Neurotensin contributes to cholestatic liver disease potentially modulating matrix metalloprotease-7.

The diagnosis and treatment of biliary atresia pose challenges due to the absence of reliable biomarkers and limited understanding of its etiology. The plasma and liver of patients with biliary atresia exhibit elevated levels of neurotensin. To investigate the specific role of neurotensin in the progression of biliary atresia, the patient's liver pathological section was employed. Biliary organoids, cultured biliary cells, and a mouse model were employed to elucidate both the potential diagnostic significance of neurotensin and its underlying mechanistic pathway. In patients' blood, the levels of neurotensin were positively correlated with matrix metalloprotease-7, interleukin-8, and liver function enzymes. Neurotensin and neurotensin receptors were mainly expressed in the intrahepatic biliary cells and were stimulated by bile acids. Neurotensin suppressed the growth and increased expression of matrix metalloprotease-7 in biliary organoids. Neurotensin inhibited mitochondrial respiration, oxidative phosphorylation, and attenuated the activation of calmodulin-dependent kinase kinase 2-adenosine monophosphate-activated protein kinase (CaMKK2-AMPK) signaling in cultured biliary cells. The stimulation of neurotensin in mice and cultured cholangiocytes resulted in the upregulation of matrix metalloprotease-7 expression through binding to its receptors, namely neurotensin receptors 1/3, thereby attenuating the activation of the CaMKK2-AMPK pathway. In conclusion, these findings revealed the changes of neurotensin in patients with cholestatic liver disease and its mechanism in the progression of the disease, providing a new understanding of the complex mechanism of hepatobiliary injury in children with biliary atresia.

Intrahepatic Cholangiolitis in Cystic Fibrosis (ICCF): An Under-Appreciated Cause of Persistent Cholestasis in Infancy.

Liver histology in infants with cystic fibrosis (CF) and persistent cholestasis is seldom reported in detail. We extend previous observation of a distinctive intrahepatic cholangiopathy (ICCF) to 3 additional infants homozygous for CFTR pathological variants and a fourth infant with a heterozygous CFTR variant, summarizing our experience in 10 infants with CFTR variants and persistent cholestasis. Cholangiograms demonstrate abnormal extrahepatic ducts in 2 infants with CF, 1 with uniform dilatation interpreted as a choledochal cyst and the other with narrow patent ducts. Liver histology in 3 CF homozygotes had prominent ductular reaction with a focally destructive cholangiolitis (inflammation of small bile ducts). The CFTR heterozygote had generalized portal edema with ductular reaction and paucity but no cholangitis. Cholestasis slowly subsided in all infants. ICCF is characterized by severe ductular reaction, prominent cholangiocyte injury, and multifocal necrotizing cholangiolitis. Local aggregates of portal ceroid might suggest previous bile leakage from damaged ducts. ICCF in liver biopsies from infants with cystic fibrosis and persistent cholestasis is unrelated to the specific CFTR genotype. Liver biopsy findings and intraoperative cholangiogram help rule out biliary atresia. ICCF is an early manifestation of CF, a likely prototype for pathogenesis of cystic fibrosis liver disease later in life.

Pathologic approach to Neonatal cholestasis with a simple scoring system for biliary atresia.

A liver biopsy is essential for the diagnostic workup of persistent neonatal cholestasis (NC). The differential diagnosis of NC is broad, including obstructive and non-obstructive causes. In addition, histologic features of certain disorders may be non-specific in the early course of the disease. To evaluate liver biopsies using a practical histopathologic approach for NC and to define a simple scoring system for biliary atresia (BA) for routine clinical practice. From June 2006 to December 2021, liver biopsy specimens from infants with persistent NC were examined by two independent pathologists. The cases diagnosed as BA were correlated with clinical, radiologic, and laboratory data to calculate the final score. Four hundred and fifty-nine cases were enrolled in the study. They had a mean age of 63.94 ± 20.62 days and were followed for a median time of 58 (1-191) months. They included 162 (35.3%) cases of BA. On multivariate analysis, portal edema, ductular proliferation, cholangiolitis, and bile duct/ductular plugs were the histopathologic predictors of BA. A liver biopsy did perform well with a 95.1% sensitivity, 91.6% specificity, 86% PPV, and 97.1% NPV. At a cutoff of 5 of the scoring system, diagnosis of BA could be done with a sensitivity of 95.1% and a specificity of 100%. We have shown detailed histopathologic features of BA with more depth to infants aged ≤ 6 weeks. We have developed a simple scoring system using a combination of liver biopsy with non-invasive methods to increase the diagnostic accuracy of BA.

