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Biomedical subjects

R Haas

Publications and source records attributed to R Haas.

At least 235 records · Page 13Linked to original sources
PubMed1994

Immunization of BALB/c mice against Helicobacter felis infection with Helicobacter pylori urease.

BACKGROUND/AIMS: Because Helicobacter pylori is a potentially dangerous human pathogen, the protective potential of oral immunization with H. pylori urease and its subunits was evaluated in an animal model. METHODS: Mice were orally immunized with H. pylori sonicate, urease, or recombinant enzymatically inactive urease subunits and then challenged with Helicobacter felis. Control mice were sham-immunized. RESULTS: H. felis colonization was present 5 days after challenge in 9 of 10 sham-immunized, 6 of 9 sonicate-immunized, and 3 of 10 urease-immunized animals (P = 0.031 vs. sham-immunized). Twelve days after challenge, urease B-immunized mice had a weaker colonization than sham-immunized controls, whereas urease A had no effect. After 70 days, most urease A- and urease B-immunized mice had cleared the colonization (10/17: P = 0.0019; 16/20: P = 0.00002 vs. sham-immunized). In urease B-immunized animals, protection was often associated with corpus gastritis. CONCLUSIONS: Oral immunization with H. pylori urease protects mice against H. felis infection. Enzymatically inactive urease A and B subunits contain protective epitopes. It is unclear whether protection depends on the development of a mononuclear inflammatory response in the gastric corpus. Our observations should encourage the development of a human vaccine.

Administration, Oral↗
PubMed1994

[The MRT of scaphoid pseudarthrosis with Gd-DTPA. Its staging and clinical correlation].

During a period of two years, 134 patients with pseudo-arthrosis of the scaphoid were examined by conventional radiography and by MRI in the course of a prospective study. The aim of the study was to define radiological staging using contrast enhanced MRI in order to improve the prognostic criteria. All MRI examinations were carried out with a 1.5 tesla scanner (SP63) using a surface coil and T1 weighted spin echo sequences in sagittal and frontal projection and frontal FLASH T2 sequences and axial spin echo T2 sequences. The T1 weighted SE sequences in frontal projection were carried out before and after iv contrast (0.1 mmol Gd-DTPA/kg KG). All sequences were compared with conventional radiographs and the operative findings. Eight patients in stage 0 showed high signal intensity of both fragments in T1 weighted SE sequences and at surgery there was good vascularisation. In 22 cases there was reduced signal intensity in at least one fragment (stage I). 45 patients with scaphoid pseudo-arthrosis showed complete signal loss but marked contrast uptake with still vital nuclei at surgery (stage II). In 22 patients, there was no increase in signal intensity after contrast and complete loss of vitality of the fragments at surgery. Staging was not possible in 37 patients because of previous operative intervention. The use of contrast enhanced MRI provides additional information compared with conventional radiography or plain MRI.

Adolescent↗
PubMed1994

Characterization of peripheral blood progenitor cells mobilized by cytotoxic chemotherapy and recombinant human granulocyte colony-stimulating factor.

The purpose of this study was to evaluate the antigenic profile of granulocyte colony-stimulating factor (G-CSF)-mobilized peripheral blood progenitor cells (PBPC) in patients with non-Hodgkin's lymphoma (NHL), Hodgkin's disease (HD), and multiple myeloma (MM). The mobilization regimens consisted of high-dose cytarabine/mitoxantrone for patients with NHL, DexaBEAM for patients with HD, and high-dose cyclophosphamide (4 or 7 g per m2) for patients with MM. Cytotoxic therapy was supported by recombinant human G-CSF (Filgrastim, 300 micrograms/day sc) to shorten the period of neutropenia and to increase the number of circulating hematopoietic progenitor cells. The mean numbers of circulating CD34+ cells/microliters during leukocyte recovery were different between patient groups, 80.5 +/- 9.8 (mean +/- SEM) in low-grade NHL and 51.2 +/- 9.7 in high-grade NHL compared with 31.3 +/- 6.9 in HD and 24.4 +/- 4.1 in patients with MM. As a result, the greatest numbers of CD34+ cells/kg collected per leukapheresis were observed in patients with NHL, whereas the collection efficiency was substantially lower in patients with HD or MM. Patients with MM had also the smallest proportion of CD34+ cells in the mononuclear cell fraction (mean 0.79 +/- 0.10% versus 2.15 +/- 0.19% in low-grade NHL) but the greatest proportion of early CD34+ HLA-DR- progenitor cells (mean 2.38 +/- 0.51 versus 0.84 +/- 14% in low-grade NHL). Patients with MM had a mean proportion of CD34/c-kit+ cells that was twofold greater than that observed in patients with high- or low-grade NHL.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1994

