Successive continuous hemodiafiltrations for severe acute pancreatitis in a patient with chronic renal failure.
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Biomedical subjects
Publications and source records attributed to H Nakao.
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The prevalence of hepatitis B virus (HBV), hepatitis C virus (HCV), and GB virus C or hepatitis G virus (GBV-C/HGV) infections was determined in 289 patients with liver disease in Ho Chi Minh City and 890 healthy inhabitants of its rural area, Dalat City, Vietnam, respectively. Serum HCV RNA and GBV-C/HGV RNA were detected by reverse transcription-polymerase chain reaction (RT-PCR). HBsAg, HCV antibodies, and GBV-C/HGV RNA were detected in 139 (47%), 69 (23%), and ten (3%) subjects, respectively, often accompanied by elevated serum levels of alanine aminotransferase. HBsAg and HCV antibodies or HCV antibodies and GBV-C/HGV RNA were detectable simultaneously in 8% and 2% of the patients, respectively. In the inhabitants, HBsAg, HCV antibodies, and GBV-C/HGV RNA were found in 51 (5.7%), nine (1.0%), and 11 (1.2%) subjects, respectively. Thus, the prevalence of HBsAg, HCV antibodies, and GBV-C/HGV RNA was significantly higher in liver disease patients than those in the general population. In the samples from 69 patients and nine inhabitants who were seropositive for HCV antibodies, HCV RNA was detectable in 42 (61%) and 4 (44%), respectively. In patients with liver disease, ten belonged to HCV genotype 1a, ten to HCV 1b, three to HCV 2a, four to HCV 2b, and two to HCV 3a by PCR with genotype-specific primers. Nine patients had mixed genotypes, and the remaining four were not classified. Of the GBV-C/HGV RNA-positive individuals, two patients and two inhabitants were positive for HBsAg, while none of the residents had HCV antibodies, although six HCV antibodies (60%) and four HCV RNA (40%) were found in patients. When a phylogenetic tree of GBV-C/HGV was constructed based on the nucleotide sequences, the 21 isolates were classified into at least two genotypes; four isolates belonged to G2, and 17 to G3. The results indicate that in Ho Chi Minh HCV infection prevails with broad distribution of genotypes together with HBV infection among patients with liver disease. This study suggests that GBV-C/HGV infection occurs independently in the two different districts in association with HCV infection.
A 63-year-old female patient with obstruction of left main bronchus due to recurrent esophageal cancer was treated by emergency Nd-YAG laser therapy under bronchoscopy. Severe dyspnea subsided dramatically and she was the given radiotherapy with a total dose of 50 Gy (2 Gy/f and 25 f/5 wks). Concurrent chemotherapy was performed at the 3rd week of radiation therapy. In this chemotherapy of CDDP plus 5-FU, CDDP (10 mg/day) was given for 5 days by intravenous and 5-FU (500 mg/day) for 5 days by continuous infusion the same week. By this treatment, a partial response (PR) was obtained, and the patient returned to normal life. But after 4 months, she had a recurrent lesion at the same place, and underwent only palliative laser therapy. Nd-YAG laser therapy for obstructive lesion of trachea due to recurrent cancer is the most useful one, but some subsequent treatment is required.
A patient on maintenance hemodialysis was infected with a recently discovered putative non-A to -E hepatitis virus designated GB virus C (GBV-C) or hepatitis G virus (HGV) by transfusion. The viral isolate was recovered from the patient soon after she turned positive for GBV-C/HGV RNA in serum (GS185) and 8.4 years thereafter (GS193), and the entire nucleotide sequences were determined. They both had a genomic length of 9391 nucleotides with a defective C gene made of only 42 nucleotides. Between GS185 and GS193, 31 (0.33%) nucleotides were different, which changed 5 (0.18%) of the encoded 2842 amino acids. Thus, GBV-C/HGV was estimated to have a mutation rate of 3.9 x 10(-4) base substitutions per site per year. Nucleotide conversions were distributed over subgenomic regions, except in the 5' untranslated region of 552 nucleotides and a defective short C gene, which were conserved in sequence. The change in the putative envelope genes (E1 and E2) was no different from that in the entire genome with only 6 (0.35%) nucleotide substitutions among the 1730, just 1 of which induced an amino acid conversion. Taken along with the comparison of the two isolates with the reported five GBV-C or HGV isolates, these results indicate that GBV-C/HGV would not have hypervariable regions and would use a strategy for viral persistence that is different from immune escape.
