Analysis of microvascular density in early gastric carcinoma using magnifying endoscopy with narrow-band imaging.

BACKGROUND AND STUDY AIMS: Intramucosal vascular density differs between differentiated and undifferentiated type gastric carcinomas. This study aimed to evaluate the microvascular density characteristics of these two types of carcinoma using magnifying endoscopy with narrow-band imaging (ME-NBI). PATIENTS AND METHODS: In total, 42 differentiated and 10 undifferentiated types were evaluated. The microvessels observed using ME-NBI were extracted from stored still images and the microvascular density in the two carcinoma types was analyzed. Histological vascular density in resected specimens was also evaluated using CD34 immunostaining. RESULTS: There were significant differences between the microvascular density in the differentiated and undifferentiated types of carcinoma (10.02 ±â€Š4.72 % vs 4.02 ±â€Š0.40 %; P < 0.001) using ME-NBI. Vascular density assessed histologically also differed significantly between differentiated and undifferentiated types in both the whole mucosal (5.81 ±â€Š3.17 % vs 3.25 ±â€Š1.21 %) and the superficial mucosal layers (0 - 100 µm) (6.38 ±â€Š3.73 % vs 3.66 ±â€Š1.46 %). However, the vascular density in the surrounding non-carcinomatous mucosa assessed using ME-NBI and histologically, was significantly lower in the differentiated than in the undifferentiated types (P < 0.001). There was good agreement between ME-NBI and histologically assessed microvascular density in both the whole (r = 0.740; P < 0.001) and superficial mucosal layers (r = 0.764; P < 0.001). White opaque substance (WOS) was seen in eight patients who had the differentiated type carcinoma. In almost all cases with WOS, the appearance of the carcinoma was discolored. CONCLUSIONS: There was a close relationship between ME-NBI assessed microvascular density and histologically assessed vascular density in the mucosal layer. Microvascular density differed significantly between the differentiated and undifferentiated types of carcinoma assessed using ME-NBI.

[Two cases of squamoid cyst of the pancreatic ducts].

Two patients were referred to our hospital with cystic lesion (diameter 5 cm) of the pancreas and elevated serum CEA and CA19-9. We diagnosed them with malignant cystic neoplasms of the pancreas and performed distal pancreatectomy. Histologically, in both cases the cysts were lined with flat, transitional, squamoid cells without keratinization. Immunohistochemical staining confirmed two rare cases of squamoid cyst of the pancreatic ducts.

Connexins regulate cell functions in pancreatic stellate cells.

OBJECTIVES: Pancreatic stellate cells (PSCs) play a pivotal role in pancreatic fibrosis associated with chronic pancreatitis and pancreatic cancer. Connexins (Cxs) allow direct intercellular communications as components of gap junction but also play important roles in the regulation of cell proliferation, cell differentiation, and tissue development. We here examined the expression of Cxs and Cx-mediated regulation of cell functions in PSCs. METHODS: Human PSCs were isolated from patients undergoing operation for chronic pancreatitis or pancreatic cancer. The expression of Cxs was examined by reverse transcription polymerase chain reaction, Western blotting, and immunofluorescent staining. The roles of Cxs in PSC functions were examined by using carbenoxolone, a broad-spectrum Cx inhibitor, and small interfering RNA for Cx43. RESULTS: Human activated PSCs expressed a variety of Cxs including Cx43 both in vitro and in vivo. Carbenoxolone inhibited platelet-derived growth factor-BB-induced proliferation and migration, and type I collagen expression in PSCs. In addition, carbenoxolone inhibited the activation of quiescent PSCs to a myofibroblastlike phenotype. Decreased Cx43 expression by small interfering RNA resulted in decreased proliferation and type I collagen expression. CONCLUSIONS: Pancreatic stellate cells expressed a variety of Cxs. Connexins, especially Cx43, might regulate the cell functions and activation of PSCs.

Response to peginterferon-alpha 2b and ribavirin in Japanese patients with chronic hepatitis C genotype 1.

