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Biomedical subjects

H Rabenau

Publications and source records attributed to H Rabenau.

At least 55 records · Page 3Linked to original sources
PubMed1994

Replication of human cytomegalovirus in a rhabdomyosarcoma cell line depends on the state of differentiation of the cells.

The replication of human cytomegalovirus (HCMV) was investigated in a new human rhabdomyosarcoma cell line (KFR) with morphological and biochemical characteristics of fetal striated muscle precursors (rhabdomyoblasts). KFR cells exhibited the unique property for spontaneous morphological transformation from a poorly-differentiated state into well-differentiated (myotube-like) rhabdomyoblasts. The poorly-differentiated rhabdomyoblasts promoted both complete viral gene expression and the production of infectious virus. In contrast, in well-differentiated rhabdomyoblasts HCMV infection was abortive. The results showed that replication of HCMV in this human rhabdomyosarcoma cell line depended on the state of cellular differentiation.

Antigens, Viral↗
PubMed1994

In vitro inhibition of human cytomegalovirus replication by desferrioxamine.

Desferrioxamine (DFO) is commonly used in therapy as a chelator of ferric ion in disorders of iron overload. We found that DFO inhibits human cytomegalovirus (HCMV) replication in infected cultures of human foreskin fibroblasts (HFF) at concentrations that have been achieved in humans with no significant adverse effects. The concentrations of DFO required for 50 and 90% reduction in the production of a HCMV-late antigen ranged for several HCMV strains from 3.1 to 4.9 microM and from 14.2 to 17.3 microM, respectively. DFO concentration of 60 microM had no significant effect on the viability of HFF cells. Inhibitory effects of DFO on HCMV replication were completely prevented by co-incubation with stoichiometric amounts of Fe3+.

Cell Division↗
PubMed1994

Evaluation of a new automated microneutralization assay for the quantitative detection of neutralizing antibodies against enteroviruses.

An automated microneutralization assay for the quantitative detection of neutralizing antibodies (NA) against polioviruses and non-polio enteroviruses (NPEV) using a pipetting roboter (Tecan RSP 5072) was established and compared to the conventional manually performed test procedure. The qualitative neutralizing antibody detection was not significantly influenced by the assay system (manual or automated assay). Concerning the quantitative antibody detection, two-fold titre differences between the two test systems were observed in only 2.3% of the 260 serum samples investigated. The intra-assay and inter-assay variability of the quantitative detection of neutralizing antibodies using the automated assay proved to be very low. The quantitative detection of neutralizing antibodies using an automated pipetting robot permitted the testing of large numbers of samples within a shorter period and with less labour intensity as compared to the manually performed assay. Therefore it represents a valuable alternative to the conventional microneutralization test, especially for the serodiagnosis of non-polio enterovirus infections in large sample collectives, assessment of immunity to polioviruses and for seroepidemiological surveys.

Animals↗
PubMed1994

Quantitative detection of neutralizing antibodies against polioviruses and non-polio enteroviruses (NPEV) using an automated microneutralization assay: a seroepidemiologic survey.

A seroepidemiological survey for the assessment of immunity against polioviruses and non-polio enteroviruses (NPEV) seroprevalence was performed using a recently established automated microneutralization assay. The seroprevalences of neutralizing antibodies against Coxsackie viruses B1-B6, and A9, echoviruses 6, 9, 11, and 30, and polioviruses 1, 2, and 3 were investigated in serum samples collected over a period of 30 months (January 1990 to June 1992). Overall, NPEV seroprevalence ranged from 8.3% (Coxsackie virus B6) to 69.8% (echovirus 30). Age-related increases of seroprevalence were only observed for Coxsackie viruses B2 and A9. Relatively low antibody titres were found, the average antibody titre being the highest for Coxsackie virus B2 (1/55.4). The overall poliovirus seropositivity rate reached 80%. The highest seroprevalence was observed in the higher age groups (> or = 50 years). A relatively high proportion (7%) of individuals did not show neutralizing antibodies against all the three poliovirus serotypes. The results of our study have shown that, using an automated microneutralization assay, it is possible to perform seroepidemiological surveys in large patient populations. Until now, no comparable study concerning the NPEV seroprevalence has been performed.

Adolescent↗
PubMed1994

Failure of antiretroviral therapy: role of viral and cellular factors.