A novel ST (Y-STc) produced by a pathogenic strain of Yersinia enterocolitica was recently purified and the mature toxin was found to include the pro-region-like sequence (Yoshino et al., FEBS Lett 362: 319-322, 1995). In the present study, we report the nucleotide sequence of the ystC gene. We also present the amino acid sequence of a ystC gene product secreted by a recombinant strain of Escherichia coli. From the nucleotide sequence of ystC gene, the Y-STc precursor protein was deduced to consist of 72 amino acid residues and to be comprised of two regions (pre- and mature toxin regions), while the known ST precursor proteins are comprised of three regions (pre-, pro-, and mature toxin regions). Therefore, we concluded that a long polypeptide corresponding to a pro-toxin of other Sts is secreted as the Y-STc mature toxin. This implies that the proteolytic processing of a pro-form appearing in the periplasm is not essential for the secretion of Yersinia-ST to the extracellular through the outer membrane. The recombinant Y-STc released by E. coli was found to be shorter and composed of C-terminal 24 amino acid residues of the native Y-STc. On the basis of these results, we propose that the cleavage site between the pro- and mature toxin regions of ST depends on both the amino acid sequence of the pro-toxin and on the bacterial protease system responsible for the maturation.
The activity of aconitate isomerase, which is involved in the biosynthesis of trans-aconitic acid as antifeedant in brown planthoppers, was detected in Pseudomonas fluorescens LRB3W1 and Pseudomonas putida MAFF301685 but not in Pseudomonas putida MAFF301684. The enzyme activity was induced in the presence of trans-aconitate, and therefore bacteria showing the enzyme activity were easily detected by their ability to grow on the minimal medium containing trans-aconitate as the sole carbon source (ACO agar medium). Experiments on growth of plant- or insect-associated bacteria on ACO agar medium showed that most of the Gram-negative bacteria displayed the aconitate isomerase activity unlike most of the Gram-positive bacteria isolated mainly from insects. Mini-Tn5 transposon derivatives of P. fluorescens LRB3W1 lacking completely or partially their ability to grow on ACO agar medium were obtained. Southern blot analysis with a mini-Tn5 DNA probe definitely showed that the genes responsible for the biosynthesis of aconitate isomerase present on chromosomal DNA. Thus, it was suggested that genes for aconitate isomerase biosynthesis are commonly present in Gram-negative plant- or insect-associated bacteria, and also the DNA fragments including the genes were detected in P. fluorescens LRB3W1.
The largest diphtheria outbreak in the developed world since the 1960s is in progress in the Russian Federation. Seventy-two Corynebacterium diphtheriae strains from throughout Russia and the Ukraine, selected for temporal and geographic diversity, and 6 reference and control strains were assayed by DNA direct sequencing, and DNA sequences of their diphtheria toxin gene, tox, and the regulatory dtxR gene, were compared to those of the Park-Williams no. 8 strain (PW8). Twenty-eight C. diphtheriae strains had entire tox sequences identical to that of the PW8 strain. Among the remaining 40 strains which were toxigenic, 4 point mutations were detected in the tox gene, one within the A and three within the B subunit gene. All four were silent mutations, indicating that diphtheria toxin is highly conserved at the amino acid sequence level; therefore, changes in the efficacy of the current vaccines would be unlikely to occur. Within the open reading frame of the regulatory dtxR gene, 35 point mutations were detected. Only 15 strains had entire dtxR sequences identical to that of the PW8 strain. Nine amino acid substitutions were found in the carboxyl half of dtxR: 22 and 25 strains differed from the PW8 strain in one and two amino acids, respectively. Given that naturally occurring variations of dtxR might be associated with increased diphtheria toxin production, studies to investigate the association of these point mutations and amino acid substitutions with quantified toxin production in the strains causing the current epidemic are under way.
