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Summary A patient died from symptoms of flaccid paralysis of extremities without showing any classical symptoms of rabies. The autopsy was perfomed, and the distribution of lesions and pathological changes in the central nervous system was similar to poliomyelitis. However, by isolating a virus and its identification, the patient was diagnosed as paralytic rabies.

The effects of human alpha and beta interferons (IFN) on the production of HBsAG by PLC/PRF/5 cells, an HBsAg-producing human hepatoma cell line, were studied in the exponential and stationary phases of cell growth. When exponential phase cells were treated with 100 or 1,000 U of IFN per ml for 48 hr. the amount of HBsAg in the culture medium decreased. The number of cells and the synthesis of DNA and proteins were also reduced by the IFN treatment. These results suggested that IFN did not affect the production of HBsAg specifically in exponential phase cells. When cells in the stationary phase were similarly treated with IFN, HbsAg production was not inhibited nor did the number of cells decrease. To examine the antiviral state induced by IFN in PLC/PRF/5, induction of 2'5'-oligo (A) synthetase and susceptibility to two kinds of viruses were examined. The 2'5'-oligo (A) synthetase activity was increased in an IFN-dose dependent manner. Susceptibility to vesicular stomatitis virus (VSV) and encephalomyocarditis virus (EMCV) was decreased by treatment with 10 and 100 U of IFN per ml for 20 hr. It was concluded that IFN-alpha and IFN-beta induce 2'5'-oligo (A) synthetase and the antiviral state, but do not inhibit HBsAg production by PLC/PRF/5 cells.

Cholera toxin inhibits human natural cell-mediated cytotoxicity in a dose- and time-dependent manner. Pretreatment of lymphocytes with 10 ng/ml of cholera toxin for 2 h almost completely inhibited cytolysis. Interferon augmented human natural cell-mediated cytolysis, but when lymphocytes were pretreated with cholera toxin before interferon treatment, no enhancement of cytolysis occurred. Cholera toxin could inhibit the enhancement of cytolysis by interferon even when lymphocytes were treated with cholera toxin after 2 h interferon treatment. Cholera toxin subunit B which binds cell surface ganglioside galactosyl-N-acetylgalactosaminyl - [N-acetylneuraminyl] - galactosylglucosylceramide (GM1) without activating adenyl cyclase had no effect either on natural cytolysis or on the enhancement of natural cytolysis by interferon, suggesting that mere binding of cholera toxin to the cellular receptor was not enough to inhibit natural cell-mediated cytolysis. Cyclic adenosine 3',5'-monophosphate (cAMP) levels increased in cholera toxin-treated lymphocytes and the time course of cAMP accumulation was similar to that of cytotoxicity inhibition. Exogenous dibutyryl-cAMP (db-cAMP) and theophylline inhibited cytolysis, while exogenous dibutyryl cyclic guanosine 3',5'-phosphate (db-cGMP) enhanced cytolysis slightly, suggesting that the process of inhibition of human natural cell-mediated cytolysis was at least partly modulated by intracellular cyclic nucleotides.

Human leukocyte interferon inhibits the proliferation of the virus-transformed human embryonic cells RSa. Incorporation of [3H]thymidine (TdR) into an intracellular pool and the activity of TdR kinase were reduced in the interferon-treated RSa cell culture. High-degree (90%) inhibition of [3H]TdR incorporation was associated with concentrations of added interferon that produced more than a twofold increase in the intracellular cyclic AMP (cAMP) level, and low-degree inhibition was associated with smaller increases in cAMP. In the IFr cell culture, which is relatively resistant to the anticellular action of interferon, considerably less inhibition of TdR incorporation and a slight increase in cAMP were observed. Extracellularly added dibutyryl-cAMP inhibited the proliferation of both RSa and IFr cells to almost the same degree. A decrease in cAMP level and the initiation of DNA synthesis of G1-phase-arrested RSa cells by serum addition were prevented when cells were pretreated with interferon. These results indicated that intracellular cAMP may mediate the inhibitory effect of interferon on DNA synthesis and cell growth.

