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Human telomerase reverse transcriptase (hTERT) has been identified as the catalytic subunit of human telomerase. To clarify the clinical significance of hTERT mRNA in hepatocellular carcinoma (HCC), we investigated the relationship between telomerase activity and hTERT mRNA in human HCC and non-HCC tissues. The hTERT mRNA was detected in 17 (89.47%) of 19 livers with HCC and in 4 (21.05%) of 19 noncancerous tissues from these livers. Telomerase activity was detected in 17 of the 19 tumor tissues (89.47%) and in 4 of the 19 nontumor tissues (21.05%). The hTERT mRNA was detected in all tissues that were telomerase-positive and it was undetected in all tissues that were telomerase-negative. The correlation between the expression of hTERT mRNA and human telomerase activity in this study indicates that hTERT mRNA could be useful to diagnose cancer. Also, as telomerase production may be under the control of hTERT mRNA, the possibility is great that noncancerous liver tissue with chronic liver diseases acquires HCC when the hTERT mRNA is positive. [ABSTRACT FROM AUTHOR]

Background/purpose: Few papers were available on the interproximal cleaning efficiency by manual toothbrushes when used alone. The aim was to investigate the efficiency of commercially available toothbrushes on interproximal cleaning and determine the key properties that would make the differences., Materials and Methods: Artificial-teeth were coated with manicure type experimental dental plaque covering mainly the interproximal surface and fixed in the jaw model of a dental simulator. A modified scrubbing technique was employed to brush out the plaque conducted by one trained dentist using 26 different toothbrushes from the equal number of separate interproximal conditions. The rate of the plaque removal (%) was calculated by measuring the plaque free areas on the post-brush images., Results: The data analysis using mixed effect modelling showed that stiffness, number of tufts and total length have effect on the rate of the plaque removable from the interproximal surfaces., Conclusion: This study indicated consideration should be given to toothbrush properties to enhance plaque removal from the interproximal surfaces., (© 2019 Association for Dental Sciences of the Republic of China. Publishing services by Elsevier B.V.)

Background: The virologic impact of adding interferon to antiviral nucleoside therapy was studied in Japanese patients having perinatally transmitted hepatitis B virus (HBV) genotype C., Methods: Sixty-four patients including 41 positive for hepatitis B e antigen (HBeAg) were assigned to receive either (1) a combination of interferon-alpha (6 million units daily for 2 weeks, then three times weekly) plus lamivudine (100 mg daily) for 24 weeks followed by lamivudine alone for 28 weeks (n = 30) or (2) 52-week lamivudine monotherapy (n = 34)., Results: The combination treatment enhanced the early virologic response, and HBV clearance was more frequent at week 8 for patients with baseline HBV DNA < or = 7 log copies/ml (90% vs. 33%, P = 0.013) and at week 24 for patients with baseline HBV DNA > 7 log copies/ml (75% vs. 40%, P = 0.080). In the combination arm, YMDD mutants emerged less often at week 52 (8% vs. 30%, P = 0.047). However, reversion of the precore mutation was more prominent with combination treatment than with monotherapy (McNemar test, P = 0.014 and P = 0.103, respectively). HBeAg seroconversion (P = 0.429) and sustained off-treatment HBV suppression to < or =5 log copies/ml (log-rank test, P = 0.195) were not improved., Conclusions: Simultaneous commencement of treatment with interferon and a nucleoside analog may be worthy as a treatment option to augment the early virologic response and prevent drug resistance in difficult-to-treat patients. Combination treatment was also shown to enhance reversion of the precore mutation. Further studies are warranted to clarify the therapeutic implications of this phenomenon.

