Unit of Internal Medicine, Department of Medicine, University of Verona, Verona, Italy.
Section of Clinical Biochemistry, University and Azienda Ospedaliera Universitaria Integrata of Verona, Verona, Italy.
Front Endocrinol (Lausanne). 2021 Aug 23;12:681974. doi: 10.3389/fendo.2021.681974. eCollection 2021.
Apparent mineralocorticoid excess (AME) is an autosomal recessive disorder caused by the 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2) enzyme deficiency, traditionally assessed by measuring either the urinary cortisol metabolites ratio (tetrahydrocortisol+allotetrahydrocortisol/tetrahydrocortisone, THF+5αTHF/THE) or the urinary cortisol/cortisone (F/E) ratio. Exosomal mRNA is an emerging diagnostic tool due to its stability in body fluids and its biological regulatory function. It is unknown whether urinary exosomal HSD11B2 mRNA is related to steroid ratio or the HSD11B2 662 C>G genotype (corresponding to a 221 A>G substitution) in patients with AME and essential hypertension (EH).
To detect and quantify HSD11B2 mRNA from urinary exosomes in samples from family members affected by AME and EH, and to evaluate the relationship between exosomal HSD11B2 mRNA, steroid ratio, 662C>G genotype, and hypertension.
In this observational case-control study, urinary steroid ratios and biochemical parameters were measured. Urinary exosomes were extracted from urine and exosomal HSD11B2 mRNA was quantified by Droplet Digital PCR (ddPCR). B2M (β-2 microglobulin) gene was selected as the reference housekeeping gene.
Among family members affected by AME, exosomal urinary HSD11B2 mRNA expression was strictly related to genotypes. The two homozygous mutant probands showed the highest HSD11B2 mRNA levels (median 169, range 118-220 copies/µl) that progressively decreased in 221 AG heterozygous with hypertension (108, range 92-124 copies/µl), 221 AG heterozygous normotensives (23.35, range 8-38.7 copies/µl), and wild-type 221 AA subjects (5.5, range 4.5-14 copies/µl). Heterozygous hypertensive subjects had more HSD11B2 mRNA than heterozygous normotensive subjects. The F/E urinary ratio correlated with HSD11B2 mRNA copy number ( < 0.05); HSD11B2 mRNA strongly decreased while THF+5αTHF/THE increased in the two probands after therapy. In the AME family, HSD11B2 copy number correlated with both F/E and THF+5αTHF/THE ratios, whereas in EH patients, a high F/E ratio reflected a reduced HSD11B2 mRNA expression.
HSD11B2 mRNA is detectable and quantifiable in urinary exosomes; its expression varies according to the 662 C>G genotype with the highest levels in homozygous mutant subjects. The HSD11B2 mRNA overexpression in AME could be due to a compensatory mechanism of the enzyme impairment. Exosomal mRNA is a useful tool to investigate HSD11B2 dysregulation in hypertension.
表观盐皮质激素过多症（AME）是一种常染色体隐性遗传病，由 11β-羟类固醇脱氢酶 2（11β-HSD2）酶缺乏引起，传统上通过测量尿皮质醇代谢物比值（四氢皮质醇+全部四氢皮质醇/四氢皮质酮，THF+5αTHF/THE）或尿皮质醇/皮质酮（F/E）比值来评估。外泌体 mRNA 是一种新兴的诊断工具，因为它在体液中稳定且具有生物学调节功能。目前尚不清楚 AME 和原发性高血压（EH）患者的尿外泌体 HSD11B2mRNA 是否与类固醇比值或 HSD11B2662C>G 基因型（对应于 221A>G 取代）相关。
检测和定量 AME 和 EH 患者家族成员尿液中外泌体 HSD11B2mRNA，并评估外泌体 HSD11B2mRNA、类固醇比值、662C>G 基因型与高血压之间的关系。
在这项观察性病例对照研究中，测量了尿类固醇比值和生化参数。从尿液中提取尿外泌体，并通过液滴数字 PCR（ddPCR）定量 HSD11B2mRNA。B2M（β-2 微球蛋白）基因被选为内参管家基因。
在 AME 患者的家族成员中，外泌体尿 HSD11B2mRNA 表达与基因型严格相关。两位纯合突变先证者表现出最高的 HSD11B2mRNA 水平（中位数 169，范围 118-220 拷贝/µl），逐渐降低至 221AG 杂合高血压患者（108，范围 92-124 拷贝/µl）、221AG 杂合血压正常者（23.35，范围 8-38.7 拷贝/µl）和野生型 221AA 受试者（5.5，范围 4.5-14 拷贝/µl）。杂合性高血压患者的 HSD11B2mRNA 多于杂合性血压正常者。F/E 尿比值与 HSD11B2mRNA 拷贝数相关（<0.05）；在两位先证者接受治疗后，HSD11B2mRNA 强烈减少，而 THF+5αTHF/THE 增加。在 AME 家族中，HSD11B2 拷贝数与 F/E 和 THF+5αTHF/THE 比值均相关，而在 EH 患者中，F/E 比值高反映 HSD11B2mRNA 表达降低。
HSD11B2mRNA 可在外泌体尿液中检测和定量；其表达根据 662C>G 基因型而变化，纯合突变患者的表达最高。AME 中 HSD11B2mRNA 的过表达可能是由于酶缺陷的代偿机制。外泌体 mRNA 是研究高血压中 HSD11B2 失调的有用工具。
