Search PubMed⌕ Search

Biomedical subjects

R Haas

Publications and source records attributed to R Haas.

At least 163 records · Page 9Linked to original sources
PubMed1997

Expression of the human immunodeficiency virus type-1 coreceptors CXCR-4 (fusin, LESTR) and CKR-5 in CD34+ hematopoietic progenitor cells.

CD34+ hematopoietic progenitor cells were assessed for mRNA expression of the human immunodeficiency virus type-1 (HIV-1) coreceptors CXCR-4, also termed fusin or LESTR, and CKR-5, also called CC-CKR-5 or CCR-5. The CD34+ cells were obtained from leukapheresis products of 17 patients after granulocyte colony-stimulating factor-supported cytotoxic chemotherapy. Using a two-step enrichment procedure including immunomagnetic bead separation and fluorescence-activated cell sorting, the CD34+ cells had a median purity of 99.8%. Assessing 9 CD34+ cell samples by polymerase chain reaction after reverse transcription (RT-PCR), CXCR-4 mRNA was found in all samples, whereas CKR-5 mRNA was only present in 3 samples, even though a nested PCR was used. Eight additional CD34+ cell samples were sorted according to CD4 expression. Based on a three-color immunofluorescence analysis, the mean relative fluorescence intensity of HLA-DR was smaller on CD34+/CD4+ cells in comparison with CD34+/ CD4- cells. CXCR-4 mRNA was found in 5 of 8 CD34+/CD4+ samples and in 7 of 8 CD34+/CD4- samples, whereas CKR-5 mRNA was detected in 2 CD34+/CD4+ samples and in 1 CD34+/CD4- cell sample. Looking at the total number of CD34+ cell samples examined, the proportion of specimens containing CXCR-4 mRNA was 84% in comparison with 24% of specimens positive for CKR-5 mRNA. These data suggest that CD34+/CD4+ hematopoietic progenitor cells, including true stem cell candidates, could be susceptible to HIV-1 infection. Considering the relatively low incidence of CD34+ cell samples containing CKR-5 mRNA, CD34+/CD4+ cells appear to be particularly prone for HIV-1 infection via the CXCR-4 coreceptor. Because this chemokine receptor allows T-cell-tropic HIV-1 strains to infect cells, CD34+ cells expressing CD4 and CXCR-4 might be infected by HIV-1 during later stages of the disease, following a viral phenotype switch from macrophage- to T-cell-tropic HIV-1 strains.

Antigens, CD34↗
PubMed1997

Lack of t(14;18) polymerase chain reaction-positive cells in highly purified CD34+ cells and their CD19 subsets in patients with follicular lymphoma.

Follicular lymphoma (FL) is characterized in a significant proportion of cases by the t(14;18) chromosomal translocation, which results in the juxtaposition of the oncogene bcl-2 to the joining region of the immunoglobulin heavy chain (IgH) gene. Molecular sequence analysis indicates that the t(14;18) rearrangement occurs in a B-lymphoid progenitor cell at the time of IgH rearrangement. We were interested whether hematopoietic stem and progenitor cells as characterized by CD34 expression bear the translocation. Bone marrow (BM)-CD34+ cells were enriched from 14 patients with FL whose BM was known to be positive for bcl-2/IgH (major breakpoint region [MBR]). Six patients were in complete remission (CR), two patients were in partial remission (PR), and six patients had active disease. Six patients had histological BM involvement when the samples were obtained. Using an immunomagnetic selection device (MINI-MACS), a mean purity of 88.7% +/- 4% CD34+ cells was achieved. The CD34+ cells were further enriched by fluorescence activated cell sorting (FACS) using CD34 fluorescein isothiocyanate (FITC)- and CD19 phycoerythrin (PE)-conjugated antibodies. The IgH gene was rearranged in the CD34+/CD19+ cell subset of all patients assessed by polymerase chain reaction (PCR). This population is thought to represent the progenitor stage at which the bcl-2/IgH translocation occurs. The unseparated BM mononuclear cell fraction from all 14 patients was positive for bcl-2/IgH using a nested PCR, but the BM-CD34+ cell fraction and the respective CD34+/CD19+ subset were negative in 13 of these 14 patients. The one patient with a positive PCR signal in the CD34+ cell subset had a relapse with BM involvement. We conclude that CD34+ progenitor cells including CD34+/CD19+ B-cell progenitors are not involved in the malignant cell clone. These data are in agreement with a transgenic mouse model, which indicates that the malignant phenotype in FL is sustained by mature B cells.