PURPOSE: Patient age and gender may be associated with response to peginterferon alpha plus ribavirin, the current standard of care (SOC) for chronic hepatitis C genotype 1. We queried whether there was an association between age, gender, and treatment response to SOC in Japanese patients infected with hepatitis C virus (HCV) genotype 1. METHODS: Between 2006 and 2009, HCV-infected Japanese patients treated with peginterferon alpha-2b plus ribavirin for 48 weeks were enrolled. Patients were allocated into four groups according to age and gender, and epidemiological data and treatment outcomes were retrospectively analyzed. HCV RNA was measured with COBAS AMPLICOR HCV Monitor Test v. 2.0. RESULTS: The overall sustained virological response (SVR) rate was 49.8%: patients aged ≤65 and >65 years, 50.9 and 44.0%, respectively; male and female, 56.5 and 39.0%. SVR rates of SOC against HCV genotype-1 females aged >65 years (19.0%) were inferior to those in males aged >65 years (57.8%) in Japan. Multivariate logistic regression analysis showed that SVR was attained independently of adherence 80/80/80 in all groups. CONCLUSIONS: Adherence to medication is also a key factor for the eradication of HCV in patients aged >65 years. As the SVR rate of patients aged ≤65 years was similar to that of patients aged >65 years, SOC could be useful for treating some of the elderly patients.

Do genetic variants in the SPINK1 gene affect the level of serum PSTI?

BACKGROUND: The serine protease inhibitor Kazal type 1 (SPINK1), also known as pancreatic secretory trypsin inhibitor (PSTI), is a peptide secreted by pancreatic acinar cells. Genetic studies have shown an association between SPINK1 gene variants and chronic pancreatitis or recurrent acute pancreatitis. The aim of this study was to clarify whether the SPINK1 variants affect the level of serum PSTI. METHODS: One hundred sixty-three patients with chronic pancreatitis or recurrent acute pancreatitis and 73 healthy controls were recruited. Serum PSTI concentrations were determined with a commercial radioimmunoassay kit. RESULTS: Ten patients with the p.N34S variant, 7 with the IVS3+2T>C variant, two with both the p.N34S and the IVS3+2T>C variants, and one with the novel missense p.P45S variant in the SPINK1 gene were identified. The serum PSTI level in patients with no SPINK1 variants was 14.3 ± 9.6 ng/ml (mean ± SD), and that in healthy controls was 10.7 ± 2.2 ng/ml. The PSTI level in patients carrying the IVS3+2T>C variant (5.1 ± 3.4 ng/ml), but not in those with the p.N34S variant (8.9 ± 3.5 ng/ml), was significantly lower than that in the patients without the SPINK1 variants and the healthy controls. The serum PSTI level in the patient with the p.P45S variant was 4.9 ng/ml. Low levels of serum PSTI (<6.0 ng/ml) showed sensitivity of 80 %, specificity of 97 %, and accuracy of 96 % in the differentiation of IVS3+2T>C and p.P45S carriers from non-carriers. CONCLUSION: Serum PSTI levels were decreased in patients with the IVS3+2T>C and p.P45S variants of the SPINK1 gene.

Pancreatic stellate cells enhance stem cell-like phenotypes in pancreatic cancer cells.

The interaction between pancreatic cancer cells and pancreatic stellate cells (PSCs), a major profibrogenic cell type in the pancreas, is receiving increasing attention. There is accumulating evidence that PSCs promote the progression of pancreatic cancer by increasing cancer cell proliferation and invasion as well as by protecting them from radiation- and gemcitabine-induced apoptosis. Recent studies have identified that a portion of cancer cells, called "cancer stem cells", within the entire cancer tissue harbor highly tumorigenic and chemo-resistant phenotypes, which lead to the recurrence after surgery or re-growth of the tumor. The mechanisms that maintain the "stemness" of these cells remain largely unknown. We hypothesized that PSCs might enhance the cancer stem cell-like phenotypes in pancreatic cancer cells. Indirect co-culture of pancreatic cancer cells with PSCs enhanced the spheroid-forming ability of cancer cells and induced the expression of cancer stem cell-related genes ABCG2, Nestin and LIN28. In addition, co-injection of PSCs enhanced tumorigenicity of pancreatic cancer cells in vivo. These results suggested a novel role of PSCs as a part of the cancer stem cell niche.

Response to peginterferon-alfa 2b and ribavirin in Japanese patients with chronic hepatitis C genotype 2.

