Abstract

Case Report

A 2-month-old boy was referred to our university hospital to receive cardiac surgery for the treatment of pulmonary artery atresia. No consanguinity was observed in the family. He was born at gestational age 34 weeks and weighed 1640 g at the time of birth. The Apgar score was estimated to be 8 points at 5 minutes after birth. He developed severe repetitive cyanosis after birth. Angiocardiographic imaging was used to establish a diagnosis of pulmonary artery atresia (Fig. 1A). A Blalock-Taussig procedure was successfully completed at age 3 months. At age 4 months, cholestatic liver disease with jaundice was diagnosed with an elevation of serum total bilirubin (19.9 mg/dL), followed by high levels of serum direct bilirubin (16.6 mg/dL), serum total bile acid (112.3 μmol/L), serum γ glutamyl peptidase (151 IU/L), serum aspartate transaminase (110 U/L) and serum alanine transaminase (75 IU/L) (Table 1). Simultaneously, dyslipidemia was also detected with a serum total cholesterol (TC) of 408 mg/dL; low-density lipoprotein cholesterol (LDL-C), 35 mg/dL; high-density lipoprotein cholesterol (HDL-C), 12 mg/dL; and triglycerides (TGs), 564 mg/dL (see Table 1). Polyacrylamide gel electrophoresis of serum revealed that intermediate-density lipoproteins (IDLs) and very-low-density lipoproteins (VLDLs; normal range, 5%-20%) values of 45% and 25%, respectively. Abdominal magnetic resonance imaging revealed hepatosplenomegaly and a normal gallbladder (Fig. 1B and ​and1C).1C). Oral UDCA was prescribed for intrahepatic cholestasis with hyperlipidemia along with fat-soluble vitamins.

Open in a separate window

Figure 1.

A, Roentgenogram shows the hypoplasia of peripheral pulmonary artery branches associated with pulmonary atresia on the catheterized examination. B and C, Abdominal magnetic resonance imaging shows liver enlargement, mild splenomegaly, and a normal gallbladder. D and E, Yellow arrows indicate xanthomas identified around multiple joints.

At age 9 months, oral pravastatin was added at a dose of 1 mg/day, because the levels of serum TC and TGs were markedly increased. This was followed by a remarkable spread of xanthomas around the joints (Fig. 1D and ​and1E).1E). At age 11 months, oral fenofibrate treatment was added because oral pravastatin and UDCA were not successful in treating the dyslipidemia. The dyslipidemia worsened after antihyperlipidemic treatment with a combination of pravastatin and fenofibrate following UDCA. At age 12 months, serum TC and TG levels were markedly elevated to 1780 mg/dL and 635 mg/dL, respectively, even though 4 mg/d of pravastatin and 16 mg/d of fenofibrate were administered. We replaced pravastatin with 7.5 mg/d of atorvastatin because the hyperlipidemic condition was refractory to the combination therapy with pravastatin and fenofibrate. Finally, the new treatment regimen improved the dyslipidemia. At age 24 months, serum TC and TG levels were approximately within the normal range (see Table 1).

The roentgenogram showed no vertebral abnormalities, and the patient’s facial appearance was not in accordance with the characteristic phenotype of AGS.

To confirm a diagnosis of cholestatic dyslipidemia, we performed genetic testing on the patient and his parents after informed consent was obtained from the parents. Genomic DNA was extracted from peripheral blood using the QIAmp Blood Midi Kit (Qiagen). The genes responsible for hereditary intrahepatic cholestasis, including JAG1, NOTCH2, ABCCX2, SLC2513, ATP8B1, ABCB11, ABCB4, TJP2, HSDB7, AKR1D, CYP7B1, BAAT, EPHX1, SLC10A1, ABCB1, SLC4A2, and SLCO1A2, were analyzed by next-generation sequencing (NGS) with targeted sequencing [9]. Genetic testing revealed that the patient had a heterozygous mutation in the JAG1 gene: NM_000214.3:c.1326G > A, p.Trp442X. This variant was not found in either of the parents. This de novo variant of the JAG1 gene has been reported to be pathogenic in AGS [10, 11].

