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The new Light Cycler technology was adapted to the detection of hepatitis C virus (HCV) RNA in clinical samples. Sera from 81 patients were tested by Light Cycler PCR, AMPLICOR HCV Monitor assay, and in-house PCR. Our data demonstrate that Light Cycler is a fast and reliable method for the detection and quantitation of HCV RNA.

Background: Genotypic resistance of Hepatitis B virus (HBV) against lamivudine evolves within months after onset of therapy., Objectives: To determine the longitudinal order in which resistance mutations appear and to compare the kinetics and pathogenicity of wild-type and resistant HBV., Study Design: In a longitudinal study, consecutive samples were drawn over a period of 28 months from a patient with chronic hepatitis B, and resistance mutations were followed by sequencing a part of the polymerase region of HBV. These data were compared with HBV copy numbers, HBsAg and ALT levels, and results of consecutive liver biopsies., Results: After 21 weeks of treatment, a silent mutation at codon 528 (CTG to TTG) occurred. Significant genotypic resistance was detectable after 68 weeks, indicated by a substitution of isoleucine for methionine at residue 552 (M552I). Nineteen weeks later, the virus exhibited additional resistance-associated mutations (L528M and I552V). The resulting high-level resistance was reflected by an increase of serum HBV copies of 4.7 log(10). The turnover of wild-type and resistant HBV was 2.6x10(6) and 1.8x10(6) virions/day, respectively. HBsAg and ALT levels were lower within the period when resistant HBV was detectable. During treatment the progress of liver fibrosis was arrested., Conclusions: The in vivo replicative capacities and dynamics of wild-type and resistant HBV were similar. However, resistant HBV seemed to exhibit reduced pathogenicity.

Positive results by cytomegalovirus (CMV) PCR of plasma are considered predictive of active CMV infection in kidney allograft recipients. To assess whether contamination with leukocyte-derived CMV DNA can distort the results, aliquots of whole-blood samples from 60 CMV immunoglobulin G-positive patients with leukocyte CMV DNAemia were stored for up to 24 h at room temperature (RT) and at 4 degrees C before plasma preparation. Native and ultrafiltered plasma samples were tested by CMV and beta-globin PCRs. Among 30 latently infected patients (negative for CMV pp65 antigens), low baseline rates (10%) and levels (median number of copies, 10 [per 10 microl]) of CMV plasma DNAemia in native plasma samples increased significantly over time (after 4 h at RT, 37% [P < 0.001]; median number of copies, 45 [P < 0.001]). Similar effects were found during storage at 4 degrees C. Ultrafiltration reduced the levels of CMV plasma DNAemia, but by 6 h of storage the levels were significantly elevated as well. CMV and beta-globin DNA kinetics in plasma were parallel. In contrast, 30 actively infected patients (pp65 positive) had high baseline rates (87% in native samples) and levels (median number of copies, 75) of CMV plasma DNAemia. No significant effects of storage or ultrafiltration and no concordance with beta-globin DNA kinetics were seen. In conclusion, delayed preparation of plasma samples bears a significant risk of false-positive CMV PCR results, probably due to leukocyte lysis. This has important implications in the clinical setting and for PCR standardization.

Cytomegalovirus (CMV) cultured from peripheral blood mononuclear cells (PBMCs) was shown to be associated more closely with clinical manifestations than infectious CMV in polymorphonuclear leukocytes (PMNLs) of renal allograft recipients with secondary CMV infection. Shell vial culture was carried out with ficoll-purified PBMCs and PMNLs of 71 CMV IgG-positive patients after kidney transplantation. Thirty-six patients experienced active CMV infections. Of these, 17 developed clinical symptoms. The diagnostic value of PMNLs and PBMCs viremia was determined in comparison to pp65 antigenemia, leukoDNAemia, plasma DNAemia, and detection of cytomegalic endothelial cells. In both PMNLs and PBMCs (with or without detectable endothelial cells), frequencies and levels of viremia were significantly higher among symptomatic patients. Regarding the occurrence of clinical CMV manifestations, the sensitivity of culture from PMNLs and from PBMCs fractions was 100%. Viremia in PBMCs, however, was far more specific (94%) than in PMNLs (74%). Cutoff values established previously for pp65 antigenemia and leukoDNAemia, standard markers in the laboratory, had similar specificity (96% each) to PBMCs viremia, but were less sensitive (88% each). Plasma DNA-emia was both less sensitive (82%) and less specific (69%) than PBMCs viremia. Detection of endothelemia showed maximal specificity (100%), but inferior sensitivity (47%). All patients had PBMCs viremia before the onset of symptoms. In conclusion, infectious CMV present in PBMCs may prove to be a determinant of clinical CMV manifestations in seropositive immunocompromised individuals. Factors involved in PBMCs tropism may help to understand the pathogenetic mechanisms of CMV dissemination in this group of patients., (Copyright 2000 Wiley-Liss, Inc.)