Antigens, CD19↗
PubMed1997

[Ambulatory therapy of multiple myeloma with vincristine, adriamycin and dexamethasone].

BACKGROUND AND OBJECTIVE: If multiple myeloma (MM) progresses in patients after chemotherapy with alkylating agents, the combination of vincristine, adriamycin and dexamethasone (VAD) can achieve a response in 40-70% of cases. Because of its low toxicity for haematopoetic stem-cells this form of chemotherapy is often undertaken before high-dose blood stem-cell transplantation. It was the objective of this study to examine effectiveness and complications of ambulant VAD treatment. PATIENTS AND METHODS: Within four years VAD chemotherapy was given to 62 ambulant MM patients, administered by microprocessor-regulated pumps via intravenously polyurethane catheters with a safety valve. Response to treatment, treatment-associated complications and infections were documented prospectively and analysed. RESULTS: VAD treatment achieved tumour reduction of more than 25% in 50 of 62 patients. This treatment had to be discontinued in two of 192 pump-infusions because of irreversible catheter occlusion. Eight patients were hospitalised because of infections and two for noninfectious complications. Severe infectious complications (> or = WHO grade III) occurred in 4% of treatment cycles. CONCLUSION: VAD chemotherapy can be performed with a low rate of infection in ambulant patients despite the need for prolonged intravenously infusion of the drugs. But to avoid complications by intravenously catheters, random prospective tests should first be done with oral alkylating agents.

Adult↗
PubMed1997

Endocrine function and bone metabolism 5 years after autologous bone marrow/blood-derived progenitor cell transplantation.

BACKGROUND: High dose chemotherapy with or without total body irradiation supported by autologous transplantation of hematopoietic progenitor cells is increasingly being used for hematologic and solid tumors. However, there is only limited information available on late toxicity. METHODS: The authors investigated endocrine function and bone metabolism in 29 patients with a median interval of 5 years after autografting. RESULTS: In accordance with an earlier report on this patient cohort, ovarian failure was observed to be unchanged, except for one woman with recovered ovarian function who gave birth to two healthy children. In two-thirds of the men, follicle-stimulating hormone levels were elevated, suggesting germinal aplasia. Determination of bone mineral density did not reveal osteopenia, despite several risk factors: prolonged immobilization, high dose corticosteroid treatment, and, in women, transient estrogen insufficiency. Frequent impairment of thyroid function has been reported in patients receiving single dose total body irradiation. Overt or subclinical hypothyroidism was not detected, most likely because the total body irradiation was hyperfractionated. CONCLUSIONS: With the exception of gonadal failure, no significant late effects on endocrine function or bone metabolism were observed in this patient cohort.

Adrenocorticotropic Hormone↗
PubMed1997

Particulate ANP-sensitive guanylyl cyclase in blood and bone marrow cells of patients with acute leukemia.

Increases in plasma cyclic GMP levels have been shown to correlate with increased plasma levels of atrial natriuretic peptide (ANP) in patients with fluid overload due to increased secretion of ANP. There is evidence that plasma cyclic GMP levels are also elevated in some patients with acute leukemia, but increased ANP secretion has not been demonstrated. To elucidate the possibility that a newly expressed guanylyl cyclase may be responsible for the increase of plasma cyclic GMP levels patients with acute and chronic leukemia as well as patients with lymphoma and healthy volunteers were studied. Plasma levels of cyclic GMP were measured and isolated peripheral blood mononuclear or bone marrow cells were incubated with increasing concentrations of ANP. The stimulation of cells was measured as cGMP accumulation in the supernatant. Furthermore guanylyl cyclase activity was measured in membrane preparations of peripheral blood mononuclear cells. While leukocytes of healthy subjects were devoid of detectable ANP-stimulated particulate guanylyl cyclase activity, ANP-sensitivity was observed in seven patients with acute lymphoblastic and in three patients with acute myelogenous leukemia. Cyclic GMP in the supernatant of cells was elevated between 2- and 132-fold of basal when cells were incubated with 1 microM ANP for 60 minutes. Like in healthy volunteers, no effect of ANP on freshly isolated mononuclear cells was observed in cases with chronic leukemia or in patients with lymphoma. Expression of ANP-sensitive particulate gunaylyl cyclase may be connected with malignant transformation of lymphocytes in patients with acute leukemia and might be useful for their diagnosis and classification.