BACKGROUND: The current standard treatment for patients infected with hepatitis C virus (HCV) of genotype 2 is the combination of peginterferon (PEG-IFN) plus ribavirin (RBV) for 24 weeks. AIMS: We assessed the sustained virological response (SVR) rates in HCV genotype 2-infected Japanese patients in relation to the duration of treatment. METHODS: Between 2006 and 2009, among 147 patients with HCV genotype 2-infection in Chiba Prefecture, 138 consecutive patients were finally enrolled. Twenty-one, 97 and 20 patients were treated with PEG-IFN-alfa 2b plus RBV for 16, 24 and 48 weeks, respectively. Epidemiological data and treatment outcomes were retrospectively evaluated. HCV RNA was measured with COBAS AMPLICOR HCV Monitor Test v. 2.0. RESULTS: The overall SVR rate was 82.6% (114 of 138): treatment-naïve patients, 86.4% (89 of 103); patients with history of previous treatment, 71.4% (25 of 35). Patients treated for 16, 24 and 48 weeks obtained SVR rates of 66.6% (14 of 21), 86.5% (84 of 97) and 80.0 (16 of 20), respectively. CONCLUSIONS: The SVR rates of PEG-IFN-alfa 2b plus RBV in Japanese patients were similar to those in previous studies. Combination treatment for 24 weeks for some patients infected with HCV genotype 2 may be superior to that for 16 weeks. More precise patient selection will be needed to shorten the combination treatment.

-651C/T promoter polymorphism in the CD14 gene is associated with severity of acute pancreatitis in Japan.

PURPOSE: This study aimed to clarify the association of the promoter variants in the CD14 gene with pancreatic diseases in Japan. METHODS: Three hundred forty-six unrelated patients with acute pancreatitis (AP) (107 with severe and 239 with mild), 263 patients with chronic pancreatitis (CP), 264 patients with pancreatic neoplasm, and 319 healthy controls were genotyped for the single nucleotide polymorphisms at positions -260 and -651 from the AUG start codon in the CD14 gene by polymerase chain reaction-restriction enzyme digestion. RESULTS: The allele and genotype frequencies of the -260C/T and -651C/T polymorphisms did not differ between controls and patients with AP. In subgroup analyses, patients with severe AP had more -651C allele than controls [P = 0.005; odds ratio (OR) 1.71; 95% confidence interval (CI) = 1.18-2.49] or patients with mild AP (P = 0.001; OR 1.95; 95% CI = 1.33-2.85). Genotype -651CC was more common (P = 0.001 vs. controls and P = 0.001 vs. mild AP), and -651CT was less (P = 0.009 vs. controls and P = 0.007 vs. mild AP) in patients with severe AP than in healthy controls or patients with mild AP. The frequencies of pseudocyst development and requirement of surgery were higher in AP patients with -651CC than in those without this genotype. The -260C/T polymorphism was not associated with the severity of AP. The allele and genotype frequencies of both polymorphisms did not differ between controls and patients with CP or pancreatic neoplasm. CONCLUSION: -651C/T promoter polymorphism in the CD14 gene was associated with severity of AP in Japan.

Effects of ethanol and its metabolites on human pancreatic stellate cells.

Pancreatic stellate cells (PSCs) play a pivotal role in pancreatic inflammation and fibrosis. In the pancreas, in addition to oxidative metabolism, ethanol can be metabolized by esterification with fatty acids to form fatty acid ethyl esters such as palmitic acid ethyl ester (PAEE). We here examined the effects of ethanol (at 20 or 50 mM), acetaldehyde (at 200 microM), or PAEE (at 100 microM), on PSCs functions. PSCs did not express mRNAs for pancreatic triglyceride lipase and carboxyl ester lipase. Ethanol and acetaldehyde, but not PAEE, induced production of procollagen type I C-peptide. Ethanol, but not acetaldehyde or PAEE, induced interleukin-8 production. PAEE activated activator protein-1, but not nuclear factor kappaB. In addition, PAEE activated extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 mitogen-activated protein kinase. Specific activation of signal transduction pathways and cell functions by ethanol and its metabolites may play a role in alcohol-induced pancreatic injury.

Ellagic acid inhibits pancreatic fibrosis in male Wistar Bonn/Kobori rats.

