Prevention and Treatment of Thrombosis Associated With Long-Term Hemodialysis Catheters

Soft, cuffed, central vein hemodialysis catheters are used in about 20% of chronic hemodialysis patients in the United States, because long-term arteriovenous blood access cannot be maintained in an aging patient population with a large proportion of diabetics. The most frequent complication of these catheters is thrombosis. The treatment of catheter-related thrombosis is difficult and expensive; thus the emphasis should be on prevention. The preferred material for a long-term catheter is silicone rubber, since it is the least thombogenic. Anticoagulation should be more vigorous during “catheter dialysis” than during “fistula dialysis.” Heparin is the least expensive and most convenient anticoagulant, suitable for over 99% of chronic dialysis patients. The dose of heparin for sufficient anticoagulation depends on many factors, varies widely, and should be established for each patient based on activated clotting time (ACT). ACT should be kept over 270 sec throughout dialysis. Recently we introduced a method of locking catheter lumina with a predetermined amount of heparin; this heparin is not discarded before the next dialysis, but serves as a loading dose. This saves a number of connections/ disconnections and decreases dialysis-associated blood losses. To prevent catheter thrombosis, over 60% of patients require warfarin in sufficient doses to keep the international normalized ratio (INR) between 1.5 and 2.5. The most common catheter-related thrombus is a periluminal fibrin sleeve. Locking the catheter with urokinase to dissolve the clot is of little value, because the bulk of the thrombus is outside the catheter. We have found a high-dose (250 000 U or more) intradialytic urokinase infusion through the venous chamber to be a very efficient and convenient method for dissolving clots. Cumulative success of up to three infusions is over 99%. This obviates the need of catheter stripping or replacement, which is more cumbersome and expensive.     

用弱化dhfr双顺反子载体在CHO细胞中高效表达低分子量尿激酶突变体

将二氢叶酸还原酶基因(dhfr)克隆至pIRES载体中弱化的脑心肌炎病毒(ECMV)内部核糖体进入位点(IRES)的下游,构建了含有弱化dhfr筛选标记和可用氨甲蝶呤(MTX)加压提高表达水平的双顺反子真核表达载体pIRES-dhfr.利用该表达载体表达了一种无糖基化和具有凝血酶抗性的低分子量尿激酶型纤溶酶原激活剂(LMW-uPA)突变体(缺失尿激酶原的1-143位氨基酸,Arg156→Lys,Asn302→Ala),用脂质体转染方法将表达载体pIRES-dhfr/LMW-UK转染CHO-dhfr-细胞后经过一轮MTX筛选,几乎所有获得的单克隆细胞株都表达LMW-UK,其中约50%为表达水平较接近(500-5000 IU/106cells/d)的高表达阳性克隆.用转瓶无血清培养表达水平约为17.5pg/cell/d的rCHO细胞系LB2-UK,收集的上清经过阳离子交换柱和凝胶过滤层析两步纯化后,获得的重组蛋白的纯度可以达到99%.用S-2444发色底物法检测,所获得的突变体双链UK比例大于98%.     

Successful Long-Term Central Venous Access; Daily Home Hemodialysis Blood Access

Successful long-term central venous access is a complex subject. The concept of “long term” implies that continued surveillance will be required. This also requires the catheter to be placed, initially, in its best configuration. To achieve long-term performance and durability, a thorough understanding of all aspects related to the catheter, catheter placement, and catheter maintenance is essential.