伊藤 弘康

Atazanavir sulfate, an azapeptide inhibitor of HIV protease, has been associated with urolithiasis. A 60-year-old man with atazanavir-induced urinary sediment crystals verified by infrared spectroscopic analysis is described. He had been receiving highly active antiretroviral therapy (HAART) for HIV infection and also had a history of urinary lithiasis and been undergoing urinalysis once every month. Needle-shaped crystals were seen in his urine sediment and infrared spectroscopic analysis revealed that these were atazanavir crystals. Because the presence of the crystals in urine do not always reveal an abnormality in the urinary test strip analysis, the urinary sediment needed to be observed microscopically in order to prevent future urolithiasis and renal failure in this HIV patient receiving atazanavir. (C) 2013 S. Karger AG, Basel

CK-MBは最も一般的な心筋マーカーであり、急性心筋梗塞(acute myocardial infarction;AMI)の診断補助として広く用いられている。本邦のCK-MBの測定方法は、従来免疫阻害による酵素活性測定法や化学発光による蛋白量測定法が主流であったが、今回ラテックス凝集比濁法(LA法)を測定原理とする蛋白量測定試薬CK-MB massの検討を行う機会を得たのでその結果を報告する。今回行った検討では再現性、最小検出感度、希釈直線性など、いずれも良好であり、十分な基本性能を有しているものと考えられる。また、AMI症例において他の心筋関連マーカーとともに時系列にて変動を確認したところ、CK-MBはAMI時の血液中への逸脱は心臓型脂肪酸結合蛋白(H-FABP)などに比べるとやや遅いが、トロポニンと同等の推奨がなされており、他の心筋マーカーと合わせて検査することで、AMIの早期診断に繋がることが期待される。本試薬は良好な基本性能を有し、CK-MBの蛋白量測定をLA法を用いて汎用生化学自動分析装置に適用することで、迅速かつ正確に測定可能であることから、臨床的有用性の高い試薬であると考えられる。(著者抄録)

Indoleamine 2,3-dioxygenase-1 (Ido), which catalyzes the first and limiting step of tryptophan catabolism, has been implicated in immune tolerance. However, the roles of Ido in systemic bacterial infection are complicated and remain controversial. To explore this issue, we examined the roles of Ido in bacterial peritonitis and sepsis after cecal ligation and puncture (CLP) in mice by using the Ido inhibitor 1-methyl-D,L-tryptophan (1-MT), by comparing Ido(+/+) and Ido(-/-) mice, or by using chimeric mice in which Ido in the bone marrow-derived cells was deficient. Ido expression in the peritoneal CD11b(+) cells and its metabolite Lkynurenine in the serum were increased after CLP. 1-MT treatment or Ido deficiency, especially in bone marrow-derived cells, reduced mortality after CLP. Compared to Ido(+/+) mice, Ido(-/-) mice showed increased recruitment of neutrophils and mononuclear cells into the peritoneal cavity and a decreased bacterial count in the blood accompanied by increased CXCL-2 and CXCL-1 mRNA in the peritoneal cells. Ido has an inhibitory effect on LPS-induced CXCL-2 and CXCL-1 production in cultured peritoneal cells. These findings indicate that inhibition of Ido reduces mortality from peritonitis and sepsis after CLP via recruitment of neutrophils and mononuclear cells by chemokine production in peritoneal CD11b(+) cells. Thus, blockade of Ido plays a beneficial role in host protection during bacterial peritonitis and sepsis.

UNLABELLED: Indoleamine 2,3-dioxygenase (IDO), an enzyme that is ubiquitously distributed in mammalian tissues and cells, converts tryptophan to kynurenine, and is also known as a key molecule that promotes apoptosis in lymphocytes and neurons. In this study, we established hepatitis B virus (HBV)-transgenic (Tg)/IDO-knockout (KO) mice and examined the influence of IDO in a murine fulminant hepatitis model induced by HBV-specific cytotoxic T lymphocytes (CTL). An increase of IDO expression in the livers of HBV-Tg/IDO-wild-type (WT) mice administered HBV-specific CTL was confirmed by real-time polymerase chain reaction, western blotting, and evaluating IDO activity. Plasma alanine aminotransferase (ALT) levels in HBV-Tg/IDO-KO mice after HBV-specific CTL injection significantly decreased compared with those in HBV-Tg/IDO-WT mice. An inhibitor of IDO, 1-methyl-d-tryptophan (1-MT), could also attenuated the observed liver injury induced by this HBV-specific CTL. The expression levels of cytokine and chemokine mRNAs in the livers of HBV-Tg/IDO-WT mice were higher than those in the livers of HBV-Tg/IDO-KO mice. The administration of kynurenine aggravated the liver injury in HBV-Tg/IDO-KO mice injected with HBV-specific CTL. Simultaneous injection of recombinant murine interferon (IFN-γ) and kynurenine also increased the ALT levels in HBV-Tg/IDO-KO mice. The liver injury induced by IFN-γ and kynurenine was improved in HBV-Tg/tumor necrosis factor-α-KO mice. CONCLUSION: Kynurenine and IFN-γ induced by the administration with HBV-specific CTL are cooperatively involved in the progression of liver injury in acute hepatitis model. Our results may lead to a new therapy for the acute liver injury caused by HBV infection.

