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Biomedical subjects

W Schraut

Publications and source records attributed to W Schraut.

At least 19 recordsLinked to original sources
PubMed2005

Minimal T-cell-stimulatory sequences and spectrum of HLA restriction of immunodominant CD4+ T-cell epitopes within hepatitis C virus NS3 and NS4 proteins.

The hepatitis C virus (HCV)-specific CD4+ T-cell response against nonstructural proteins is strongly associated with successful viral clearance during acute hepatitis C. To further develop these observations into peptide-based vaccines and clinical immunomonitoring tools like HLA class II tetramers, a detailed characterization of immunodominant CD4+ T-cell epitopes is required. We studied peripheral blood mononuclear cells from 20 patients with acute hepatitis C using 83 overlapping 20-mer peptides covering the NS3 helicase and NS4. Eight peptides were recognized by > or = 40% of patients, and specific CD4+ T-cell clones were obtained for seven of these and three additional, subdominant epitopes. Mapping of minimal stimulatory sequences defined epitopes of 8 to 13 amino acids in length, but optimal T-cell stimulation was observed with 10- to 15-mers. While some epitopes were presented by different HLA molecules, others were presented by only a single HLA class II molecule, which has implications for patient selection in clinical trials of peptide-based immunotherapies. In conclusion, using two different approaches we identified and characterized a set of CD4+ T-cell epitopes in the HCV NS3-NS4 region which are immunodominant in patients achieving transient or persistent viral control. This information allows the construction of a valuable panel of HCV-specific HLA class II tetramers for further study of CD4+ T-cell responses in chronic hepatitis C. The finding of immunodominant epitopes with very constrained HLA restriction has implications for patient selection in clinical trials of peptide-based immunotherapies.

Adolescent↗
PubMed1999

Virus-specific lymphokine production differs quantitatively but not qualitatively in acute and chronic hepatitis B infection.

Cytokines that are secreted as a response to viral antigen not only have direct antiviral properties but also crucially influence immune reactions determining the outcome of infection. As an advantageous alternative to the study of cytokines present in the supernatants of antigen-specific T cell clones and lines, we have used ELISPOT assays to determine the number of interferon-gamma (IFN-gamma)- and IL4-producing cells generated by peripheral blood mononuclear cells from patients with acute hepatitis B (AHB) and chronic hepatitis B (CHB) infection in response to HBcAg in a short-term culture (48 h). In response to HBcAg IFN-gamma was predominantly produced. In contrast to the results obtained in acute hepatitis B, the typical lymphokine pattern in CHB was characterized by a weak or absent antigen-specific IFN-gamma production. A predominance of IL-4-producing cells was not observed in either AHB or CHB. A significant number of IFN-gamma-producing cells was usually detectable during phases of viral elimination and the quality of the lymphokine response seemed to be epitope independent. Comparison of the results obtained in proliferation assays and ELISPOT assays clearly shows that lymphokine production upon stimulation with viral protein is totally independent of T cell proliferation and more sensitively reflects antiviral reactivity.

Acute Disease↗
PubMed1999

Outcome analysis of external coloanal anastomosis.

BACKGROUND: To evaluate the safety and efficacy of treating low-lying rectal lesions with resection and primary repair using a pull-through technique with rectal stump eversion and external coloanal anastomosis with immediate reintroduction into the pelvis. METHODS: All coloanal anastomoses with the above technique on the Gastrointestinal Surgery Service at the University of Pittsburgh from March 1990 to September 1995 were evaluated. RESULTS: Fifty-two patients underwent coloanal anastomoses with the above technique, and follow-up was available for 96% (50 of 52) of patients. Rectal lesions in the 50 patients included cancer (n = 34), rectal adenomas (n = 13), and other lesions (n = 3). Mean follow-up period was 29.6 +/- 21.8 months (28.5 months for patients with carcinoma). Fecal continence was normal or good in 88% (44 of 50) of patients. Moderate or complete incontinence was present in 12% (6 of 50) of patients. The local recurrence rate of rectal cancer was 0%. Morbidity occurred in 22% (11 of 50) of patients. Survival was 90% (45 of 50 patients). CONCLUSIONS: Coloanal anastomosis with this technique provides effective treatment for low-lying malignant or benign rectal lesions and has an acceptable complication rate.

