Morphological and biochemical re‐evaluation of the process of cavitation in the rat knee joint: cellular and cell strata alterations in the interzone

Abstract

To assess the contribution of apoptosis to the mechanism of synovial joint cavitation, and to clarify morphological cellular changes during cavitation, we investigated the development of the rat knee joint by light and electron microscopy, TUNEL methods, and electrophoresis of DNA fragments. Although cavitation occurred within the interzone, which consists of 2 outer and a middle layer termed the intermediate zone, no morphological or biochemical signs of cell death, in particular apoptosis, were seen in the interzone at any embryonic stage. Microscopic and ultrastructural alterations affecting cell differentiation were clearly observed in the interzone, i.e. mesenchymal cells gradually showed elongation, cytoplasmic vacuolation and pyknosis in the intermediate zone where the elongated cells were arranged in parallel in some strata. Some of these cells were further flattened into spindle cells and the number of strata decreased to 2. The rest of the cells were incorporated secondarily into the outer layers, becoming chondroblasts. Collagen fibrils were arranged in a network structure in the outer layers, which obviously differed from the directional pattern parallel to the long axis of elongated cells in the intermediate zone. In addition, the density of collagen fibrils was higher in the outer layers than in the intermediate zone. During cavitation, the initial separation was detected between the elongated cells in the intermediate zone in paraffin sections at E16n5 and the spindle cells in epoxy sections at E18n5. The spindle cells lining the cavity, namely, the surfaces of the epiphysis and meniscus, finally became chondrocytes. The diminution of proteoglycans and collagen fibrils and the synthesis of hyaluronan in the extracellular matrix are now generally believed to be parts of the mechanism for cavitation based on the concept of ' loss of cohesion '. The microscopic and ultrastructural alterations in the interzone seemed to reflect differences in the arrangement and density of collagen fibrils and the developmental condition of the extracellular matrix between layers. Also it did not seem likely that these alterations inhibit the synthesis of hyaluronan at the presumptive joint line because this synthesis takes place at the plasma membrane. Separation between spindle cells should therefore represent the mechanism for developmentally programmed cavitation. Reorganization of the extracellular matrix is probably necessary for the cellular metamorphoses in the interzone involved in the process of cavitation.

Development of the articular cavity in the rat temporomandibular joint with special reference to the behavior of endothelial cells and macrophages

“…In spite of lack of detailed information, these hypotheses have been directly adopted on the mechanism of cavity formation in the TMJ, which is categorized in secondary cartilaginous joints such as sternoclavicular and acromioclavicular joints. For instance, many researchers have failed to find apoptosis at the cavity-forming area in long bone joints (Ballard and Holt, 1968;Murray and Drachman, 1969;Mitrovic, 1977Mitrovic, , 1978Mori et al, 1995;Nalin et al, 1995;Kimura and Shiota, 1996;Kavanagh et al, 2002), while some studies have detected it (Abu-Hijleh et al, 1997;Ito and Kida, 2000). Matsuda et al (1997) revealed no detection of apoptosis in this area by biochemical and histochemical analyses with the TUNEL method in TMJ development, consistent with the present observations.…”

Birth and death of cells in limb development: A mapping study

“…Recently, the possibility of a role of cell death in joint formation has been questioned (Ito and Kida, 2000;Archer et al, 2003).…”

Formation of synovial joints and articular cartilage

“…This view was challenged by several authors who assumed that articular cartilage is formed by the cells lying at the periphery of the cavitated joint and derived from the cells of interzone [4,8,28]. The authors of these publications showed that the interzone is ultrastructurally distinguishable into 2 outer layers contiguous to the epiphyseal ends and a thin central intermediate zone.…”

“…WNT14 expression seems to be sufficient to direct joint development as judged by its ability to induce and/or maintain a panel of gene interzone by clearing up the cells with chondroblastic potentialities. In another study, however, cell death was not observed during cavitation of the developing joints in rat or rabbit knee [28,31]. Macrophages were present prior to cavitation in the periphery of joint interzones but not at the presumptive joint line in the central region of interzone [14].…”