Biomedical subjects

Annette Ohnemus

Publications and source records attributed to Annette Ohnemus.

3 recordsLinked to original sources

PubMed2004

Synthesis of IFN-beta by virus-infected chicken embryo cells demonstrated with specific antisera and a new bioassay.

Transcripts of interferon-alpha (IFN-alpha) and IFN-beta genes are present in virus-infected chicken cells, but because of a lack of appropriate assays and reagents, it was unclear if biologically active IFN-beta is secreted. We have established a nonviral bioassay for the sensitive detection of chicken IFN (ChIFN). This assay is based on a quail cell line that carries a luciferase gene that is controlled by the IFN-responsive chicken Mx promoter. Luciferase activity was strongly stimulated when the indicator cells were incubated with ChIFN-alpha, ChIFN-beta, or ChIFN-gamma but not with chicken interleukin-1beta (ChIL-1beta). Unlike the classic antiviral assay that preferentially detects ChIFN-alpha, the Mx-luciferase assay detected ChIFN-alpha and ChIFN-beta with similar sensitivity. With the help of this novel assay and with rabbit antisera specific for either IFN-alpha or IFN-beta, we analyzed the composition of IFN in supernatants of virus-infected chicken embryo cells. Virtually all IFN produced in response to Newcastle disease virus (NDV) was IFN-alpha. However, IFN produced in response to influenza A or vaccinia virus (VV) was a mixture of usually more than 80% IFN-alpha and up to 20% IFN-beta. Thus, IFN-alpha and IFN-beta both contribute to the cytokine activity in supernatants of virus-infected chicken cells. Furthermore, the infecting virus appears to determine the IFN subtype composition.

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PubMed2003

A sensitive bioassay for chicken interleukin-18 based on the inducible release of preformed interferon-gamma.

Conventional tests for measuring the biological activity of chicken interleukin (IL)-18 require primary chicken spleen cells. We now describe a sensitive bioassay that is based on interleukin-18-induced release of interferon (IFN)-gamma by a permanent chicken cell line. In B19-2D8 cells, cytoplasmically stored interferon-gamma is quickly secreted in response to interleukin-18 exposure.

Animals↗

PubMed2003

Truncated chicken interleukin-1beta with increased biologic activity.

Chicken interleukin-1beta (ChIL-1beta) is synthesized as a precursor molecule that unlike its mammalian counterpart, lacks a typical caspase-1 cleavage site. Therefore, it was unclear if proteolytic cleavage of ChIL-1beta can occur and if cleavage might modulate the biologic activity of this cytokine. Using an avian indicator cell line that carries an NF-kappaB-regulated luciferase reporter gene, we established a sensitive and highly specific bioassay for ChIL-1beta. Experiments with a rabbit antiserum indicated that the NF-kappaB-stimulating activity in supernatants of lipopolysaccharide (LPS)-treated chicken HD-11 macrophages is largely due to IL-1beta and that proteolytic processing of natural and recombinant ChIL-1beta is not very efficient. Functional analyses further revealed that cDNAs for either full-length or N-terminally truncated chicken ChIL-1beta yielded active cytokine. A truncated molecule that closely resembled putative mature ChIL-1beta exhibited more than 100-fold enhanced biologic activity after expression in mammalian cells, indicating that precursor cleavage is indeed of critical importance for maximal activity.

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