Undetected human papillomavirus DNA and uterine cervical carcinoma: Association with cancer recurrence.

BACKGROUND: The time course of human papillomavirus (HPV) DNA clearance was studied in patients with carcinoma of the cervix during follow-up after primary radical radiotherapy (RT). This study investigated the relationship between timing of HPV clearance and RT effectiveness. PATIENTS AND METHODS: A total of 71 consecutive patients who were treated for cervical cancer with primary radical radiotherapy and high-dose rate intracavitary brachytherapy with or without chemotherapy were enrolled in the study. Samples for HPV DNA examination were taken before (1) treatment, (2) every brachytherapy, and (3) every follow-up examination. The times when HPV DNA was undetected were analyzed for association with recurrence-free survival. RESULTS: HPV DNA was not detected in 13 patients (18 %) before RT. Of the 58 patients with HPV DNA detected before treatment, HPV DNA was not detected in 34 % during treatment and in 66 % after the treatment. Within 6 months after RT, HPV DNA was detected in 0 % of all patients. The patients were followed up for a median period of 43 months (range 7-70 months). In all, 20 patients were found to develop recurrence. The 3-year cumulative disease-free survival (DFS) rate was 71 ・} 5.4 % for all 71 patients. In multivariate analysis, DFS was significantly associated with HPV (detected vs. not detected) with a hazard ratio of 0.07 (95 % confidence interval 0.008-0.6, p = 0.009). CONCLUSION: In this study, patients in whom HPV was not detected had the worst prognosis. Six months after RT, HPV DNA was detected in 0 % of the patients. Patients in whom HPV DNA could not be detected before treatment need careful follow-up for recurrence and may be considered for additional, or alternative treatment.

CD1d, a sentinel molecule bridging innate and adaptive immunity, is downregulated by the human papillomavirus (HPV) E5 protein: a possible mechanism for immune evasion by HPV.

CD1d and CD1d-restricted natural killer T (NKT) cells serve as a natural bridge between innate and adaptive immune responses to microbes. CD1d downregulation is utilized by a variety of microbes to evade immune detection. We demonstrate here that CD1d is downregulated in human papillomavirus (HPV)-positive cells in vivo and in vitro. CD1d immunoreactivity was strong in HPV-negative normal cervical epithelium but absent in HPV16-positive CIN1 and HPV6-positive condyloma lesions. We used two cell lines for in vitro assay; one was stably CD1d-transfected cells established from an HPV-negative cervical cancer cell line, C33A (C33A/CD1d), and the other was normal human vaginal keratinocyte bearing endogenous CD1d (Vag). Flow cytometry revealed that cell surface CD1d was downregulated in both C33A/CD1d and Vag cells stably transfected with HPV6 E5 and HPV16 E5. Although the steady-state levels of CD1d protein decreased in both E5-expressing cell lines compared to empty retrovirus-infected cells, CD1d mRNA levels were not affected. Confocal microscopy demonstrated that residual CD1d was not trafficked to the E5-expressing cell surface but colocalized with E5 near the endoplasmic reticulum (ER). In the ER, E5 interacted with calnexin, an ER chaperone known to mediate folding of CD1d. CD1d protein levels were rescued by the proteasome inhibitor, MG132, indicating a role for proteasome-mediated degradation in HPV-associated CD1d downregulation. Taken together, our data suggest that E5 targets CD1d to the cytosolic proteolytic pathway by inhibiting calnexin-related CD1d trafficking. Finally, CD1d-mediated production of interleukin-12 from the C33A/CD1d cells was abrogated in both E5-expressing cell lines. Decreased CD1d expression in the presence of HPV E5 may help HPV-infected cells evade protective immunological surveillance.

Utility of quantitative 99mTc-phytate scintigraphy to diagnose early-stage non-alcoholic steatohepatitis.

