shell-less culture. To examine whether BMP-2 is involved in osteogenic or chondrogenic differentiation, mandibular ectomesenchyme from HH stage 22 embryos was cultured in the absence of mandibular epithelium, but in the presence of BMP-2 or BMP-2 and/or type IV collagen. Chondrogenesis and osteogenesis were examined by histological, histochemical and immunohistochemical methods. Implantation of BMP-2-containing beads in vivo retarded mandibular growth and morphogenesis in a dose-dependent manner. BMP-2 induced localized death of ectomesenchymal cells in the vicinity of the implanted bead and in proportion to the concentration of BMP-2 applied. Neither BMP-2 alone, nor BMP-2+collagen type IV, was sufficient to initiate osteogenesis in vitro in the absence of epithelium. BMP-2 inhibited chondrogenesis both in vivo and in vitro. Cartilage morphology was rod-like in the absence of BMP-2 but nodular in ectomesenchyme cultured in the presence of BMP-2. These results are discussed in relation to the stimulatory and inhibitory effects of BMPs on skeletal development.

Animals↗

Paralysis and long bone growth in the chick: growth shape trajectories of the pelvic limb.

Growth of chick embryonic femora, tibiotarsi and first phalanges of digit three were measured at one day intervals from day 6 through 16 of incubation. Normal controls were compared to embryos paralyzed at 5 days of incubation. Over the 10 day study period, length of the paralyzed femora, length and width of the paralyzed tibiotarsi and differences in length of the phalanges were observed. Growth in length of phalanx one of digit three was most affected by paralysis over this period. Changes in shape of these bones also occurred during growth. Normal long bones undergo changes in shape as differential growth in length and width occurs. Such changes in shape can be considered as the bone's normal growth "trajectory". Paralyzed bones displayed a different growth trajectory than normal bones. As expected, the long bones of paralyzed embryos were shorter than age-matched controls. Contrary to expectations, however, paralyzed long bones were relatively more stout than age-matched controls.

Animals↗

In vitro differentiation potential of the periosteal cells from a membrane bone, the quadratojugal of the embryonic chick.

The quadratojugal (QJ) is a neural crest-derived membrane bone in the maxillary region of the avian head. In vivo its periosteum undergoes both osteogenesis to form membrane bone and chondrogenesis to form secondary cartilage. This bipotential property, which also exists in some other membrane bones, is poorly understood. The present study used cell culture to investigate the differentiation potential of QJ periosteal cells. Three cell populations were enzymatically released from QJ periostea and plated at different densities. Cell density greatly affected phenotypic expression and differentiation pathways. We found two culture conditions that favored osteogenesis and chondrogenesis, respectively. In micromass culture, the periosteal cells produced a layer of osteogenic cells that expressed alkaline phosphatase (APase) and secreted bony extracellular matrix (ECM). In contrast, low-density monolayer culture elicited chondrogenesis. Cells with pericellular refractile ECM and round shape appeared at 7 to 8 days and formed colonies later. The chondrogenic phenotype of these cells was confirmed by immunolocalization of type II collagen and Alcian blue staining of ECM. This result demonstrated that a fully expressed chondrogenic phenotype can be achieved from membrane bone periosteal cells in primary monolayer culture. Chondrogenesis requires a cell density lower than confluence and cannot be initiated in confluent cultures. Among the three cell populations, those cells from the outer layer have the highest growth rate and require the lowest initial plating density (below 5 x 10(3) cells/ml) to achieve chondrogenesis. Cells from the inner layer have the slowest growth rate and chondrify at the highest initial density (below 5 x 10(4) cells/ml). Chondrocytes from all populations express distinct phenotypic markers-APase and type I collagen-from initial chondrogenesis, but are not hypertrophic morphologically. Furthermore, the fact that chondrocytes arise within the same colony as APase-positive polygonal cells suggests that chondrocytes may differentiate from precursors related to the osteogenic cell lineage. This cell culture approach mimics secondary cartilage and membrane bone formation in vivo.

Alkaline Phosphatase↗

FRA3B extends over a broad region and contains a spontaneous HPV16 integration site: direct evidence for the coincidence of viral integration sites and fragile sites.

