Evaluation of IAEA Clearance Concept for Low-level Radioactive Waste from a Radioisotope Research Institute.

UNLABELLED: The clearance of solid low-level radioactive laboratory waste (LLRW) after decay-in-storage (DIS) obtained from a research institute and thoroughly separated using the separation and classification protocols presented in this study was evaluated. METHOD: The radioisotope (RI) content of incinerated LLRW from the specified RI research group (group A); the RI content of LLRW obtained in fiscal year 2000, which contained radionuclides with half-lives of less than 164 d (LLRW2); and the RI content of the LLRW reported in group A's disposal records were compared. The LLRW2 and LLRW of group A were incinerated after 2 y of decay-in-storage and immediately after storage, respectively. RESULTS: The highest ratio of the RI of incinerated LLRW to the value in the disposal records was 2.52 for 5¹Cr. The radioactivities of radionuclides in both the LLRW2 and LLRW for ³5S, 45Ca, 5¹Cr, ¹²5I, ³²P, ³³P, and 99mTc and the incinerated ash after 1 y later of decay-in-storage were below the clearance level defined by the RS-G-1.7 of the International Basic Safety Standard without contamination by ³H and ¹4C. These remains contained very small amounts of some long-half-life radionuclides of natural origin after 7 y of decay-in-storage. CONCLUSION: This LLRW separation protocol was effective for the separation of ³H and ¹4C. LLRW2 after 2 years of DIS and its incinerated ash after one year later of DIS were below the clearance level for radioactivity and radioactivity concentration.

IGFBP-3 expression in hepatocellular carcinoma involves abnormalities in TGF-beta and/or Rb signaling pathways.

Insulin-like growth factor binding protein-3 (IGFBP-3) is a mediator of growth suppression signals and a putative tumor suppressor gene. The growth suppression mechanisms of IGFBP-3 have not been well clarified. We examined the expression of IGFBP-3 transcripts in human hepatocellular carcinoma (HCC) and the relationship between IGFBP-3 expression and the transforming growth factor-beta (TGF-beta) and/or retinoblastoma (Rb) signaling pathways. In situ hybridization revealed IGFBP-3 transcripts in cancer cells in 6 of 57 (10%) HCCs, including moderately and poorly differentiated HCCs with intrahepatic metastasis. In contrast, all lung metastatic nodules of 4 HCCs showed IGFBP-3 transcripts in cancer cells. The cDNA microarray showed that genes for the TGF-beta pathway and Rb were up-regulated in IGFBP-3-expressing HCCs. In 6 HCCs presenting IGFBP-3, immunohistochemical analyses showed abnormalities in the TGF-beta and/or Rb pathways; the loss of phosphorylated-Smad2 was observed in 2, and overexpression of phosphorylated-Rb was observed in the remaining 4 HCCs. The present study suggests that IGFBP-3 mediates growth suppression signals via the TGF-beta and/or Rb pathways in HCC.

Decreased expression of B7 costimulatory molecules and major histocompatibility complex class-I in human hepatocellular carcinoma.

BACKGROUND AND AIM: We analyzed the expression of antigen-processing and antigen-presenting molecules in surgically resected fresh samples of human hepatocellular carcinoma (HCC) tissue to elucidate a mechanism of immune escape. We also examined the expression of interleukin (IL)-10 protein, which might act to downregulate expression of antigen-processing and antigen-presenting molecules. METHODS: Twenty-eight HCC samples obtained by surgical resection were analyzed for the expression of beta2-microglobulin, heat-shock protein (HSP)-70, human leukocyte antigen (HLA) class-I, CD80 (B7-1), CD86 (B7-2) and IL-10 by immunostaining. RESULTS: Beta2-microglobulin and HSP-70 were preserved in all samples. In contrast, the expression of HLA class-I molecules was significantly reduced according to lowering in the histological grading of tumor differentiation (P = 0.024). Furthermore, B7-1 and B7-2 expression was reduced in tumor cells compared with corresponding areas of liver tissue without malignant involvement irrespective of the histological grading of tumors (21% and 36%, respectively). Although IL-10 protein was expressed in 54% of HCC, no relationship between the expression of IL-10 and downregulation of B7-1, B7-2, and HLA class-I was evident. CONCLUSION: These findings suggest the potential role of B7 co-stimulatory molecules and HLA class-I molecules in facilitating HCC escape from immune surveillance without the involvement of IL-10.

