Department of Orthopaedic Surgery, Zhongnan Hospital of Wuhan University, Hubei Provincial Key Laboratory of Developmentally Originated Disease, Wuhan, China.
Department of Pharmacology, Basic Medical School of Wuhan University, Hubei Provincial Key Laboratory of Developmentally Originated Disease, Wuhan, China.
Folia Morphol (Warsz). 2021;80(4):1005-1019. doi: 10.5603/FM.a2020.0137. Epub 2020 Nov 10.
At present, studies on osteochondral morphogenesis only focus on a certain period of time or only provide a pattern diagram, but lack of dynamic tracking observation from the initiation of development to maturity. This study was aimed to dynamically observe the changes of skeleton morphology and structure from embryo to adult, to provide research data for enriching the knowledge of bone and cartilage tissue structure.
In the intrauterine experiment, 5 normal pregnant Wistar rats were sacrificed under anaesthesia at gestational day (GD) 14, 17, 20, respectively. One of their offspring was randomly selected, and a total of 5 offspring were obtained at each time point. In the postnatal experiment, on the 7th and 10th day after birth and at postnatal weeks (PW) 2, 3, 6, 12, 28, 5 offspring rats from 5 different pregnant Wistar rats were randomly selected and sacrificed under anaesthesia at each time point. After obtaining the above offspring, the soft tissue was removed, and the tibia of hind limbs was retained for paraffin-embedded section. After stained with Safranin-O-fast-green and haematoxylin, the morphological development of the tibia was observed under an optical microscope.
At GD14, there was no obvious joint space, the whole hind limb was cartilage and bone tissue was not visible. At GD17, visible joint space was seen and the chondrocytes in the centre region appeared to hypertrophy. At GD20, the primary ossification centre was obvious, and a typical epiphysis growth plate structure could be seen. On the 7th day after birth, the chondrocytes in the centre of epiphysis cartilage were hypertrophic and differentiated, the cartilage canal grows from the cartilage surface toward the centre of the epiphysis cartilage, at postnatal day 14, the secondary ossification centre was formed. At this time, the tibia had typical morphological characteristics of the metaphysis, however, there was no obvious layered structure of articular cartilage; the stratified structure of articular cartilage could be seen at PW6, but its mature marker (tidemark) was still not visible; however, at PW12, typical 4 layers of articular cartilage appeared, and the tidemark was visible. The growth plates were clearly visible at PW2, 6 and 12. At PW28, growth plates could still be observed, but its morphology is abnormal.
Our results, for the first time, dynamically observed the morphological changes of osteochondral at critical period of development from embryo to adult, especially the process of cartilage canal participating in the formation of secondary ossification centre.
目前，关于骺软骨形态发生的研究仅关注某一特定时期，或者仅提供模式图，而缺乏从发育起始到成熟的动态跟踪观察。本研究旨在动态观察胚胎到成年骨骼形态和结构的变化，为丰富骨软骨组织结构知识提供研究数据。
在子宫内实验中，在妊娠第 14、17、20 天分别麻醉处死 5 只正常妊娠 Wistar 大鼠，每个时间点随机选择 1 只后代，共获得 5 只后代。在产后实验中，在出生后第 7、10 天和生后周（PW）2、3、6、12、28 时，分别从 5 只不同妊娠 Wistar 大鼠中随机选择 5 只后代大鼠并麻醉处死，每个时间点处死 1 只。获得上述后代后，去除软组织，保留后肢胫骨进行石蜡包埋切片。经番红 O-快绿和苏木精染色后，在光学显微镜下观察胫骨的形态发育。
在妊娠第 14 天，没有明显的关节间隙，整个后肢都是软骨，看不到骨组织。在妊娠第 17 天，可见关节间隙，中心区的软骨细胞出现肥大。在妊娠第 20 天，初级骨化中心明显，可见典型的骺板生长板结构。出生后第 7 天，骺软骨中心的软骨细胞肥大分化，软骨管从软骨表面向骺软骨中心生长，生后第 14 天形成次级骨化中心。此时，胫骨具有典型的干骺端形态特征，但关节软骨无明显分层结构；PW6 时可见分层结构，但成熟标志物（骺板线）仍不可见；然而，在 PW12 时，出现典型的 4 层关节软骨，可见骺板线。PW2、6 和 12 时生长板清晰可见。PW28 时仍可见生长板，但形态异常。
本研究首次动态观察胚胎到成年骺软骨在关键发育时期的形态变化，特别是软骨管参与次级骨化中心形成的过程。
