Kanazawa University Hospital

Abstract The Tokyo Guidelines 2013 (TG13) for acute cholangitis and cholecystitis were globally disseminated and various clinical studies about the management of acute cholecystitis were reported by many researchers and clinicians from all over the world. The 1 st edition of the Tokyo Guidelines 2007 ( TG 07) was revised in 2013. According to that revision, the TG 13 diagnostic criteria of acute cholecystitis provided better specificity and higher diagnostic accuracy. Thorough our literature search about diagnostic criteria for acute cholecystitis, new and strong evidence that had been released from 2013 to 2017 was not found with serious and important issues about using TG 13 diagnostic criteria of acute cholecystitis. On the other hand, the TG 13 severity grading for acute cholecystitis has been validated in numerous studies. As a result of these reviews, the TG 13 severity grading for acute cholecystitis was significantly associated with parameters including 30‐day overall mortality, length of hospital stay, conversion rates to open surgery, and medical costs. In terms of severity assessment, breakthrough and intensive literature for revising severity grading was not reported. Consequently, TG 13 diagnostic criteria and severity grading were judged from numerous validation studies as useful indicators in clinical practice and adopted as TG 18/ TG 13 diagnostic criteria and severity grading of acute cholecystitis without any modification. Free full articles and mobile app of TG18 are available at: http://www.jshbps.jp/modules/en/index.php?content_id=47 . Related clinical questions and references are also included.

A. Khosroshahi, Z. S. Wallace, J. L. Crowe, T. Akamizu, A. Azumi, M. N. Carruthers, S. T. Chari, E. Della-Torre, L. Frulloni, H. Goto, P. A. Hart, T. Kamisawa, S. Kawa, M. Kawano, M. H. Kim, Y. Kodama, K. Kubota, M. M. Lerch, M. L€ ohr, Y. Masaki, S. Matsui, T. Mimori, S. Nakamura, T. Nakazawa, H. Ohara, K. Okazaki, J. H. Ryu, T. Saeki, N. Schleinitz, A. Shimatsu, T. Shimosegawa, H. Takahashi, M. Takahira, A. Tanaka, M. Topazian, H. Umehara, G. J. Webster, T. E. Witzig, M. Yamamoto, W. Zhang, T. Chiba, and J. H. Stone

Although the diagnostic and severity grading criteria on the 2013 Tokyo Guidelines (TG13) are used worldwide as the primary standard for management of acute cholangitis (AC), they need to be validated through implementation and assessment in actual clinical practice. Here, we conduct a systematic review of the literature to validate the TG13 diagnostic and severity grading criteria for AC and propose TG18 criteria. While there is little evidence evaluating the TG13 criteria, they were validated through a large-scale case series study in Japan and Taiwan. Analyzing big data from this study confirmed that the diagnostic rate of AC based on the TG13 diagnostic criteria was higher than that based on the TG07 criteria, and that 30-day mortality in patients with a higher severity based on the TG13 severity grading criteria was significantly higher. Furthermore, a comparison of patients treated with early or urgent biliary drainage versus patients not treated this way showed no difference in 30-day mortality among patients with Grade I or Grade III AC, but significantly lower 30-day mortality in patients with Grade II AC who were treated with early or urgent biliary drainage. This suggests that the TG13 severity grading criteria can be used to identify Grade II patients whose prognoses may be improved through biliary drainage. The TG13 severity grading criteria may therefore be useful as an indicator for biliary drainage as well as a predictive factor when assessing the patient's prognosis. The TG13 diagnostic and severity grading criteria for AC can provide results quickly, are minimally invasive for the patients, and are inexpensive. We recommend that the TG13 criteria be adopted in the TG18 guidelines and used as standard practice in the clinical setting. Free full articles and mobile app of TG18 are available at: http://www.jshbps.jp/modules/en/index.php?content_id=47. Related clinical questions and references are also included.

UNLABELLED: Immunoglobin G (IgG) 4-related sclerosing pancreatitis and cholangitis (autoimmune pancreato-cholangitis [AIPC]) are recently recognized disease entities characterized by high serum IgG4 concentrations and sclerosing inflammation with numerous IgG4-positive plasma cells, although the underlining immune mechanism remains only speculative. In this study, the immunopathogenesis of AIPC was examined with respect to the production of cytokines in situ and the possible involvement of regulatory T cells (Tregs) using fresh (5 cases) and formalin-fixed (28 cases) specimens of AIPC and related extra-pancreatobiliary lesions. Quantitative real-time polymerase chain reaction revealed that AIPC and extra-pancreatobiliary lesions had significantly higher ratios of interleukin (IL)-4/interferon-gamma (IFN-gamma) (45.8-fold), IL-5/IFN-gamma (18.7-fold), IL-13/interferon (IFN)-gamma (20.7-fold), IL-10/CD4 (45.3-fold), and tumor growth factor (TGF)-beta/CD4 (39.4-fold) than did primary sclerosing cholangitis (PSC) and primary biliary cirrhosis (PBC). Lymphocytes with signals for IL-4 and IL-10 were frequently found in AIPC by in situ hybridization. The expression of Foxp3 messenger RNA, a transcription factor specific for naturally arising CD4(+)CD25(+) Tregs, was significantly increased in AIPC and extra-pancreatobiliary lesions in comparison to PSC and PBC (36.4-fold). Immunohistochemically, CD4(+)CD25(+)Foxp3(+) cells were frequently found in AIPC, while few were found in PSC and other disease controls. Taken together, AIPC could be characterized by the over-production of T helper (Th) 2 and regulatory cytokines. Tregs might be involved in the in situ production of IL-10 and TGF-beta, which could be followed by IgG4 class switching and fibroplasia. CONCLUSION: AIPC is a unique inflammatory disorder characterized by an immune reaction predominantly mediated by Th2 cells and Tregs.

OBJECTIVE: IgG4-related disease (IgG4-RD) can cause fibroinflammatory lesions in nearly any organ. Correlation among clinical, serologic, radiologic, and pathologic data is required for diagnosis. This work was undertaken to develop and validate an international set of classification criteria for IgG4-RD. METHODS: An international multispecialty group of 86 physicians was assembled by the American College of Rheumatology (ACR) and the European League Against Rheumatism (EULAR). Investigators used consensus exercises, existing literature, derivation and validation cohorts of 1,879 subjects (1,086 cases, 793 mimickers), and multicriterion decision analysis to identify, weight, and test potential classification criteria. Two independent validation cohorts were included. RESULTS: A 3-step classification process was developed. First, it must be demonstrated that a potential IgG4-RD case has involvement of at least 1 of 11 possible organs in a manner consistent with IgG4-RD. Second, exclusion criteria consisting of a total of 32 clinical, serologic, radiologic, and pathologic items must be applied; the presence of any of these criteria eliminates the patient from IgG4-RD classification. Third, 8 weighted inclusion criteria domains, addressing clinical findings, serologic results, radiology assessments, and pathology interpretations, are applied. In the first validation cohort, a threshold of 20 points had a specificity of 99.2% (95% confidence interval [95% CI] 97.2-99.8%) and a sensitivity of 85.5% (95% CI 81.9-88.5%). In the second, the specificity was 97.8% (95% CI 93.7-99.2%) and the sensitivity was 82.0% (95% CI 77.0-86.1%). The criteria were shown to have robust test characteristics over a wide range of thresholds. CONCLUSION: ACR/EULAR classification criteria for IgG4-RD have been developed and validated in a large cohort of patients. These criteria demonstrate excellent test performance and should contribute substantially to future clinical, epidemiologic, and basic science investigations.

