Showing papers by "Masashi Mizokami published in 2019"

Effects of bowel preparation on the human gut microbiome and metabolome

Abstract: Large bowel preparation may cause a substantial change in the gut microbiota and metabolites. Here, we included a bowel prep group and a no-procedure control group and evaluated the effects of bowel prep on the stability of the gut microbiome and metabolome as well as on recovery. Gut microbiota and metabolome compositions were analyzed by 16S rRNA sequencing and capillary electrophoresis time-of-flight mass spectrometry, respectively. Analysis of coefficients at the genus and species level and weighted UniFrac distance showed that, compared with controls, microbiota composition was significantly reduced immediately after the prep but not at 14 days after it. For the gut metabolome profiles, correlation coefficients between before and immediately after the prep were significantly lower than those between before and 14 days after prep and were not significantly different compared with those for between-subject differences. Thirty-two metabolites were significantly changed before and immediately after the prep, but these metabolites recovered within 14 days. In conclusion, bowel preparation has a profound effect on the gut microbiome and metabolome, but the overall composition recovers to baseline within 14 days. To properly conduct studies of the human gut microbiome and metabolome, fecal sampling should be avoided immediately after bowel prep.

Peroxiredoxin 1, a Novel HBx-Interacting Protein, Interacts with Exosome Component 5 and Negatively Regulates Hepatitis B Virus (HBV) Propagation through Degradation of HBV RNA.

Abstract: Hepatitis B virus (HBV) infection is a major risk factor for the development of chronic liver diseases, including cirrhosis and hepatocellular carcinoma (HCC). A growing body of evidence suggests that HBV X protein (HBx) plays a crucial role in viral replication and HCC development. Here, we identified peroxiredoxin 1 (Prdx1), a cellular hydrogen peroxide scavenger, as a novel HBx-interacting protein. Coimmunoprecipitation analysis coupled with site-directed mutagenesis revealed that the region from amino acids 17 to 20 of the HBx, particularly HBx Cys17, is responsible for the interaction with Prdx1. Knockdown of Prdx1 by siRNA significantly increased the levels of intracellular HBV RNA, HBV antigens, and extracellular HBV DNA, whereas knockdown of Prdx1 did not increase the activities of HBV core, enhancer I (Enh1)/X, preS1, and preS2/S promoters. Kinetic analysis of HBV RNA showed that knockdown of Prdx1 inhibited HBV RNA decay, suggesting that Prdx1 reduces HBV RNA levels posttranscriptionally. The RNA coimmunoprecipitation assay revealed that Prdx1 interacted with HBV RNA. The exosome component 5 (Exosc5), a member of the RNA exosome complexes, was coimmunoprecipitated with Prdx1, suggesting its role in regulation of HBV RNA stability. Taken together, these results suggest that Prdx1 and Exosc5 play crucial roles in host defense mechanisms against HBV infection.IMPORTANCE Hepatitis B virus (HBV) infection is a major global health problem. HBx plays important roles in HBV replication and viral carcinogenesis through its interaction with host factors. In this study, we identified Prdx1 as a novel HBx-binding protein. We provide evidence suggesting that Prdx1 promotes HBV RNA decay through interaction with HBV RNA and Exosc5, leading to downregulation of HBV RNA. These results suggest that Prdx1 negatively regulates HBV propagation. Our findings may shed new light on the roles of Prdx1 and Exosc5 in host defense mechanisms in HBV infection.

Systematic review with meta-analysis: extrahepatic manifestations in chronic hepatitis C virus-infected patients in East Asia.