Effective therapy of human immunodeficiency virus (HIV) infection is mainly based on inhibition of reverse transcriptase by nucleoside analogues such as zidovudine (azidothymidine; AZT), didanosine, and zalcitabine. A major problem associated with long-term AZT therapy is the waning efficacy ('clinical resistance') over time. Clinical isolates of HIV-1 with reduced susceptibility to AZT can be recovered from HIV-infected individuals under prolonged treatment. However, the clinical importance of AZT resistance is uncertain. Other factors such as increased virus burden, increased virulence, and AZT toxicity could contribute, singly or in combination, to the loss of therapeutic benefit. Recent observations based on experimental models and clinical trials suggest that cellular mechanisms ('cellular resistance') may account for clinical resistance to antiviral agents. In vitro experiments demonstrated that in analogy to antitumoral therapy, the acquisition of multidrug resistance, i.e., resistance of cells to multiple, structurally unrelated chemotherapeutic agents, may play a role in the failure of long-term antiretroviral therapy. The 'cellular resistance' may contribute directly to the failure of antiviral therapy by the generation of subtherapeutic levels of antiviral compounds and/or their active forms. Indirectly, such subtherapeutic concentrations of active substances which permit limited replication of virus may represent a selective pressure for emergence and development of a resistant virus population. Hence it is of great importance to investigate the role of cellular factors in 'clinical resistance' to AZT and other anti-HIV agents. More detailed knowledge of cellular interactions and antiviral agents could help to improve or develop new strategies for antiviral therapy regimens.

Antiviral Agents↗
PubMed1994

Suspension culture of HeLa cells in protein-free medium: sensitivity to human pathogenic viruses.

Human adherent HeLa-PF cells grown for 5 years in a protein-free 1:1 nutrient mixture of Dulbecco's modified MEM and Ham's F12 (DMEM-F12) were established in suspension culture. The cells grew in protein-free DMEM-F12 (using magnetically stirred flasks) as a monodisperse suspension with a population doubling time of 28 h. The cells were infected with poliovirus types 2 and 3, herpes simplex virus types 1 and 2 (HSV-1 and HSV-2), respiratory syncytial virus (RSV), echovirus 6 and adenoviruses 3 and 7. Polioviruses replicated in suspension culture of HeLa-PF cells to a similar extent as in protein-free and serum-supplemented adherent cultures. RSV and HSV developed significantly lower titers while adenoviruses and echovirus 6 developed significantly higher titers in suspension than in adherent cultures. The results show that suspension culture of HeLa-PF may both provide the advantage of a high virus yield and enable cultivation of viruses which are not contaminated by serum or other proteins usually added to the culture medium.

Cell Culture Techniques↗
PubMed1994

Detection of primary HIV infection by a second-generation HIV(p24) antigen test.

We studied the course of a primary HIV infection in a 54-year-old woman. Probably the source of infection was sexual intercourse, since other risks, such as i.v. drug use, acupuncture and transfusion were excluded. On admission she presented with fever, a maculopapular rash, and moderate enlargement of nuchal lymph nodes. At that time the anti-HIV 1,2 enzyme immunoassay was negative. However, the HIV-p24 Ag test, which was performed in every HIV screening in our laboratory, was positive. The suspicion of an acute HIV infection was supported by a positive HIV-cDNA-PCR and confirmed by Western blot after seroconversion. As additional finding, the blood smear showed abnormal white cell differential count, indicating viral infection. Aminotransferases were slightly increased, and antibodies to hepatitis B surface and core antigens demonstrated former hepatitis B infection. It is concluded that in this case the HIV-p24 Ag test proved its suitability for early diagnosis of an acute HIV infection. In case of testing blood donors, none of the compulsory serological screening methods would have detected the HIV infection.

AIDS Serodiagnosis↗
PubMed1994

In vitro anti-human cytomegalovirus activity of liposome-encapsulated foscarnet.

Foscarnet is a pyrophosphate analogue that possesses substantial activity against human cytomegalovirus (HCMV) in patients with CMV retinitis. In this in vitro study we tested whether liposome encapsulation of foscarnet would provide the drug with anti-HCMV activity comparable with that of nonencapsulated foscarnet. The drug was entrapped in large unilamellar vesicles of homogeneous size at a concentration of 10 mg/ml. The vesicles were prepared by the method of fast and controlled dialysis of mixed detergent-lipid micelles using N-Octyl--D-glucopyranoside as the detergent. The activity of drugs against two laboratory HCMV strains (AD169 and Towne) was tested in a plaque-reduction assay using monolayers of human foreskin fibroblasts. The dose of encapsulated or nonencapsulated foscarnet required for a 50% reduction in the number of plaques (IC50) for the AD169 strain was 106 and 113 mumol/l, respectively. For the HCMV Towne strain the dose was 112 and 109 mumol/l, respectively. Treatment with concentrations of up to 400 mumol/l (maximal concentration tested) demonstrated that foscarnet is released at efficient doses from liposomes and that it is not toxic in the range of concentrations tested.

Cells, Cultured↗
PubMed1993

In vitro differentiation of human neuroblastoma cells induced by sodium phenylacetate.

