Role of transduodenal endoscopic ultrasound-guided fine-needle aspiration/biopsy (EUS-FNA/FNB) for diagnosis of retroperitoneal fibrosis (Ormond's disease).

BACKGROUND: Retroperitoneal fibrosis (RPF), often referred to as Ormond's disease when it is of idiopathic origin, is a rare disease characterized by the presence of inflammatory infiltrates and periaortic masses in the retroperitoneum. For a definite diagnosis, a biopsy and subsequent pathological examination is required. Currently accepted methods for retroperitoneal biopsy include open, laparoscopic, or CT-guided approaches. However, transduodenal endoscopic ultrasound-guided fine-needle aspiration/biopsy (EUS-FNA/FNB) for diagnosis of RPF has attracted only little attention in the literature. CASE REPORTS: We report two male patient cases who presented with leukocytosis, elevated C-reactive protein, and a suspicious retroperitoneal mass of unknown origin on computed tomography. One patient also reported left lower quadrant pain, whereas the other patient suffered from back pain and weight loss. In both patients, idiopathic RPF was successfully diagnosed by using transduodenal EUS-FNA/FNB with 22- and 20-gauge aspiration needles. Histopathology revealed dense lymphocytic infiltrates and fibrosis. The procedures lasted approximately 25 and 20 minutes, respectively, and in both patients no serious adverse events occurred. Treatment included steroid therapy and administration of Azathioprine. CONCLUSION: We demonstrate that using EUS-FNA/FNB to diagnose RPF is a feasible, fast, and safe method, which should always be considered as a first-line diagnostic modality. Hence, this case report emphasizes that gastrointestinal endoscopists are likely to play an important role in the setting of suspected RPF.

A Synthetic Glycopeptide Vaccine for the Induction of a Monoclonal Antibody that Differentiates between Normal and Tumor Mammary Cells and Enables the Diagnosis of Human Pancreatic Cancer.

In studies within the realm of cancer immunotherapy, the synthesis of exactly specified tumor-associated glycopeptide antigens is shown to be a key strategy for obtaining a highly selective biological reagent, that is, a monoclonal antibody that completely differentiates between tumor and normal epithelial cells and specifically marks the tumor cells in pancreas tumors. Mucin MUC1, which is overexpressed in many prevalent cancers, was identified as a promising target for this strategy. Tumor-associated MUC1 differs significantly from that expressed by normal cells, in particular by altered glycosylation. Structurally defined tumor-associated MUC1 cannot be isolated from tumor cells. We synthesized MUC1-glycopeptide vaccines and analyzed their structure-activity relationships in immunizations; a monoclonal antibody that specifically distinguishes between human normal and tumor epithelial cells was thus generated.

Multicenter randomized controlled trial comparing the performance of 22 gauge versus 25 gauge EUS-FNA needles in solid masses.

BACKGROUND AND AIMS: Few randomized studies have assessed the clinical performance of 25-gauge (25G) needles compared with 22-gauge (22G) needles during endoscopic ultrasound-guided fine needle aspiration (EUS-FNA) biopsy of intra-abdominal lesions. We aimed to compare the diagnostic yield, as well as performance characteristics of 22G versus 25G EUS biopsy needles by determining their diagnostic capabilities, the number of needle passes as well as cellularity of aspirated tissue specimen. METHODS: The study is a prospective, randomized, multicenter study. Patients were referred between January 2009 and January 2010 for diagnostic EUS including EUS-guided FNA of different lesions adjacent to the upper GI tract. All patients were randomized to EUS-FNA performed with either a 22G or 25G aspiration needle. RESULTS: EUS-FNA was performed in 135 patients (62 patients with a 22G needle). Sensitivity and specificity of the 22G needle was 94.1% and 95.8%, respectively, and for the 25G needle 94.1% and 100%, respectively. Investigators reported better visualization and performance for the 22G needle compared to the 25G (p < 0.0001). The number of tissue slides obtained was higher for the 22G needle during the second and third needle passes (p < 0.05). We did not observe significant differences between the number and preservation status of obtained cells (p > 0.05). CONCLUSIONS: A significant difference was found between the two types of needles in terms of reduced visualization of the 25G needle and suboptimal performance rating. However, this did not impact on overall results since both needles were equally successful in terms of a high diagnostic yield and overall accuracy.

Loss of 13q is associated with genes involved in cell cycle and proliferation in dedifferentiated hepatocellular carcinoma.