Abstract: Background Although the prevalence of extrahepatic manifestations of chronic hepatitis C virus (HCV) infection has been reported from Western countries, their prevalence in East Asian countries is not well known. Aim To perform a systematic review to quantify the prevalence of selected extrahepatic manifestations of HCV among patients from East Asia. Methods Medline, CINAHL, EMBASE, Cochrane and country-specific databases were reviewed according to standard guidelines for meta-analyses. Only articles with patients from East Asian countries were included. Results After review, 34 articles were selected (Japan = 9; China = 5, Korea = 3, Taiwan = 16, multiple countries = 1) with 646 228 subjects, 66 436 with HCV. Mean age for HCV-infected patients was 56 years (range 41-72 years), 50% were male (range 26%-73%). The pooled prevalence of type 2 diabetes in East Asian HCV patients was 19.0% (95% confidence interval 15.6%-22.9%) (n = 19 studies) with an increased risk when compared to non-HCV: odds ratio (OR) 1.58 (1.28-1.94). The prevalence of chronic kidney disease in HCV was 9.2% (5.0%-16.2%) (n = 7 studies), also with a significantly increased risk: OR=1.98 (1.41-2.77). Pooled prevalence of cardiovascular disease in HCV was 8.6% (3.5%-19.9%) (n = 6 studies), also with an increased risk: OR = 1.55 (1.21-1.98). The prevalence of lichen planus in HCV was 8.9% (3.6%-20.6%) (n = 6 studies) while the prevalence of rheumatoid arthritis was 4.5% (0.6%-25.7%) (n = 4 studies). Conclusions These data show increased risk of developing extrahepatic manifestations in East Asian patients with HCV. Both hepatic and extrahepatic manifestations of the infection should be used to accurately determine the total burden of the disease in the region.

Construction of complete Tupaia belangeri transcriptome database by whole-genome and comprehensive RNA sequencing.

Abstract: The northern tree shrew (Tupaia belangeri) possesses high potential as an animal model of human diseases and biology, given its genetic similarity to primates. Although genetic information on the tree shrew has already been published, some of the entire coding sequences (CDSs) of tree shrew genes remained incomplete, and the reliability of these CDSs remained difficult to determine. To improve the determination of tree shrew CDSs, we performed sequencing of the whole-genome, mRNA, and total RNA and integrated the resulting data. Additionally, we established criteria for the selection of reliable CDSs and annotated these sequences by comparison to the human transcriptome, resulting in the identification of complete CDSs for 12,612 tree shrew genes and yielding a more accurate tree shrew genome database (TupaiaBase: http://tupaiabase.org ). Transcriptome profiles in hepatitis B virus infected tree shrew livers were analyzed for validation. Gene ontology analysis showed enriched transcriptional regulation at 1 day post-infection, namely in the “type I interferon signaling pathway”. Moreover, a negative regulator of type I interferon, SOCS3, was induced. This work, which provides a tree shrew CDS database based on genomic DNA and RNA sequencing, is expected to serve as a powerful tool for further development of the tree shrew model.

Serologic testing of randomly selected children after hepatitis B vaccination: a cross-sectional population-based study in Lao People’s Democratic Republic

Abstract: Population immunity against hepatitis B virus (HBV) in Lao People’s Demographic Republic (PDR) has not been examined since the national HBV vaccination program was started in 2002. Vaccine has been observed to be frozen at times during cold-chain transport in vaccination programs in Lao PDR and other developing countries, which will inactivate the vaccine. Therefore, this study used post-vaccination serologic testing to evaluate the effects of HBV immunization in Lao PDR. A cross-sectional serologic study was conducted among children (age range, 5–9 years) and mothers (15–45 years) who were randomly selected using probability-proportional-to-size sampling from central Lao PDR. Blood samples were collected as dried blood spots (DBS) and analyzed using chemiluminescent microparticle immunoassay to detect anti-hepatitis B surface (HBs) titers. We also evaluated the correlation between anti-HBs levels measured in DBS and serum among healthy healthcare workers in Vientiane. Anti-HBs titers from DBS were strongly correlated with serum levels (correlation coefficient = 0.999) in all 12 healthcare workers evaluated. A linear regression model showed that 10 mIU/mL of serum anti-HBs was equivalent to 3.45 mIU/mL (95% CI: 3.06–3.85) of DBS. Among 911 mother-child pairs tested, 171 children had documentation of vaccination. Of the 147 children who had received ≥3 doses of the hepatitis B vaccine, 1 (0.7%) was positive for anti-HBs. The remaining 24 children received the hepatitis B vaccine only twice, once or no dose. The results showed extremely low positivity for anti-HBs among vaccinated children in central Lao PDR. Therefore, post-vaccination serologic testing is important to evaluate population immunity against HBV infection. DBS testing is a potential low-cost tool to evaluating the effectiveness of HBV vaccination programs.