DNA alterations during multi-step development of human hepatocellular carcinomas revealed by laser capture microdissection.

In order to clarify early molecular events involved in liver carcinogenesis, we analyzed 53 liver-cirrhosis nodules (LCNs) from five patients and 13 micro-hepatocellular carcinoma (HCC) nodules from one patient and looked for alterations of microsatellites in genomic DNA after carefully preparing the tissue samples by laser-capture microdissection (LCM). Allelotyping was done with 20 markers corresponding to anonymous microsatellites and 13 corresponding to tumor suppressor genes (TSGs) that had shown significant alterations in HCCs. We detected both loss of heterozygosity (LOH) and microsatellite shifts (MS). Overall, 24 of 53 (47%) of LCNs showed LOH with any of the informative markers used in the study, reflecting that proportion of LCNs with clonal growth. The fractional allelic loss (FAL) index, an indication of total genomic complexity, was not significantly different between LCN and micro-HCC nodules, but their profiles of alteration were different. These profiles were classified into three groups: (1) LCN profile-allelic loss at chromosomal arms 1q and 14q, TBP and BRCA1; (2) HCC profile-LOH at 4q, 6q, 7q, 17p, NF1, IGFIIr and p53 in micro-HCC nodules; these changes in early lesions were identical to those seen in mature HCCs; (3) Common profile-LOH at NF1 and 6q, including IGFIIr, common to both LCN and HCC. No LCN showed LOH at p53 and Rb, loci that are generally altered in HCCs. However, 12 intra-tumoral nodules examined had lost p53 in all informative cases, although the loss of Rb was a late event. These results suggest that early genomic profiles confined to LCNs, and additional profiles that can be observed when liver tissue undergoes malignant transformation, support a model of multi-step development of HCC.

Quantitative analysis of p40/p46 and p69/p71 forms of 2',5'-oligoadenylate synthetase mRNA by competitive PCR and its clinical application.

BACKGROUND: 2',5'-Oligoadenylate synthetases (2-5AS) are type I interferon (IFN)-induced proteins with antiviral capacity. Three major forms of 2-5AS with distinct enzymatic activities have been described in IFN-treated human cells. We measured distinct forms of 2-5AS mRNA to analyze the relationship with its enzymatic activity and response to IFN therapy in chronic hepatitis C. METHODS: We established a method to quantify p40/p46 and p69/p71 forms of 2-5AS mRNA by use of reverse transcription followed by competitive PCR. The 2-5AS mRNA concentrations were measured in peripheral blood mononuclear cells from 40 patients with chronic hepatitis C and 28 control individuals. RESULTS: Reconstitution experiments and comparison with Northern blot analyses revealed that our method accurately and linearly quantified 2-5AS mRNA. 2-5AS mRNA concentrations and 2-5AS enzymatic activity were correlated (P <0.03). Our data demonstrated a correlation in 2-5AS mRNA between p40/p46 and p69/p71 (P <0.02), indicating a similar regulation of the expression of these genes. Our data also demonstrated that pretreatment concentrations of 2-5AS mRNA correlated with responses to IFN therapy in chronic hepatitis C. CONCLUSIONS: Our method for measuring 2-5AS mRNA concentrations could provide an important marker for selecting patients for IFN therapy and may be useful for the development of more effective therapeutic strategies for chronic hepatitis C.

Genome aberrations observed by restriction landmark genome scanning analysis of chromosomal DNA in various types of primary hepatocellular carcinoma.