Dedifferentiation of hepatocellular carcinoma implies aggressive clinical behavior and is associated with an increasing number of genomic alterations, eg deletion of 13q. Genes directly or indirectly deregulated due to these genomic alterations are mainly unknown. Therefore this study compares array comparative genomic hybridization and whole genome gene expression data of 23 well, moderately, or poorly dedifferentiated hepatocellular carcinoma, using unsupervised hierarchical clustering. Dedifferentiated carcinoma clearly branched off from well and moderately differentiated carcinoma (P<0.001 chi(2)-test). Within the dedifferentiated group, 827 genes were upregulated and 33 genes were downregulated. Significance analysis of microarrays for hepatocellular carcinoma with and without deletion of 13q did not display deregulation of any gene located in the deleted region. However, 531 significantly upregulated genes were identified in these cases. A total of 6 genes (BIC, CPNE1, RBPMS, RFC4, RPSA, TOP2A) were among the 20 most significantly upregulated genes both in dedifferentiated carcinoma and in carcinoma with loss of 13q. These genes are involved in cell-cycle control and proliferation. Of 33 downregulated genes in the dedifferentiated subgroup, 4 metallothioneins had the lowest fold change, most probably mediated through inactivation of C/EBPalpha by the PI3K/AKT cascade. In conclusion dedifferentiation of hepatocellular carcinoma is associated with upregulation of genes involved in cell-cycle control and proliferation. Notably, a significant portion of these genes is also upregulated in carcinoma with deletion of 13q. As no downregulated genes were identified and microRNAs (mir-621, mir-16-1, mir-15a) are located within the deleted region of 13q and may be lost, we speculate that these miRNAs may induce the upregulation of critical cell-cycle control genes.

Losses of chromosome arms 4q, 8p, 13q and gain of 8q are correlated with increasing chromosomal instability in hepatocellular carcinoma.

OBJECTIVE: Chromosomal instability is a key feature in hepatocellular carcinoma (HCC). Array comparative genomic hybridization (aCGH) revealed recurring structural aberrations, whereas fluorescence in situ hybridization (FISH) indicated an increasing number of numerical aberrations in dedifferentiating HCC. Therefore, we examined whether there was a correlation between structural and numerical aberrations of chromosomal instability in HCC. METHODS AND RESULTS: 27 HCC (5 well, 10 moderately, 12 lower differentiated) already cytogenetically characterized by aCGH were analyzed. FISH analysis using probes for chromosomes 1, 3, 7, 8 and 17 revealed 1.46-4.24 signals/nucleus, which correlated with the histological grade (well vs. moderately,p < 0.0003; moderately vs. lower, p < 0.004). The number of chromosomes to each other was stable with exceptions only seen for chromosome 8. Loss of 4q and 13q, respectively, were correlated with the number of aberrations detected by aCGH (p < 0.001, p < 0.005; Mann-Whitney test). Loss of 4q and gain of 8q were correlated with an increasing number of numerical aberrations detected by FISH (p < 0.020, p < 0.031). Loss of 8p was correlated with the number of structural imbalances seen in aCGH (p < 0.048), but not with the number of numerical changes seen in FISH. CONCLUSION: We found that losses of 4q, 8p and 13q were closely correlated with an increasing number of aberrations detected by aCGH, whereas a loss of 4q and a gain of 8q were also observed in the context of polyploidization, the cytogenetic correlate of morphological dedifferentiation.

Distinct methylation patterns of benign and malignant liver tumors revealed by quantitative methylation profiling.

PURPOSE: A comparative quantitative methylation profiling of hepatocellular carcinoma and the most frequent benign liver tumor, hepatocellular adenoma, was set up for the identification of tumor-specific methylation patterns. EXPERIMENTAL DESIGN: The quantitative methylation levels of nine genes (RASSF1A, cyclinD2, p16INK4a, DAP-K, APC, RIZ-1, HIN-1, GSTpi1, SOCS-1) were analyzed in hepatocellular carcinoma and adjacent normal tissue (n = 41), hepatocellular adenoma and adjacent normal tissue (n = 26), focal nodular hyperplasia (n = 10), and unrelated normal liver tissue (n = 28). Accumulated methylation data were analyzed using various statistical algorithms, including hierarchical clustering, to detect tumor-specific methylation patterns. RESULTS: Cluster analysis revealed that hepatocellular adenoma displays a methylation profile much more similar to that found in normal liver tissue and focal nodular hyperplasia than to that found in hepatocellular carcinoma. Many characteristic differences were not detected when using mere qualitative methylation assays. The cyclinD2 gene was identified as a new and frequent target for aberrant hypermethylation in hepatocellular carcinoma (68%). In the control group of 28 liver specimens from healthy donors, a clear correlation between age of patient and frequency and level of aberrant methylation was seen, which could not be detected in the group of hepatocellular carcinoma specimens. CONCLUSIONS: Methylation profiling can clearly contribute to the unequivocal classification of suspicious lesions, but only if done in a quantitative manner applying cell type and gene-specific thresholds. In hepatocellular carcinoma, the altered methylation patterns accompanying malignant transformation override the age-dependent increase in gene methylation.