BACKGROUND: The purpose of the study was to survey the epidemiology of recently reported non-A through -E hepatitis virus designated hepatitis G virus (HGV) and its strain variant, the GB agent (GBV-C). STUDY DESIGN AND METHODS: Pilot samples from 2461 blood donors in Japan, randomly selected to form cohorts with different levels of alanine aminotransferase (ALT) and markers of hepatitis B virus or hepatitis C virus (HCV) infection, were tested for RNA of HGV/GBV-C by reverse transcription-polymerase chain reaction with nested primers deduced from the 5'-noncoding region. RESULTS: HGV/GBV-C RNA was detected in 23 (7.4%) of the 361 donors with anti-HCV and HCV RNA. This detection is more frequent than that in donors without elevated ALT levels (< or = 45 U/L) or markers of HCV or hepatitis B virus infection (15/1303; 1.2%) (p < 0.001), donors with ALT values between 46 and 99 U per L (0/108) (p < 0.002), donors with ALT values > or = 100 U per L (5/361; 1.4%), and donors with anti-HCV but without detectable HCV RNA (1/93; 1.1%) (p < 0.05). CONCLUSION: More than 1 percent of Japanese blood donors were infected with HGV/GBV-C, and the prevalence was much higher in those with HCV RNA. Should persistent infection with HGV/GBV-C induce any hepatotoxic sequelae, either alone or in concert with the other hepatitis viruses, screening of blood units for HGV/GBV-C would deserve consideration.
RNA of a putative non-A, -B, -C, -D, or -E hepatitis virus named GB virus C (GBV-C) was detected by reverse transcription-polymerase chain reaction with primers deduced from the 5' untranslated region in 15 (24%) of 63 men with hemophilia in Japan at a frequency higher (P < .001) than that in 2 (0.6%) of 337 controls. By phylogenetic analysis, GBV-C isolates from some patients were similar in sequence, indicating infection with closely related strains, and those from certain patients resembled sequences reported from foreign countries. All patients were infected with hepatitis C virus, and genotypes that are rare in Japan were detected in 36 (57%) of them. These results indicate that patients with hemophilia in Japan would be at increased risk for infection with GBV-C and hepatitis C virus, some of which would have been transmitted via imported coagulation factor concentrates in the past.
Recently, putative viral agents responsible for human non-A to E hepatitis have been independently reported by two groups of investigators and designated GB virus C (GBV-C) and hepatitis G virus (HGV), respectively. The entire nucleotide sequences were determined for two viral genomes isolated from Japanese blood donors with GBV-C RNA. One of them (GT230) had a total genomic length of 9390 nucleotides (nt) with 5' and 3' untranslated regions of 551 and 313 nt, while the other (GT110) had genomic lengths of 9395, 281 and 315 nt, respectively. They both had a single long open reading frame, encoding 2842 amino acids (aa) in GT230 and 2933 aa in GT110. Surprisingly, they both lacked a clearly identifiable core gene, and possessed the E1/E2 gene with only four potential N-linked glycosylation sites. Pairwise comparison and phylogenetic analysis of the entire sequence indicated that the prototype GBV-C and two HGV isolates reported, as well as GT230 and GT110, are the same virus possibly of different genotypes. The five GBV-C/HGV isolates were variable up to 13.8% in the genomic nucleotide sequence, and contained deletions and insertions within the 5'-terminal 518-593 nt, which resulted in four different sizes of predicted polyproteins encoded by genomes of individual isolates. By contrast, the 3' untranslated region was well conserved. The high degree of sequence conservation within this region would favour it as a target for sensitive detection of GBV-C/HGV RNA.
PCR has proved to be a reliable tool for the detection of the diphtheria toxin gene, tox, and its use has allowed for the rapid differentiation between toxigenic and nontoxigenic strains. In this study, this PCR was further developed, evaluated, and standardized to detect this gene directly from clinical specimens. Optimal conditions for collection, transport, and storage of the clinical specimens and isolation and purification of DNA from the clinical specimens were defined. With two sets of primers that detect the A and B subunits of the diphtheria toxin gene, sensitivity levels of 50 and 500 CFU/PCR mixture, respectively, were achieved. This PCR was evaluated with 162 clinical samples collected from patients with diphtheria and other upper respiratory tract infections, as well as from healthy individuals.
Gnathostoma doloresi parasitizes the gastric wall of wild (boars) and domestic (pigs) swine (Sus scrofa). Its larvae cause cutaneous larva migrans in humans. Amphibians, reptiles and a freshwater fish are infected with the advanced 3rd stage larvae. Prevalence of G. doloresi larvae were surveyed in several snakes, especially in a common frog-eating snake (Rhabdophis tigrinus). All species of snakes examined were infected with G. doloresi larvae suggesting that snakes are important reservoir hosts. Prevalence of G. doloresi larvae in frog-eating snakes was lower than that found in mammal-eating snakes. Thus, as a source of infection to snakes, small mammals may be more important than frogs in the natural life cycle of G. doloresi in Japan.