Differentiating biliary atresia from other causes of infantile cholestasis: An appraisal of the histomorphological changes on liver biopsy.

Background: Cholestatic disorders are a significant cause of morbidity and mortality in infants. Characterization of these disorders and differentiating biliary atresia (BA) from other causes of intrahepatic cholestasis is an age-old problem. Objectives: To study the spectrum of different infantile cholestatic disorders in our population, to differentiate BA from other causes of neonatal cholestasis (NC) on a liver biopsy, and validation of the available scoring system for the characterization of these disorders. Materials and Methods: This is an observational cross-sectional study performed over a period of 3 years between 2018 and 2021, done on neonates and infants presenting with cholestatic jaundice. The changes on liver biopsy were evaluated by different histological parameters and available scoring systems to differentiate BA from non-BA causes. Correlation with clinical, biochemical, and imaging findings was done in all cases. Results: This study included 87 cases of NC, of which BA comprised 28 cases (32%), whereas idiopathic neonatal hepatitis (INH) comprised only 12 cases (14%). Portal neutrophilic inflammation (P = 0.000053), ductal cholestasis (P < 0.001), neoductular bile plugs (P < 0.001) and bile ductular proliferation (P < 0.0001) were significantly more in BA, whereas lobular lymphocytic inflammation (P = 0.001) and giant cell transformation of hepatocytes (P = 0.0024) were more frequent in the non-BA group. Using the Lee and Looi scoring system, a histologic score ≥7 was helpful in identifying BA with 85.7% sensitivity, 92.6% specificity, and 90.6% accuracy. Conclusion: BA is the commonest cause of NC in neonates, whereas the frequency of INH is declining. Detailed histomorphologic analysis of liver biopsy, aided with IHC, is the cornerstone for the diagnosis of these disorders.

Performance of two-dimensional shear wave elastography for detecting advanced liver fibrosis and cirrhosis in patients with biliary atresia: a systematic review and meta-analysis.

BACKGROUND: Two-dimensional shear wave elastography (2D-SWE) has been proposed for detecting liver fibrosis in biliary atresia. OBJECTIVES: To assess the performance of 2D-SWE for detecting advanced liver fibrosis and cirrhosis in patients with biliary atresia. MATERIALS AND METHODS: Five electronic databases were searched to identify studies investigating the performance of 2D-SWE for diagnosing liver fibrosis in biliary atresia in children. We constructed the summary receiver operating characteristic (SROC) curves of 2D-SWE for detecting advanced liver fibrosis and cirrhosis, and then calculated the area under the SROC curves (AUROCs). RESULTS: Six studies with 470 patients (ages 55 days to 6.6 years) were included. The median correlation coefficient of 2D-SWE with pathological liver fibrosis stages was 0.779 (range: 0.443‒0.813). The summary AUROCs for advanced liver fibrosis and cirrhosis were 0.929 and 0.883, respectively. The summary sensitivity and specificity of 2D-SWE for advanced liver fibrosis were 88% (95% confidence interval [CI]: 80‒94%) and 85% (95% CI: 77‒91%) with I values of 0% and 45.6%, respectively, and for cirrhosis were 80% (95% CI: 72‒87%) and 82% (95% CI: 77‒86%) with I values of 12.9% and 0%, respectively. The diagnostic odds ratio (DOR) of 2D-SWE for advanced liver fibrosis and cirrhosis were 40.3 (95% CI: 18.2‒89.4) and 18.9 (95% CI: 11.2‒31.7), respectively. For preoperative detection of cirrhosis, the pooled AUROC, sensitivity, specificity, and DOR based on the four 2D-SWE studies were 0.877, 79% (95% CI: 71‒86%), 82% (95% CI: 77‒86%), and 17.58 (95% CI: 10.35‒29.85), respectively. CONCLUSIONS: Results show that 2D-SWE has potential as a non-invasive tool for detecting advanced liver fibrosis and cirrhosis in patients with biliary atresia.