Aim: To develop a new sensitive and inexpensive hepatitis C virus (HCV) combination test (HCV Guideline test) that enables the determination of HCV genotypes 1, 2 and 3, and simultaneous determination of HCV viral load using commercial Amplicor GT HCV MONITOR test v2.0 (microwell version)., Methods: The HCV Guideline test used the PCR product generated in commercial Amplicor GT HCV Monitor test v2.0 for viral load measurement using microwell plate version of Amplicor HCV Monitor and also captured on separate plates containing capture probes and competitive oligonucleotide probes specific for HCV genotypes 1, 2 and 3, The HCV genotype was subsequently determined using the biotin-labeled PCR product and five biotin-labeled HCV-specific probes., Results: The sensitivity of the HCV Guideline test was 0.5 KIU/mL. Specificity of the HCV Guideline test was confirmed by direct sequencing of HCV core region and molecular evolutionary analyses based on a panel of 31 samples. The comparison of the HCV Guideline test and an in-house HCV core genotyping assay using 252 samples from chronic hepatitis C patients indicated concordant results for 97.2% of samples (59.5% genotype 1, 33.7% genotype 2, 6.0% genotype 3, and 0.8% mixed genotypes). Similarly, the HCV Guideline test showed concordance with a serological test, and the serological test failed to assign any serotype in 12.7% of the samples, indicating a better sensitivity of the HCV Guideline test., Conclusion: Clinically, both viral load and genotypes (1, 2 and 3) have been found to be major predictors of antiviral therapy outcome regarding chronic hepatitis C based on guidelines and they are, in normal circumstances, performed as separate stand-alone assays. The HCV Guideline test is a useful method for screening large cohorts in a routine clinical setting for determining the treatment regimen and for predicting the outcome of antiviral therapy of chronic hepatitis C.

Clinical relevance of occult hepatitis C virus (HCV) and/or hepatitis B virus (HBV) infection(s) remains uncertain years after interferon (IFN) therapy for chronic hepatitis C. By 1993, 38 sustained virological responders (SVRs) showing HCV RNA clearance at 6 months post-treatment and 37 biochemical responders (BRs) with end-of-treatment alanine aminotransferase (ALT) normalization and subsequent 6-month stabilization within 2 x the upper limit of normal (ULN) were enrolled. They were monitored for 4.4-12 years (median 6.8), then 15 SVRs and 15 BRs underwent paired liver biopsies. Biopsy samples were tested for positive and negative HCV RNA strands, and HBV DNA surface and X sequences. All SVRs showed sustained serum HCV RNA clearance during follow-up, but hepatocellular carcinoma (HCC) developed in 4 (11%) SVRs. On paired liver biopsies, histological improvement was significant, but mild inflammation persisted in 87% of SVRs. Nonetheless, no HCV RNA sequence was amplified from liver tissues, and HBV DNA sequences were found in only one SVR. As for BRs, biochemical flare-up of >2 x ULN occurred at a 5-year risk of 41% (95% CI 24.7-56.4). The event was unpredictable but controllable by retreatment in 70%. Liver tissues after follow-up contained positive and negative HCV RNA strands, but no HBV DNA sequence was amplified. These results suggest that SVRs, albeit free of occult HCV and/or HBV infection(s) over a decade, retain mild liver inflammation and the risk of HCC. Occult HBV was also shown uninvolved in flare-up during follow-up of BRs.

Rationale: The serotonergic (5-hydroxytryptamine, 5-HT) system is extensively implicated in feeding behavior. In recent years, 5-HT receptors have been classified into 14 subtypes, and activation of 5-HT1B and 5-HT2C receptors inhibits food intake in rats. However, the precise functions in local brain areas of these receptor subtypes are unclear., Objectives: Frontal cortex (FC), lateral hypothalamic area (LH), or ventromedial hypothalamic nucleus (VMH) are involved in control of feeding behavior. We investigated the effects of 5-HT1B and 5-HT2C receptor stimulations in the three local brain areas on feeding behavior and on 5-HT metabolism., Methods: We perfused mCPP, 5-HT(1B/2C) agonist, at multiple doses via a microdialysis probe into the three local brain areas and observed food intake. Extracellular concentrations of 5-HT and 5-HIAA were measured simultaneously., Results: Perfusion of 1 mM mCPP into VMH, but not into LH nor FC at any dose, induced significant reduction of food intake compared with control. The extracellular concentrations of 5-HT were markedly increased in all three areas, but the concentrations of 5-HIAA were not changed by mCPP perfusions., Conclusions: These results indicate that the effects of 5-HT1B or 5-HT2C receptor activation on feeding behaviors depended on the brain regions, and that 5-HT1B or 5-HT2C receptors in VMH, but not in FC or in LH, play important roles in the regulation of food intake. The results also suggested that mCPP acts not only as a 5-HT(1B/2C) agonist, but also as a 5-HT releaser or as a re-uptake inhibitor. Further studies using antagonists should be conducted.