Acute Disease↗
PubMed1997

Expression and purification of recombinant human apolipoprotein A-I in Chinese hamster ovary cells.

The expression of apolipoprotein A-I (apoA-I) has been shown to be very difficult due to its amphiphilic character, autoaggregation, and degradation. We have expressed apoA-I using CHO cells, Baculovirus, and Escherichia coli [Schmidt et al., J. Biol. Chem. (1995) 270, 469-475]. Here we report about optimized conditions for the expression of proapoA-I in CHO cells, testing various serum-free media. We were able to yield apoA-I expression up to 80 micrograms/ml, by far the highest ever reported. However, immunoblot analysis revealed degraded apoA-I. The best apoA-I expression testing various conditions was about 20-30 micrograms/ml without any evidence of degradation. Interestingly, the apoA-I expression resulted in reproducible apoA-I fragments of 26 and 14 kDa. These fragments are consistent with already reported in vivo findings, in which carboxy-terminal proteolysis was suggested. The use of the protease inhibitors pepstatin and chymostatin, both carboxy-peptidase inhibitors, did result in contrast to other studied protease inhibitors in increased apoA-I yield. Therefore, limited carboxy-terminal proteolysis contributes to the degradation of CHO cell-secreted apoA-I. In addition, we evaluated various purification methods for the preparative isolation of recombinant apoA-I. In our hands we obtained the best recovery and no degradation with reversed-phase chromatography using a FPLC system.

Animals↗
PubMed1997

[Malignant lymphoma. Pathology, diagnosis, therapy].

Malignant lymphomas can be subdivided into Hodgkin's disease and low- or high-grade non-Hodgkin's lymphoma (NHL). The principal therapeutic options are polychemotherapy and radiotherapy. Besides the histological classification, staging of the disease with particular regard to risk factors is an essential prerequisite for the therapeutic decision. Diagnostic imaging modalities such as computer tomography, magnetic resonance imaging, and ultrasonography have improved the accuracy of clinical staging such that invasive pathological staging is only necessary in exceptional cases. A novel therapeutic approach is high-dose chemotherapy with autologous haematopoietic stem-cell support. This treatment improves the survival of patients with relapsed high-grade NHL. The place of high-dose therapy as the primary therapeutic option in malignant lymphoma is now being assessed in prospective studies following encouraging results from single-centre studies, including those involving the treatment of low-grade lymphoma. The effects of antibodies directed against lymphatic cells are currently being examined in experimental treatments. An assessment of the viability and rate of proliferation of lymphoma tissue on completion of therapy using sensitive radiological and nuclear medical methods is an important aim for the future.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1997

[Ultrasound criteria for staging and follow-up of malignant lymphoma].

Ultrasonography is an integral part of the radiological imaging of malignant lymphomas for staging, therapy planning, monitoring, follow-up, post-therapeutic care, and detection of recurrences, as well as the diagnostic differentiation of benign lesions and metastasis of solid tumors. None of the imaging methods, including sonography, however, can safely determine the stage (UICC certainty level C4) in agreement with the pathohistological examination. Also considering the clinical appearance the sonographic criteria in the synoptic evaluation will nevertheless provide important indications for diagnosis and therapy planning. Not only changes in size and shape, but also the echogenicity (gray-scale histogram) and vascularization (color Doppler) of nodal and extranodal processes are decisive criteria for estimating the therapeutic response rates and detecting relapses.

Follow-Up Studies↗
PubMed1997

Cloning and functional characterization of the genes encoding 3-dehydroquinate synthase (aroB) and tRNA-guanine transglycosylase (tgt) from Helicobacter pylori.