Human alpha interferon (IFN-alpha) and beta interferon (IFN-beta) showed antiviral and anticellular effects on human lymphoblastoid Daudi and P3HR-1 cells, but up to 1,000 units/ ml of gamma interferon (IFN-gamma) showed no such effect. Though a fairly high level of dsRNA-dependent 2'-5'-oligoadenylate synthetase (2-5A synthetase) was found in Daudi cells, treatment of these cells with IFN-alpha and beta enhanced the enzyme level in cells at least four-fold, but IFN-gamma did not show any such effect. Lymphoblastoid Raji cells were insensitive to the antiviral and anticellular activities of IFN-alpha, beta and gamma, but 2-5A synthetase was induced in cells by the treatment with IFN-alpha and beta, though the enzyme level was lower than that found in interferon-treated Daudi cells. Human leukemic K562 cells were completely insensitive to IFN-alpha, beta and at the same time to IFN-gamma with regard to the antiviral, anticellular activities and to the induction of 2-5A synthetase.

A human leukocyte interferon preparation suppressed DNA synthesis and subsequent mitosis of synchronously growing virus-transformed human RSa cells. DNA synthesis was inhibited completely when interferon was added to cell cultures in late G1 phase, but was not so inhibited when interferon was added in G1/S boundary phase. Overall protein synthesis was reduced only slightly by interferon treatment. The findings suggested that critical events leading to the inhibition of DNA synthesis and subsequent mitosis by interferon action take place in late G1 phase or S phase.

Sensitivities of human transformed cell line RSa and its variant cell line IFr to the cytotoxicity of Con A were compared. IFr cells were more resistant than RSa to Con A. Con A-resistant cell lines, Con Ar-1 and Con Ar-3, were isolated from RSa, and they were slightly more sensitive than RSa cells to the cell growth-inhibitory actions of interferons. Agglutinability of RSa, IFr, and Con Ar cells by Con A was compared and found to be almost equal. The combined effects of Con A and interferon upon growth and viability of these cell lines were tested. When RSa and IFr cells were treated simultaneously with Con A and Le-IF, growth of the cells was suppressed more markedly than when treatment was with Con A or Le-IF alone. To clarify the mechanism of this phenomenon, binding of 125I-labeled Con A was examined. Though ther wee some differences, both leukocyte and fibroblast interferon enhanced the binding of Con A to RSa cells and also in Con Ar cells but, in interferon-resistant IFr and HEC cells, enhancement of Con A binding was low or not observed. Therefore, the combined effect of Con A and interferon on the inhibition of cell growth is not considered to be merely due to the enhanced binding of Con A by interferon action. Successive treatment of RSa or Con Ar cells with Con A and interferon did not enhance the antiviral action of interferon at all. On the contrary, simultaneous treatment with Con A and interferon suppressed the antiviral action of interferon, depending on the concentration of Con A used. Thus, the effect of Con A on the antiviral and cell growth-inhibitory actin of interferon seems rather different.

Human rhabdomyosarcoma cells chronically infected with retroviruses were examined for their responses to human interferon (HuIFN-α). Production of baboon endogenous retrovirus (M7) from A204 cells and feline endogenous retrovirus (RD114) from RD114 cells and from subclone RD114-Cl cells in each case was highly sensitive to the antiviral action of HuIFN-α. However, the antiviral responses of the cells after interferon treatment against encephalomyocarditis (EMC) virus and vesicular stromatitis virus (VSV) were different for each cell strain used. In A204 cells, replications of EMC virus and VSV were sensitive to interferon, but resistant in RD114 and RD114-Cl cells. Both 2′-5′-oligo(A) (2–5A) synthetase and dsRNA-dependent protein kinase were markedly increased in A204 cells after HuIFN-α treatment but no significant increase was observed in RD114 and RD114-Cl cells. In all these cells, HuIFN-a efficiently induced an anti-cell fusion state which was determined by inhibition of syncytium formation induced by uv-inactivated Sendai virus. These results indicate that the mechanisms underlying the anti-retrovirus and the anti-cell fusion activities of interferon may be closely related, and that they are different from those of antiviral action against exogenous virus infections.

The anti-viral and anti-cell fusion actions of human gamma interferon (IFN) were examined on human rhabdomyosarcoma cells and compared with the actions of IFN-alpha. Treatment of A204 and RD114-C1 cells with IFN-gamma resulted in significant inhibition of retrovirus production and cell fusions which were induced by Sendai virus, but IFN-gamma did not induce 2'-5' oligoadenylate (2-5A) synthetase or dsRNA-dependent protein kinase, and failed to inhibit EMC virus replication in RD114-C1 cells as previously observed on IFN-alpha treatment (Tomita, Y. et al. (1982) Virology 120, 258-263). Although IFN-gamma induced 56K protein more strongly than IFN-alpha in human transformed HEp-2, HeLa, RSa, IFr, and A204 cells, no significant induction of this protein was observed in RD114-C1 cells after IFN-alpha or IFN-gamma treatment. Specific bindings of 125I-labeled human IFN-alpha A to HeLa, A204 and RD114-C1 cell surfaces showed that the numbers of the binding sites on RD114-C1 cells were reduced to less than 22% of those on A204 cells. These results suggest that RD114-C1 cells exhibit a reduced number of receptors for IFN on the cell surface and that the receptors are functional for the expression of the anti-retrovirus and anti-cell fusion actions of IFN, but are not enough in number for expression of the anti-EMC virus action of IFN.