The key pathological features of chronic pancreatitis are chronic inflammation, acinar atrophy, and pancreatic fibrosis. We have previously shown that ellagic acid, a plant-derived polyphenol found in fruits and nuts, inhibited activation of pancreatic stellate cells, a major profibrogenic cell type in the pancreas, in vitro. Here we examined whether ellagic acid inhibited the development of pancreatic fibrosis in vivo. Ellagic acid was administered orally in the diet to ten-week-old male Wistar Bonn/Kobori rats, an experimental model of spontaneous chronic pancreatitis, for ten weeks. Ellagic acid (100 mg/kg body weight/day) attenuated pancreatic inflammation and fibrosis. The protective effects were confirmed by an increase in pancreatic weight and decreases in myeloperoxidase activity (an index of neutrophil infiltration), collagen content, transforming growth factor-beta1 expression, and the number of alpha-smooth muscle actin-positive cells (activated pancreatic stellate cells) and ED-1-positive cells (macrophages/monocytes). Ellagic acid inhibited the production of reactive oxygen species in pancreatic stellate cells in response to transforming growth factor-beta1 or platelet-derived growth factor. Our results suggest that ellagic acid might be a candidate for treatment of chronic pancreatitis.

Hydrogen peroxide activates activator protein-1 and mitogen-activated protein kinases in pancreatic stellate cells.

Activated pancreatic stellate cells (PSCs) are implicated in the pathogenesis of pancreatic inflammation and fibrosis, where oxidative stress is thought to play a key role. Reactive oxygen species such as hydrogen peroxide (H(2)O(2)) may act as a second messenger to mediate the actions of growth factors and cytokines. But the role of reactive oxygen species in the activation and regulation of cell functions in PSCs remains largely unknown. We here examined the effects of H(2)O(2) on the activation of signal transduction pathways and cell functions in PSCs. PSCs were isolated from the pancreas of male Wistar rats, and used in their culture-activated, myofibroblast-like phenotype unless otherwise stated. Activation of transcription factors was examined by electrophoretic mobility shift assay and luciferase assay. Activation of mitogen-activated protein (MAP) kinases was assessed by Western blotting using anti-phosphospecific antibodies. The effects of H(2)O(2) on proliferation, alpha(1)(I)procollagen gene expression, and monocyte chemoattractant protein-1 production were evaluated. The effect of H(2)O(2) on the transformation of freshly isolated PSCs in culture was also assessed. H(2)O(2) at non-cytotoxic concentrations (up to 100 microM) induced oxidative stress in PSCs. H(2)O(2) activated activator protein-1, but not nuclear factor kappaB. In addition, H(2)O(2) activated three classes of MAP kinases: extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 MAP kinase. H(2)O(2) induced alpha(1)(I)procollagen gene expression but did not induce proliferation or monocyte chemoattractant protein-1 production. H(2)O(2) did not initiate the transformation of freshly isolated PSCs to myofibroblast-like phenotype. Specific activation of these signal transduction pathways and collagen gene expression by H(2)O(2) may play a role in the pathogenesis of pancreatic fibrosis.

Curcumin blocks activation of pancreatic stellate cells.

Activated pancreatic stellate cells (PSCs) play a pivotal role in the pathogenesis of pancreatic fibrosis and inflammation. Inhibition of activation and cell functions of PSCs is a potential target for the treatment of pancreatic fibrosis and inflammation. The polyphenol compound curcumin is the yellow pigment in curry, and has anti-inflammatory and anti-fibrotic properties. We here evaluated the effects of curcumin on the activation and cell functions of PSCs. PSCs were isolated from rat pancreas tissue and used in their culture-activated, myofibroblast-like phenotype unless otherwise stated. The effects of curcumin on proliferation, alpha-smooth muscle actin gene expression, monocyte chemoattractant protein (MCP)-1 production, and collagen expression were examined. The effect of curcumin on the activation of freshly isolated cells in culture was also assessed. Curcumin inhibited platelet-derived growth factor (PDGF)-induced proliferation, alpha-smooth muscle actin gene expression, interleukin-1beta- and tumor necrosis factor (TNF)-alpha-induced MCP-1 production, type I collagen production, and expression of type I and type III collagen genes. Curcumin inhibited PDGF-BB-induced cyclin D1 expression and activation of extracellular signal-regulated kinase (ERK). Curcumin inhibited interleukin-1beta- and TNF-alpha-induced activation of activator protein-1 (AP-1) and mitogen-activated protein (MAP) kinases (ERK, c-Jun N-terminal kinase (JNK), and p38 MAP kinase), but not of nuclear factor-kappaB (NF-kappaB). In addition, curcumin inhibited transformation of freshly isolated cells to myofibroblast-like phenotype. In conclusion, curcumin inhibited key cell functions and activation of PSCs.