Contribution of ADD3 and the HLA Genes to Biliary Atresia Risk in Chinese.

Nonsyndromic biliary atresia (BA) is a rare polygenic disease, with autoimmunity, virus infection and inflammation thought to play roles in its pathogenesis. We conducted a genome-wide association study in 336 nonsyndromic BA infants and 8900 controls. Our results validated the association of rs17095355 in ADD3 with BA risk (odds ratio (OR) = 1.70, 95% confidence interval (95% CI) = 1.49-1.99; p = 4.07 × 10-11). An eQTL analysis revealed that the risk allele of rs17095355 was associated with increased expression of ADD3. Single-cell RNA-sequencing data and immunofluorescence analysis revealed that ADD3 was moderately expressed in cholangiocytes and weakly expressed in hepatocytes. Immuno-fluorescent staining showed abnormal deposition of ADD3 in the cytoplasm of BA hepatocytes. No ADD3 auto-antibody was observed in the plasma of BA infants. In the HLA gene region, no variants achieved genome-wide significance. HLA-DQB1 residue Ala57 is the most significant residue in the MHC region (OR = 1.44, 95% CI = 1.20-1.74; p = 1.23 × 10-4), and HLA-DQB1 was aberrantly expressed in the bile duct cells. GWAS stratified by cytomegalovirus (CMV) IgM status in 87 CMV IgM (+) BA cases versus 141 CMV IgM (-) BA cases did not yield genome-wide significant associations. These findings support the notion that common variants of ADD3 account for BA risk. The HLA genes might have a minimal role in the genetic predisposition of BA due to the weak association signal. CMV IgM (+) BA patients might not have different genetic risk factor profiles compared to CMV IgM (-) subtype.

Frequency of infiltrating regulatory T-cells in the portal tract of biliary atresia.

PURPOSE: Immunological abnormalities have been hypothesized as a pathogenesis of biliary atresia (BA). We previously investigated the frequency and function of circulating regulatory T-cells (Tregs) and reported no differences compared to controls. However, the local Treg profile remains uncertain. We aimed to investigate the frequency of Tregs in BA liver tissues. METHODS: The number of lymphocytes, CD4+ cells, and CD4+FOXP3+ Tregs infiltrating the portal tract and the percentage of Tregs among CD4+ cells of BA and control patients were visually counted. The correlation between these data and clinical indicators was also examined. RESULTS: The number of lymphocytes, CD4+ cells, and CD4+FOXP3+ Tregs was higher in the BA group. However, the percentage of Tregs among CD4+ cells was similar in both groups. Each parameter was correlated with serum γ-GTP, but there was no clear association with liver fibrosis, jaundice clearance, and native liver survival. CONCLUSION: The number of Tregs infiltrating the portal tract was higher in BA patients. However, the infiltration of lymphocytes was also generally increased. Tregs appear to be unsuccessful in suppressing progressive inflammation in BA patients, despite recruitment to local sites. Investigation of Treg function in the local environment is warranted.

A fetal wound healing program after intrauterine bile duct injury may contribute to biliary atresia.