The Clinical Practice Guidelines for the Management of Hepatocellular Carcinoma proposed by the Japan Society of Hepatology was updated in June 2014 at a consensus meeting of the Liver Cancer Study Group of Japan. Three important items have been updated: the surveillance and diagnostic algorithm, the treatment algorithm, and the definition of transarterial chemoembolization (TACE) failure/refractoriness. The most important update to the diagnostic algorithm is the inclusion of gadolinium-ethoxybenzyl-diethylenetriamine pentaacetic acid-enhanced magnetic resonance imaging as a first line surveillance/diagnostic tool. Another significant update concerns removal of the term "lipiodol" from the definition of TACE failure/refractoriness.

Liver cancer is an aggressive disease with a poor outcome. Several hepatic stem/progenitor markers are useful for isolating a subset of liver cells with stem cell features, known as cancer stem cells (CSCs). These cells are responsible for tumor relapse, metastasis, and chemoresistance. Liver CSCs dictate a hierarchical organization that is shared in both organogenesis and tumorigenesis. An increased understanding of the molecular signaling events that regulate cellular hierarchy and stemness, and success in defining key CSC-specific genes, have opened up new avenues to accelerate the development of novel diagnostic and treatment strategies. This Review highlights recent advances in understanding the pathogenesis of liver CSCs and discusses unanswered questions about the concept of liver CSCs.

A cellular disintegrin and metalloproteinase (ADAM) is a new family of genes with structural homology to the snake venom metalloproteinases and disintegrins. We screened genes which were selectively expressed in the cachexigenic colon 26 adenocarcinoma subline in vivo. It was found that one novel cDNA clone, identified as a cachexigenic tumor selective gene, encodes a cysteine-rich protein which shows a sequence similarity to that of both the snake venom metalloproteinases and thrombospondins. We named this cDNA clone A disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS-1). ADAMTS1 consists of six domains, 1) a pro- and 2) a metalloproteinase, 3) a disintegrin-like, 4) a thrombospondin (TSP) homologous domain containing TSP type I motif, 5) a spacer region, and 6) COOH-terminal TSP submotifs. Unlike other ADAMs, ADAMTS-1 does not possess a transmembrane domain and is a putative secretory protein. Therefore, ADAMTS-1 is a new type of ADAM family protein with TSP type I motifs. We demonstrated that the TSP homologous domain containing the TSP type I motif of ADAMTS-1 is functional for binding to heparin. ADAMTS-1 mRNA could be induced by stimulating colon 26 cells with an inflammatory cytokine, interleukin-1, in vitro. Moreover, intravenous administration of lipopolysaccharide in mice selectively induced ADAMTS-1 mRNA in kidney and heart. These data suggest that ADAM-TS-1 may be a gene whose expression is associated with various inflammatory processes as well as development of cancer cachexia.

Telomerase activation is thought to be a critical step in cellular immortalization and carcinogenesis. The human telomerase catalytic subunit (hTERT) is a rate limiting determinant of the enzymatic activity of human telomerase. In the previous study, we identified the proximal 181 bp core promoter responsible for transcriptional activity of the hTERT gene. To identify the regulatory factors of transcription, transient expression assays were performed using hTERT promoter reporter plasmids. Serial deletion assays of the core promoter revealed that the 5'-region containing the E-box, which binds Myc/Max, as well as the 3'-region containing the GC-box, which binds Sp1, are essential for transactivation. The mutations introduced in the E-box or GC-box significantly decreased transcriptional activity of the promoter. Overexpression of Myc/Max or Sp1 led to significant activation of transcription in a cell type-specific manner, while Mad/Max introduction repressed it. However, the effects of Myc/Max on transactivation were marginal when Sp1 sites were mutated. Western blot analysis using various cell lines revealed a positive correlation between c-Myc and Sp1 expression and transcriptional activity of hTERT. Using fibroblast lineages in different stages of transformation, we found that c-Myc and Sp1 were induced to a dramatic extent when cells overcame replicative senescence and obtained immortal characteristics, in association with telomerase activation. These findings suggest that c-Myc and Sp1 cooperatively function as the major determinants of hTERT expression, and that the switching functions of Myc/Max and Mad/Max might also play roles in telomerase regulation.

