Enomoto H, Yamashita S, Usa T, Namba H, Ohtsuru A, Villadolid M C, Tsukazaki T, Matsumoto T, Iwasaki K
Department of Orthopaedic Surgery, Nagasaki University School of Medicine, Japan.
Biochem Biophys Res Commun. 1993 Mar 31;191(3):1261-9. doi: 10.1006/bbrc.1993.1353.
Parathyroid hormone-related peptide (PTHrP) may be synthesized in or near its target tissues and acts by autocrine and/or paracrine fashions. The rat clonal strain of the osteoblast-like cell, ROS 17/2.8-5, can express low levels of PTHrP in the cytoplasm; however, the autocrine function of PTHrP in ROS cells has not yet been clarified. We created PTHrP expression vectors and transfected them into cells in order to elucidate the functional role of PTHrP on their own target cells. Sense and antisense rat PTHrP expression vectors (pSV2 neo-ECE-rPLP) were transfected into ROS cells independently and cultured for 72 hours. Cells overexpressing PTHrP were detected by immunocytochemical analysis and confirmed by Northern blot analysis, respectively. These transfected cells demonstrated mitogenic activity as determined by BrdU uptake staining. These findings suggest the functional role of PTHrP on their own PTHrP expressing cells via an autocrine/paracrine fashion. These PTHrP-overexpressing ROS cells provide a model in vitro system to clarify the mechanism by which PTHrP acts in an autocrine/paracrine fashion.
甲状旁腺激素相关肽（PTHrP）可能在其靶组织内或附近合成，并以自分泌和/或旁分泌方式发挥作用。成骨细胞样细胞的大鼠克隆株ROS 17/2.8-5可在细胞质中表达低水平的PTHrP；然而，PTHrP在ROS细胞中的自分泌功能尚未阐明。我们构建了PTHrP表达载体并将其转染到细胞中，以阐明PTHrP对其自身靶细胞的功能作用。将有义链和反义链大鼠PTHrP表达载体（pSV2 neo-ECE-rPLP）分别独立转染到ROS细胞中，并培养72小时。通过免疫细胞化学分析检测过表达PTHrP的细胞，并分别通过Northern印迹分析进行确认。通过BrdU摄取染色确定，这些转染细胞表现出促有丝分裂活性。这些发现表明PTHrP通过自分泌/旁分泌方式对其自身表达PTHrP的细胞发挥功能作用。这些过表达PTHrP的ROS细胞提供了一个体外系统模型，以阐明PTHrP以自分泌/旁分泌方式发挥作用的机制。