Endothelin-1 stimulates contraction and migration of rat pancreatic stellate cells.

AIM: Activated pancreatic stellate cells (PSCs) are implicated in the pathogenesis of pancreatic fibrosis and inflammation. Endothelin-1 (ET-1), which acts through G-protein coupled ET(A) and ET(B) receptors, has several biological activities. We here examined the ability of ET-1 to affect the cell functions of PSCs and the underlying molecular mechanisms. METHODS: PSCs were isolated from the pancreas of male Wistar rats after perfusion with collagenase, and cells between passages two and five were used. Expression of ET-1 and ET receptors was assessed by reverse transcription-PCR and immunostaining. Phosphorylation of myosin regulatory light chain (MLC), extracellular-signal regulated kinase (ERK), and Akt was examined by Western blotting. Contraction of PSCs was assessed on hydrated collagen lattices. Cell migration was examined using modified Boyden chambers. Cell proliferation was assessed by measuring the incorporation of 5-bromo-2'-deoxyuridine. RESULTS: Culture-activated PSCs expressed ET(A) and ET(B) receptors, and ET-1. ET-1 induced phosphorylation of MLC and ERK, but not Akt. ET-1 induced contraction and migration, but did not alter proliferation of PSCs. ET-1-induced contraction was inhibited by an ET(A) receptor antagonist BQ-123 and an ET(B) receptor antagonist BQ-788, whereas migration was inhibited by BQ-788 but not by BQ-123. A Rho kinase inhibitor Y-27632 abolished both contraction and migration. CONCLUSION: ET-1 induced contraction and migration of PSCs through ET receptors and activation of Rho-Rho kinase. ET(A) and ET(B) receptors play different roles in the regulation of these cellular functions in response to ET-1.

Ellagic acid blocks activation of pancreatic stellate cells.

Activated pancreatic stellate cells (PSCs) play a pivotal role in the pathogenesis of pancreatic fibrosis and inflammation. Ellagic acid is a plant-derived polyphenol found in fruits and nuts, and has anti-oxidant and anti-inflammatory properties. But, little is known about the effects of ellagic acid on PSCs as well as on the activation of signal transduction pathways. We here evaluated the effects of ellagic acid on the activation and cell functions of PSCs. PSCs were isolated from rat pancreas tissue and used in their culture-activated, myofibroblast-like phenotype unless otherwise stated. Ellagic acid inhibited platelet-derived growth factor (PDGF)-BB-induced proliferation and migration, interleukin (IL)-1beta- and tumor necrosis factor (TNF)-alpha-induced monocyte chemoattractant protein-1 production, and expression of alpha-smooth muscle actin and collagen genes. Ellagic acid inhibited PDGF-BB-induced tyrosine phosphorylation of PDGF beta-receptor and the downstream activation of extracellular signal-regulated kinase and Akt. Ellagic acid inhibited IL-1beta- and TNF-alpha-induced activation of activator protein-1 and mitogen-activated protein kinases (extracellular signal-regulated kinase, c-Jun N-terminal kinase and p38 mitogen-activated protein kinase), but not of nuclear factor-kappaB. In addition, ellagic acid inhibited transformation of freshly isolated cells to an activated, myofibroblast-like phenotype. In conclusion, ellagic acid inhibited key cell functions and activation of PSCs.

Mutations in the serine protease inhibitor Kazal Type 1 (SPINK1) gene in Japanese patients with pancreatitis.

BACKGROUND/AIMS: Recent studies have shown an association between the N34S mutation in the serine protease inhibitor Kazal type 1 (SPINK1) gene and chronic pancreatitis (CP). We here examined the prevalence of SPINK1 mutations in Japanese patients with pancreatitis. METHODS: Genomic DNA was prepared from 80 Japanese patients with CP, 36 patients with acute pancreatitis (AP), and 165 healthy controls. All exons and the promotor region of the SPINK1 gene were amplified by the polymerase chain reaction, and directly sequenced. RESULTS: We found four types of mutation (N34S, IVS1-37T>C, -215G>A, and IVS3 + 2T>C) and two types of polymorphism (-253T>C and 272C>T). The N34S mutation cosegregated with IVS1-37T>C, and was present in 8 CP and 1 AP patients. The -215G>A mutation was in a complete linkage with IVS3 + 2T>C, and was present in 8 CP and 1 AP patients. The prevalences of [N34S; IVS1-37T>C] and [-215G>A; IVS3 + 2T>C] were significantly higher in patients with familial pancreatitis (38 and 13%, respectively) and with idiopathic CP (13 and 16%) than normal subjects (0.6 and 0%). In addition, the frequency of [N34S; IVS1-37T>C] mutation was higher in patients with autoimmune CP (33%). CONCLUSION: The SPINK1 gene mutations were associated with pancreatitis also in Japan.