The common fragile site at 3p14.2 (FRA3B) is the most sensitive site on normal human chromosomes for the formation of gaps and breaks when DNA replication is perturbed by aphidicolin or folate stress. Although rare fragile sites are known to arise through the expansion of CCG repeats, the mechanism responsible for common fragile sites is unknown. Beyond being a basic component of chromosome structure, no biological effects of common fragile sites have been convincingly shown, although suggestions have been made that breakage and recombination at these sites may sometimes be mechanistically involved in deletions observed in many tumors and in constitutional deletions. In an observation related to the high rate of recombination at fragile sites, a number of studies have shown a statistical association between the integration of transforming DNA viruses and chromosomal fragile sites. Using FISH analysis we recently identified a 1.3 Mb YAC spanning both FRA3B and the t(3;8) translocation associated with hereditary RCC. Here we report the further localization of FRA3B within this YAC. Using lambda subclones of the YAC as FISH probes, gaps and breaks were found to occur over a broad region of at least 50 kb. Neither CCG nor CAG repeats were found in this region suggesting a different mechanism for fragility than seen with rare fragile sites. We further show that an area of frequent gaps and breaks within FRA3B, defined by a lambda contig, coincides with a previously characterized site of HPV16 integration in a primary cervical carcinoma. The HPV16 integration event gave rise to a short chromosomal deletion limited to the local FRA3B region within 3p14.2. Interestingly, 3p14.2 lies within the smallest commonly deleted region of 3p in cervical cancers, which are often HPV16 associated. To our knowledge this is the first molecular characterization of an in vivo viral integration event within a confirmed fragile site region, supporting previous cytogenetic observations linking viral integration sites and fragile sites.

Bacteriophage lambda↗

Signal transduction and TGF-beta superfamily receptors.

The TGF-beta superfamily includes a large number of related growth and differentiation factors expressed in virtually all phyla. Superfamily members bind to specific cell surface receptors that activate signal transduction mechanisms to elicit their effects. Candidate receptors fall into two primary groups, termed type I and type II receptors. Both types are serine/threonine kinases. Upon activation by the appropriate ligand, type I and type II receptors physically interact to form hetero-oligomers and subsequently activate intracellular signaling cascades, ultimately regulating gene transcription and expression. In addition, TGF-beta binds to a third receptor class, type III, a membrane-anchored proteoglycan lacking the kinase activity typical of signal transducing molecules. Type III receptors appear to regulate ligand availability to type I and type II receptors. Although a number of transduction mechanisms may be available to TGF-beta superfamily members, evidence gathered through the use of specific kinase and G-protein inhibitors and through assays measuring activation and levels of signaling intermediates suggests that at least one signaling pathway interacts with Ras and Raf proteins via a G-protein intermediate. Raf begins the cytoplasmic kinase cascade that leads to gene regulation. The myriad responses regulated by TGF-beta superfamily members makes the understanding of signal transduction mechanisms utilized by these proteins of great interest to a wide range of biological disciplines.

Animals↗

Selection of hybrids by affinity capture (SHAC): a method for the generation of cDNAs enriched in sequences from a specific chromosome region.

We have established a method for preparing cDNA sublibraries enriched in sequences from specific chromosome regions, called selection of hybrids by affinity capture (SHAC). This procedure can be described in two stages. In the first stage, a particular chromosome region, in this study mouse chromosome 11, was microdissected, followed by PCR amplification with a universal degenerate primer. This material is referred to as the "target" DNA. In the second stage, a mouse liver cDNA library with unique linker-adapter ends, referred to as the "source" cDNA, was hybridized to the biotin-labeled target DNA prepared during the first stage. The resulting DNA duplexes were captured by streptavidin-coated magnetic beads. The cDNAs were released from their biotin-labeled target homologs by alkaline denaturation and recovered by PCR amplification. These cDNAs were referred to as the SHACcDNAs. Specificity of the SHACcDNA to chromosome 11 was verified by FISH analysis. To examine representation of the SHACcDNA, we confirmed the presence of seven genes or single-copy DNA segments known to be localized on mouse chromosome 11, using a dot blot assay. In addition, a second round of SHAC was performed to achieve even higher specificity for the resulting chromosome 11 SHACcDNA. The SHAC technology should facilitate construction of cytogenetically defined cDNA libraries and should assist in the fields of gene discovery and genome mapping.