BACKGROUND & AIMS: Biliary atresia (BA) is an obstructive cholangiopathy that initially affects the extrahepatic bile ducts (EHBDs) of neonates. The etiology is uncertain, but evidence points to a prenatal cause. Fetal tissues have increased levels of hyaluronic acid (HA), which plays an integral role in fetal wound healing. The objective of this study was to determine whether a program of fetal wound healing is part of the response to fetal EHBD injury. METHODS: Mouse, rat, sheep, and human EHBD samples were studied at different developmental time points. Models included a fetal sheep model of prenatal hypoxia, human BA EHBD remnants and liver samples taken at the time of the Kasai procedure, EHBDs isolated from neonatal rats and mice, and spheroids and other models generated from primary neonatal mouse cholangiocytes. RESULTS: A wide layer of high molecular weight HA encircling the lumen was characteristic of the normal perinatal but not adult EHBD. This layer, which was surrounded by collagen, expanded in injured ducts in parallel with extensive peribiliary gland hyperplasia, increased mucus production and elevated serum bilirubin levels. BA EHBD remnants similarly showed increased HA centered around ductular structures compared with age-appropriate controls. High molecular weight HA typical of the fetal/neonatal ducts caused increased cholangiocyte spheroid growth, whereas low molecular weight HA induced abnormal epithelial morphology; low molecular weight HA caused matrix swelling in a bile duct-on-a-chip device. CONCLUSION: The fetal/neonatal EHBD, including in human EHBD remnants from Kasai surgeries, demonstrated an injury response with prolonged high levels of HA typical of fetal wound healing. The expanded peri-luminal HA layer may swell and lead to elevated bilirubin levels and obstruction of the EHBD. IMPACT AND IMPLICATIONS: Biliary atresia is a pediatric cholangiopathy associated with high morbidity and mortality rates; although multiple etiologies have been proposed, the fetal response to bile duct damage is largely unknown. This study explores the fetal pathogenesis after extrahepatic bile duct damage, thereby opening a completely new avenue to study therapeutic targets in the context of biliary atresia.

The association of immune-related genes and the potential role of IL10 with biliary atresia.

BACKGROUND: Biliary atresia (BA) is a severe immune-related disease that is characterized by biliary obstruction and cholestasis. The etiology of BA is unclear, our aim was to explore the relationship between biliary tract inflammation and immune-related genes. METHODS: We selected 14 SNPs in 13 immune-related genes and investigated their associations with BA by using a large case‒control cohort with a total of 503 cases and 1473 controls from southern China. RESULTS: SNP rs1518111 in interleukin10 (IL10) was identified as associated with BA (P = 5.79E-03; OR: 0.80; 95% CI: 0.68-0.94). The epistatic effects of the following pairwise interactions among these SNPs were associated with BA: signal transducer and activator of transcription 4 (STAT4) and chemokine (C-X-C motif) ligand 3 (CXCL3); STAT4 and damage-regulated autophagy modulator1 (DRAM1); CXCL3 and RAD51 paralog B (RAD51B); and interferon gamma (IFNG) and interleukin26 (IL26). Furthermore, we explored the potential role of IL-10 in the pathogenesis of the neonatal mouse model of BA. IL-10 effectively prevented biliary epithelial cell injury and biliary obstruction in murine BA as well as inhibit the activation of BA-related immune cells. CONCLUSIONS: In conclusion, this study provided strong evidence implicating IL10 as a susceptibility gene for BA in the southern Chinese population. IMPACT: This study provided strong evidence implicating IL10 as a susceptibility gene for BA in the southern Chinese population. This study could infer that IL-10 may play a protective role in BA mouse model. We found that four SNPs (rs7574865, rs352038, rs4622329, and rs4902562) have genetic interactions.

Plasticity between type 2 innate lymphoid cell subsets and amphiregulin expression regulates epithelial repair in biliary atresia.

