Department of Internal Medicine, University of Michigan, Ann Arbor, MI, USA.
Department of Chemistry and Biochemistry, The Ohio State University at Marion, Marion, OH, USA.
J Lipid Res. 2024 Jul;65(7):100574. doi: 10.1016/j.jlr.2024.100574. Epub 2024 Jun 9.
Bis(monoacylglycerol)phosphate (BMP) is an acidic glycerophospholipid localized to late endosomes and lysosomes. However, the metabolism of BMP is poorly understood. Because many drugs that cause phospholipidosis inhibit lysosomal phospholipase A2 (LPLA2, PLA2G15, LYPLA3) activity, we investigated whether this enzyme has a role in BMPcatabolism. The incubation of recombinant human LPLA2 (hLPLA2) and liposomes containing the naturally occurring BMP (sn-(2-oleoyl-3-hydroxy)-glycerol-1-phospho-sn-1'-(2'-oleoyl-3'-hydroxy)-glycerol (S,S-(2,2',C)-BMP) resulted in the deacylation of this BMP isomer. The deacylation rate was 70 times lower than that of dioleoyl phosphatidylglycerol (DOPG), an isomer and precursor of BMP. The release rates of oleic acid from DOPG and four BMP stereoisomers by LPLA2 differed. The rank order of the rates of hydrolysis were DOPG>S,S-(3,3',C)-BMP>R,S-(3,1',C)-BMP>R,R-(1,1',C)>S,S-(2,2')-BMP. The cationic amphiphilic drug amiodarone (AMD) inhibited the deacylation of DOPG and BMP isomers by hLPLA2 in a concentration-dependent manner. Under these experimental conditions, the ICs of amiodarone-induced inhibition of the four BMP isomers and DOPG were less than 20 μM and approximately 30 μM, respectively. BMP accumulation was observed in AMD-treated RAW 264.7 cells. The accumulated BMP was significantly reduced by exogenous treatment of cells with active recombinant hLPLA2 but not with diisopropylfluorophosphate-inactivated recombinant hLPLA2. Finally, a series of cationic amphiphilic drugs known to cause phospholipidosis were screened for inhibition of LPLA2 activity as measured by either the transacylation or fatty acid hydrolysis of BMP or phosphatidylcholine as substrates. Fifteen compounds demonstrated significant inhibition with ICs ranging from 6.8 to 63.3 μM. These results indicate that LPLA2 degrades BMP isomers with different substrate specificities under acidic conditions and may be the key enzyme associated with BMP accumulation in drug-induced phospholipidosis.
双(单酰基甘油)磷酸酯 (BMP) 是一种酸性甘油磷脂，定位于晚期内体和溶酶体。然而，BMP 的代谢过程还不太清楚。由于许多导致磷脂病的药物抑制溶酶体磷脂酶 A2（LPLA2，PLA2G15，LYPLA3）的活性，因此我们研究了这种酶是否在 BMP 分解代谢中起作用。重组人 LPLA2（hLPLA2）与含有天然 BMP（sn-(2-油酰基-3-羟基）-甘油-1-磷酸-sn-1'-(2'-油酰基-3'-羟基）-甘油（S，S-(2,2',C）-BMP）的脂质体孵育导致这种 BMP 异构体的去酰化。这种 BMP 异构体的去酰化速率比二油酰基磷脂酰甘油（DOPG）低 70 倍，DOPG 是 BMP 的异构体和前体。LPLA2 从 DOPG 和四种 BMP 立体异构体中释放油酸的速率不同。水解速率的顺序为 DOPG>S，S-(3,3'，C）-BMP>R，S-(3,1'，C）-BMP>R，R-(1,1'，C）>S，S-(2,2'）-BMP。阳离子两亲性药物胺碘酮（AMD）以浓度依赖的方式抑制 hLPLA2 对 DOPG 和 BMP 异构体的去酰化。在这些实验条件下，AMD 诱导的四种 BMP 异构体和 DOPG 抑制的 IC50 均小于 20 μM 和大约 30 μM。在 AMD 处理的 RAW 264.7 细胞中观察到 BMP 积累。用外源性活性重组 hLPLA2 处理细胞可显著减少积累的 BMP，但用二异丙基氟磷酸失活的重组 hLPLA2 处理则不能。最后，对一系列已知可引起磷脂病的阳离子两亲性药物进行了筛选，以测定 LPLA2 活性，方法是使用 BMP 或磷脂酰胆碱作为底物的转酰基或脂肪酸水解。十五种化合物表现出显著的抑制作用，IC50 范围为 6.8 至 63.3 μM。这些结果表明，LPLA2 在酸性条件下以不同的底物特异性降解 BMP 异构体，可能是与药物诱导的磷脂病中 BMP 积累相关的关键酶。