The molecular mechanism underlying the interaction between myeloma cells and stromal cells was investigated by using a human myeloma cell line (OPM-2) and human umbilical vein endothelial cells (HUVECs). Adhesion of OPM-2 cells to HUVECs was found to be significantly augmented with treatment of OPM-2 cells with an alpha-glycosidase inhibitor, castanospermine (CSP). The treatment of OPM-2 cells with CSP resulted in alteration of oligosaccharide structures of cell surface glycoproteins particularly at molecular weight of 220 kD (GP220). To determine if GP220 was involved in the adhesion of OPM-2 cells to HUVECs, cell surface glycoproteins of HUVECs were labeled by biotin and were incubated with the PVDF membrane to which cell surface glycoproteins of OPM-2 cells were blotted. The biotinylated glycoproteins at the plasma membrane of HUVECs specifically bound to GP220 of OPM-2 cells. Purification and partial amino acid sequencing of GP220 revealed that GP220 had a structure homologous to cation-independent mannose 6-phosphate/insulin-like growth factor-II (CIM6P/IGF-II) receptor. Furthermore, an antibody against CIM6P/IGF-II receptor was reactive with GP220, indicating that GP220 was a CIM6P/IGF-II receptor. The adhesion of OPM-2 cells to HUVECs was inhibited by mannose 6-phosphate. Moreover, M6P was found to suppress the adhesion of human myeloma cell lines, OPM-2 and RPMI 8226, to bone marrow stromal cells that was established from the patients with multiple myeloma. In addition, proliferation of OPM-2 was stimulated in response to IGF-II. These results suggest that CIM6P/IGF-II receptor may be functional in terms of supporting cell adhesion and proliferation of myeloma cells.
Diabetes-prone (DP) BB rats (RT1(u), RT6.1) spontaneously develop insulin-dependent diabetes mellitus (IDDM) and the disease manifestation resembles that in human IDDM. DP rats are immunodeficient with severe T lymphocytopenia due to the absence of T cells expressing the RT6 differential alloantigen, which have immunoregulatory functions. MHC- and non-MHC-compatible Wistar Furth (WF; RT1(u), RT6.2) pancreases were transplanted into DP rats. WF pancreas grafts were destroyed by IDDM recurrence (insulitis), but not by rejection, with a mean survival time of 65.3 +/- 21.7 days. To prevent the recurrence of IDDM in the grafts, monoclonal antibodies to intercellular adhesion molecule-1 and leukocyte function-associated antigen-1 were administered. WF pancreas grafts were indefinitely accepted (>108.0 +/- 26.8 days) in monoclonal antibody-treated DP recipients. The number of T cells was increased and cellular immune responses restored only in the DP rats that had accepted grafts. The increased number of T cells was due to the peripheral appearance of donor-type RT6.2+ T cells, which represented 34.3 +/- 7.0% of total splenic T cells. The cytotoxicity of splenic T cells to WF islet cells was suppressed in the presence of RT6+ T cells in vitro. These findings demonstrated that stable macrochimerism of donor-derived RT6+ T cells could restore the immune responses and prevent the recurrence of IDDM in the DP recipients.
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The detection of hepatitis C virus (HCV) in blood donors and patients with acute and chronic hepatitis has brought to the fore another virus or viruses which can be transmitted parenterally and induce liver disease. The RNA of a candidate virus designated GB virus C (GBV-C) was determined by the polymerase chain reaction with primers deduced from a helicase-like region in 229 leprous patients in Japan. GBV-C RNA was detected in 12 (5.2%) patients, and HCV RNA in 41 (18%). Three patients were coinfected with GBV-C and HCV. The nine patients infected with GBV-C alone had aminotransferase levels lower than the three patients with the mixed infection or the 38 patients infected with HCV only (P < 0.001). Sequence comparison within 100 base pairs in the helicase-like region suggested that two, three and three patients, respectively, would have been infected with three distinct strains of GBV-C. These results indicate that patients with leprosy are at increased risk for infection not only with HCV, but also with GBV-C, and that the infection with GBV-C alone would not induce hepatic injuries as severe as HCV infection.
We sequenced the entire genome of an Italian isolate of hepatitis C virus: the first full-length sequence for the genotype 2c. We report hereby its characteristics and differential detection of 2c isolates using PCR.