MicroRevolution in understanding primary biliary cholangitis pathophysiology
Authors: Marta B. Afonso, Cecília M.P. Rodrigues
Journal: Hepatology (2017)

Potential conflict of interest: Nothing to report. See Article On Page 1420 Primary biliary cholangitis (PBC) is a chronic inflammatory autoimmune cholestatic liver disease characterized by nonsuppurative small duct inflammation and destruction, ductal proliferation, and interface hepatitis. PBC incidence is rising, although it is unclear whether this is attributed to increased awareness, improved diagnostic tools, or exposure to novel environmental factors. PBC may progress to biliary cirrhosis, portal hypertension, and liver failure and associate with variceal hemorrhage, ascites, or hepatic encephalopathy. Liver transplantation (LT) is the standard of care for patients with decompensated cirrhosis; despite being a rare disease, PBC is a leading cause of LT in middle‐aged women.1 Our current understanding of PBC pathogenesis has made major progress, despite its complex and still largely unknown underlying genetic and environmental factors.3 Specific loss of tolerance to pyruvate dehydrogenase complex‐E2 (PDC‐E2), an inner membrane mitochondrial protein, is a common characteristic of PBC and is associated with variceal hemorrhage. Indeed, a serological signature reflected by high‐titer antimitochondrial antibodies (AMAs) is a diagnostic criterion of PBC found in 90%‐95% of patients. Ursodeoxycholic acid (UDCA), a naturally occurring hydrophilic bile acid, is the first‐line pharmacotherapy for PBC patients. The mechanisms by which UDCA improves PBC rely, at least in part, in its choleretic and hepatoprotective effects. Of note, while exhibiting minimal side effects, UDCA treatment results in incomplete response in 30%‐40% of PBC patients. In 2016, obeticholic acid (OCA) was approved in Europe and the United States for the treatment of PBC when combined with UDCA, in adults with inadequate response to UDCA, or as monotherapy, in patients unable to tolerate UDCA. OCA is a semisynthetic bile acid–derivative and potent agonist of the farnesoid X receptor regulating hepatic bile acid synthesis and excretion. OCA improves relevant markers of active cholestatic liver injury, which translate into positive clinical outcomes in PBC patients. Still, long‐term outcomes of OCA in PBC remain to be elucidated. Given the complex clinical phenotype, natural history, and response to therapy among PBC patients, future studies should continue to dissect the intricate pathophysiological basis of disease to aid in the discovery of new‐generation treatments.1 MicroRNAs (miRNAs) are a class of conserved, small noncoding RNAs that induce posttranscriptional gene silencing in eukaryotes by targeting the 3'‐untranslated region (UTR) of specific mRNAs. To date, more than 2,500 miRNAs have been reported in humans (miRBase 21), which are involved in epigenetic regulation of virtually all cellular processes. As such, altered miRNA expression associates with the triggering and progression of numerous diseases. In addition, their stability in blood circulation and other body fluids, easy quantification, and differential expression in extracellular vesicles makes them attractive noninvasive biomarkers. Interestingly, recent evidence indicates that miRNA deregulation may be associated with cholangiopathies.4 Particularly, miR‐506 is increased in intrahepatic bile ducts of PBC patients and modulates fluid secretion in cholangiocytes by negatively targeting Cl−/ HCO3− anion exchanger 2 (AE2).5 AE2 mediates biliary‐bicarbonate excretion in cholangiocytes and hence reduces bile toxicity. The pivotal role of AE2 dysfunction in the pathogenesis of PBC was highlighted by the clinical observation that AE2 is down‐regulated in the liver of PBC patients, with possible implications in cholangiocyte damage, whereas UDCA restores AE2 expression and bicarbonate secretion. Furthermore, at 15 months of age, Ae2‐deficient mice show partial hepatobiliary morphological and immunological features resembling human PBC, including production of AMAs that are reactive to PDC‐E2.6 Likewise, type III inositol 1,4,5‐trisphosphate receptor (InsP3R3), an intracellular calcium release channel in endoplasmic reticulum and a major driver of bile duct secretion, is also a validated target of miR‐506.7 In the current issue of Hepatology, Erice et al.8 found that miR‐506 overexpression in human cholangiocytes associates with aberrant expression of PDC‐E2. Proteomic analysis of human cholangiocytes revealed that up‐regulation of miR‐506 impacts on proteins involved in several fundamental biological processes, particularly mitochondrial metabolism. Moreover, overexpression of miR‐506 augmented the mitochondrial respiration rate during