OBJECTIVE: In patients with non-alcoholic fatty liver disease (NAFLD), liver biopsy remains the only reliable method to differentiate simple steatosis from non-alcoholic steatohepatitis (NASH). The objective of this study was to evaluate the efficacy of non-invasive (99m)Tc-phytate scintigraphy in the diagnosis of NASH. MATERIAL AND METHODS: Thirty-seven patients with suspected NAFLD at the time of liver biopsy also underwent (99m)Tc-phytate scintigraphy. Signal intensities of regions of interest (ROI) in the liver, spleen, and heart were measured. We also examined scintigraphic features in a nutritional model of NASH in rats fed a methionine- and choline-deficient (MCD) diet. RESULTS: The liver/spleen uptake ratio determined by scintigraphy was significantly decreased in patients with NASH in comparison with patients with simple steatosis. The liver/spleen ratio was an independent predictor distinguishing NASH from simple steatosis. The decrease was observed for all stages of NASH, including the early stage (stages 1 and 0). In animal studies, the liver/spleen uptake ratio was significantly decreased in rats after 8 weeks of MCD dietary feeding in comparison with control diet-fed rats. CONCLUSIONS: The non-invasive (99m)Tc-phytate scintigraphy test is a reliable tool to differentiate NASH from simple steatosis.

Immunosuppressant FK506 inhibits matrix metalloproteinase-9 induction in TNF-alpha-stimulated human hepatic stellate cells.

Activated hepatic stellate cells (HSCs) play a key role in the pathogenesis of hepatic inflammation and fibrosis through the production of matrix metalloproteinases (MMPs). Cytokines and growth factors are thought to activate HSCs. TNF-alpha has pleiotropic functions in hepatitis, but its role in liver fibrosis remains elusive. In this study we investigated the regulation of tumor necrosis factor-alpha (TNF-alpha) in the expression of MMPs by HSCs. We also examined whether the immunosuppressant FK506 influences the MMPs expression in human HSCs. Human HSCs, LI90, were treated with TNF-alpha in the presence of FK506. Release of MMPs into culture media, levels of MMP-9 mRNA and activation of NF-kappaB were compared between the cells cultured with or without FK506. Stimulation of human HSCs, LI90 cells, with TNF-alpha caused the induction of pro-MMP-9. Further, TNF-alpha stimulation induced the degradation of IkappaB-alpha and resulted in the transcriptional activation of NF-kappaB. FK506 suppressed this TNF-alpha-induced NK-kappaB activation, alone with pro-MMP-9 mRNA and protein induction, in HSC. TNF-alpha contributes to the perpetuation of liver fibrosis through MMP-9 production from HSCs and that FK506 inhibits the induction of MMP-9 through NF-kappaB pathway suggesting the anti-inflammatory properties of FK506.

Peroxynitrite-mediated matrix metalloproteinase-2 activation in human hepatic stellate cells.

To investigate whether hepatic stellate cells (HSCs) alter their expression of MMPs after exposure to nitrogen oxide intermediate (NOI), a human hepatic stellate cell line, LI90 cells, was stimulated with an NO donor, SNAP, or a peroxynitrite donor, SIN-1, and the culture supernatants were analyzed by gelatin zymography or anti-MMPs immunoblot. Although SIN-1 did not enhance the secretions of MMP-1 and 13, SIN-1 induced the NF-kappaB activation, MT1-MMP expression and the secretion of activated MMP-2 in LI90 cells. These results suggest that peroxynitrite may contribute to the remodeling of the extracellular matrix in liver by activating pro-MMP-2.

Effect of 3-O-octanoyl-(+)-catechin on the responses of GABA(A) receptors and Na+/glucose cotransporters expressed in xenopus oocytes and on the oocyte membrane potential.

Recently, 3-O-octanoyl-(+)-catechin (OC) was synthesized from (+)-catechin (C) by incorporation of an octanoyl chain into C in the light of (-)-epicatechin gallate (ECg) and (-)-epigallocatechin gallate (EGCg), which are the major polyphenols found in green tea and have strong physiological activities. OC was found to inhibit the response of ionotropic gamma-aminobutyric acid (GABA) receptors (GABA(A) receptors) and Na+/glucose cotransporters expressed in Xenopus oocytes in a noncompetitive manner more strongly than does C. OC also induced a nonspecific membrane current and decreased the membrane potential of the oocyte, and thus death of the oocyte occurred even at lower concentrations than that induced by C or EGCg. Although EGCg produced H2O2 in aqueous solution, OC did not. This newly synthesized catechin derivative OC possibly binds to the lipid membrane more strongly than does C, Ecg, or EGCg and as a result perturbs the membrane structure.

