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Lung adenocarcinoma (LAC) is a common and aggressive form of lung cancer that is increasing in incidence among never smokers at a younger age. Current treatment of patients with LAC is insufficient and there is a need for identification of effective biomarkers and development of therapeutic targets. These demands require also improved models for in vivo and in vitro experimentation. In this study, we describe the establishment of two LAC cell lines, named LuCa-3 and LuCa-6. Both were derived from pleural effusion (PE) cells of LAC patients (L3 and L6) and readily propagated as tumor xenografts in immunodeficient mice. PE cells from the patient L6 exhibited also the capacity for in vitro growth and were cultured in two forms: (i) as a suspension growing cell population, labeled LuCa-6S, composed of non-clumping single cells; and (ii) as a monolayer-like culture, labeled LuCa-6A, exhibiting tight cell-to-cell and to culture surface adherence. Unique features of these two sublines and their cell clones are the capacity to convert from a non-clumping single-cell suspension into the adherent growth pattern and vice versa. Immunostaining of patients' tumor tissue xenografts and cultured subline cells displayed markers specific for the phenotype of human LAC. LuCa-6S and LuCa-6A cells did not reveal a noticeable disparity in quantitative growth characteristics. However, a number of differences were detected between these two cell populations manifested in detection or intensities of antigen expressions on the cell surface (CD133, SFTPC) and in the nucleus (TTF-1) including pluripotent (OCT-4, SOX-2, NANOG) genes in cancer stem-like cells (CSCs). Dissimilarities between these two sublines were also detected in N-glycan profiles and in the sensitivity to natural killer cells. Salient features of these subline cell populations are responsiveness to selective upregulation of the pluripotent genes in subsets of CSCs via conversion of their growth patterns and/or by using culture stem media with growth factors. The described in vivo/in vitro model enables broader experimental approaches in studies of lung adenocarcinoma.

The major internal protein of bovine leukemia virus (BLV p24) was isolated using ion exchange chromatography on phosphocellulose and gel filtration. The specificity of the BLV p24 isolated was checked by both the radioimmunoprecipitation (RIP) and the competitive radioimmunoassay (RIA). No cross reactivity between BLV p25 and the mammalian viruses of type C and D and the retrovirus isolated from human myeloma cells RPMI8226 [8, 17] was detected. The analysis of 429 leukemia-suspected bovine blood sera resulted in the detection of 165 (38.5%) positive and 264 (61.5%) negative blood sera. A correlation of the results of radioimmunoprecipitation reaction of major internal protein p24 and immunodiffusion test on the glycoprotein antigen of BLV was observed.

Human sarcoma associated antigens (HSAA) have previously been identified by indirect immune fluorescence in human sarcoma cells in culture using sera from patients bearing different types of sarcoma. To further characterize these HSAA, surface proteins of cultured cells were labeled with 125Iodine, [3H]-glucosamine and [35S]-methionine and solubilized. After immunoprecipitation labeled proteins were detected in immune complexes by SDS polyacrylamide gel electrophoresis and autoradiography, which allowed comparison with antigens described by other groups. A surface protein (Mr 96 000) was precipitated with sera from sarcoma bearing patients, and two glycoproteins (Mr 115 000 and 85 000) were preferentially precipitated with antisera from rabbits immunized with membranes from two human sarcoma cell lines. At least two of these proteins were found in each of five human sarcoma cell lines studied (U-4SS, U-3930S, U-20S, B-5GT and B-6FS). None of the proteins were precipitated with three human control sera, and only occasionally a faint band was observed in immunoprecipitates from control cells (B-25F, B-41B, B-42FC, U-2S, and U-393S with the immune sera. These proteins are probably some of the antigens responsible for the immune fluorescence observed in determination of HSAA. However, purification of the proteins and competition experiments are needed before this can be finally established.

A comparison was made of the cell surface glycoproteins of four human cell lines, namely a giant tumor of bone cell line, an osteosarcoma line, a fibrosarcoma line and a human fibroblast line. The cells were labeled by lactoperoxidase catalyzed iodination and the glycoproteins extracted by 0.5% Triton X-100 were bound to lentil-lectin and subsequently analyzed by SDS gel electrophoresis. While the cell lines examined shared a series of common glycoproteins, it was found that the giant cell tumor line and the fibrosarcoma lines exhibited a higher degree of homology than the other cell lines.

The establishment and cultivation of two human neoplastic cell lines is described. The cell line B-5GT was derived from bone giant cell tumor and B-6FS from poorly differentiated fibrosarcoma. In comparison to the normal skin fibroblasts both cell lines have a potential for "indefinite" multiplication in vitro and they exhibit growth properties which are associated with malignant transformation. The parameters investigated included cell morphology, chromosome characteristics, terminal cell density, growth pattern, residual DNA synthesis and growth in soft agar. Both cell lines exhibited human karyotype with aneuploidy and differed in their karyotype from each other.