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.

We have studied neutralization of the antiviral activity of human fibroblast (IFN-beta) and immune interferon (IFN-gamma) by incubation with glycolipids (including the gangliosides GM1, GM2 and GM3, as well as various other glycolipids) and with the sialoglycoprotein, glycophorin. When 100 units/ml of IFN-beta were preincubated with 30-80 microM of the gangliosides, all antiviral activity was abolished. Similarly, 120 microM of glycophorin completely reversed the antiviral activity of 100 units/ml of IFN-beta. Glycolipids containing more than two sugars also showed moderate inhibitory effects. GM2 at a concentration of 200 microM almost completely inhibited the antiviral activity of 100 units/ml of IFN-gamma, but GM1, GM3 and glycophorin had only a moderate inhibitory effect. These results suggest that the terminal N-acetylneuraminic acid (NANA) residues of gangliosides and of glycophorin play an important role in the inhibition of IFN-beta, and that they may be similarly involved in the inhibition of IFN-gamma.

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.

A human nongestational choriocarcinoma cell line of ovarian origin (IMa) was established in vitro. This cell line had been subcultured serially more than 22 times over 18 months. Small polygonal cells with a prominent nucleus were dominant and a sparsity of cytoplasmic organelles was an ultrastructural characteristic of the IMa cells. The production and secretion of human chorionic gonadotropin and its subunits were identified by radioimmunoassay. The IMa cells were transplantable in the hamster cheek pouch and the histological diagnosis was choriocarcinoma. A newly established ovarian choriocarcinoma cell line can be considered useful for clarifying the biological differences between nongestational and gestational choriocarcinoma cells.

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.

The effect of retinoic acid (RA) on the level of interferon (IFN)-induced 2–5 oligoadenylate (2-5A) synthetase activity was examined in human histiocytic lymphoma U937 cells and WISH cells ** in order to ascertain the role of this polymerase in interaction between IFNs and RA. Cultures containing both IFNs (1–100 U/ml) and RA (0.1–10 μ M ) consistently had higher levels of enzyme activity than corresponding cells treated with IFN alone and this was true for all three types of IFNs in both cell lines. The potentiating effect of RA was dose- and time-dependent and under optimal conditions, the induction of the synthetase was synergistic between IFN-β (10–100 U/ml) and RA (0.1–10 μ M ). Furthermore, pretreatment (but not posttreatment) with RA followed by subsequent treatment with IFNs preferentially induced higher levels of enzyme activity in U937 cells but not in WISH cells. In addition, our results indicated that the modulating effect of RA on IFNs did not involve interaction at the receptor level and the level of enhancement of 2–5A synthetase activity was not in parallel with either cell-growth arrest or promotion of differentiation. Lastly, the present study raises the possibility that interactions between IFNs and RA, in either a synergistic or antagonistic manner, may be mediated through amplification of the 2–5A system.

Two cell lines derived from human choriocarcinomas (HCCM-5 and BeWo) are resistant to several biological effects of human interferon such as inhibition of VSV multiplication and inhibition of cell growth, but they develop a normal antiviral activity against EMCV. Nevertheless, in both cell lines, 2–5A synthetase and protein kinase are induced by IFN. 2–5A-dependent endonuclease can be measured by two independent methods and 2–5A itself is detected at least in poly(rI):poly(rC)- and IFN-treated BeWo cells. This is another example of two cell lines that are partially, with respect to the antiviral effect toward VSV, and totally, with respect to the anticellular effect, refractory to IFN treatment, although all the known elements of the 2–5A system are present.

Human enbryo cells were successively transformed by the Schmidt-Ruppin strain of Rous sarcoma virus (SR-RSV) and simian virus 40 (SV40) in vitro, and the double transformant HuE 13 RS was established. From this cell line the two clonal cell lines RSa and RSb were isolated. In both, presence of SV40 T antigens was demonstrated by the fluorescent antibody technique, and the presence of RSV genomes was verified in one RSb clone by focus formation after fusion with chick embryo cells. Growth of these cells was affected by dibutyryl cAMP without marked morphologic changes. Cells were extremely sensitive to the anticellular action of human leukocyte interferon.