The apoptosis-inducing Fas receptor has been shown to be down-regulated in various types of tumors, while its ligand (FasL) appears to be frequently up-regulated. Here we provide evidence that there is a strong selective pressure in vivo against Fas-expressing, tumorigenic NIH3T3 cells, favoring survival, proliferation and eventually tumor formation by Fas-negative cells. Importantly, re-expression of Fas in these cells results in either the complete abolishment of tumor development, or in a significant extenuation of the latency period of tumor outgrowth. In addition, we found that environmental conditions which prevail during tumorigenesis, such as limiting amounts of survival factors and the lack of cell adhesion, are markedly sensitizing tumor cells to Fas-mediated suicide. Our data suggest that in addition to T cell-mediated immune responses, mechanisms of Fas-dependent tissue turnover are also centrally implicated in tumor cell clearance.

Introduction: The aim of the study was to evaluate the modern principles of surgery in diverticulitis, e.g. early elective resection and primary anastomosis., Methods: The data of 445 consecutive patients were retrospectively analysed after classifying all cases in four subgroups according to a modified Hinchey classification (stages 0-III)., Results: Within our study group the morbidity was 26.5% (n = 118) and the mortality was 1.6% (n = 7). In 96% (n = 425) of all cases and in 64% (21/33) of patients with perforated diverticulitis and peritonitis (stage III), a primary anastomosis was performed. Four patients of the study group showed insufficient anastomosis (0.9%). No leakage was observed from any of the anastomoses performed in stage III diverticulitis. Stage of inflammation and age of patient correlate with morbidity and mortality. Some 53% (94/177) of the patients in stage II and 67% (22/33) of the patients in stage III had never showed symptoms of diverticulitis before., Conclusion: Prophylactic surgery to avoid life-threatening situations, including abscess formation or perforation, is not possible in many cases. However, especially patients at risk (age, coexisting illness) should undergo early surgery. Primary anastomosis can be performed safely even at an advanced stage.

To investigate the vertical transmission of the newly described TT virus (TTV), serum and breast milk samples from 46 women as well as sera from their 47 newborns were examined for the presence of TTV DNA by PCR. TTV DNA was detected in 47.8% (n = 22) of the women. All but one child born to these women were also viremic for TTV from the first sample onward. TTV DNA was found in 73.9% (n = 17) of the breast milk samples derived from TTV viremic mothers. The one TTV-negative child born to a viremic mother remained negative during follow-up, although it was breast-fed. Our data show that TTV is highly effectively transmitted from mothers to their children during pregnancy. Although the majority of breast milk samples from viremic mothers are positive by TTV PCR, there is no need to discourage women from breast-feeding, because most children are TTV viremic even before breast-feeding begins.

To evaluate the risk of hepatitis C virus (HCV) transmission via breast milk, we collected 76 samples of breast milk from 73 chronically HCV-infected women and serum samples from their 76 perinatally HCV-exposed children. Enzyme immunoassay and strip immunoblot assay were used for detection of antibodies to HCV, and reverse transcriptase-polymerase chain reaction analysis was used for detection of HCV RNA. None of the 76 samples of breast milk contained HCV RNA, whereas 37 (59.7%) of 62 mothers tested for HCV RNA had HCV viremia. Only 1 of the 76 breast-fed infants had evidence of HCV infection. Because HCV infection in this child was detected 1 month after birth, it seems unlikely that it was transmitted by breast-feeding. These results indicate that HCV infection in pregnant women should not be a contra-indication for breast-feeding.

Bis(imidazole)sulfur difluoride (4), bis(pyrazole)sulfur difluoride (6), and bis(1,2,4-triazole)sulfur difluoride (8) are formed in the reactions of N-(trimethylsilyl)imidazole, N-(trimethylsilyl)pyrazole, and 1-(trimethylsilyl)-1,2,4-triazole with SF(4), in high yield. The ring systems in these three molecules occupy equatorial positions in the pseudo-trigonal-bipyramidal coordination sphere of the central sulfur atoms. The angles between the planes of the ring substituents and the FSF axis for 4 and 6 are in the range 23.6-35.3 degrees, and in 8 the triazole rings are almost parallel (deviation 1.7 degrees ). The interaction between the heterocyclic substituents and the sulfur centers and their influence on the axial and equatorial bonds is discussed.