Sodium phenylacetate (NaPA) at concentrations ranging from 2 to 6 mM stimulated morphological differentiation of two human neuroblastoma cell lines IMR-32 and UKF-NB-3. These concentrations inhibited growth and DNA synthesis of the cells in a dose dependent manner without significant effect on cell viability. The differentiated cells showed pseudoganglia formation and extension of cellular processes. The morphological differentiation in both cell lines was accompanied by decreased expression of N-myc oncoprotein. These results suggest that NaPA at concentrations, which have been achieved in humans with no significant adverse effects, promotes differentiation of cultured human neuroblastoma cells in association with the reduced expression of the malignant phenotype.

Cell Differentiation↗
PubMed1993

Detection of neutralizing antibodies against human cytomegalovirus: influence of strain variation.

The influence of human cytomegalovirus (HCMV) strain variation on neutralizing antibody titers was investigated in sequential sera obtained from 12 organ transplant recipients (11 renal transplant recipients, 1 liver transplant patient) suffering from primary or secondary HCMV infection. Cross-neutralization assays using either the international HCMV reference strain AD169 or individual clinical isolates from patients showed congruent results. Restriction enzyme analysis of the hypervariable alpha-sequence of the L-S junction of AD169 and HCMV isolates amplified by polymerase chain reaction (PCR) were carried out to confirm the expected strain differences. At least 6 groups of clinical strains were differentiated. The results of this study demonstrated that despite the strain heterogenicity of HCMV, the neutralization assay using AD169 permitted a reliable and quantitative serologic detection of neutralizing antibodies against HCMV.

Antibodies, Viral↗
PubMed1993

Contamination of genetically engineered CHO-cells by epizootic haemorrhagic disease virus (EHDV).

The characterization of a contaminating virus which was detected in genetically-engineered Chinese hamster ovary (CHO) cells during the production of biologicals is described in the present paper. Under electron microscopy, the contaminating virus had a morphology resembling that of an orbivirus. The relationship was confirmed by nucleic acid analysis which showed a RNA segment pattern characteristic of orbiviruses. With an immunoperoxidase staining of monolayer cells and through sero-neutralization tests the virus was identified as being identical to Epizootic Haemorrhagic Disease Virus (EHDV), isolate 318 (untyped) from Bahrain. Potential sources of the contaminating virus and feasible procedures to avoid adventitous virus infections in cell cultures are discussed.

Animals↗
PubMed1993

Protein-free culture of Vero cells: a substrate for replication of human pathogenic viruses.

A protein-free chemically defined medium designated PFEK-1 was developed for culture of VERO cells on polyvinyl formal (PVF) culture surface without serum or other macromolecular supplements. VERO cells proliferated in PFEK-1 medium on PVF surface to a similar extent as cells in serum-supplemented medium without previous adaptation from serum-containing conditions. The protein-free culture infected with coxsackievirus B4, herpes simplex virus types 1 and 2, measles virus and poliovirus types 1, 2 and 3 developed viral titers comparable to those found in conventionally grown cells. The results demonstrated that VERO cells in protein-free culture provide a sensitive substrate for the production of human pathogenic viruses which are not contaminated by serum or other protein factors usually added to a culture medium.

Animals↗
PubMed1993

Decreased anti-human immunodeficiency virus type-1 activities of 2',3'-dideoxynucleoside analogs in MOLT-4 cell sublines resistant to 2',3'-dideoxynucleoside analogs.

Human T-lymphoid MOLT-4 cells were grown continuously for more than 1 year in medium containing either 3'-azido-2',3'-dideoxythymidine (AZT), 2',3'-dideoxyinosine (ddI) or 2',3'-dideoxycytidine (ddC) at concentrations similar to peak plasma levels found in clinical trials in patients with AIDS. To test antiviral activities of the nucleoside analogs against HIV-1 in the cell sublines designated MOLT-4r-AZT, MOLT-4r-ddI and MOLT-4r-ddC, the number of infected cells, p24 HIV-1 antigen in culture medium and syncytium formation of infected cultures were determined. The results showed that anti-HIV-1 activities of AZT, ddI and ddC were significantly decreased in the resistant MOLT-4 cell sublines grown continuously with the respective nucleoside analog, probably due to the development of cell populations resistant to the drugs.

Cell Line↗
PubMed1993

In vitro anti-human immunodeficiency virus activity of 2',3'-dideoxynucleosides and their effect on clonal growth of hemopoietic cells from human bone marrow.