Green tea polyphenol epigallocatechin-3-gallate blocks PDGF-induced proliferation and migration of rat pancreatic stellate cells.

AIM: To clarify the effects of epigallocatechin-3-gallate (EGCG) on the platelet-derived growth factor (PDGF)-BB-induced proliferation and migration of pancreatic stellate cells (PSCs). METHODS: PSCs were isolated from rat pancreas tissue and used in their culture-activated, myofibroblast-like phenotype. Cell proliferation was assessed by measuring the incorporation of 5-bromo-2'-deoxyuridine. Cell migration was assessed using modified Boyden chambers. Cyclin D1, p21Waf1, and p27Kip1 expression and phosphorylation of PDGF beta-receptor, extracellular signal-regulated kinase, and Akt were examined by Western blotting. Activation of phospha-tidylinositol 3-kinase was examined by kinase assay using phosphatidylinositol as a substrate. Cell cycle was assessed by flow cytometry after staining with propidium iodide. RESULTS: EGCG at non-cytotoxic concentrations inhibited PDGF-induced proliferation and migration. This effect was associated with the inhibition of cell cycle progression beyond the G1 phase, decreased cyclin D1 and increased p27Kip1 expression. EGCG inhibited tyrosine phosphorylation of PDGF beta-receptor and downstream activation of extracellular signal-regulated kinase and phosphatidylinositol 3-kinase/Akt pathways. CONCLUSION: EGCG inhibited PDGF-BB-induced proliferation and migration of PSCs through the inhibition of PDGF-mediated signaling pathways.

Activation of JAK-STAT pathway is required for platelet-derived growth factor-induced proliferation of pancreatic stellate cells.

AIM: To clarify the role of Janus kinase-signal transducers and activators of transcription (JAK-STAT) pathway in platelet-derived growth factor (PDGF) induced proliferation in activated pancreatic stellate cells (PSCs). METHODS: PSCs were isolated from rat pancreas tissue, and used in their culture-activated, myofibroblast-like phenotype. STAT-specific binding activity was assessed by electrophoretic mobility shift assay. Activation of Src, JAK2, STAT1, STAT3, and ERK was determined by Western blotting using anti-phospho-specific antibodies. Cell proliferation was assessed by measuring the incorporation of 5-bromo-2'-deoxyuridine. RESULTS: PDGF-BB induced STAT-specific binding activity, and activation of Src, JAK2, STAT1, STAT3, and ERK. Ethanol and acetaldehyde at clinically relevant concentrations decreased basal activation of JAK2 and STAT3. PDGF-induced activation of STAT1 and STAT3 was inhibited by a Src inhibitor PP1 and a JAK2 inhibitor AG490, whereas PDGF-induced activation of ERK was inhibited by PP1, and not by AG490. PDGF-induced proliferation was inhibited by PP1 and AG490 as well as by STAT3 antisense oligonucleotide. CONCLUSION: PDGF-BB activated JAK2-STAT pathway via Src-dependent mechanism. Activation of JAK2-STAT3 pathway, in addition to ERK, may play a role in PDGF-induced proliferation of PSCs.

Protease-activated receptor-2-mediated proliferation and collagen production of rat pancreatic stellate cells.

