Cartilage canals in the distal intermediate ridge of the tibia of fetuses and foals are surrounded by different types of collagen

Abstract

Some epiphyseal growth cartilage canals are surrounded by a ring of hypereosinophilic matrix consisting of collagen type I. Absence of the collagen type I ring may predispose canal vessels to failure and osteochondrosis, which can lead to fragments in joints (osteochondrosis dissecans). It is not known whether the ring develops in response to programming or biomechanical force. The distribution that may reveal the function of the ring has only been described in the distal femur of a limited number of foals. It is also not known which cells are responsible for producing the collagen ring. The aims of the current study were to examine fetuses and foals to infer whether the ring forms in response to biomechanical force or programming, to describe distribution and to investigate which cell type produces the ring. The material consisted of 46 fetuses and foals from 293 days of gestation to 142 days old, of both sexes and different breeds, divided into three groups, designated the naïve group up to and including the day of birth, the adapting group from 2 days up to and including 14 days old, and the loaded group from 15 days and older. The distal tibia was sawn into parasagittal slabs and the cranial half of the central slab from the intermediate ridge was examined by light microscopy and immunohistochemical staining for collagen type I. Presence, completeness and location of the collagen ring was compared, as was the quantity of perivascular mesenchymal cells. An eosinophilic ring present on HE-stained sections was seen in every single fetus and foal examined, which corresponded to collagen type I in immunostained sections. A higher proportion of cartilage canals were surrounded by an eosinophilic ring in the naïve and adapting groups at 73 and 76%, respectively, compared with the loaded group at 51%. When considering only patent canals, the proportion of canals with an eosinophilic ring was higher in the adapting and loaded than the naïve group of foals. The ring was present around 90 and 81% of patent canals in the deep and middle layers, respectively, compared with 58% in the superficial layer, and the ring was more often complete around deep compared with superficial canals. The ring was absent or partial around chondrifying canals. When an eosinophilic ring was present around patent canals, it was more common for the canal to contain one or more layers of perivascular mesenchymal cells rather than few to no layers. It was also more common for the collagen ring to be more complete around canals that contained many as opposed to few mesenchymal cells. In conclusion, the proportion of cartilage canals that had an eosinophilic ring was similar in all three groups of fetuses and foals, indicating that the presence of the collagen ring was mostly programmed, although some adaptation was evident. The ring was more often present around deep, compared with superficial canals, indicating a role in preparation for ossification. The collagen ring appeared to be produced by perivascular mesenchymal cells.

Keywords: cartilage canal; collagen type I; epiphyseal growth cartilage; foal; histology; immunohistochemical staining; osteochondrosis; tibia.

Figure 1

Collection method. (A) The distal tibia was sawn parallel to the ridge in a slightly oblique parasagittal plane. The cranial half of the most centrally positioned slab (asterisk) was selected for further analysis. (B) For descriptive purposes, the tissue in each section was divided into cranial, apical and articular aspects and the thickness of the articular‐epiphyseal cartilage complex was divided into three equally thick layers of superficial, middle and deep. SOC, secondary centre of ossification.

Figure 2

Patent cartilage canals in epiphyseal growth cartilage from the distal intermediate ridge of the tibia. (A) Three‐day‐old male Standardbred, HE, 100×. Patent canal without an eosinophilic ring. (B) Fetus at 295 days of gestation, female Standardbred, HE, 50×. Patent cartilage canals surrounded by a complete eosinophilic ring (between arrows), articular aspect, deep layer. (C) Same foal and area inside stippled box in (B), immunostained for collagen type I, 100×. The eosinophilic ring in HE‐stained sections corresponded to extracellular matrix, immunostained with polyclonal antibodies against equine collagen type I. (D) The same section as (D), control‐stained with non‐immune serum (isotype).

Figure 3

Chondrifying canals in epiphyseal growth cartilage from the distal intermediate ridge of the tibia. (A) Thirty‐eight day‐old male Warmblood, HE, 50×. Partial eosinophilic ring (arrows) in the late chondrifying part of the canal (asterisk) in the middle layer of the cartilage does not have an eosinophilic ring, whereas the early chondrifying part in the deep layer of the cartilage is surrounded by a weak‐staining eosinophilic ring (arrowheads). (B) Forty‐five day‐old male Quarter Horse, HE, 100×. This late chondrifying canal does not have an eosinophilic ring.

Figure 4

Perivascular mesenchymal cells. (A) Female Warmblood Riding Horse fetus at 293 days of gestation, HE, 100×. When an eosinophilic ring was present around patent cartilage canals (asterisk), it was more common for the canal to contain one or more layers than few to no layers of perivascular mesenchymal cells separating the blood vessels from the surrounding cartilage. (B) Higher power magnification of (A), HE, 400×, showing perivascular mesenchymal cells (arrows) immediately adjacent to eosinophilic ring.