Animals↗

Mouse chromosome-specific painting probes generated from microdissected chromosomes.

Using degenerate primer amplification of chromosomes microdissected from banded cytogenetic preparations, we constructed both whole chromosome painting probes for mouse Chromosomes (Chrs) 1, 2, 3, and 11 and a centromere probe that strongly paints most mouse centromeres. We also amplified a Robertsonian translocation chromosome microdissected from unstained preparations to construct a painting probe for Chrs 9 and 19. The chromosome probes uniformly painted the respective chromosomes of origin. We demonstrated the utility of the Chr 11 probe in aberration analysis by staining mutants that we had previously identified as containing a Chr 11 translocation, and in some mutant cell lines we observed chromosome rearrangements not previously detected in stained cytogenetic preparations. The technology of microdissection and amplification applies to all mouse chromosomes or to specific subchromosomal regions and will be useful in mouse genetics, in aberration analysis, and for chromosome identification.

Animals↗

Relationships between cellular condensation, preosteoblast formation and epithelial-mesenchymal interactions in initiation of osteogenesis.

Initiation of osteogenesis or bone formation is dependent on cell and tissue interactions. We investigated the events between 4 and 7 days of incubation that translate epithelial-mesenchymal signalling into overt differentiation of osteoblasts and deposition of bone in the mandibles of chick embryos. Condensation of mandibular mesenchyme (the membranous skeleton), visualized with PNA-lectin, occurred at H.H. mid-26 (5.75 days), lasted 12 h and preceded osteoblast differentiation by 1.5 days. As determined from 3D-reconstruction all mandibular membrane bones arose from a single condensation closely associated with the stomodeal epithelium. The finding that the osteogenic condensation in the mandibular arch is a major branch of a common condensation that provides osteogenic mesenchyme to both maxillary and mandibular arches establishes a closer link between mechanisms controlling development of the skeleton in these two arches than previously suspected. Preosteoblasts (alkaline phosphatase-positive cells) form in the mandible at H.H. early 25, which is before condensation but after the epithelial-mesenchymal interaction upon which preosteoblast formation and condensation depend--neither form in isolated mesenchyme, whereas both form after recombination of mesenchyme and epithelium. Tenascin was present in the mandibular epithelium only at H.H. stage 19 but not in the mesenchyme at any age. Therefore, the epithelial-mesenchymal interaction controls initiation of osteogenesis at the preosteoblast stage. Preosteoblasts then condense, transform into osteoblasts and deposit bone matrix. Differentiation of preosteoblasts precedes condensation which amplifies their number. This is in contrast with chondrogenesis where condensation triggers prechondroblast differentiation.

Animals↗

Differential expression of neural cell adhesion molecule (NCAM) during osteogenesis and secondary chondrogenesis in the embryonic chick.

Progenitor cells in the periosteum-perichondrium of the posterior hook of the quadratojugal (QJ, a membrane bone) in the embryonic chick are bipotential for osteogenesis and chondrogenesis. These cells switch from osteogenesis to chondrogenesis between 10 to 11 days in normal (mobile) embryos but not in paralyzed (immobile) embryos. Expression of neural cell adhesion molecule (NCAM) was studied using a monoclonal antibody in QJ hooks from normal and paralyzed chick embryos between 10 and 21 days of incubation. NCAM is expressed in osteoprogenitor cells and osteoblasts but not in chondroprogenitor cells, chondroblasts, or chondrocytes. The switch of progenitor cell differentiation from an osteogenic to a chondrogenic pathway between 10 and 11 days of incubation coincides with down-regulation of NCAM expression. Both initiation of secondary chondrogenesis and down-regulation of NCAM depend on biomechanical stimulation. In embryos paralyzed at 9 days, secondary cartilage fails to form and progenitor cells remain positive for NCAM. Furthermore, paralysis influences NCAM expression in progenitor cells before secondary chondrogenesis morphologically begins, indicating that NCAM may play a role in the initiation of secondary chondrogenesis. In 15-day normal embryos, NCAM-positive cells accumulate between the perichondrium and secondary cartilage in a position that prevents further cartilage formation in the hook. In 19-day embryos, these cells lose their NCAM expression and restart chondrogenesis in a second phase of differentiation, forming an articular cartilage. Loss of NCAM expression in this cell layer and re-commencement of chondrogenesis do not occur in embryos paralyzed at 13 days, and therefore also require biomechanical stimulation. Hence, down-regulation of NCAM expression correlates with two phases of secondary chondrogenesis in embryonic life, both of which are dependent upon embryonic movement.