Telomerase is a cellular RNA-dependent DNA polymerase that serves to maintain the tandem arrays of telomeric TTAGGG repeats at eukaryotic chromosome ends. One of the human telomerase components is hTERT, which has three alternative spliced sites that introduce eight isoforms of hTERT mRNA. The expression of these isoforms in gastrointestinal cell lines is unknown. We developed a PCR-based assay for detecting these splicing variants. In gastric and hepatocellular carcinoma cell lines, the gamma deletion variant and its combination variants, alpha- and gamma-, beta- and gamma-, and alpha-, beta- and gamma-deletion variants were frequently detected, while they were not detected in colorectal carcinoma cell lines. Our results provide important information of use for more detailed studies on the regulation of telomerase activity.

The presence of circulatory metastasis is one of the most significant factors for poor-prognosis in patients with several types of cancer. To establish a sensitive reverse transcription PCR assay to detect micrometastasis in blood containing several cancer types, we first investigated Uroplakin II (UP II), a novel molecular marker for human transitional cell carcinoma of the bladder, in 25 types of normal organs. In our study, UP II mRNA was detected in 10 types of organs, including bladder, kidney, lung and pancreas, but was not detected in normal lymph nodes or leukocytes. The data indicated evidence of UP II expression in various types of normal tissues by RT-nested PCR analysis. UP II mRNA was detected in 2 of 11 (18.2%) peripheral blood samples from lung cancer patients with no metastasis, and in 5 of 12 (41.7%) peripheral blood samples of lung cancer patients with metastasis. UP II was also detected in 6 of 16 (37.5%) peripheral blood samples of patients with pancreatic cancer. The data are particularly important in that the molecular detection of micrometastasis in the blood by means of UP II mRNA identification is feasible for UP II-positive neoplasms, including lung and pancreatic cancers.

The human progesterone receptor (PR) is a ligand-activated nuclear transcription factor that mediates progesterone action in target tissues. We found a novel alternatively spliced variant (ASV) of the PR mRNA in breast cancer tissues. The deleted transcript was characterized by an out-of-frame deletion of 52 bp in exon 6 (PR delta6/2 ASV). The PR delta6/2 ASV mRNA results in a partial defect in the region of the ligand-binding domain of the hormone receptor, where conserved residues are missing from the core of the protein. To clarify the clinical significance of the PR delta6/2 ASV, we investigated the expression of this ASV in noncancerous and cancerous tissues from patients with breast cancer using RT-PCR. The novel PR delta6/2 mRNA was detected in 24 of 39 (61.5%) cancerous tissues and in 3 of 39 (7.7%) noncancerous tissues from patients with breast cancer. PR delta6/2 ASV mRNA was expressed more frequently in breast cancer tissues than in noncancerous tissues (p < 0.0001), which suggests a possible relationship between the expression of PR delta6/2 and breast cancer. Our observations may provide a novel strategy for the genetic diagnosis of breast cancer., (Copyright 2003 Wiley-Liss, Inc.)

The long-term impact of acute self-limited hepatitis B on the liver is unknown. Fourteen patients were recalled at a median of 4.2 years (range, 1.8-9.5 years) after the onset of acute hepatitis B. All showed clinical and serologic recovery with circulating hepatitis B surface antigen (HBsAg) clearance. Antibody to HBsAg (anti-HBs) had developed in 12 patients. Nine underwent liver biopsies at a median of 7.2 years, and histologic findings were evaluated using Ishak scores. Serum samples and frozen liver tissue were subjected to real-time detection polymerase chain reaction (PCR) to quantify the surface and X regions of the hepatitis B virus (HBV) genome and qualitative PCR to detect the covalently closed circular (ccc) HBV DNA replicative intermediate. Three patients had low levels of circulating HBV DNA up to 8.9 years after the onset, whereas both HBV DNA surface and X regions were found in the liver of all 9 patients examined, including 7 negative for serum HBV DNA. Liver viral loads assessed by the 2 regions showed a significant correlation (r = 0.946; P =.008), and all patients tested positive for ccc HBV DNA. Liver fibrosis and mild inflammation persisted in 8 patients. The fibrosis stage had relation to peak serum HBV DNA in the acute phase (P =.046) but not to liver viral loads in the late convalescent phase. In conclusion, occult HBV infection persists in the liver and is accompanied by abnormal liver histology for a decade after complete clinical recovery from acute self-limited hepatitis B.