Discussion

We encountered an infantile case of AGS presenting with severe dyslipidemia. Our case showed extreme hypercholesterolemia and hypertriglyceridemia, associated with high levels of IDL and VLDL and low levels of LDL-C and HDL-C. Reportedly, patients with AGS have abnormal lipoprotein patterns, which differ depending on the degree of hyperbilirubinemia [12]. The lipid profile of patients with AGS and mild hyperbilirubinemia shows high levels of LDL-C and HDL-C, with increased levels of serum apoprotein A-I and apoprotein A-II. However, patients diagnosed with AGS who have severe hyperbilirubinemia show dyslipidemia with lower levels of LDL-C and HDL-C, which is associated with low levels of apoprotein A-I and apoprotein A-II [12]. LDL and HDL are cholesterol-rich lipoproteins. One of the pivotal functions of the hepatic triglyceride lipase (HTGL) is to convert IDL to LDL, and the main function of lipoprotein lipase (LPL) is to convert VLDL to IDL. IDL and VLDL are TG-rich lipoproteins. Thus, HTGL plays an important role in TG level regulation in the blood by maintaining steady levels of IDL, HDL, and LDL. In that context, our patient’s condition theoretically indicated that LPL activity and/or HTGL activity might have decreased, depending on the severity of cholestasis.

In contrast, a previous report documented that the SRB1 gene, the gene of the HDL receptor, was upregulated in the liver of patients with AGS [13]. Upregulated SRB1 messenger RNA (mRNA) in the liver is helpful in scavenging HDL-C converted from IDL and VLDL with cholesteryl ester transfer protein, which is a compensatory mechanism in patients with AGS that might be protective against atherosclerosis [14]. However, our case suggests the possibility that premature HDL can accumulate because of a decrease in HTGL activity and/or a mass due to cholestatic liver damage, thus indicating that mature HDL levels could be decreased by decreased HDL metabolism in the present patient [15].

This was the first reported case of the therapeutic use of atorvastatin for infants with AGS and cholestatic dyslipidemia. A similar case in terms of severity was previously described by Hannoush et al [16]. They reported that cholestatic dyslipidemia was successfully treated with UDCA [16]. The case reported by Hannoush and colleagues [16] might have been a case of spontaneous recovery of dyslipidemia. However, the index case was refractory to UDCA therapy, pravastatin, and fenofibrate therapy. We emphasize that atorvastatin therapy was effective and safe in terms of lowering serum TC and TG levels in refractory cases, with no adverse effects. Atorvastatin is widely used as a statin in the treatment of hypercholesterolemia and has proven beneficial in hyperlipidemic patients. Atorvastatin is well known as a statin with great effectiveness in lowering circulating LDL-C levels. Statins upregulate the expression of hepatic LDL receptors at the transcriptional level by activating sterol regulatory element-binding protein 2 (SREBP2), whereas it reduces intrinsic cholesterol production by the inhibition of HMG-CoA reductase, the rate-limiting enzyme of cholesterol synthesis. IDL (remnant VLDL) is also captured by the apoprotein B receptor and LDL receptor in the liver. Therapeutic uses of atorvastatin in infants have not been reported in the literature, although the efficacy and safety of atorvastatin treatment in adolescents and children with familial hypercholesterolemia have been reported [6]. Its safety and efficacy in infants with AGS should be verified in future clinical studies.