The aroB gene from Helicobacter pylori strain P1 was cloned and further characterized by sequence analysis and by functional complementation of the aroB mutation in Escherichia coli. The aroB gene encodes the enzyme 3-dehydroquinate synthase which catalyzes one of the early steps in the shikimate pathway. This pathway, which creates aromatic molecules from sugar precursors, is present in prokaryotes, fungi and plants but is absent from mammalian cells. The predicted amino acid sequence of the H. pylori aroB gene product showed significant homology (30-40% identity and 50-60% similarity) to 3-dehydroquinate synthases from various other prokaryotes and eukaryotes. The single gene on a plasmid was biologically active in E. coli. It suppressed the specific phenotype of aroB mutants by restoring the shikimate pathway-dependent synthesis of aromatic amino acids and the production of the siderophore enterobactin. Two other reading frames were found adjacent to the aroB gene. The first, designated as orf1, had no significant homology to proteins and genes present in databases, whereas the second was found to share a significant degree of homology with the tgt gene encoding tRNA-guanine transglycosylase from a variety of other bacteria (40-50% identity and 60-70% similarity). The function of the tgt gene was confirmed by heterologous complementation. The gene on a plasmid was shown to complement the queuosine biosynthesis defect in a genetically defined tgt- strain of E. coli. The presence of the aroB gene and the putative tgt homologue in unrelated H. pylori strains was confirmed by Southern blot hybridization and by polymerase chain reaction with specific primers.

Amino Acid Sequence↗
PubMed1997

MALT-type lymphoma of the stomach is associated with Helicobacter pylori strains expressing the CagA protein.

BACKGROUND & AIMS: Helicobacter pylori is considered to be involved in the pathogenesis of gastric lymphoma of mucosa-associated lymphoid tissue (MALT) type. Strains expressing the CagA protein (CagA+ strains) have been strongly associated with severe gastritis, duodenal ulceration, and gastric adenocarcinoma. The aim of this study was to determine the presence of H. pylori as well as incidence of CagA+ strains in gastric MALT-type lymphoma. METHODS: Sera of 68 patients with gastric MALT-type lymphoma (22 with low grade, 36 with high grade, and 10 with secondary high grade) were obtained, and the serological response to CagA was studied by immunoblotting using a purified recombinant CagA protein, a CagA+ strain, and the corresponding isogenic CagA- mutant. RESULTS: Of the patients with MALT-type lymphoma, 98.5% (67 of 68 patients) were H. pylori seropositive. In the only seronegative patient, the bacterium was detected histologically by Warthin-Starry staining. Of the seropositive patients, 95.5% had serum immunoglobulin G antibodies to CagA compared with 67% of an H. pylori-positive control group (33 of 49 patients; P = 0.000037) with chronic active gastritis. CONCLUSIONS: These results indicate infection of almost all patients with MALT-type lymphoma by CagA+ H. pylori strains. Strains expressing the CagA protein seem to play a crucial role in the pathogenesis of gastric MALT-type lymphoma.

Adult↗
PubMed1997

Ultra-sensitive FISH is a useful tool for studying chronic HIV-1 infection.

Using an ultra-sensitive fluorescence in situ hybridization (FISH) protocol, integration sites of the human immunodeficiency virus-1 (HIV-1) were mapped in two model cell lines for chronic HIV infection. In the promonocytic cell line U1/HIV-1, the two integrated copies of proviral HIV-DNA were located on the short arm of one copy of chromosome X (band Xp21) and on a rearranged chromosome 6. In the T-cell line 8E5/LAV, the single copy of proviral HIV-DNA was integrated into chromosomal bands 13q14-q21, which are known to contain common fragile sites. In addition, ultra-sensitive FISH was used for the detection of HIV provirus sequences in interphase cells, achieving a high hybridization efficiency. This application allows the analysis of the HIV status on a single cell level and thus may become a useful tool for the study of HIV infection.

Cell Line↗
PubMed1997

Disinfection of cell-associated and extracellular HIV-1 by PUVA treatment.

