Yumi Matsushima

Autoimmune-related pancreatitis is associated with autoantibodies and a Th1/Th2-type cellular immune response

BACKGROUND & AIMS Although autoimmunity may be involved in some cases of pancreatitis, the mechanism is still unknown. To clarify this, we studied serum autoantibodies, subsets of lymphocytes, and the Th1/Th2 balance of cellular immune responses in patients with autoimmune-related pancreatitis (AIP). METHODS Seventeen patients with AIP (8 men and 9 women; age, 53.2 +/- 13.0 years) were studied. Autoantibodies including antilactoferrin (ALF) or carbonic anhydrase II antibody (ACA-II) were examined using the enzyme-linked immunosorbent assay (ELISA) or the indirect fluorescein antibody method. Intracellular cytokines (interferon gamma and interleukin 4) and subtypes of peripheral blood lymphocytes were examined by flow cytometry and ELISA. RESULTS More than one autoantibody was observed in all 17 patients. Serum antinuclear antibody was detected in 13 of 17 patients, ALF antibody in 13, ACA-II antibody in 10, rheumatoid factor in 5, and anti-smooth muscle antibody in 3, but antimitochondrial antibody in none. The serum levels of ACA-II and LF antibody were not correlated. HLA-DR(+)CD8(+) and HLA-DR(+)CD4(+) cells were significantly increased in peripheral blood (P < 0.05). CD4(+) cells producing interferon gamma and the secreted levels were significantly increased compared with those in controls (P < 0.05), but interleukin 4 was not increased. CONCLUSIONS An autoimmune mechanism against CA-II or LF, and Th1-type immune response, may be involved in AIP.

Experimental immune-mediated pancreatitis in neonatally thymectomized mice immunized with carbonic anhydrase II and lactoferrin

We previously reported that autoantibodies against carbonic anhydrase II and lactoferrin are frequently identified in patients with autoimmune-related pancreatitis. To clarify the role of carbonic anhydrase II and lactoferrin, we created animal models of autoimmune pancreatitis by immunizing neonatally thymectomized mice with carbonic anhydrase II and lactoferrin and also by transferring immunized spleen cells to nude mice. Neonatally thymectomized BALB/c mice were immunized with carbonic anhydrase II or lactoferrin followed by three booster injections (n = 10 in each group). We transferred whole, CD4+, or CD8+ spleen cells prepared from immunized neonatally thymectomized mice to nude mice (n = 5 in each group). Gene expression of IFN-γ and IL-4 was investigated using semiquantitative reverse transcription-polymerase chain reaction. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling staining was used to examine apoptosis. In immunized neonatally thymectomized mice, the prevalence of inflammation was significantly higher in the pancreas. Inflammation was present in all mice receiving whole or CD4+ cells. There was no change in any of the mice receiving CD8+ cells or nonimmunized spleen cells. Carbonic anhydrase II or lactoferrin-immunized mice had apoptotic duct cells or acinar cells, respectively. Expression of the IFN-γ gene was up-regulated in each group. Similar findings were observed in the salivary glands and liver. An immunologic mechanism against carbonic anhydrase II or lactoferrin is involved in the pathogenesis of these pancreatitis models, in which the effector cells are Th1-type CD4+ T cells.

Gene expression of keratinocyte and hepatocyte growth factors during the healing of rat gastric mucosal lesions

BACKGROUND & AIMS The factors that stimulate the growth of gastric mucosal cells during gastric mucosal healing are not completely understood. This study was designed to investigate the gene expression of keratinocyte growth factor (KGF) and hepatocyte growth factor (HGF) in the healing of gastric mucosal lesions. METHODS Northern blot analysis and reverse-transcription polymerase chain reaction were used to show HGF and KGF messenger RNA. RESULTS Transcripts of HGF and KGF were not shown in rat gastric mucosal epithelium but were found in submucosal and muscular layers under normal conditions. When acute gastric mucosal lesions were induced by indomethacin treatment, expression of HGF messenger RNA was augmented in submucosal, muscular, or serosal layers, whereas the transcript of KGF was not influenced. When rat gastric mucosal epithelial cell line RGM1 and rat gastric mucosal primary culture cells were incubated with HGF or KGF, their proliferation was enhanced. CONCLUSIONS The results showed increased gene expression of HGF together with constant production of KGF during gastric mucosal healing.

Presence of Prostaglandin EP4 Receptor Gene Expression in a Rat Gastric Mucosal Cell Line

RGM-1 is an epithelial cell line established from gastric mucosa of adult Wistar rats. In this study, we characterized this newly established cell line by Northern blot analysis. We also investigated deoxyribonucleic acid (DNA) synthesis and hexosamine production in RGM-1 by PGE2. Northern blot analysis did not detect any transcript of proton pump, gastrin receptor, histidine decarboxylase, somatostatin and pepsinogen 1, indicating the absence of characteristics of parietal, ECL, D and chief cells in RGM-1 cells. However, this periodic acid-Schiff (PAS)-positive cell line expressed prostaglandin EP4 receptor mRNA but not EP1 and EP3 receptor mRNAs. [3H]-thymidine incorporation into DNA of the cells was not increased by PGE2. In contrast, PGE2 increased hexosamine content in RGM-1 cells. These results suggest that RGM-1 may be a useful model of gastric mucosal cells and that PGE2 plays a role on mucin synthesis in RGM-1 cells possibly via EP4 receptors.