Anal Canal↗
PubMed1998

A vigorous virus-specific CD4+ T cell response may contribute to the association of HLA-DR13 with viral clearance in hepatitis B.

A strong virus-specific CD4+ and CD8+ T lymphocyte response to hepatitis B virus (HBV) has been associated with viral clearance, but little is known about factors determining the individual's ability to mount such a T cell response. Recently a strong association between the HLA class II allele DR13 and a self-limited course of HBV infection has been described. In the present study of 33 patients with acute hepatitis B we show that individuals carrying HLA-DR13 mount a more vigorous CD4+ T cell response to HBV core (5706 ct/min (25th/75th percentile 3239 ct/min; 10,552 ct/min)) than patients without HLA-DR 13 (1365 ct/min (490 ct/min; 5334 ct/min); P = 0.006). However, peptide epitopes aa 50-69, aa 61-85, and aa 81-105 were recognized most frequently by both patient groups. Moreover, among 14 HBV core-specific CD4+ T cell clones from two patients with HLA-DR13, only one T cell clone was HLA-DR13-restricted. Our data suggest that the beneficial effect of the HLA-DR13 alleles on the outcome of HBV infection could be explained by a more vigorous HBV core-specific CD4+ T cell response, which may either be due to more proficient antigen presentation by the HLA-DR13 molecules themselves or a linked polymorphism in a neighbouring immunoregulatory gene.

Acute Disease↗
PubMed1998

Increased immune response elicited by DNA vaccination with a synthetic gp120 sequence with optimized codon usage.

DNA vaccination elicits humoral and cellular immune responses and has been shown to confer protection against several viral, bacterial, and parasitic pathogens. Here we report that optimized codon usage of an injected DNA sequence considerably increases both humoral and cellular immune responses. We recently generated a synthetic human immunodeficiency virus type 1 gp120 sequence in which most wild-type codons were replaced with codons from highly expressed human genes (syngp120). In vitro expression of syngp120 is considerably increased in comparison to that of the respective wild-type sequence. In BALB/c mice, DNA immunization with syngp120 resulted in significantly increased antibody titers and cytotoxic T-lymphocyte reactivity, suggesting a direct correlation between expression levels and the immune response. Moreover, syngp120 is characterized by rev-independent expression and a low risk of recombination with viral sequences. Thus, synthetic genes with optimized codon usage represent a novel strategy to increase the efficacy and safety of DNA vaccination.

AIDS Vaccines↗
PubMed1998

Meta-analyses of studies on bone marrow micrometastases: an independent prognostic impact remains to be substantiated.

PURPOSE: In 1997, the immunocytologic detection of isolated tumor cells in bone marrow, termed micrometastasis, will be optionally included in the tumor-node-metastasis (TNM) classification indicated M1(i). In the present meta-analyses, 20 studies, which included 2,494 patients, regarding the prognostic influence of a positive bone marrow micrometastases (BMM) status on relapse-free and/or overall survival were analyzed. MATERIALS AND METHODS: The literature search included the Medline and Current Contents bibliographic data bases from August 1980 to June 1997. The statistical evaluation considered the prognostic influence of the prevalence of micrometastatic cells in bone marrow on relapse-free and/or overall survival. The comparable effect estimate and its corresponding 95% confidence interval (CI) were calculated with the Mantel-Haenszel method using the originally published data of the retrieved studies. RESULTS: The presence of epithelial cells in bone marrow was detectable in all carcinoma types, with a median prevalence of approximately 35%. Fourteen of 20 studies found a positive correlation between positive BMM status and reduced relapse-free survival by univariate analysis, but only five of 11 studies confirmed positive BMM status as an independent predictor of short disease-free survival. Regarding overall survival, positive BMM status was identified univariately in five of 12 studies, but multivariately in only two studies, as an independent factor of poor survival. Despite the heterogeneity of the studies, calculation of the relative risk (RR) for reduced relapse-free survival was possible for breast cancer, which resulted in a Mantel-Haenszel RR (RR(MH)) of 1.34 (95% CI, 1.27 to 1.42). CONCLUSION: In conclusion, the results suggest that the prognostic impact of epithelial cells in bone marrow remains to be substantiated by further studies using standardized methodic protocols before its entrance in the TNM classification.