Serological determination of hepatitis C virus (HCV) subtypes has been hampered by the lack of suitable assays. Therefore, a recombinant immunoblot assay has been established for serological differentiation of HCV subtypes 1a, 1b, 2a, 2b, 3a, and 4a. It consists of recombinant HCV proteins from the NS-4 region propagated in Escherichia coli. To confirm the serotyping assay results, the results were compared with those obtained by nucleotide sequencing of the NS-5 region. Sera from 157 patients with chronic HCV infection were examined by this assay, and specific antibodies could be detected in 86% (n = 135) of them. The HCV genotype was determined correctly in all but one sample, and the subtypes determined by the serotyping assay corresponded to the HCV subtypes detected by nucleotide sequencing for 95% (n = 128) of the samples. These data indicate that HCV subtypes can be distinguished serologically. The assay that is described provides an easier means of identification of infection with different HCV subtypes for wider clinical and epidemiological applications.

Background: It is still unclear how many patients with hepatitis C virus (HCV) antibodies have viremia and hence are infectious., Objectives: To determine the chronicity of HCV infection by correlation of HCV antibodies with presence of viremia in long-term follow-up., Study Design: In a longitudinal study sera of 4110 patients were analyzed with second generation HCV-enzyme immunoassay (EIA) and polymerase chain reaction (PCR). Only those patients were included in this study in whom sequential serum samples over a period of 2 years were available. To avoid preanalytical and analytical failures, we used a transport solution to prevent RNA degradation and a four-antigen recombinant immunoblot assay, established in our laboratory, for confirmation of antibody reactivity., Results: Of 2815 patients with confirmed HCV antibodies 2784 (98.9%) were also positive in HCV-PCR assay. False reactive EIA results were detected in 177 (13.7%) individuals as shown by confirmatory assay and PCR. Only one patient (0.04%) spontaneously lost detectable HCV viremia and subsequently HCV-specific antibodies., Conclusions: Our study clearly demonstrates that presence of confirmed HCV-specific antibodies correlates significantly (98.9%; P < 0.001) with HCV viremia, and that spontaneous loss of viremia is a very rare event in HCV infection. We also found that elimination of HCV infection is not sufficiently predicted by the loss of detectable viremia in PCR, but can be concluded from the disappearance of virus-specific antibodies.

CD95/APO-1 ligand (FasL) is implicated in the maintenance of immune privileged sites by inducing apoptosis of activated infiltrating T lymphocytes. Therefore, progressive tumors might express high levels of FasL and develop as immune privileged sites. In this study, we investigated the expression of FasL and CD95/APO-1 (Fas, the FasL-receptor) in vitro in rat adenocarcinoma cell lines and the localization in situ in normal human kidney and colon and in their adenocarcinomas. The rat cell line PROb (a progressive tumor in vivo) expressed a higher level of FasL than the sister cell line REGb (a regressive tumor in vivo), as detected by flow cytometry. The 2 cell lines expressed the same level of Fas, but were resistant to FasL-induced apoptosis. In human tissue, both kidney and colon extracts expressed FasL by Western blot. Further investigations, using immunohistochemical staining of paraffin sections, showed that normal colon mucosa expressed Fas and FasL in crypt epithelial cells in the subnuclear compartment. Normal kidney showed Fas and FasL labeling mostly restricted to epithelial cells of proximal tubules and Henlé's loop, showing that this expression is not uniform throughout the organ. Smooth-muscle cells of muscularis propria and blood vessels in and around the tumors were also intensely but more uniformly labeled. In colon-cancer cells, FasL expression remained strong, whereas Fas expression was significantly reduced. A similar reduction in Fas expression was noted in renal-cancer cells. Tumor-infiltrating immune cells of the macrophage lineage do not express FasL. Our results show that smooth-muscle cells of muscularis propria and blood vessels are able to express FasL and to a slight extent Fas. In normal epithelial cells of colon and kidney, Fas and FasL are often co-expressed. The reduced expression of Fas in corresponding cancer cells in combination with the ability to express FasL might facilitate immune escape.

Until now there have been few seroepidemiological data for hepatitis G virus/GB virus type C (HGV/GBV-C). A four-antigen HGV/GBV-C immunoblot was established to examine 446 serum specimens from healthy individuals without risk factors for parenteral viral transmission. These individuals were divided into seven groups according to age. Seroprevalence rates were low for children and adolescents (5.6%) and increased for the age groups assumed to be the most sexually active (15.3 to 26.8%). Remarkably, none of the 80 individuals who tested positive for HGV/GBV-C antibodies were simultaneously positive for HGV/GBV-C viremia. From our data we conclude that HGV/GBV-C infection is widespread in the general population (16 to 25%). The development of an antibody response is associated with clearance of HGV/GBV-C viremia. Due to the lack of risk factors for HGV/GBV-C infection of blood, other efficient transmission routes must exist. It must be assumed that HGV/GBV-C transmission may be linked to sexual activity.