Interestingly, atorvastatin upregulates cholesterol 7 alpha-hydroxylase (CYP7A1) by downregulating the small heterodimer partner (SHP) gene in the rodent liver. CYP7A1 is the first rate-limiting enzyme in the bile acid synthesis pathway in the liver [17]. Reportedly, atorvastatin has a stronger effect on fecal excretion of bile acids than rosuvastatin and lovastatin [18]. In this context, atorvastatin has a unique effect of lowering circulating cholesterol by cholesterol excretion into the intestine by increasing hepatic bile acid production. Reportedly, hepatic mRNA of CYP7A1 was statistically significantly reduced in AGS patients, indicating that a reduction in the cholesterol-to-bile acid conversion contributed to the high cholesterol content in the liver of patients with AGS [13]. The major upregulated proteins in hepatic cholesterol production, including farnesoid X receptor, SHP, and liver X receptor α, might be increased in patients with AGS, indicating that impairment of the negative feedback mechanism accelerates hepatic cholesterol accumulation [13]. In this context, we expect that atorvastatin could have desirable effects against hyperlipidemia in patients with AGS by upregulating hepatic CYP7A1 mRNA through SHP downregulation at the transcriptional level, in addition to enzymatic inhibition of HMG-CoA reductase, compared with pravastatin.

Huang et al [19] reported that combination therapy with atorvastatin and fenofibrate reduced the levels of circulating TC and TGs in rats that were fed a high-fructose diet, followed by upregulation of hepatic mRNA of peroxisome proliferator-activated receptor-α (PPARα) better than sole treatment with atorvastatin. The TG-lowering effects of fenofibrate were mediated by upregulation of apoprotein A-V (ApoA-V) by PPARα in the liver, whereas fenofibrate effectively upregulates hepatic PPARα at the transcriptional level. PPARα plays a crucial role in the homeostasis of lipid and lipoprotein metabolism at the transcriptional levels [20]. Activated PPARα induces a reduction in serum TGs and an increase in HDL-C [20]. Activation of PPARα increases the production of LPL and ApoA-V in the liver, while it decreases the circulating apoprotein C-III, which inhibits LPL activity, thereby enhancing the catabolism of TG-rich lipoproteins and reducing serum TG levels [20-22]. PPARα activation also upregulates the expression of genes involved in β-oxidation pathways. Fatty acid levels in the liver are decreased through enhanced β-oxidation and increased expression of hepatic acyl-CoA synthase [23]. Thus, the hepatic production of VLDL particles is attenuated by PPARα activation [24]. In addition, atorvastatin directly upregulates ApoA-V production in the liver. ApoA-V is an important factor in the circulating TG-lowering mechanism [25]. In this context, atorvastatin has an additive TG-lowering effect based on PPARα upregulated by fenofibrate in the liver, although atorvastatin itself is not able to upregulate PPARα effectively [26]. We hypothesized that fenofibrate was necessary to treat the present patient diagnosed with AGS because atorvastatin is less effective than PPARα agonists in terms of lowering TG levels.

We detected a pathogenic variant of the JAG1 gene in a patient with AGS using NGS. Pediatricians usually face difficulties in diagnosing infantile cholestatic liver diseases. In particular, our patient showed an atypical phenotype of AGS, since he neither had vertebral abnormalities nor specific facial characteristics. Biliary atresia is a frequently encountered cholestatic hepatic disease during infancy [27]. The differential diagnosis of AGS and biliary atresia is usually difficult for pediatricians and pediatric surgeons. We suggest using genetic analysis using NGS, a noninvasive and effective tool, for a prompt diagnosis by differentiating infantile cholestatic liver diseases. This diagnostic tool is recommended before exploratory laparotomy as a less invasive method for children with cholestatic liver disease. Diagnosis based on genetic testing also helps determine the proper treatment policy and follow-up plan.

Conclusion

We report a case of infantile AGS successfully treated with atorvastatin, based on the combination of UDCA and fenofibrate. Our experience suggests that add-on atorvastatin therapy could be effective for the improvement of refractory dyslipidemia in patients diagnosed with AGS by accelerating bile acid production and excretion, inhibiting cholesterol production, and upregulating lipoprotein scavenging. We believe that a clinical trial involving treatment with atorvastatin is necessary for infant and early-childhood AGS with severe dyslipidemia. We start in earnest the discussion on strong statin treatment in hyperlipidemic AGS patients with unique lipid metabolism.