Lipopolysaccharide signaling induces serum amyloid A (SAA) synthesis in human hepatocytes in vitro.

To investigate the role of lipopolysaccharide (LPS) in hepatocyte activation, we examined the expression of Toll-like receptor 4 (TLR4), the putative receptor for LPS in human hepatocytes. TLR4 mRNA and protein expression was confirmed in human hepatocytes. Stimulation of human hepatocytes with LPS results in rapid degradation of IkappaB-alpha and mitogen activated protein kinase activation. Human hepatocytes stimulated by LPS produced serum amyloid A protein. Our data suggest that human hepatocytes utilize components of TLR4 signal transduction pathways in response to LPS and these direct LPS-mediated effects on hepatocytes may contribute to liver inflammation and injury.

Polyphenol-induced inhibition of the response of na(+)/glucose cotransporter expressed in Xenopus oocytes.

To study the effects of polyphenols on the Na(+)/glucose cotransporter (SGLT1) response, SGLT1 was expressed in Xenopus oocytes by injecting cRNA synthesized from the cloned cDNA of the small intestine cotransporter of rats, and the electrical response elicited by glucose or galactose was measured by a voltage clamping method. Most phenol derivatives had no effect on the response. However, the polyphenols (+)-catechin, (-)-epicatechin gallate (ECg), and (-)-epigallocatechin gallate (EGCg), which are components of green tea, caused an inhibition of the response, which was almost independent of glucose concentration. The inhibition constants were estimated to be 2.3 mM for (+)-catechin and 0.45 mM for both ECg and EGCg, assuming the noncompetitive inhibition mechanism. Saponin prepared from tea seeds also inhibited the response significantly. Tannic acid and aqueous extracts of teas induced nonspecific electrical responses in both cRNA-injected and noninjected oocytes at lower concentrations than those that caused an inhibition of the SGLT1 response when their dose-dependent effects were examined. These results are possibly helpful in the development of a dietary supplement for diabetic patients.

Oral vaccination against HPV E7 for treatment of cervical intraepithelial neoplasia grade 3 (CIN3) elicits E7-specific mucosal immunity in the cervix of CIN3 patients.

BACKGROUND: Cervical intraepithelial neoplasia grade 3 (CIN3) is a mucosal precancerous lesion caused by high-risk human papillomavirus (HPV). Induction of immunological clearance of CIN3 by targeting HPV antigens is a promising strategy for CIN3 therapy. No successful HPV therapeutic vaccine has been developed. METHODS: We evaluated the safety and clinical efficacy of an attenuated Lactobacillus casei expressing modified full-length HPV16 E7 protein in patients with HPV16-associated CIN3. Ten patients were vaccinated orally during dose optimization studies (1, 2, 4, or 6 capsules/day) at weeks 1, 2, 4, and 8 (Step 1). Seven additional participants were only tested using the optimized vaccine formulation (Step 2), giving a total of 10 patients who received optimized vaccination. Cervical lymphocytes (CxLs) and peripheral blood mononuclear cells (PBMCs) were collected and E7 specific interferon-γ-producing cells were counted (E7 cell-mediated immune responses: E7-CMI) by ELISPOT assay. All patients were re-evaluated 9 weeks after initial vaccine exposure using cytology and biopsy to assess pathological efficacy. RESULTS: No patient experienced an adverse event. E7-CMI in both CxLs and PBMCs was negligible at baseline. All patients using 4-6 capsules/day showed increased E7-CMI in CxLs, whereas patients using 1-2 capsules/day did not. No patient demonstrated an increase in E7-CMI in their PBMCs. In comparison between patients of cohorts, E7-CMI at week 9 (9 wk) in patients on 4 capsules/day was significantly higher than those in patients on 1, 2, or 6 capsules/day. Most patients (70%) taking the optimized dose experienced a pathological down-grade to CIN2 at week 9 of treatment. E7-CMI in CxLs correlated directly with the pathological down-grade. CONCLUSIONS: Oral administration of an E7-expressing Lactobacillus-based vaccine can elicit E7-specific mucosal immunity in the uterine cervical lesions. We are the first to report a correlation between mucosal E7-CMI in the cervix and clinical response after immunotherapy in human mucosal neoplasia.