To inactivate cell-associated and extracellular HIV-1 while preserving cellular surface antigens, a procedure was used based on PUVA treatment, i.e. addition of psoralen to cell suspensions followed by irradiation with UVA light. T-lymphoid MT-4 cells were infected with HIV-1 strain NL4-3, 4'-aminomethyl-4,5',8-trimethylpsoralen was added, and the cell suspension was irradiated with 20 mW/cm2 UVA light for 3, 4 and 5 min. To evaluate virus inactivation, cells and supernatants were diluted serially and cocultured with uninfected MT-4 cells. Infectious HIV-1 was detected by cytopathic effects, immunofluorescence and p24 antigen ELISA. UVA irradiation at 3.6 J/cm2 (3 min 20 mW/cm2) reduced the amounts of both cell-associated and extracellular infectious HIV-1 by more than five orders of magnitude. Even at more stringent conditions of PUVA treatment (10 min 20 mW/cm2 UVA irradiation), conformational cellular surface epitopes remained detectable by flow cytometry.

Anti-HIV Agents↗
PubMed1997

Tandem high-dose therapy with ifosfamide, epirubicin, carboplatin and peripheral blood stem cell support is an effective adjuvant treatment for high-risk primary breast cancer.

We evaluated the therapeutic efficacy and toxicity of a tandem high-dose therapy with peripheral blood stem cell (PBSC) support in 40 patients with high-risk, primary breast cancer (stage II-III) and involvement of ten or more positive axillary lymph nodes. Their median age was 44 years (range 23-56). Two cycles of cytotoxic chemotherapy with ifosfamide (10000 mg/m2) and epirubicin (100 mg/m2) were administered. Granulocyte colony-stimulating factor (G-CSF) was given to hasten neutrophil reconstitution and to mobilise PBSC during marrow recovery. Leukaphereses were performed following the first and/or second cycle. Tandem high-dose therapy consisted of two cycles with ifosfamide (15 or 12 g/m2) and epirubicin (150 mg/m2), while carboplatin (900 mg/m2) was added for the last 24 patients included. Using an immunocytochemical method, two of 11 patients had cytokeratin-positive tumour cells in three leukapheresis products that were collected following the first G-CSF-supported cycle with ifosfamide and epirubicin, whereas only two harvests obtained following the second cycle in 26 patients contained cytokeratin-positive tumour cells. The number of CD34+ cells/kg re-infused following both high-dose cycles was similar (4.20 +/- 0.29 x 10(6), first cycle and 5.25 +/- 0.63 x 10(6), second cycle), and no notable difference was noted in the speed of haematological reconstitution. An absolute neutrophil count (ANC) of 0.5 x 10(9)/1 was reached after a median time of 13 days, while an unsupported platelet count of 20.0 x 10(9)/1 was achieved after a median time of 8 (first cycle) and 9 (second cycle) days post-transplantation. Patients autografted with more than 7.5 x 10(6) CD34+ cells/kg had platelet counts above 20 x 10(9)/1 within less than 10 days. 6 patients relapsed between 7 and 11 months (median 8 months) post-transplantation. 37 patients are alive and in remission with a median follow-up time of 11 months (range 1-38). This translates into a probability of disease-free survival (DFS) of 77% (95% CI 32-95%) at 38 months. The probability of overall survival is 85%, since 3 patients with local relapse achieved a second complete remission following surgery and involved-field radiotherapy. In conclusion, a sequential high-dose therapy including ifosfamide, epirubicin, carboplatin and PBSC support is well tolerated and effective in patients with high-risk primary breast cancer. Involved-field irradiation should be performed post-transplantation to reduce the risk of local relapse.

Adult↗
PubMed1997

High-dose therapy with peripheral blood progenitor cell transplantation in multiple myeloma.

BACKGROUND: The objective of our study was to evaluate the efficacy and toxicity of a high-dose melphalan-based (HD-Mel) therapy with or without total body irradiation (TBI) followed by peripheral blood progenitor (PBPC1 transplantation in patients with multiple myeloma (MM). PATIENTS AND METHODS: Between June 1992 and May 1996, 100 patients (67 males 33 females with a median age of 51 years range 30-65) were transplanted at our centre. PBPC were collected during G-CSF-enhanced leukocyte recovery following high-dose chemotherapy. Fifty patients were treated with TBI + melphalan 140 mg m2, while 50 patients received melphalan 200 mg/m2. RESULTS: Following PBPC autografting, the median time to reach platelets > or = 20 x 10(9) 1 and neutrophils > or = 0.5 x 10(9)/1 was 11 and 14 days with no difference between the treatment groups. In the TBI group significantly longer periods of total parenteral nutrition were required due to severe mucositis. Two patients from the TBI group died due to transplantation-related complications. Following high-dose treatment, remission state improved in 43 out of 98 patients. No statistically significant advantage in reaching CR or PR was observed with TBI + HD-Mel compared to the treatment with HD-Mel alone. CONCLUSION: Dose-escalated treatments, with particular regard to the inclusion or omission of TBI, should be prospectively investigated to find the best high-dose regimen.