Acknowledgments

Glossary

References

1. Alagille D, Odièvre M, Gautier M, Dommergues JP. Hepatic ductular hypoplasia associated with characteristic facies, vertebral malformations, retarded physical, mental and sexual development, and cardiac murmur. J Pediatr. 1975;86(1):63-71. [Abstract] [Google Scholar]

2. Alagille D, Estrada A, Hadchouel M, Gautier M, Odièvre M, Dommergues JP. Syndromic paucity of interlobular bile ducts (Alagille syndrome or arteriohepatic dysplasia): review of 80 cases. J Pediatr. 1987;110(2):195-200. [Abstract] [Google Scholar]

3. Kronsten V, Fitzpatrick E, Baker A. Management of cholestatic pruritus in paediatric patients with Alagille syndrome: the King’s College Hospital experience. J Pediatr Gastroenterol Nutr. 2013;57(2):149-154. [Abstract] [Google Scholar]

4. Larrosa-Haro A, Sáenz-Rivera C, González-Ortiz M, Coello-Ramírez P, Vázquez-Camacho G. Lack of cholesterol-lowering effect of graded doses of cholestyramine in children with Alagille syndrome: a pilot study. J Pediatr Gastroenterol Nutr. 2003;36(1):50-33. [Abstract] [Google Scholar]

5. Gilbert MA, Bauer RC, Rajagopalan R, et al. . Alagille syndrome mutation update: comprehensive overview of JAG1 and NOTCH2 mutation frequencies and insight into missense variant classification. Hum Mutat. 2019;40(12):2197-2220. [Europe PMC free article] [Abstract] [Google Scholar]

6. Karapostolakis G, Vakaki M, Attilakos A, et al. . The effect of long-term atorvastatin therapy on carotid intima-media thickness of children with dyslipidemia. Angiology. 2021;72(4):322-3311. [Abstract] [Google Scholar]

7. Canas JA, Ross JL, Taboada MV, et al. . A randomized, double blind, placebo-controlled pilot trial of the safety and efficacy of atorvastatin in children with elevated low-density lipoprotein cholesterol (LDL-C) and type 1 diabetes. Pediatr Diabetes. 2015;16(2):79-89. [Abstract] [Google Scholar]

8. Langslet G, Breazna A, Drogari E. A 3-year study of atorvastatin in children and adolescents with heterozygous familial hypercholesterolemia. J Clin Lipidol. 2016;10(5):1153-1162.e3. [Abstract] [Google Scholar]

9. Togawa T, Sugiura T, Ito K, et al. . Molecular genetic dissection and neonatal/infantile intrahepatic cholestasis using targeted next-generation sequencing. J Pediatr. 2016;171:171-177.e1-e4. [Abstract] [Google Scholar]

10. Giannakudis J, Röpke A, Kujat A, et al. . Parental mosaicism of JAG1 mutations in families with Alagille syndrome. Eur J Hum Genet. 2001;9(3):209-216. [Abstract] [Google Scholar]

11. Ohashi K, Togawa T, Sugiura T, et al. . Combined genetic analyses achieve efficient diagnostic yields for subjects with Alagille syndrome and incomplete Alagille syndrome. Acta Paediatr. 2017;106(11):1817-1824. [Abstract] [Google Scholar]

12. Davit-Spraul A, Pourci ML, Atger V, et al. . Abnormal lipoprotein pattern in patients with Alagille syndrome depends on icterus severity. Gastroenterology. 1996;111(4):1023-1032. [Abstract] [Google Scholar]

13. Miyahara Y, Bessho K, Kondou H, et al. . Negative feedback loop of cholesterol regulation is impaired in the livers of patients with Alagille syndrome. Clin Chim Acta. 2015;440:49-54. [Abstract] [Google Scholar]