Visceral adiposity in obesity causes excessive free fatty acid (FFA) flux into the liver via the portal vein and may cause fatty liver disease and hepatic insulin resistance. However, because animal models of insulin resistance induced by lipid infusion or a high fat diet are complex and may be accompanied by alterations not restricted to the liver, it is difficult to determine the contribution of FFAs to hepatic insulin resistance. Therefore, we treated H4IIEC3 cells, a rat hepatocyte cell line, with a monounsaturated fatty acid (oleate) and a saturated fatty acid (palmitate) to investigate the direct and initial effects of FFAs on hepatocytes. We show that palmitate, but not oleate, inhibited insulin-stimulated tyrosine phosphorylation of insulin receptor substrate 2 and serine phosphorylation of Akt, through c-Jun NH2-terminal kinase (JNK) activation. Among the well established stimuli for JNK activation, reactive oxygen species (ROS) played a causal role in palmitate-induced JNK activation. In addition, etomoxir, an inhibitor of carnitine palmitoyltransferase-1, which is the rate-limiting enzyme in mitochondrial fatty acid β-oxidation, as well as inhibitors of the mitochondrial respiratory chain complex (thenoyltrifluoroacetone and carbonyl cyanide m-chlorophenylhydrazone) decreased palmitate-induced ROS production. Together, our findings in hepatocytes indicate that palmitate inhibited insulin signal transduction through JNK activation and that accelerated β-oxidation of palmitate caused excess electron flux in the mitochondrial respiratory chain, resulting in increased ROS generation. Thus, mitochondria-derived ROS induced by palmitate may be major contributors to JNK activation and cellular insulin resistance. Visceral adiposity in obesity causes excessive free fatty acid (FFA) flux into the liver via the portal vein and may cause fatty liver disease and hepatic insulin resistance. However, because animal models of insulin resistance induced by lipid infusion or a high fat diet are complex and may be accompanied by alterations not restricted to the liver, it is difficult to determine the contribution of FFAs to hepatic insulin resistance. Therefore, we treated H4IIEC3 cells, a rat hepatocyte cell line, with a monounsaturated fatty acid (oleate) and a saturated fatty acid (palmitate) to investigate the direct and initial effects of FFAs on hepatocytes. We show that palmitate, but not oleate, inhibited insulin-stimulated tyrosine phosphorylation of insulin receptor substrate 2 and serine phosphorylation of Akt, through c-Jun NH2-terminal kinase (JNK) activation. Among the well established stimuli for JNK activation, reactive oxygen species (ROS) played a causal role in palmitate-induced JNK activation. In addition, etomoxir, an inhibitor of carnitine palmitoyltransferase-1, which is the rate-limiting enzyme in mitochondrial fatty acid β-oxidation, as well as inhibitors of the mitochondrial respiratory chain complex (thenoyltrifluoroacetone and carbonyl cyanide m-chlorophenylhydrazone) decreased palmitate-induced ROS production. Together, our findings in hepatocytes indicate that palmitate inhibited insulin signal transduction through JNK activation and that accelerated β-oxidation of palmitate caused excess electron flux in the mitochondrial respiratory chain, resulting in increased ROS generation. Thus, mitochondria-derived ROS induced by palmitate may be major contributors to JNK activation and cellular insulin resistance. Insulin is the major hormone that inhibits gluconeogenesis in the liver. Visceral adiposity in obesity causes hepatic steatosis and insulin resistance. In an insulin-resistant state, impaired insulin action allows enhancement of glucose production in the liver, resulting in systemic hyperglycemia (1Saltiel A.R. Kahn C.R. Nature. 2001; 414: 799-806Crossref PubMed Scopus (3870) Google Scholar) and contributing to the development of type 2 diabetes. In addition, we have demonstrated experimentally that insulin resistance accelerated the pathology of steatohepatitis in genetically obese diabetic OLETF rats (2Ota T. Takamura T. Kurita S. Matsuzawa N. Kita Y. Uno M. Akahori H. Misu H. Sakurai M. Zen Y. Nakanuma Y. Kaneko S. Gastroenterology. 2007; 132: 282-293Abstract Full Text Full Text PDF PubMed Scopus (197) Google Scholar). In contrast, lipid-induced oxidative stress caused steatohepatitis and hepatic insulin resistance in mice (3Matsuzawa N. Takamura T. Kurita S. Misu H. Ota T. Ando H. Yokoyama M. Honda M. Zen Y. Nakanuma Y. Miyamoto K. Kaneko S. Hepatology. 2007; 46: 1392-1403Crossref PubMed Scopus (403) Google Scholar). In fact, steatosis of the liver is an independent predictor of insulin resistance in patients with nonalcoholic fatty liver disease (4Sakurai M. Takamura T. Ota T. Ando H. Akahori H. Kaji K. Sasaki M. Nakanuma Y. Miura K. Kaneko S. J. Gastroenterol. 2007; 42: 312-317Crossref PubMed Scopus (46) Google Scholar). It remains unclear whether hepatic steatosis causally contributes to insulin resistance or whether it is merely a resulting pathology. Excessive dietary free fatty acid (FFA) 2The abbreviations used are: FFA, free fatty acid; IRS, insulin receptor substrate; JNK, c-Jun NH2-terminal kinase; ER, endoplasmic reticulum; ROS, reactive oxygen species; NAC, N-acetyl-l-cysteine; H2DCFDA, 2′,7′-dichlorofluorescin diacetate; OXPHOS, oxidative phosphorylation; PVDF, polyvinylidene difluoride.2The abbreviations used are: FFA, free fatty acid; IRS, insulin receptor substrate; JNK, c-Jun NH2-terminal kinase; ER, endoplasmic reticulum; ROS, reactive oxygen species; NAC, N-acetyl-l-cysteine; H2DCFDA, 2′,7′-dichlorofluorescin diacetate; OXPHOS, oxidative phosphorylation; PVDF, polyvinylidene difluoride. flux into the liver via the portal vein may cause fatty liver disease and hepatic insulin resistance. Indeed, elevated plasma FFA concentrations correlate with obesity and decreased target tissue insulin sensitivity (5Boden G. Diabetes. 1997; 46: 3-10Crossref PubMed Scopus (0) Google Scholar). Experimentally, lipid infusion or a high fat diet that increases circulating FFA levels promotes insulin resistance in the liver. Candidate events linking FFA to insulin resistance in vivo are the up-regulation of SREBP-1c (6Ide T. Shimano H. Yahagi N. Matsuzaka T. Nakakuki M. Yamamoto T. Nakagawa Y. Takahashi A. Suzuki H. Sone H. Toyoshima H. Fukamizu A. Yamada N. Nat. Cell Biol. 2004; 6: 351-357Crossref PubMed Scopus (278) Google Scholar), inflammation caused by activation of c-Jun amino-terminal kinase (JNK) (7Hirosumi J. Tuncman G. Chang L. Gorgun C.Z. Uysal K.T. Maeda K. Karin M. Hotamisligil G.S. Nature. 2002; 420: 333-336Crossref PubMed Scopus (2612) Google Scholar) or IKKβ (8Boden G. She P. Mozzoli M. Cheung P. Gumireddy K. Reddy P. Xiang X. Luo Z. Ruderman N. Diabetes. 2005; 54: 3458-3465Crossref PubMed Scopus (413) Google Scholar), endoplasmic reticulum (ER) stress (9Ozcan U. Cao Q. Yilmaz E. Lee A.H. Iwakoshi N.N. Ozdelen E. Tuncman G. Gorgun C. Glimcher L.H. Hotamisligil G.S. Science. 2004; 306: 457-461Crossref PubMed Scopus (2950) Google Scholar), ceramide (10Kim J.K. Fillmore J.J. Chen Y. Yu C. Moore I.K. Pypaert M. Lutz E.P. Kako Y. Velez-Carrasco W. Goldberg I.J. Breslow J.L. Shulman G.I. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 7522-7527Crossref PubMed Scopus (585) Google Scholar, 11Turinsky J. O'Sullivan D.M. Bayly B.P. J. Biol. Chem. 1990; 265: 16880-16885Abstract Full Text PDF PubMed Google Scholar), and TRB3 (12Du K. Herzig S. Kulkarni R.N. Montminy M. Science. 2003; 300: 1574-1577Crossref PubMed Scopus (707) Google Scholar). However, which event is the direct and initial target of FFA in the liver is unclear. Insulin resistance induced by lipid infusion or a high fat diet is complex and may be accompanied by alterations not restricted to the liver, making it difficult to determine the contribution of FFAs to hepatic insulin resistance. For example, hyperinsulinemia and hyperglycemia secondary to the initial event also may contribute to the development of diet-induced insulin resistance in vivo (6Ide T. Shimano H. Yahagi N. Matsuzaka T. Nakakuki M. Yamamoto T. Nakagawa Y. Takahashi A. Suzuki H. Sone H. Toyoshima H. Fukamizu A. Yamada N. Nat. Cell Biol. 2004; 6: 351-357Crossref PubMed Scopus (278) Google Scholar). To address the early event(s) triggering the development of high fat diet- or obesity-induced insulin resistance, we investigated the molecular mechanism(s) underlying the direct action of FFA on hepatocytes to cause insulin resistance in vitro, using the rat hepatocyte cell line H4IIEC3. We found that mitochondria-derived reactive oxygen species (ROS) were a cause of palmitate-induced insulin resistance in hepatocytes. Materials—The antibody against IRS-2 was purchased from Upstate Biotechnology, Inc. (Lake Placid, NY). Antibodies against phosphotyrosine, Akt, phospho-Akt (Ser473), stress-activated protein kinase/JNK, phospho-stress-activated protein kinase/JNK (Thr183/Tyr185), and phospho-GSK (glycogen synthase kinase)-3 (Ser21/9) were purchased from Cell Signaling Technology (Beverly, MA). Antibodies against GSK-3 and phospho-c-Jun were from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA). Insulin from porcine pancreas, sodium palmitate, sodium oleate, myriocin, N-acetyl-l-cysteine, rotenone, thenoyltrifluoroacetone, cyanide m-chlorophenylhydrazone, oxypurinol, etomoxir, and tunicamycin was obtained from Sigma. SP600125 and apocynin were from Calbiochem. dl-α-tocopherol and 2′,7′-dichlorofluorescin diacetate (H2DCFDA) were from Wako (Osaka, Japan). Cell Culture and Fatty Acid Treatment—Studies were performed in the rat hepatoma cell line H4IIEC3, purchased from the American Type Culture Collection (Manassas, VA). Cells were cultured in Dulbecco's modified Eagle's medium (Invitrogen) supplemented with 10% fetal bovine serum (Invitrogen), penicillin (100 units/ml), and streptomycin (0.1 mg/ml; Invitrogen). The cells were cultured at 37 °C in a humidified atmosphere containing 5% CO2, with medium changes three times a week. All studies were conducted using 80–90% confluent cells, which were treated with the indicated concentrations of FFAs in the presence of 2% FFA-free bovine serum albumin (Sigma). Cell Harvest and Western Blot Analysis—H4IIEC3 hepatocytes, grown to 80–90% confluence in 6-well plates, were treated with the indicated reagents for 16 h in Dulbecco's modified Eagle's medium. After treatment, the cells were stimulated with insulin (1 ng/ml) for 15 min. Then the cells were washed with ice-cold phosphate-buffered saline and lysed in buffer containing 20 mm Tris-HCl (pH 7.5), 5 mm EDTA, 1% Nonidet P-40, 2 mm Na3VO4, 100 mm NaF, and a protease inhibitor mixture (Sigma). After sonication with a Bioruptor (Cosmo Bio, Tokyo, Japan), the lysates were centrifuged to remove insoluble materials. The supernatants (10 μg/lane) were separated by SDS-PAGE and transferred onto polyvinylidene difluoride membranes (Millipore, Billerica, MA). For detection of phosphotyrosine insulin receptor and phosphotyrosine IRS-2, the supernatants (400 μgof protein) were immunoprecipitated with a phosphotyrosine antibody and protein G beads for 2 h at 4 °C before SDS-PAGE. The membranes were blocked in a buffer containing 5% nonfat milk, 50 mm Tris (pH 7.6), 150 mm NaCl, and 0.1% Tween 20 (TBS-T) for 1 h at room temperature. They were then incubated with specific primary antibodies and subsequently with horseradish peroxidase-linked secondary antibodies. Signals were detected with a chemiluminescence detection system (ECL Plus Western blotting detection reagents; GE Healthcare). Densitometric analysis was conducted directly on the blotted membrane, using a CCD camera system (LAS-3000 Mini; Fujifilm, Tokyo, Japan) and Scion Image software. Quantitative Real Time PCR—Total RNA was extracted from cultured H4IIEC3 hepatocytes using an RNeasy mini kit (Qiagen, Germantown, MD), according to the manufacturer's protocol. The cDNA was synthesized from total RNA (100 ng) using random hexamer primers, N6, and a high capacity cDNA reverse transcription kit (Applied Biosystems, Foster City, CA). Quantitative real time PCR was performed with an ABI Prism 7900HT (Applied Biosystems). The set of specific primers and TaqMan probes in the present study was obtained from Applied Biosystems. The PCR conditions were one cycle at 50 °C for 2 min and 95 °C for 10 min, followed by 40 at 95 °C for 15 and °C for 1 min. of RNA was extracted from H4IIEC3 hepatocytes, and cDNA was synthesized as The cDNA was with a of primers and to the rat The PCR conditions were initial at °C for min, followed by of °C for °C for °C for and a at °C for min. The PCR were separated by of of ROS was detected using the H2DCFDA, according to a T. S. T. Y. M. Nature. PubMed Scopus Google Scholar). H4IIEC3 hepatocytes, grown to confluence in plates, were treated with the indicated reagents in Dulbecco's modified Eagle's medium for After treatment, the cells were washed with phosphate-buffered with 10 H2DCFDA, and incubated for min at 37 The was using a MA). of cellular of containing carbonyl that with to the was using a protein carbonyl kit according to the manufacturer's and as N. Takamura T. Ando H. S. Kurita S. Misu H. Ota T. Yokoyama M. Honda M. Miyamoto K. Kaneko S. Full Text Full Text PDF PubMed Scopus Google Scholar). are as were using were by analysis of All were performed with for Insulin chain fatty were for the palmitate, a saturated fatty and oleate, a monounsaturated fatty To whether FFAs impaired insulin signal transduction in H4IIEC3 hepatocytes, we the of FFAs on insulin-stimulated tyrosine phosphorylation of IRS-2 and serine phosphorylation of and with mm palmitate inhibited insulin-stimulated tyrosine phosphorylation of IRS-2 by in H4IIEC3 of IRS-2, insulin-stimulated serine phosphorylation of and were also inhibited by mm palmitate treatment, by and an insulin-resistant However, the protein levels of total IRS-2, Akt, and GSK-3 were by we that palmitate, but not oleate, impaired insulin-stimulated serine phosphorylation in the hepatoma cell line JNK by to Insulin a stress-activated protein to and at serine T. L. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, G. W. Lee Karin M. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). phosphorylation of tyrosine to a in insulin studies have the role of JNK in insulin resistance in (7Hirosumi J. Tuncman G. Chang L. Gorgun C.Z. Uysal K.T. Maeda K. Karin M. Hotamisligil G.S. Nature. 2002; 420: 333-336Crossref PubMed Scopus (2612) Google Scholar, Y. H. M. T. Y. M. Y. M. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, H. S. J.L. T. J. J. Biol. Chem. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar). Thus, we the of FFAs on JNK activation and in insulin but not oleate, increased JNK and c-Jun and inhibitor of JNK, SP600125 Sasaki W. A. S. Y. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: PubMed Scopus Google Scholar), the palmitate-induced phosphorylation of c-Jun that palmitate To whether palmitate-induced JNK activation cellular insulin resistance, we inhibited the JNK with SP600125 insulin-stimulated serine phosphorylation of and GSK-3 in H4IIEC3 hepatocytes to palmitate that JNK activation by palmitate to palmitate-induced insulin of a JNK inhibitor on palmitate-induced alterations in insulin-stimulated phosphorylation of and GSK-3 in H4IIEC3 hepatocytes. H4IIEC3 cells were incubated in the presence or of palmitate and the JNK inhibitor SP600125 for 16 h to with insulin (1 15 cell lysates were by transferred to a membrane, and with the indicated antibodies. was by are the from of or independent were to the of total or GSK-3 and as the palmitate Image for SREBP-1c and in JNK and Insulin in H4IIEC3 SREBP-1c to a role in diet-induced insulin resistance in (6Ide T. Shimano H. Yahagi N. Matsuzaka T. Nakakuki M. Yamamoto T. Nakagawa Y. Takahashi A. Suzuki H. Sone H. Toyoshima H. Fukamizu A. Yamada N. Nat. Cell Biol. 2004; 6: 351-357Crossref PubMed Scopus (278) Google Scholar) found that high diet-induced hyperglycemia and hyperinsulinemia hepatic of to of IRS-2 at the However, in the present palmitate the of SREBP-1c in H4IIEC3 hepatocytes with the and protein levels of IRS-2 were by Thus, palmitate not to cause insulin resistance in hepatocytes via the SREBP-1c stress is induced in insulin-resistant as obesity and type 2 and in stress to to the of insulin through of JNK (9Ozcan U. Cao Q. Yilmaz E. Lee A.H. Iwakoshi N.N. Ozdelen E. Tuncman G. Gorgun C. Glimcher L.H. Hotamisligil G.S. Science. 2004; 306: 457-461Crossref PubMed Scopus (2950) Google Scholar). excessive FFAs have to stress in E. C. L. T. M. A. PubMed Scopus Google Scholar), we whether palmitate caused stress in H4IIEC3 hepatocytes. stress the of which the transcription of molecular protein) M. Cell Biol. 2002; PubMed Scopus Google Scholar). at mm not the of or the of an used to stress we the of palmitate and tunicamycin on insulin-stimulated signal transduction and JNK activation The of tunicamycin on insulin-stimulated serine phosphorylation of was and not with that of the in JNK by tunicamycin was and not with that of that stress played a role in palmitate-induced JNK activation and cellular insulin resistance in H4IIEC3 hepatocytes. ROS to increased cellular ROS levels are to phosphorylation of JNK H. Y. S. 2004; PubMed Scopus Google Scholar). Indeed, ROS levels are increased in conditions with insulin resistance, as and type 2 S. T. M. M. Yamada Y. Y. M. M. J. 2004; PubMed Scopus Google Scholar). FFAs have to ROS in cells, as cells C. N. PubMed Google Scholar), Y. Ruderman 2005; PubMed Scopus Google Scholar), and S. T. M. M. Yamada Y. Y. M. M. J. 2004; PubMed Scopus Google Scholar). Thus, we that palmitate increased ROS production and JNK, to the impaired insulin To H4IIEC3 hepatocytes were incubated with H2DCFDA, a to ROS, with or was elevated by with mm palmitate for and palmitate induced ROS production with the of protein a of oxidative increased in hepatocytes with cells that palmitate, cause ROS production and oxidative stress in H4IIEC3 hepatocytes. Insulin to whether palmitate-induced ROS a causal role in insulin resistance by whether and also as insulin and palmitate-induced ROS at 10 mm and at mm ROS production by 50 and In with decreased ROS the the insulin-stimulated phosphorylation impaired by at 10 mm and at mm the phosphorylation by 40 and palmitate-induced JNK phosphorylation; at 10 mm and at mm it by and that palmitate increased ROS levels in H4IIEC3 hepatocytes and JNK, resulting in insulin of on palmitate-induced alterations in insulin-stimulated serine phosphorylation of in H4IIEC3 hepatocytes. H4IIEC3 cells were incubated in the presence or of palmitate and for 16 h to with insulin (1 15 cell lysates were by transferred to a membrane, and with the indicated antibodies. was by are the from of or independent were to the of total protein and as the palmitate NAC, N-acetyl-l-cysteine; Image of on palmitate-induced JNK activation in H4IIEC3 hepatocytes. H4IIEC3 cells were incubated in the presence or of palmitate and for 16 cell lysates were by transferred to a membrane, and with the indicated antibodies. was by are the from of independent were to the of total JNK protein and as the palmitate Image ROS in the of ROS induced by palmitate in H4IIEC3 hepatocytes, we the cellular in ROS and ROS production was by rotenone, an inhibitor of mitochondrial respiratory chain complex thenoyltrifluoroacetone, an inhibitor of mitochondrial respiratory chain complex and carbonyl cyanide m-chlorophenylhydrazone, an of oxidative phosphorylation In contrast, ROS production in H4IIEC3 cells was not by an inhibitor of or oxypurinol, an inhibitor of that the mitochondrial respiratory chain is in palmitate-induced ROS in H4IIEC3 hepatocytes. ROS through the Fatty Acid are in the mitochondrial fatty acid β-oxidation which the mitochondrial respiratory chain with of the respiratory chain may cause of to ROS production. Thus, we whether palmitate-induced ROS production was on mitochondrial fatty acid is the rate-limiting enzyme in mitochondrial fatty acid etomoxir, a decreased palmitate-induced ROS by palmitate, but not oleate, increased of the up-regulation may contribute to palmitate-induced ROS because the accelerated β-oxidation cause excessive electron flux in the respiratory In the present we investigated the direct action of fatty on insulin in hepatocytes. The saturated fatty acid palmitate, but not the fatty acid oleate, impaired tyrosine phosphorylation of IRS-2, serine phosphorylation of Akt, and serine phosphorylation of of which are of insulin resistance in cultured H4IIEC3 hepatocytes in vivo findings (6Ide T. Shimano H. Yahagi N. Matsuzaka T. Nakakuki M. Yamamoto T. Nakagawa Y. Takahashi A. Suzuki H. Sone H. Toyoshima H. Fukamizu A. Yamada N. Nat. Cell Biol. 2004; 6: 351-357Crossref PubMed Scopus (278) Google Scholar), the of the SREBP-1c was by palmitate to cultured H4IIEC3 hepatocytes, which is a of a for fatty acid and IRS-2 protein levels were insulin resistance in cells, as H. S. J.L. T. J. J. Biol. Chem. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar) and cells 2003; PubMed Scopus Google Scholar). with the present that FFA inhibits insulin at the of tyrosine phosphorylation of of cell to the findings in H. S. J.L. T. J. J. Biol. Chem. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar) and primary hepatocytes and G. W. Lee Karin M. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar), the activation of JNK, a of the tyrosine phosphorylation of was in tyrosine phosphorylation of IRS-2 in cultured H4IIEC3 hepatocytes. a JNK palmitate-induced of the insulin JNK activation to a major role in the development of palmitate-induced insulin resistance in H4IIEC3 hepatocytes. in vivo findings that JNK is in the liver of an animal of obesity and in which FFA into the liver is elevated (9Ozcan U. Cao Q. Yilmaz E. Lee A.H. Iwakoshi N.N. Ozdelen E. Tuncman G. Gorgun C. Glimcher L.H. Hotamisligil G.S. Science. 2004; 306: 457-461Crossref PubMed Scopus (2950) Google Scholar, Y. H. K. M. K. S. M. Y. M. J. Biol. Chem. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar). The of JNK in liver in hepatic insulin resistance at the of tyrosine and the of a of JNK in the liver accelerated hepatic insulin Y. H. M. T. Y. M. Y. M. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). that JNK is by of cellular Full Text Full Text PDF PubMed Scopus Google Scholar), we for a palmitate and JNK activation in H4IIEC3 hepatocytes. stress was to palmitate-induced insulin resistance in H4IIEC3 hepatocytes, because palmitate caused insulin resistance independent of tunicamycin caused stress insulin we found that palmitate-induced ROS insulin resistance. ROS are one of to have a role in insulin resistance J.L. 2002; PubMed Scopus Google Scholar, N. Nature. PubMed Scopus Google Scholar). ROS reactive as and which are as of mitochondrial oxidative phosphorylation Thus, as a increased mitochondrial flux to increased of ROS M. Nature. 2001; 414: PubMed Scopus Google Scholar, H. J. PubMed Scopus Google Scholar). ROS also be β-oxidation of fatty as a of H. J. PubMed Scopus Google Scholar). ROS be by as S. Gastroenterology. Full Text Full Text PDF PubMed Scopus Google Scholar), present in cells, ROS are an of cellular specific inhibitors of ROS, we mitochondrial as an of palmitate-induced ROS in H4IIEC3 hepatocytes. FFAs mitochondrial with through mitochondrial fatty acid of fatty is in the acid In the of fatty acid β-oxidation and the acid and are and excessive for NAC, a of ROS, in cells However, by was to palmitate-induced insulin resistance. the of and not reverse JNK activation and and palmitate-induced insulin resistance and Therefore, may also be in insulin resistance caused by ceramide is a contributing to palmitate-induced JNK activation. from saturated fatty to JNK and phosphorylation in G. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar, S. A. E. J. 2004; PubMed Scopus Google Scholar, C. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). In our palmitate increased the of ceramide in H4IIEC3 hepatocytes at the the of ceramide was not blocked by a inhibitor of serine at the in ceramide ceramide was not blocked was used in with an inhibitor of ceramide synthase not Therefore, we the that ceramide contributes to palmitate-induced insulin resistance in H4IIEC3 hepatocytes. studies are to the role of the ceramide in palmitate-induced insulin resistance in hepatocytes. In the present etomoxir, an inhibitor of decreased palmitate-induced ROS production. palmitate, but not oleate, increased the of the which may for the in insulin action palmitate and it was that fatty acid may be a in insulin sensitivity C.R. Nat. 2007; PubMed Scopus Google Scholar, H. K. Hotamisligil G.S. Full Text Full Text PDF PubMed Scopus Google Scholar). In we investigated the of on insulin in hepatocytes. palmitate-induced ROS and JNK phosphorylation and palmitate-induced phosphorylation of and T. at the molecular underlying the and of FFAs are In study mitochondrial ROS as a in palmitate-induced hepatic insulin resistance. may excess for mitochondrial OXPHOS, and ROS then the insulin by JNK, tyrosine phosphorylation of IRS-2, and hepatic insulin resistance The that an initial event in high diet-induced or obesity-induced insulin resistance in the liver is mitochondrial ROS which be a In to JNK inhibitors or mitochondrial as cyanide m-chlorophenylhydrazone, may a for by ROS generation. We and and for on Western analysis of with

