Although the histology of post -foetal osteogenesis is
well recognised, much uncertainty still remains about the
fundamental problems of the causal genesis, the differential
potencies and activities, and the metabolism of the osteogenic cells in bone growth and repair.
In the past, the histochemistry of osteogenesis has
been limited to the inorganic phase of mineralisation and
there has been remarkably little work on the preceding phase
of the stimulus to cellular differentiation, and on the
organic phase which is concerned with the pre-osseous
cellular metabolism and elaboration of the bone matrix and is
the precursor stage for the inorganic phase of new bone
formation. An important intercellular substance of this
organic phase is the sulphated mucopolysaccharide complex,
chondroitin sulphuric acid, which is secreted by the osteogenic cells - the osteoblast and the chondroblast.
In the present study the importance of this chemical
substance is emphasised because it controls and indeed
identifies the morphological appearance of the cartilage cell
and hence their possible origin, and because is is closely
related to the subsequent deposition of the calcium phosphate
complexes by acting as a "calcium- phosphate acceptor" in the
bone matrix.
A histochemical method is described of identifying
chondroitin sulphuric acid in undecalcified bone sections
which were obtained by an original technique. The histochemical localisation of this substance was obtained by
staining for metachromasia and by "labelling" the sulphate
radicle in the complex with radioactive sulphur with the
process of autoradiography. The two techniques were
complementary to each other and provided a high degree of
specificity of localisation of chondroitin sulphuric acid.
The accuracy of this method is discussed.
By serial sacrifice of groups of rats in which a uniform fracture had been inflicted, the chondroitin
sulphuric acid complex was studied histochemically in vivo
during epiphyseal growth and repair of the fracture. Further
analysis of the normal pattern of this substance was obtained
by producing abnormal hormonal states by the administration o oestradiol dipropionate, parathormone and thyroxin; an abnormal nutritional state of hypervitaminosis A; altered
vascularity by the use of a local haemostatic substance
Adrenoxyl and by ligation; a mechanical effect of internal
fixation with use of an intrame dullary nail; and an enzymatic
action by the local application of testicular hyaluronidase.
The varying effects are recorded, correlated and discussed,
especially from the morphogenetic aspect.
The significance of the accumulation of mast cells
in the vicinity of the fracture as the source of secretion of
histamine to act as the stimulus for cellular differentiation
was studied after the local injection of a histamine- liberat.:
substance - Compound 48/80.
The problem of resorption of the organic phase of
bone structure was studied histochemically after the injection of a known chelating agent - ethylene -diamine tetra - acetic acid - and the negative results are considered.
During the last two hundred years a large volume of
experimental work has been undertaken to solve the fundamental problems of osteogenesis by studying the transplantation of osseous tissue. However, such work has been of
little value because of the lack of histochemical information
about the osteogenic cells and also because of the inability
to define and to discriminate accurately between the relative
osteogenetic activities and properties of the tissue transferred and those of the host or the environment of the
tissue transfer.
In the present study there are recorded for the first
time histochemical results from tissue culture in vivo of
various forms of osteogenic cells whose activity was clearly
defined by a physical and cellular barrier from the tissue
of the host. From Algire's original diffusion chamber
technique, a new method of tissue culture in vivo of post - embryonic tissues, which consisted of a diffusion chamber made of Millipore filter membranes, was developed and is
described in detail. The physico -chemical properties of
these filter membranes were accurately evaluated.
The histochemistry of post-foetal osteogenesis with
particular reference to the metabolise of chondroitin
sulphuric acid was studied during the processes of endochondral and intramembranous ossification resulting from the
survival and differentiation of osteogenic cells of a fracture callus and of periosteum by this tissue culture
technique in vivo. The osteogenetic potency and activity
of various osseous transfers including cortical bone were
studied and the cellular contents of the haversian systems
in cortical bone were shown by their metabolism of radioactive
chondroitin sulphuric acid to survive for at least seven days
after transplantation.
To demonstrate the process of induction, an attempt
was made to discover the possible secretion and passage into
the surrounding host tissues of a specific osteogenic
inductor substance from cells which exhibited osteogenetic
activity within a diffusion chamber. Since the process of
induction was not observed to take place, the necessity for
the presence of a physico - chemical pathway for the transmission of any specific inductor substance is emphasised and
the general reaction of the relative properties of the host
tissue to those of the tissue transfers is discussed.
Although the results from the present study are
not immediately applicable to clinical problems, their
consideration will lead to further understanding of the
host -tissue transfer relationship as well as of the histochemical nature and importance of the organic phase of
post-foetal osteogenesis.