Molecular profiling of human hepatocellular carcinoma defines mutually exclusive interferon regulation and insulin-like growth factor II overexpression.

Molecular subtyping of human hepatocellular carcinoma (HCC) with potential mechanistic and therapeutic impact has not been achieved thus far. We have analyzed the mRNA expression patterns of 43 different human HCC samples and 3 HCC cell lines in comparison with normal adult liver using high-density cDNA microarrays. Two main groups of HCC, designated group A (65%) and group B (35%), were distinguished based on clustering of the most highly varying genes. Group A HCCs were characterized by induction of a number of interferon (IFN)-regulated genes, whereas group B was characterized mainly by down-regulation of several apoptosis-relevant and IFN-regulated genes. The number of apoptotic tumor cells and tumor-infiltrating lymphocytes was significantly higher in tumors of group A as compared with those of group B. Based on the expression pattern, group B was further subdivided into two subgroups, designated subgroup B1 (6 of 43 tumors, 14%) and subgroup B2 (9 of 43 tumors, 21%). A prominent characteristic of subgroup B1 was high overexpression of insulin-like growth factor (IGF)-II. All tested HCC cell lines expressed equally high concentrations of IGF-II transcripts and co-segregated with group B1 in clustering. IGF-II overexpression and induction of IFN-related genes were mutually exclusive, even when analysis was extended to other cancer expression profile studies. Moreover, IFN-gamma treatment substantially reduced IGF-II expression in HCC cells. In conclusion, cDNA microarray analyses provided subtyping of HCCs that is related to intratumor inflammation and tumor cell apoptosis. This profiling may be of mechanistic and therapeutic impact because IGF-II overexpression has been linked to reduced apoptosis and increased proliferation and may be accessible to therapeutic intervention.

Frequent epigenetic inactivation of the RASSF1A gene in hepatocellular carcinoma.

Aberrant promoter methylation is a fundamental mechanism of inactivation of tumor suppressor genes in cancer. The Ras association domain family 1A gene (RASSF1A) is frequently epigenetically silenced in several types of human solid tumors. In this study, we have investigated the expression and methylation status of the RASSF1A gene in hepatocellular carcinoma (HCC). In two HCC cell lines (HepG2 and Hep3B) RASSF1A was inactivated and treatment of these cell lines with a DNA methylation inhibitor reactivated the transcription of RASSF1A. The methylation status of the RASSF1A promoter region was analysed in 26 primary liver tissues including HCC, hepatocellular adenoma (HCA), liver fibrosis, hepatocirrhosis. Out of 15, 14 (93%) HCC were methylated at the RASSF1A CpG island and hypermethylation was independent of hepatitis virus infection. RASSF1A was also methylated in two out of two fibrosis and in three (75%) out of four cirrhosis; the latter carries an increased risk of developing HCC. Additionally, we analysed the methylation status of p16(INK4a) and other cancer-related genes in the same liver tumors. Aberrant methylation in the HCC samples was detected in 71% of samples for p16, 25% for TIMP3, 17% for PTEN, 13% for CDH1, and 7% for RARbeta2. In conclusion, our results demonstrate that RASSF1A and p16(INK4a) inactivation by methylation are frequent events in hepatocellular carcinoma, but not in HCA, which is in contrast to HCC without cirrhosis, viral hepatitis, storage diseases, or genetic background. Therefore, this study gives additional evidence against a progression of adenoma to carcinoma in the liver. Thus, RASSF1A hypermethylation could be useful as a marker of malignancy and to distinguish between the distinct forms of highly differentiated liver neoplasm.

Hepatocyte telomere shortening and senescence are general markers of human liver cirrhosis.