Recently, attention has been drawn to papillary neoplasm of the pancreatobiliary systems. In the pancreas, the disease entity of intraductal papillary mucinous neoplasm (IPMN-P) is widely recognized. In contrast, the pathological characteristics of biliary papillary tumors, such as biliary papilloma(tosis) and papillary cholangiocarcinoma, have not yet been well documented. In this study, we compared the pathological features and post-operative prognosis among biliary papillary tumors (10 cases of biliary papilloma(tosis) and 22 cases of papillary cholangiocarcinoma), conventional non-papillary cholangiocarcinoma (15 cases), and IPMN-P (31 cases). Macroscopically, all biliary papillary tumors were characterized by the prominent intraductal papillary proliferation, and macroscopic mucin-hypersecretion was seen in 9 of 32 cases (28%). Histologically, biliary papillary tumors consisted of three types of tumor cells (pancreaticobiliary, intestinal and gastric types), whereas only the pancreaticobiliary type was observed in non-papillary cholangiocarcinoma. Immunohistochemically, biliary papillary tumors were characterized by the common expression of MUC2, CDX2 and cytokeratin 20. In addition, biliary papillary tumors could be associated with two types of invasive lesions: tubular adenocarcinoma (9 cases) and mucinous carcinoma (5 cases). Patients with tubular adenocarcinoma had a poor prognosis compared to non-invasive papillary tumor or papillary tumor with mucinous carcinoma. These pathological characteristics and the survival status of biliary papillary tumors were different from those of non-papillary cholangiocarcinoma, and rather closely resembled those of IPMN-P. In conclusion, biliary papillary tumors may be the biliary counterpart (intraductal papillary neoplasm of the bile duct) of IPMN-P.

