Jean R. Starkey

Inhibition of natural killer cell activity in vitro by alcohols.

Abstract The effect of the addition of small quantities of alcohols to cocultures of natural killer effector cells from C57BL 6 mice and BDIX rats with YAC-1 tumor cell targets has been studied. The order of inhibition of NK cell-mediated killing is 1-butanol > 1-propanol > 2-propanol > ethanol > methanol. The inhibition of killing due to the addition of alcohol correlates with decreases of effector-target cell binding. Therefore caution should be exercised in interpreting results of cellular experiments in which these alcohols have been used to solubilize inhibitors.

Cell Surface Thiols, but not Intracellular Glutathione, are Essential for Cytolysis by a Cloned Murine Natural Killer Cell Line

Cell surface thiols are required for a line of cloned murine natural killer lymphocytes to bind to and lyse tumor target cells. These lymphocytes neither bound to nor killed YAC-1 or G1Tc cells when the effector lymphocyte cell surface thiols were covalently coupled with the non-penetrating reagent, monobromotrimethylammoniobimane (qBBr). A limited number of thiol-bearing proteins were identified by gel electrophoresis on the cell surface using the fluorescence of the group that remains associated with the sulfur molecule. These results indicate that either one or more of these reactive proteins or different cell surface thiol-bearing molecules present at low frequencies are crucial to lymphocyte binding and killing. In contrast, we found little evidence that intracellular thiols are required for natural killer cell activity. Killing was relatively unimpaired when over 90% of lymphocyte glutathione was depleted with DL buthionine-S,R-Sulfoximine (BSO). Blocking the intracellular or the extracellular thiols of tumor targets had no effect on their ability to be lysed. Based on these data, we suggest that infrequently expressed extracellular thiols are required either for the conformation or for the disulfide crosslinking of proteins that participate in lymphocyte-mediated cytolysis.

Immunoselection of tumor variants resistant to antibody-mediated cytotoxicity - Their immunologic and metastatic characterization

SummaryThe immunological characteristics of two series of metastatic variants of restricted genetic origin were related to their lung-colony-forming potential. A series of metastatic variants was isolated from a tumor-cell population in which heterogeneity appeared following short-term in vivo passage, while a second series of variants were immunoselected in vitro for resistance to antibody-complement-mediated cell lysis. In the case of the first series, the sensitivity of the individual variants to cell-mediated and antibody-complement-mediated cytotoxicity appeared to be partly determined by the number and rate of loss of antibody-binding sites on the cell surface. These characteristics also correlated with the efficiency of experimental metastasis in the animal. We compared the results reported herein with our previous studies of nonimmune parameters for this series of metastatic variants, and we conclude that immunological differences can be important to the efficiency of lung-colony formation. However, in the case of the second series of variants, despite selection in vitro for resistance to antibody-complement-mediated cell lysis, the behavior of these variants in the lung colony assay could not be predicted by the immunologic parameters examined.

Natural killer cell activity in fawn-hooded rats

Fawn-hooded (FH) rats were shown to lack the genetically conditioned defect of natural killer (NK) activity hypothesized to be present by analogy with the Chediak-Higashi syndrome (CHS) in mice and human beings. In 4-h51Cr release assays, splenic NK cells from FH rats killed YAC-1, RL♂ 1 and G1-TC tumor targets without deficiency based upon comparison with cells from BD-IV, BD-IX and NBR inbred rat strains. Progeny of BD×FH F1 rats backcrossed to FH failed to reveal a correlation of reduced NK activity and dilute coat color. From these, and other data presented, it is concluded that despite similarities in coat color dilution and platelet storage pool deficiency, FH rats do not closely resemble CHS mice or human beings in having deficient NK activity and cannot be considered the rat homolog of the CHS.

Immunologic enhancement of experimental metastasis in the rat

SummaryUsing a series of immunologically cross-reactive metastatic tumor variants, we demonstrate that serum from animals bearing pulmonary tumor colonies possesses enhancing properties in the experimental metastasis (lung colony) assay. Enhancement is produced by chronic serum administration and promotes the growth of tumor cells arrested in the lungs which would not otherwise proliferate to form grossly detectable lung nodules. Tumor-bearer serum from animals with lung colonies derived from the most highly metastatic variant examined is shown to possess enhancing properties in both BD-IX(H-1d) and BD-IV(H-1d) rat strains, while tumor-bearer serum from animals with lung colonies derived from the less metastatic parent tumor cell line possesses enhancing properties in the BD-IX rat strain only. Removal of immunoglobulin from enhancing serum by affinity column chromatography simultaneously removes the enhancing factor(s), and enhancing activity correlates with the presence of increased levels of Clq-binding immune complexes in the serum. Serum levels of immune complexes are shown to be more elevated in serum from animals bearing lung colonies derived from the most highly metastatic variant. The enhancing moieties are shown to bind to concanavalin A, but not to staphylococcal protein A, and the active fraction elutes from concanavalin A-Sepharose with α-methyl-mannoside. Consideration of immunoprecipitation studies on whole and fractionated enhancing sera, along with studies on affinity purified isotype fractions reveals that the activity resides with antibodies of IgG2b subclass.