Biomedical subjects
Publications and source records attributed to S Pan.
Cell lines transformed by simian virus 40 mutant F8dl (deleted from 0.168 to 0.424 map units, corresponding to the carboxy-terminal 62% of the wild-type simian virus 40 large tumor antigen) are tumorigenic in nude mice. Four of five C3H10T1/2 cell lines transformed by F8dl were tumorigenic in nude mice, whereas two of two wild-type transformants were tumorigenic.
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The human small cell (oat cell) carcinoma line, SHP-77, established by Fisher and Paulson in 1977 and originally described as a "large cell variant of oat cell cancer" has been evaluated by several different parameters and shown even after more than 200 passages to retain properties described for the original cell line. Karyotypic, histological, and biochemical features are retained, as well as tumorigenicity in nude mice. The original authors' suggestion that this is a propitious cell line for both in vitro and in vivo studies is supported by this report. Modulation of growth characteristics in vivo (in xenografts) emphasizes the plasticity of this unique line which serves as a valuable model for basic as well as therapeutic studies. SHP-77 can serve as an in vitro target in 51Cr and 111In release cytotoxicity assays as well as in in vivo nude mouse assays for evaluating immune reactivity of cells and serum from lung cancer patients. The potential histological variability of SHP-77, despite its biochemical stability, calls attention to the inadequacy of histological criteria for lung tumor classification.
Simian virus 40 (SV40), which transforms mouse cells in vitro, has not been previously observed to cause tumors when injected in immunocompetent mice. We have investigated both the fate of the injected virion in mice and several immunological parameters as potential factors controlling tumorigenicity. We find that although SV40 does not replicate in mouse cells, the viral DNA can persist for many months postinjection; the majority of the viral DNA is found in the cytoplasm, but a small amount of the viral DNA is integrated at multiple sites in the host nuclear DNA. The persistence of the viral genome is independent of the ability of the mouse to mount an SV40 TSTA specific cytotoxic T-cell response and may be attributed to the cytoplasmic location of the majority of the viral genome. However, in long-term studies of SV40-injected mice, genetically identical except for the major histocompatibility complex, we find that tumors were induced in some mice of the H-2d (low cytotoxic T-lymphocyte responder to SV40 TSTA) but not of the H-2k (high responder to SV40 TSTA) haplotype. Thus, a combination of inefficient disposal of the injected virion and inefficient immunological surveillance and elimination of cells containing nuclear SV40 DNA can eventually result in SV40-induced tumors at multiple sites in mice.
The recognition by cytotoxic T lymphocytes (CTL) of herpes simplex virus (HSV) glycoprotein(s) in association with the H-2K and the H-2D gene products of the H-2b complex was examined by using cell lines derived from H-2 recombinant mice as target cells, and by using H-2 recombinant mouse strains for the generation of HSV-specific CTL populations. CTL from H-2b HSV-immunized mice were found to lyse HSV-infected B6/WT-3 (KbDb) and K5RSV (KbDd) cells, but not KHTGSV (KdDb) cells. Unlabeled HSV-infected K5RSV cells were as efficient in competing for specific CTL lysis of 51Cr labeled HSV-infected B6/WT-3 cells as unlabeled B6/WT-3 cells themselves, whereas infected KHTGSV cells were ineffective. Furthermore, CTL generated in H-2 recombinant mice containing the H-2Kb allele (KbDd) effectively lysed infected B6/WT-3 cells; no specific lysis was observed with immune lymphocytes from those mice containing the H-2Db (KkDb) allele. Limiting dilution analysis of the interleukin 2 (IL 2)-dependent, antigen-independent, CTL precursor populations showed that CTL precursors giving rise to H-2Kb-restricted progeny were present at a relatively high frequency, whereas H-2Db-restricted progeny were present at low frequency or were undetectable. Target cells carrying mutations in the H-2Kb glycoprotein (H-2Kbm1 and H-2Kbm8) infected with HSV-1 were found to be drastically reduced in their ability to be lysed by anti-HSV-1 CTL. HSV-1-infected H-2Kbm5 cells provided a good target for anti-HSV CTL. We conclude that the HSV-specific glycoprotein(s) are recognized by CTL primarily in association with the H-2Kb gene product.
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Limit-dilution cultures were used to select vaccinia-immune T-cell populations from bm1 and bm3 mutant mice that were not lytic for virus-infected targets expressing the Kb and Db MHC glycoprotein. Approximately 30% of virus-immune CTL were restricted in each case to Kbm1 and Kbm3, rather than to Db. Evidence of extensive cross-reactivity was found for these virus-immune CTL. Bm3 and bm11 mice sharing one amino acid mutation from wild-type but differing by a second mutation seen only in bm3 are the most cross-reactive pair in their presentation of vaccinia. The bm1 and bm10 pair with dissimilar mutations from wild-type affecting the same CNBr fragment are also largely cross-reactive. However, 30% cross-reactivity is also found for bm1 and bm3, which differ in separate CNBr fragments. That mutants expressing amino acid substitutions in the same region of the peptide tend to show more evidence of cross-reactivity does not necessarily mean the T cells see linear arrays of amino acids on the MHC glycoprotein. For instance, Kbm1 and Kbm10 differ for three amino acids, but bm1 T cells are highly lytic for bm10 virus-infected targets. However, there is no cross-reactivity for Kbm1 and Kb, which differ at only two amino acids. The key to further understanding may rest with defining the nature of the conformational differences among the Kbm1, Kbm10, and Kb glycoproteins.