The retrovirus designated RPMI8226V (isolated from human myeloma cells RPMI8226) has been characterized with respect to its morphological, biochemical and immunological properties as well as its propagation in various animal and human cells. The myeloma cells RPMI8226 produce intracytoplasmatic A-type particles and extracellular particles. The extracellular particles have been classified as immature particles with translucent core center, typical mammalian C-type virus particles and C-type particles with intermediate membrane. However, the budded particles in secondarily infected human neoplastic cells contained complete doughnut-shaped nucleoids. This type of budding is rather characteristic for B-type particles. The 3H-uridine labeled RPMI8226 viral particles have a buoyant density 1.17 g/ml in sucrose gradient containing high molecular weight RNA and the distribution of viral structural proteins in SDS-PAGE is characteristic for oncornaviruses. The internal structural proteins according to MW are ranged from 13 000 to 30 000 daltons. The virus contains a magnesium-dependent reverse transcriptase. The cellular homogenate and viral concentrate from RPMI8226 cultures do not react with antibodies against ALSV, MuLV, FeLV, RD114, MP-MV and SiSLV. The only reaction was scored with anti BLV antibodies. However, anti BLV serum inhibiting the reverse transcriptase activity of BLV to 60% does not cross-react with the reverse transcriptase of RPMI8226V. In contrast to BLV concentrates, neither XC nor KC cells show syncytia formation by RPMI8226V. The RPMI8226V replication is restricted to human tumor and normal human glia-like cells. The possible origin of the virus is discussed.

The plasminogen activator (PA) production and the capacity to inhibit embryonic neural retina (NR) cell aggregation by human normal and neoplastic cell lines have been studied. The PA production was detected by both iodinated fibrin and casein lysis assays, and by changes in cell morphology at the presence of activated PA, using dog serum. Since the casein lysis assay and morphological changes proved to be less sensitive than 125I-fibrin lysis assay, a good correlation between these three assays could be observed provided that PA production measured by fibrinolysis exceeded 10--20%. The neoplastic cell lines exhibited the PA production to quite a large extent. The highest fibrinolytic activity (78%) was found in the case of bladder carcinoma cells T24, while the B-5GT cells from giant cell tumor of bone failed to produce any detectable amount of the PA. The cells from synovial sarcoma and both glioma lines exhibited fibrinolytic activity of about 10% and four sarcoma cell lines over the range 20--50%. Out of 13 normal cell lines tested, 7 were negative or exhibited very low fibrinolysis not exceeding 3% of total radioactivity. Four cell lines derived from kidneys, lungs, intestines, and from mixed embryonic tissues showed a marked fibrinolytic activity of about 10--37%, a slightly elevated fibrinolysis being found in embryonic lung cells LEP and cells from fetal skin tissue only at the presence of dog serum. The fibrinolysis detected in the neoplastic cloned cell populations showed considerable differences in the PA production between individual cell clones isolated from the same parental cell line. Unlike the normal fibroblastic cells B-41FB derived from bone, all neoplastic cell lines tested possess the capability to inhibit embryonic NR cell aggregation significantly. The results suggest the effect not to be dependent upon the PA production.

Murine xenotropic virus, designated 698/X, was recovered by implantation of human lymphosarcoma-derived cells U-698M into nude mice of Giovanella's colony. The budded and extracellular particles revealed typical type-C morphology, the latter possessing reverse transcriptase (RT) activity and exhibiting a buoyant density 1.17 g/ml in sucrose gradient. In competitive radioimmunoassay using iodinated p30 of Rauscher MuLV, the 698X viral concentrate and cell extracts of both implanted lymphosarcoma cells (U-698M-N-1 and U-715M-N-1) were as effective as Gross MuLV, thus indicating the murine origin of the virus. The propagation of the 698/X virus in five human, four mouse (permissive for N- and B-tropic MuLV), two rat and one bovine cell lines was followed by RT, XC syncytia assays and EM investigations. The replication of the 698/X virus seems to be restricted mainly to both human lymphosarcoma-derived cell lines U-698M and U-715M. The new recovery of the virus from the nude mouse by implantation of U-175M cells has asserted its high tropism to human lymphosarcoma cells and its murine origin. The comparative response of mouse, rabbit and rat cells exposed to both NZB and 698/X xenotropic murine viruses exhibited host range differences between these viruses. The rabbit SIRC and rat embryonic cells REC were fully permissive for the murine xenotropic NZB virus, while low viral production was detected by RT assay only in 698/X virus infected SIRC cells.

The infection of cat embryonic kidney cells (CRC) with Gardner and Snyder-Theilen strains of feline sarcoma viruses (Ga-FeSV and ST-FeSV) do not lead to transformation and focus formation. The focus forming insusceptibility is shown to be stable and nontransmissable property of these cells. However, the CRC cells are permissive for ST-FeSV, and the viral production is 10 to 100 folds lower than in focus forming susceptible cat cells. Produced ST-FeSV(c) by CRC cells induced on cat embryonic cells FE in higher proportion round and fusiform (rf) foci and fusiform(f) foci. The f foci were not previously observed in original stock of ST-FeSV tested directly on FE cells. Attempts to detect some differences in morphology or in growth behavior between ST-FeSV infected and non-infected CRC cell population gave negative results.