Development of a therapeutic anti-marinobufagenin antibody

在PNX大鼠中，我们研究了3E9抗MBG单抗对血压及心肌肥厚和纤维化的影响。在PNX大鼠中，血浆MBG水平升高四倍与高血压、心脏碳化蛋白水平增加、心肌肥厚、心脏胶原合成负调控因子Fli-1的表达减少以及胶原-1水平增加有关。一次给PNX大鼠腹腔注射3E9单抗可使血压降低59毫米汞柱，持续7天，并显著降低心脏重量和心脏氧化应激水平。这些效应与Fli-1表达的增加和心脏纤维化的减少有关。因此，在慢性肾功能衰竭中，MBG通过抑制Fli-1促进高血压和诱导心脏纤维化，这是一个潜在的治疗靶点。 由于先前在尿毒症大鼠中，主动免疫MBG可减少心肌纤维化，但对血压的影响最小，因此我们假设MBG通过非BP依赖机制诱导血管纤维化。我们测定了盐负荷雄性Wistar链脲佐菌素诱导的2型糖尿病大鼠(DM-NaCl)和正常对照大鼠的血压、血、尿MBG、主动脉胶原-1和血管功能。本实验观察了胸主动脉环对硝普钠(SNP)的反应性。糖尿病-氯化钠组大鼠在不改变血压的情况下，MBG排泄量增加3.5倍，胸主动脉胶原-1水平增加2.5倍。与对照组相比，糖尿病-氯化钠大鼠的主动脉环对硝普钠的松弛反应减弱。体内注射3E9单抗对大鼠血压无明显影响，但可降低主动脉中胶原-1的水平，恢复主动脉环对硝普钠的敏感性。 因此，MBG能够在不影响血压的情况下增加血管硬度。

In PNx rats, we studied effects of 3E9 anti-MBG monoclonal antibody (mAb) on BP and cardiac hypertrophy and fibrosis. In PNx rats, a four-fold elevation in plasma MBG levels was associated with hypertension, increased cardiac levels of carbonylated protein, cardiac hypertrophy, a reduction in cardiac expression of a nuclear transcription factor which is a negative regulator of collagen synthesis, Fli-1, and an increase in the levels of collagen-1. A single intraperitoneal administration of 3E9 mAb to PNx rats reduced BP by 59 mmHg for 7 days and significantly reduced cardiac weight and cardiac levels of oxidative stress. These effects were associated with an increase in the expression of Fli-1, and a reduction in cardiac fibrosis. Thus, in chronic renal failure MBG contributes to hypertension and induces cardiac fibrosis via suppression of Fli-1, representing a potential target for therapy. Because previously in uremic rats active immunization against MBG reduced cardiac fibrosis but minimally affected BP, we hypothesized that MBG induces vascular fibrosis via BP-independent mechanism. We determined BP, plasma and urinary MBG, aortic collagen-1, and vascular function in NaCl-loaded male Wistar rats with streptozotocin-induced type 2 diabetes mellitus (DM-NaCl) and in control animals. Isolated rings of thoracic aortae were tested for their responsiveness to sodium nitroprusside (SNP) following endothelin-1-induced constriction. DM-NaCl rats exhibited a 3.5-fold increase in MBG excretion and 2.5-fold increase in levels of collagen-1 in thoracic aortae without changes of BP. As compared to control, aortic rings from DM-NaCl rats exhibited impaired response to the relaxant effect of SNP. In vivo administration of 3E9 mAb to DM-NaCl rats did not affect BP, but reduced aortic levels of collagen-1, and restored sensitivity of aortic rings to SNP. Thus, MBG is capable to increase vascular stiffness without affecting BP.