Three kinds of human choriocarcinoma cell lines (BeWo, HCCM-5, and NUC-1) were used for examining the antiviral and antiproliferative activities of human interferons (IFNs) in vitro and in vivo. All of the cell lines showed only low sensitivity to the antiviral action of every IFN-alpha, IFN-beta, and IFN-gamma against vesicular stomatitis virus infection. However, 2-5A synthetase was normally induced by IFN-alpha in all of the cell lines. The [3H]thymidine incorporation of both BeWo and HCCM-5 cells was suppressed in dose-dependent manner at 48 hr after treatment with 1 to 1,000 units (U)/ml of IFN-alpha or IFN-beta and the growth of them was also slightly inhibited when treated continuously with 1,000 U/ml for 6 days in vitro. Another cell line NUC-1 was the least sensitive to these IFNs among the three cell lines. IFN-gamma did not show any antiproliferative effect on these cell lines. The intraperitoneal administration of 5000 or 10,000 U of IFN-beta suppressed the growth of xenografts developed in hamster cheek pouches and subcutis of nude mice when its administration was initiated on the first day of cell inoculation. These results indicate that although some heterogeneities exist among the cell lines choriocarcinoma cells are weakly sensitive to the antiproliferative activity of human IFNs.

Cultivation of Tsutsugamushi disease rickettsia was performed in the yolk sac of developing chick embryo by using 8 strains which were isolated in the endemic areas of Niigata, Japan. All these strains were possible to make passages in the yolk sac, but failed to demonstrate rickettsiae in the smears by staining except one strain, AX.From yolk sac infected with the AX strain, antigen for complement-fixation test was prepared by ether-or benzene-extraction method. Both of these antigens belong to soluble antigen and antigenicity was not demonstrable in the sediment from 10, 000r.p.m. centrifugation.Crude yolk sac vaccines were prepared from yolk sac heavily infected with rickettsia. Such vaccines had some degree of immunogenicity for mice but lost it after ether extraction.It was suggested that there might be antigenic heterogeneity among strains of rickettsia isolated at Niigata in the same year.

Characteristics of quinone-induced carcinoma and fructose-induced sarcoma were described from their susceptibility to neurotropic viruses. Fixed rabies virus did not propagate at all in either tumor. Since Col-SK and WEE viruses multiplied well in the carcinoma, their growth pattern in the tumor was analyzed, together with their invasiveness into the central nervous system.

By passing Col-SK virus serially from muscle to muscle in mice increase in myotropism and decrease of neurotropism is observed. The peripheral inoculation of the myotropic variant causes widespread necrosis of skeletal muscle, which, rather than the brain, is the principal site of viral multiplication. The virus neither multiplies nor produces histological change in the heart. Passage of the myotropic variant from brain to brain results in a reversion to the original neurotropism and a corresponding decrease in myotropism. The question, of whether this reversible change in pathogenicity of the virus is due to a forward-and-back mutation or to tissue-induced variation, is discussed.

Sarcomas were induced in AKR mice by Schmidt-Ruppin strain of Rous sarcoma virus (RSV). Sarcoma cells were passagedin vivo andin vitro, and their virogenicities were examined. They did not produce any infectious RSV, but when sarcoma cell suspensions were inoculated into wing webs of chickens, sarcomas were induced with high frequencies. Moreover, it was verified by physical and biological methods that clonal line (K3b) of AKR sarcoma cells cultivatedin vitro, release murine leukemia virus. These results suggest that RSV and murine leukemia virus genomes coexist in the K3b cells.

Interferon exerts antiviral and various non-antiviral activities on cells. For the development of the antiviral state in cells against exogenous infection of cytolytic viruses such as vesicular stomatitis virus and encephalomyocarditis virus (EMCV), induction of 2'-5'-oligo(A) synthetase or double-stranded (ds)-RNA-dependent protein kinase, or both, has been shown to have a crucial role (Revel 1979). However, in some cases, the antiviral effects of interferon are virus-specific (Nilsen et al. 1980; Samuel & Knutson 1981). Recently, different effects of mouse interferon on retrovirus production and on EM CV replication in a clonal cell line from NIH / 3T3 cells were reported (Gzarniecki et al. 1981).