Adult↗
PubMed1997

Elimination of bacteria on different implant surfaces through photosensitization and soft laser. An in vitro study.

Microbiologic examinations of implants have shown that certain microorganisms described as periodontal pathogens may have an influence on the development and the progression of peri-implant disease. This experimental study aimed to examine the bactericidal effect of irradiation with a soft laser on bacteria associated with peri-implantitis following exposure to a photosensitizing substance. Platelets made of commercially pure titanium, either with a machined surface or with a hydroxyapatite or plasma-flame-sprayed surface or with a corundum-blasted and etched surface, were incubated with a pure suspension of Actinobacillus actinomycetemcomitans or Porphyromonas gingivalis or Prevotella intermedia. The surfaces were then treated with a toluidine blue solution and irradiated with a diode soft laser with a wave length of 905 nm for 1 min. None of the smears obtained from the thus treated surfaces showed bacterial growth, whereas the smears obtained from surfaces that had been subjected to only one type of treatment showed unchanged growth of every target organism tested (P < 0.0006). Electron microscopic inspection of the thus treated platelets revealed that combined dye/laser treatment resulted in the destruction of bacterial cells. The present in vitro results indicate that lethal photosensitization may be of use for treatment of peri-implantitis.

Aggregatibacter actinomycetemcomitans↗
PubMed1997

Evidence of linkage between the serotonin transporter and autistic disorder.

The serotonin transporter gene (HTT) is a primary candidate in autistic disorder based on efficacy of potent serotonin transporter inhibitors in reducing rituals and routines. We initiated a candidate gene study of HTT in trios consisting of probands with autistic disorder and both parents. Preliminary transmission/disequilibrium test (TDT) analysis with 86 families revealed no evidence for linkage or linkage disequilibrium between autistic disorder and a polymorphism in the second intron of HTT. However, preferential transmission of a short variant of the HTT promoter was found in the same 86 trios (TDT chi 2 = 4.69, 1 d.f., P = 0.030). In further analyses, we considered haplotypes of the HTT promoter variant and second intron locus as alleles in a multiallelic TDT. Results confirmed the significance of the effect of this region (TDT chi 2 = 11.85, 4 d.f., P = 0.018). This provides preliminary evidence of linkage and association between HTT and autistic disorder.

Adolescent↗
PubMed1997

Immunomagnetic selection of CD34+ peripheral blood stem cells for autografting in patients with breast cancer.

Contamination of transplants with tumour cells may contribute to relapse after peripheral blood stem cell transplantation (PBSCT). We studied the feasibility of CD34+ cell selection from blood-derived autografts obtained following G-CSF-supported cytotoxic chemotherapy in a group of 25 patients with breast cancer (10 with high-risk stage II/III and 15 with stage IV without bone or bone marrow involvement). Using immunomagnetic beads (Isolex 300 SA. Baxter) CD34+ cells were enriched and released by chymopapain resulting in a median purity of 95% (range 82-99%) and a median recovery of 80% (range 27-132%). The enrichment procedure did not change the proportion of CD34+ subsets coexpressing HLA-DR, CD38 and Thy-1, while L-selectin was removed from the cell surface following selection. Using a sensitive immunocytological technique with a cocktail of epithelial-specific antibodies (anti-cytokeratin 8, 18 and 19; HEA125; BM7 and BM8), five leukaphereses products contained epithelial cells, whereas the selected CD34+ cell fraction was free of tumour cells. A neutrophil count of 0.5 x 10(9)/l and a platelet count of 20 x 10(9)/l was reached after a median time of 14 and 10d following 40 high-dose chemotherapy (HDC) cycles. Our results indicate that immunomagnetic selection of CD34+ cells yields highly purified autografts devoid of tumour cells whereas the engraftment ability of the progenitor and stem cells is fully retained.

Adult↗