Seven 2',3'-dideoxynucleosides synthesized by substitution of nucleosides using nucleoside deoxyribosyltransferase from Lactobacillus leichmanii were tested for their anti-human immunodeficiency virus (HIV) activity. Two of them, including 2,6-diaminopurine-2',3'-dideoxyriboside (DAPDDR) and 6-chlorpurine-2',3'-dideoxyriboside (CPDDR) demonstrated high antiviral activity against several strains of HIV-1 and one strain of HIV-2. The selectivity index of the drugs (SI; ratio of the drug concentration required for 50% of cell killing to drug concentration required to inhibit 50% of virus-induced cell killing) was established by application of tetrazolium (MTT) colorimetric assay. SI ranged for different HIV strains from 501 to 850 and from 60 to 118 for DAPDDR and CPDDR, respectively. Both DAPDDR and CPDDR retained their antiviral activity against HIV-1 strain D148/88 which was resistant to Zidovudine (3'-azido-3'-deoxythymidine, AZT). Assays for clonal growth of human bone marrow cells in semisolid fibrin clot culture medium demonstrated that DAPDDR possesses significantly lower inhibitory activity for erythroid (BFU-E), multipotent (GEMM-CFC) and granulocyte-monocyte (GM-CFC) bone marrow progenitor cells than CPDDR or AZT. These results suggest that DAPDDR is a nucleoside analog which should be further tested as an anti-HIV compound especially in combination with other anti-retroviral drugs.

Antiviral Agents↗
PubMed1992

Aphidicolin selectively kills neuroblastoma cells in vitro.

Aphidicolin is a tetracyclic diterpene antibiotic which is known to inhibit the growth of eucaryotic cells by reversible binding to DNA polymerase alpha without significant effect on cell viability in most common human cell lines. We observed that aphidicolin at a concentration of 5 x 10(-7) M kills all cells of four human neuroblastoma cell lines. In contrast, viability of normal human embryonal cells and of human continuous cell lines including HeLa, H9, A549 and Caco-2 was influenced only moderately by aphidicolin. In addition, neuroblastoma cells were killed after treatment with 5 x 10(-7) M aphidicolin in cocultures with normal embryonal cells which continued to proliferate after removal of aphidicolin. These results show that aphidicolin provides an agent which selectively kills neuroblastoma cells in vitro.

Antineoplastic Agents↗
PubMed1992

Comparison of shell viral culture and serology for the diagnosis of human cytomegalovirus infection in neonates and immunocompromised subjects.

The present retrospective study compares the laboratory diagnosis of cytomegalic inclusion disease (CID) by the use of "shell vial culture" [i.e., immunoperoxidase staining of human cytomegalovirus (HCMV) early antigen in human fibroblasts 24 h postinoculation] to the results of serology (i.e. immunoglobulins IgG, IgM, and IgA HCMV antibody testing) in 21 infants with congenital or postnatally acquired HCMV infection, 5 patients with lymphoproliferative disorders, 35 human immunodeficiency virus (HIV)-seropositive patients who met the Centers for Disease Control (CDC) criteria for stages IVA and IVB of HIV infection, and 115 patients suffering from the acquired immunodeficiency syndrome, AIDS (stages IVC-IVE according to CDC criteria). HCMV infection was diagnosed by means of the shell vial culture inoculated with patient samples (e.g., urine, bronchoalveolar lavage, induced sputum, etc.) and serology in 163 (92.6%) and 65 (36.9%) patients, respectively. Viral shedding was detected by shell vial culture in 100% of the neonates, 80% of the patients suffering from lymphoproliferative disorders, 100% of the AIDS related complex (ARC) and 89.6% of the AIDS patients. In contrast, serologic testing for HCMV-specific antibodies was positive in only 28.6%, 42.9%, and 34.8% of the neonates, ARC, and AIDS patients, respectively. In lymphoma patients, serologic testing gave identical results (80%) to the shell vial culture technique. With the use of the shell vial procedure, active HCMV infection in immunocompromised subjects and neonates can be recognized more reliably than by serologic testing. Nevertheless, in a low percentage of patients (7.4%), virus isolation by the shell vial culture may fail to detect HCMV infection.

Acquired Immunodeficiency Syndrome↗
PubMed1992

Increased HIV-1 production in chronically infected H9 cells grown in protein-free medium.

Human T cell line H9 was established in a protein-free 1:1 mixture of Ham's F-12 and IMDM. After 230 passages (3 years) in protein-free medium, the cells designated H9-PF were infected with HIV-1. The infectivity titers of HIV-1 in cell culture medium were monitored by determining the median tissue culture infectious doses (TCID50). Additionally, the production of viral antigen in cells was measured by an immunoenzymatical alkaline phosphatase anti-alkaline phosphatase (APAAP) method using a monoclonal antibody against HIV-1-p24 antigen. In acutely infected H9-PF and H9 cultures similar TCID50 values and percentage of cells positive for p24 antigen were found. In contrast, both TCID50 values and percentage of cells positive for p24 antigen were by far greater in chronically infected H9-PF than in H9 cultures.

Cell Line↗