We pharmacokinetically examined the effect of gamma-butyrobetaine, a precursor of L-carnitine, on the change of fatty acid metabolism in juvenile visceral steatosis (JVS) mice, which have systemic L-carnitine deficiency due to lack of L-carnitine transporter activity. The concentrations of total free fatty acid (FFA), palmitic acid and stearic acid in the liver of JVS mice were significantly higher than those in wild-type mice. After intravenous administration of gamma-butyrobetaine (50 mg kg(-1)), the concentration of L-carnitine in the plasma of JVS mice reached about twice that of the control level and levels in the brain, liver and kidney were also significantly increased, whereas those in wild-type mice hardly changed. Although the plasma concentrations of FFA in both types of mice were unchanged after administration of gamma-butyrobetaine, the concentrations of palmitic acid and stearic acid were significantly decreased. In particular, the liver concentration of FFA in JVS mice was decreased to the wild-type control level, accompanied by significant decreases in long-chain fatty acids, palmitic acid and stearic acid, whereas those in wild-type mice were not changed. These results suggest that gamma-butyrobetaine can be taken up into organs, including the liver, of JVS mice, and transformed to L-carnitine. Consequently, administration of gamma-butyrobetaine may be more useful than that of L-carnitine itself for treatment of primary deficiency of carnitine due to a functional defect of the carnitine transporter.