Genetic analysis of the Helicobacter pylori vacuolating cytotoxin: structural similarities with the IgA protease type of exported protein.

The human gastric bacterial pathogen Helicobacter pylori has been implicated in type B gastritis, peptic ulceration and gastric adenocarcinoma. Here we report on the cloning and genetic characterization of an H. pylori gene named vacA, which encodes the vacuolating cytotoxin VacA, a novel type of antigenic bacterial toxin that induces the formation of intracellular vacuoles in epithelial cells. The vacuolating cytotoxin activity is expressed by a subset of clinical isolates (Vac+), all of which produce the 87 kDa cytotoxin antigen, but strains which produce neither the activity nor the cytotoxin protein (Vac-) also carry the gene. Isogenic H. pylori mutants in vacA generated by transposon shuttle mutagenesis produce neither the VacA antigen nor a vacuolating activity in a cell culture model. The vacA gene itself encodes a precursor protein of 139.6 kDa consisting of a 33-amino acid signal sequence, the 87 kDa cytotoxin and a 50 kDa C-terminal domain with features typical of a bacterial outer membrane protein. The VacA precursor shows no significant primary sequence homology with any previously reported protein, but its structural organization closely resembles the IgA protease-type of exoprotein produced by pathogenic Neisseriae and Haemophilus species. Our current data support a model for secretion of the cytotoxin through the two bacterial membranes which involves the 50 kDa domain for outer membrane translocation with subsequent proteolytic cleavage and release of the mature 87 kDa cytotoxin into the extracellular environment.

Amino Acid Sequence↗
PubMed1994

Dexa-BEAM in patients with Hodgkin's disease refractory to multidrug chemotherapy regimens: a trial of the German Hodgkin's Disease Study Group.

PURPOSE: A prospective phase II study was conducted to evaluate the efficacy of dexamethasone, carmustine, etoposide, cytarabine, and melphalan (Dexa-BEAM) as salvage chemotherapy for patients with Hodgkin's disease. PATIENTS AND METHODS: Fifty-five patients progressing on or relapsing after eight- or 10-drug chemotherapy (cyclophosphamide, vincristine, procarbazine, and prednisone plus doxorubicin, bleomycin, vinblastine, and dacarbazine [COPP+ABVD] or COPP+ABV+ifosfamide, methotrexate, etoposide, and prednisone [IMEP]) were treated with Dexa-BEAM. Patients who responded after two cycles of Dexa-BEAM either continued treatment for another two to three cycles or received high-dose chemotherapy/autologous bone marrow transplantation (HDCT/ABMT) with cyclophosphamide, etoposide, and carmustine (BCNU) (CVB) as conditioning regimen. RESULTS: Seventeen patients (31%) achieved a complete remission and 16 (29%) a partial remission, resulting in a response rate of 60% (95% confidence interval, 46% to 73%). Progressive disease developed in 18 patients. Toxicity of Dexa-BEAM was acceptable with pronounced, but temporary World Health Organization (WHO) grade III/IV granulocytopenia and thrombocytopenia occurring in more than 90% of all courses. Two patients died of sepsis during granulocytopenia. Three prognostic subgroups could be distinguished: (1) patients progressing on initial chemotherapy, (2) patients relapsing within 12 months, and (3) patients with late relapses. The response rates for these groups were 52%, 60%, and 83%, and the median survival duration 12, 29, and 40+ months, respectively. In a nonrandomized comparison, the survival of patients who responded to two cycles of Dexa-BEAM and had additional cycles of Dexa-BEAM (n = 14) was not different from those responding patients who underwent HDCT/ABMT (n = 19). However, the power to detect a 20% survival difference was only 33% in this comparison. CONCLUSION: Dexa-BEAM is an effective salvage treatment for patients with Hodgkin's disease who fail to respond to multidrug chemotherapy. Efficacy and toxicity are comparable to HDCT/ABMT and underline the need for prospective randomized trials to define better the role of HDCT with and without ABMT in these patients.