Cytotoxic T lymphocytes (CTLs) are thought to be major effectors involved in viral clearance during acute infections, including hepatitis B virus (HBV) infection. A persistent HBV infection is characterized by a lack of or a weak CTL response to HBV, which may be reflective of tolerance to HBV. Efficient induction of HBV-specific CTLs leads to the clearance of HBV in patients with a chronic HBV infection. Previously, we reported that alpha-galactosylceramide (alpha-GalCer), a specific natural killer T (NKT) cell agonist, enhanced the induction of HBV surface antigen (HBsAg)-specific CTLs. In the present study, we found that inhibition of indoleamine 2,3-dioxygenase (IDO) activity enhanced the induction of HBsAg-specific CTLs after immunization with HBsAg and alpha-GalCer. The administration of HBsAg and alpha-GalCer increased the production of interleukin-2 and interleukin-12b, which are crucial for the induction of HBsAg-specific CTLs. The production of these cytokines was more strongly enhanced in IDO knockout mice compared with wild-type mice. In addition, alpha-GalCer induced the production of IDO in CD11b(+) cells, and these cells inhibited proliferation of HBsAg-specific CTLs. Our results lead to strategies for improving the induction of HBsAg-specific CTLs.

臨床検査データからみた2型糖尿病患者の末梢神経障害についてアルブミン尿とABIの関連性から検討した。末梢神経伝達速度検査の依頼のあった2型糖尿病患者126例を対象とした。合併症は、末梢神経障害73例、腎症35例、網膜症48例、高血圧症74例、大血管障害28例、脂質異常症71例であった。罹病期間はMCV・SCV、腎機能検査のA/CとSCVに有意な負の相関を認めた。動脈硬化の指標であるABIと神経伝達速度間に有意な正の相関を認めた。A/Cで有意な関連性を認めた項目は、罹病期間、BUN、CRE、ABI、後脛骨神経のMCVとLatency、腓腹神経のSCVとLatencyであった。下肢のMCV・SCVはA/Cの上昇に反比例して有意に低下し、Latencyは増加した。末梢神経障害、大血管障害において有意な関連性を認めた。

マトリックスメタロプロテイナーゼ-3(matrix metalloproteinase-3;MMP-3)は関節リウマチ(rheumatoid arthritis;RA)の予後予測因子や治療効果判定に広く用いられている。今回我々はラッテクス凝集比濁法を測定原理とする栄研化学社のLZテスト'栄研'MMP-3(Eiken)の検討を行った。その結果、良好な再現性や直線性を確認し、ラテックス凝集比濁法を測定原理とする他社試薬やELISA法とも良好な相関性が得られたことより、十分な基本性能を有していることが考えられる。MMP-3測定値はRA患者群に比べ変形性関節症(OA)患者群で有意に低値であり、また他の自己免疫性疾患患者群でも上昇することを確認した。ステロイド薬の使用群では未使用群に比べ有意に高値となった。さらに、女性の腎機能低下群は正常群に比べ有意に高値であった。MMP-3が高値を示した際には、これらの影響を考慮に入れて判断する必要がある。本試薬は良好な基本性能を有して、汎用測定装置で測定可能であることから、RA診療において臨床的有用性が高いと考えられる。(著者抄録)

超音波検査(US)による肩腱板断裂診断能についてMRI所見と比較検討した。関節鏡視下手術を行い、術前のUS、MRI結果を確認できた70例を対象とした。腱板断裂(RCT)は48例で、完全断裂44例、不完全断裂4例であった。RCT陰性22例では、Bankrt損傷2例、石灰性腱炎6例、上方関節唇損傷2例、拘縮肩2例、脱臼7例、変形性関節症2例、傍関節唇嚢胞1例であった。術中所見Gold standardとすると、USでは感度95.8%、特異度90.9%、正診率94.3%、MRIでは感度97.9%、特異度86.4%、正診率94.3%で、両者に有意差は認めなかった。上腕二頭筋長頭腱所見と腱板断裂サイズおよび肩蜂下滑液包内水腫や腱板付着面の不整と腱板断裂サイズに有意な関連性を認めた。最も関連性の高かった所見は腱板表面の性状で、断裂サイズの増大に伴い、凹凸・平坦化や欠損所見の出現頻度が増加した。

