Supplementary Material for: Perlecan Diversely Regulates the Migration and Proliferation of Distinct Cell Types in vitro

Perlecan is a multifunctional component of the extracellular matrix. It shows different effects on distinct cell types, and therefore it is thought to show potential for therapies targeting multiple cell types. However, the full range of multifunctionality of perlecan remains to be elucidated. We cultured various cell types, which were derived from epithelial/endothelial, connective and muscle tissues, in the presence of either antiserum against perlecan or exogenous perlecan, and examined the effects of perlecan on cell migration and proliferation. Cell migration was determined using a scratch assay. Blocking of perlecan by anti-perlecan antiserum inhibited the migration of vascular endothelial cells (VECs) and bone marrow-derived mesenchymal stem cells, and exogenous perlecan added to the culture medium promoted the migration of these cell types. The migration of other cell types was inhibited or was not promoted by exogenous perlecan. Cell proliferation was measured using a water-soluble tetrazolium dye. When cells were cultured at low densities, perlecan blocking inhibited the proliferation of VECs, and exogenous perlecan promoted the proliferation of keratinocytes. In contrast, the proliferation of fibroblasts, pre-adipocytes and vascular smooth muscle cells cultured at low densities was inhibited by exogenous perlecan. When cells were cultured at high densities, perlecan blocking promoted the proliferation of most cell types, with the exception of skeletal system-derived cells (chondrocytes and osteoblasts), which were inhibited by exogenous perlecan. Our results provide an overview of the multiple functions of perlecan in various cell types, and implicate a potential role of perlecan to inhibit undesirable activities, such as fibrosis, obesity and intimal hyperplasia.

串珠蛋白聚糖（Perlecan）是细胞外基质的多功能组分，其对不同细胞类型具有差异化调控效应，因此被认为具备靶向多种细胞类型的治疗潜力。然而，串珠蛋白聚糖完整的多功能调控谱系仍有待阐明。本研究培养了源自上皮/内皮、结缔组织及肌肉组织的多种细胞，分别施加抗串珠蛋白聚糖抗血清或外源性串珠蛋白聚糖进行干预，以此考察串珠蛋白聚糖对细胞迁移与增殖的影响。细胞迁移能力通过划痕实验（scratch assay）测定。实验结果表明，抗串珠蛋白聚糖抗血清阻断内源性串珠蛋白聚糖后，血管内皮细胞（vascular endothelial cells, VECs）与骨髓源性间充质干细胞的迁移受到抑制；而在培养基中添加外源性串珠蛋白聚糖，则可促进这两类细胞的迁移。其余细胞类型的迁移则受到外源性串珠蛋白聚糖的抑制，或未被其促进。细胞增殖能力采用水溶性四唑盐染料法检测。当细胞以低密度培养时，阻断串珠蛋白聚糖会抑制血管内皮细胞的增殖，而外源性串珠蛋白聚糖可促进角质形成细胞的增殖。与之相反，低密度培养的成纤维细胞、前脂肪细胞及血管平滑肌细胞的增殖，则会被外源性串珠蛋白聚糖抑制。当细胞以高密度培养时，阻断串珠蛋白聚糖可促进多数细胞类型的增殖，仅骨骼系统来源细胞（软骨细胞与成骨细胞）除外——外源性串珠蛋白聚糖会抑制这两类细胞的增殖。本研究结果全面梳理了串珠蛋白聚糖在多种细胞类型中的多重功能，并提示其可通过抑制不良细胞活动发挥潜在治疗作用，例如纤维化、肥胖及内膜增生。