Ameri A, Kuppuswamy M N, Basu S, Bajaj S P
Department of Medicine, St Louis University School of Medicine, MO.
Blood. 1992 Jun 15;79(12):3219-26.
We recently proposed that endothelium may represent the primary physiologic site of synthesis of the tissue factor pathway inhibitor (TFPI). In support of this conclusion, we have now found that the poly(A)+ RNAs obtained from rabbit and bovine lung tissues contain abundant amounts of TFPI messenger RNAs (mRNAs), whereas the poly(A)+ RNAs obtained from the liver of these animals contain less than 5% of that found in the lung tissues. Because inflammatory mediators are known to upregulate tissue factor (TF) expression by the endothelium, we have examined the effect of these agents on the TFPI expression by the cultured endothelial cells. When cultured human umbilical vein endothelial cells were stimulated (in 10% fetal bovine serum) with phorbol myristate acetate (PMA), endotoxin, interleukin-1, or tumor necrosis factor-alpha, the TF mRNA increased approximately 7- to 10-fold within 2 to 4 hours. Unstimulated cells constitutively expressed TFPI mRNA and its levels either did not change or increased slightly (up to 1.5-fold) upon stimulation with these inflammatory agents. TF mRNA abruptly declined to a negligible level and the TFPI mRNA returned essentially to the basal level at approximately 24 hours. The membrane-bound TF clotting activity of induced cells peaked between 4 and 8 hours, and finally declined. The cumulative TFPI activity secreted into the media was either unchanged or slightly higher in the induced cell cultures as compared with that present in the noninduced cultures. Endothelial cells were also cultured in 10% heat-inactivated human serum derived from plasma or whole blood. TFPI secreted into the media containing whole blood serum was consistently higher (approximately 1.5-fold at 8 hours) than that secreted into the media supplemented with serum obtained from plasma lacking the formed elements; these cells also expressed similarly increased levels of TFPI mRNA. Moreover, PMA-stimulated cells cultured in whole blood serum expressed modestly increased levels of TFPI mRNA (approximately 1.5-fold); supernatants from these cells also contained similarly increased TFPI activity. Cumulatively, our data indicate that, unlike thrombomodulin and fibrinolytic enzymes synthesized by the endothelial cells, TFPI synthesis is not downregulated and may be slightly upregulated during an inflammatory response. Inspection of the 5' flanking region of the TFPI gene showed a conserved GATA-binding motif located approximately 400 bp upstream of the proposed transcription initiation site(s). This motif by binding to the GATA-2 transcriptional factor may keep the endothelium in an 'on' state for constitutive expression of TFPI.(ABSTRACT TRUNCATED AT 400 WORDS)
我们最近提出，内皮可能是组织因子途径抑制剂（TFPI）合成的主要生理部位。为支持这一结论，我们现已发现，从兔和牛肺组织获得的聚腺苷酸（poly(A)+）RNA含有大量TFPI信使RNA（mRNA），而从这些动物肝脏获得的poly(A)+ RNA所含的量不到肺组织中的5%。由于已知炎症介质可上调内皮细胞的组织因子（TF）表达，我们研究了这些介质对培养的内皮细胞TFPI表达的影响。当用佛波酯肉豆蔻酸酯乙酸盐（PMA）、内毒素、白细胞介素-1或肿瘤坏死因子-α刺激培养的人脐静脉内皮细胞（在10%胎牛血清中）时，TF mRNA在2至4小时内增加约7至10倍。未刺激的细胞组成性表达TFPI mRNA，在用这些炎症介质刺激后，其水平要么不变，要么略有增加（高达1.5倍）。TF mRNA在约24小时时突然降至可忽略不计的水平，而TFPI mRNA基本恢复到基础水平。诱导细胞的膜结合TF凝血活性在4至8小时达到峰值，最终下降。与未诱导的细胞培养物相比，诱导细胞培养物中分泌到培养基中的TFPI累积活性要么不变，要么略高。内皮细胞也在10%热灭活的源自血浆或全血的人血清中培养。分泌到含有全血血清的培养基中的TFPI始终高于分泌到补充有不含有形成分的血浆获得的血清的培养基中的TFPI（8小时时约为1.5倍）；这些细胞也表达类似增加水平的TFPI mRNA。此外，在全血血清中培养的PMA刺激细胞表达适度增加水平的TFPI mRNA（约1.5倍）；这些细胞的上清液也含有类似增加的TFPI活性。总体而言，我们的数据表明，与内皮细胞合成的血栓调节蛋白和纤溶酶不同，TFPI的合成在炎症反应期间不会下调，反而可能略有上调。对TFPI基因5'侧翼区域的检查显示，在拟转录起始位点上游约400 bp处有一个保守的GATA结合基序。该基序通过与GATA-2转录因子结合，可能使内皮处于TFPI组成性表达的“开启”状态。（摘要截短至400字）