We identified two novel deletion variants of the thymidylate synthase transcript in gastric cell lines. Sequence analyses indicate that none of these variants results in introduction of a premature stop-codon or a frame shifts. In 39 gastric cancer samples, both the full-length and one-deletion variant messages were detected in cancerous as well as non-cancerous tissues. However, another isoform was found in only seven of 39 cancerous tissues. Our results provide important information to assist more detailed studies on the regulation of thymidylate synthase activity.

We identified a novel alternatively spliced isoform of PR mRNA in breast cancer tissues. The deleted transcript was characterized by an out-of-frame deletion of 437 bp, corresponding to the complete loss of exons 4 and 6 (PR delta4+6 ASV). PR delta4+6 ASV will result in a partial defect in the region of the ligand-binding domain of hormone receptors, suggesting that the conserved residues are missing from the core of the protein. In the limited number of samples studied, a novel PR delta4+6 mRNA was detected in 1 of 45 (2.2%) non-cancerous tissues of patients with breast cancer, in 5 of 45 (11.1%) cancerous tissues of patients with breast cancer. Loss of both exons 4 and 6 will be induced by incomplete splicing and/or repair mechanism. Further studies are necessary to establish the biological significance of this alternative splicing. The expressions of ASVs that induced the mimic PR transcripts need to be considered when designing strategies for regulation analysis of the PR gene.

The long-term histological and virological outcomes of spontaneous circulating hepatitis C virus (HCV) clearance were studied in chronic liver disease. Between 1979 and 1984, three patients underwent laparoscopy for chronic non-A, non-B liver disease, and two were found to have cirrhosis and one with chronic active hepatitis. After HCV assays became available in 1990, they were positive persistently for HCV antibody without serum HCV RNA. Reductions of antibody levels to HCV core and/or nonstructural proteins were observed, and liver biopsies were undertaken between 1995 and 2000. Liver biopsies at 11-19 years after laparoscopy disclosed marked alleviation of liver inflammation and fibrosis in each case although a low grade of inflammation remained. The two patients with cirrhosis no longer showed histological features of cirrhosis, and the poor liver function in one patient had been ameliorated. Liver specimens from two patients were subjected to polymerase chain reaction to detect positive and negative HCV RNA strands and hepatitis B virus DNA. Only the positive HCV RNA strand was detected for one patient who had previously cirrhosis. Liver specimens were examined from another six nonviremic HCV-seropositive individuals without chronic liver disease. Five patients displayed low-grade liver inflammation without evident fibrosis, but none had any viral genome in the liver. These findings suggest that spontaneous circulating HCV clearance in chronic liver disease confers favorable liver histological outcome, although occult HCV infection persists. J. Med. Virol. 69:41-49, 2003., (Copyright 2003 Wiley-Liss, Inc.)

Genetic testings are commonly employed in various fields of clinical medicine and the test items performed at clinical laboratories are increasing rapidly in number. They are utilized to make early and/or definite diagnoses of infectious diseases, leukemia, cancers and molecular inherited diseases and also to monitor the progress of the diseases. However, these genetic testings except for infectious diseases have been developed independently at each clinical laboratory and the test results obtained at each laboratory are not always compatible each other. Under these situations it is widely expected to construct advanced genetic testing systems that can supply standardized data at any of domestic and international clinical laboratories. For the period from April, 1999 to March, 2002 three major clinical laboratories, SRL, Inc., BML, Inc. and MBC, Inc., were consigned by JBA (Japan Bioindustry Association) to collaborate in standardizing the evaluation methods for genetic testing systems among the clinical laboratories. The aim of the study is to develop the standardized genetic testing systems and to propose them as international standard operational procedures to the ISO/TC212 working group. Although one of the most important issues for standardization is the external quality assessment, they have not been carried out in reality. In this study we evaluated the difference of the genetic testing results obtained during the year of 2000 and 2001 among the clinical laboratories. The genetic testings for hematopoietic tumor, CML were selected to be evaluated since they are widely accepted as clinically useful tests.

