Tsuguo Kuwata

Effects of interferon on cell and virus growth in transformed human cell lines.

The anticellular and antiviral effects of human leukocyte interferons were studied in vitro in the transformed human embryonic cell lines. RSa and RSb. The growth of these cells was inhibited and they began to deteriorate about 48 h after treatment with 500 units/ml of interferon. When interferon was washed out within 48 h, their growth recovered gradually. The effects of interferon on cell growth depended on the amount of interferon added per cell. A subline, named IFr, was isolated which grows in the presence of 2000 units/ml of interferon, whereas growth of vesicular stomatitis virus in these cells is suppressed by 10 units/ml of interferon, just as in the parent cells. The anticellular and antiviral effects of interferon on IFr cells are discussed in relation to cell surface receptors.

Enzymatic activities induced by interferon in human fibroblast cell lines differing in their sensitivity to the anticellular activity of interferon.

Abstract IF r , a subline of transformed human fibroblast cells, which is sensitive to the antiviral but resistant to the anticellular activity of interferon, was found to be equally well inducible as its parental cell line RSa for the two major interferon-mediated double-stranded RNA-dependent enzymatic activities, 2–5A synthetase and 73K phosphoprotein kinase. The induction of 2-5A synthetase as a function of interferon dose, the specific activity of the 2-5A synthetase, the nature of the 2-5A oligonucleotide products, and the activity of the 2-5A-activated endonuclease were essentially the same for both cell lines. The 73K phosphoprotein kinase was induced at a similar rate of activity, whether detected in solution or after immobilization on poly(I)·poly(C)-Sepharose. Our observations thus suggest that the induction of these two enzymatic activities are not sufficient for the anticellular activity of interferon.

Enhancement by interferon of natural cytotoxic activities of lymphocytes from human cord blood and peripheral blood of aged persons

Abstract Effects of interferon on natural (or “spontaneous”) cytotoxicity of lymphocytes from cord blood and peripheral blood of aged persons were tested in a chromium-release assay against RSb target cells. These natural cytotoxic activities were enhanced by leukocyte and fibroblast interferon as shown in adult lymphocytes.

Effects of Interferon on the Human Clonal Cell Line, RSa: Inhibition of Macromolecular Synthesis

Multiplication of the human clonal cell line, RSa, is completely inhibited by human leukocyte interferon preparations. Synthesis of DNA and protein is markedly reduced in these cells in proportion to the concentration of interferon applied. Interferon treatment leads to accumulation of cells of an epithelial morphology which do not enter the division cycle. It is suggested that the growth inhibitory effects of interferon on RSa cells may result from these effects.

2-5A synthetase activity induced by interferon α, β, and γ in human cell lines differing in their sensitivity to the anticellular and antiviral activities of these interferons

Abstract Two human cell lines (HEC-1, IFr) which are resistant to the anticellular and/or antiviral action of HuIFN-α or -β have been tested with respect to the anticellular and antiviral activities of HuIFN-γ. HEC-1 cells were totally resistant to the antiviral activity of the IFN-γ, with either EMC or VSV as challenge virus. Per antiviral unit HuIFN-γ exerted a much stronger inhibitory effect on the growth of the interferon-sensitive cell line RSa than did IFN-α. IFr cells were more resistant to the anticellular effect of IFN-γ than the wild-type cells (RSa), and HEC-1 cells were totally resistant to the anticellular effect of HuIFN-γ. 2-5A synthetase was present at a high basic level in untreated HEC-1 cells, as previously reported. This activity was not influenced by either HuIFN-β or -γ. Similar levels of 2-5A synthetase were induced by HuIFN-γ in IFr and RSa cells; however, this induction was 10 times lower than that obtained for HuIFN-β. A 2′-phosphodiesterase activity was detected in control extracts from RSa and was not induced by either HuIFN-β or -γ. There was no correlation between the amplitude of 2-5A synthetase induction and extent of anticellular effect of either HuIFN-γ in different cell lines or of different interferons in a given cell line. Finally, HEC-1 represents the first example of a human cell line which is resistant to all three types of interferon (HuIFN-α, -β, and -γ).

