Shibata Y, Takiguchi H, Abiko Y
Department of Biochemistry, Nihon University School of Dentistry at Matsudo, Chiba, Japan.
J Periodontol. 1999 Oct;70(10):1158-65. doi: 10.1902/jop.1999.70.10.1158.
Matrix metalloproteinases (MMPs), produced by both infiltrating and resident cells of the periodontium, play a role in physiologic and pathologic events. It is recognized that an imbalance between activated MMPs and their endogenous inhibitors leads to pathologic breakdown of the extracellular matrix during periodontitis. Although it is known that pro-MMPs are activated by the plasminogen activator (PA)/plasmin system, and that the activated MMPs are inactivated by tissue inhibitor of metalloproteinases (TIMPs), participation of TIMPs in the PA/plasmin system has not been defined.
We investigated the effects of the antisense oligonucleotide, consisting of a 21-base sequence from the human TIMP-1 gene including the first ATG initiation codon, on PA/plasmin activities in the cultured medium of periodontal ligament (PDL) fibroblastic cells. Antisense or sense oligonucleotides were directly added into cell-cultured medium, and enzyme activities from the PDL cells were measured.
Antisense TIMP-1 oligonucleotide specifically stimulated the PA activity dose-dependently. Other oligonucleotides, sense TIMP-1 or antisense TIMP-2, did not affect PA activity in PDL cells. The PA activity increased by antisense TIMP-1 oligonucleotide was due to an increase of urokinase-type PA (uPA) protein, but not that of tissue-type PA by means of immunoblotting. Furthermore, the stimulation of PA activity in the conditioned medium by adding antisense oligonucleotide for TIMP-1 was not due to the decreasing levels of PA inhibitor-1, an inhibitor of PA.
TIMP-1 controls the synthesis of uPA in the PDL cells. Control of the TIMP-uPA system is important in inflammatory periodontal ligament healing.
基质金属蛋白酶（MMPs）由牙周组织中的浸润细胞和常驻细胞产生，在生理和病理过程中发挥作用。人们认识到，活化的MMPs与其内源性抑制剂之间的失衡会导致牙周炎期间细胞外基质的病理性破坏。虽然已知前MMPs由纤溶酶原激活物（PA）/纤溶酶系统激活，且活化的MMPs被金属蛋白酶组织抑制剂（TIMPs）灭活，但TIMPs在PA/纤溶酶系统中的参与情况尚未明确。
我们研究了由人TIMP-1基因包含第一个ATG起始密码子的21个碱基序列组成的反义寡核苷酸对牙周膜（PDL）成纤维细胞培养基中PA/纤溶酶活性的影响。将反义或正义寡核苷酸直接添加到细胞培养基中，并测量PDL细胞的酶活性。
反义TIMP-1寡核苷酸剂量依赖性地特异性刺激PA活性。其他寡核苷酸，正义TIMP-1或反义TIMP-2，对PDL细胞中的PA活性没有影响。通过免疫印迹法，反义TIMP-1寡核苷酸增加的PA活性是由于尿激酶型PA（uPA）蛋白增加，而非组织型PA增加。此外，添加TIMP-1反义寡核苷酸对条件培养基中PA活性的刺激并非由于PA抑制剂-1（PA的一种抑制剂）水平降低。
TIMP-1控制PDL细胞中uPA的合成。TIMP-uPA系统的调控在炎症性牙周膜愈合中很重要。