Hepatic immune regulation is associated with the progression from simple steatosis to non-alcoholic steatohepatitis, a severe condition of inflamed fatty liver. Indoleamine 2,3-dioxygenase (IDO), an intracellular enzyme that mediates the catabolism of L-tryptophan to L-kynurenine, plays an important role in hepatic immune regulation. In the present study, we examined the effects of IDO gene silencing on high-fat diet (HFD)-induced liver inflammation and fibrosis in mice. After being fed a HFD for 26 weeks, the IDO-knockout (KO) mice showed a marked infiltration of inflammatory cells, especially macrophages and T lymphocytes, in the liver. The expression levels of F4/80, IFN gamma, IL-1 beta, and IL-6 mRNA in the liver and the expression levels of F4/80 and TNF-alpha mRNA in the white adipose tissue were significantly increased in IDO-KO mice, although hepatic steatosis, the accumulation of intrahepatic triglycerides, and the amount of oxidative stress were lower than those in IDO-wild-type mice. IDO-KO mice also developed marked pericellular fibrosis in the liver, accumulated hepatic hydroxyproline, and exhibited increased expression levels of hepatic TGF-beta 1 mRNA. These findings suggest that IDO-KO renders the mice more susceptible to HFD-induced hepatic inflammation and fibrosis. Therefore, IDO may have a protective effect against hepatic fibrosis, at least in this HFD-induced liver injury model.

ミュータスワコーブラームスPCT(以下μTAS)およびブラームスPCT-Q(以下PCT-Q)の基礎的検討とPCTの有用性評価を行い、μTASは良好な基本性能を有していることを確認した。μTASとPCT-Qの一致率は75.4%であった。PCT-Qは簡便に測定可能であるが、カットオフ値付近での不一致検体の他にRFや他のグロブリンによるPCT-Qの非特異的反応が原因と推測される不一致検体が存在した。敗血症患者の他の炎症マーカーと同時にPCT値を時系列で観察し、敗血症のマーカーとして有用であることを確認した。また、検出される菌種によりPCTのピーク値が異なることが推測され、高感度測定が可能なμTASの測定意義が高いことがわかった。(著者抄録)

Chronic kidney disease (CKD) significantly contributes to the increased number of dialysis patients with end stage renal disease. Anew CKD risk classification (KDIGO 2009) established in 2011, which is defined by albuminuria and estimated glomerular filtration rate (eGFR) values, demonstrates the relative risks of CKD in great detail. In this study, we evaluated the clinical significance of urinary casts by categorizing a risk Group 1 to 5 according to the KDIGO 2009 classification. In the high risk CKD group (risk group 3 and over), we found a significantly higher number of patients who had>100 hyaline casts/whole field (WF) in their urine than those that had<100 hyaline casts/WF. Further, we determined the diagnostic accuracy for the high risk CKD group when the cutoff value for the number of hyaline casts was set at>or = 100 hyaline casts/WF (sensitivity: 44.7%, specificity: 96.5%). The eGFR value was significantly lower in the group with>or = 100 hyaline casts/WF, particularly in hypertensive patients, than that in the group with<100 hyaline casts/ WF. Of interest is that the eGFR value was significantly lower in patients with 100-999 hyaline casts/WF and>or = 1,000 hyaline cas

前立腺癌の診断には前立腺特異抗原(Prostate specific antigen:PSA)の測定が有効な補助診断として用いられており,特異抗体も作成されて多く測定系試薬が確立している.また,近年,標準物質の設定や等モル反応系(Equimolar反応)の測定系への統一がなされ,標準化への試みが進んでいる.今回,新たにシスメックスより発売された全自動免疫化学発光測定機器HISCL-2000i専用試薬「HISCL PSA試薬」の基礎的検討を行ったので報告する.HISCL PSA試薬の再現性は極めて高性能であり,希釈直線性も良好であった.また,従来法のルミパルス,アーキテクトとの相関は両者とも良好であり,最小検出感度は0.0006ng/mLと高感度を有している.必要検体量は10μLと微量であることから,日常の健診検体においても有用性が高い試薬であると考えられる.(著者抄録)

Vα14 natural killer T (iNKT) cells activated by alpha- galactosylceramide (GalCer) secrete a large amount of cytokines. Toll-like receptors (TLRs) play a critical role in the innate immune responses via the recognition of pathological antigen. Previously we demonstrated that the iNKT cells activated by GalCer augmented LPS-induced NO production in peritoneal cells. In this study, we examined the effect of GalCer and TLR agonists by IFN-γ production from splenocytes. Splenocytes pretreated with GalCer induced TLR3, 4, 7/8, and 9 agonists in vitro, resulting in the enhancement of IFN-γ mRNA expression. In particular, IFN-γ stimulated by GalCer and LPS was increased in NK cells and CD8 T cells, and inhibited by a neutralizing anti-IL-12 antibody. Pretreatment with GalCer enhanced the phosphorylation of IκB-α induced by LPS stimulation. The present study showed that co-stimulation of GalCer and TLR agonists powerfully induced the production of IFN-γ from splenocytes.

