Raymond Purves Bone and Joint Research Laboratories, Level 10, Kolling Institute of Medical Research B6, The Royal North Shore Hospital, St. Leonards, NSW, 2065, Australia.
Graduate School of Biomedical Engineering, University of New South Wales, Sydney, NSW, 2052, Australia.
J Mol Histol. 2019 Jun;50(3):285-294. doi: 10.1007/s10735-019-09823-1. Epub 2019 Apr 16.
The aim of this study was to ascertain whether, like many cell types in cartilaginous tissues if type XI collagen was a pericellular component of annulus fibrosus (AF) cells and chondrocytes. Fine fibrillar networks were visualised which were perlecan, HS (MAb 10E4) and type XI collagen positive. Heparitinase-III pre-digestion abolished the type XI collagen and 10E4 localisation in these fibrillar assemblies demonstrating a putative HS mediated interaction which localised the type XI collagen. Type XI collagen was confirmed to be present in the Heparitinase III treated AF monolayer media samples by immunoblotting. Heparitinase-III generated ΔHS stub epitopes throughout these fibrillar networks strongly visualised by MAb 3-G-10. Monolayers of murine hip articular chondrocytes from C57BL/6 and Hspg2 exon 3 null mice also displayed pericellular perlecan localisations, however type XI collagen was only evident in the Wild type mice. Perlecan was also immunolocalised in control and murine knee articular cartilage from the two mouse genotypes subjected to a medial meniscal destabilisation procedure which induces OA. This resulted in a severe depletion of perlecan levels particularly in the perlecan exon 3 null mice and was consistent with OA representing a disease of the pericellular matrix. A model was prepared to explain these observations between the NPP type XI collagen domain and HS chains of perlecan domain-I in the pericellular matrix of AF cells which likely contributed to cellular communication, tissue stabilization and the regulation of extracellular matrix homeostasis.
本研究旨在确定型胶原是否像软骨组织中的许多细胞类型一样，是纤维环 (AF) 细胞和软骨细胞的细胞周成分。观察到细纤维网络，其对硫酸乙酰肝素（MAb 10E4）和型胶原呈阳性。肝素酶-III 预处理消除了这些纤维组装物中的型胶原和 10E4 定位，表明存在潜在的 HS 介导的相互作用，该相互作用将型胶原定位。通过免疫印迹证实型胶原存在于肝素酶 III 处理的 AF 单层培养基样品中。肝素酶-III 在这些纤维网络中生成 ΔHS 短突表位，MAb 3-G-10 强烈可视化。来自 C57BL/6 和 Hspg2 外显子 3 缺失小鼠的 hip 关节软骨单层也显示细胞周层粘连蛋白定位，然而仅在野生型小鼠中可见型胶原。层粘连蛋白也在对照和两种小鼠基因型的膝关节关节软骨中免疫定位，这两种小鼠基因型均接受内侧半月板不稳定程序处理，该程序可诱导 OA。这导致层粘连蛋白水平严重耗竭，特别是在外显子 3 缺失的小鼠中，这与 OA 代表细胞周基质疾病一致。制备了一个模型来解释这些在 AF 细胞细胞周基质中的 NPP 型胶原结构域和层粘连蛋白结构域-I 的 HS 链之间的观察结果，这可能有助于细胞通讯、组织稳定和细胞外基质动态平衡的调节。