Telomere shortening limits the number of cell divisions of primary human cells and might affect the regenerative capacity of organ systems during aging and chronic disease. To test whether the telomere hypothesis applies to human cirrhosis, the telomere length was monitored in cirrhosis induced by a broad variety of different etiologies. Telomeres were significantly shorter in cirrhosis compared with noncirrhotic samples independent of the primary etiology and independent of the age of the patients. Quantitative fluorescence in situ hybridization showed that telomere shortening was restricted to hepatocytes whereas lymphocytes and stellate cells in areas of fibrosis had significantly longer telomere reserves. Hepatocyte-specific telomere shortening correlated with senescence-associated beta-galactosidase staining in 84% of the cirrhosis samples, specifically in hepatocytes, but not in stellate cells or lymphocytes. Hepatocyte telomere shortening and senescence correlated with progression of fibrosis in cirrhosis samples. This study demonstrates for the first time that cell type-specific telomere shortening and senescence are linked to progression of human cirrhosis. These findings give a novel explanation for the pathophysiology of cirrhosis, indicating that fibrotic scarring at the cirrhosis stage is a consequence of hepatocyte telomere shortening and senescence. The data imply that future therapies aiming to restore regenerative capacity during aging and chronic diseases will have to ensure efficient targeting of specific cell types within the affected organs.

Donor origin of de novo hepatocellular carcinoma in hepatic allografts.

BACKGROUND: Hepatocellular carcinomas (HCC) that originate de novo in liver transplants without preceding HCC in the explanted organ have only rarely been reported. Because recent data demonstrated a mixed hepatocellular chimerism caused by the integration of host-derived stem cells, a study was conducted on the origin of tumor cells in de novo HCC. METHODS: From two cases of de novo HCC arising in liver transplants after hepatitis B reinfection, tumor cells and nonneoplastic liver cells from the patient's own liver and donor liver were isolated by laser microdissection and highly polymorphic short tandem DNA repeats (STR) were investigated. RESULTS: Isolated tumor cells revealed donor-specific STR genotypes that could clearly be discriminated from the genotype of the host. CONCLUSIONS: Hepatitis B virus-associated de novo HCC in liver transplants is of donor but not host origin. The new technique described here can also discriminate between true recurrence of the original tumor and new recipient tumors.

Donor origin of de novo hepatocellular carcinoma in hepatic allografts.

BACKGROUND: Hepatocellular carcinomas (HCC) that originate de novo in liver transplants without preceding HCC in the explanted organ have only rarely been reported. Because recent data demonstrated a mixed hepatocellular chimerism caused by the integration of host-derived stem cells, a study was conducted on the origin of tumor cells in de novo HCC. METHODS: From two cases of de novo HCC arising in liver transplants after hepatitis B reinfection, tumor cells and non-neoplastic liver cells from the patient's own liver and donor liver were isolated by laser microdissection, and highly polymorphic short tandem DNA repeats (STR) were investigated. RESULTS: Isolated tumor cells revealed donor-specific STR genotypes that could clearly be discriminated from the genotype of the host. CONCLUSIONS: Hepatitis B virus-associated de novo HCC in liver transplants is of donor but not of host origin. The new technique described here can also discriminate between true recurrence of the original tumor and new recipient tumors.

Detection of chromosomal imbalances in hepatocellular carcinoma.

Hepatocellular carcinoma is the most common malignant tumor of the liver. The discrimination between well-differentiated hepatocellular carcinoma and reactive lesions and benign tumors may be difficult, especially when performed on the basis of needle biopsies. A promising means of solving this problem is provided by chromosomal analysis of imbalances in hepatocellular carcinoma. This article describes the different approaches to ascertain differential diagnosis by chromosomal studies in a reliable and cost-effective manner. It is shown that in situ hybridization techniques provide a reliable means of defining chromosome alterations. These techniques allow the detection of genetic gains and losses of defined chromosomes in a histopathological context and can serve as a helpful tool in establishing diagnosis of liver cell proliferation.

Diagnostic potential of circulating TIMP-1 and MMP-2 as markers of liver fibrosis in patients with chronic hepatitis C.