BACKGROUND AND AIMS: Although a dysregulated type 1 immune response is integral to the pathogenesis of biliary atresia, studies in both humans and mice have uncovered a type 2 response, primarily driven by type 2 innate lymphoid cells. In nonhepatic tissues, natural type 2 innate lymphoid cell (nILC2s) regulate epithelial proliferation and tissue repair, whereas inflammatory ILC2s (iIlC2s) drive tissue inflammation and injury. The aim of this study is to determine the mechanisms used by type 2 innate lymphoid cell (ILC2) subpopulations to regulate biliary epithelial response to an injury. APPROACH AND RESULTS: Using Spearman correlation analysis, nILC2 transcripts, but not those of iILC2s, are positively associated with cholangiocyte abundance in biliary atresia patients at the time of diagnosis. nILC2s are identified in the mouse liver through flow cytometry. They undergo expansion and increase amphiregulin production after IL-33 administration. This drives epithelial proliferation dependent on the IL-13/IL-4Rα/STAT6 pathway as determined by decreased nILC2s and reduced epithelial proliferation in knockout strains. The addition of IL-2 promotes inter-lineage plasticity towards a nILC2 phenotype. In experimental biliary atresia induced by rotavirus, this pathway promotes epithelial repair and tissue regeneration. The genetic loss or molecular inhibition of any part of this circuit switches nILC2s to inflammatory type 2 innate lymphoid cell-like, resulting in decreased amphiregulin production, decreased epithelial proliferation, and the full phenotype of experimental biliary atresia. CONCLUSIONS: These findings identify a key function of the IL-13/IL-4Rα/STAT6 pathway in ILC2 plasticity and an alternate circuit driven by IL-2 to promote nILC2 stability and amphiregulin expression. This pathway induces epithelial homeostasis and repair in experimental biliary atresia.

Biliary Atresia in an Infant Presenting With Kabuki Syndrome: An Autopsy Report and Review of the Literature.

Biliary atresia (BA) is an inflammatory obliterative cholangiopathy which is very common during neonatal and infancy period. We present an autopsy report of a BA in an infant suffering from a genetic syndrome.

Necroptosis of macrophage is a key pathological feature in biliary atresia via GDCA/S1PR2/ZBP1/p-MLKL axis.

Biliary atresia (BA) is a severe inflammatory and fibrosing neonatal cholangiopathy disease characterized by progressive obstruction of extrahepatic bile ducts, resulting in cholestasis and progressive hepatic failure. Cholestasis may play an important role in the inflammatory and fibrotic pathological processes, but its specific mechanism is still unclear. Necroptosis mediated by Z-DNA-binding protein 1 (ZBP1)/phosphorylated-mixed lineage kinase domain-like pseudokinase (p-MLKL) is a prominent pathogenic factor in inflammatory and fibrotic diseases, but its function in BA remains unclear. Here, we aim to determine the effect of macrophage necroptosis in the BA pathology, and to explore the specific molecular mechanism. We found that necroptosis existed in BA livers, which was occurred in liver macrophages. Furthermore, this process was mediated by ZBP1/p-MLKL, and the upregulated expression of ZBP1 in BA livers was correlated with liver fibrosis and prognosis. Similarly, in the bile duct ligation (BDL) induced mouse cholestatic liver injury model, macrophage necroptosis mediated by ZBP1/p-MLKL was also observed. In vitro, conjugated bile acid-glycodeoxycholate (GDCA) upregulated ZBP1 expression in mouse bone marrow-derived monocyte/macrophages (BMDMs) through sphingosine 1-phosphate receptor 2 (S1PR2), and the induction of ZBP1 was a prerequisite for the enhanced necroptosis. Finally, after selectively knocking down of macrophage S1pr2 in vivo, ZBP1/p-MLKL-mediated necroptosis was decreased, and further collagen deposition was markedly attenuated in BDL mice. Furthermore, macrophage Zbp1 or Mlkl specific knockdown also alleviated BDL-induced liver injury/fibrosis. In conclusion, GDCA/S1PR2/ZBP1/p-MLKL mediated macrophage necroptosis plays vital role in the pathogenesis of BA liver fibrosis, and targeting this process may represent a potential therapeutic strategy for BA.

A Mouse Model of Chronic Liver Fibrosis for the Study of Biliary Atresia.

Biliary atresia (BA) is a fatal disease involving obstructive jaundice, and it is the most common indication for liver transplantation in children. Due to the complex etiology and unknown pathogenesis, there are still no effective drug treatments. At present, the classic BA mouse model induced by rhesus rotavirus (RRV) is the most commonly used model for studying the pathogenesis of BA. This model is characterized by growth retardation, jaundice of the skin and mucosa, clay stools, and dark yellow urine. The histopathology shows severe liver inflammation and obstruction of the intrahepatic and extrahepatic bile ducts, which are similar to the symptoms of human BA. However, the livers of end-stage mice in this model lack fibrosis and cannot fully simulate the characteristics of liver fibrosis in clinical BA. The presented study developed a novel BA mouse model of chronic liver fibrosis by injecting 5-10 µg of anti-Ly6G antibody four times, with gaps of 2 days after each injection. The results showed that some of the mice successfully formed chronic BA with typical fibrosis after the time lapse, meaning these mice represent a suitable animal model for the virus-induced liver fibrosis mechanistic study of BA and a platform for developing future BA treatments.