Chronic sclerosing sialadenitis (CSS) is a cryptogenic tumor-like condition of the salivary gland(s). While immune-mediated processes are suspected in its pathogenesis, and CSS is occasionally reported to be associated with sclerosing pancreatitis, an IgG4-related disease, the exact immunopathologic processes of CSS remain speculative. In this study, we examined the clinicopathologic findings of CSS (12 cases) in comparison with sialolithiasis (8 cases) and Sjogren's syndrome (13 cases), and tried to clarify whether CSS is an IgG4-related disease or not. Submandibular gland(s) were affected in all cases of CSS. CSS cases could be divided into two types: 5 cases were associated with sclerosing lesions in extrasalivary glandular tissue (systemic type), while only salivary gland(s) were affected in the remaining 7 cases (localized type). In the former type, which showed male predominance, bilateral salivary glands were frequently affected, and eosinophilia and elevations of gamma-globulin and IgG in serum were frequently found. Histologically, all cases of CSS showed marked lymphoplasmacytic infiltration admixed with fibrosis and the destruction of glandular lobules. Obliterative phlebitis was found in the affected salivary glands in all cases of CSS. Immunohistochemically, the proportion of IgG4/IgG-positive plasma cells was more than 45% in CSS, while it was less than 5% in controls. The resemblance of the clinicopathologic features of CSS with those of sclerosing pancreatitis suggests the participation of a similar immunopathologic process with IgG4 disturbance in CSS. The abundance of IgG4-positive plasma cells in the lesions would be useful for distinguishing CSS from other forms of sialadenitis.