When human transformed RSa cells were treated simultaneously with potent tumor promoter 12-O-tetradecanoyl-phorbol-13 acetate (TPA) and interferon (IFN) (20-100 IU/ml) at the concentration which suppresses the multiplication of cells slightly, distinct synergistic suppression of cell growth was observed. Synthesis of DNA in these cells was reduced in proportion to the concentration of IFN and it was more strongly reduced by combined treatment with IFN and TPA. When IFN was pretreated with anti-IFN serum, such synergistic effect was not observed. Highly purified IFN (specific activity, 2 X 10(8) U/ml) together with TPA elicited almost the same synergistic effect on RSa cells. Similar concentration of phorbol-12,13-didecanoate (PDD) also produced synergistic effects with IFN on RSa cells. In contrast, phorbol (PHR), which lacks tumor promoting activity, did not produce such synergistic effects even at a concentration of as high as 500 ng/ml. In the case of HeLa cells and IFN-resistant IFr cells, only an additive effect was observed. Effect of TPA on the antiviral action of IFN against VSV was tested and there was almost no influence of TPA on IFN action. In accordance with these results, 2-5A synthetase activity was not induced by TPA treatment and the level of enzyme action did not change significantly by the combined treatment with TPA and IFN.

Publisher Summary This chapter discusses the assay of effect of interferon on virus-induced cell fusion. Interferon (IFN) is known to exert multiple effects on cells. Suppression by IFN of virus-induced cell fusion is one of such pleiotropic actions of IFN. These anti-cell fusion actions are observed with human alpha, beta, and gamma IFNs, cloned human IFN-αA and mouse interferon, but all molecular species of IFNs from various animals probably exhibit this action. Paramyxoviruses and other enveloped viruses are known to fuse cultured cells in vitro . Cell-to-cell fusion of tissue culture cells is mediated by virus adsorbed to cells (fusion from without) or by viral glycoproteins expressed on the surface of infected cells (fusion from within). IFN seems to inhibit both types of cell fusion. Sendai virus is known as a most potent cell-fusing virus, and such cell fusion is known as a typical fusion from without. The chapter describes methods for the assay of effect of interferon on cell fusion induced by Sendai virus and several retroviruses. Retroviruses from various species of animals, including humans induce formation of syncytia in certain indicator cells. Cocultivation of retrovirus releasing cells with indicator cells also produces syncytia, but for quantitative experiments, use of cell-free virus preparations is desirable.

Summary 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.

Summary When transformed human embryonic cells, RSa and RSb, were treated for 48 h with both 10-7 m-ouabain and 1000 units/ml of human leucocyte interferon, there was an additive suppression of cell growth. In contrast, the antiviral action of the human interferon was inhibited by this concentration of ouabain. Similar effects of combined treatment with interferon and ouabain were found with IFr cells (which are relatively resistant to the anticellular effect of interferon), and with mouse cells of the K3b line.

We examined the sensitivity of four human germ-cell-tumor cell lines exhibiting different stages of differentiation to human interferons (IFNs) in vitro. The cell lines were derived from two embryonal carcinomas (NEC 8 and NEC 14), a choriocarcinoma (IMa), and a yolk-sac tumor (HUOT). Treatment with poly I:C induced IFN production in IMa and HUOT cells, but not in NEC-8 and NEC-14 cells. In the two embryonal-carcinoma cell lines, the addition of IFN-alpha, IFN-beta, and IFN-gamma did not prevent infection by vesicular stomatitis virus and encephalomyocarditis virus. Also, in these two lines, 2-5A synthetase was not induced by the addition of IFN-alpha. In contrast, both IMa and HUOT showed sensitivity to the antiviral action of IFN-alpha and IFN-beta against the two viruses, and 2-5A synthetase was induced by IFN-alpha. IFNs added at doses of up to 1000 IU/ml had no antiproliferative effect on NEC 8, NEC 14, and HUOT, whereas colony formation by IMa cells was greatly suppressed by all three forms of IFN. These results indicate that the production of and sensitivity to IFN are developmentally regulated and are related to the level of differentiation of human germ-cell stem cells.

The effect of retinoic acid (RA) on the antiviral activity of interferons (IFNs) alpha and beta in the U937 and WISH cells was examined to ascertain whether or not RA could reduce the effectiveness of IFN-induced resistance to viral infection. Our results indicate that in the U937 cells, RA (0.1-1.0 microM) had neither enhancing nor suppressive effect on the antiviral activity of IFN-alpha or beta against the Semliki Forest virus (SFV). However, in the WISH cells, RA had different effects on IFNs alpha and beta. Thus, while RA (0.1-50 microM) invariably suppressed the activity of IFN-alpha, it enhanced the action of IFN-beta at low dose (0.1-1.0 microM) but became suppressive at higher concentrations (greater than or equal to 10 microM). Furthermore, higher antiviral activity was consistently obtained when RA (0.1-10 microM) was added prior to either IFN-alpha or IFN-beta comparing to cultures with IFN alone. In addition, direct correlation between antiviral activity and the amplitude of 2-5 oligoadenylate (A) synthetase activity was not observed. These results suggest that modulation of IFN antiviral activity by RA varies with different systems and is dependent on the sequence of treatment.