RT-nested PCR has been introduced as a highly specific and sensitive assay method to detect the prostate-specific membrane antigen (PSM) mRNA in peripheral blood. However, appreciable percentages of false-positive cases have been reported. Additionally, primer sets reported previously could not discriminate between PSM and PSM', an alternatively spliced variant, mRNA. These isoforms can be produced from a single gene. Switches in alternative splicing patterns are often controlled with strict cell-type or developmental-stage specificity. Therefore, it is most important to discriminate between PSM mRNA and PSM' mRNA. Using our highly specific primer sets, PSM mRNA was detected in 3 of 24 peripheral blood samples of normal male volunteers (12.5%) and was not detected in peripheral blood of 11 normal female volunteers. PSM' mRNA was detected in 5 of 24 peripheral blood samples of normal male volunteers (20.8%) and in 4 of 11 of normal female volunteers (36.4%). PSM' mRNA induced false-positive results, it is important for genetic diagnosis of prostate cancer to discriminate between PSM and PSM' using our primer sets with high specificity. The advances in the uniquely designed primer sets may allow researchers to detect a real PSM mRNA without PSM' mRNA.

Mutations in the LDL receptor (LDLR) cause familial hypercholesterolaemia (FH) in an autosomal dominant manner. The condition frequently progresses to coronary atherosclerosis. We describe a patient with FH, but without ischaemic heart disease, who had a novel frameshift mutation (327insC) in exon 4 of the LDLR gene. This mutation introduced a premature termination codon (TGA, codon 158). The patient was a 59-year-old man who had presented with hypercholesterolaemia and a plasma total cholesterol (TC) concentration of 12.2 mmol/L at age 44 years. The mutation 327insC in this patient was heterozygous and hypercholesterolaemia was common within his family. Despite taking lipid-lowering medications (probucol and pravastatin) for more than 20 years, his TC concentration hardly fell below 7.8 mmol/L. However, neither the patient nor anyone else in his family developed characteristic symptoms of ischaemic heart disease or xanthoma. This patient was discovered by an intensive mutation survey among 22 unrelated Japanese with FH mainly in the Kanto area of Japan, suggesting a low incidence of the mutation in the area.

We developed a method to measure the quantity of TTV-DNA. This measurement is based on the principle of the real time PCR method using the TaqMan probe. By measuring the change of the fluorescent intensity caused by FRET, we could detect the amount of TTV-DNA. This method has the characteristics that the possibility of the contamination is very rare when it is compared with the usual PCR method, because the reaction system contains UNG and dUTP. This quantification method is useful for the future research of TTV to study the relationship between this virus and diseases.

We examined the positive rate in healthy medical workers about TT virus (TTV) which was a new hepatitis virus reported in 1997. The healthy medical workers showed a positive rate of 24%. Because there was no significant difference of the positive rate between the medical workers and general healthy persons we could not conclude that the positive rate was influenced by their profession. Generally it is known that a positive rate changes according to PCR primers used for the measurement. As for TTV as well, it is reported that a positive rate is greatly dependent on the selected region of primer. Therefore, we must pay attention to the evaluation of the positive rate for each assay system, especially using different primers.

Background: Pretransplantation injection of freshly heparinized donor blood (donor-specific blood transfusion, or DST) significantly prolongs the survival of hepatic allografts from ACI(RT1a) to LEW(RT1l) rats. We investigated hepatocyte growth factor (HGF) expression in rat hepatic allografts of recipients pretreated with or without DST., Methods: The levels of HGF mRNA and protein in hepatic allografts were determined after transplantation. The localization of HGF+ cells was identified with a rat anti-HGF monoclonal antibody., Results: Plasma HGF concentrations in transplanted rats treated with DST were significantly and persistently increased compared to untreated rats with hepatic allografts. The number of HGF+ cells in hepatic allografts of recipients pretreated with DST on day 14 was significantly greater than that in allografts of untreated recipients on day 7. HGF+ cells were also found in the marginal zone and red pulp of recipient spleens. Northern blot analysis revealed the presence of three HGF+ cell phenotypes: HGF+ED1+, HGF+ED2+, and HGF+ED1-ED2-. Most HGF+ cells were ED1-ED2-. In situ hybridization demonstrated HGF mRNA in the mononuclear cells in the portal and sinusoidal areas as well as the marginal zone and red pulp in both DST-treated and untreated recipient spleens., Conclusions: Enhanced HGF expression in rat hepatic allografts is associated with immunologic unresponsiveness induced by DST.