Biliary Atresia Remnants Revisited: Myogenesis, Hepatic Duct-Like Structures, and Fate of Peribiliary Glands.

PURPOSE AND CONTEXT: Proximal levels of excised remnants from youngest infants may reveal early features of biliary atresia (BA). METHOD: A targeted IHC survey was applied to 34 most proximal 2 levels in 17 BA remnants excised at age 10-74 days including 7 = <30 days old and 6 control hepatic ducts (HD). KEY RESULTS: Severity of inflammation and extent of active fibroplasia do not distinguish proximal remnants in younger (n = 7) and older (n = 10) infants. In 27/34 levels of 14/17 remnants, reactive stroma is focally SM-MHC-2 (+), marking smooth muscle myosin, termed reactive myogenesis (RM), that is absent in controls. RM facilitates identification of 3 novel hepatic duct remnants (HDR): an HD-like collagen collar lined by degenerating cholangiocytes (n = 5); erosion defects in loose reactive stroma (n = 14); solitary foci of hyperplastic squamoid epithelium (n = 4). Peribiliary glands are either hyperplastic or atretic and typically lack RM. CONCLUSION: Minimally inflammed end-stage lesions in BA remnants occur at youngest ages favoring prenatal onset. Three novel HDR are defined. RM, a useful surrogate for HDR, is a prevalent inappropriate stromal reaction in proximal remnants of uncertain biological significance. RM is the source of mature smooth muscle in BA remnants.

Liver-restricted deletion of the biliary atresia candidate gene Pkd1l1 causes bile duct dysmorphogenesis and ciliopathy.

BACKGROUND AND AIMS: A recent multicenter genetic exploration of the biliary atresia splenic malformation syndrome identified mutations in the ciliary gene PKD1L1 as candidate etiologic contributors. We hypothesized that deletion of Pkd1l1 in developing hepatoblasts would lead to cholangiopathy in mice. APPROACH AND RESULTS: CRISPR-based genome editing inserted loxP sites flanking exon 8 of the murine Pkd1l1 gene. Pkd1l1Fl/Fl cross-bred with alpha-fetoprotein-Cre expressing mice to generate a liver-specific intrahepatic Pkd1l1 -deficient model (LKO). From embryonic day 18 through week 30, control ( Fl/Fl ) and LKO mice were evaluated with standard serum chemistries and liver histology. At select ages, tissues were analyzed using RNA sequencing, immunofluorescence, and electron microscopy with a focus on biliary structures, peribiliary inflammation, and fibrosis. Bile duct ligation for 5 days of Fl/Fl and LKO mice was followed by standard serum and liver analytics. Histological analyses from perinatal ages revealed delayed biliary maturation and reduced primary cilia, with progressive cholangiocyte proliferation, peribiliary fibroinflammation, and arterial hypertrophy evident in 7- to 16-week-old LKO versus Fl/Fl livers. Following bile duct ligation, cholangiocyte proliferation, peribiliary fibroinflammation, and necrosis were increased in LKO compared with Fl/Fl livers. CONCLUSIONS: Bile duct ligation of the Pkd1l1 -deficient mouse model mirrors several aspects of the intrahepatic pathophysiology of biliary atresia in humans including bile duct dysmorphogenesis, peribiliary fibroinflammation, hepatic arteriopathy, and ciliopathy. This first genetically linked model of biliary atresia, the Pkd1l1 LKO mouse, may allow researchers a means to develop a deeper understanding of the pathophysiology of this serious and perplexing disorder, including the opportunity to identify rational therapeutic targets.