Tumor growth is the result of deregulated tissue homeostasis which is maintained through the delicate balance of cell growth and apoptosis. One of the most efficient inducers of apoptosis is the death receptor Fas. We report here that oncogenic Ras (H-Ras) downregulates Fas expression and renders cells of fibroblastic and epitheloid origin resistant to Fas ligand-induced apoptosis. In Ras-transformed cells, Fas mRNA is absent. Inhibition of DNA methylation restores Fas expression. H-Ras signals via the PI 3-kinase pathway to downregulate Fas, suggesting that the known anti-apoptotic effect of the downstream PKB/Akt kinase may be mediated, at least in part, by the repression of Fas expression. Thus, the oncogenic potential of H-ras may reside on its capacity not only to promote cellular proliferation, but also to simultaneously inhibit Fas-triggered apoptosis.

GB Virus C/Hepatitis G Virus (GBV-C/HGV) was identified recently and only two assays, consisting of a single recombinant protein, have been described for determination of the seroprevalence of this virus. An immunoblot assay was devised, which contains four recombinant GBV-C/HGV proteins. In this study, serum samples from 154 patients on maintenance hemodialysis were examined to assess the rate of seroreactivity against GBV-C/HGV. All sera were tested for the presence of antibodies by an in-house recombinant immunoblot assay, for GBV-C/HGV viremia by RT-PCR, and for HCV infection by PCR and by serological assays. Antibody reactivity against GBV-C/HGV was detected in 20.8% (n = 32) and viremia was found in 6.5% (n = 10) of the patients. In no case were viremia and GBV-C/HGV antibodies detected in parallel. HCV infection was observed in 15.6% (n = 24) by RT-PCR. In 20 of these patients, HCV antibodies were detected by enzyme immuno assay (EIA) and immunoblot assay. However, four of the HCV PCR-positive patients were negative by both serological tests. Only two patients were viremic for GBV-C/HGV and HCV in parallel. It is concluded that antibody reactivity against GBV-C/HGV is common among patients on maintenance hemodialysis. In contrast to HCV, parallel occurrence of GBV-C/HGV viremia and GBV-C/HGV seroreactivity was not observed. This suggests that GBV-C/HGV infection might be self-limiting.

The rate of false-positive hepatitis C virus enzyme immunoassay results was determined to be at least 10% among 1,814 reactive serum samples based on (i) negative results in an independent confirmation assay, (ii) negative PCR results, and (iii) no patients developing clinical or biochemical signs of hepatitis during a 1-year follow-up.

FLICE-inhibitory protein, FLIP (Casper/I-FLICE/FLAME-1/CASH/CLARP/MRIT), which contains two death effector domains and an inactive caspase domain, binds to FADD and caspase-8, and thereby inhibits death receptor-mediated apoptosis. Here, we characterize the inhibitory effect of FLIP on a variety of apoptotic pathways. Human Jurkat T cells undergoing Fas ligand-mediated apoptosis in response to CD3 activation were completely resistant when transfected with FLIP. In contrast, the presence of FLIP did not affect apoptosis induced by granzyme B in combination with adenovirus or perforin. Moreover, the Fas ligand, but not the perforin/granzyme B-dependent lytic pathway of CTL, was inhibited by FLIP. Apoptosis mediated by chemotherapeutic drugs (i.e., doxorubicin, etoposide, and vincristine) and gamma irradiation was not affected by FLIP or the absence of Fas, indicating that these treatments can induce cell death in a Fas-independent and FLIP-insensitive manner.

Members of the tumor necrosis factor (TNF) family induce pleiotropic biological responses, including cell growth, differentiation, and even death. Here we describe a novel member of the TNF family designated APRIL (for a proliferation-inducing ligand). Although transcripts of APRIL are of low abundance in normal tissues, high levels of mRNA are detected in transformed cell lines, and in human cancers of colon, thyroid, and lymphoid tissues in vivo. The addition of recombinant APRIL to various tumor cells stimulates their proliferation. Moreover, APRIL-transfected NIH-3T3 cells show an increased rate of tumor growth in nude mice compared with the parental cell line. These findings suggest that APRIL may be implicated in the regulation of tumor cell growth.