Summary Human leucocyte interferon has antiviral activity and anticellular effects on the transformed human cell lines, RSa and RSb. Treatment of the cells with cycloheximide or puromycin at 0·5 to 5·0 µg/ml suppressed the antiviral action of interferon but increased its anticellular effects. Interferon also has antiviral activity in IFr cells, but this is relatively resistant to its anticellular action. Nevertheless, both drugs suppressed the antiviral activity of interferon and increased its anticellular action.

F-IFr cells, which are more resistant to the anticellular and antiviral action of human alpha and beta interferons (IFN-alpha and IFN-beta) than the parental RSa cells, were also found to be more resistant to both the anticellular and antiviral effect of IFN-gamma. A high level of 2-5A synthetase was induced by treatment with IFN-alpha or -beta, but induction of 2-5A synthetase was not detected after IFN-gamma treatment of the cells. F-IFr cells had less than half the number of specific binding sites for IFN-alpha than the parental RSa cells.

Human cells doubly transformed by Rous sarcoma virus and SV40 (RSb cells) formed syncytia by cocultivation with Mason-Pfizer monkey virus (MPMV)-producing cells. This cell fusion was blocked by anti-MPMV serum indicating that the phenomenon is MPMV specific. The RSb cells were successfully used for MPMV infectivity assay in the same manner as KC cells.

When uterine cavities of pregnant rats were treated with pellets of 7,12-dimethylbenz[a]anthracene, histologically different types of malignant tumors were induced during a 12 month observation period. Tumors produced at 219 and 314 days after treatment, which were diagnosed histologically as adenocarcinoma, were cultivated in vitro. After 2 months of tissue culture, colonies consisting of epithelial cells appeared and showed active proliferation. They were transplantable to isologous hosts, and metastases to visceral and pulmonary regions were observed. Histology of the tumors and metastases produced was adenocarcinoma in all the cases as observed in original tumors. From this parent culture, two apparently distinct types of cloned cells were obtained: a high tumor-producing clone (HTP clone) and a low tumor-producing one (LTP clone). HTP clone was isolated from the colony in a semisolid agar medium, and LTP clone was isolated from the colony in liquid medium. They were different in colony-forming ability in semisolid agar medium in vitro and tumorigenicity to isologous hosts in vivo.

In contrast with the Schmidt-Ruppin strain of RSV, Bryan strains are not oncogenic for ducks. From the Bryan standard strain (BS-RSV) a variant virus oncogenic for ducks was isolated by passing BS-RSV through cortisone-treated ducklings together with chick embryonic tissues. From the Bryan high titer strain (BH-RSV) such a variant virus could not be isolated. The variant virus (BK-RSV) is also oncogenic for mice and hamsters, whereas parent strain lacks such oncogenicity for mammals. Thus, there may be some correlation between oncogenicity for ducks and mammals. Mouse or hamster tumors induced by the BK strain did not produce infectious RSV eitherin vivo orin vitro. However, when hamster tumor cells in appropriate amounts were inoculated in the wing web of chickens, sarcomas were induced yielding infectious RSV. Thus, the RSV genome persists in BK-induced mammalian tumor cells and is able to produce infectious virus in the presence of chicken cells.

The duck-adapted variant of the Rous sarcoma virus, strain 14(d)7, is oncogenic in chickens and ducks to the same degree, and this even after passages in chickens, ducks or chick embryo-grafted hamsters. Heat-inactivated 14(d)7 virus has no influence upon oncogenicity of Rous sarcoma virus in either species of birds.

Trombicula (Leptotrombidium) fuji n. sp. Belonging to the "tsutsugamushi group" of chigger mites, this species is distinguished by its very sparsely punctate, unusually small scutum with the sensillary bases well behind the line of the postero-lateral setae, the latter situated in advance of the median line, the sensillae feathered on the outer two-thirds, barbed on the distal half of the shank, and a bare seta on palpal segment III which exceeds the tip of the palpal claw in length (Fig 1).