DataSheet1_The tendon interfascicular basement membrane provides a vascular niche for CD146+ cell subpopulations.docx

Introduction: The interfascicular matrix (IFM; also known as the endotenon) is critical to the mechanical adaptations and response to load in energy-storing tendons, such as the human Achilles and equine superficial digital flexor tendon (SDFT). We hypothesized that the IFM is a tendon progenitor cell niche housing an exclusive cell subpopulation.
Methods: Immunolabelling of equine superficial digital flexor tendon was used to identify the interfascicular matrix niche, localising expression patterns of CD31 (endothelial cells), Desmin (smooth muscle cells and pericytes), CD146 (interfascicular matrix cells) and LAMA4 (interfascicular matrix basement membrane marker). Magnetic-activated cell sorting was employed to isolate and compare in vitro properties of CD146+ and CD146− subpopulations.
Results: Labelling for CD146 using standard histological and 3D imaging of large intact 3D segments revealed an exclusive interfascicular cell subpopulation that resides in proximity to a basal lamina which forms extensive, interconnected vascular networks. Isolated CD146+ cells exhibited limited mineralisation (osteogenesis) and lipid production (adipogenesis).
Discussion: This study demonstrates that the interfascicular matrix is a unique tendon cell niche, containing a vascular-rich network of basement membrane, CD31+ endothelial cells, Desmin+ mural cells, and CD146+ cell populations that are likely essential to tendon structure and/or function. Contrary to our hypothesis, interfascicular CD146+ subpopulations did not exhibit stem cell-like phenotypes. Instead, our results indicate CD146 as a pan-vascular marker within the tendon interfascicular matrix. Together with previous work demonstrating that endogenous tendon CD146+ cells migrate to sites of injury, our data suggest that their mobilisation to promote intrinsic repair involves changes in their relationships with local interfascicular matrix vascular and basement membrane constituents.

引言：束间基质（interfascicular matrix, IFM；又称腱内膜（endotenon））对储能肌腱（如人类跟腱与马浅指屈肌腱（superficial digital flexor tendon, SDFT））的力学适应及载荷应答至关重要。本研究提出假说：束间基质是肌腱祖细胞龛，容纳一类独特的细胞亚群。方法：本研究通过对马浅指屈肌腱进行免疫标记，定位CD31（内皮细胞）、结蛋白（Desmin，平滑肌细胞与周细胞）、CD146（束间基质细胞）及层黏连蛋白α4（LAMA4，束间基质基底膜标志物）的表达模式，以此鉴定束间基质龛。采用磁活化细胞分选术分离CD146阳性与CD146阴性细胞亚群，并对比二者的体外特性。结果：通过标准组织学方法及大型完整三维样本的三维成像对CD146进行标记，结果显示存在一类独特的束间细胞亚群，其定位于形成广泛互连血管网络的基膜附近。分离得到的CD146阳性细胞仅表现出微弱的矿化（成骨）与脂质生成（成脂）能力。讨论：本研究证实，束间基质是一类独特的肌腱细胞龛，包含富含血管的基底膜网络、CD31阳性内皮细胞、结蛋白阳性壁细胞及CD146阳性细胞群，这些组分可能对肌腱结构与/或功能至关重要。与本研究初始假说相悖的是，束间CD146阳性细胞亚群并未表现出干细胞样表型。反之，本研究结果表明CD146可作为肌腱束间基质内的泛血管标志物。结合此前研究证实内源性肌腱CD146阳性细胞可迁移至损伤部位，本研究数据提示，此类细胞动员以促进固有修复的过程，涉及它们与局部束间基质血管及基底膜组分之间相互关系的改变。