Bone Marrow Neoplasms↗
PubMed1998

[Expression of Lewis blood group antigens in stomach carcinoma induces metastatic potential].

In a series of gastric cancer patients twenty-eight percent of the primary tumors expressed high levels of the Lewis carbohydrate. This correlated significantly with clinicopathological parameters of advanced disease (tumor size of > 50 mm, M1 stage, UICC stage IIIB/IV). In multivariate analysis, high Lewis expression was found to be a new independent factor of poor prognosis. In addition, Lewis was expressed on solid metastases and micrometastatic cells originating from Lewis positive primary tumors. The data suggest that Lewis positive tumor cells may have an advantage in penetrating secondary organs because, like leukocytes, they can specifically adhere to activated vascular endothelia.

Animals↗
PubMed1997

TNF gene expression in monocytes of low and high responder individuals.

Individuals with a consistently lower immune response may be more susceptible to infection but less prone to autoimmune disease or severe sepsis. The molecular mechanisms determining the low responder status are, however, unclear. We have screened 60 male donors for tumour necrosis factor (TNF) protein levels after stimulation of monocytes with lipopolysaccharide (LPS). Among these we identified three donors each that consistently had a level of less than 20% (low responders; LR) or of more than 80% (high responders; HR) of the maximum response seen in this population. Northern blot analysis of TNF mRNA after LPS stimulation revealed lower transcript levels in LR. Half life determination after actinomycin D blockade showed a similar decay rate for LR and HR and after blockade of degradation by cycloheximide treatment mRNA levels increased but LR remained lower compared to HR. These data indicate that the lower TNF mRNA levels in LR are not due to a more rapid mRNA degradation but rather a result of lower transcription. Transcripts for interleukin 6 (IL-6) were also low in LPS-stimulated monocytes of LR. Because expression of the LPS receptor CD14 was similar in LR and HR monocytes, our data suggest that differences in signal transduction account for the LR and HR phenotype.

Blotting, Northern↗
PubMed1997

The Rhesus D-negative phenotype is an independent predictor of poor prognosis in curatively (RO) resected gastric cancer patients.

Among gastric cancer patients, the Rhesus D-negative phenotype correlated with increased tumour recurrence [all patients, n = 83, P = 0.026; curatively (R0) resected patients, n = 51, P = 0.093] and reduced overall survival time (all patients, log-rank P = 0.0028; R0 patients, log-rank P = 0.0003) and was identified in multivariate analysis as the most important independent prognostic marker in the R0 patient group (relative risk 9.1, P = 0.0013).

Aged↗
PubMed1996

Pyruvate prevents ischemia-reperfusion mucosal injury of rat small intestine.

BACKGROUND: Since reactive oxygen intermediates (ROI, or free radicals) have been implicated in the pathogenesis of ischemia-reperfusion injury of the small bowel, we evaluated the pretreatment effect of pyruvate, a 3-carbon compound recently shown to inhibit superoxide production, on reperfusion mucosal injury in the rat. METHODS: The small bowel of the ACI rat (n = 6) was divided into 2 5-cm segments, and 10 mL of a liquid diet containing pyruvate (0.32 g) or placebo (0.26 g) was instilled into the lumen of one of the segments for 10 minutes. The bowel was then made completely ischemic for 45 minutes by clamping the superior mesenteric artery, which was followed by 60 minutes of reperfusion. RESULTS: The production of ROI in bowel biopsy samples, estimated by luminol-enhanced chemiluminescence, was at least 80% decreased in the segment containing pyruvate compared with placebo immediately after ischemia (time 0), and compared with 30 and 60 minutes of reperfusion (P < 0.05 for each time point). After 60 minutes of reperfusion, the bowel segment containing the placebo diet showed villus sloughing with destruction of lamina propria and crypts, and mucosal neutrophil infiltration had increased by 80%. Electron microscope evaluation revealed a reduction in number and size of microvilli, dilatation of intercellular spaces, and intracellular vacuoles. The bowel segment containing pyruvate showed the villi and crypts to be intact, without enhanced neutrophil infiltration. CONCLUSION: Pyruvate pretreatment of the rat small bowel inhibits postischemic reperfusion mucosal histologic injury, neutrophil infiltration, and ROI production.