Adult↗
PubMed1994

Sequential high-dose therapy with peripheral-blood progenitor-cell support in low-grade non-Hodgkin's lymphoma.

PURPOSE: To evaluate the feasibility of a sequential high-dose therapy with peripheral-blood progenitor-cell (PBPC) support in patients with follicular lymphoma. PATIENTS AND METHODS: Since July 1991, we have included 30 patients (17 men and 13 women) with a median age of 41 years (range, 26 to 55) in the study. At the time of study entry, 17 patients were in first and six in second or higher remission. Another six patients had relapse of disease and one had tumor progression. PBPC were collected during filgrastim-supported leukocyte recovery following high-dose cytarabine (ara-C)/mitoxantrone (HAM). RESULTS: A median of two leukaphereses (range, one to seven) resulted in a median of 5.7 x 10(6) CD34+ cells/kg (range, 2.9 to 23.7 x 10(6). A distinct population of B-lymphoid progenitors (CD34+/CD19+) was not detectable in the autografts, and the content of CD19+ B cells was remarkably low, comprising a median of 0.07% of the mononuclear cells. Using the polymerase chain reaction (PCR) assay for the major breakpoint regions (MBR) of the bcl-2/immunoglobulin H (IgH) translocation, 22 patients had autografts positive for the t(14;18) translocation, whereas seven patients had PCR-negative transplants. The autograft of one patient could not be assessed. Following myeloablative therapy, hematologic recovery was rapid without cytokine support. The median times to reach a platelet count > or = 20 x 10(9)/L and neutrophil count > or = 0.5 x 10(9)/L were 11 and 13 days, respectively. Nonhematologic toxicity was moderate. Twenty-nine patients were alive in remission after a median follow-up duration of 6 months (range, 1 to 18). Of 22 patients autografted with t(14;18)-positive harvests, 11 had PCR-detectable cells in bone marrow and/or peripheral blood as long as 16 months posttransplantation. In contrast, six patients became PCR-negative between 3 and 16 months after reinfusion. Follow-up examinations with PCR data for the remaining five patients are not yet available. CONCLUSION: Conversion to PCR negativity in patients autografted with PCR-positive harvests suggests that the myeloablative regimen is effective and that any reinfused t(14;18)-positive cells may not be sustained. Because conventional chemotherapy provides no cure, we believe that high-dose therapy including total-body irradiation (TBI) should be explored in these particularly radiosensitive lymphomas.

Adult↗
PubMed1994

Filgrastim post-chemotherapy mobilizes more CD34+ cells with a different antigenic profile compared with use during steady-state hematopoiesis.