Virological Factors Associated With the Occurrence of Hepatitis B Virus (HBV) Reactivation in Patients With Resolved HBV Infection Analyzed Through Ultradeep Sequencing.

Abstract: Background Hepatitis B virus reactivation (HBVr) is an important complication of immunosuppressive drug therapy. It can occur via both virological and host factors; however, the underlying mechanisms remain largely unknown. Methods We examined serum samples derived from patients with HBVr and those with acute hepatitis B (AHB). The targeted nucleic acid molecule in hepatitis B virus deoxyribonucleic acid was amplified and analyzed by next-generation sequencing. Results The percentage of patients infected with genotype Bj among the HBVr patients was significantly higher than that in the AHB patients. The frequency of mutation sites in the whole HBV genome, especially in the envelope region, in the HBVr was significantly higher than that in the AHB. The prevalence of the S3N amino acid substitution in the envelope protein and mutations at positions G1896A and G1899A in the precore region were significantly higher in the HBVr compared with AHB. The population of S3N amino acid substitution and nucleotide G1896A and G1899A mutations in each individual showed a similar percentage of occurrence. Conclusions We identified specific virological factors in patients with HBVr through ultradeep sequencing. Our findings could be beneficial for the elucidation of mechanisms underlying HBVr development and for disease control.

Full-Genome Analysis of Hepatitis C Virus in Japanese and Non-Japanese Patients Coinfected With HIV-1 in Tokyo.

Abstract: Background:Acute hepatitis C virus (HCV) infection is increasing among HIV-1–infected individuals in Tokyo. Appropriate clinical management is needed.Setting:To delineate the epidemiological status of HCV transmission, we analyzed stocked plasma samples of HCV/HIV-1–coinfected patients seen at the l

Validity of administrative database detection of previously resolved hepatitis B virus in Japan.

Abstract: The risk of hepatitis B virus (HBV) reactivation has increased owing to advances in the immunosuppressive therapy field. However, the HBV reactivation incidence among patients with previously resolved HBV (prHBV) infection during immunosuppressive therapy for rheumatoid arthritis (RA) remains unclear. The objective of this work is to describe the validity of detecting prHBV infection from administrative data through comparisons with chart abstraction and determine the incidence of HBV reactivation during immunosuppressive therapy for RA in Japan. In this retrospective cohort study, data on selected patients were extracted from administrative claims data. To identify patients with prHBV infection and de novo hepatitis, and HBsAg carriers, we conducted chart abstraction. The incidence rate of de novo hepatitis was 1.23 of 100 person-years. The positive predictive value (PPV) and its 95% confidence interval (CI) of administrative data for the identification of suspected prHBV infections was 85.8% (95% CI: 81.7%-89.3%). This study evaluated the PPV of the algorithm of HBV-DNA testing with immunosuppressive therapy performed four times or more per year for the detection of prHBV infection from administrative data. Additionally, we determined the incidence rate of HBV reactivation among preHBV infections during immunosuppressive therapy for RA to be 1.23 of 100 person-years.

Histopathological factors affecting the extraction of high quality genomic DNA from tissue sections for next-generation sequencing.