Animals↗
PubMed1994

Tolerance to lipopolysaccharide involves mobilization of nuclear factor kappa B with predominance of p50 homodimers.

Stimulation of the human monocytic cell line Mono Mac 6 with lipopolysaccharide (LPS) leads to rapid and transient expression of cytokines like tumor necrosis factor (TNF). When such cells are precultured for 2 days with a low dose of LPS (20 ng/ml) followed by stimulation with a high dose of LPS (1 microgram/ml), expression of the TNF gene is minimal, i.e. the cells are tolerant. In nuclear run-on analysis, such tolerant cells show only a low degree of transcription, indicating that tolerance operates at or upstream of the transcription level. The CD14 LPS receptor is, however, up-regulated (not down-regulated) in tolerant cells, and LPS can, in fact, still lead to activation of tolerant cells as evidenced by mobilization of the transcription factor nuclear factor kappa B (NF-kappa B). Resolution of the NF-kappa B complex in gel shift analysis shows that the binding protein, mobilized in naive Mono Mac 6 cells, consists mainly of p50-p65 heterodimers, while in tolerant cells, the p50 homodimer is predominant. This increase in p50 homodimers coincides with an increase in p105 mRNA, suggestive of a transcriptional up-regulation of p50. Reporter gene analysis reveals that the NF-kappa B complex mobilized in tolerant cells is functionally inactive in that NF-kappa B-dependent luciferase constructs containing the human immunodeficiency virus long terminal repeat or the TNF 5'-region show only minimal transactivation after LPS stimulation. Similar to Mono Mac 6 cells, primary blood monocytes, when precultured with a low dose of LPS, also become tolerant and produce little TNF after LPS stimulation. The tolerant blood monocytes also up-regulate CD14, and they mobilize NF-kappa B with a predominance of p50 homodimers. Taken together, these results demonstrate that tolerance to LPS is determined by post-receptor mechanisms that involve an altered composition of the NF-kappa B complex.

Antigens, CD↗
PubMed1994

Distinct patterns of differentiation induced in the monocytic cell line Mono Mac 6.