For the mobilization of CD34+ peripheral blood progenitor cells (PBPC) filgrastim (R-metHuG-CSF) can be administered either during steady-state hematopoiesis or following cytotoxic chemotherapy. We compared both mobilization modalities intra-individually in seven patients with breast cancer. The number of circulating CD34+ cells was increased after filgrastim-supported chemotherapy compared with filgrastim administration during steady-state (on average, 129 vs. 19/microliters), resulting in a sevenfold higher yield of CD34+ cells per leukapheresis (5.73 vs. 0.79 x 10(6)/kg bodyweight). CD34+ PBPC harvested post-chemotherapy comprised a smaller proportion of early progenitor cells (CD34+/HLA-DR- or CD34+/CD38-) compared with filgrastim treatment alone. However, the absolute number of these early progenitor cells harvested was fivefold higher. The filgrastim-supported rebound after chemotherapy was characterized by a greater proportion of CD34+/CD33+ cells. Correspondingly, CD34+ PBPC mobilized post-chemotherapy contained a higher proportion of CFU-GM compared with filgrastim treatment during steady-state, while the cloning efficiency of CD34+ cells for BFU-E tended to be lower. Of note, the proportion of CD34+/CD19+ lymphoid progenitor and B cells was reduced after chemotherapy. In cancer patients, filgrastim mobilizes higher numbers of CD34+ cells when administered post-chemotherapy, compensating for the smaller proportion of early hematopoietic progenitors.

Adult↗
PubMed1994

Complete remission of metastatic breast cancer by non-specific immunostimulation with a Nocardia opaca cell-wall extract after failure of conventional therapy.

In a male patient with recurring lung metastases of infiltrating breast carcinoma (T2NOMO), primary treatment mastectomy, axillar dissection and local irradiation) the tumor metastases developed progressively despite of repeated surgical removement, polychemotherapy, and antiestrogen treatment. Active unspecific immunotherapy with a Nocardia opaca cell-wall extract, started in the final stage of the disease with deterioration of the performance status, resulted within several months in a complete remission without serious side effects. The reported result suggest that is is worth while to try such bacterial immunomodulators in therapy of advanced cancer in spite of many disappointing clinical trials.

Adjuvants, Immunologic↗
PubMed1993

TnMax--a versatile mini-transposon for the analysis of cloned genes and shuttle mutagenesis.

A series of Tn1721-based mini-transposons (TnMax) has been developed which are suitable for the targeting of cloned genes, shuttle mutagenesis, identification of protein export signals and the rescue of chromosomal markers. TnMax mini-Tn consist of two 38-bp inverted repeats (IR) flanking a resolution site (res), a suicide replication origin (orifd), and a gene conferring resistance to either chloramphenicol (TnMax1) or erythromycin (TnMax2). Other versions of TnMax (TnMax3 and TnMax4) carry a promoterless alkaline phosphatase-encoding gene (phoA') useful for the identification of protein export signals. The various mini-Tn cartridges are fused on the same plasmid with a common transposition control unit (TCU) comprising the tnpA (transposase) and tnpR (resolvase) genes under control of the inducible Ptrc promoter. This configuration causes a high frequency of transposition in Escherichia coli (approx. 10(-2) events/copy of target plasmid) and a minimum size of the mini-Tn. Like Tn1721, TnMax variants prefer random insertion into plasmids rather than into the E. coli chromosome, thus representing superb tools for the insertion mutagenesis of cloned genes. The TnMax-borne orifd simplifies the identification of targeted plasmids and facilitates shuttle mutagenesis, i.e., suicide delivery of a mutated gene with subsequent allelic replacement of a corresponding resident gene, in a variety of microorganisms. Rescue of such targeted chromosomal genes is easily accomplished by the excision of TnMax plus flanking segments using appropriate restriction endonucleases, ligation, and transformation of an E. coli host permissive for orifd-directed replication.

Base Sequence↗
PubMed1993

Development of an immunoassay to detect hemoglobin adducts formed by benzene exposure.