Aim The use of sodium glucose co‐transporter 2 (SGLT2) inhibitors in patients with type 2 diabetes mellitus (T2DM) and chronic kidney disease (CKD) has been limited, primarily because glycaemic efficacy is dependent on kidney function. We performed a systematic review and meta‐analysis to assess the efficacy and safety of SGLT2 inhibitors in patients with T2DM and CKD, defined as estimated glomerular filtration rate (eGFR) <60 mL/min/1.73 m 2 . Materials and methods We searched MEDLINE, EMBASE and the Cochrane Library until 7 August 2018 and websites of the US, European and Japanese regulatory authorities until 27 July 2018 for data from randomized controlled trials of SGLT2 inhibitors that included reporting of effects on biomarkers, cardiovascular, renal or safety outcomes in individuals with T2DM and CKD. Random effects models and inverse variance weighting were used to calculate relative risks with 95% confidence intervals. Results Data were obtained from 27 studies with up to 7363 participants involved. In patients with T2DM and CKD, SGLT2 inhibitors lowered glycated haemoglobin (−0.29%; 95% CI, −0.39 to −0.19) as well as blood pressure, body weight and albuminuria. SGLT2 inhibition reduced the risk of cardiovascular death, nonfatal myocardial infarction or nonfatal stroke (RR, 0.81; 95% CI, 0.70‐0.94) and heart failure (RR, 0.61; 95% CI, 0.48‐0.78), without a clear effect on all‐cause mortality (HR, 0.86; 95% CI, 0.73‐1.01). These agents also attenuated the annual decline in eGFR slope (placebo‐subtracted difference of 1.35 mL/1.73 m 2 /y; 95% CI, 0.78‐1.93) and reduced the risk of the composite renal outcome (HR, 0.71; 95% CI, 0.53‐0.95). There was no evidence of additional risks with SGLT2 inhibition in CKD beyond those already known for the class, although heterogeneity was observed across individual agents for some safety outcomes. Conclusion Currently available data suggest that, despite only modest reductions in glycated haemoglobin, SGLT2 inhibitors reduce the risk of cardiovascular and renal outcomes in patients with T2DM and CKD, without clear evidence of additional safety concerns.

Abstract Cases of non–small-cell lung cancer (NSCLC) carrying the somatic mutation of epidermal growth factor receptor (EGFR) have been shown to be hyperresponsive to the EGFR tyrosine kinase inhibitor gefitinib (IRESSA). If EGFR mutations can be observed in serum DNA, this could serve as a noninvasive source of information on the genotype of the original tumor cells that could influence treatment and the ability to predict patient response to gefitinib. Serum genomic DNA was obtained from Japanese patients with NSCLC before first-line gefitinib monotherapy. Scorpion Amplified Refractory Mutation System technology was used to detect EGFR mutations. Wild-type EGFR was detected in all of the 27 serum samples. EGFR mutations were detected in 13 of 27 (48.1%) patients and two major EGFR mutations were identified (E746_A750del and L858R). The EGFR mutations were seen significantly more frequently in patients with a partial response than in patients with stable disease or progressive disease (P = 0.046, Fisher's exact test). The median progression-free survival was significantly longer in patients with EGFR mutations than in patients without EGFR mutations (200 versus 46 days; P = 0.005, log-rank test). The median survival was 611 days in patients with EGFR mutations and 232 days in patients without EGFR mutations (P > 0.05). In pairs of tumor and serum samples obtained from 11 patients, the EGFR mutation status in the tumors was consistent with those in the serum of 8 of 11 (72.7%) of the paired samples. Thus, EGFR mutations were detectable using Scorpion Amplified Refractory Mutation System technology in serum DNA from patients with NSCLC. These results suggest that patients with EGFR mutations seem to have better outcomes with gefitinib treatment, in terms of progression-free survival, overall survival, and response, than those patients without EGFR mutations.

