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Biomedical subjects

D M Lawton

Publications and source records attributed to D M Lawton.

10 recordsLinked to original sources

Mature osteoblasts in human non-union fractures express collagen type III.

AIMS: High levels of collagen type III are biochemically detectable in biopsies of non-uniting fractures, and in the serum of patients suffering from this condition. The aim of this study was to determine whether the expression of collagen type III was limited to fibrous tissue in non-unions, or whether some was present in bone. METHODS: Biopsies from normally healing human fractures and non-unions were examined using in situ hybridisation and immunohistochemistry. RESULTS: The mesenchymal cell population, which includes fibroblast and osteoblast precursors, expressed mRNA for collagen type III. However, mature osteoblasts on the surface of woven bone varied profoundly between normally healing fractures (in which they were negative or occasionally weakly positive) and non-unions (in which they were strongly positive). Areas of woven bone that had osteoblasts positive for collagen type III mRNA also immunostained positively for the protein. CONCLUSIONS: This study shows that non-union fracture callus osteoblasts on the surfaces of woven bone exhibit an unusual phenotype: they express collagen type III, a molecule characteristic of an earlier stage of osteoblast differentiation, which is not expressed by osteoblasts on woven bone surfaces of bone that develops normally. This finding may be useful in developing an early clinical test for impending non-union.

Adolescent

The biological reality of the interlacunar network in the embryonic, cartilaginous, skeleton: a thiazine dye/absolute ethanol/LR White resin protocol for visualizing the network with minimal tissue shrinkage.

Third toe phalanges of chicks aged 8-13 days in ovo and 7-day post-natal rat femoral growth plate were examined to determine whether the interlacunar network (IN), a structure with no lipoprotein membrane component or cytoplasmic organelles, is a genuine component of young growth cartilage. In chick phalanges dehydrated by 70% (v/v) ethanol and LR White resin, variable metachromatic staining of the interlacunar network by toluidine blue and red staining by picro-Sirius red indicate the presence of glycosaminoglycans and collagen. The network in phalanges dehydrated by 80% (v/v) ethanol appears little different; however, the network is much less widely detectable in phalanges dehydrated by 90% (v/v) ethanol and, after dehydration by absolute ethanol, is almost completely undetectable. In contrast, when the young cartilage is permeated by a thiazine dye such as toluidine blue, using a solution of dye in the aldehyde fixative, the network is widely detectable, following dehydration by absolute ethanol, both in chick phalanges and in rat growth plate. Comparison of projected areas shows that the extent to which whole chick feet are found to have shrunk, by the time that they are photographed under LR White resin, is determined principally by the extent of dehydration, by 70% (v/v) or absolute ethanol; post-shrinkage areas are 33% or 35% of areas measured in buffer for 70% (v/v) ethanol/LR White resin and 71% or 75% for absolute ethanol/LR White resin (the higher value in each is for the toluidine blue treatment). The network is thus present in radically shrunk tissue, but, significantly, is also fully represented in tissue shrunk by only a conventional margin and is therefore not produced as an artefact by exceptional tissue shrinkage as has been suggested.

Animals

New histological observations in spontaneously developing osteoarthritis in the STR/ORT mouse questioning its acceptability as a model of human osteoarthritis.

Naturally developing spontaneous osteoarthritis (OA) is seen in a number of small animals one of which is the STR/ORT mouse, an accepted model of human OA. A histological evaluation of the patello-femoral joints of 37 male STR/ORT mice has shown features that are inconsistent with the disease in human joints. These include the presence of a prominent acute and chronic synovial inflammatory infiltrate. Such findings call into question the proposed aetiology of the arthropathy in this strain of mouse and its acceptability as a model of human primary OA.

Acute Disease

Low temperature scanning electron microscopy of the superficial envelope of canine chondrocytes in culture.

Low temperature scanning electron microscopy (LTSEM) has been employed to examine the surface morphology of chondrocyte cultures on ceramic granules. Well-established cultures on porous hydroxyapatite consist of ceramic cores overlaid and interspersed with a cellular matrix of collagen and proteoglycan (Cheung, 1985); of especial interest is the superficial layer of cells. These cells are believed, on the basis of immuno-light microscopy (Gardner et al., 1987), to be coated by an hydrated porous envelope of collagen/proteoglycan which is likely to obscure cell outlines. This relationship is confirmed by enzymic digestion of the superficial material. Post-digestion LTSEM examination of the fully hydrated preparations establishes the existence of arrays of rounded structures identified as superficial cells.

Animals

Biocompatibility of hydroxyapatite ceramic: response of chondrocytes in a test system using low temperature scanning electron microscopy.

An investigation is described of R37W, one of a number of porous ceramics being developed for maxillofacial surgery and the restoration of periodontal bone defects. Sensitive and precise methods are needed to assess the biocompatibility of these new materials. Mammalian chondrocytes are known to form colonies on and within porous ceramics; therefore, the tissue formed has been evaluated to gauge the response of these proliferating mesenchymal cells to the hydroxyapatite. Cell colonies, grown on R37W, have been rapidly frozen at -210 degrees C (63K) in nitrogen slush and examined by low temperature scanning electron microscopy (SEM). This method enables unfixed, fully hydrated cells to be viewed in detail and demonstrates the three-dimensional surface structure of chondrocytes in a life-like state. Features such as complex pericellular fenestrations and papilliform surface processes are believed to indicate cell viability and normality: they are not detectable by the light microscopy and SEM of fixed, dehydrated preparations. This recently recognized fine structure, together with the determination of rates of increase of cell numbers and histochemical and immunological tests of cell synthetic and secretory behaviour, provides a new guide to biocompatibility. It is concluded that the low temperature SEM of chondrocytes grown on ceramics is a valuable addition to the procedures available for the testing of hard materials before their adoption in oral surgery.

Animals

The apparent reflexion coefficient of the leaky corneal endothelium to sodium chloride is about one in the rabbit.

1. Rabbit corneal thickness changes were measured after some of the NaCl in the bathing Ringer solution was substituted by a neutral sugar. 2. The response had three phases which could be closely modelled by three exponentials of different amplitudes and rate constants, originating from the time of the substitution. 3. The first, fastest, phase was interpreted as being driven by the pure osmotic pressure difference developed across the corneal endothelium by the difference between the removed NaCl and the added sugar in the bathing Ringer solution; the second was driven by the diffusion of NaCl out of the stroma; and the third, slowest, phase was driven by the diffusion of the added sugar into the stroma. 4. Consistent with the interpretation, only the third, slowest, phase had its rate constant dependent upon the nature of the substituting neutral sugar. 5. The amplitude of the pure osmotic phase was a linear function of the added sugar. Its amplitude was zero when an equal osmolarity of sugar was substituted for NaCl. 6. It was concluded that the reflexion coefficient of rabbit corneal endothelium to NaCl is the same as that to sucrose, raffinose and stachyose, i.e. about 1. 7. The calculated hydraulic conductivity of the endothelium plus stroma was about the same as that of stroma alone, and it was concluded that the hydraulic conductivity of corneal endothelium is large compared to corneal stroma. 8. It is proposed that most of the hydraulic flow in response to osmotic gradients passes through the cells, whereas salt diffuses through the paracellular route, resulting in an apparent reflexion coefficient of 1. 9. Thus, corneal endothelium is a 'leaky' (12 omega cm2) salt-permeable epithelium and, according to the present study, simultaneously a semi-permeable membrane. To resolve the terminology, we suggest the term 'bi-permeable' for such epithelia which have a high salt permeability but such a very high water permeability as to give apparent reflexion coefficients of 1.

Animals