A 41-year-old man was given a diagnosis with of acute promyelocytic leukemia (APL) in August 1994. A chromosome analysis showed 46, XY, t(15; 17) and 47, XY, idem, +8 at that time. Because initial induction chemotherapy (BHAC-DMP) has not been successful, the patient was given 45 mg/m2 of all-trans retinoic acid (ATRA) and achieved complete remission (CR) after 26 days on this regimen. Following intensified chemotherapy, he received an autologous peripheral blood stem cell transplant (PBSCT) with high-dose busulfan and cyclophosphamide in April 1995. Competitive RT-PCR for PML-RAR alpha mRNA did not find any of APL cells in the collected stem-cell fraction. Although the patient remained in CR without therapy, a myeloblastoma was found in his left external auditory canal in August 1996. Recurrence in bone marrow, moreover, was discovered the following month. A chromosome analysis of bone marrow cells showed 47, XY, t(15; 17), +8 at this time. Thus, the extramedullary relapse developed after autologous PBSCT. This case provides information linking ATRA to the development of extramedullary relapse in patients with APL.

Adenine phosphoribosyltransferase (APRT) is a purine metabolic enzyme and a homozygous deficiency in this enzyme causes 2,8-dihydroxyadenine urolithiasis. Various germline abnormalities have been described, but we report here a unique type of germline mutation in a homozygous individual (SY) who had excreted 2,8-dihydroxyadenine crystals. In SY, TCA was substituted for the physiological stop codon TGA. This base substitution generates a new HinfI restriction site, and, using the polymerase chain reaction and subsequent digestion by this enzyme, it was confirmed that SY is homozygous for the base substitution. This base change is unique in that it generates an open reading frame that extends to the poly(A) addition site. The amount of mRNA in transformed B cells from SY was approximately a quarter of that in control subjects and no APRT proteins were detected. In eukaryotes, unlike in prokaryotes, no rescue systems for defective polypeptide termination caused by a missing stop codon have been found. Therefore, the outcome of the defect of SY is unclear from present knowledge about termination of polypeptide synthesis. Investigations into the mechanisms of the absence of protein in the cells of SY may lead to a better understanding of the physiological and nonphysiological termination of polypeptide synthesis in eukaryotic cells.

Phenotypic conversion from acute myeloid leukemia (AML) to acute lymphoblastic leukemia (ALL) is rare. A 38-year-old man was initially diagnosed as having AML (FAB-M2) associated with the t(8;21)(q22;q22) chromosomal abnormality. The blasts showed myeloperoxidase (MPO) activity and CD13 antigen expression. He showed complete remission after standard chemotherapy for AML. However, the patient relapsed with blasts showing ALL morphology (FAB-L1), MPO negativity, and CD19 antigen expression 33 months after cessation of AML therapy. Cytogenetic analysis at relapse was unsuccessful. Molecular analysis of ALL blasts revealed immunoglobulin heavy-chain gene and MLL gene rearrangements but no AML1 gene. MLL gene rearrangement or the 11q23 chromosomal abnormality has been associated with therapy-related leukemia. The subsequent ALL in our patient may have been induced by the chemotherapy including daunorubicin, known as a topoisomerase II inhibitor.

We studied the prevalence of GB virus C/hepatitis G virus (GBV-C/HGV) infection among 112 patients with liver disease and 121 blood donors in Ulaanbaatar, Mongolia. Reverse transcription and polymerase chain reaction were employed to detect GBV-C/HGV RNA using the specific primers derived from the 5'-untranslated region (5'-UTR) of the GBV-C/HGV genome. Nucleotide sequences of all positive samples for GBV-C/HGV RNA were determined. The sequences were analyzed by a molecular evolutionary method. Twenty-five (10.7%) of 233 people were positive for GBV-C/HGV RNA. Eight (6.6%), 11 (9.1%), and 30 (24.8%) blood donors were positive for GBV-C/HGV RNA, HBsAg, and anti-HCV, respectively, although 17 (15.2%), 65 (58.0%), and 64 (54.5%) patients with liver disease were positive for each viral marker. The prevalences of GBV-C/HGV RNA, HBV, and HCV in the patients were significantly higher than those in blood donors (P < 0.05). There was no significant difference in the prevalence of anti-HCV among people with and without GBV-C/HGV RNA, while the prevalence of HBsAg among people with GBV-C/HGV RNA was significantly higher than among those without GBV-C/HGV RNA (P < 0.05). The molecular evolutionary tree showed that GBV-C/HGV was a heterogeneous virus and all strains could be divided into 2 types. One is the same phylogenetic type as HGV, and the other is a new type that is different from GBV-C and HGV.