BACKGROUND: Circulating levels of tissue inhibitor of metalloproteinase (TIMP)-1 and matrix metalloproteinase (MMP)-2 are investigated as parameters for the diagnosis of fibrosis in chronic liver disease. We evaluated their diagnostic potential in comparison to hepatic histology, serum hyaluronate and standard liver function tests. METHODS: Commercially available ELISA assays were used to study circulating values of TIMP-1 and MMP-2 (Bindazyme, Biotrak, Quantikine) in patients with chronic hepatitis C (CAH; n=59), hepatitis C virus-induced cirrhosis (n=19) and 30 healthy controls. Hepatic histology was evaluated using the Hepatitis-Activity-Index according to Ishak et al. [J. Hepatol., 22 (1995) 696-699], quantifying separately inflammatory activity and fibrosis. RESULTS: Normal ranges for TIMP-1 and MMP-2 values differed for the different assays. Nevertheless, the various assays showed similar diagnostic ability and linear correlation. MMP-2 values were similar in controls and in CAH patients with and without fibrosis, but increased significantly in cirrhosis. TIMP-1 values showed a steady increase from normal to CAH without fibrosis, hepatitis with fibrosis, and cirrhosis. The diagnostic potential of serum MMP-2 to detect fibrosis was low with a sensitivity of 7% in the two assays used and an overall diagnostic efficiency of 56% and 58%. The potential of circulating MMP-2 to detect cirrhosis was higher with sensitivities of 74% and 83% and specificities of 96% and 100%, resulting in a diagnostic efficiency of 92% in the different assays. Plasma TIMP-1 values detect fibrosis with a sensitivity of 52% and 67% and a specificity of 68% and 88% resulting in overall efficiency rates of 68% and 71%, respectively. TIMP-1 values detect cirrhosis with 100% sensitivity but only 56% and 75% specificity. The diagnostic potential of circulating TIMP-1 was similar to that of hyaluronate and better than that of enzymes or albumin values. CONCLUSION: Plasma values of TIMP-1 and MMP-2 are able to detect cirrhosis with high sensitivity. TIMP-1 values also detect fibrosis with comparable efficiency. Regular determinations of both TIMP-1 and MMP-2 in CAH patients may be used as indicators of increasing fibrosis and the development of cirrhosis.

Liver resection using heat coagulative necrosis: indications and limits of a new method.

BACKGROUND: A new approach towards achieving bloodless liver resection is the use of heat coagulative necrosis. The latest stage of this technique is a four-probe device (Habib Sealer), which we used for a variety of resections to find the best indications for the method. METHODS: Between 2005 and 2006 we performed 28 liver resections in 20 consecutive patients. The most common indication was metastatic colorectal cancer (75%). We treated a heterogeneous patient collective in terms of tumour localization and extent of resection. Resection was performed after creating a necrotic zone. The device achieved an area of coagulation of 1-cm width in which even larger vessels and bile ducts were safely sealed. RESULTS: Operative spectrum covered atypical resections (8), one- or bisegmentectomies at different locations (15), hemihepatectomies (4) and one extended right hepatectomy. With one exception intra-operative blood loss was lower than 100 mL. Four patients (20%) developed operation-related complications comprising abscess formation at the resection site. Follow-up shows tumour-free survival for 13 of 18 patients 12 months after resection. CONCLUSION: Liver resection using the sealer device seems safe. In proximity of hilar structures or large vessels the method is not favourable for the fear of thermal damage. Extended resections are possible but not parenchyma saving. Good indications are atypical (deep) resections - especially in Segment IVb.

Gene expression profiling in hepatocellular carcinoma: upregulation of genes in amplified chromosome regions.

Cytogenetics of hepatocellular carcinoma and adenoma have revealed gains of chromosome 1q as a significant differentiating factor. However, no studies are available comparing these amplification events with gene expression. Therefore, gene expression profiling was performed on tumours cytogenetically well characterized by array-based comparative genomic hybridisation. For this approach analysis was carried out on 24 hepatocellular carcinoma and 8 hepatocellular adenoma cytogenetically characterised by array-based comparative genomic hybridisation. Expression profiles of mRNA were determined using a genome-wide microarray containing 43,000 spots. Hierarchical clustering analysis branched all hepatocellular adenoma from hepatocellular carcinoma. Significance analysis of microarray demonstrated 722 dysregulated genes in hepatocellular carcinoma. Gene set enrichment analysis detected groups of upregulated genes located in chromosome bands 1q22-42 seen also as the most frequently gained regions by comparative genomic hybridisation. Comparison of significance analysis of microarray and gene set enrichment analysis narrowed down the number of dysregulated genes to 18, with 7 genes localised on 1q22 (SCAMP3, IQGAP3, PYGO2, GPATC4, ASH1L, APOA1BP, and CCT3). In hepatocellular adenoma 26 genes in bands 11p15, 11q12, and 12p13 were upregulated. However, the respective chromosome bands were not gained in hepatocellular adenoma. Expression analysis and comparative genomic hybridisation identified an upregulation of genes in amplified regions of 1q. These results may serve to further narrow down the number of candidate driver genes in hepatocarcinogenesis.