The human Mono Mac 6 cell line exhibits many characteristics of mature blood monocytes including expression of the CD14 molecule and production of cytokines, such as interleukin-1 (IL-1), IL-6, and tumor necrosis factor. To determine whether these cells can be further differentiated, we treated the cells for up to 3 days with either prostaglandin E2 (PGE2; 10(-5) or 10(-6) M), lipopolysaccharide (LPS; 10-20 ng/ml), or tetradecanoylphorbol-13-acetate (TPA; 10-50 ng/ml). All three reagents reduced proliferation and expression of the early myelomonocytic antigen CD33, and all increased phagocytosis of staphylococci and constitutive expression of mRNA for the macrophage colony-stimulating factor (M-CSF) receptor. By contrast, with respect to CD23 (Fc epsilon RII) expression, CD14 expression, and production of O2-, the three reagents induced distinct responses. Expression of CD23 (Fc epsilon RII) on Mono Mac 6 cells (36%) was not increased by LPS and TPA but was increased by PGE2 treatment to 48%, with a 50% increase of fluorescence intensity. The CD14 antibody My4 stained more than 75% of untreated Mono Mac 6 cells with a specific mean fluorescence intensity of 87.5 channels. This staining was increased more than twofold by both PGE2 and LPS. Staining with the CD14 antibody UCHM1 (6%) was increased to 43% by PGE2 and to 43% by LPS. This increase in CD14 cell surface expression was accompanied by a rise in soluble CD14 and enhancement of CD14 mRNA. By contrast, TPA treatment resulted in a twofold decrease of CD14 cell surface staining with no significant change in sCD14, while CD14 mRNA was transiently down-regulated. Secretion of O2- (stimulated by TPA) was already detectable in untreated Mono Mac 6 cells (6.1 mmol/10(6) cells/30 min), and this response was enhanced 10-fold by pretreatment with LPS but not with PGE2 or TPA. The kinetics of M-CSF receptor mRNA, CD14 expression, and O2- production revealed that these monocytic features started to increase at 6-24 h and were maximal at 2 days. These data suggest that the three reagents induce maturation of the Mono Mac 6 cells to different levels or into different branches of the monocyte system with the notable differences that PGE2 enhances CD23 expression, LPS enhances O2- secretion, and TPA down-regulates CD14.

Antigens, CD↗
PubMed1993

Erythema induratum of Bazin. Evidence of T-lymphocyte hyperresponsiveness to purified protein derivative of tuberculin: report of two cases and treatment.

BACKGROUND AND DESIGN: Erythema induratum of Bazin, a chronic form of nodular vasculitis, may be associated with chronic infections by Mycobacterium tuberculosis. However, the true origin of the disease is a subject of speculation and remains elusive. Two female patients (58 years old and 33 years old) with a minimum 10-year history of chronic tender ulcerating nodules on the lower aspects of the legs were studied both clinically and in the response of their peripheral T cells to purified protein derivative of tuberculin. RESULTS: Both patients with no previous history of tuberculosis had strongly positive skin test results at a Mantoux 10(-4) dilution (1 unit of purified protein derivative). In response to full-course triple-agent (isoniazid, rifampicin, and ethambutol) chemotherapy, a complete remission of clinical symptoms was seen in both cases and no relapse occurred after discontinuation of therapy. A marked increase in peripheral T-lymphocyte response to purified protein derivative was found before onset of and during successful therapy. CONCLUSIONS: The present clinical observations together with the pronounced cellular response to purified protein derivative suggest a tuberculous origin of erythema induratum of Bazin.

Adult↗
PubMed1993

The novel subset of CD14+/CD16+ blood monocytes exhibits features of tissue macrophages.

The CD14+/CD16+ cells account for about 10% of all blood monocytes. They are characterized by a low level expression of the CD14 molecule and a high level expression of the CD16 (Fc gamma R III) molecule. Polymerase chain reaction analysis of mRNA prevalence in CD14+/CD16+ cells (compared to the regular CD14++ blood monocytes) demonstrates low levels of CD14 transcripts and high levels of CD16 transcripts, suggestive of a transcriptional control for both of these proteins. Analysis of additional cell surface molecules in three-color immunofluorescence reveals that CD14+/CD16+ cells express the Fc gamma R II in all, and Fc gamma R I and ICAM-1 in some donors. Furthermore, class II antigens are expressed at fourfold higher levels, while both, CD11b and CD33 cell surface proteins, are decreased by a factor of two. Transcript levels were reduced in CD14+/CD16+ cells for all three cell surface molecules. Since these phenotypic markers of the CD14+/CD16+ blood monocytes are reminiscent of tissue macrophages, we performed a comparative analysis with alveolar macrophages (AM). These cells are similar to the CD14+/CD16+ monocytes in that they show low levels of CD14 and strong expression of CD16. Furthermore, similar to the CD14+/CD16+ cells, the AM also exhibit higher levels of class II and lower levels of CD11b and CD33 when compared to the regular CD14++ blood monocytes. In vitro induction of maturation of blood monocytes by 5 day culture of peripheral blood mononuclear cells in 10% human serum will result in decreased CD14 and increased CD16 cell surface expression on the monocyte derived macrophages. At the same time, these cells acquire increased levels of class II and decreased levels of CD11b and CD33. Taken together, these data show that CD14+/CD16+ monocytes, while still in circulation, have acquired features in common with mature tissue macrophages.

