Department of Pharmacology and Toxicology, Michigan State University, East Lansing, MI 48824, USA.
Int J Mol Sci. 2021 Mar 5;22(5):2649. doi: 10.3390/ijms22052649.
Transglutaminases (TGs) are crosslinking enzymes best known for their vascular remodeling in hypertension. They require calcium to form an isopeptide bond, connecting a glutamine to a protein bound lysine residue or a free amine donor such as norepinephrine (NE) or serotonin (5-HT). We discovered that perivascular adipose tissue (PVAT) contains significant amounts of these amines, making PVAT an ideal model to test interactions of amines and TGs. We hypothesized that transglutaminases are active in PVAT. Real time RT-PCR determined that Sprague Dawley rat aortic, superior mesenteric artery (SMA), and mesenteric resistance vessel (MR) PVATs express TG2 and blood coagulation Factor-XIII (FXIII) mRNA. Consistent with this, immunohistochemical analyses support that these PVATs all express TG2 and FXIII protein. The activity of TG2 and FXIII was investigated in tissue sections using substrate peptides that label active TGs when in a catalyzing calcium solution. Both TG2 and FXIII were active in rat aortic PVAT, SMAPVAT, and MRPVAT. Western blot analysis determined that the known TG inhibitor cystamine reduced incorporation of experimentally added amine donor 5-(biotinamido)pentylamine (BAP) into MRPVAT. Finally, experimentally added NE competitively inhibited incorporation of BAP into MRPVAT adipocytes. Further studies to determine the identity of amidated proteins will give insight into how these enzymes contribute to functions of PVAT and, ultimately, blood pressure.
转谷氨酰胺酶（TGs）是一种交联酶，其在高血压中的血管重塑作用最为人所知。它们需要钙才能形成异肽键，将谷氨酰胺连接到结合赖氨酸残基的蛋白质上，或连接游离胺供体，如去甲肾上腺素（NE）或 5-羟色胺（5-HT）。我们发现血管周围脂肪组织（PVAT）中含有大量这些胺，这使得 PVAT 成为测试胺和 TGs 相互作用的理想模型。我们假设 TGs 在 PVAT 中是活跃的。实时 RT-PCR 确定，Sprague Dawley 大鼠主动脉、肠系膜上动脉（SMA）和肠系膜阻力血管（MR）PVAT 表达 TG2 和凝血因子 XIII（FXIII）mRNA。与此一致，免疫组织化学分析支持这些 PVAT 均表达 TG2 和 FXIII 蛋白。使用标记催化钙溶液中活性 TGs 的底物肽在组织切片中研究了 TG2 和 FXIII 的活性。TG2 和 FXIII 在大鼠主动脉 PVAT、SMAPVAT 和 MRPVAT 中均具有活性。Western blot 分析确定，已知的 TG 抑制剂半胱胺减少了实验添加的胺供体 5-（生物素氨基）戊基胺（BAP）掺入 MRPVAT。最后，实验添加的 NE 竞争性抑制 BAP 掺入 MRPVAT 脂肪细胞。进一步的研究确定酰胺化蛋白的身份将深入了解这些酶如何促进 PVAT 的功能，最终促进血压。
