Guanghua Rong

Epithelial Cell Specificity and Apotope Recognition by Serum Autoantibodies in Primary Biliary Cirrhosis

A major enigma of primary biliary cirrhosis (PBC) is the selective targeting of biliary cells. Our laboratory has reported that after apoptosis, human intrahepatic biliary epithelial cells (HiBECs) translocate the E2 subunit of the pyruvate dehydrogenase complex immunologically intact into apoptotic bodies, forming an apotope. However, the cell type and specificity of this reaction has not been fully defined. To address this issue, we investigated whether the E2 subunit of the pyruvate dehydrogenase complex, the E2 subunit of the branched chain 2‐oxo acid dehydrogenase complex, the E2 subunit of the oxo‐glutarate dehydrogenase complex, four additional inner mitochondrial enzymes, and four nuclear antigens remain immunologically intact with respect to postapoptotic translocation in HiBECs and three additional control epithelial cells. We report that all three 2‐oxo acid dehydrogenase enzymes share the ability to remain intact within the apotope of HiBECs. Interestingly, the E2 subunit of the branched chain 2‐oxo acid dehydrogenase complex also remained intact in the other cell types tested. We extended the data, using sera from 95 AMA‐positive and 19 AMA‐negative patients with PBC and 76 controls, by testing for reactivity against the seven mitochondrial proteins studied herein and also the ability of AMA‐negative sera to react with HiBEC apotopes. Sera from 3 of 95 AMA‐positive sera, but none of the controls, reacted with 2,4‐dienoyl coenzyme A reductase 1, an enzyme also present intact only in the HiBEC apotope, but which has not been previously associated with any autoimmune disease. Finally, the specificity of HiBEC apotope reactivity was confined to AMA‐positive sera. Conclusion: We submit that the biliary specificity of PBC is secondary to the unique processes of biliary apoptosis. (HEPATOLOGY 2011)

Deletion of Interleukin-6 in Mice With the Dominant Negative Form of Transforming Growth Factor β Receptor II Improves Colitis but Exacerbates Autoimmune Cholangitis

The role of interleukin‐6 (IL‐6) in autoimmunity attracts attention because of the clinical usage of monoclonal antibodies to IL‐6 receptor (IL‐6R), designed to block IL‐6 pathways. In autoimmune liver disease, activation of the hepatocyte IL‐6/STAT3 (signal transducer and activator of transcription 3) pathway is associated with modulating pathology in acute liver failure, in liver regeneration, and in the murine model of concanavalin A–induced liver inflammation. We have reported that mice expressing a dominant negative form of transforming growth factor β receptor II (dnTGFβRII) under control of the CD4 promoter develop both colitis and autoimmune cholangitis with elevated serum levels of IL‐6. Based on this observation, we generated IL‐6–deficient mice on a dnTGF‐βRII background (dnTGFβRII IL‐6−/−) and examined for the presence of antimitochondrial antibodies, levels of cytokines, histopathology, and immunohistochemistry of liver and colon tissues. As expected, based on reports of the use of anti–IL‐6R in inflammatory bowel disease, dnTGFβRII IL‐6−/− mice manifest a dramatic improvement in their inflammatory bowel disease, including reduced diarrhea and significant reduction in intestinal lymphocytic infiltrates. Importantly, however, autoimmune cholangitis in dnTGFβRII IL‐6−/− mice was significantly exacerbated, including elevated inflammatory cytokines, increased numbers of activated T cells, and worsening hepatic pathology. Conclusion: The data from these observations emphasize that there are distinct mechanisms involved in inducing pathology in inflammatory bowel disease compared to autoimmune cholangitis. These data also suggest that patients with inflammatory bowel disease may not be the best candidates for treatment with anti–IL‐6R if they have accompanying autoimmune liver disease and emphasize caution for therapeutic use of anti–IL‐6R antibody. HEPATOLOGY 2010

Fine phenotypic and functional characterization of effector cluster of differentiation 8 positive T cells in human patients with primary biliary cirrhosis

In primary biliary cirrhosis (PBC), patients develop a multilineage response to a highly restricted peptide of the E2 component of pyruvate dehydrogenase (PDC‐E2) involving autoantibody and autoreactive cluster of differentiation (CD)4+ and CD8+ T‐cell responses. Recent data from murine models have suggested that liver‐infiltrating CD8+ cells play a critical role in biliary destruction in PBC. We hypothesized that chronic antigen stimulation of CD8+ T cells alters effector memory T cell (TEM) frequency and function similar to that seen with chronic viral infections, including failure to terminally differentiate and relative resistance to apoptosis. We have rigorously phenotyped CD8+ T‐cell subpopulations from 132 subjects, including 76 patients with PBC and 56 controls, and report a higher frequency of TEM cells characterized as CD45ROhighCD57+CD8high, but expressing the gut homing integrin, α4β7, in peripheral blood mononuclear cells of PBC. These CD8high TEM cells have reduced expression of Annexin V after TCR stimulation. Consistent with a TEM phenotype, CD45ROhighCD57+CD8high T cells express higher levels of granzyme A, granzyme B, perforin, CCR5 and α4β7, and lower levels of CCR7 and CD28 than other CD8high T cells. Furthermore, interleukin (IL)‐5 produced by CD8+CD57+ T lymphocytes upon in vitro T‐cell receptor stimulation are increased in PBC. Histologically, CD8+CD57+ T cells accumulate around the portal area in PBC. Moreover, CD8+CD57+ T cells respond specifically to the major histocompatibility class I epitope of PDC‐E2. Conclusion: In conclusion, our data demonstrate that CD45ROhighCD57+CD8high T cells are a subset of terminally differentiated cytotoxic TEM cells, which could play a critical role in the progressive destruction of biliary epithelial cells. (HEPATOLOGY 2011;54:1293–1302)