Background: Screening for occult blood by means of guaiac tests has an unsatisfactory sensitivity for the detection of colorectal neoplasms. To increase sensitivity and specificity the immunological determination of human hemoglobin and albumin in feces has been developed. The validity of analyzing only two samples from one bowel movement of either test is not known., Methods: An immunological determination of human fecal hemoglobin and albumin using luminescence immunoassays (LIA) was performed in 739 patients with gastrointestinal complaints before scheduled colonoscopy. Each patient collected two 1 ml samples from one stool. There were no dietary restrictions., Results: The sensitivity for detecting colorectal carcinomas was 95.3% (95% confidence interval 84.2-99.4%) with hemoglobin and 67.4% (95% confidence interval 51.2-80.9%) with albumin. The sensitivity for detecting large adenomatous polyps was 62.9% (95% confidence interval 50.5-74.1%) with hemoglobin and 32.9% (95% confidence interval 22.1-45.1%) with albumin. The specificity was 97% for hemoglobin, 96% for albumin and 94% for the combined test., Conclusions: The immunological determination of fecal hemoglobin is superior to albumin and has a better sensitivity for the detection of colorectal neoplasms than that reported for guaiac tests, even if two samples from one bowel movement are examined. The immunological determination of fecal hemoglobin should therefore be evaluated for use in colorectal cancer screening.

Until now, no serological assay has been available for the differentiation of HCV subtypes. Since there is evidence that the subtypes differently influence the clinical course of HCV infection and the outcome of interferon therapy, we established a strip immunoblot assay (NS-4 IBA) with recombinant HCV proteins of the nonstructural 4 (NS-4) region propagated in Escherichia coli. Using this NS-4 IBA, we were able to distinguish HCV subtypes 1a and 1b, which are the most prevalent subtypes in Europe and the U.S.A. The results of the serotyping assay were compared with those obtained by nucleotide sequencing from the NS-5 region. Concordant results were observed to match 94.9% (n=100) by the NS-4 IBA and nucleotide sequencing. Discrepant results were obtained in only 5.1% (n=6). These data indicate that HCV subtypes can be serologically distinguished, providing the possibility for easier identification of infection with different HCV subtypes.

The short-term kinetics of infectious HIV titers, HIV copy numbers and p24-antigen during the first 28 days of AZT monotherapy were evaluated. In three of four patients, infectious HIV was culturable and infectious titers rose 2- and 4-fold compared to baseline values. This increase was neither associated with mutations conferring resistance to AZT nor a switch from NSI to SI phenotypes. Two patients showed an increase of plasma infectivity associated with a reduction of HIV copies and p24-antigen. We conclude that transient dissociations of plasma infectivity and HIV copy numbers occur during early AZT monotherapy.

Normal or perforin-deficient C57BL/6 mice are able to heal spontaneously cutaneous lesions induced by Leishmania major. In this report, we show that syngeneic gld and lpr mice, lacking a functional Fas system, fail to heal their lesions. This inability to control infection could not be attributed either to a failure to mount a protective CD4+ Th1 response or to an unresponsiveness of their macrophages to the activating signals of IFN-gamma. The observation showing that administration of exogenous recombinant Fas ligand (FasL) to FasL-deficient mice resulted in the resolution of cutaneous lesions demonstrates the importance of the Fas-FasL pathway in the elimination of parasites. Furthermore, macrophages infected with L. major in vitro up-regulate their surface expression of Fas in response to IFN-gamma and as a result become susceptible to CD4+ T cell-induced apoptotic death. These results suggest that the CD4+ T cell-induced apoptotic death of MHC class II-expressing antigen-presenting cells, observed in vitro and operating through the Fas (Apo-1/CD95) pathway, is relevant in vivo.

A new virus named hepatitis G virus (HGV) has been detected recently. Until now, no assays for the detection of antibodies against different HGV proteins have been commercially available. Therefore, a strip immunoblot assay has been established to investigate seroreactivity against recombinant structural (core) and nonstructural proteins (NS3 and NS4) of HGV produced in Escherichia coli. Seropositivity for HGV was evaluated and concordanced with HGV polymerase chain reaction (PCR) results in 709 subjects. These individuals were classified into a nonrisk or a risk group, on the basis of infection with human immunodeficiency virus (HIV) or hepatitis C virus (HCV) or frequent parenteral exposure, including hemophilia, intravenous drug addiction, receipt of blood transfusion, or hemodialysis. The nonrisk group consisted of 257 healthy blood donors with normal alanine transaminase (ALT) levels (ALT < 30 U/L) and 154 patients with suspected non-A-E hepatitis (ALT > 45 U/L). In the group of healthy blood donors, 1.9% (5 of 257) had detectable HGV viremia and 15.9% (41 of 257) showed antibody response to HGV. In the collective of patients with suspected non-A-E hepatitis, results from 1.9% of patients (3 of 154) were positive by HGV PCR, and 15.6% of patients (24 of 154) showed seropositivity against the recombinant HGV proteins. In six groups of patients (n = 298) with different risk factors, the prevalence of both HGV viremia (V) and serological reactivity (SR) was higher compared with that of the nonrisk group: V, 6.80%-35.2%; serological reactivity (SR), 25.4%-52.9%. The following conclusions can be derived from our data. HGV infection is widespread in the general population. The prevalence of antibodies against HGV or detectable HGV viremia is higher in patients with risk factors for parenteral viral transmission than in those without risk factors. The majority of HGV infections (70.2%) is self-limiting and not persistent in our collective of patients. We found no correlation between HGV viremia and clinical or biochemical signs of hepatitis in individuals without risk factors for acquiring parenterally transmitted agents.