Activated pancreatic stellate cells (PSCs) play a pivotal role in the pathogenesis of pancreatic inflammation and fibrosis. Trypsin and tryptase, which are agonists for protease-activated receptor-2 (PAR-2), are involved in the pathogenesis of pancreatitis. Here, we examined whether PSCs expressed PAR-2 and its agonists affect the cell functions of PSCs. PSCs were isolated from rat pancreas tissue. Expression of PAR-2 was examined by Western blotting and reverse transcription-polymerase chain reaction. Trypsin, activating peptide (SLIGRL-NH(2), corresponding to the PAR-2 tethered ligand), and tryptase were tested for their ability to affect proliferation, chemokine production, and collagen synthesis in culture-activated PSCs. Activation of mitogen-activated protein (MAP) kinases was assessed by Western blotting using antiphosphospecific antibodies. The effect of PAR-2 agonists on the activation of freshly isolated PSCs in culture was also examined. PAR-2 expression was observed in culture-activated PSCs, whereas it was undetectable in freshly isolated PSCs. PAR-2 agonists activated activator protein-1 and MAP kinases (extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 MAP kinase) but not nuclear factor kappaB. PAR-2 agonists induced proliferation of PSCs through the activation of extracellular signal-regulated kinase. PAR-2 agonists increased collagen synthesis through the activation of c-Jun N-terminal kinase and p38 MAP kinase. PAR-2 agonists did not induce the production of monocyte chemoattractant protein-1 and cytokine-induced neutrophil chemoattractant-1 or initiate the transformation of freshly isolated PSCs in culture. Taken together, our results suggest a role of PAR-2 in the sustenance of pancreatic fibrosis through the increased proliferation and collagen production in PSCs.

High sustained virologic response rate after interferon monotherapy in Japanese hepatitis C patients with a low HCV RNA titer and/or HCV genotype 2. A prospective study.

OBJECTIVE: Hepatitis C virus (HCV) RNA titer and HCV genotype are considered to be major determinants of the outcome of interferon monotherapy. To clarify whether interferon monotherapy is really effective in patients with the appropriate viral parameters, we prospectively examined these parameters and treated the patients with interferon monotherapy. METHODS: Sixty-four patients with an HCV RNA titer <100 kIU/ml and/or HCV genotype 2 were enrolled in the study. Eighteen patients with an HCV RNA titer >100 kIU/ml and genotype 1 were also enrolled as controls. All patients were treated with 10 megaunits of interferon-alpha2b every day for 2 weeks and then 3 times a week for 24 weeks. RESULTS: Of the 64 patients with either HCV RNA <100 kIU/ml and/or genotype 2, seven dropped out from the study. Of the remaining 57 who completed the treatment, 48 (84%) showed a virologic sustained response. In contrast, only 4 of the 18 patients (22%) with HCV RNA >100 kIU/ml and genotype 1 were virologic sustained responders (p < 0.001). CONCLUSION: Our current study showed that the patients with HCV RNA <100 kIU/ml and/or HCV genotype 2 are good candidates for interferon monotherapy.

4-hydroxy-2, 3-nonenal activates activator protein-1 and mitogen-activated protein kinases in rat pancreatic stellate cells.

AIM: Activated pancreatic stellate cells (PSCs) are implicated in the pathogenesis of pancreatic inflammation and fibrosis, where oxidative stress is thought to play a key role. 4-hydroxy-2,3-nonenal (HNE) is generated endogenously during the process of lipid peroxidation, and has been accepted as a mediator of oxidative stress. The aim of this study was to clarify the effects of HNE on the activation of signal transduction pathways and cellular functions in PSCs. METHODS: PSCs were isolated from the pancreas of male Wistar rats after perfusion with collagenase P, and used in their culture-activated, myofibroblast-like phenotype unless otherwise stated. PSCs were treated with physiologically relevant and non-cytotoxic concentrations (up to 5 micromol/L) of HNE. Activation of transcription factors was examined by electrophoretic mobility shift assay and luciferase assay. Activation of mitogen-activated protein (MAP) kinases was assessed by Western blotting using anti-phosphospecific antibodies. Cell proliferation was assessed by measuring the incorporation of 5-bromo-2'-deoxyuridine. Production of type I collagen and monocyte chemoattractant protein-1 was determined by enzyme-linked immunosorbent assay. The effect of HNE on the transformation of freshly isolated PSCs in culture was also assessed. RESULTS: HNE activated activator protein-1, but not nuclear factor kappaB. In addition, HNE activated three classes of MAP kinases: extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 MAP kinase. HNE increased type I collagen production through the activation of p38 MAP kinase and c-Jun N-terminal kinase. HNE did not alter the proliferation, or monocyte chemoattractant protein-1 production. HNE did not initiate the transformation of freshly isolated PSCs to myofibroblast-like phenotype. CONCLUSION: Specific activation of these signal transduction pathways and altered cell functions such as collagen production by HNE may play a role in the pathogenesis of pancreatic disorders.