Human intrahepatic biliary epithelial cells engulf blebs from their apoptotic peers

The phagocytic clearance of apoptotic cells is critical for tissue homeostasis; a number of non‐professional phagocytic cells, including epithelial cells, can both take up and process apoptotic bodies, including the release of anti‐inflammatory mediators. These observations are particularly important in the case of human intrahepatic biliary cells (HiBEC), because such cells are themselves a target of destruction in primary biliary cirrhosis, the human autoimmune disease. To address the apoptotic ability of HiBECs, we have focused on their ability to phagocytize apoptotic blebs from autologous HiBECs. In this study we report that HiBEC cells demonstrate phagocytic function from autologous HiBEC peers accompanied by up‐regulation of the chemokines CCL2 [monocyte chemotactic protein‐1 (MCP‐1)] and CXCL8 [interleukin (IL)‐8]. In particular, HiBEC cells express the phagocytosis‐related receptor phosphatidylserine receptors (PSR), implying that HiBECs function through the ‘eat‐me’ signal phosphatidylserine expressed by apoptotic cells. Indeed, although HiBEC cells acquire antigen‐presenting cell (APC) function, they do not change the expression of classic APC function surface markers after engulfment of blebs, both with and without the presence of Toll‐like receptor (TLR) stimulation. These results are important not only for understanding of the normal physiological function of HiBECs, but also explain the inflammatory potential and reduced clearance of HiBEC cells following the inflammatory cascade in primary biliary cirrhosis.

Long-term outcomes of percutaneous cryoablation for patients with hepatocellular carcinoma within Milan criteria.

Background Accumulating evidences have suggested that percutaneous cryoablation could be a valuable alternative ablation therapy for HCC but there has been no large cohort-based analysis on its long-term outcomes. Methods A series of 866 patients with Child-Pugh class A-B cirrhosis and HCC within Milan criteria who underwent percutaneous cryoablation was long-term followed. The safety, efficacy, 5-year survival, and prognostic factors of percutaneous cryoablation in the treatment of HCC were analyzed. Results A total of 1197 HCC lesions were ablated with 1401 cryoablation sessions. Complete response (CR) was achieved in 1163 (97.2%) lesions and 832 (96.1%) patients with 34 (2.8%) major complications, but no treatment-related mortality. After a median of 30.9 months follow-up, 502 (60.3%) patients who achieved CR developed different types of recurrence. The cumulative local tumor recurrence rate was 24.2% at 5-years. Multiple tumor lesions, tumor size > 3 cm, and repeated ablation of same lesion were independent risk factors associated with local recurrence. The 5-year overall survival (OS) rates were 59.5%. Age < 36 years, HCC family history, baseline hepatitis B virus DNA >106 copies/ml, and three HCC lesions were independently and significantly negative predictors to the post-cryoablation OS. Conclusions Percutaneous cryoablation is an effective therapy for patients with HCC within Milan criteria, with comparable efficacy, safety and long-term survival to the reported outcomes of radiofrequency ablation.

Tumour seeding after percutaneous cryoablation for hepatocellular carcinoma

AIM To assess the rate and risk factors for tumour seeding in a large cohort of patients. METHODS Over an 8-year period, 1436 hepatocellular carcinoma (HCC) patients with 2423 tumour nodules underwent 3015 image-guided percutaneous cryoablation sessions [1215 guided by ultrasonography and 221 by spiral computed tomography (CT)]. Follow-up CT or magnetic resonance imaging was performed every 3 mo. The detailed clinical data were recorded to analyse the risk factors for seeding. RESULTS The median follow-up time was 18 (range 1-90) mo. Seeding was detected in 11 patients (0.76%) at 1-24 (median 6.0) mo after cryoablation. Seeding occurred along the needle tract in 10 patients and at a distant location in 1 patient. Seeded tumours usually showed similar imaging and histopathological features to the primary HCCs. Univariate analyses identified subcapsular tumour location and direct subcapsular needle insertion as risk factors for seeding. Multivariate analysis showed that only direct subcapsular needle insertion was an independent risk factor for seeding (P = 0.017; odds ratio 2.57; 95%CI: 1.47-3.65). Seeding after cryoablation occurred earlier in patients with poorly differentiated HCC than those with well or moderately differentiated HCC [1.33 ± 0.577 mo vs 11.12 ± 6.896 mo; P = 0.042; 95%CI: (-19.115)-(-0.468)]. CONCLUSION The risk of seeding after cryoablation for HCC is small. Direct puncture of subcapsular tumours should be avoided to minimise seeding.