The accuracy of magnetic resonance cholangiopancreatography in the diagnosis of biliary atresia in preterm infants with cholestasis.

BACKGROUND: Magnetic resonance cholangiopancreatography (MRCP) is a useful and non-invasive method to diagnose biliary atresia (BA) in term infants, however few studies have investigated its use in preterm infants. This study aimed to evaluate the accuracy of MRCP in the diagnosis of BA in preterm infants with cholestasis. METHODS: Infants aged less than 6 months who received MRCP for cholestasis at a tertiary medical center were enrolled from 2011 to 2020. Demographic and laboratory data were retrospectively obtained. One pediatric radiologist reviewed the MRCP images. The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and accuracy of MRCP to diagnose BA based on surgical proof or at least 6 months of follow-up were assessed. RESULTS: A total of 80 infants (36 preterm and 44 term) were analyzed. The mean post-chronological age was 1.8 months, and the female-to-male ratio was 0.78. Six (16.7%) preterm and 16 (36.4%) term infants were confirmed to have BA. BA was obscured by a choledochal cyst preoperatively in two term infants. In the preterm infants, the sensitivity, specificity, PPV, NPV, and accuracy of MRCP to diagnose BA were 100%, 77%, 46%, 100%, and 81%, respectively, compared to 81%, 86%, 76%, 89%, and 84% in the term infants. Using MRCP to differentiate BA from other cholestasis in the preterm infants had superior sensitivity (100% vs. 81%) and NPV (100% vs. 89%), and lower specificity (77% vs. 86%) and PPV (46% vs. 76%) than in the term infants. CONCLUSIONS: Negative MRCP findings can be used to exclude BA in preterm infants with cholestasis based on a favorable NPV.

Liver Explants of Biliary Atresia Patients Transplanted in Adulthood Show Features of Obliterative Portal Venopathy: Case Series and Guidelines for Pathologic Reporting of Adult Explants.

CONTEXT.­: Biliary atresia (BA) patients can have portal vein (PV) abnormalities. OBJECTIVE.­: To investigate the explant pathology of BA patients transplanted in adulthood with a focus on portal venous abnormalities. DESIGN.­: Adult BA liver explants were reviewed, along with prior biopsies, Kasai portoenterostomy (KP), and relevant medical records. RESULTS.­: Three explants were identified; all patients were female, with age at diagnosis, KP, and liver transplantation (LT) as follows: (1) less than 1 week, 8 days, and 25 years; (2) 15 weeks, 16 weeks, and 32 years; and (3) 7 weeks, 8 weeks, and 33 years, respectively, with normalization of conjugated bilirubin within 6 months of KP and development of portal hypertension (PHTN) within 3 years of KP for all 3. The first 2 had recurrent cholangitis. Duration of pre-LT PHTN was 22, 29, and 30 years, and that of pre-LT cholangitis was 9, 3, and 0 years, respectively. All 3 explants showed hilar and extrahepatic fibromyxoid intimal hyperplasia of the PV with parenchymal hepatoportal sclerosis. Cholestasis was limited to those with a history of cholangitis. Patient 3, without cholangitis, showed delicate septal fibrosis with peripheral accentuation without biliary cirrhosis. CONCLUSIONS.­: In the context of a functioning KP, cholestasis and biliary cirrhosis are likely related to recurrent cholangitis, which may or may not occur after KP. In the absence of biliary cirrhosis, PHTN may be secondary to obliterative venopathy. Adult BA explants should be sampled thoroughly, with a focus on hilar/perihilar connective tissue to include PV branches. Explants may not show biliary cirrhosis and should be reported with appropriate clinicopathologic correlation.

Biliary atresia & choledochal malformation--Embryological and anatomical considerations.