BACKGROUND/AIMS: The genes responsible for hepatocellular carcinoma have not been identified. To identify the relevant genes of hepatocellular carcinoma, detailed and comprehensive information of genomic aberrations must be obtained. To reveal the chromosomal aberrations in hepatocellular carcinoma, we carried out a restriction landmark genome scanning analysis of various types of hepatocellular carcinoma. METHODOLOGY: Samples of various types of hepatocellular carcinoma, including two with multinodular-hepatocellular carcinomas, one hepatocellular carcinoma showing nodules in a nodule pattern, one hepatocellular carcinoma metastasized to different tissues, three small (< 2.0 cm) hepatocellular carcinomas and four large (> 5.0 cm) hepatocellular carcinomas were examined by the restriction landmark genome scanning method with corresponding non-hepatocellular carcinoma tissues. Restriction enzyme Not I was used as a landmark enzyme, Eco RV was used as a fragmentation enzyme, and Hin fI was used as a digestion enzyme in the gel for two-dimensional electrophoresis. RESULTS: We observed spot aberrations with different origins. Frequently observed spot aberrations originated from the change in the methylation status of repetitive sequences. No clear correlation between the pathological grade and the number or type of spot aberrations was observed. CONCLUSIONS: We demonstrated that major aberrations of restriction landmark genome scanning spots originated from the change of methylation status in hepatocellular carcinoma.

Serum gamma-interferon-inducing factor (IL-18) and IL-10 levels in patients with acute hepatitis and fulminant hepatic failure.

BACKGROUND AND AIMS: The aim was to determine the role of T-helper (Th)1/Th2 cytokine responses in the clinical outcome of patients with acute liver injury. METHODS: The serum levels of the cytokines, interleukin (IL)-18, gamma-interferon (IFN-gamma), IL-10 and IL-4 were measured in 20 fulminant hepatic failure (FHF), 18 acute hepatitis (AH), 30 chronic viral hepatitis and 20 liver cirrhosis (LC) patients. Thirteen cases were from the intensive care unit (ICU) and there were 21 healthy volunteers. Immunohistochemical staining of liver biopsies for IL-18 expression was also performed. RESULTS: Serum IL-18 levels in patients with FHF were significantly more elevated than in patients with other liver diseases, ICU cases and healthy volunteers. Furthermore, serum IFN-gamma levels in patients with FHF were also significantly higher than in patients with chronic viral hepatitis, LC and healthy volunteers. We found a positive correlation between the levels of IL-18 and IFN-gamma. However, no relationship was observed between these and clinical outcome. In immunohistochemical staining, CD68+ macrophage cells and IL-18-positive cells were observed in portal zones. Elevated serum IL-10 levels were restricted to patients presenting with FHF, and were significantly higher in surviving cases (P < 0.01). Furthermore, serum IL-10 levels, but not IL-4 levels, were inversely correlated with serum total bilirubin concentrations (P = 0.045) and the death rate (p) outlined in Japan (P = 0.030). CONCLUSION: These results suggest that IL-18 and IFN-gamma are involved in the pathogenesis of acute hepatic injury in humans, and that, in particular, elevated serum levels of IL-10 may be predictive of improved outcomes for these patients.

Reduced expression of insulin-like growth factor binding protein-3 and its promoter hypermethylation in human hepatocellular carcinoma.

Insulin-like growth factor binding protein-3 (IGFBP-3) is postulated to be a mediator of growth suppression signals. Reduced expression of the IGFBP-3 was observed in nine out of 12 human hepatocellular carcinomas (HCC) (75%). Promoter hypermethylation of the IGFBP-3 was detected in four out of 12 HCCs (33%) although mutations were not identified. The expression of IGFBP-3 was restored by the demethylating agent 5-aza-2'-deoxycytidine in HCC cell line with promoter hypermethylation (HepG2). As IGFBP-3 functions like a tumor suppressor gene, it may be used as a therapeutic target for HCC.