Yasuo Matsubara

Pancreas duct abnormalities in patients with ulcerative colitis. A magnetic resonance pancreatography study

Background: The presumed etiology and prevalence of pancreatic abnormalities in patients with ulcerative colitis (UC) have been controversial. We conducted a controlled (cross‐sectional) study of patients with UC compared with non‐UC controls to determine the prevalence of pancreatic duct abnormalities in patients with UC and to determine if these are specific to UC using magnetic resonance cholangiopancreatography. Methods: Magnetic resonance cholangiopancreatography was performed on 79 unselected patients with UC and 45 non‐UC controls, without a history of pancreatitis attack, between February 2000 and May 2003. Results: Among patients with UC, the prevalence of pancreatic duct abnormalities was 16.4% (95% confidence interval, 8.3%‐24.6%): coexisting dilatation and narrowing of the main pancreatic duct were found in 5; diffuse narrowing of pancreatic ducts in 5; and dilatation of pancreatic ductal branches in 3. These abnormalities were compatible with chronic pancreatitis. Among the controls, no pancreatic duct abnormality was found. Conclusions: Changes in the pancreatic duct were found in approximately one‐sixth of patients with UC, none of whom had a history of overt pancreatitis. These pancreatic duct abnormalities are likely to be specific to patients with UC.

Novel chemoimmunotherapeutic strategy for hepatocellular carcinoma based on a genome-wide association study

Pharmacotherapeutic options are limited for hepatocellular carcinoma (HCC). Recently, we identified the anti-tumor ligand MHC class I polypeptide-related sequence A (MICA) gene as a susceptibility gene for hepatitis C virus-induced HCC in a genome-wide association study (GWAS). To prove the concept of HCC immunotherapy based on the results of a GWAS, in the present study, we searched for drugs that could restore MICA expression. A screen of the FDA-approved drug library identified the anti-cancer agent vorinostat as the strongest hit, suggesting histone deacetylase inhibitors (HDACis) as potent candidates. Indeed, the HDACi-induced expression of MICA specific to HCC cells enhanced natural killer (NK) cell-mediated cytotoxicity in co-culture, which was further reinforced by treatment with an inhibitor of MICA sheddase. Similarly augmented anti-tumor activity of NK cells via NK group 2D was observed in vivo. Metabolomics analysis revealed HDACi-mediated alterations in energy supply and stresses for MICA induction and HCC inhibition, providing a mechanism for the chemoimmunotherapeutic actions. These data are indicative of promising strategies for selective HCC innate immunotherapy.

miR-425 regulates inflammatory cytokine production in CD4+ T cells via N-Ras upregulation in primary biliary cholangitis

BACKGROUND & AIMS Primary biliary cholangitis (PBC) is an autoimmune liver disease of unknown pathogenesis. Consequently, therapeutic targets for PBC have yet to be identified. CD4+ T cells play a pivotal role in immunological dysfunction observed in PBC, and therefore, microRNA (miRNA) and mRNA expression were analysed in CD4+ T cells, to investigate PBC pathogenesis and identify novel therapeutic targets. METHODS Integral miRNA and mRNA analysis of 14 PBC patients and ten healthy controls was carried out using microarray and quantitative real-time polymerase chain reaction (qRT-PCR), with gene set enrichment analysis. The functional analyses of miRNA were then assessed using reporter and miRNA-overexpression assays. RESULTS The integral analysis of miRNA and mRNA identified four significantly downregulated miRNAs (miR-181a, -181b, -374b, and -425) related to the T cell receptor (TCR) signalling pathway in CD4+ T cells of PBC. N-Ras, a regulator of the TCR signalling pathway, was found to be targeted by all four identified miRNAs. In addition, in vitro assays confirmed that decreased miR-425 strongly induced inflammatory cytokines (interleukin [IL]-2 and interferon [IFN]-γ) via N-Ras upregulation in the TCR signalling pathway. CONCLUSION The decreased expression of four miRNAs that dysregulate TCR signalling in PBC CD4+ T cells was identified. miR-425 was demonstrated as an inflammatory regulator of PBC via N-Ras upregulation. Therefore, the restoration of decreased miR-425 or the suppression of N-Ras may be a promising immunotherapeutic strategy against PBC. LAY SUMMARY Primary biliary cholangitis (PBC) is an autoimmune liver disease, but the causes are unknown. MicroRNAs are molecules known to regulate biological signals. In this study, four microRNAs were identified as being decreased in PBC patients, leading to activation of T cell receptor signalling pathways, involved in inflammation. One particular target, N-Ras, could be an attractive and novel immunotherapeutic option for PBC. TRANSCRIPT PROFILING Microarray data are deposited in GEO (GEO accession: GSE93172).

Cowden's disease with a defined genetic alteration-chromosomal duplication at 15q11-q13

Cowden’s disease, multiple hamartoma syndrome, is a dominantly inherited disorder characterized by multiple hamartomas of ectodermal, endodermal, and mesodermal origin and also by a high incidence of malignant tumors. Despite many efforts to identify the genetic alterations responsible for the syndrome, the molecular mechanism remains unclear. We report a case of Cowden’s disease in which karyotype analysis revealed a small duplication (about 1 Mb) at 15q11–q13. This part of the genome is a region that is deleted in the Prader-Willi/Angelman syndrome and is a “hot spot” of chromosomal duplication.