Animals↗

Divide, accumulate, differentiate: cell condensation in skeletal development revisited.

Cell condensation is a pivotal stage in skeletal development. Although prechondrogenic condensations normally exist for some 12 h, duration can vary. Variation is seen both between condensations for different cartilages (Meckel's vs. elastic ear cartilage) and within a single condensation from which more than one skeletal element will form, as in the three components of the single first arch chondrogenic condensation. Understanding how duration of the condensation phase is established--how the condensation phase is entered and exited during cell differentiation--remains a major area for future study. During chondrogenesis, cell-specific products such as collagen types II and IX and cartilage proteoglycan appear concomitant with condensation. Therefore, during chondrogenesis, condensation precedes commitment of cells as prechondroblasts. During osteogenesis, however, differentiation of preosteoblasts precedes condensation. Therefore, during osteogenesis, condensation amplifies the number of committed osteogenic cells. Further comparative analysis of skeletogenesis should provide us with a more rigorous understanding of cell commitment, when differentiation is initiated, how commitment and differentiation are measured and the relationship of condensation to onset of differentiation. Current knowledge of molecules characteristic of condensations focused attention on extracellular matrix and cell surface components on the one hand, and on growth factors homeobox genes and transcription factors on the other. We have drawn together the molecular data for pre-chondrogenic condensations in diagrammatic form in Figure 2. Three major phases of chondrogenesis are identified: (a) epithelial-mesenchymal interactions that precede condensation, (b) condensation itself, and (c) cell differentiation. Although we label the third phase differentiation, it is important to recognize that phases a and b also constitute aspects of chondroblast cell differentiation (see Dunlop and Hall, 1995 for a discussion of this point. The pre-condensation phase is characterized by expression of Hox genes, growth factors (TGF-beta and BMP-2) and the cell surface proteoglycan receptor, syndecan-1. Expression of Msx-1 and Msx-2, growth factors and syndecan continues into the condensation phase. Other molecules, such as versican, syndecan-3 and tenascin, present in low concentrations before condensation, are up-regulated during condensation. Yet other molecules--Hox genes, transcription factors, growth factors (activin, BMP-4 and -5, GDF-5), cell adhesion molecules and proteoglycans--are only expressed during the condensation phase, while the transcription factor Pax-1, fibronectin, hyaluronan and hyaladherin are expressed both during and after condensation. During condensation mRNAs for collagen types II and IX and for the core protein of cartilage proteoglycan are up-regulated. Late in condensation and increasingly thereafter, the protein products of these genes accumulate as chondroblasts differentiate (see Fig. 2 for details). Not all the molecules present before, during of after condensation can be placed into causal sequences. Some however can. In Figure 3 we summarize the causal sequences discussed in this paper as they relate to initiation of condensation and to transit from condensation to overt differentiation during chondrogenesis. Condensations form following activation of at least three pathways: (1) Initiation of epithelial-mesenchymal interactions by tenascin, BMP-2, TGF beta-1 and Msx-1 and -2. (2) Up-regulation of N-CAM by activin. (3) Up-regulation of fibronectin by TGF-beta, further enhancing N-CAM accumulation (Fig. 3). It is by these three pathways that condensations are initiated and grow. Transition from condensation to overt cell differentiation is under both positive and negative control (Fig. 3). Syndecan blocks fibronectin and so blocks N-CAM accumulation, preventing accumulation of additional cell

Animals↗

Multicolor FISH mapping of YAC clones in 3p14 and identification of a YAC spanning both FRA3B and the t(3;8) associated with hereditary renal cell carcinoma.