Abstract: To enable the widespread application of genomic medicine, the extraction of genomic DNA from thin sections of archived formalin-fixed and paraffin-embedded (FFPE) tissue blocks for next-generation sequencing (NGS) is often necessary. However, there are currently no guidelines available on which specific regions of the microtome sections to use for macrodissection with respect to the histopathological factors observed under microscopic examination. The aim of this study was to clarify the relationship between histopathological factors and DNA quality, and to standardize the macrodissection method for more efficient implementation of NGS. FFPE tissue specimens of 218 patients from the Biomarker Research for Anti-EGFR Monoclonal Antibodies by Comprehensive Cancer Genomics study were used to investigate the relationship between 15 histopathological factors and the quantitative ratio of double-stranded DNA (dsDNA) to total nucleic acids, as well as the ∆ crossing point value of each tissue specimen. Multivariate logistic regression analysis revealed that specimen storage of ≥3 years was negatively associated with dsDNA quality (P=0.0007, OR: 4.30, 95% CI: 1.85-10.04). In contrast, the presence of a mucus pool was positively associated with dsDNA quality (P=0.0308, OR: 0.23, 95% CI: 0.06-0.87). Metastatic tumors and longer specimen storage periods were significantly associated with lower ∆Cp values (P=0.0007, OR: 4.43, 95% CI: 1.87-10.49; and P=0.0003, OR: 5.51, 95% CI: 2.18-13.95, respectively). Therefore, macrodissection should not be performed on specimens exhibiting histopathological factors associated with poor DNA quality. In particular, the use of tissue blocks with a storage period of <3 years allows the extraction of genomic DNA suitable for NGS.

Response to the letter by Dr. Naoya Yamada, and Dr. Koichi Mizuta regarding our manuscript: "Mac-2 binding protein glycan isomer (M2BPGi) is a new serum biomarker for assessing liver fibrosis: more than a biomarker of liver fibrosis".

Abstract: Dear Editor, We thank Drs. Yamada and Mizuta for their interest in our study and for sharing their experience with a patient with biliary atresia (BA). Serum markers of liver fibrosis in patients with BA are extremely valuable, because patients are typically children, and acquiring a liver biopsy may be difficult. Drs. Yamada and Mizuta demonstrated the clinical significance of serum levels of Mac-2 binding protein glycosylation isomer (M2BPGi) for assessing liver fibrosis in patients with BA [1]. They note the dramatic changes in serum levels of M2BPGi in their female infant with BA. During cholangitis, the serum level of M2BPGi was high and decreased to the normal range after treatment. The half-life of M2BPGi in human serum has not been established. Nevertheless, in patients who undergo liver transplantation for end-stage liver cirrhosis, preoperative serum levels of M2BPGi are typically high and decrease to the normal range within 24 h (unpublished data). Therefore, the levels of M2BPGi rapidly decrease if its production is stopped. There, apparently, is no explanation for the elevation of M2BPGi levels during cholangitis. We previously discovered the unique features of M2BPGi as follows: (1) The cut-off values differ, even for the same stage of fibrosis independent of cause. (2) The levels of M2BPGi rapidly decrease after achieving a sustained antiviral response to hepatitis C virus [2]. Recently, Bekki Y et al. [3] found that hepatic stellate cells (HSCs) are the source of M2BPGi in subpopulations of other liver-derived cells such as Kupffer cells, endothelial cells, biliary epithelial cells, and hepatocytes. The unique features of M2BPGi may be explained if it is specifically produced by activated HSCs. Therefore, M2BPGi levels may reflect the extent of liver fibrosis as well as the acceleration of liver fibrosis in certain chronic liver diseases. The mechanism of the activation of HSCs during cholangitis is unknown. Nevertheless, endotoxin present in obstructive jaundice and obstructive jaundice itself severely impair hepatic circulation [4, 5]. Furthermore, refractory and recurrent cholangitis can cause liver fibrosis. HSCs regulate fibrogenesis in the liver and in the hepatic circulation [6, 7]. Together, these findings suggest that HSCs might be activated during cholangitis. Further studies are required to better understand the significance of elevated levels of M2BPGi in patients with BA and the role of HSCs in this process. New clinical methods are available for predicting the activation of HSCs. We agree with the suggestion of Drs. Yamada and Mizuta that the pathophysiology of certain types of liver disease may be revealed by measuring the serum levels of M2BPGi. This reply refers to the comment available at: https://doi.org/10.1007/ s00535-018-1523-6.