Adjuvant effect of Japanese herbal medicines on the mucosal type 1 immune responses to human papillomavirus (HPV) E7 in mice immunized orally with Lactobacillus-based therapeutic HPV vaccine in a synergistic manner.

The Japanese herbal medicines, Juzen-taiho-to (JTT) and Hochu-ekki-to (HET), have been shown to enhance humoral immune responses to vaccine antigen when used as adjuvants for prophylactic vaccines. However, their adjuvant effect on mucosal cellular immune responses remains unstudied. The precursor lesion of cervical cancer, high-grade CIN that expresses HPV E7 oncoprotein ubiquitously is a target for HPV therapeutic vaccines that elicit mucosal E7-specific type 1 T cell responses. We have demonstrated that oral immunization with recombinant Lactobacillus casei expressing HPV16 E7 (LacE7) is more effective in eliciting mucosal E7-specific IFNγ-producing cells than subcutaneous or intramuscular antigen delivery. Here we report the synergistic effect of an oral Lactobacillus-based vaccine and Japanese herbal medicines on mucosal immune responses. Oral immunization of mice with LacE7 plus either a Japanese herbal medicine (JTT or HET) or a mucosal adjuvant, heated-labile enterotoxin T subunit (LTB), promotes systemic E7-specific type 1 T cell responses but not mucosal responses. Administration of LacE7 plus either Japanese herbal medicine and LTB enhanced mucosal E7-specific type 1 T cell response to levels approximately 3-fold higher than those after administration of LacE7 alone. Furthermore, secretion of IFNγ and IL-2 into the intestinal lumen was observed after oral administration of LacE7 and was enhanced considerably by the addition of Japanese herbal medicines and LTB. Our data indicated that Japanese herbal medicines, in synergy with Lactobacillus and LTB, enhance the mucosal type 1 immune responses to orally immunized antigen. Japanese herbal medicines may be excellent adjuvants for oral Lactobacillus-based vaccines and oral immunization of LacE7, HET and LTB may have the potential to elicit extremely high E7-specific mucosal cytotoxic immune response to HPV-associated neoplastic lesions.

Characterization of gut-derived intraepithelial lymphocyte (IEL) residing in human papillomavirus (HPV)-infected intraepithelial neoplastic lesions.

PROBLEM: Mucosal T cells are the most likely direct effectors in host anti-human papillomavirus adaptive immunity and regression of cervical intraepithelial neoplasia (CIN) lesions. There are no studies addressing intraepithelial lymphocytes (IELs) in CIN lesions. METHOD OF STUDY: Cervical lymphocytes were collected using cytobrushes from patients with CIN and analyzed by FACS analysis. Comparisons were made between populations of cervical T cells in CIN regressors and non-regressors. RESULTS: A median of 74% of cervical lymphocytes were CD3(+) T cells. Populations of integrin αEß7(+) IEL in CIN lesions varied markedly among patients (6-57%). Approximately half of integrin ß7(+) T cells were CD45RA-negative memory T cells. The number of integrin αEß7(+) cells among cervical T cells was significantly higher in CIN regressors when compared to non-regressors. CONCLUSION: Higher cervical IEL numbers are associated with spontaneous regression of CIN. Accumulation of cervical integrin αEß7(+) IEL may be necessary for local adaptive effector functions.

Oral immunization with a Lactobacillus casei vaccine expressing human papillomavirus (HPV) type 16 E7 is an effective strategy to induce mucosal cytotoxic lymphocytes against HPV16 E7.