Epidemiologic data suggest that non-alcoholic fatty liver disease (NAFLD) have an increased tendency to occur in patients who are associated with metabolic conditions such as obesity, type 2 diabetes mellitus, and dyslipidemia. NAFLD represents a wide spectrum of conditions ranging from fatty liver, which in general follows a benign, no-progressive clinical course, to non-alcoholic steatohepatitis (NASH), a more serious form of NAFLD that may progress to cirrhosis and end stage liver disease. However, currently the diagnosis of NASH requires an invasive liver biopsy. The aim of this study is to evaluate whether Cytokeratin 18 (CK-18) has potential non-invasive diagnostic capability for the NASH. Serum levels of alanine aminotransferase, alkaline phosphatase, gamma-glutamyl transpeptidase, adiponectin, leptin, and CK-18 were measured in 27 patients (8 patients with simple fatty liver, 6 fatty liver with fibrosis patients, 13 patients with NASH) and 23 healthy controls. Regarding gender difference in the control group, although both adiponectin and leptin significantly increased in the female compared with the male (p<0.002 and p<0.01, respectively), there were no significant gender

A 91-year-old dog-owning woman with a history of hypertension and femoral neck fracture consulted our hospital with fever and femur pain with redness. Laboratory test results showed leukocytosis with 85 % neutrophils and high values of C-reactive protein and procalcitonin. In addition, growth of Gram-positive streptococcus was observed in two independent blood culture sets. The isolated bacterium was identified as Streptococcus canis on the basis of biochemical properties and sequencing analyses of the 16S rRNA gene. The patient recovered completely without critical illness following prompt antimicrobial treatment with ceftriaxone. S. canis, a β-hemolytic Lancefield group G streptococcus, is in general isolated from various animal sources, but its isolation from a human clinical sample is extremely rare. Since β-hemolytic streptococci can cause severe infectious diseases such as necrotizing fasciitis, it is absolutely necessary to start antimicrobial treatment immediately. It is necessary to identify pathogenic bacteria carefully and to obtain information on a patient's background, including history of contact with an animal, when S. canis is isolated. © 2013 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases.

ヒト免疫不全ウイルス(HIV)は治療や2次感染予防において早期診断が重要である。今回われわれはウインドウ期を20日間前後に短縮した第4世代HIVスクリーニング試薬「HISCL HIV Ag+Ab試薬」の基礎的検討を行った。その結果、再現性や既存の第2世代試薬、第4世代試薬との一致率において良好な成績であり、化学発光を測定原理とする他社第4世代試薬と同等の感度を有していた。また、陰性分布では、偽陽性率0.09%となり、非常に特異性が高いことが確認できた。本試薬は良好な基本性能を有し、さらに測定時間17分と迅速測定が可能であることより、緊急検査および診療前検査に大きく貢献できるものと考えられる。(著者抄録)

Immune escape, the ability of tumor cells to avoid tumor-specific immune responses, occurs during the development and progression of several types of human malignancies, including colorectal cancer (CRC). Indoleamine 2,3-dioxygenase (IDO), the tryptophan catabolic enzyme, plays a significant role in regulating the immune response and provides tumor cells with a potent tool to evade the immune system. In the present study, we examined the effects of (-)-epigallocatechin gallate (EGCG), the major catechin in green tea, on the inhibition of IDO expression induced by interferon (IFN)-gamma in human CRC cells. We found that IFN-gamma increased the expression levels of IDO protein and m RNA in HT29 and SW837 CRC cell lines. Treatment of SW837 cells with EGCG significantly decreased IFN-gamma-induced expression of IDO protein and m RNA in a dose-dependent manner. Enzymatic activity of IDO, determined by the concentration of L-kynurenine in the culture medium, was also significantly inhibited by EGCG treatment. Phosphorylation of signal transducer and activator of transcription 1 (STAT1) induced by IFN-gamma was also significantly inhibited by EGCG. Reporter assays indicated that EGCG inhibited the transcriptional activities of IDO promoters, IFN-stimulated response element and IFN-gamma activation sequence, activated by STAT I phosphorylation. These findings suggest that EGCG may exert antitumor effects on CRC, at least in part, by inhibiting the expression and function of IDO through the suppression of STAT I activation. EGCG may, thus, serve as a potential agent for antitumor immunotherapy and be useful in the chemoprevention and/or treatment of CRC.

The escape of preneoplastic cells from the immune system, which is caused by immune tolerance, occurs during the development of several types of tumors. Indoleamine 2,3-dioxygenase (IDO) plays a critical role in the induction of immune tolerance. In the present study we investigated the effects of 1-methyltryptophan (1-MT), an IDO inhibitor, and (-1 )-epigallocatechin gallate (EGCG), the major catechin in green tea, on the development of azoxymethane (AOM)-induced colonic preneoplastic lesions by focusing on the inhibition of IDO. To induce colonic premalignant lesions, male F344 rats were injected with AOM (20 mg/kg body weight, s.c.) once a week for 2 weeks. They also received 0.2% 1-MT or 0.1% EGCG in their drinking water for 4 weeks, starting 1 week before the first dose of AOM. Both 1-MT and EGCG significantly decreased the total number of aberrant crypt foci and β-catenin-accumulated crypts, which overexpressed IDO protein. Treatment with EGCG decreased IDO mRNA expression in both the colonic epithelium and stroma of rats induced by AOM. The AOM-induced increase in cyclooxygenase-2 mRNA expression in the colonic stroma was significantly decreased by EGCG. Furthermore, AOM-induced increases in IDO activity in the serum and stroma were significantly inhibited by 1-MT and EGCG. Inhibition of IDO activity by 1-MT and EGCG was also observed in cell-free assays. These findings suggest that upregulation of IDO activity is observed in the early stages of colon carcinogenesis and that the use of IDO inhibitors, such as 1-MT and EGCG, which suppress the occurrence of colonic preneoplastic lesions, could be a novel strategy for the chemoprevention of colon cancer. © 2012 Japanese Cancer Association.