The widely expressed protein Fas is a member of the tumour necrosis factor receptor family which can trigger apoptosis. However, Fas surface expression does not necessarily render cells susceptible to Fas ligand-induced death signals, indicating that inhibitors of the apoptosis-signalling pathway must exist. Here we report the characterization of an inhibitor of apoptosis, designated FLIP (for FLICE-inhibitory protein), which is predominantly expressed in muscle and lymphoid tissues. The short form, FLIPs, contains two death effector domains and is structurally related to the viral FLIP inhibitors of apoptosis, whereas the long form, FLIP(L), contains in addition a caspase-like domain in which the active-centre cysteine residue is substituted by a tyrosine residue. FLIPs and FLIP(L) interact with the adaptor protein FADD and the protease FLICE, and potently inhibit apoptosis induced by all known human death receptors. FLIP(L) is expressed during the early stage of T-cell activation, but disappears when T cells become susceptible to Fas ligand-mediated apoptosis. High levels of FLIP(L) protein are also detectable in melanoma cell lines and malignant melanoma tumours. Thus FLIP may be implicated in tissue homeostasis as an important regulator of apoptosis.

Viruses have evolved many distinct strategies to avoid the host's apoptotic response. Here we describe a new family of viral inhibitors (v-FLIPs) which interfere with apoptosis signalled through death receptors and which are present in several gamma-herpesviruses (including Kaposi's-sarcoma-associated human herpesvirus-8), as well as in the tumorigenic human molluscipoxvirus. v-FLIPs contain two death-effector domains which interact with the adaptor protein FADD, and this inhibits the recruitment and activation of the protease FLICE by the CD95 death receptor. Cells expressing v-FLIPs are protected against apoptosis induced by CD95 or by the related death receptors TRAMP and TRAIL-R. The herpesvirus saimiri FLIP is detected late during the lytic viral replication cycle, at a time when host cells are partially protected from CD95-ligand-mediated apoptosis. Protection of virus-infected cells against death-receptor-induced apoptosis may lead to higher virus production and contribute to the persistence and oncogenicity of several FLIP-encoding viruses.

The distribution of hepatitis C virus (HCV) genotypes was determined in isolates of 447 chronically HCV-infected German patients by nucleotide sequencing. Of these, 206 (46.1%) were infected with the subtype 1a, 215 (48.1%) with subtype 1b, 2 (0.4%) with subtype 1c, 9 (2.0%) with subtype 3a, and 15 (3.4%) with subtype 4a. Subtype 1a was predominant in those < 40 years old (62.6%) and was associated with the risk factor of intravenous drug addiction and with shorter duration of disease. Conversely, subtype 1b was more frequent in patients > 50 years old (84.7%; P < .001) and was associated with the risk factor of blood transfusions and with longer duration of disease. These data suggest that a shift from subtype 1b to subtype 1a occurred in the population studied. An increase in HCV infection with subtype 1a and a diminution of subtype 1b in the future can be expected.

Among patients undergoing maintenance hemodialysis, a high prevalence of hepatitis C virus (HCV) infection can be observed. In a prospective study, sera of 273 patients were examined for the presence of HCV infection by serological tests and by PCR. Thirty-five patients (12.8%) were HCV antibody positive, and in 31 of them HCV RNA could be detected by PCR. Among the 238 seronegative patients HCV infection was detected in 12 cases (5.0%) exclusively by PCR. Only in 1 of these patients seroconversion could be observed within the 18-month follow-up period. These findings demonstrate that in hemodialysis patients PCR is necessary for the diagnosis of HCV infection.

