Alonso-Peña Marta, Espinosa-Escudero Ricardo, Herraez Elisa, Briz Oscar, Cagigal Maria Luisa, Gonzalez-Santiago Jesus M, Ortega-Alonso Aida, Fernandez-Rodriguez Conrado, Bujanda Luis, Calvo Sanchez Marta, D Avola Delia, Londoño Maria-Carlota, Diago Moises, Fernandez-Checa Jose C, Garcia-Ruiz Carmen, Andrade Raul J, Lammert Frank, Prieto Jesus, Crespo Javier, Juamperez Javier, Diaz-Gonzalez Alvaro, Monte Maria J, Marin Jose J G
Experimental Hepatology and Drug Targeting, Institute for Biomedical Research, University of Salamanca, Salamanca, Spain.
Gastroenterology and Hepatology Department, Clinical and Translational Research in Digestive Diseases, Valdecilla Research Institute (IDIVAL), Marqués de Valdecilla University Hospital, Santander, Spain.
Hepatology. 2022 Nov;76(5):1259-1274. doi: 10.1002/hep.32517. Epub 2022 Jul 1.
A variant (p.Arg225Trp) of peroxisomal acyl-CoA oxidase 2 (ACOX2), involved in bile acid (BA) side-chain shortening, has been associated with unexplained persistent hypertransaminasemia and accumulation of C27-BAs, mainly 3α,7α,12α-trihydroxy-5β-cholestanoic acid (THCA). We aimed to investigate the prevalence of ACOX2 deficiency-associated hypertransaminasemia (ADAH), its response to ursodeoxycholic acid (UDCA), elucidate its pathophysiological mechanism and identify other inborn errors that could cause this alteration.
Among 33 patients with unexplained hypertransaminasemia from 11 hospitals and 13 of their relatives, seven individuals with abnormally high C27-BA levels (>50% of total BAs) were identified by high-performance liquid chromatography-mass spectrometry. The p.Arg225Trp variant was found in homozygosity (exon amplification/sequencing) in two patients and three family members. Two additional nonrelated patients were heterozygous carriers of different alleles: c.673C>T (p.Arg225Trp) and c.456_459del (p.Thr154fs). In patients with ADAH, impaired liver expression of ACOX2, but not ACOX3, was found (immunohistochemistry). Treatment with UDCA normalized aminotransferase levels. Incubation of HuH-7 hepatoma cells with THCA, which was efficiently taken up, but not through BA transporters, increased reactive oxygen species production (flow cytometry), endoplasmic reticulum stress biomarkers (GRP78, CHOP, and XBP1-S/XBP1-U ratio), and BAXα expression (reverse transcription followed by quantitative polymerase chain reaction and immunoblot), whereas cell viability was decreased (tetrazolium salt-based cell viability test). THCA-induced cell toxicity was higher than that of major C24-BAs and was not prevented by UDCA. Fourteen predicted ACOX2 variants were generated (site-directed mutagenesis) and expressed in HuH-7 cells. Functional tests to determine their ability to metabolize THCA identified six with the potential to cause ADAH.
Dysfunctional ACOX2 has been found in several patients with unexplained hypertransaminasemia. This condition can be accurately identified by a noninvasive diagnostic strategy based on plasma BA profiling and ACOX2 sequencing. Moreover, UDCA treatment can efficiently attenuate liver damage in these patients.
过氧化物酶体酰基辅酶A氧化酶2（ACOX2）的一种变体（p.Arg225Trp）参与胆汁酸（BA）侧链缩短，与不明原因的持续性高转氨酶血症以及C27 - BA的蓄积有关，主要是3α,7α,12α - 三羟基 - 5β - 胆甾烷酸（THCA）。我们旨在研究ACOX2缺乏相关高转氨酶血症（ADAH）的患病率、其对熊去氧胆酸（UDCA）的反应，阐明其病理生理机制，并识别可能导致这种改变的其他先天性代谢缺陷。
在来自11家医院的33例不明原因高转氨酶血症患者及其13名亲属中，通过高效液相色谱 - 质谱法鉴定出7例C27 - BA水平异常高（>总BA的50%）的个体。在2例患者和3名家庭成员中通过外显子扩增/测序发现纯合的p.Arg225Trp变体。另外2例无亲缘关系的患者是不同等位基因的杂合携带者：c.673C>T（p.Arg225Trp）和c.456_459del（p.Thr154fs）。在ADAH患者中，发现肝脏ACOX2的表达受损，但ACOX3未受损（免疫组织化学）。UDCA治疗使转氨酶水平恢复正常。用THCA孵育HuH - 7肝癌细胞，THCA能被有效摄取，但不是通过BA转运体，这增加了活性氧的产生（流式细胞术）、内质网应激生物标志物（GRP78、CHOP和XBP1 - S/XBP1 - U比值）以及BAXα的表达（逆转录随后进行定量聚合酶链反应和免疫印迹），而细胞活力降低（基于四唑盐的细胞活力试验）。THCA诱导的细胞毒性高于主要的C24 - BA，且不能被UDCA预防。通过定点诱变产生了14种预测的ACOX2变体，并在HuH - 7细胞中表达。用于确定它们代谢THCA能力的功能测试鉴定出6种具有导致ADAH的潜力。
在数例不明原因高转氨酶血症患者中发现了功能失调的ACOX2。这种情况可以通过基于血浆BA谱分析和ACOX2测序的非侵入性诊断策略准确识别。此外，UDCA治疗可有效减轻这些患者的肝损伤。