We investigated the clinical significance of a minor population of paroxysmal nocturnal hemoglobinuria (PNH)-type blood cells in patients with acquired aplastic anemia (AA). We quantified CD55-CD59- granulocytes and red blood cells (RBCs) in peripheral blood from 122 patients with recently diagnosed AA and correlated numbers of PNH-type cells and responses to immunosuppressive therapy (IST). Flow cytometry detected 0.005% to 23.1% of GPI-AP- cells in 68% of patients with AA. Sixty-eight of 83 (91%) patients with an increased proportion of PNH-type cells (PNH+) responded to antithymocyte globulin (ATG) + cyclosporin (CsA) therapy, whereas 18 of 39 (48%) without such an increase (PNH-) responded. Failure-free survival rates were significantly higher (64%) among patients with PNH+ than patients with PNH- (12%) at 5 years, although overall survival rates were comparable between the groups. Numbers of PNH-type and normal-type cells increased in parallel among most patients with PNH+ who responded to IST, suggesting that these cells are equally sensitive to immune attack. These results indicate that a minor population of PNH-type cells represents a reliable marker of a positive IST response and a favorable prognosis among patients with AA. Furthermore, immune attack against hematopoietic stem cells that allows PNH clonal expansion might occur only at the onset of AA.

Management bundles that define items or procedures strongly recommended in clinical practice have been used in many guidelines in recent years. Application of these bundles facilitates the adaptation of guidelines and helps improve the prognosis of target diseases. In Tokyo Guidelines 2013 (TG13), we proposed management bundles for acute cholangitis and cholecystitis. Here, in Tokyo Guidelines 2018 (TG18), we redefine the management bundles for acute cholangitis and cholecystitis. Critical parts of the bundles in TG18 include the diagnostic process, severity assessment, transfer of patients if necessary, and therapeutic approach at each time point. Observance of these items and procedures should improve the prognosis of acute cholangitis and cholecystitis. Studies are now needed to evaluate the dissemination of these TG18 bundles and their effectiveness. Free full articles and mobile app of TG18 are available at: http://www.jshbps.jp/modules/en/index.php?content_id=47. Related clinical questions and references are also included.

C-C motif chemokine receptor (CCR)2 and its ligand, monocyte chemoattractant protein (MCP)-1, are pivotal for adipose tissue macrophage (ATM) recruitment and the development of insulin resistance. However, other chemokine systems also may play a role in these processes. In this study, we investigated the role of CCR5 in obesity-induced adipose tissue inflammation and insulin resistance. We analyzed expression levels of CCR5 and its ligands in white adipose tissue (WAT) of genetically (ob/ob) and high-fat (HF) diet-induced obese (DIO) mice. Furthermore, we examined the metabolic phenotype of Ccr5(-/-) mice. CCR5 and its ligands were markedly upregulated in WAT of DIO and ob/ob mice. Fluorescence-activated cell sorter analysis also revealed that DIO mice had a robust increase in CCR5(+) cells within ATMs compared with chow-fed mice. Furthermore, Ccr5(-/-) mice were protected from insulin resistance, glucose intolerance, and hepatic steatosis induced by HF feeding. The effects of loss of CCR5 were related to both reduction of total ATM content and an M2-dominant shift in ATM polarization. It is noteworthy that transplantation of Ccr5(-/-) bone marrow was sufficient to protect against impaired glucose tolerance. CCR5 plays a critical role in ATM recruitment and polarization and subsequent development of insulin resistance.

UNLABELLED: Recent evidence suggests that hepatocellular carcinoma (HCC) is organized by a subset of cells with stem cell features (cancer stem cells; CSCs). CSCs are considered a pivotal target for the eradication of cancer, and liver CSCs have been identified by the use of various stem cell markers. However, little information is known about the expression patterns and characteristics of marker-positive CSCs, hampering the development of personalized CSC-targeted therapy. Here, we show that CSC markers EpCAM and CD90 are independently expressed in liver cancer. In primary HCC, EpCAM+ and CD90+ cells resided distinctively, and gene-expression analysis of sorted cells suggested that EpCAM+ cells had features of epithelial cells, whereas CD90+ cells had those of vascular endothelial cells. Clinicopathological analysis indicated that the presence of EpCAM+ cells was associated with poorly differentiated morphology and high serum alpha-fetoprotein (AFP), whereas the presence of CD90+ cells was associated with a high incidence of distant organ metastasis. Serial xenotransplantation of EpCAM+ /CD90+ cells from primary HCCs in immune-deficient mice revealed rapid growth of EpCAM+ cells in the subcutaneous lesion and a highly metastatic capacity of CD90+ cells in the lung. In cell lines, CD90+ cells showed abundant expression of c-Kit and in vitro chemosensitivity to imatinib mesylate. Furthermore, CD90+ cells enhanced the motility of EpCAM+ cells when cocultured in vitro through the activation of transforming growth factor beta (TGF-β) signaling, whereas imatinib mesylate suppressed TGFB1 expression in CD90+ cells as well as CD90+ cell-induced motility of EpCAM+ cells. CONCLUSION: Our data suggest the discrete nature and potential interaction of EpCAM+ and CD90+ CSCs with specific gene-expression patterns and chemosensitivity to molecular targeted therapy. The presence of distinct CSCs may determine the clinical outcome of HCC.

Multicentric Castleman disease (MCD) describes a heterogeneous group of disorders involving systemic inflammation, characteristic lymph node histopathology, and multi-organ dysfunction because of pathologic hypercytokinemia. Whereas Human Herpes Virus-8 (HHV-8) drives the hypercytokinemia in a cohort of immunocompromised patients, the etiology of HHV-8-negative MCD is idiopathic (iMCD). Recently, a limited series of iMCD cases in Japan sharing a constellation of clinical features, including thrombocytopenia (T), anasarca (A), fever (F), reticulin fibrosis (R), and organomegaly (O) has been described as TAFRO syndrome. Herein, we report clinicopathological findings on 25 patients (14 males and 11 females; 23 Japanese-born and two US-born), the largest TAFRO syndrome case series, including the first report of cases from the USA. The median age of onset was 50 years old (range: 23-72). The frequency of each feature was as follows: thrombocytopenia (21/25), anasarca (24/25), fever (21/25), organomegaly (25/25), and reticulin fibrosis (13/16). These patients frequently demonstrated abdominal pain, elevated serum alkaline phosphatase levels, and acute kidney failure. Surprisingly, none of the cases demonstrated marked hypergammoglobulinemia, which is frequently reported in iMCD. Lymph node biopsies revealed atrophic germinal centers with enlarged nuclei of endothelial cells and proliferation of endothelial venules in interfollicular zone. 23 of 25 cases were treated initially with corticosteroids; 12 patients responded poorly and required further therapy. Three patients died during the observation period (median: 9 months) because of disease progression or infections. TAFRO syndrome is a unique subtype of iMCD that demonstrates characteristic clinicopathological findings. Further study to clarify prognosis, pathophysiology, and appropriate treatment is needed.