A novel member of the tumor necrosis factor (TNF) receptor family, designated TRAMP, has been identified. The structural organization of the 393 amino acid long human TRAMP is most homologous to TNF receptor 1. TRAMP is abundantly expressed on thymocytes and lymphocytes. Its extracellular domain is composed of four cysteine-rich domains, and the cytoplasmic region contains a death domain known to signal apoptosis. Overexpression of TRAMP leads to two major responses, NF-kappaB activation and apoptosis. TRAMP-induced cell death is inhibited by an inhibitor of ICE-like proteases, but not by Bcl-2. In addition, TRAMP does not appear to interact with any of the known apoptosis-inducing ligands of the TNF family.

A compartment model of the transport of oxygen from the alveoli to the tissues is described. In patients with both pulmonary shunt and alveolar resistance to diffusion of oxygen, the model is used to simulate their response to variations in the inspired oxygen fraction. These simulation results are compared to the responses from a patient with respiratory malfunction, indicating that the method can identify patients where not only a pulmonary shunt but also a high alveolar resistance to diffusion of oxygen is clinically significant. Estimation of pulmonary shunt and oxygen diffusion resistance can be done in two different implementations of the model. In the first implementation the estimates are generated by numerical solution of the equations of the compartment model. In the second implementation the equations have been used to construct a causal probabilistic net where biological uncertainties and uncertainties in the measurements can be represented.

To determine the distribution of hepatitis C virus (HCV) genotypes in German isolates, nucleotide sequences of the viral nonstructural 5 (NS5) genome domains were analyzed in isolates from 107 chronically HCV-infected patients. Of these 107 patients, 46 (43.0% were infected with subtype 1a and 47 (43.9%) with subtype 1b. Six patients (5.6%) with a history of intravenous drug abuse were infected with subtype 3a. Eight patients (7.5%) who had acquired their HCV infection in Egypt carried subtype 4a. Forty-three of the 107 patients were treated with alpha-interferon. Of these 43 patients, 16 (37.2%) were infected with subtype 1a and 27 patients (62.8%) with subtype 1b. Three patients infected with HCV-subtype 1a (18.7%) and four patients infected with subtype 1b (14.8%) showed a sustained complete response after interferon therapy. The HCV genotype 1 with its subtypes 1a and 1b was the most common source of HCV infection in this group of patients. There was no significant difference in response to alpha-interferon treatment of HCV infection with the subtypes 1a or 1b.

Cytotoxic T cells (CTL) induce cell death of their target cells either by the surface interaction between Fas ligand and Fas or by the release of perforin and granzymes. Both lytic pathways induce apoptosis yet it is not known whether identical or distinct apoptotic pathways are activated. The protooncogene bcl-2 is known to protect various hematopoietic cells from apoptosis induced by diverse agents. Here we show that overexpression of the Bcl-2 protein in the murine mastocytoma line P815 or in concanavalin A-activated splenocytes suppresses apoptotic cell death induced by allospecific primary cytotoxic T lymphocytes (CTL) in which only the Fas lytic pathway was functional. Bcl-2 also reduced target cell killing induced by CTL whose lytic activity was dependent on the perforin/granzyme pathway only. These data provide evidence that, in the target cells studied here, both perforin/granzyme and Fas apoptotic pathways are modulated by Bcl-2 and suggest that these two pathways converge at a step prior to Bcl-2 inhibition.

Mice homozygous for either the gld or lpr mutation develop autoimmune diseases and progressive lymphadenopathy. The lpr mutation is characterized by the absence of functional Fas, whereas gld mice exhibit an inactive FasL due to a point mutation proximal to the extracellular C-terminus. The structural repercussions of this amino acid substitution remain unknown. Here we report that FasL is expressed at similar levels on the surface of activated T lymphocytes from gld and wild-type mice. Using a polyclonal anti-FasL antibody, indistinguishable amounts of a 40 kDa protein are detected in both gld and wild-type splenocytes. The molecular model of FasL, based on the known structure of TNF-alpha, predicts that the Phe --> Leu gld mutation is located at the protomer interface which is close to the FasR interaction site. We conclude that the gld mutation allows normal FasL biosynthesis, surface expression and oligomerization, but induces structural alterations to the Fas binding region leading to the phenotypic changes observed.

With access to new sources the author reconstructs the conditions and circumstances leading in 1912 to what in the literature has come to be known as Freud's "secret committee". Schröter's sociological vantage enables him to pinpoint the mechanisms that made it possible for Freud to seek a resolution of the conflict smouldering between himself and Jung by staging a "palace revolution" which dethroned the institutionalized powerholder (Jung was president of the IPV and editor of the Jahrbuch für psychoanalytische und psychopathologische Forschungen) and established the Viennese group-centering around Freud and standing for his interest in the survival of his work-as an informal, secret body wielding power collectively and thus making it unnecessary for Freud himself to take over direct, personal "rule". At the same time, the author contends, the differences between Vienna and Zurich also need to be understood in terms of local and historical factors. Whereas Freud and Vienna represent a monarchic understanding of power in which power may be delegated but is never shared or relinquished, Jung and Zurich stand for democratic, liberal-bourgeois attitude towards power stemming from a long tradition of anti-monarchism in Switzerland.