The nucleic acids of the helical and coccoid forms of Helicobacter pylori were studied to determine if the coccoid forms are "viable (capable of growing) but nonculturable." Using a reference strain (NCTC 11638) and five clinical strains, the nucleic acid contents, DNA integrity, and results of PCR and reverse transcription-PCR (RT-PCR) were compared for helical H. pylori and coccoid forms induced using glycochenodeoxycholic acid or bismuth citrate. The DNA and RNA contents of the coccoid forms were respectively 6.8- and 8.1-fold lower than those of helical H. pylori after 3 days of induction and 11.5- and 14.7-fold lower after 7 days. Agarose gel electrophoresis of DNA extracted from the coccoid forms after 3 days of induction showed a smear pattern indicating DNA cleavage, whereas DNA from helical H. pylori showed a single band with a high molecular mass. After 12 days of induction, all RNA samples from 100% coccoid cultures were negative for the mRNA of urease A or the 26-kDa species-specific protein by RT-PCR. However, most RNA samples obtained after 3 or 7 days of induction were positive at low levels despite the lack of recovery from these cultures. These results suggest that the coccoid form of H. pylori has impaired genomic DNA and is in the process of cellular degeneration, thus being still alive but nonincreasable.

A 18-year old female with acute myelogenous leukemia (AML), M2 had translocation: t(6;9) (p23; q34). The patient entered into hematological complete remission after two courses of BHAC-DMP chemotherapy with disappearance of cytogenetic abnormality. However, minimal residual disease (MRD) detected with DEK/CAN chimeric m-RNA by reverse transcription polymerase chain reaction (RT-PCR) was continuously observed, although decreased quantitatively, following several courses of consolidation and intensification chemotherapies. MRD was detected also in the harvested peripheral blood stem cells (PBSC). Leukemia relapsed with the reappearance of t(6;9) 2 months after the subsequent peripheral blood stem cell transplantation (PBSCT). Leukemia became refractory to chemotherapy, and the patient died 5 months thereafter.

Myotonic dystrophy (DM) is genetically characterized by abnormal expansion of an unstable CTG trinucleotide repeat, located in the 3'-untranslated region of mRNA encoding the family of serine-threonine protein kinases. DNA extracted from various organs of patients with DM was analyzed by the Southern blotting method. We identified differently expanded bands in DNAs from various tissues from patients with DM. In studying the length of the CTG repeat in different regions of the brain, we found a noticeably small increase in repeat length in the cerebellum compared with other tissues. While this phenomenon has been reported in other triplet repeat diseases such as Huntington disease, spinocerebellar ataxia type 1, and dentatorubral-pallidoluysian atrophy, we are the first to describe it in DM. Although the mechanism of expansion of the triplet repeat remains to be defined, the tissue-dependent somatic mosaicism suggests that its occurrence may depend on the differentiated state of each tissue.

In order to detect herpes virus group DNA including that of the Epstein-Barr virus (EBV) in patients with Vogt-Koyanagi-Harada disease (VKH), the authors employed the polymerase chain reaction (PCR) procedure using DNA from cerebrospinal fluid (CSF) obtained from patients with VKH. Method. Seven CSF samples were obtained from six definite, active VKH cases and DNA was isolated. DNA fragments containing parts of herpes simplex virus (HSV), herpes zoster virus (VZV), cytomegalo virus (CMV), EBV and human herpes virus type 6 (HHV-6) sequences were amplified by PCR. Results. No DNA fragment corresponding to the DNA sequence of the herpes virus group was detected. Conclusion. Our results suggest that the herpes virus group does not have a close association with the cause of VKH.