The lysis of SV40-transformed target cells by a cloned cytotoxic T cell line (CTB6) that is SV40 TASA-specific and H-2Kb-restricted is blocked by a monoclonal antibody reactive with the SV40 T-antigen. The blocking is maximized at high antibody concentrations and low effector-to-target cell ratios. These data indicate that at least one antigenic determinant of the SV40 T-antigen molecule is expressed at the cell surface and is either recognized by this cytotoxic T lymphocyte or is proximate to that determinant recognized by this cytotoxic T lymphocyte.
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The cytotoxic T lymphocyte (CTL) responses directed towards SV40 tumor-associated specific antigen (TASA) in nine strains of spontaneously arising Kb mutant mice were analyzed. All nine mutants generated normal levels of H-2Db-restricted response, but the K-end-restricted CTL response varied. B6.C-H-2bm1 (bm1) did not produce K-end-restricted SV40 TASA-specific CTL upon immunization, and SV40-transformed bm1 cells were not lysed by intra-H-2 recombinant Kb [B10.A(5R)] CTL. Nonreciprocal cross-reactive lysis was seen between B6-H-2bm8 (bm8) and B10.A(5R). Strain B6-H-2bm8 mice produce highly specific Kbm8-restricted CTL that lyse SV40-transformed bm8 cells (Kbm8SV) but not B10.A(5R) target cells (K5RSV), although Kbm8SV targets can be partially lysed by B10.A(5R) CTL. The other seven Kb mutants cross-react with B10.A(5R). These experiments definitively show that genes mapping to the K and/or D region directly control the H-2-restricted CTL response to SV40 TASA.
BN rats were irradiated with 500 rad of 60Co 1 day before immunization with 0.1 mg of ovalbumin in alum hydroxide gel. The onset of reaginic and haemagglutinating antibody synthesis was suppressed to a non-detectable level for at least 3-4 weeks. When these irradiated rats were injected intraperitoneally with 10(8) viable or sonicated thymocytes 1 day after irradiation, the suppressed reaginic and haemagglutinating antibody synthesis was successfully restored. This suggests that: (i) thymocytes can restore the radiation-induced immunosuppression, but viability of thymocytes is not essential in this immune restoration; (ii) active biological molecules exist in the cytoplasmic pool of the normal thymocytes which can restore the radiation-induced immunosuppression; and (iii) the thymus not only plays a central role in the normal morphological development of the lymphoid system and its functional immunological maturation, but may also play an important role in the reversal of radiation-induced immunosuppression. To investigate tissue specificity of the factors that are actively engaged in the restoration of the suppressed immune response, peripheral lymphocytes, spleen cells, bone marrow cells and kidney cells were also tested. The results demonstrated that both peripheral lymphocytes and spleen cells could restore the suppressed immune response, but not bone marrow cells or kidney cells. This suggests that the active factors may be present only in T lymphocytes. Since the active factors could be found in the tissues other than thymocytes and non-stimulated lymphocytes, they appear to be different from thymosin, transfer factor or lymphokines derived from sensitized lymphocytes, and they were most likely not synthesized de novo.
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Murine embryonal carcinoma cells (ECCs) do not express antigens of the major histocompatibility complex (H-2), but do express cell-surface molecules shared with early embryos. ECCs are also characterized by their insusceptibility to infection by various oncogenic viruses, and their ability to differentiate into a variety of adult cell types. Differentiation of ECCs in vitro can occur spontaneously or can be induced. On exposure to retinoic acid the ECC line F9 (ref. 13) differentiates into cells which have the characteristics of parietal endoderm. When ECCs are exposed to simian virus 40 (SV40), the SV40 tumour (T) antigen is not expressed, although the virus genome reaches the nucleus, and a primary transcript of the SV40 A gene is made. However, following exposure to retinoic acid, the differentiated cells, like most mouse somatic cells, are susceptible to SV40 abortive infection and synthesize large T and small t antigens. To monitor the molecular events associated with the expression of the SV40 A gene on differentiation, we have constructed an ECC line (F9 12-1) containing a single integrated copy of the SV40 genome. This was accomplished by introducing a recombinant plasmid consisting of pBR322 linked to the herpes simplex type 1 thymidine kinase gene and SV40 genome into a thymidine kinase-deficient F9 cell line. We report here that in F9 12-1 cells exposed to retinoic acid, synthesis of the SV40 A gene product(s), T and tumour-associated specific antigens (TASA), parallels the appearance of the normal hallmarks of differentiation in this cell line, H-2 antigens and the basement membrane protein laminin.
The interrelationships among fatness measures, plasma triglycerides and high density lipoproteins (HDL) were examined in 131 normal adult subjects: 38 men aged 27-46, 40 men aged 47-66, 29 women aged 27-46 and 24 women aged 47-66. None of the women were taking estrogens or oral contraceptive medication. The HDL concentration was subdivided into HDL2b, HDL2a and HDL3 by a computerized fitting of the total schlieren pattern to reference schlieren patterns. Anthropometric measures employed included skinfolds at 3 sites. 2 weight/height indices and 2 girth measurements. A high correlation was found among the various fatness measures. These measures were negatively correlated with total HDL, reflecting the negative correlation between fatness measures and HDL2 (as the sum of HDL2a and 2b). Fatness measures showed no relationship to HDL3. There was also an inverse correlation between triglyceride concentration and HDL2. No particular fatness measure was better than any other for demonstrating the inverse correlation with HDL but multiple correlations using all of the measures of obesity improved the correlations. Partial correlations controlling for fatness did not reduce any of the significant correlations between triglycerides and HDL2 to insignificance. The weak correlation between fatness and triglycerides was reduced to insignificance when controlled for HDL2.
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