Although many clinical trials on human papillomavirus (HPV) therapeutic vaccines have been performed, clinical responses have not been consistent. We have addressed mucosal cytotoxic cellular immune responses to HPV16 E7 after oral immunization of mice with recombinant Lactobacillus casei expressing HPV16 E7 (LacE7). C57BL/6 mice were orally exposed to 0.1-100mg/head of attenuated LacE7 or vehicle (Lac) vaccines at weeks 1, 2, 4, and 8. Responses to subcutaneous or intramuscular injection of an HPV16 E7 fusion protein using the same timing protocol were used for comparison. Oral immunization with LacE7 elicited E7-specific IFN gamma-producing cells (T cells with E7-type 1 immune responses) among integrin alpha 4 beta 7(+) mucosal lymphocytes collected from gut mucosa. An induction of E7-specific granzyme B-producing cells (E7-CTL) exhibiting killer responses toward HPV16 E7-positive cells was also observed. The induction of T cells with specific mucosal E7-type 1 immune responses was greater after oral immunization with LacE7 when compared to subcutaneous or intramuscular antigen delivery. Oral immunization with Lactobacillus-based vaccines was also able to induce mucosal cytotoxic cellular immune responses. This novel approach at a therapeutic HPV vaccine may achieve more effective clinical responses through its induction of mucosal E7-specific CTL.

Increased expression of Toll-like receptor 3 in intrahepatic biliary epithelial cells at sites of ductular reaction in diseased livers.

Background Toll-like receptors (TLRs) may play active roles in both innate and adaptive immune responses in human intrahepatic biliary epithelial cells (HIBECs). The role of TLR3 expressed by HIBECs, however, remains unclear. Methods We determined the in vivo expression of TLRs in biopsy specimens derived from diseased livers immunohistochemically using a panel of monoclonal antibodies against human TLRs. We then examined the response of cultured HIBECs to a TLR3 ligand, polyinosinic-polycytidylic acid (polyI:C). Using siRNAs specific for Toll-IL-1R homology domain-containing adaptor molecule 1 (TICAM-1) and mitochondrial antiviral signaling protein (MAVS), we studied signaling pathways inducing IFN-beta expression. Results The expression of TLR3 was markedly increased in biliary epithelial cells at sites of ductular reaction in diseased livers, including primary biliary cirrhosis (PBC), autoimmune hepatitis (AIH), and chronic viral hepatitis (CH) as compared to nondiseased livers. Although cultured HIBECs constitutively expressed TLR3 at both the protein and mRNA levels in vitro, the addition of polyI:C to culture media induced only minimal increases in IFN-beta mRNA. In contrast, transfection of HIBECs with polyI:C induced a marked increase in mRNAs encoding a variety of chemokines/cytokines, including IFN-beta, IL-6, and TNF-alpha. The induction of IFN-beta mRNA was efficiently inhibited by an siRNA against MAVS but not against TICAM-1, indicating that the main signaling pathway for IFN-beta induction following polyI:C transfection is via retinoic acid-inducible gene I (RIG-I)/melanoma differentiation-associated gene 5 (MDA5) in HIBECs. Conclusions TLR3 expression by biliary epithelial cells increased at sites of ductular reaction in diseased livers; further study will be necessary to characterize it's in vivo physiological role.

Anti-gp210 and anti-centromere antibodies are different risk factors for the progression of primary biliary cirrhosis.

UNLABELLED: The predictive role of antinuclear antibodies (ANAs) remains elusive in the long-term outcome of primary biliary cirrhosis (PBC). The progression of PBC was evaluated in association with ANAs using stepwise Cox proportional hazard regression and an unconditional stepwise logistic regression model based on the data of 276 biopsy-proven, definite PBC patients who have been registered to the National Hospital Organization Study Group for Liver Disease in Japan (NHOSLJ). When death of hepatic failure/liver transplantation (LT) was defined as an end-point, positive anti-gp210 antibodies (Hazard ratio (HR) = 6.742, 95% confidence interval (CI): 2.408, 18.877), the late stage (Scheuer's stage 3, 4) (HR = 4.285, 95% CI:1.682,10.913) and male sex (HR = 3.266, 95% CI: 1.321,8.075) were significant risk factors at the time of initial liver biopsy. When clinical progression to death of hepatic failure/LT (i.e., hepatic failure type progression) or to the development of esophageal varices or hepatocellular carcinoma without developing jaundice (Total bilirubin < 1.5 mg/dL) (i.e., portal hypertension type progression) was defined as an end-point in the early stage (Scheuer's stage 1, 2) PBC patients, positive anti-gp210 antibodies was a significant risk factor for hepatic failure type progression [odds ratio (OR) = 33.777, 95% CI: 5.930, 636.745], whereas positive anti-centromere antibodies was a significant risk factor for portal hypertension type progression (OR = 4.202, 95% CI: 1.307, 14.763). Histologically, positive anti-gp210 antibodies was most significantly associated with more severe interface hepatitis and lobular inflammation, whereas positive anticentromere antibodies was most significantly associated with more severe ductular reaction. CONCLUSION: These results indicate 2 different progression types in PBC, hepatic failure type and portal hypertension type progression, which may be represented by positive-anti-gp210 and positive-anticentromere antibodies, respectively.

