Fibrous Structures.

The fibrils of ’unknown origin’ are an interesting feature of some patients (C) with 01. They have a close association with type I collagen fibrils and in some micrographs appear to be continuous and ’splay’ out from type I collagen.

This would indicate that they are indeed unravelled type I collagen fibrils or possibly the type III widely reported to associate with type I.

Haebara et al (1969) reported thin, delicate fibres, but without a striation periodicity and o f thickness 40nm. Parry (1977) reported the presence of subfibrils in rat

tail tendon. Some type I collagen appeared to have ’exploded’ into subfibrils. In both transverse and longitudinal section, these subfibrils had a uniform diameter of 140A. They reported that the percentage of fibrils which had ’exploded’ was very low, the frequency of observation was high due to the large number of fibrils observed in a typical section. This was a feature of the OI bone, low percentage with higher frequency. In longitudinal section thay reported that the subfibrils sometimes appeared to emanate from a D-periodic collagen but more commonly the parent fibril showed no obvious D period. It was thought that the subfibrils might not be coUageneous but were in fact elastic fibres as reported by Ross and Bomstein (1969). However the collagen ous nature of the subfibrils was suggested by the following evidence :

a) The microfibrillar component of an elastic fibre is tubular, beaded and lOOA in diameter. Subfibrils reported by Parry and Craig (1977) were 140A and the fibrils reported here were much larger and were not beaded;

b) In transverse section, the subfibrils were composed of yet smaller units approximately 10-20 nm in diameter, with many hundreds of these units per subfibril. Parry and Craig correctly surmised that the evidence for these subfibrils being collagen ous was not conclusive and that elastic fibres should not be ruled out. A note added at the proof stage of their publication concluded that these fibrils were indeed elastic as demonstrated by elastin-specific staining. However, the evidence for these subfibrils being elastin in nature is still not convincing. Amianthoid fibrils, elastin, oxytalin or elaunin fibres (Smith et al. 1982) are still discounted from this study by the criteria of ultrastrucural characteristics.

Campo and Phillips (1973) reported fraying or unravelling of collagen fibrils in bovine costal cartilage, periodicity was evident at the point of fraying. They concluded that this fraying represented a disaggregation of collagen into smaller constituent fibrils. The incorporation of an abnormal type I collagen trimer into a type I collagen fibril could result in the formation of an unstable fibril which is degraded into its sub-units. These ’fibres of unknown origin’ may represent these breakdown products.

Patients (A and B) demonstrated a more amorphous fibrous structure which were large enough to be seen at the light microscopic level after staining with toluidine blue. No unequivocal explanation can be offered as to the nature of these fibres. Again, there was an indication in certain sections that these fil^s were continuous with type I collagen. Immunohistochemistry is undoubtedly the next step, but to simply ’throw antibodies at sections and hope they will stick’ will only confuse the nature of this material further. Firstly, some idea about the nature of these fibrous structures is required and this is best obtained by further ultrastructural analysis of more biopsies. Certainly, suggestions as to the nature of the fibres can be offered. At the present time the most likely candidate is an altered type I collagen formed de novo or a breakdown product. To prove this is more difficult than first predicted. Because of the close association between these fibres and ultrastructurally normal type I collagen the chances of non-specific labelling of these fibres by an anti type I collagen antibody are very high, this would give a wrong conclusion.

However, as with the problem of detecting type III collagen in association with type I collagen, the amount of protein is very low and may be ’buried’ in the fibrillar structure, making access to the antigen difficult. Equally, detecting the presence of an altered type I collagen protein by methods such as electrophoresis will not yield an unequivocal answer. Some of the type I collagen fibrils that appear normal will undoubtedly have altered collagen chains incorporated and these could appear on the electrophoretogram as abnormal bands which could be mistaken as having come from these fibrous structures. 2 .4 .6 . Growth Plate

Subtle changes can occur in the epiphyseal growth plate in the more severe lethal disease where the normal distribution of the cartilagen ous columns can be distorted and diminished.

In the 3 specimens of neonatal or foetal growth plate examined in this study, attenuated bony trabeculae and relatively disordered cartilage epiphyseal columns were found when compared to normal controls. It was from these specimens that the trabeculae demonstrated the stromal calcification. Woven bone was also a common feature, however, it was difficult to distinguish pathology from immaturity. The obvious pathology associated with lethal, perinatal type II 01 makes it difficult to correctly assess ultrastructural changes that are a direct result of the molecular disease and not simply part of the rapidly changing picture of foetal/neonatal morphology.

However, it is widely accepted that the overall type I collagen abnormality in the connective tissue is the cause of death and not any specific abnormality of bone. The reduced production of type I collagen undoubtedly accounts for the attenuated, bony trabeculae. No real explanation can be given for the partly distorted cartilagen ous columns seen in some patientsr^wever, the inconsistency of observation may hold some clue to this. Many previous reports, although not all (see below) have indicated that the cartilagen'^csus component of 01 bone is normaf^erhaps these abnormalities are patient specific, resulting from physical trauma in utero or postnatal trauma.