The two main biliary pathologies in paediatric practice, biliary atresia and choledochal malformations (CM), have their origins within prenatal life. Nevertheless, the actual mechanisms remain elusive with many unanswered questions. The extrahepatic bile duct develops as a funnel-like structure emerging from the foregut from about 3-4 weeks of gestation into the mesenchyme of the septum transversum. The cranial elements of this contain hepatoblasts - the precursors to the two key cell lines that will become hepatocytes and biliary epithelial cells. The intrahepatic bile ducts develop separately and emerge from a complex process involving the ductal plate surrounding the in-growing portal venous system from about the 7-8th week of gestation. A developmental defect at some point(s) in this process may be the cause of at least some variants of BA - the Biliary Atresia Splenic Malformation syndrome particularly - though evidence in the more common isolated BA is much more circumstantial. Similarly, some types of choledochal malformation, specifically the cystic type of CM, are invariably present during prenatal life although again an actual aetiological mechanism remains elusive.

Human poliovirus receptor contributes to biliary atresia pathogenesis by exacerbating natural-killer-cell-mediated bile duct injury.

BACKGROUND AND AIMS: Natural killer (NK) cells play an important role in biliary atresia (BA) pathogenesis; human poliovirus receptor (PVR) is an important NK-cell modulator. Here, we explored the role of PVR in BA pathogenesis. METHODS: Poliovirus receptor expression and NK cell-associated genes were detected in human BA samples and a rotavirus-induced BA mouse model using quantitative PCR and immunofluorescence staining. Chemically modified small interfering RNA silenced PVR expression in the BA model, and its effects on the population and function of intrahepatic NK cells were investigated using flow cytometry (FCM). The effects of PVR overexpression and knockdown on proliferation, apoptosis and NK-cell-mediated lysis of cultured human cholangiocytes were analysed using FCM and cell viability assays. Serum PVR, high-mobility group box 1 (HMGB1), and interleukin-1beta (IL-1beta) levels were measured in a cohort of 50 patients using ELISA. RESULTS: Poliovirus receptor expression was upregulated in the biliary epithelium of BA patients and BA model and was positively correlated with the population and activation of intrahepatic NK cells. Silencing of PVR expression impaired the cytotoxicity of NK cells, reduced inflammation and protected mice from rotavirus-induced BA. Activation of the TLR3-IRF3 signalling pathway induced PVR expression in cultured cholangiocytes. PVR overexpression promoted proliferation and inhibited the apoptosis of cholangiocytes but exacerbated NK cell-mediated cholangiocyte lysis. Serum PVR levels were elevated in BA patients and were positively correlated with HMGB1 and IL-1beta levels. CONCLUSIONS: Poliovirus receptor contributes to BA pathogenesis by regulating NK cell-mediated bile duct injury; PVR has the value as a biomarker of BA.

Extrahepatic Bile Duct and Gall Bladder Dissection in Nine-Day-Old Mouse Neonates.

The dissection of murine neonatal bile ducts has been described as difficult. The main aim of the described standard operating procedure is the isolation of the extrahepatic bile duct (EBD) in mouse neonates without damaging the bile duct during preparation. Because of its exceptionally close preparation compared to the cholangiocytes cell line and harvesting of the entire extrahepatic bile duct system (EBDS), the described approach is extremely useful in researching animal models of newborn bile duct disorders, such as biliary atresia. After euthanasia, the peritoneal cavity was accessed, and the bile duct system, duodenum, and liver were extracted with the unique En-bloc-Resection (EbR). The extracted sample is placed on a foam mat, and the EBD is dissected from contaminating cells atraumatically without necessary touch. The dissection of the entire EBDS is a significant advantage of this method. Caution must be taken due to the small size and amount of bile duct tissue. Using the described technique, there is no damage to the cholangiocytes. Further, the purity of the technique is reproducible (n = 10). Therefore, optimally comparable samples can be harvested. Furthermore, no bile duct tissue is harmed, because any contact with the bile duct system can be avoided during preparation, leaving the bile fluid inside the gall bladder. Most importantly, while performing the final gall bladder and bile duct dissection, atraumatic microinstruments were used only slightly lateral of the bile duct without squeezing it. This is the key to a clean and intact sample, and essential for further histological investigation or the isolation of cholangiocytes. To summarize, the described innovative dissection technique enables especially inexperienced operators with the necessary equipment to isolate the EBDS as cleanly as possible.