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Biomedical subjects

R M Welsh

Publications and source records attributed to R M Welsh.

152 records · Page 9Linked to original sources
PubMed1972

Arenaviruses: cellular response to long-term in vitro infection with parana and lymphocytic choriomeningitis viruses.

Persistent infections were established in suspension cultures of BHK21/13S cells with both Parana and lymphocytic choriomeningitis viruses. Four generations after infection with either virus, more than 90% of the cells scored as infective centers, with concomitant peaks in extracellular virus yields. In both cultures the synthesis of detectable plaque-forming units (PFU) ceased about the 50th generation postinfection, and this condition was maintained until the 350th cell generation when the cultures were discontinued. The generation time of each culture was identical to that of uninfected parent controls, and at no time were cytopathic effects evident. In spite of the absence of infectivity, over 90% of the cells sampled at various times contained viral antigen demonstrable by immunofluorescence. When either of these persistently infected cell lines was substituted for normal cells in the standard plaque assay, very low efficiencies of plating were observed for homotypic and heterotypic viruses. Plaque formation by several heterologous viruses was virtually unaffected. The mechanism of homotypic plaque exclusion in both cell lines was shown to occur beyond the virion adsorption stage. The original infecting virus genome persisted in both cell lines after standard virus was no longer detectable. This was shown with the lymphocytic choriomeningitis virus-infected cells after storage in liquid nitrogen. After thawing, such cells were found to synthesize standard virus for a brief period. Although the Parana virus-infected cells did not behave this way, the growth medium from these cells would initiate PFU synthesis in normal cells within 36 hr after infection.

Adsorption↗
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Window of vulnerability of vaccinia virus-infected cells to natural killer (NK) cell-mediated cytolysis correlates with enhanced NK cell triggering and is concomitant with a decrease in H-2 class I antigen expression.

A time course study was performed in order to determine if vaccinia virus (VV)-infected targets were more susceptible to murine natural killer (NK) cell-mediated lysis during a discrete period of time postinfection. Activated NK cells were used in short-term (e.g. 4 h) assays in order to avoid a further in vitro activation of the NK cells by interferon (IFN) and to test the innate susceptibility of target cells to lysis. The sensitivity of VV-infected L929 cells to lysis by NK cells increased as the infection progressed, reached a peak at approximately 24 h postinfection, and subsequently declined to levels lower than that of uninfected cells. This window of vulnerability was not due to an increase in the number of effector/target cell conjugates, which continually decreased as the VV infection progressed. Triggering of NK cells was measured by the influx of 45Ca2+. Target cells treated with IFN induced less 45Ca2+ uptake, whereas cycloheximide treatment of targets caused a greater influx of 45Ca2+ into the effector cells. When L929 cells were infected with VV for various time intervals and used in the triggering assays, an enhanced triggering of the effectors corresponding to the time of enhanced susceptibility of the target cells to lysis was detected. Quantitative decreases in H-2Kk and Dk class I antigens were observed following VV infection of target cells as measured by FACS analysis using alloantibodies. Qualitative changes in H-2 class I antigens were also observed, as detected by a loss in VV-infected target cell susceptibility to lysis by allospecific cytotoxic T lymphocytes (CTL) at a time when they were highly sensitive to killing by NK cells and VV-specific CTL. These results show that virus-infected targets may become innately more sensitive to lysis by NK cells at discrete time points after infection and that the susceptibility to lysis correlates with enhanced triggering of NK cells and reduced H-2 class I antigen expression.

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The biological effects of allogeneic effect factor (AEF) on T lymphocytes. III.--Interferon does not contribute to the biological activities displayed by AEF on both T and B lymphocytes.

The studies reported here were designed to evaluate the potential contribution of interferon (IF) to the biological activities mediated by allogeneic effect factor (AEF), a soluble product of allogeneic cell interactions, on responses of T and B lymphocytes. AEF supernatants were found to contain varying levels of IF, predominantly of type II (immune). AEF preparations which were practically free of--or very low in--IF activity were obtained by bovine serum albumin-Sepharose chromatography of AEF, or by using strain combinations involving K-only or I-only differences within the H-2 complex for the production of AEF. Such preparations retained their biological activities in three in vitro assays characteristic of AEF: (a) induction of primary self-H-2-reactive cytotoxic T-lymphocyte responses, (b) mitogenicity for normal T cells, and (c) induction of plaque-forming cell antibody responses to a T-dependent antigen in T-cell-depleted spleen cultures. These results demonstrate that IF does not play any significant role in the biological activities medicated by AEF.

Animals↗
PubMedDate not supplied

Antiviral activity of NK 1.1+ natural killer cells in C57BL/6 scid mice infected with murine cytomegalovirus.

The activation, proliferation, and antiviral effects of natural killer (NK) cells were examined in a newly developed stock of mice, C57BL/6JSz mice homozygous for the severe combined immunodeficiency (scid) mutation. These mice lack functional T and B cells and express the NK 1.1 alloantigen. Such NK 1.1 expression facilitates the analysis of NK cells and their depletion in vivo with a monoclonal anti-NK 1.1 antibody. These mice, therefore, provide an excellent model to examine unambiguously the interactions between viral infections and NK cells in a system devoid of adaptive immune response mechanisms. Here we show that murine cytomegalovirus (MCMV) and lymphocytic choriomeningitis virus (LCMV) infections resulted in profound levels of NK cell activation. NK cells also proliferated greatly in response to LCMV but generally to a lesser degree in response to MCMV. Depletion of the NK cell activity in vivo caused substantial increases in MCMV synthesis and MCMV-induced pathology. These results further support the concept that NK cells are major regulators of MCMV pathogenesis.

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PubMedDate not supplied

B cells induce apoptosis via a novel mechanism in fibroblasts infected with mouse hepatitis virus.

B cells from nonimmune mice mediate the cytolysis of fibroblasts infected with the coronavirus, mouse hepatitis virus (MHV), strain A59. In this investigation, we report that splenic B cells and a B cell hybridoma induced the fragmentation of MHV-infected target cell DNA into a nucleosomal ladder pattern, characteristic of apoptosis. To determine the mechanism by which B cells mediated this killing event, we used criteria previously established for the killing of target cells by cytotoxic T lymphocytes (CTLs) and compared this B-cell-mediated killing to lymphocytic choriomeningitis virus (LCMV)-specific CTL killing of LCMV-infected target cells. Unlike CTL-mediated cytotoxicity, B cells efficiently lysed and induced the fragmentation of the DNA in their target cells in the presence of EGTA, arguing against a Ca(2+)-dependent granule exocytosis model for killing. In addition, paraformaldehyde-fixed B cells were able to kill MHV-infected targets. We were unable to detect TNF-alpha-associated cytotoxicity via bioassay with nonimmune effector B cells against the TNF-sensitive cell line, LM, or the TNF-alpha-resistant subline, L929.w, infected with MHV. Serine esterase inhibitors (benzamidine hydrochloride and N alpha-p-tosyl-L-arginine methyl ester) blocked CTL-induced 51Cr release and DNA fragmentation. In contrast, the inhibitors did not block the B-cell-induced 51Cr release, but did cause an inhibition in the fragmentation of the DNA of the target cell. These data indicate that B cells are capable of inducing the lysis and DNA fragmentation of MHV-infected target cells similar to CTL-induced apoptosis. However, we show that the mechanism(s) by which these processes are induced by B cells is distinct from CTL-mediated cytotoxicity.

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