Human chromosome band 3p14 contains two tightly linked cytogenetic markers of broad interest, FRA3B and the t(3;8) breakpoint associated with hereditary renal cell carcinoma (RCC). The common fragile site at 3p14.2 (FRA3B) is the most sensitive site on normal human chromosomes to breakage when DNA replication is perturbed by aphidicolin or folate stress. The t(3;8)(p14.2;q24.1) translocation segregates with RCC in a large family and could mark the location of a tumor suppressor gene involved in renal cancers. In studies aimed at positional cloning of FRA3B and the t(3;8) breakpoint, we have used multicolor fluorescence in situ hybridization analysis (FISH) on metaphase spreads and interphase nuclei to order 14 yeast artificial chromosomes (YACs) in 3p14. The YACs used in this study were identified by a group of unordered lambda clones that had been previously localized to the 3p14 region and mapped proximal or distal to the t(3;8) breakpoint. FISH analysis was used to order the YACs and to map them in relation both to the t(3;8) translocation breakpoint and to FRA3B induced on normal chromosomes by treatment with aphidicolin. YACs that closely flanked both the t(3;8) translocation breakpoint and the fragile site were identified. A YAC walk from the closest distal YAC allowed the identification of a 1.3-Mb YAC derived from the CEPH large insert YAC library that spans both the FRA3B and the t(3;8) breakpoint. The order of the YACs and cytogenetic landmarks in 3p14 is cen-(126E1/230B9)-181H6-B15-D20F4-258B7-++ +280D2-70E12-168A8- 403B2-143C5-413C6-468B10-[850A6/t(3;8)/ FRA3B]-74B2.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

TP53 mutations are frequent in malignant NF1 tumors.

Neurofibromatosis type I (NFI) is a common autosomal dominant disorder with an increased risk for developing benign and malignant tumors. The NFI gene has been cloned and maps to 17q11.2, and the gene product acts as a tumor suppressor gene. Here we analyzed the role of mutations in TP53 in four malignant NFI tumors. Mutations were found in 3 out of 4 tumors. One of these mutations is a common missense mutation in codon 278 in one of the previously identified hot spots for mutations. The two other are hitherto unreported mutations, including a splice mutation of exon 3 and a nonsense mutation in exon 4. In addition, these four tumors also showed loss of heterozygosity (LOH) for markers on chromosome 17 in the region of TP53. Malignant NFI tumors are initiated by a somatic inactivation of the second NFI allele. Tumor progression, however, occurs by accumulation of additional genetic abnormalities, such as homozygous inactivation of TP53, as demonstrated in this paper.

Base Sequence↗

Ectopic ossicles associated with metacercariae of Apophallus brevis (Trematoda) in yellow perch, Perca flavescens (Teleostei): development and identification of bone and chondroid bone.

This paper describes the development and tissues in mineralized ossicles in the musculature of Perca flavescens infected with metacercariae of the trematode Apophallus brevis. Analysis involved light microscopy, transmission and scanning electron microscopy, X-ray scanning electron microprobe analysis, and tetracycline labelling. Two to 14 days post-infection, fibroblast-like host cells stream towards the parasite cyst forming a fusiform cellular capsule. By 14 days post-infection the capsule differentiates into an inner hypertrophied layer, an extensive middle layer of fibroblast-like cells, and a thin outer layer of flattened fibroblast-like cells forming a fibrous sheath at the capsule/muscle interface. From 21-35 days post-infection, a bony tissue is deposited periosteally in an equatorial ring around the cyst. With time, additional tissue is secreted over the ring increasing its thickness and advancing the matrix front towards the poles of the ossicle. Plump osteoblast-like cells cover the developing ossicle and may become trapped within the matrix in lacunae encapsulated by collagen. By 63 days post-infection, medium-sized ossicles are morphologically similar to large cysts from perch captured in the wild; ovoid with two polarized canals, but lacking acellular or lamellar bone-like tissue. Mineralized ossicles contain calcium, phosphorus and oxygen. Large ossicles retrieved from perch given multiple doses of tetracycline revealed discrete fluorescent bands, indicative of incremental growth. Fully developed ossicles are composed of two skeletal tissues, an inner region of chondroid bone and an outer region of acellular, lamellar bone.

Animals↗