The activity of IDO that catalyzes the degradation of tryptophan (Trp) into kynurenine (Kyn) increases after diseases caused by different infectious agents. Previously, we demonstrated that IDO has an important immunomodulatory function in immune-related diseases. However, the pathophysiological role of IDO following acute viral infection is not fully understood. To investigate the role of IDO in the L-Trp-Kyn pathway during acute viral myocarditis, mice were infected with encephalomyocarditis virus, which induces acute myocarditis. We used IDO-deficient (IDO -/-) mice and mice treated with 1-methyl-D,L-Trp (1-MT), an inhibitor of IDO, to study the importance of Trp-Kyn pathway metabolites. Postinfection with encephalomyocarditis virus infection, the serum levels of Kyn increased, whereas those of Trp decreased, and IDO activity increased in the spleen and heart. The survival rate of IDO -/- or 1-MT-treated mice was significantly greater than that of IDO +/+ mice. Indeed, the viral load was suppressed in the IDO -/- or 1-MT-treated mice. Furthermore, the levels of type I IFNs in IDO -/- mice and IDO -/- bone marrow-transplanted IDO +/+ mice were significantly higher than those in IDO +/+ mice, and treatment of IDO -/- mice with Kyn metabolites eliminated the effects of IDO -/- on the improved survival rates. These results suggest that IDO has an important role in acute viral myocarditis. Specifically, IDO increases the accumulation of Kyn pathway metabolites, which suppress type I IFNs production and enhance viral replication. We concluded that inhibition of the Trp-Kyn pathway ameliorates acute viral myocarditis. © 2012 by The American Association of Immunologists, Inc.

Secondary bacterial infection in humans is one of the pathological conditions requiring clinical attention. In this study, we examined the effect of lipopolysaccharide (LPS) on encephalomyocarditis virus (EMCV) infected mice. All mice inoculated with EMCV at 5 days before LPS challenge died within 24 h. LPS-induced TNF-alpha mRNA expression was significantly increased in the brain and heart at 5 days after EMCV infection. CD11b(+)/TLR4(+) cell population in the heart was remarkably elevated at 5 days after EMCV infection, and sorted CD11b(+) cells at 5 days after EMCV infection produced a large amount of TNF-alpha on LPS stimulation in vivo and in vitro. In conclusion, we found that the infiltration of CD11b(+) cells into infected organs is involved in the subsequent LPS-induced lethal shock in viral encephalomyocarditis. This new experimental model can help define the mechanism by which secondary bacterial infection causes a lethal shock in viral encephalomyocarditis.

Nonalcoholic fatty liver disease is one of the most common liver diseases. L-Tryptophan and its metabolite serotonin are involved in hepatic lipid metabolism and inflammation. However, it is unclear whether L-tryptophan promotes hepatic steatosis. To explore this issue, we examined the role of L-tryptophan in mouse hepatic steatosis by using a high fat and high fructose diet (HFHFD) model. L-Tryptophan treatment in combination with an HFHFD exacerbated hepatic steatosis, expression of HNE-modified proteins, hydroxyproline content, and serum alanine aminotransaminase levels, whereas L-tryptophan alone did not result in these effects. We also found that L-tryptophan treatment increases serum serotonin levels. The introduction of adenoviral aromatic amino acid decarboxylase, which stimulates the serotonin synthesis from L-tryptophan, aggravated hepatic steatosis induced by the HFHFD. The fatty acid-induced accumulation of lipid was further increased by serotonin treatment in cultured hepatocytes. These results suggest that L-tryptophan increases the sensitivity to hepatic steatosis through serotonin production. Furthermore, L-tryptophan treatment, adenoviral AADC introduction, and serotonin treatment induced phosphorylation of the mammalian target of rapamycin (mTOR), and a potent mTOR inhibitor rapamycin attenuated hepatocyte lipid accumulation induced by fatty acid with serotonin. These results suggest the importance of mTOR activation for the exacerbation of hepatic steatosis. In conclusion, L-tryptophan exacerbates hepatic steatosis induced by HFHFD through serotonin-mediated activation of mTOR. © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.