Human intrahepatic biliary epithelial cells function in innate immunity by producing IL-6 and IL-8 via the TLR4-NF-kappaB and -MAPK signaling pathways.

BACKGROUND: Human intrahepatic biliary epithelial cells (HIBECs) may play active roles in both the innate and adaptive immune responses. Little is known, however, about the role of toll-like receptors (TLRs) on HIBECs in inflammatory cholangiopathies. METHODS: The expression of TLR1-9 and the biological responses to their ligands, lipopolysaccharide (LPS) or lipoteichoic acid (LTA), were studied in cultured HIBECs by reverse transcription-polymerase chain reaction, immunoblotting, and enzyme-linked immunosorbent assay. RESULTS: HIBECs constitutively expressed transcripts encoding TLR1-6 and 9, as well as myeloid differentiation factor 88 (MyD88), MD2, and CD14. Stimulation of HIBECs with LPS resulted in translocation of NF-kappaB subunits from the cytoplasmic to the nuclear fraction, followed by increased secretion of a variety of chemokines/cytokines, including interleukin-8 (IL-8), monocyte chemotactic protein-1 (MCP-1), and IL-6. Treatment with BAY11-7082 efficiently inhibited the LPS-induced transcription and secretion of these chemokines/cytokines. In HIBECs, the mitogen-activated protein kinases (MAPKs) were also activated by LPS stimulation. These results indicated that LPS activates HIBECs via a TLR4-MyD88-dependent pathway. Stimulation of HIBECs with LTA induced the secretion of a similar profile of cytokines/chemokines via a TLR2-MyD88-dependent pathway. CONCLUSIONS: In HIBECs, at least TLR2 and 4 are capable of mediating innate immune system function in vitro. This result, in conjunction with our recent finding that TLR4 expression is increased in biliary epithelial cells in primary biliary cirrhosis, suggests the involvement of TLRs in the development of chronic inflammatory cholangiopathies.

Increased expression of nuclear envelope gp210 antigen in small bile ducts in primary biliary cirrhosis.

The sustained antibody response to nuclear envelope gp210 antigen indicates a group of primary biliary cirrhosis (PBC) patients at high risk for the progression to end-stage hepatic failure. To address this issue, we immunohistochemically studied the expression of gp210 antigen in needle liver biopsy specimens from PBC patients using a monoclonal antibody specific for gp210 antigen. The specimens from autoimmune hepatitis (AIH), chronic viral hepatitis B (CHB) and C (CHC) patients served as disease controls. The expression of gp210 antigen was apparently increased on the nuclear envelope of biliary epithelial cells (BECs) of small bile ducts in almost all specimens from PBC. In contrast, the expression of gp210 antigen was negative in BECs of small bile ducts in normal liver, while relatively weak anti-gp210 immunostaining was observed in AIH, CHC and CHB. In addition, the degree of gp210 expression in BECs of small bile ducts was positively correlated to that of portal inflammation, interface hepatitis and lobular inflammation in PBC. These results indicate that the increased expression of gp210 in small bile ducts, which is probably associated with damage to BECs by inflammation, is possibly involved in autoimmune response to gp210 leading to the progression to end-stage hepatic failure in PBC.

Hepatic expression of toll-like receptor 4 in primary biliary cirrhosis.

