Biochemistry Department, Yorkhill Hospital, Yorkhill, Glasgow, United Kingdom.
Clin Chim Acta. 2012 Aug 16;413(15-16):1207-10. doi: 10.1016/j.cca.2012.03.019. Epub 2012 Mar 29.
Cholesterol ester storage disease (CESD) and Wolman Disease (WD) are due to deficiency of lysosomal acid lipase (LAL). A new method is described for the measurement of LAL in dried blood spots (DBS) using Lalistat 2 an inhibitor of LAL.
LAL activity in DBS extracts was measured using the substrate 4-methylumbelliferyl palmitate. LAL activity was determined by measuring total lipase activity and lipase activity in the presence of Lalistat 2. The specificity of Lalistat 2 was investigated using human recombinant LAL (hrLAL) and human pancreatic lipase (hPL).
Lalistat 2 inhibited hrLAL with 1% residual activity at 1 μM inhibitor but had no effect on hPL up to 10 μM. LAL activity in DBS samples obtained from normal controls (n=140) was 0.50-2.30 nmol/punch/h and in patients with CESD was <0.03 nmol/punch/h (n=11). Activity in carriers showed intermediate activity: 0.15-0.40 nmol/punch/h (n=15).
Measurement of LAL using DBS is made difficult by the presence of other lipases in whole blood. Lalistat 2 is a specific inhibitor of LAL which allows the determination of LAL in DBS. Results show the method differentiates clearly between normal controls, carriers and affected cases.
胆固醇酯贮积病（CESD）和 Wolman 病（WD）是由于溶酶体酸性脂肪酶（LAL）缺乏所致。本文描述了一种使用 Lalistat 2（LAL 的抑制剂）测定干血斑（DBS）中 LAL 的新方法。
使用 4-甲基伞形酮棕榈酸酯作为底物，测定 DBS 提取物中的 LAL 活性。通过测量总脂酶活性和 Lalistat 2 存在下的脂酶活性来确定 LAL 活性。使用人重组 LAL（hrLAL）和人胰腺脂肪酶（hPL）研究 Lalistat 2 的特异性。
Lalistat 2 以 1μM 抑制剂抑制 hrLAL，残留活性为 1%，但对 hPL 无影响，最高可达 10μM。从正常对照者（n=140）获得的 DBS 样本中的 LAL 活性为 0.50-2.30 nmol/打孔/h，CESD 患者的活性<0.03 nmol/打孔/h（n=11）。携带者的活性显示出中间活性：0.15-0.40 nmol/打孔/h（n=15）。
全血中存在其他脂酶使 DBS 中 LAL 的测定变得困难。Lalistat 2 是 LAL 的特异性抑制剂，可用于测定 DBS 中的 LAL。结果表明，该方法可清楚地区分正常对照者、携带者和受影响者。