Polyclonal murine antibodies that recognize the adducts formed by benzene metabolites in hemoglobin (Hb) were prepared and used to develop immunoassays. In competitive inhibition assays, the concentration of competitor needed to reduce the signal by 50% (IC50) was less than 10 pmoles for hydroquinone-hemoglobin (HQHb) adducts and less than 1 pmole for 1,2,4 trihydroxybenzene-hemoglobin (TriOH Hb). Hemoglobin (Hb) incubated with either phenol or catechol (CAT) did not elicit antibodies suitable for quantitative immunoassays. The metabolite-directed immunoassays were tested using hemoglobin from mice previously administered [C14] benzene for two to four weeks. The most sensitive assay for hydroquinone measured 0.49 pmoles adduct/40 pmoles Hb (191 pmoles adduct/mg Hb) in mice treated with 200 mg/kg benzene (P < 0.05, Student's t test). TriOH Hb adducts were not detected.

Animals↗
PubMed1993

Reactivity of an active center analog of Cu2Zn2superoxide dismutase in murine model of acute and chronic inflammation.

The antiinflammatory efficacy of CuPu(Py)2 ([[N,N'-bis(2-pyridylmethylene)-1,4-butanediamine] (N,N',N'',N''')]-Cu2+), a serum stable active center analog of Cu2Zn2superoxide dismutase (SOD), was tested in vitro and in vivo in male Wistar rats suffering from potassium peroxochromate-induced inflammation. Parameters including 99mTc gamma-scintigraphic imaging, the arthritis score, the plasma superoxide dismutase activity, the inhibition of plasma sulfhydryl depletion as well as mitogenic and phagocytic responses were used to quantify the disease activity. All parameters improved impressively during the treatment with CuPu(Py)2 and resembled those of healthy animals after 21 days. The arthritis score was inhibited by 80% (P > 0.001) and the plasma SOD activity enhanced by 380% (P > 0.001). The depletion of plasma sulfhydryls and the leukocytic responses to concanavalin A, tetradecanoylphorbolacetate, and lipopolysaccharide were significantly reduced (P > 0.001) and correlated well with the arthritis score. The collapse of antioxidant defenses in human plasma as well as the depolymerization of hyaluronic acid was mimicked in vitro and successfully inhibited by CuPu(Py)2. Oxidant-induced injury of plasma components during the aqueous decay of potassium peroxochromate were demonstrated to activate the oxidative burst of phagocytes in human blood. The role of impaired pro- and antioxidant balances in the etiology of inflammatory and autoimmune rheumatic diseases is discussed.

Acute Disease↗
PubMed1993

Hematopoietic growth factors for the mobilization of peripheral blood stem cells.

Hematopoietic growth factors can be used for the mobilization of peripheral blood stem cells that have the proliferative capacity to restore long-term hematopoiesis after myeloablative therapy. An association between specific cytokines and the composition of the blood-derived progenitor cells has not yet emerged. It appears that yield and composition of PBSC are influenced far more by the individual than by the use of specific growth factors. Our future studies will focus on how many and what kind of stem cells are needed for high-dose regimens with different myelotoxicity. In parallel, autografts will be assessed for contaminating tumor cells, if disease- or clone-specific markers are available. This approach may then provide the rationale for the increasing use of PBSC for autografting.

Blood Cells↗
PubMed1993

Expression of adhesion molecules and c-kit on CD34+ hematopoietic progenitor cells: comparison of cytokine-mobilized blood stem cells with normal bone marrow and peripheral blood.