Toll-like receptor 4 (TLR4) is a receptor for bacterial lipopolysaccharide, which is suggested to be involved in the pathogenesis of disease of hepatobiliary tracts. To explore a possible role for this receptor in primary biliary cirrhosis (PBC), we investigated the expression of TLR4 in liver tissues from PBC patients. We studied liver biopsy sections from 62 PBC patients and 41 patients with chronic hepatitis C (CHC). Expression of TLR4 in paraffin-embedded sections was analyzed by immunohistochemistry. The bile duct epithelial cells (BECs) of PBC liver tissues markedly expressed TLR4, whereas BECs of CHC liver tissues barely expressed TLR4. The TLR4 expression was also observed in periportal hepatocytes of PBC liver tissues and its expression was extended to interlobular hepatocytes in advanced stage PBC. Although periportal hepatocytes of CHC liver tissues expressed TLR4, its expression levels were not correlated with the fibrosis stage. Our data demonstrated that TLR4 was expressed in BECs and periportal hepatocytes in PBC livers, suggesting the possible involvement of bacterial pathogens and TLR4 in the inflammatory processes of PBC.

Antibody titer to gp210-C terminal peptide as a clinical parameter for monitoring primary biliary cirrhosis.

BACKGROUND/AIMS: The presence of antibodies to the 210-kDa glycoprotein of the nuclear pore complex (gp210) is highly indicative of primary biliary cirrhosis (PBC). However, the significance of anti-gp210 antibody titers for monitoring PBC remains unresolved. METHODS: We used an ELISA with a gp210 C-terminal peptide as an antigen to assess serum antibody titers in 71 patients with PBC. RESULTS: Patients were classified into three groups: Group A in whom anti-gp210 titers were sustained at a high level, Group B in whom anti-gp210 status changed from positive to negative under ursodeoxycholic acid (UDCA) therapy, Group C in whom anti-gp210 antibodies were negative at the time of diagnosis. The rate of progression to end-stage hepatic failure was significantly higher in group A (60%) as compared to groups B (0%) and C (4.2%). The sustained antibody response to gp210 was closely associated with the severity of interface hepatitis. The significance of anti-gp210 antibody was confirmed by National Hospital Organization Study Group for Liver Disease in Japan. CONCLUSIONS: The serial quantitation of serum anti-gp210-C-terminal peptide antibodies is useful for monitoring the effect of UDCA and for the early identification of patients at high risk for end-stage hepatic failure.

Enhanced expression of type I interferon and toll-like receptor-3 in primary biliary cirrhosis.

The pathogenesis of primary biliary cirrhosis (PBC) remains enigmatic. In order to address this issue, we analyzed by laser capture microdissection and real-time reverse transcription-polymerase chain reaction the site-specific expression of messenger RNA (mRNA) for cytokines (interferon (IFN)-alpha, -beta, -gamma, interleukin (IL)-1beta, -4, -6, -10, -12p40, -18, tumor necrosis factor-alpha) and toll-like receptors (TLRs) (TLR-2, -3, -4, -7, -9) in portal tract and liver parenchyma from patients with early-stage PBC. Expression of IFN-alpha, -beta and TLR-3 proteins was also studied by immunohistochemistry. Autoimmune hepatitis (AIH) and chronic hepatitis C (CHC) served as disease controls. The expression levels of type I IFN (IFN-alpha, -beta) and TLR-3 mRNAs, which are known to induce type I IFN, were significantly higher in portal tract and liver parenchyma as compared to AIH and CHC. A strong positive correlation between the mRNA levels of type I IFN and TLR-3 was also seen in both areas. Immunohistologically, IFN-alpha is present in the mononuclear cells in portal tract and sinusoidal cells. Macrophages in portal tract and hepatocytes expressed IFN-beta and TLR-3. Furthermore, the level of IFN-alpha mRNA in the portal tract was positively correlated with serum alkaline phosphatase. In conclusion, these data indicate that TLR-3 and type I IFN signaling pathways are active in both the portal tract and liver parenchyma of early-stage PBC, and form the basis for our hypothesis that these signaling pathways are involved in the pathophysiology of PBC.