It is commonly assumed that thiazide diuretics are ineffective in patients with advanced renal failure (GFR < 30 ml/min/1.73 m2). Thiazides act on the nephron segment distal to the ascending thick loop of Henle, that is, the site of action of loop diuretics. Blockade of sodium reabsorption in the thiazide-sensitive segment should therefore obliterate the compensatory increase in sodium reabsorption seen after administration of loop diuretics and thus potentiate the natriuretic efficacy of loop diuretics even in advanced renal failure. In a single-blind, randomized, placebo controlled crossover study we compared the natriuretic and chloruretic effect of the loop diuretic, torasemide, given alone or in combination with the thiazide diuretic, butizid, in 10 patients with advanced renal failure (mean CIn 13.1 +/- 5.9 ml/min/1.73 m2). For two weeks patients adhered to a diet containing a standardized amount of Na+ and K+. On the 6th and 13th study days, two sham infusions were given to patients in order to assess basal 24-hour urinary electrolyte excretion. On the 7th and 14th days they were randomly allocated to receive either 50 mg i.v. torasemide in combination with a sham infusion or torasemide in combination with 20 mg i.v. butizid. Administration of torasemide alone significantly (P < 0.01) increased mean cumulative 24-hour excretion of sodium (from 154 +/- 30 to 232 +/- 59 mmol/24 hr) and chloride (from 128 +/- 21 to 233 +/- 84 mmol/24 hr) as compared with baseline.(ABSTRACT TRUNCATED AT 250 WORDS)

Why did Freud, in his Leonardo study, speak of a vulture instead of a kite (in Italian nibbio)? It can be shown that contemporary dictionaries rendered nibbio by Hühnergeier; that this designation of the kite was unfamiliar in Standard German; but that it was automatically understood as referring to a sub-species of Geier (vulture). Freud had an anachronistic idea about non-marital sex relations in quattrocento Italy. Such liaisons were not uncommon between higher-ranking males and lower-ranking females, their offspring were assigned to the paternal family. Leonardo's mother probably served as wetnurse for her illegitimate son, returning him to his father and being married to a man of equal standing after the customary time of weaning (ca. 18 months). Concerning Leonardo's psycho-sexual development, the corrected data shift the emphasis from the oedipal level (Freud) to an earlier, ambivalent mother-child relation (Eissler). Both notes warn against some pitfalls of psychobiography.

The theory of civilization proposed by the eminent sociologist Norbert Elias, who died in 1990, has its roots in Freud's theory of culture. Elias always denied the importance of personal factors in connection with his work, but as has been demonstrated in connection with artists such as Leonardo da Vinci and indeed Freud himself, this does not make it any the less legitimate to go in search of those infantile/unconscious motifs and affective sources powering Elias' prodigious scholarly creativity and informing his work, in however sublimated a form. Proceeding from the observation that successful scholarly or artistic production represents a synthesis of the forces of id, ego and super-ego, i.e. of affect, knowledge and conscience, the author demonstrates that concerns of central moment to Elias' thinking--commitment and non-involvement, establishment and outsider, the restraining of aggression in the civilizational process, interdependence of individuals--are partly the product of early experiences which Elias was able to turn to fruitful account in his work.

Transition protein 1 (TNP1) is a highly basic nuclear protein of 54 amino acids that is found in haploid spermatogenic cells during the period of transition of histones to protamines. Using the cDNA clone for human TNP1, we have isolated the gene encoding human TNP1 from human genomic libraries. The gene contains an intron of 200 bp; 1104 bp of the 5'- and 276 bp of the 3'-noncoding region have been sequenced. Comparison with the rat TNP1 gene yielded a similarity of 77% over the region between the transcription start point and the polyadenylation signal. The gene contains typical CAAT and TATAA boxes at conventional distances from the transcriptional start site. Using a series of human-rodent somatic cell hybrids containing variant complements of human chromosomes, the TNP1 gene was found to cosegregate with human chromosome 2. By in situ hybridization, the gene was assigned to the q35 and q36 bands of the long arm of chromosome 2. This chromosomal region encodes several genes, including TNP1, that are located on murine chromosome 1.

The clinical picture of electrolyte-losing villous tumours localized in the rectum or colon is represented by own observations and the literature. Diagnostic errors arising from disturbances of water and electrolyte balance are emphasized. In order to find out the optimal therapy, classification of the tumour according to clinical criteria is recommended.