Kim K T
Department of Oral Biochemistry, Kanagawa Dental College, Japan.
Bull Kanagawa Dent Coll. 1990 Sep;18(2):89-98.
The periodontium, especially the periodontal ligament and alveolar bone, are tissues constantly subjected to physical stress such as occlusion and mastication. This study was designed to explore the effect of the pulsed electromagnetic fields (PEMF) on the cell attachment and the spread of human periodontal ligament fibroblasts (HPLF) and rat osteoblasts (ROB). PEMF are categorized as one type of mechanical stress. HPLF were obtained by the explantation method described by Saito et al. They were then subcultured in Dulbecco's modified Eagle's medium (D-MEM) and supplemented with 2 mg/ml dialyzed fetal calf serum protein (FCSP), 50 micrograms/ml ascorbic acid and penicillin/streptomycin after trypsinization. ROB were isolated from a two-day-old rat calvaria by the sequential bacterial collagenase digestion method described by Dziak and Brand and were subcultured in D-MEM supplemented with FCSP, ascorbic acid and penicillin/streptomycin. After the confluent HPLF were cultured with serum-free MCDB 107 medium, the quiescent HPLF were exposed with or without PEMF for 24 hr. This was followed by the collection of the control conditioned medium (C-CM) and PEMF exposed conditioned medium (PEMF-CM). The cell attachment assay was performed so that the hydrophobic 24 multiwells were coated with the whole conditioned medium or fractionated conditioned medium by a PO-60K column. After coating, heat inactivated BSA blocked nonspecific sites for cell adhesion, and 3H-TdR labeled HPLF or ROB were cultured on the precoated wells. The activity of cell attachment and spreading was determined by the radioactivity of 3H-TdR using a scintillation counter. The characters of cell attachment factors derived from HPLF were hydrophobic, heat labile and proteolytic enzyme digestible. In addition, the fractionated PEMF-CM enhanced the spreading activity of ROB. PEMF induced the 10 KDa which can enhance the HPLF and ROB spreading. Therefore, the cell attachment and spreading factors secreted by HPLF exposed with PEMF may regulate HPLF and also ROB.
牙周组织，尤其是牙周膜和牙槽骨，是不断受到诸如咬合和咀嚼等物理应力作用的组织。本研究旨在探讨脉冲电磁场（PEMF）对人牙周膜成纤维细胞（HPLF）和大鼠成骨细胞（ROB）的细胞附着及铺展的影响。PEMF被归类为一种机械应力。HPLF通过Saito等人描述的组织块培养法获得。然后将其在胰蛋白酶消化后，接种于添加了2mg/ml透析胎牛血清蛋白（FCSP）、50μg/ml抗坏血酸以及青霉素/链霉素的杜氏改良 Eagle 培养基（D-MEM）中进行传代培养。ROB通过Dziak和Brand描述的连续细菌胶原酶消化法从2日龄大鼠颅骨中分离出来，并在添加了FCSP、抗坏血酸以及青霉素/链霉素的D-MEM中传代培养。将汇合的HPLF用无血清MCDB 107培养基培养后，使静止的HPLF在有或无PEMF的条件下暴露24小时。随后收集对照条件培养基（C-CM）和暴露于PEMF的条件培养基（PEMF-CM）。进行细胞附着试验，通过PO-60K柱将疏水的24孔板用全条件培养基或分级条件培养基包被。包被后，用热灭活的牛血清白蛋白封闭细胞黏附的非特异性位点，并将3H-TdR标记的HPLF或ROB接种于预先包被的孔中。使用闪烁计数器通过3H-TdR的放射性来测定细胞附着和铺展的活性。源自HPLF的细胞附着因子的特性是疏水、热不稳定且可被蛋白酶消化。此外，分级的PEMF-CM增强了ROB的铺展活性。PEMF诱导产生了可增强HPLF和ROB铺展的10KDa蛋白。因此，暴露于PEMF的HPLF分泌的细胞附着和铺展因子可能对HPLF以及ROB均有调节作用。
