肽酶与平滑肌细胞血管紧张素II受体药理学

Cohen R B, Webb M L, Dickinson K E

Department of Cardiovascular Biochemistry, Bristol-Myers Squibb Pharmaceutical Research Institute, Princeton, NJ 08643-4000.

Peptides. 1993 Mar-Apr;14(2):345-52. doi: 10.1016/0196-9781(93)90051-h.

Angiotensin (A) II receptors on rat aortic smooth muscle (RASM) cell membranes were characterized using the radioligand [125I][Sar1Ile8]AII ([125I]SI-AII). Angiotensin I, AII, and AIII inhibited specific [125I]SI-AII binding, and their rank order of potencies, and Ki values (nM) were: AII (3.7) > AI (32.5) > or = AIII (54.0), which differed from that observed for rat adrenal cortex: AII (0.85) > AIII (3.3) >> AI (100). Similar results were observed for RASM membranes in the presence of guanine nucleotides, and for intact cells in the absence or presence of an internalization inhibitor. Lowering the incubation temperature from 37 degrees C to 4 degrees C, or inclusion of PMSF (1 mM), and preparing membranes in the presence of EGTA (1 mM) altered the rank order of potencies and Ki values (nM) of the angiotensin peptides to: AII (1.1) > AIII (7.0) >> AI (144). [125I]Angiotensin I was metabolized completely over the course of 90 min to small (<tetrapeptide) fragments as measured by HPLC. There was no evidence for formation of AII or AIII from AI, which would have explained the unusually high potency of AI. [125I]Angiotensin I metabolism could be attenuated by inhibitors of serine proteases PMSF, aprotinin, and chymostatin. The beneficial effects of PMSF and EGTA suggested that serine protease(s) and metalloproteases contribute to the observed anomalous pharmacological characteristics of AI and AIII, respectively. The RASM cell membranes contained a homogeneous population of binding sites for losartan, and its Ki value differed in the absence (50 nM) or presence (16 nM) of protease inhibitors, which suggests that the receptor may also be a target for these peptidases.(ABSTRACT TRUNCATED AT 250 WORDS)

利用放射性配体[125I][Sar1Ile8]AII（[125I]SI-AII）对大鼠主动脉平滑肌（RASM）细胞膜上的血管紧张素（A）II受体进行了表征。血管紧张素I、AII和AIII抑制特异性[125I]SI-AII结合，其效价顺序和Ki值（nM）为：AII（3.7）>AI（32.5）≥AIII（54.0），这与大鼠肾上腺皮质中观察到的情况不同：AII（0.85）>AIII（3.3）>>AI（100）。在存在鸟嘌呤核苷酸的情况下，对RASM膜以及在不存在或存在内化抑制剂的情况下对完整细胞观察到了类似结果。将孵育温度从37℃降至4℃，或加入苯甲基磺酰氟（PMSF，1 mM），以及在乙二醇双（2-氨基乙基醚）四乙酸（EGTA，1 mM）存在下制备膜，改变了血管紧张素肽的效价顺序和Ki值（nM），变为：AII（1.1）>AIII（7.0）>>AI（144）。通过高效液相色谱法测定，[125I]血管紧张素I在90分钟内完全代谢为小的（<四肽）片段。没有证据表明AI可形成AII或AIII，否则就可以解释AI异常高的效价。[125I]血管紧张素I的代谢可被丝氨酸蛋白酶抑制剂PMSF、抑肽酶和糜蛋白酶抑制剂减弱。PMSF和EGTA的有益作用表明，丝氨酸蛋白酶和金属蛋白酶分别导致了观察到的AI和AIII异常药理学特征。RASM细胞膜含有一群对氯沙坦结合位点均一的结合位点，其Ki值在不存在（50 nM）或存在（16 nM）蛋白酶抑制剂时有所不同，这表明该受体可能也是这些肽酶的作用靶点。（摘要截短于250字）