14. Jansen H, Verhoeven AJM, Sijbrands EJG. Hepatic lipase: a pro- or anti-atherogenic protein? J Lipid Res. 2002;43(9):1352-1362. [Abstract] [Google Scholar]

15. Yancey PG, Asztalos BF, Stettler N, et al. . SR-BI- and ABCA1-mediated cholesterol efflux to serum from patients with Alagille syndrome. J Lipid Res. 2004;45(9):1724-1732. [Abstract] [Google Scholar]

16. Hannoush ZC, Puerta H, Bauer MS, Goldberg RB. New JAG1 mutation causing Alagille syndrome presenting with severe hypercholesterolemia: case report with emphasis on genetics and lipid abnormalities. J Clin Endocrinol Metab. 2017;102(2):350-353. [Abstract] [Google Scholar]

17. Fu ZD, Cui JY, Klaassen CD. Atorvastatin induces bile acid-synthetic enzyme Cyp7a1 by suppressing FXR signaling in both liver and intestine in mice. J Lipid Res. 2014;55(12):2576-2586. [Europe PMC free article] [Abstract] [Google Scholar]

18. Schonewille M, de Boer JF, Mele L, et al. . Statins increase hepatic cholesterol synthesis and stimulate fecal cholesterol elimination in mice. J Lipid Res. 2016;57(8):1455-1464. [Europe PMC free article] [Abstract] [Google Scholar]

19. Huang XS, Zhao SP, Bai L, Hu M, Zhao W, Zhang Q. Atorvastatin and fenofibrate increase apolipoprotein AV and decrease triglycerides by up-regulating peroxisome proliferator-activated receptor-alpha. Br J Pharmacol. 2009;158(3):706-712. [Europe PMC free article] [Abstract] [Google Scholar]

20. Yamashita S, Matsuda D, Matsuzawa Y. Pemafibrate, a new selective PPARα modulator: concept and its clinical applications for dyslipidemia and metabolic diseases. Curr Atheroscler Rep. 2020;22(1):5. [Europe PMC free article] [Abstract] [Google Scholar]

21. Dubois V, Eeckhoute J, Lefebvre P, Staels B. Distinct but complementary contributions of PPAR isotypes to energy homeostasis. J Clin Invest. 2017;127(4):1202-1214. [Europe PMC free article] [Abstract] [Google Scholar]

22. Prieur X, Coste H, Rodriguez JC. The human apolipoprotein AV gene is regulated by peroxisome proliferator-activated receptor-alpha and contains a novel farnesoid X-activated receptor response element. J Biol Chem. 2003;278(28):25468-25480. [Abstract] [Google Scholar]

23. Cariello M, Piccinin E, Moschetta A. Transcriptional regulation of metabolic pathways via lipid-sensing nuclear receptors PPARs, FXR, and LXR in NASH. Cell Mol Gastroenterol Hepatol. 2021;11(5):1519-1539. [Europe PMC free article] [Abstract] [Google Scholar]

24. Fruchart JC. Peroxisome proliferator-activated receptor-alpha (PPARalpha): at the crossroads of obesity, diabetes and cardiovascular disease. Atherosclerosis. 2009;205(1):1-8. [Abstract] [Google Scholar]

25. Pennacchio LA, Rubin EM. Apolipoprotein A5, a newly identified gene that affects plasma triglyceride levels in humans and mice. Arterioscler Thromb Vasc Biol. 2003;23(4):529-534. [Abstract] [Google Scholar]

26. Zao W, Zao SP. Atorvastatin might inhibit insulin resistance induced by insulin through the triglyceride-lowering role of apolipoprotein AV. Eur Rev Med Pharmacol Sci. 2015;19(20):3895-3903. [Abstract] [Google Scholar]

27. Hwang SM, Jeon TY, Yoo SY, et al. . Alagille syndrome candidates for liver transplantation: differentiation from end-stage biliary atresia using preoperative CT. PLoS One. 2016;11(2):e0149681. [Europe PMC free article] [Abstract] [Google Scholar]