Cryotherapy for cirrhosis-based hepatocellular carcinoma: a single center experience from 1595 treated cases

Cryoablation is a less prevalent percutaneous ablative therapy for hepatocellular carcinoma (HCC), and current evidence about its usefulness is limited. We report our experience in treating 1595 HCC cases with percutaneous cryoablation to give a comprehensive profile about the effectiveness, safety and long-term outcome of this therapy. From January 2003 to December 2013, 1595 patients with 2313 HCC nodules were ablated with 2958 cryoablation sessions in our center. Complete ablation was achieved in 1294 patients for 1893 nodules with a mean diameter of 3.4 ± 2.2 cm. The complete ablation rate was 81.2%, 99.4%, 94.4%, and 45.6% in all tumors, tumors < 3 cm, tumors < 5 cm, and tumors > 5 cm, respectively. Major complications were observed after 80 (3.4%) of the 2958 cryoablations and minor complications were observed after 330 cryoablations with no treatment-related deaths. After a median follow-up of 33.4 months, 937 patients developed different types of recurrence. The 5- and 10-year overall survival was 25.7% and 9.2%, respectively. Cryoablation showed reliable safety and efficacy and should be considered as a promising technique, particularly when a large zone of ablation is required.

Blocking and reversing hepatic fibrosis in patients with chronic hepatitis B treated by traditional Chinese medicine (tablets of biejia ruangan or RGT): study protocol for a randomized controlled trial

BackgroundChronic hepatitis B (CHB) can progress to cirrhosis, hepatocellular carcinoma (HCC) and ultimately liver-related death. Although oral antiviral therapy for patients with CHB reduces the risk of such complications, once cirrhosis is established, the benefits of antiviral therapy are not robustly demonstrated. According to traditional Chinese medicine (TCM), some Chinese herbal medicines promote blood circulation and soften hard masses, and therefore they may block and reverse hepatic fibrosis. The aim of this study is to evaluate the effects of TCM tablets of the compound biejia ruangan (RGT) administered for fibrosis, and entecavir (ETV), on the development of HCC in patients with CHB or hepatitis B virus (HBV)-related compensated cirrhosis.Methods/designThis multicenter, centrally randomized, double-blind, placebo-controlled, parallel-group study is planned to complete within 5 years. For the study, 1,000 with CHB or HBV-related compensated cirrhosis are randomly assigned in a 1:1 ratio to a treatment group (0.5 mg ETV once daily; 2 g RGT three times daily) or a control group (0.5 mg ETV once daily; 2 g RGT dummy agent three times daily). The primary end points are the development of HCC and liver-related death. Secondary end points include disease progression and overall survival.DiscussionAlthough antiviral therapy can achieve sustained suppression of HBV replication, thereby preventing cirrhosis, patients with CHB treated with nucleos(t)ide analogs (NUCs) retain a higher risk for HCC compared with patients with inactive disease. Although previous clinical trials with RGT have confirmed the efficacy of blocking and reversing hepatic fibrosis in patients with CHB or compensated cirrhosis, the long-term risk for HCC or disease progression in these patients treated with combination of RGT and NUCs compared with NUCs alone is unclear. Therefore, it is necessary to investigate the effects of the RGT blockade and reversal of hepatic fibrosis on the development of HCC in patients with CHB or HBV-related compensated cirrhosis in large, prospective, multicenter, double-blind, randomized, controlled trials in China.Trial registrationClinicalTrials.gov Identifier: NCT01965418. Date registered: 17 October 2013

The "Vessel through Strait" Sign is a Signature Radiological Sign for the Diagnosis of Left Hepatic Artery Variation.

An aberrant artery (AA) can frequently be observed coursing through the fissure for the ligamentum venosum (FLV) which was termed the “vessel through strait” sign (VTSS) by us. Fundamental data including the incidence, anatomical composition and clinical significance of VTSS and the AAs composing VTSS are still lacking. We sought to give a systematic demonstration on this issue in the present study. VTSS was respectively analyzed in 2,275 patients and was observed in 357 of them. Interestingly, 319 (89.4%) out of the 357 patients exhibiting VTSS were proved to have left hepatic artery variation (LHAV) (247 with replaced left hepatic artery, 64 with accessory left hepatic artery and 8 with variant common hepatic artery). We therefore hypothesized that VTSS could be a sign that strongly associated with LHAV and could be used for its diagnosis. In the following validating analysis, VTSS gained a sensitivity of 96.3% and a specificity of 98.3% for the diagnosis of LHAV in another bicenter cohort consisted of 1,329 patients. In conclusion, VTSS is a signature radiological sign of LHAV which could be used as an easy and specific method for the diagnosis of LHAV.