Epigenetic defects of hepatocellular carcinoma are already found in non-neoplastic liver cells from patients with hereditary haemochromatosis.

Gene silencing through aberrant CpG island methylation is a frequent epigenetic defect in hepatocellular carcinoma (HCC). However, nothing is known as yet whether aberrant hypermethylation occurs already in non-neoplastic liver cells from patients with hereditary haemochromatosis who have a clearly elevated risk for developing HCC. Therefore, quantitative real-time PCR-based methylation analysis of six genes frequently hypermethylated in HCC (RASSF1A, cyclinD2, p16(INK4a), GSTpi1, SOCS-1, APC) was performed for liver biopsies from patients with hereditary haemochromatosis. For genotyping of the HFE gene restriction enzyme analysis and Pyrosequencing were used. Transcriptional repression of hypermethylated genes was assessed using real-time RT-PCR. Eighty-four percent of all samples with severe hepatic iron overload and a mutated HFE gene (but without HCC) had at least one gene hypermethylated. All six genes tested were affected by aberrant hypermethylation, albeit to a different extent: RASSF1A 55%, cyclinD2 45%, p16(INK4a) 32%, GSTpi1 10%, SOCS-1 6%, APC 8%. Concomitant transcriptional down-regulation was shown for RASSF1A, cyclinD2, GSTpi1 and SOCS-1. Biopsies from haemochromatosis patients showed significantly more aberrant hypermethylation than normal liver tissue or benign liver tumours (P < 0.001) and also to a higher degree. This effect is independent of patient age, cirrhosis or hepatitis infection. This is the first report demonstrating that longstanding severe iron overload is frequently associated with epigenetic defects characteristic of HCC, which reflects the increased risk of these lesions to progress to HCC. Thus, changes in DNA methylation patterns are an early event preceding morphological alterations of malignant transformation and represent promising targets for early detection.

Liver resection for metastasis due to malignant mesenchymal tumours.

While liver resection for colorectal metastases has shown promising long-term survival, data for metastasectomy in sarcoma and leiomyosarcoma patients have not yielded the same optimism. Due to the rarity of the tumour entity it has always been difficult to provide significant data. Advances in tumour classification suggest that most of the metastases formerly classified to be of sarcomatoid and especially leiomyosarcomatoid origin are actually metastases of GISTs (gastro-intestinal stromal tumours). Neoadjuvant/adjuvant imatinib therapy might improve overall survival and enable surgeons to provide resections in previously unresectable patients. Only R0 resection has been proven to prolong survival so far, with a long disease-free interval as the only independent predictor of outcome.

Expression of the interferon-induced Mx proteins in biliary atresia.

Biliary atresia (BA) is a rare disease of the newborn for which the Kasai procedure is curative only for a few of the patients. The dilemma is that all therapeutic attempts to cure the disease are symptomatic because the etiology is still unclear. One theory suggests a progressive inflammatory process, possibly induced by a viral infection. The aim of the present study was to investigate the activity of type I interferons (IFNs) in the livers of patients with BA. Mx proteins, which mediate an early innate immune response, are a very sensitive marker for type I IFN activity (eg, to viral infection). Liver biopsies were taken during the Kasai procedure from 13 newborns with BA who were serologically negative for hepatotropic viruses. Age-matched controls originated from 7 patients with neonatal cholestasis (eg, inspissated bile syndrome), 3 aborted fetuses, and a 10-year-old child. The immunostaining procedure (alkaline phosphatase anti-alkaline phosphatase) was performed with Mx-specific monoclonal antibody. Immunostaining for Mx proteins was positive in the hepatocytes of all newborns with BA, whereas the intrahepatic bile ducts were positive in all but one. In the control group, 8 of 11 liver samples were Mx-negative. This is the first study dealing with the detection of type I IFN activity in the liver of patients with BA. This observation supports the etiologic consideration of type I IFN-mediated immune response. Although positive findings of viruses in patients with BA are still inconsistent, the present study retraces the progressive inflammatory process in BA one more step toward its beginning.