Acid sphingomyelinase (ASM) regulates the homeostasis of sphingolipids, including ceramides and sphingosine-1-phosphate (S1P). Because sphingolipids regulate AKT activation, we investigated the role of ASM in hepatic glucose and lipid metabolism. Initially, we overexpressed ASM in the livers of wild-type and diabetic db/db mice by adenovirus vector (Ad5ASM). In these mice, glucose tolerance was improved, and glycogen and lipid accumulation in the liver were increased. Using primary cultured hepatocytes, we confirmed that ASM increased glucose uptake, glycogen deposition, and lipid accumulation through activation of AKT and glycogen synthase kinase-3(β. In addition, ASM induced up-regulation of glucose transporter 2 accompanied by suppression of AMP-activated protein kinase (AMPK) phosphorylation. Loss of sphingosine kinase-1 (SphK1) diminished ASM-mediated AKT phosphorylation, but exogenous S1P induced AKT activation in hepatocytes. In contrast, SphK1 deficiency did not affect AMPK activation. These results suggest that the SphK/S1P pathway is required for ASM-mediated AKT activation but not for AMPK inactivation. Finally, we found that treatment with high-dose glucose increased glycogen deposition and lipid accumulation in wild-type hepatocytes but not in ASM-cells. This result is consistent with glucose intolerance in ASM-mice. In conclusion, ASM modulates AKT activation and AMPK inactivation, thus regulating glucose and lipid metabolism in the liver. © The Author(s).

Indoleamine 2,3-dioxygenase (IDO) exerts immunomodulatory effects due to enzymatic activities catalyzing the essential amino acid L-tryptophan. IDO activity might play an important role in regulating immune responses exerted by antigen-presenting cells as a potent tool to help escape from assault by the immune system. In this study, we performed immunohistochemical analysis for IDO expression using mouse anti-human IDO monoclonal antibody in 119 tissue samples of diffuse large B-cell lymphoma (DLBCL) obtained before treatment with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP). Not only the lymphoma cells themselves but also dendritic cells (DCs) expressed IDO. Positive IDO expression in lymphoma cells was found in 38 cases (32%). Complete remission rates in patients with IDO-positive DLBCL and IDO-negative DLBCL were 55.3% and 79.0% (p=0.008), while 3-year overall survival rates were 49.8% and 78.8%, respectively (p=0.0003). IDO activity might thus play an important role in DLBCL and cells that express IDO appear important for determining outcomes after R-CHOP treatment. IDO might represent a candidate therapeutic target for DLBCL patients who show resistance to chemotherapy.

Acid sphingomyelinase (ASM) regulates the homeostasis of sphingolipids, including ceramides and sphingosine-1-phosphate (S1P). Because sphingolipids regulate AKT activation, we investigated the role of ASM in hepatic glucose and lipid metabolism. Initially, we overexpressed ASM in the livers of wild-type and diabetic db/db mice by adenovirus vector (Ad5ASM). In these mice, glucose tolerance was improved, and glycogen and lipid accumulation in the liver were increased. Using primary cultured hepatocytes, we confirmed that ASM increased glucose uptake, glycogen deposition, and lipid accumulation through activation of AKT and glycogen synthase kinase-3β. In addition, ASM induced up-regulation of glucose transporter 2 accompanied by suppression of AMP-activated protein kinase (AMPK) phosphorylation. Loss of sphingosine kinase-1 (SphK1) diminished ASM-mediated AKT phosphorylation, but exogenous S1P induced AKT activation in hepatocytes. In contrast, SphK1 deficiency did not affect AMPK activation. These results suggest that the SphK/S1P pathway is required for ASM-mediated AKT activation but not for AMPK inactivation. Finally, we found that treatment with high-dose glucose increased gl

IDO converts tryptophan to L-kynurenine, and it is noted as a relevant molecule in promoting tolerance and suppressing adaptive immunity. In this study, we examined the effect of IDO in α-galactosylceramide (α-GalCer)-induced hepatitis. The increase in IDO expression in the liver of wild-type (WT) mice administered α-GalCer was confirmed by real-time PCR, Western blotting, and IDO immunohistochemical analysis. The serum alanine aminotransferase levels in IDO-knockout (KO) mice after α-GalCer injection significantly increased compared with those in WT mice. 1-Methyl-D-tryptophan also exacerbated liver injury in this murine hepatitis model. In α-GalCer-induced hepatitis models, TNF-α is critical in the development of liver injury. The mRNA expression and protein level of TNF-α in the liver from IDO-KO mice were more enhanced compared with those in WT mice. The phenotypes of intrahepatic lymphocytes from WT mice and IDO-KO mice treated with α-GalCer were analyzed by flow cytometry, and the numbers of CD49b+ and CD11b+ cells were found to have increased in IDO-KO mice. Moreover, as a result of the increase in the number of NK cells and macrophages in the liver of IDO-KO mice injected with α-GalCer, TNF-α secretion in these mice was greater than that in WT mice. Deficiency of IDO exacerbated liver injury in α-GalCer-induced hepatitis. IDO induced by proinflammatory cytokines may decrease the number of TNF-α-producing immune cells in the liver. Thus, IDO may suppress overactive immune response in the α-GalCer-induced hepatitis model. Copyright © 2010 by The American Association of Immunologists, Inc.