We assessed the expression of the adhesion molecules leukocyte function antigen-1 (LFA-1, CD11a), intercellular adhesion molecule-1 (ICAM-1, CD54), homing-associated cell adhesion molecule (H-CAM, CD44), and c-kit (stem cell factor receptor) on the CD34+ progenitor population from the leukapheresis products of 23 patients (LP CD34+). For blood stem cell collection granulocyte colony-stimulating factor (G-CSF) or interleukin-3/granulocyte-macrophage colony-stimulating factor (IL-3/GM-CSF) was administered after cytotoxic chemotherapy. Furthermore, bone marrow- and blood-derived CD34+ progenitor cells from 6 normal volunteers (BM and PB CD34+) were analyzed. LFA-1 expression was higher on PB CD34+ (88.2 +/- 2.5%, mean +/- SEM) than on BM CD34+ (75.3 +/- 4.3%). Following cytokine administration, LFA-1 was expressed on only 59.7 +/- 3.7% of LP CD34+ at a low fluorescence intensity, suggesting that down-regulation of LFA-1 may facilitate the egress of cells from the bone marrow and prolong their circulation. In contrast, ICAM-1 was weakly positive on CD34+ cells from all sources. CD44 was expressed on the vast majority of CD34+ cells (> 95%) in all samples studied. The highest proportion of CD34+ cells costaining for c-kit was found in normal bone marrow (32.2 +/- 3.3%). In normal peripheral blood and after cytokine mobilization, fewer of the CD34+ cells weakly expressed c-kit (< 15%). The low percentage and level of c-kit expression may indicate that the majority of cytokine-mobilized CD34+ cells are lineage-committed progenitor cells, as reflected by the coexpression pattern for CD38, HLA-DR, and CD33.

Antigens, CD↗
PubMed1993

Aflagellated mutants of Helicobacter pylori generated by genetic transformation of naturally competent strains using transposon shuttle mutagenesis.

Three out of 10 Helicobacter pylori clinical isolates were found to be naturally competent for genetic transformation to streptomycin resistance by chromosomal DNA extracted from a spontaneous streptomycin-resistant H. pylori mutant. The frequency of transformation varied between 5 x 10(-4) and 4 x 10(-6), depending on the H. pylori isolate used. Transposon shuttle mutagenesis based on this natural competence was established using the flagellin gene flaA as the target. The cloned flaA gene was interrupted by insertion of TnMax1, a mini-Tn1721 transposon carrying a modified chloramphenicol-acetyltransferase gene, the catGC cassette. Natural transformation of competent H. pylori strains with plasmid constructs harbouring a catGC-inactivated flaA gene resulted in chloramphenicol-resistant transformants at an average frequency of 4 x 10(-5). Southern hybridization experiments confirmed the replacement of the chromosomal H. pylori flaA gene by the cat-inactivated cloned gene copy via homologous recombination resulting in allelic exchange. Phenotypic characterization of the mutants demonstrated the absence of flagella under the electron microscope and the loss of bacterial motility. Immunoblots of cell lysates of the H. pylori mutants with an antiserum raised against the C-terminal portion of recombinant H. pylori major flagellin (FlaA) confirmed the absence of the 54 kDa FlaA protein. This efficient transposon shuttle mutagenesis procedure for H. pylori based on natural competence opens up new possibilities for the genetic assessment of putative H. pylori virulence determinants.

Bacterial Proteins↗
PubMed1993

Identification of Actinobacillus actinomycetemcomitans by leukotoxin gene-specific hybridization and polymerase chain reaction assays.

Eleven strains of Actinobacillus actinomycetemcomitans isolated from cases of systemic infections, local abscesses, and periodontitis were identified by genetic assays using the leukotoxin gene as the target. We have developed a polymerase chain reaction (PCR) assay, based on the leukotoxin structural gene of this pathogen, which clearly identified all tested strains of A. actinomycetemcomitans and separated them from the closely related Haemophilus aphrophilus as well as other bacterial species. Furthermore, DNA-DNA hybridization was performed with the cloned partial leukotoxin structural gene (lktA) as a probe, which again clearly distinguished A. actinomycetemcomitans from H. aphrophilus, parts of the normal oral flora, and species harboring RTX (repeats in toxin) family-related cytotoxins. The PCR fragment amplified from the leukotoxin structural gene gave results similar to those given by the cloned leukotoxin gene when used as a probe in hybridization experiments. The hybridization and PCR assays described here are fundamental improvements for the identification of A. actinomycetemcomitans.

Aggregatibacter actinomycetemcomitans↗