Cross-sensitivity between interferon and uv in human cell strains: IFr, HEC-1, and CRL1200

Correlation between susceptibility to the anticellular effect of human interferon (HuIFN) and ultraviolet (uv) lethality was examined in a set of isogeneous human cell lines (RSa and IFr cells), a human endometrial cancer cell line (HEC-1 cells), and a Xeroderma pigmentosum-derived fibroblast cell line ( CRL1200 cells). IFr cells, previously established as a HuIFN-alpha-resistant subline by exposing HuIFN-alpha- and uv-sensitive RSa cells to HuIFN-alpha preparations, appeared more refractory to uv than did the parent RSa cells in the cell proliferation and colony-formation studies. The extent of recovery from uv-inhibited total cellular DNA synthesis and uv-induced DNA-repair synthesis was enhanced to a greater extent in IFr cells than in RSa cells. The preexposure of RSa cells to HuIFN-alpha enhanced uv-induced DNA-repair replication activities. HEC-1 cells, which are reportedly totally refractory to HuIFN actions, appeared most resistant to uv, in all the tests. The uv-sensitive CRL1200 cells appeared highly susceptible to HuIFN-beta, in both cell proliferation and DNA-synthesis inhibition tests. These results support and extend our previous finding (N. Suzuki, J. Nishimaki , and T. Kuwata (1982). Mutat . Res. 106, 357-376) that susceptibility to the anticellular effect of HuIFN closely relates with uv lethality.

Resistance to interferon of a human adenocarcinoma cell line, HEC-1, and its sensitivity to natural killer cell action.

A human endometrial cancer cell line, HEC-1, was found to be resistant to the antiviral and anticellular action of interferon. However, HEC-1 cells were susceptible to the cytotoxicity of natural killer (NK) cells, and interferon enhanced such NK activity. When HEC-1 cells were treated with interferon, sensitivity of HEC cells to the cytotoxicity of NK cells was not suppressed.

Effect of glycolipids detectable in transformed human cells on interferon activities.

Summary Human transformed cell lines, RSa, RSb, and IF r , were analyzed for their glycolipid composition. Ganglioside GM 2 , GM 3 , and 4 neutral glycolipids were identified as the component of these 3 cell lines. Antiviral action of human interferon was neutralized by pretreatment with GM 3 or GM 2 . Lactosylceramide also neutralized antiviral action of human interferon. Thus, these glycolipids are considered to be the components of the receptor for interferon in these cells. However, when GM 3 or ganglioside mixture was tested for the anticellular action of human interferon, they showed no inhibitory action on the growth suppressive activity of interferon.

Comparison of the Suppression of Cell and Virus Growth in Transformed Human Cells by Leukocyte and Fibroblast Interferon

The effects of human leukocyte interferon (Le-IF) and fibroblast interferon (F-IF) on the growth of the transformed human cell lines, RSa and RSb, were compared. Both were considerably more sensitive to F-IF than to Le-IF. Two cell lines, IFr and F-IFr, derived from the RSa cell line, were resistant to the anticellular effects of Le-IF, but less resistant to those of F-IF.

Suppressive Effects of Interferon on Cell Fusion by Sendai Virus

Suppressive effects of human IFN-alpha, IFN-beta and mouse-IFN on syncytium formation in human and mouse transformed cells have been studied using u.v.-irradiated Sendai virus (UV-Sendai virus). After treatment of human RSa cells with 500 units/ml human IFN-alpha for 16 h, the syncytium formation induced by UV-Sendai virus was reduced to less than 10% of controls. The suppressive effect was dependent on the amount of interferon added, and it was blocked by the addition of cycloheximide. Suppression of syncytium formation was also observed on human IFr cells, which are partially resistant to the anticellular effect of interferon but are sensitive to the antiviral effect of interferon. Human IFN-beta also had a suppressive effect on syncytium formation in human transformed cells, and human IFN-alpha, IFN-beta and mouse IFN showed species specificity in their effect on fusion. This anti-cell fusion activity was developed in IFr cells from about 5 h after addition, of IFN-alpha and when the cells were treated with IFN-alpha for 16 h, the resistant state of cell continued for over 20 h after removal of IFN-alpha.