Predominance of regorafenib over sorafenib: restoration of membrane-bound MICA in hepatocellular carcinoma cells

The multi‐kinase inhibitor regorafenib (REG) was recently demonstrated to be effective in patients with sorafenib (SOR)‐resistant hepatocellular carcinoma (HCC). Interestingly, SOR is known to enhance the accumulation of membrane‐bound MHC class I polypeptide‐related sequence A (mMICA) in HCC cells and to block the production of soluble MICA (sMICA), an immunological decoy. In addition, MICA is associated with HCC in patients with chronic hepatitis C. We have now compared the impact of REG and SOR on MICA in HCC cells, as well as the immunotherapeutic implications thereof.

Enzymatic inhibition of MICA sheddase ADAM17 by lomofungin in hepatocellular carcinoma cells: Lomofungin suppresses ADAM17 in hepatocellular carcinoma cells

In our previous study on hepatocellular carcinoma (HCC) susceptibility genes in chronic hepatitis patients, we identified the MHC class I polypeptide‐related sequence A (MICA). Natural killer cells eliminate various cancer cells, including HCC, by suppressing MICA shedding. Therefore, we investigated MICA sheddases and inhibitors for HCC immunotherapy. In this study, HepG2, PLC/PRF/5, and Hep3B were treated with the siRNA of a disintegrin and metalloproteases (ADAMs) and matrix metalloproteases to measure the concentration of soluble MICA (sMICA) by ELISA to detect the therapeutic target. Furthermore, an FDA‐approved drug library was tested for the enzymatic inhibition of the targeted enzyme in an in vitro drug screening assay system. ADAM17 knockdown reduced sMICA levels and increased membrane‐bound MICA (mMICA) expression in HCC cells. In an in vitro drug screen using an FDA‐approved drug library, lomofungin, an antifungal drug, was found to strongly decrease ADAM17 activity. In HCC cells, mMICA expression was induced and sMICA production was inhibited in a dose‐dependent manner. These effects were cancelled upon ADAM17 knockdown, suggesting that lomofungin targeted ADAM17. Analysis of lomofungin analogs revealed the responsible functional groups. In summary, we suggest lomofungin to be an attractive agent for the immunological control of HCC, via the suppression of ADAM17.

Tu1830 Specifically Expressed MicroRNAs in CD4+ T Cells Participate With the Pathogenesis of Primary Biliary Cirrhosis to Regulate T Cell Signaling Pathway

A S L D A b st ra ct s 1.Circulating Th17 cells were elevated in PBC compared to HC(P<0.0001), and higher than those in chronic hepatic B(CHB) (P<0.0001). IL-17-positive cells that infiltrated the liver were higher in PBC compared to HC. 2.The progenitor of Th17 cells, CD4+CD161+ cells is increased in PBC(P<0.0001). And stimulated with IL-23 and IL-1 β, they expressed more IL-17(P=0.0556).The serum IL-1β and IL-23 were higher in PBC(P=0.023). 3.Early PBC presented with more Th17 cells in periphery blood than advanced PBC(P=0.0351). Whereas, IL-17 was higher in the liver in advanced PBC. The serum CCL20 levels were higher in PBC patients(P=0.001), especially in advanced disease(P=0.001). The CCL20 levels were more obvious in liver in PBC. Whereas, CD4+CCR6+ cells were not statistically different between PBC and HC(P=0.975). 4.IL-17 can promote the proliferation of HSCs in a dose dependent way(P<0.001), and increase the IL-8 expression in both dose and time dependent way(P<0.001). However, α-SMA production may not be influenced by IL-17(P=0.0847). Anti-IL-17 neutralizes above reactions. Conclusions: 1.Compared to HC and CHB, the levels of circulating Th17 cells and IL-17-positive cells infiltrated liver are higher in PBC. 2.CD4+CD161+ cells produce more IL-17 after stimulated with IL-23 and IL-1 βin PBC, therefore CD4+CD161+ cells are a source of increased Th17 cells in PBC. 3.The increased Th17 population is greater in circulation and less in liver in early PBC patients. Enhanced serum and hepatic CCL20 induce Th17 cells to migrate to liver, therefore Th17 cells are lower in circulation and enhanced in liver in advanced patients. 4.IL-17 can promote proliferation of HSCs in a dose dependent way and increase the IL-8 expression in both dose and time dependent way. Therefore, anti-IL-17 is a promising therapy for PBC fibrosis.

Hepatocyte growth factor region-specifically stimulates gastro-intestinal epithelial growth in primary culture

Hepatocyte growth factor (HGF) stimulated the growth of fetal rat gastrointestinal epithelial cells in primary culture with a clear dose-response relationship. The epithelial response to the mitogenic activity of HGF was different among the region of the gastro-intestinal tract; glandular stomach most responsive, followed by intestine and forestomach. The interaction of HGF with other growth factors in inducing the epithelial growth was also different depending on the type of the epithelial cells, indicating a region-specific growth regulation in the gastro-intestinal tract. Analyses using Northern blot and RT-PCR revealed that HGF mRNA was expressed only in mesenchymes but not in epithelia of the gastro-intestinal tract while expression of c-met (HGF receptor) gene was observed in both tissues. These results suggest that gastro-intestinal mesenchymes secrete HGF which stimulates the growth of attaching epithelial cells by a paracrine mechanism, and that the epithelial response to HGF is controlled by a region-specific growth regulatory mechanism.