mitochondrial stress tests. Still, adenosine triphosphate (ATP) synthesis was not perfectly coupled to oxygen consumption because of proton leak across the mitochondrial inner membrane, leading to decreased ATP production in miR‐506‐overexpressing cholangiocytes. Up‐regulation of miR‐506 in cholangiocytes further triggered a metabolic switch reflected by increased glycolysis, oxidative and endoplasmic reticulum stress, and DNA damage. In addition, bile duct cell phenotype was affected, namely cholangiocyte dedifferention depicted by decreased expression of biliary and epithelial markers; induction of mesenchymal, proinflammatory, and profibrotic markers; and diminished cholangiocyte adhesion and proliferation. These events provide a noxious milieu that sensitizes cholangiocytes to apoptosis triggered by toxic bile acids during stasis (Fig. 1).Figure 1: Schematic model depicting the deleterious effects of miR‐506 up‐regulation in cholangiocytes, resembling those observed in PBC. Abbreviations: ER, endoplasmic reticulum; OXPHOS, oxidative phosphorylation; ROS, reactive oxygen species.Overall, up‐regulation of miR‐506 in cholangiocytes induces a robust PBC‐like phenotype. A key follow‐up query is to understand whether targeting miR‐506 might constitute a novel and effective therapeutic intervention for this cholestatic liver disease. Of note, multiple miRNAs can target an individual mRNA or, at least, regulate the same cellular event and thereby act redundantly. Indeed, multiplicity and cooperativity of miRNA‐target interactions have been shown in the liver of PBC patients, representing inflammatory response and calcium homeostasis key biological processes controlled by up‐regulated miRNAs.9 As such, it may be that several miRNAs have to be simultaneously targeted in order to improve outcomes in PBC patients or, alternatively, combined with other therapeutic approaches. Furthermore, does miR‐506 modulate human direct cholestasis‐related symptoms, including fatigue and pruritus, in addition to extrahepatic characteristics of the disease? PBC patients often manifest features of other autoimmune conditions, such as Sjogren's syndrome and autoimmune Hashimoto's thyroiditis. Here, the researchers show that coculturing miR‐506‐overexpressing cholangiocytes with peripheral blood mononuclear cells from a PBC patient or a normal donor induces activation and proliferation of immunocytes, suggesting an important role of cholangiocyte miR‐506 up‐regulation in PBC‐related immune activation.8 Immune activation together with miR‐506‐associated PDC‐E2 expression in cholanciocytes, and subsequent development of autoantibodies, prompts to speculate that deregulation of miR‐506 expression in PBC might impact on concomitant autoimmune diseases. On the other hand, because miR‐506 functions as either oncogene or tumor suppressor, in a context‐dependent manner, further studies are needed to better evaluate the likelihood of off‐target effects associated with a putative pharmacological inhibition of miR‐506. Given that UDCA is usually administered to PBC patients for life, the safety of long‐term miR‐506 targeting is also worth considering. To address some of the above questions, rigorous preclinical testing in proof‐of‐concept studies will be key. Still, animal models of PBC are lacking, because the complex interaction of immunological injury and localized tissue response is difficult to recapitulate. Furthermore, given that the specific binding site of miR‐506 to the 3'‐UTR region of human AE2 is absent in the rodent homolog, inhibition of miR‐506 in mice may not completely reflect its therapeutic potential. Thus, future translation of this research will be challenging, but certainly rewarding. Another important issue that needs addressing is how to pharmacologically modulate miR‐506 levels and/or activity. Erice et al.8 showed that different proinflammatory cytokines, increased in the liver of PBC patients, induce miR‐506 promoter activity in human cholangiocytes, whereas UDCA is known to modulate miRNA expression.(10) However, neither UDCA nor other bile acids impacted on miR‐506 promoter activity experiments. As such, antisense oligonucleotides targeted against miR‐506 with high affinity and specificity should be designed and tested. Of note, uptake of miRNA‐based therapeutics is enhanced in the liver, with clinical trials ongoing for liver diseases. These new findings shed light on our understanding of miR‐506 function that relies on a complex transcription network of fine‐tuning cholangiocyte homeostasis. The therapeutic potential of targeting miR‐506 remains to be seen; however, this study opens potential avenues for a novel and attractive field of research in PBC treatment. Author names in bold designate shared co‐first authorship.