Indoleamine 2,3-dioxygenase, the L-tryptophan-degrading enzyme, plays a key role in the powerful immunomodulatory effects on several different types of cells. Because modulation of IDO activities after viral infection may have great impact on disease progression, we investigated the role of IDO following infection with LP-BM5 murine leukemia virus. We found suppressed BM5 provirus copies and increased type I IFNs in the spleen from IDO knockout (IDO -/-) and 1-methyl-D-L-tryptophan-treated mice compared with those from wild-type (WT) mice. Additionally, the number of plasmacytoid dendritic cells in IDO-/- mice was higher in the former than in the WT mice. In addition, neutralization of type I IFNs in IDO-/- mice resulted in an increase in LP-BM5 viral replication. Moreover, the survival rate of IDO -/- mice or 1-methyl-D-L-tryptophan-treated mice infected with LP-BM5 alone or with both Toxoplasma gondii and LP-BM5 was clearly greater than the survival rate of WT mice. To our knowledge, the present study is the first report to observe suppressed virus replication with upregulated type I IFN in IDO-/- mice, suggesting that modulation of the IDO pathway may be an effective strategy for treatment of virus infection.

Objectives Serum creatinine (Scr) is not a reliable marker of renal function in critically ill patients because of an enhancement of protein catabolism, which makes it difficult to adjust the dosage of renally eliminated drugs such as antibiotics. This study aimed to investigate whether serum cystatin C (Scys-C) could be used as a reliable marker of renal function. Methods We investigated whether Scys-C was a reliable marker of renal function in 56 critically ill patients. Subsequently, the usefulness of Scys-C to determine the initial loading and the maintenance dose of vancomycin was examined in 18 patients. Crea- tinine clearance (Ccr) was assessed from Scr and creatinine in urine collected over 24 h (24-h Ccr). Key findings There was a good correlation between 24-h Ccr and 1/Scys-C (r2 = 0.616), whereas less marked correlation was observed between 24-h Ccr and 1/Scr (r2 = 0.221). On the other hand, vancomycin concentration was predicted from population pharmacokinetic parameters based on a two-compartment linear model. There were significant correlations between real trough concentrations of vancomycin and the values predicted from Scys-C using various equations (r 2 = 0.416-0.

Serum creatinine (Scr) is not a reliable marker of renal function in critically ill patients because of an enhancement of protein catabolism, which makes it difficult to adjust the dosage of renally eliminated drugs such as antibiotics. This study aimed to investigate whether serum cystatin C (Scys-C) could be used as a reliable marker of renal function.We investigated whether Scys-C was a reliable marker of renal function in 56 critically ill patients. Subsequently, the usefulness of Scys-C to determine the initial loading and the maintenance dose of vancomycin was examined in 18 patients. Crea- tinine clearance (Ccr) was assessed from Scr and creatinine in urine collected over 24 h (24-h Ccr).There was a good correlation between 24-h Ccr and 1/Scys-C (r(2) = 0.616), whereas less marked correlation was observed between 24-h Ccr and 1/Scr (r(2) = 0.221). On the other hand, vancomycin concentration was predicted from population pharmacokinetic parameters based on a two-compartment linear model. There were significant correlations between real trough concentrations of vancomycin and the values predicted from Scys-C using various equations (r(2) = 0.416-0.488), while less pronounced r

Early detection of nerve dysfunction is important to provide appropriate care for patients with diabetic polyneuropathy. The aim of this study was to assess the echo intensity of the peripheral nerve and to evaluate the relationship between nerve conduction study results and sonographic findings in patients with type 2 diabetes mellitus.Thirty patients with type 2 diabetes (mean +/- SD, 59.8 +/- 10.2 years) and 32 healthy volunteers (mean, 53.7 +/- 13.9 years) were enrolled in this study. The cross-sectional area (CSA) and echo intensity of the peripheral nerve were evaluated at the carpal tunnel and proximal to the wrist (wrist) of the median nerve and in the tibial nerve at the ankle.There was a significant increase in the CSA and hypoechoic area of the nerve in diabetic patients compared with controls (wrist, 7.1 +/- 2.0 mm(2), 62.3% +/- 3.0%; ankle, 8.9 +/- 2.8 mm(2), 57.6% +/- 3.9%; and wrist, 9.8 +/- 3.7 mm(2), 72.3% +/- 6.6%; ankle, 15.0 +/- 6.1 mm(2), 61.4% +/- 5.3% in controls and diabetic patients, respectively; P<.05). Cross-sectional areas were negatively correlated with reduced motor nerve conduction velocity and delayed latency.These results suggest that sonographic e

Introduction of rituximab has largely improved the prognosis of patients with diffuse large B-cell lymphoma(DLBCL). Such change in therapeutic outcome necessitates the identification of additional prognostic factors to conventional indexes that have been validated for CHOP without rituximab. Indoleamine 2,3-dioxygenase (IDO) exerts intense immunomodulatory effects because of enzymatic activities that catalyze the breakdown of the essential amino acid L-tryptophan. The activity of IDO can be estimated by measuring the serum concentration of L-kynurenine. Here, we investigated the role of L-kynurenine as a prognostic marker in R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisolone) therapy. Experimental design: Data from 73 consecutive patients treated with eight cycles of R-CHOP or R-THP (tetrahydropyranyl adriamycin)-COP between December 2002 and March 2007 were analyzed. L-kynurenine concentrations in serum samples obtained at admission were measured by high-performance liquid chromatography. Results: The median serum L-kynurenine level was 1.575 microm (range 0.537-9.588). The complete response (CR) rates of patients with L-kynurenine<1.5 and>or =1.5 microm w