GASTRIN RECEPTOR GENES ARE EXPRESSED IN GASTRIC PARIETAL AND ENTEROCHROMAFFIN-LIKE CELLS OF MASTOMYS NATALENSIS

Although gastric enterochromaffin-like (ECL) carcinoid tumors are known to develop in patients with long-standing hypergastrinemia, the expression of the gastrin receptor gene in ECL cells has not yet been demonstrated. Therefore, this study was designed to examine gastrin receptor gene expression in ECL cells.Mastomys gastric mucosal cells isolated by enzyme dispersion were separated into 10 fractions (F1–10) by centrifugal elutriation. Each fraction was examined histologically to determine whether they contained ECL and/or parietal cells and Northern blot analysis was used to confirm the presence of histidine decarboxylase and H+, K+-ATPase gene expression. ECL cells were found only in fractions 1 and 2, whereas parietal cells were detected in fractions 6–10. Gastrin receptor gene expression was demonstrated in both parietal cell-rich and ECL cell-rich fractions. In addition, the gastrin receptor cDNA sequences obtained from the two of the fractions (F1 and 8) were identical. These results suggest that gastrin receptor genes are expressed in ECL cells as well as in parietal cells and that these receptors are identical.

Erythropoietin Stimulates Proliferation of Rat-Cultured Gastric Mucosal Cells

Most anemic patients with chronic renal failure have gastric mucosal lesions. However, these gastric lesions are often improved after the administration of recombinant human erythropoietin (rHuEPO). We have used the rat gastric mucosal cell line RGM-1, to examine the possibility that rHuEPO might directly stimulate the growth of gastric mucosal cells in vitro. Our results show that rHuEPO dose-dependently increased [3H]thymidine incorporation into RGM-1 cells and their expression of c-myc gene. In addition, 125I-rHuEPO specifically bound to RGM-1 cells, and moreover, erythropoietin receptor gene expression was detected by RT-PCR. We conclude that rHuEPO has a direct growth-promoting effect on RGM-1 cells, suggesting possible usefulness of rHuEPO administration for the treatment of gastric mucosal damage in patients with chronic renal failure.

Relationship between severity and symptoms of reflux oesophagitis in elderly patients in Japan

Since information concerning reflux oesophagitis in the elderly is limited, particularly in Japan, the severity and symptomatic profiles of reflux oesophagitis in elderly patients were investigated. One hundred and nineteen patients with reflux oesophagitis found among 2278 endoscopy cases between 1993 and 1996 were investigated in this study. The patients were divided into two groups, elderly and non‐elderly. The severity of reflux oesophagitis was estimated by the Los Angeles classification. The presence or absence of typical symptoms (heartburn and regurgitation) was determined by interview. Reflux oesophagitis was not only more frequently found in the elderly group, but was more severe than in the non‐elderly. Although the degree of manifestation of typical symptoms was similar between the elderly and the non‐elderly with high‐grade oesophagitis, the elderly patients with mild reflux oesophagitis were less symptomatic than the non‐elderly. Mild reflux oesophagitis in the elderly may be missed due to its rarity of typical reflux symptoms and a substantial number of elderly persons might have subclinical reflux oesophagitis.

Expression of Protooncogene c-kit and Its Ligand Stem Cell Factor (SCF) in Gastric Carcinoma Cell Lines

We examined 13 human gastric carcinoma celllines for the expression of both c-kit and stem cellfactor (SCF). Expression of mRNAs was detected by bothNorthern blot analysis and reversetranscriptase-polymerase chain reaction (RT-PCR), and expression oftranslated proteins was detected by western blotting.Using RT-PCR we confirmed the expression of c-kit infive (ECC12, TMK1, MKN7, GCIY, and HGC27) cell lines. Northern blot analysis showed coexpression ofboth c-kit and SCF in ECC12 and expression of SCF infive other (MKN74, MKN1 OKAJIMA, KATOIII, and TMK1) celllines. SCF stimulated both tyrosine phosphorylation of c-kit and growth of ECC12, whereas it didnot stimulate those of GCIY. The sizes of c-kittranscript and protein in GCIY were slightly smallerthan those of the reported ones, suggesting the presence of a biologically inactive truncated form ofc-kit in GCIY. The present study suggests that c-kit/SCFsystem might play an important role in thecarcinogenesis and tumor growth of ECC12 and that thetruncated form of c-kit in GCIY might not be associatedwith malignant transformation.

Diagnostic accuracy of a new non‐invasive enzyme immunoassay for detecting Helicobacter pylori in stools after eradication therapy

Helicobacter pylori eradication therapy has been commonly performed for patients with peptic ulcer. An inexpensive, reliable, non‐invasive test would be useful for evaluation of the effectiveness of eradication therapy.

Establishment of primary epithelial cell culture from elutriated rat gastric mucosal cells

Proliferating cells in the gastric mucosal epithelium were successfully enriched by counterflow elutriation in a medium-sized cell fraction. When inoculated on culture plates coated with E-C-L cell attachment matrix, these cells differentiated into mucus-producing cells after reaching confluence. Northern blot analysis did not detect any transcript of the proton pump, histidine decarboxylase, somatostatin, or pepsinogen I, indicating the absence of parietal, ECL, D, and chief cells in the confluent monolayer. These mucus-producing cell monolayers that respond to various growth factors may be a suitable model with which to investigate the function of gastric mucus cells in vitro.