Antigens, CD↗
PubMed1993

Increased frequency of CD8+ CD45R0+ memory T lymphocytes in acute hepatitis B virus infection.

CD8+ lymphocytes of the memory subset (= CD45R0+) encompass antigen-specific effector cells, which are believed to be decisive for virus elimination in several viral infections. To determine whether this can be extended to HBV infection naive and memory T cells were studied among CD(4+)- and CD(8+)-lymphocytes and used monoclonal antibodies in two-color flow cytometric analysis to quantitate functional T cell subsets in peripheral blood of patients with acute hepatitis B (n = 11), chronic hepatitis B (n = 24) and healthy individuals (n = 26). Compared to CD4+ populations of healthy individuals the number of total CD4+ lymphocytes in patients with both acute or chronic hepatitis was significantly reduced. In contrast CD8+ cells did not significantly change in either acute and chronic hepatitis. Analysis of naive and memory subsets demonstrated, however, a significant rise in CD45R0+ memory cells from 5 to 15% (70% of all CD8+ cells) in acute hepatitis. These changes within the CD8+ population were, however, restricted to the acute phase of hepatitis in that the frequency of CD8+CD45R0+ decreased within weeks post infection. Furthermore, patients with chronic hepatitis did exhibit normal values of CD8+ memory cells (30% of all CD8+ cells). These findings suggest that enrichment of CD8+CD45R0+ memory cells reflects an accumulation of functional effector cells, which may be specifically activated by viral antigens and determine the outcome of infection.

Acute Disease↗
PubMed1992

Nitric oxide production in host-versus-graft and graft-versus-host reactions in the rat.

The present study was designed to determine whether .N = O produced in vivo during the rejection of histoincompatible tissues might permit serum NO2-/NO3- levels to serve as markers of a rejection reaction. Rat syngeneic and allogeneic liver, heart, bone marrow/spleen cell, small bowel, skin, and sponge matrix grafts were performed and the stable end-products of .N = O, NO2-/NO3-, were serially assayed in the serum of the grafted animals. A significant rise of serum NO2-/NO3- levels in the allografted animals preceded the onset of clinical signs of rejection or graft-versus-host disease, with the exception of the skin and sponge matrix graft models, where elevated serum NO2-/NO3- levels were never observed. In all transplant models, normal serum NO2-/NO3- levels were observed at all times in animals that received syngeneic grafts. Furthermore, treatment of allograft recipients with the immunosuppressive agents FK 506 or cyclosporine A inhibited .N = O production. Determination of serum creatinine levels demonstrated that the elevated serum NO2-/NO3- levels were not caused by kidney dysfunction. Serum NO2-/NO3- levels might be useful early serum markers of the initiation of a rejection reaction or graft-versus-host disease when functional markers of graft dysfunction are not apparent.

Animals↗
PubMed1991

Hepatitis B virus antigen-specific T-cell activation in patients with acute and chronic hepatitis B.

Since the hepatitis B virus is noncytopathic, it is generally believed that the individual specific immune response determines the course of infection. The lack of data about hepatitis B virus-specific T-cell reactions in acute infection led us to investigate the specific cellular immune response of infected individuals in terms of proliferation, and gamma-interferon and lymphotoxin production. Our results demonstrate that peripheral blood mononuclear cells (PBMNC) from patients with acute and chronic hepatitis B respond weakly to HBsAg. In contrast, patients with acute hepatitis show a vigorous response to the nucleocapsid antigen (HBcAg) in terms of proliferation and lymphokine production, while only few chronic virus carriers gave a proliferative response. Either of the antigens could activate lymphocytes to produce gamma-interferon and lymphotoxin, cytokines which may modulate antiviral immune response.

Acute Disease↗