Autoantibodies against cyclic citrullinated peptide (anti-CCP) are sensitive and highly specific markers for rheumatoid arthritis (RA). We evaluated the analytical and diagnostic accuracy of chemiluminescenceenzyme immunoassay (CLEIA) for anti-CCP antibodies, and compared it with that of ELISA.Ninety-nine RA patients who were diagnosed according to the American College of Rheumatology criteria, 16 patients with osteoarthritis, and 94 healthy subjects were included. Sera were used to assess the precision, functional sensitivity, and linearity of anti-CCP antibody determination by CLEIA and the correlation of anti-CCP antibody values between CLEIA and ELISA.For anti-CCP antibodies by CLEIA, the total CV was 4.0 and 5.3% at 21.17 and 90 U/ml, respectively, and the lower limit of detection was 0.1 U/ml. The correlation of CLEIA (x) with ELISA (y) for anti-CCP was: y=1.08x+4.171, r=0.9178 (p<0.0001). No difference was observed in the sensitivity and specificity between CLEIA and ELISA.The automated CLEIA processing system for determining anti-CCP antibodies showed a good analytical performance, and suggested that the CLEIA system has a potential to provide clinically useful data within

Kupffer cells, resident tissue macrophages of the liver, play a key role in the regulation of hepatic inflammation, hepatocyte death, and fibrosis that characterize liver diseases. However, it is controversial whether Kupffer cells promote or protect from liver injury. To explore this issue we examined the role of Kupffer cells in liver injury, cell death, regeneration, and fibrosis on cholestatic liver injury in C57BL/6 mice using a model of partial bile duct ligation (BDL), in which animals do not die and the effects of BDL can be compared betweeninjured ligated lobes and nonligated lobes. In cholestatic liver injury, the remaining viable cells represented tolerance for tumor necrosis factor alpha (TNF-alpha)-induced hepatocyte apoptosis and regenerative features along with AKT activation. Inhibition of AKT by adenovirus expressing dominant-negative AKT abolished the survival and regenerative properties in hepatocytes. Moreover, Kupffer cell depletion by alendronate liposomes increased hepatocyte damage and the sensitivity of TNF-alpha-induced hepatocyte apoptosis in ligated lobes. Kupffer cell depletion decreased hepatocyte regeneration and liver fibrosis with reduced AKT activa

Cytotoxic T lymphocytes (CTLs) play a crucial role in the infections and the antitumor immunity. Induction and activation of antigen-specific CTLs is an important strategy in immunotherapy for various diseases, and several researchers have focused on the modulation of CTL induction and function. Natural killer T (NKT) cells are an important focus area of researchers studying immunomodulatory responses to tumors and infectious diseases. CD1d-restricted NKT cells consist of type I NKT cells and type II NKT cells. -galactosylceramide ( -GalCer)-activated type I NKT cells secrete both Th1 (e.g., IFN- ) and Th2 cytokines, affect the expression of costimulatory molecules in immune cells, and regulate the host immune system. Type II NKT cells, however, are stimulated by sulfatide, a self-glycolipid derived from myelin, and play an immunosuppressive role in animal model of autoimmune diseases. CTL generation, activation, and suppression are strongly affected by activated type I and type II NKT cells. Thus, the regulation of these NKT cells leads to the modification of CTL function. CTLs contribute to antimicrobial responses, antitumor immune and autoimmune responses. Understanding the role of NKT cells in the regulation of CTL generation, activation, and suppression enable the development of novel treatment strategies for these diseases. Copyright © 2010 H. Ito and M. Seishima.

Aim: Statins are effective in lowering cholesterol levels, but cause fatal rhabdomyolysis in susceptible individuals. Because it has been hypothesized that muscle damage could result from alterations in Ca2+ homeostasis in muscle cells, we tested whether measuring statin-induced changes in intracellular calcium ([Ca2+](i)) is useful for predicting susceptibility to statin-muscle damage, using human CD19+ primary B lymphocytes. Methods: Statin-induced alterations in [Ca2+](i) were studied using the human THP-1 cell line and CD19+ primary B lymphocytes. Changes in [Ca2+](i) were measured directly in fluo-3- loaded cells using either single or dual-color flow cytometry. Results: The Ca2+ release study suggested that statin-induced changes in [Ca2+](i) were due to Ca2+ release from ryanodine-sensitive Ca2+ stores and mitochondrial compartments. Further, statin users who experienced elevated creatine kinase (n=8) exhibited significantly greater statin-induced Ca2+ release in B cells than healthy volunteers (n=45) and statin users without elevated creatine kinase (n=16), while no difference was seen between the latter two groups. Conclusion: Statin-induced Ca2+ release from ryanodine-sensitive stores and mitochondria may contribute to myotoxicity. The laboratory test for Ca2+ release using CD19+ primary B lymphocytes may be useful to predict susceptibility to statin-induced muscle toxicity prior to statin use.